PK"|9,refs.MYD;~?(Deng, M. Lancto, C. A. Abrahamsen, M. S.2004JCryptosporidium parvum regulation of human epithelial cell gene expression73-82Int J Parasitol341JaniCryptosporidium parvum is an obligate intracellular protozoan capable of causing life-threatening diarrhoeal disease in immunocompromised individuals. Efforts to develop novel therapeutic strategies have been hampered by the lack of understanding of the pathogenesis of infection. To better understand the host response to C. parvum infection, gene expression profiles of infected human ileocecal adenocarcinoma cells were analysed by using Affymetrix oligonucleotide microarrays containing probe sets for 12,600 human genes. Statistical analysis of expression data from three independent experiments identified 223 genes whose expression was reproducibly regulated by C. parvum infection at 24 h post-inoculation (125 up-regulated and 98 down-regulated), 13 of which were validated by quantitative reverse transcriptase polymerase chain reaction analysis. This analysis revealed the consistent up-regulation of host heat-shock genes and genes for pro-inflammatory chemokines IL-8, RANTES, and SCYB5. Multiple genes for host actin and tubulin genes were up-regulated whereas genes for actin binding proteins were down-regulated, confirming previous observations of host cytoskeleton rearrangement in response to C. parvum infection. In addition, host genes associated with cell proliferation and apoptosis were differentially regulated, reflecting the complexity of host-parasite interaction. Together, this study demonstrated that C. parvum infection results in significant changes in host biochemical pathways and provides new insights into specific biological processes of infectious disease caused by an intracellular protozoan parasite.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=147115920020-7519 Journal Article14711592Department of Veterinary PathoBiology, College of Veterinary Medicine, University of Minnesota, 1988 Fitch Avenue, St Paul, MN 55108, USA.~?jGomez-Couso, H. Freire-Santos, F. Amar, C. F. Grant, K. A. Williamson, K. Ares-Mazas, M. E. McLauchlin, J.2004YDetection of Cryptosporidium and Giardia in molluscan shellfish by multiplexed nested-PCR279-88Int J Food Microbiol913Mar 15A multiplexed nested-PCR procedure (ABC-PCR) previously developed to detect Cryptosporidium spp. and Giardia duodenalis assemblages A and B in whole human faeces was applied to DNA extracted from filter-feeding molluscs. Species of Cryptosporidium and G. duodenalis were identified by restriction fragment analysis of the PCR products and by DNA sequencing. The extraction and ABC-PCR procedures were shown to be suitable for application to shellfish by amplification of specific target sequences using DNA from Cryptosporidium parvum genotype 2 and G. duodenalis assemblages A and B which were spiked into DNA extracted from mussels. Using 49 molluscan shellfish specimens (18 clam, 22 mussel and 9 oyster samples) from Spain, cryptosporidial oocysts were detected in 56% by immunofluorescence microscopy, and in 44% by ABC-PCR. For detection of Cryptosporidium, there was a significant association, but not total agreement, between the results of microscopy and PCR. G. duodenalis assemblage B was detected from one oyster sample by PCR. Amongst 38 specimens (20 mussel and 18 cockle samples) collected in the UK and tested by the ABC-PCR, G. duodenalis was not detected, and Cryptosporidium was detected in 11% of the samples. Overall, the 26 samples where Cryptosporidium was detected, C. hominis/C. parvum genotype 1 was detected in 1, C. parvum genotype 2 in 22, and the remaining three samples contained either sequences similar to C. parvum genotype 2 or heterogeneous mixtures of Cryptosporidium species. There was no significant association between the level of Escherichia coli detected by conventional microbiological methods and the presence of Cryptosporidium detected by ABC-PCR.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=149847750168-1605 Journal Article14984775Laboratorio de Parasitologia, Departamento de Microbiologia y Parasitologia/Instituto de Acuicultura, Universidad de Santiago de Compostela, 15782 Santiago de Compostela, A Coruna, Spain.H~?YGomez-Couso, H. Freire-Santos, F. Martinez-Urtaza, J. Garcia-Martin, O. Ares-Mazas, M. E.2003hContamination of bivalve molluscs by Cryptosporidium oocysts: the need for new quality control standards97-105Int J Food Microbiol871-2Animals Consumer Product Safety Cryptosporidium/*isolation & purification/physiology European Union *Food Contamination Food Parasitology Mollusca/*parasitology Oocysts/growth & development Quality Control Seasons Shellfish/*parasitology/*standards Spain Support, Non-U.S. Gov'tOct 15A yearlong study was carried out to investigate the presence and viability of Cryptosporidium oocysts in 203 samples of cultured shellfish from Galicia (NW Spain) and 38 samples imported from other European Union (EU) countries. Shellfish samples included mussels, oysters, clams and cockles. Cryptosporidium oocysts were detected, using a direct immunofluorescence antibody test (IFAT), in 34.4% of the samples analyzed; use of the fluorogenic dye propidium iodide (PI) revealed viable potentially infective oocysts in 53.0% of these samples. There was no relation between the presence of Cryptosporidium oocysts and the microbiological contamination detected in the samples expressed as Most-Probable-Number (MPN) of fecal coliforms, the different species of mollusc, or the month of sampling. One important finding was that the depuration process was ineffective in totally removing oocyst contamination. Furthermore, the existence of viable oocysts in samples with microbiological contamination levels lower than 300 fecal coliforms/100 g, which in accordance with current legislation are considered suitable for human consumption, suggests the need to include parasitological analyses in the quality control for these molluscs.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=129277110168-1605 Journal Article12927711Laboratorio de Parasitologia, Departamento de Microbiologia y Parasitologia, Instituto de Acuicultura, Universidad de Santiago de Compostela, 15782 Santiago de Compostela, La Coruna, Spain. ~?Moss, D. M. Arrowood, M. J.2001Quantification of Cryptosporidium parvum oocysts in mouse fecal specimens using immunomagnetic particles and two-color flow cytometry406-12 J Parasitol872Animals Cattle Cattle Diseases/diagnosis/parasitology Cell Separation/methods/veterinary Cryptosporidiosis/diagnosis/parasitology/*veterinary Cryptosporidium parvum/*isolation & purification Feces/*parasitology Flow Cytometry/methods/*veterinary Immunomagnetic Separation/methods/*veterinary Mice Mice, Inbred BALB C Parasite Egg Count/methods/*veterinary Rodent Diseases/diagnosis/parasitologyAprAlthough single-color flow cytometry has been shown to be more sensitive than fluorescence microscopy for the quantification of Cryptosporidium parvum oocysts, this method has not been optimized. Monoclonal antibody OW50, specific to the cell wall of oocysts, was conjugated to superparamagnetic particles, to fluorescein isothiocyanate, and to r-phycoerythrin. The oocysts were then double stained with the fluorochrome-labeled OW50 and were placed in tubes with known numbers of highly fluorescent polystyrene beads, allowing quantification of the oocysts without dependence on acquired sample volume by flow cytometry. Data from 2-color flow cytometry using logical gating of the oocysts and beads showed a linear relationship between dilutions of a purified oocyst suspension and the mean numbers of oocysts detected (r2 = 1.00). An average of 15 purified oocysts/ml were counted in a dilution with a theoretical concentration of 12 oocysts/ml. Known numbers of purified oocysts were seeded into normal mouse fecal specimens, captured by OW50-labeled immunomagnetic particles, eluted with 5% potassium dichromate at low pH, and double stained with fluorochrome-labeled OW50. By flow cytometry, the mean recovery was 43.1% (+/-8.3%), and as few as 133 oocysts were detected. The captured and eluted oocysts were infective in neonatal BALB/c mice. This 2-color flow cytometry method, used in conjunction with the capture and elution of oocysts by and from immunomagnetic particles, provides a powerful tool for not only the quantification and purification of C. parvum oocysts from different sources but also for the characterization of oocysts in vitro and in vivo.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11318573,0022-3395 Evaluation Studies Journal Article11318573iDivision of Parasitic Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia, 30341, USA. ~?<Call, J. L. Arrowood, M. Xie, L. T. Hancock, K. Tsang, V. C.2001[Immunoassay for viable Cryptosporidium parvum oocysts in turbid environmental water samples203-10 J Parasitol871iAnimals Antibodies, Monoclonal/immunology Antibodies, Protozoan/immunology Antigens, Protozoan/analysis Cryptosporidiosis/parasitology Cryptosporidium parvum/*growth & development/immunology/*isolation & purification/pathogenicity Environmental Monitoring/*methods Fresh Water/*parasitology Human Immunoassay/methods Sensitivity and Specificity *Water PollutionFeblCryptosporidium parvum oocysts in drinking water have been implicated in outbreaks of diarrheal disease. Current methods for monitoring environmental exposures to C. parvum only account for total number of oocysts without regard for the viability of the parasite. Measurement of oocyst viability, as indicated by an oocyst's ability to excyst, is useful because over time oocysts lose the ability to excyst and become noninfective. Thus, correlating the number of viable oocysts in drinking water with incidence and risk for disease should be more reliable than using the total number of oocysts. We have developed a quantitative assay capable of detecting low numbers of excystable, sporozoite-releasing C. parvum oocysts in turbid water samples. Monoclonal (CP7) and polyclonal antibodies have been developed against a sporozoite antigen released only during excystation or when the oocyst is mechanically disrupted. CP7 is specific for C. parvum and does not react with C. baileyi, C. muris, C. serpentis, Giardia spp., Eimeria spp., or E. nieschulzi. In this assay, oocysts in the test sample are first excysted and then centrifuged. The soluble sporozoite antigen is captured by CP7 attached to a magnetic bead. The captured antigen is then detected by ruthenium-labeled polyclonal antibodies via electrochemiluminescence. The CP7 viability assay can detect as few as 50 viable oocysts in a 1-ml assay sample with a turbidity as high as 200 Nephelometric turbidity units. This sensitive, turbidity-tolerant assay for oocyst viability may permit a better assessment of the disease risk associated with the presence of environmental oocysts.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11227892,0022-3395 Evaluation Studies Journal Article11227892National Center for Infectious Diseases, Centers for Disease Control and Prevention, Public Health Service, U.S. Department of Health and Human Services, Atlanta, Georgia 30341, USA.~?Eberhard, M. L. Ortega, Y. R. Hanes, D. E. Nace, E. K. Do, R. Q. Robl, M. G. Won, K. Y. Gavidia, C. Sass, N. L. Mansfield, K. Gozalo, A. Griffiths, J. Gilman, R. Sterling, C. R. Arrowood, M. J.2000ZAttempts to establish experimental Cyclospora cayetanensis infection in laboratory animals577-82 J Parasitol863Animals Animals, Newborn Chickens Coccidiosis/*immunology *Disease Models, Animal Disease Susceptibility Dogs Ducks Eucoccidiida/*pathogenicity Feces/parasitology Female Ferrets Haplorhini Human Male Rabbits Rodentia Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S. SwineJunAttempts were made to develop an animal model for Cyclospora cayetanensis to identify a practical laboratory host for studying human cyclosporiasis. Oocysts collected from stool of infected humans in the United States, Haiti, Guatemala, Peru, and Nepal were held in potassium dichromate solution to allow development of sporozoites. The following animal types were inoculated: 9 strains of mice, including adult and neonatal immunocompetent and immune-deficient inbred and outbred strains, rats, sandrats, chickens, ducks, rabbits, jirds, hamsters, ferrets, pigs, dogs, owl monkeys, rhesus monkeys, and cynomolgus monkeys. Most animals were inoculated by gavage, although some of the primates were fed oocysts on food items. The animals were examined for signs of infection, particularly diarrhea, and stool samples were examined for 4-6 wk after inoculation. None of the animals developed patent infections or signs of infection. We conclude that none of the animals tested is susceptible to infection with C. cayetanensis.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=108642570022-3395 Journal Article10864257Division of Parasitic Diseases, Centers for Disease Control and Prevention, U.S. Department of Health and Human Services, Atlanta, Georgia 30341, USA.~?hKhramtsov, N. V. Chung, P. A. Dykstra, C. C. Griffiths, J. K. Morgan, U. M. Arrowood, M. J. Upton, S. J.2000UPresence of double-stranded RNAs in human and calf isolates of Cryptosporidium parvum275-82 J Parasitol862Animals Base Sequence Blotting, Northern Cattle Cattle Diseases/*parasitology Cryptosporidiosis/*parasitology Cryptosporidium parvum/classification/*genetics/isolation & purification Feces/parasitology Genotype Human Molecular Sequence Data Phylogeny Polymorphism, Restriction Fragment Length RNA, Double-Stranded/*chemistry/isolation & purification RNA, Protozoan/*chemistry/isolation & purification Reverse Transcriptase Polymerase Chain Reaction Sequence Alignment Support, U.S. Gov't, P.H.S.AprWe examined the occurrence of 2 virus-like double-stranded (ds)RNAs in human and calf isolates of Cryptosporidium parvum senso latu and other microorganisms, including 7 other members of the genus. A total of 32 isolates of C. parium, 16 from humans (5 from acquired immune deficiency syndrome patients) and 16 from calves, were analyzed. Ethidium bromide staining, or Northern blot analysis, or reverse transcription/polymerase chain reaction, or all 3 methods, revealed that both genotype 1 and genotype 2 isolates of C. parvum possessed these dsRNAs. No other Cryptosporidium spp. or other organisms examined possessed these dsRNAs. Comparison analysis of partial cDNA sequences of dsRNAs from human and calf isolates revealed a high degree of similarity (>92% and >93% identical nucleotides for large and small dsRNAs, respectively). Slight, consistent differences in nucleotide sequences could be seen at select sites and were associated with an isolate being either genotype 1 or 2. Because of the widespread distribution of the dsRNAs, the similarity of these molecules between isolates, and high host specificity, these nucleic acids may prove to represent species-specific molecular markers for C. parvum. Evidence also suggests that the dsRNA can be utilized for molecular genotyping of C. parvum.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=107805450022-3395 Journal Article10780545CDivision of Biology, Kansas State University, Manhattan 66506, USA.~?3Sulaiman, I. M. Lal, A. A. Arrowood, M. J. Xiao, L.1999]Biallelic polymorphism in the intron region of beta-tubulin gene of Cryptosporidium parasites154-7 J Parasitol851]Alleles Animals Base Sequence Cattle Cryptosporidium/*genetics Cryptosporidium parvum/*genetics DNA Primers/chemistry DNA, Protozoan/chemistry Genotype Human Introns/*genetics Molecular Sequence Data Polymerase Chain Reaction *Polymorphism (Genetics) Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S. Tubulin/*genetics Variation (Genetics)FebXNucleotide sequencing of polymerase chain reaction amplified intron region of the Cryptosporidium parvum beta-tubulin gene in 26 human and 15 animal isolates revealed distinct genetic polymorphism between the human and bovine genotypes. The separation of 2 genotypes of C. parvum is in agreement with our previous genotyping data based on the thrombospondin-related adhesion protein (TRAP-C2) gene, indicating these genotype characteristics are linked at 2 genetic loci. Characterization of Cryptosporidium muris and Cryptosporidium serpentis has further shown that non-parvum Cryptosporidium parasites have beta-tubulin intron sequences identical to bovine genotype of C. parvum. Thus, results of this study confirm the lineage of 2 genotypes of C. parvum at 2 genetic loci and suggest a need for extensive characterization of various Cryptosporidium spp.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=102073870022-3395 Journal Article10207387mDivision of Parasitic Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 30341-3717, USA.~? 'Arrowood, M. J. Hurd, M. R. Mead, J. R.1995nA new method for evaluating experimental cryptosporidial parasite loads using immunofluorescent flow cytometry404-9 J Parasitol813Animals Cryptosporidiosis/*parasitology Cryptosporidium parvum/*growth & development Feces/*parasitology Flow Cytometry Fluorescent Antibody Technique Mice Mice, SCID Parasite Egg Count/methods Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.JunA flow cytometric method for the quantification of Cryptosporidium parvum oocysts in stool specimens was developed to replace conventional microscopic immunofluorescent assays. Fecal pellets were collected from control (uninfected) severe combined immune-deficient mice, suspended in 2.5% potassium dichromate at a ratio of 400 microliter per pellet, and homogenized by vortexing. Purified oocytes were added to the samples (10(5), 10(4), 10(3), and 10(2)/ml). Aliquots (200 microliters) of the vortexed samples were centrifuged over microscale discontinuous sucrose gradients. The oocyst-containing fractions were collected, washed, and incubated with an oocyst-specific monoclonal antibody (labeled with fluorescein isothiocyanate) for 30 min at 37 degrees C. Sample volumes were adjusted to 600 microliters with phosphate-buffered saline and assayed by using logical gating of forward/side scatter and fluorescence signal on a flow cytometer. Seeded samples showed a linear correlation with the number of oocysts recovered from the gradients. Analyses of stool samples from chronically infected mice demonstrated that the flow cytometry method was approximately 10 times more sensitive than conventional immunofluorescent assays.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=77761250022-3395 Journal Article7776125hParasitic Diseases Branch, Centers for Disease Control and Prevention, Atlanta, Georgia 30341-3724, USA.~? -Arrowood, M. J. Sterling, C. R. Healey, M. C.1991jImmunofluorescent microscopical visualization of trails left by gliding Cryptosporidium parvum sporozoites315-7 J Parasitol772Animals Antibodies, Monoclonal/immunology Antigens, Protozoan/*analysis Antigens, Surface/analysis Cryptosporidium/immunology/*physiology Fluorescent Antibody Technique Movement Support, Non-U.S. Gov'tApruCryptosporidium parvum sporozoites that exhibited gliding motility in vitro were examined by immunofluorescence with anticryptosporidial monoclonal antibodies (Mabs) for surface antigen deposition on poly-L-lysine-coated glass microscope slides. The Mabs that revealed trails are specific for an immunodominant 23-kDa antigen previously localized to the sporozoite surface.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20108650022-3395 Journal Article2010865FDepartment of Animal Science, Utah State University, Logan 84322-5600.~? :Mead, J. R. Arrowood, M. J. Current, W. L. Sterling, C. R.1988[Field inversion gel electrophoretic separation of Cryptosporidium spp. chromosome-sized DNA366-9 J Parasitol743Animals Chromosomes/*analysis Coccidia/*genetics Cryptosporidium/classification/*genetics DNA/*analysis Electrophoresis, Agar Gel/methods Fluorescent Antibody Technique Support, Non-U.S. Gov'tJuntChromosomal DNA from 5 isolates of Cryptosporidium parvum and 1 of C. baileyi were compared by field-inversion gel electrophoresis (FIGE). FIGE analyses of parasite DNA prepared from purified sporozoites versus intact oocysts showed no observable differences. Chromosomal DNA migration patterns of the 5 C. parvum isolates were indistinguishable, whereas similar but distinct differences were evident between C. baileyi and the isolates of C. parvum. Oocyst-reactive monoclonal antibodies differentiated oocysts of C. parvum from those of C. baileyi but were unable to distinguish oocysts of 1 isolate of C. parvum from another.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=32887380022-3395 Journal Article3288738@Department of Microbiology, University of Arizona, Tucson 85721.~? +Mead, J. R. Arrowood, M. J. Sterling, C. R.1988`Antigens of Cryptosporidium sporozoites recognized by immune sera of infected animals and humans135-43 J Parasitol741Animals Antibodies, Monoclonal Antibodies, Protozoan/*analysis/immunology Antigens, Protozoan/*immunology Antigens, Surface/immunology Cattle Cattle Diseases/immunology Coccidia/*immunology Cryptosporidiosis/*immunology Cryptosporidium/*immunology Electrophoresis, Polyacrylamide Gel Fluorescent Antibody Technique Horse Diseases/immunology Horses Human Immunoassay Peptides/immunology Support, Non-U.S. Gov'tFebThe humoral response of humans, calves, and horses to Cryptosporidium sporozoite antigens was evaluated using a western blot technique. Sera from calves, humans, and horses were obtained at various times following the detection of infection. Sera were reacted with detergent-solubilized, sodium dodecyl sulfate polyacrylamide gel-electrophoresed (SDS-PAGE) sporozoite antigens. The number of antigens recognized by immune sera from humans and animals increased with time postinfection. A 20-kDa antigen appears to be a major sporozoite surface determinant labeled via membrane protein biotinylation and recognized by mouse monoclonal antibodies using indirect immunofluorescence and western blotting. Detectable recognition of the 20-kDa band occurred in 3-wk postinfection (PI) sera from all species tested. Reactivity to the 20-kDa band diminished significantly in sera 5 mo PI or longer from infected humans with no known recurrence of cryptosporidial diarrhea. In contrast, 12-mo PI sera from an individual constantly exposed to oocysts under working conditions was as strongly reactive as the 3-wk convalescent sera. Serum reactivity to the 20-kDa antigen appears to be a good indicator of exposure to Cryptosporidium.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=32820490022-3395 Journal Article3282049@Department of Microbiology, University of Arizona, Tucson 85721.~? FMadore, M. S. Rose, J. B. Gerba, C. P. Arrowood, M. J. Sterling, C. R.1987UOccurrence of Cryptosporidium oocysts in sewage effluents and selected surface waters702-5 J Parasitol734Animals Arizona Coccidia/*isolation & purification Cryptosporidium/*isolation & purification Fresh Water *Sewage Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. *WaterAugAn existing method for the detection of Cryptosporidium oocysts in water was modified to investigate oocyst prevalence in large volumes of water. Surface waters and sewage effluents were filtered, eluted from the filter, and concentrated using centrifugation. The resultant pellet was then homogenized, sonicated, and placed on a sucrose gradient to separate oocysts from the sediment. The uppermost gradient layer was then examined by immunofluorescence using a labeled monoclonal antibody. Using this technique, average numbers of oocysts detected in raw and treated sewage were 5.18 X 10(3) and 1.30 X 10(3)/L, respectively. Filtered sewage effluents had significantly lower numbers of oocysts (10.0/L). These data show that sand filtration may reduce the concentrations of this parasite in waste waters. Highly variable oocyst numbers were encountered in surface waters. Since Cryptosporidium oocysts are frequently present in environmental waters, they could be responsible for waterborne outbreaks of disease.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=36254240022-3395 Journal Article3625424~?Arrowood, M. J. Sterling, C. R.1987pIsolation of Cryptosporidium oocysts and sporozoites using discontinuous sucrose and isopycnic Percoll gradients314-9 J Parasitol732Animals Cattle Centrifugation, Density Gradient Centrifugation, Isopycnic Coccidia/*isolation & purification Cryptosporidium/cytology/*isolation & purification Feces/*parasitology Support, U.S. Gov't, Non-P.H.S.AprTechniques for the large-scale isolation of Cryptosporidium oocysts and sporozoites, obtained from the feces of experimentally infected Holstein calves, were developed employing discontinuous sucrose gradients and isopycnic Percoll gradients. The oocyst recovery method utilized 2 sequential discontinuous sucrose gradients followed by 1 Percoll gradient. Recovered oocysts were essentially free of debris and bacteria and represented 34% of the original oocyst suspension. Sporozoites were recovered from excystation mixtures on a single Percoll gradient. Sixty-three percent of the original sporozoites were recovered with 2.2% contamination by intact oocysts and virtually no oocyst walls.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=35856260022-3395 Journal Article3585626 ~?-Rulofson, F. C. Atwill, E. R. Holmberg, C. A.2001Fecal shedding of Giardia duodenalis, Cryptosporidium parvum, Salmonella organisms, and Escherichia coli O157:H7 from llamas in California637-42 Am J Vet Res624[Age Factors Animal Husbandry Animals California/epidemiology Camelids, New World/*microbiology/*parasitology Cryptosporidiosis/epidemiology/microbiology/veterinary Cryptosporidium parvum/*isolation & purification Escherichia coli Infections/epidemiology/microbiology/*veterinary Escherichia coli O157/*isolation & purification Feces/microbiology/parasitology/virology Female Giardia/*isolation & purification Giardiasis/epidemiology/microbiology/veterinary Male Prevalence Salmonella/*isolation & purification Salmonella Infections, Animal/epidemiology/*microbiology/parasitology Support, Non-U.S. Gov'tAprOBJECTIVE: To evaluate fecal shedding of Giardia duodenalis, Cryptosporidium parvum, Salmonella organisms, and Escherichia coli O157:H7 from llamas in California with respect to host factors and management practices. ANIMALS: 354 llamas from 33 facilities. PROCEDURE: Fecal specimens were collected and examined for G. duodenalis and C. parvum by means of immunofluorescent microscopy. Salmonella organisms were cultured by placing feces into selenite enrichment broth followed by selective media. Escherichia coli O157:H7 was cultured by use of modified tryptocase soy broth followed by sorbitol MacConkey agar, with suspect colonies confirmed by means of immunofluorescent microscopy. RESULTS: 12 of 354 fecal specimens (3.4%) had G. duodenalis cysts. Younger llamas (crias) were more likely to be shedding cysts, compared with older llamas. Farm-level factors that increased the risk of shedding were large numbers of yearlings on the property (> 10), smaller pen sizes, large numbers of crias born during the previous year (> 10), and large pen or pasture populations (> 20). None of the 354 fecal specimens had C. parvum oocysts. Seventy-six (from 7 facilities) and 192 (from 22 facilities) llamas were tested for Salmonella organisms and E. coli O157:H7, respectively. All fecal specimens had negative results for these bacteria. CONCLUSIONS AND CLINICAL RELEVANCE: Shedding of G. duodenalis was primarily limited to crias 1 to 4 months old. Llamas from properties with large numbers of crias born in the previous year, resulting in large numbers of yearlings in the current year, were at greater risk of infection. In addition, housing llamas in smaller pens or pastures and managing llamas and crias in large groups also increased the risk of G. duodenalis shedding.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=113274780002-9645 Journal Article11327478BUniversity of California Cooperative Extension, Sonora 95370, USA. d~?DHoar, B. R. Atwill, E. R. Elmi, C. Utterback, W. W. Edmondson, A. J.1999Comparison of fecal samples collected per rectum and off the ground for estimation of environmental contamination attributable to beef cattle1352-6 Am J Vet Res6011zAnimals Bacterial Infections/transmission/veterinary Campylobacter/*isolation & purification Campylobacter jejuni/isolation & purification Cattle Cattle Diseases/microbiology Cryptosporidium parvum/*isolation & purification Feces/*microbiology Giardia/*isolation & purification Human Rectum/microbiology Reproducibility of Results Support, Non-U.S. Gov't Water Supply/*standardsNovOBJECTIVES: To determine whether sampling feces off the ground replicates prevalence estimates for specific pathogens obtained from fecal samples collected per rectum of adult cows, and to determine characteristics of feces on the ground (fecal pats) that are associated with subsequent identification of Campylobacter spp, Cryptosporidium parvum, and Giardia duodenalis. ANIMALS: A random sample of adult beef cattle from 25 herds located throughout California. PROCEDURE: 1,115 rectal and ground fecal samples were obtained. Samples were submitted for culture of Campylobacter spp and examined, using a direct fluorescent antibody assay, to detect C parvum oocysts and G duodenalis cysts. Characteristics of fecal pats, such as volume and consistency, were recorded. RESULTS: Prevalence of Campylobacter spp was 5.0% (20/401) for rectal fecal samples, which was significantly greater than prevalence determined for ground fecal samples (2/402; 0.5%). Most isolates were C jejuni subsp jejuni. Prevalence of C parvum was higher in rectal fecal samples (6/557; 1.1%) than in ground fecal samples (1/558; 0.2%), but this difference was not significant. Prevalence of G duodenalis did not differ for rectal (36/557; 6.5%) versus ground (26/558; 4.7%) fecal samples. CONCLUSIONS AND CLINICAL RELEVANCE: Evaluation of ground fecal samples may not accurately indicate the prevalence of Campylobacter spp or C parvum in cattle but may reflect prevalence of G duodenalis. Differences in prevalence estimates between the 2 methods suggest inactivation of pathogens in feces after cattle have defecated. Prevalence estimates generated by evaluation of ground fecal samples, however, may more accurately estimate environmental pathogen burden.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=105668070002-9645 Journal Article10566807|Department of Population Health and Reproduction, School of Veterinary Medicine, University of California, Davis 95616, USA. ~?Atwill, E. R. Johnson, E. Klingborg, D. J. Veserat, G. M. Markegard, G. Jensen, W. A. Pratt, D. W. Delmas, R. E. George, H. A. Forero, L. C. Philips, R. L. Barry, S. J. McDougald, N. K. Gildersleeve, R. R. Frost, W. E.1999pAge, geographic, and temporal distribution of fecal shedding of Cryptosporidium parvum oocysts in cow-calf herds420-5 Am J Vet Res604Age Factors Animals California/epidemiology Cattle Cattle Diseases/epidemiology/*parasitology Cryptosporidiosis/epidemiology/*veterinary Cryptosporidium parvum/*isolation & purification Feces/*parasitology Geography Parasite Egg Count Support, Non-U.S. Gov't Time FactorsApr(OBJECTIVE: To evaluate fecal shedding of Cryptosporidium parvum from California cow-calf herds with respect to age, geographic region, temporal effects, and association with watery feces. ANIMALS: Cows and calves from 38 beef cow-calf operations. PROCEDURE: Fecal specimens were collected and examined for C parvum oocysts, using immunofluorescent microscopy. Associations between age, geographic region, month of collection, watery feces, and likelihood of shedding C parvum were evaluated. RESULTS: 3.9% of cattle were shedding C parvum oocysts. Prevalence of shedding among calves ranged from 0 to 13%, and was 0.6% among cattle > or = 12 months old. The odds of shedding C parvum among 2-month-old calves were 41 times greater than among cattle > 4 months old. The odds of shedding C parvum among cattle tested in May were 8.7 times greater than among cattle tested during June, July, or August. The odds of infected individuals having watery feces were 3 to 4 times greater than for noninfected individuals, but the etiologic fraction was only 8 to 9%. CONCLUSIONS AND CLINICAL RELEVANCE: Substantial fecal shedding of C parvum by cow-calf herds was limited to calves 1 to 4 months old, with low prevalence detected in older animals. Risk of contamination of watersheds with C parvum was limited to those periods when young calves were in the herd. Although the odds of having watery feces were greater for animals infected with C parvum than for noninfected animals, the low etiologic fraction suggests that most calves with watery feces were not infected with C parvum.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=102116830002-9645 Journal Article10211683Veterinary Medicine Teaching and Research Center, School of Veterinary Medicine, University of California-Davis, Tulare 93274, USA. ~?HAtwill, E. R. Harp, J. A. Jones, T. Jardon, P. W. Checel, S. Zylstra, M.1998|Evaluation of periparturient dairy cows and contact surfaces as a reservoir of Cryptosporidium parvum for calfhood infection1116-21 Am J Vet Res599xAnimals Animals, Newborn/*parasitology California/epidemiology Cattle Cattle Diseases/epidemiology/*parasitology Cryptosporidiosis/epidemiology/parasitology/*veterinary *Cryptosporidium parvum Disease Reservoirs/*veterinary Feces/parasitology Fluorescent Antibody Technique, Direct/veterinary Parasite Egg Count/veterinary Prevalence Support, Non-U.S. Gov't Surface PropertiesSepOBJECTIVE: To determine whether periparturient cows or contact surfaces to which newborn calves are exposed are reservoirs of Cryptosporidium parvum oocysts. ANIMALS: Periparturient cows and their calves. PROCEDURE: Using direct fluorescent antibody (DFA) and acid-fast (AF) assays, fecal samples taken before and after calving from periparturient cows were tested for C parvum oocysts. Fecal samples from calves were collected every other day from age 7 to 21 days and were tested by use of the AF assay. Topsoil from close-up and maternity pens and scrapings from wooden walls and floors of calf hutches were tested for C parvum oocysts by use of DFA assay. RESULTS: None of the 384 fecal samples obtained 1 to 21 days before or after calving or on the day of calving from 154 periparturient cows contained detectable C parvum oocysts. Despite this lack of detectable periparturient shedding, the period prevalence of calfhood infection was 92% (123/134) from age 7 to 21 days. Soil samples from the close-up and maternity pens where newborn calves spend the first 12 hours of life also were negative for C parvum oocysts. Wood scrap ings from the outer 2 mm of the walls and floors of empty and cleaned calf hutches that were ready to receive calves were C parvum oocyst-positive. CONCLUSIONS: Conditional on sensitivity of DFA, periparturient cows did not appear to shed detectable C parvum oocysts. In contrast, the floors and walls of wooden calf hutches contained detectable C parvum oocysts on the surface.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=97363870002-9645 Journal Article9736387Veterinary Medicine Teaching and Research Center, School of Veterinary Medicine, University of California, Davis, Tulare, CA 93274, USA.~?UHarp, J. A. Jardon, P. Atwill, E. R. Zylstra, M. Checel, S. Goff, J. P. De Simone, C.1996tField testing of prophylactic measures against Cryptosporidium parvum infection in calves in a California dairy herd1586-8 Am J Vet Res5711Animals California Cattle Cattle Diseases/parasitology/*prevention & control Cryptosporidiosis/prevention & control/*veterinary *Cryptosporidium parvum Diarrhea/parasitology/prevention & control/veterinary Protozoan Vaccines/*therapeutic use Random Allocation Support, Non-U.S. Gov'tNovOBJECTIVE: To test the ability of oral vaccination or probiotic treatment with lactic acid-producing bacteria to protect calves from Cryptosporidium parvum infection under field conditions. ANIMALS: 134 Holstein calves born on a dairy farm where cryptosporidiosis was endemic. PROCEDURE: Calves were randomly assigned to 1 of 3 treatment groups at birth. Calves in the vaccine group received an oral dose of C parvum vaccine within several hours of birth. Calves in the bacteria group received an oral dose of lactic acid-producing bacteria daily for the first 10 days after birth. Control calves were not treated. All calves were monitored for diarrhea and fecal shedding of C parvum oocysts for 3 weeks. RESULTS: There were no significant differences in the incidence of diarrhea and oocyst shedding among the 3 groups. CONCLUSIONS: Neither vaccination nor probiotic treatment was effective in preventing C parvum infection in calves under field conditions. High numbers of C parvum in the environment may have overwhelmed any potential benefits of these regimens. Further work is necessary to develop effective prophylaxis against C parvum under field conditions.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=89154340002-9645 Journal Article8915434^USDA, Agricultural Research Service, National Animal Disease Center, Ames, IA 50010-0070, USA. ~?+Atwill, E. R. Johnson, E. M. Pereira, M. G.1999Association of herd composition, stocking rate, and duration of calving season with fecal shedding of Cryptosporidium parvum oocysts in beef herds1833-8J Am Vet Med Assoc21512fAge Factors Animal Husbandry Animals California Cattle Cattle Diseases/*parasitology/transmission Cross-Sectional Studies Cryptosporidiosis/parasitology/transmission/*veterinary Cryptosporidium parvum/*physiology Feces/parasitology Female Fluorescent Antibody Technique, Direct/veterinary Logistic Models Multivariate Analysis Seasons Support, Non-U.S. Gov'tDec 15OBJECTIVE: To evaluate the association of herd demographics, parturition variables, stocking rate, and rotational grazing practices with the probability of fecal shedding of Cryptosporidium parvum from beef cow-calf herds in California. DESIGN: Cross-sectional study. SAMPLE POPULATION: 38 beef cow-calf operations. PROCEDURE: Fecal specimens were collected and examined for C parvum oocysts, using immunofluorescent microscopy. Association between various demographic and management factors and the probability of shedding C parvum were statistically evaluated. RESULTS: Adjusted for age and month of collection of a fecal sample, cattle from herds with a high number of young calves (< or = 2 months old) on the day of sample collection, a high stocking rate (No. of cattle/acre/mo), or a longer calving season were more likely to shed C parvum oocysts, compared with cattle from herds with fewer young calves, a lower stocking rate, or a shorter calving season. Cattle from herds with a higher number of older calves (> 2 months old) on the day of sample collection were less likely to shed C parvum oocysts, compared with cattle from herds with fewer older calves. Using our multivariate model, rotational grazing systems or season of onset of calving were not associated with shedding status for C parvum oocysts. CONCLUSIONS AND CLINICAL RELEVANCE: Reproductive management that would result in a shorter calving season and use of a lower stocking rate for cattle may be associated with reduced risk of C parvum shedding. Intensive rotational grazing systems and time of year for onset of calving season apparently have little effect on reducing prevalence of oocyst shedding.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=106132180003-1488 Journal Article10613218Veterinary Medicine Teaching and Research Center, School of Veterinary Medicine, University of California-Davis, Tulare 93274, USA.~??Hou, L. Li, X. Dunbar, L. Moeller, R. Palermo, B. Atwill, E. R.2004qNeonatal-mouse infectivity of intact Cryptosporidium parvum oocysts isolated after optimized in vitro excystation642-6Appl Environ Microbiol701JanWe reexamined the finding of Neumann et al. that intact Cryptosporidium parvum oocysts obtained after in vitro excystation were infectious for neonatal CD-1 mice. We used both established excystation protocols and our own protocol that maximized excystation. Although intact oocysts isolated after any of three protocols were infectious for neonatal CD-1 mice, the infectivity of intact oocysts isolated with our optimized excystation protocol was significantly lower than the infectivity of intact oocysts isolated after established protocols or from fresh oocysts. Excystation should not be considered a valid measure of C. parvum viability, given that it is biologically implausible for oocysts to be nonviable and yet infectious.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=147117040099-2240 Journal Article14711704Veterinary Medicine Teaching and Research Center, School of Veterinary Medicine, University of California-Davis, Tulare, California 93274, USA. \~?WAtwill, E. R. Hoar, B. das Gracas Cabral Pereira, M. Tate, K. W. Rulofson, F. Nader, G.2003Improved quantitative estimates of low environmental loading and sporadic periparturient shedding of Cryptosporidium parvum in adult beef cattle4604-10Appl Environ Microbiol698Animals Cattle/*parasitology Cryptosporidium parvum/*isolation & purification Feces/*parasitology Oocysts/isolation & purification Support, Non-U.S. Gov'tAugPOur primary goal was to generate an accurate estimate of the daily environmental loading rate of Cryptosporidium parvum oocysts for adult beef cattle, using immunomagnetic separation coupled with direct immunofluorescence microscopy for a highly sensitive diagnostic assay. An additional goal was to measure the prevalence and intensity of fecal shedding of C. parvum oocysts in pre- and postparturient cows as an indicator of their potential to infect young calves. This diagnostic method could detect with a > or = 90% probability oocyst concentrations as low as 3.2 oocysts g of feces(-1), with a 54% probability of detecting just one oocyst g of feces(-1). Using this diagnostic method, the overall apparent prevalence of adult beef cattle testing positive for C. parvum was 7.1% (17 of 240), with 8.3 and 5.8% of cattle shedding oocysts during the pre- and postcalving periods, respectively. The mean intensity of oocyst shedding for test-positive cattle was 3.38 oocysts g of feces(-1). The estimated environmental loading rate of C. parvum ranged from 3,900 to 9,200 oocysts cow(-1) day(-1), which is substantially less than a previous estimate of 1.7 x 10(5) oocysts cow(-1) day(-1) (range of 7.7 x 10(4) to 2.3 x 10(5) oocysts cow(-1) day(-1)) (B. Hoar, E. R. Atwill, and T. B. Farver, Quant. Microbiol. 2:21-36, 2000). Use of this highly sensitive assay functioned to detect a greater proportion of low-intensity shedders in our population of cattle, which reduced the estimated mean intensity of shedding and thereby reduced the associated environmental loading rate compared to those of previous studies.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=129022480099-2240 Journal Article12902248Veterinary Medicine Teaching and Research Center, School of Veterinary Medicine, University of California-Davis, Tulare, California 93274, USA. ratwill@vmtrc.ucdavis.edu4~?IAtwill, E. R. Hou, L. Karle, B. M. Harter, T. Tate, K. W. Dahlgren, R. A.2002oTransport of Cryptosporidium parvum oocysts through vegetated buffer strips and estimated filtration efficiency5517-27Appl Environ Microbiol6811nAnimals Buffers Cryptosporidium parvum/*physiology Filtration Oocysts/*physiology Rain Support, Non-U.S. Gov'tNovVegetated buffer strips were evaluated for their ability to remove waterborne Cryptosporidium parvum from surface and shallow subsurface flow during simulated rainfall rates of 15 or 40 mm/h for 4 h. Log(10) reductions for spiked C. parvum oocysts ranged from 1.0 to 3.1 per m of vegetated buffer, with buffers set at 5 to 20% slope, 85 to 99% fescue cover, soil textures of either silty clay (19:47:34 sand-silt-clay), loam (45:37:18), or sandy loam (70:25:5), and bulk densities of between 0.6 to 1.7 g/cm(3). Vegetated buffers constructed with sandy loam or higher soil bulk densities were less effective at removing waterborne C. parvum (1- to 2-log(10) reduction/m) compared to buffers constructed with silty clay or loam or at lower bulk densities (2- to 3-log(10) reduction/m). The effect of slope on filtration efficiency was conditional on soil texture and soil bulk density. Based on these results, a vegetated buffer strip comprised of similar soils at a slope of or=3 m should function to remove >or=99.9% of C. parvum oocysts from agricultural runoff generated during events involving mild to moderate precipitation.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=124067450099-2240 Journal Article12406745Veterinary Medicine Teaching and Research Center, School of Veterinary Medicine, University of California-Davis, Tulare, California 93274, USA. ratwill@vmtrc.ucdavis.edu~?uAtwill, E. R. Camargo, S. M. Phillips, R. Alonso, L. H. Tate, K. W. Jensen, W. A. Bennet, J. Little, S. Salmon, T. P.2001wQuantitative shedding of two genotypes of Cryptosporidium parvum in California ground squirrels (Spermophilus beecheyi)2840-3Appl Environ Microbiol676<Animals California Cryptosporidiosis/transmission/*veterinary Cryptosporidium parvum/*classification/genetics/isolation & purification *Environment Feces/*parasitology Female Genotype Male Polymerase Chain Reaction Polymorphism, Restriction Fragment Length Sciuridae/*parasitology Sex Factors Support, Non-U.S. Gov'tJunSixteen percent of California ground squirrels (Spermophilus beecheyi) were found to be shedding an average of 53,875 Cryptosporidium parvum oocysts/g of feces. Male squirrels had a higher prevalence and higher intensity of shedding than did female squirrels. The majority of C. parvum isolates matched a bovine-murine genotype, with a few isolates resembling a porcine genotype. Higher intensities of shedding by males may enhance dissemination and genotypic mixing of this protozoa given males' proclivity to disperse to nonnatal colonies.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=113752040099-2240 Journal Article11375204Veterinary Medicine Teaching and Research Center, School of Veterinary Medicine, University of California-Davis, Tulare, California 93274, USA. ratwill@vmtrc.ucdavis.edu~?&Pereira, M. D. Atwill, E. R. Jones, T.1999Comparison of sensitivity of immunofluorescent microscopy to that of a combination of immunofluorescent microscopy and immunomagnetic separation for detection of Cryptosporidium parvum oocysts in adult bovine feces3236-9Appl Environ Microbiol657]Animals Cattle Cattle Diseases/*parasitology Comparative Study Cryptosporidiosis/parasitology/*veterinary Cryptosporidium parvum/*isolation & purification Feces/*parasitology Fluorescent Antibody Technique, Direct *Immunomagnetic Separation Microscopy, Fluorescence Sensitivity and Specificity Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S.JulA direct immunofluorescence assay (DFA) (Merifluor; Meridian Diagnostics, Inc., Cincinnati, Ohio) was compared to an immunomagnetic separation (IMS) assay (Dynabeads; Dynal, Inc., Lake Success, N.Y.) coupled with immunofluorescent microscopy (Waterborne, Inc., New Orleans, La.) for their ability to detect low concentrations of Cryptosporidium parvum oocysts in adult bovine fecal material. IMS-DFA resulted in a 2-log-unit increase in sensitivity (10 oocysts/g) compared to DFA alone (1,000 oocysts/g). The higher sensitivity obtained with IMS-DFA resulted from testing 2 g of fecal material instead of the 13 to 19 mg of fecal material tested in the DFA; the increased sensitivity was not attributable to a higher percent recovery.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=103887280099-2240 Journal Article10388728Veterinary Medicine Teaching and Research Center, School of Veterinary Medicine, University of California, Davis, Tulare, California 93274, USA.~?@Pereira, M. das G. Atwill, E. R. Crawford, M. R. Lefebvre, R. B.1998MDNA sequence similarity between California isolates of Cryptosporidium parvum1584-6Appl Environ Microbiol644Animals Animals, Domestic/parasitology Animals, Wild/parasitology Base Sequence California Cattle/parasitology Comparative Study Cryptosporidium parvum/classification/*genetics/*isolation & purification DNA Primers/genetics DNA, Protozoan/*genetics/isolation & purification Goats/parasitology Horses/parasitology Human Molecular Sequence Data Polymerase Chain Reaction Sequence Homology, Nucleic Acid Sewage/parasitology Species Specificity Support, Non-U.S. Gov't Swine/parasitologyAprWe evaluated whether nucleic acid amplification with primers specific for Cryptosporidium parvum followed by automated DNA sequence analysis of the PCR amplicons could differentiate between California isolates of C. parvum obtained from livestock, humans, and feral pigs. Almost complete sequence identity existed among the livestock isolates and between the livestock and human isolates. DNA sequences from feral pig isolates differed from those from livestock and humans by 1.0 to 1.2%. The reference sequence obtained by Laxer et al. (M. A. Laxer, B. K. Timblin, and R. J. Patel, Am. J. Trop. Med. Hyg. 45:688-694, 1991.) differed from California isolates of C. parvum by 1.8 to 3.2%. These data suggest that DNA sequence analysis of the amplicon of Laxer et al. does not allow for differentiation between various strains of C. parvum or that our collection of isolates obtained from various hosts from across California was limited to one strain of C. parvum.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=95461950099-2240 Journal Article9546195Veterinary Medicine Teaching and Research Center, School of Veterinary Medicine, University of California, Davis, Tulare 93274, USA. D~?VAtwill, E. R. Sweitzer, R. A. Pereira, M. G. Gardner, I. A. Van Vuren, D. Boyce, W. M.1997Prevalence of and associated risk factors for shedding Cryptosporidium parvum oocysts and Giardia cysts within feral pig populations in California3946-9Appl Environ Microbiol6310IAnimals Animals, Wild/*parasitology California Cattle Cross-Sectional Studies Cryptosporidium parvum/growth & development/*isolation & purification Disease Reservoirs Giardia/growth & development/*isolation & purification Risk Factors Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Swine/*parasitology Water/parasitologyOctPopulations of feral pigs (Sus scrofa) may serve as an environmental reservoir of Cryptosporidium parvum oocysts and Giardia sp. cysts for source water. We conducted a cross-sectional study to determine the prevalence of and associated demographic and environmental risk factors for the shedding of C. parvum oocysts and Giardia sp. cysts. Feral pigs were either live-trapped or dispatched from 10 populations located along the coastal mountains of western California, and fecal samples were obtained for immunofluorescence detection of C. parvum oocysts and Giardia sp. cysts. We found that 12 (5.4%) and 17 (7.6%) of 221 feral pigs were shedding C. parvum oocysts and Giardia sp. cysts, respectively. The pig's sex and body condition and the presence of cattle were not associated with the probability of the shedding of C. parvum oocysts. However, younger pigs (< or = 8 months) and pigs from high-density populations (> 2.0 feral pigs/km2) were significantly more likely to shed oocysts compared to older pigs (> 8 months) and pigs from low-density populations (< or = 1.9 feral pigs/km2). In contrast, none of these demographic and environmental variables were associated with the probability of the shedding of Giardia sp. cysts among feral pigs. These results suggest that given the propensity for feral pigs to focus their activity in riparian areas, feral pigs may serve as a source of protozoal contamination for surface water.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=93275600099-2240 Journal Article9327560Veterinary Medicine Teaching and Research Center, School of Veterinary Medicine, University of California, Davis 93274, USA. ratwill@vmtrc.ucdavis.edu z~?iQuilez, J. Vergara-Castiblanco, C. A. Ares-Mazas, M. E. Sanchez-Acedo, C. del Cacho, E. Freire-Santos, F.2002sSerum antibody response and Cryptosporidium parvum oocyst antigens recognized by sera from naturally infected sheep187-97 Vet Parasitol1043Age Factors Animals Animals, Newborn Antibodies, Protozoan/*blood/immunology Antigens, Protozoan/*immunology Cryptosporidiosis/immunology/parasitology/*veterinary Cryptosporidium parvum/*immunology Disease Outbreaks/veterinary Enzyme-Linked Immunosorbent Assay/methods/veterinary Feces/parasitology Female Immune Sera/immunology Immunoblotting/methods/veterinary Immunoglobulins/blood/immunology Male Oocytes Sheep Sheep Diseases/epidemiology/immunology/*parasitology Spain/epidemiology Support, Non-U.S. Gov'tMar 20The response of specific serum immunoglobulins (IgG, IgM and IgA) and the major antigens of Cryptosporidium parvum recognized by these isotypes were investigated by using enzyme-linked immunosorbent assay and immunoblot techniques in lambs and ewes naturally infected throughout an outbreak of cryptosporidiosis. Serum samples were collected from 20 lambs the first day they showed diarrhoea (D1), and Days 11 and 22, in addition to single serum samples from 17 of their dams. Serum anti-C. parvum IgG, IgM and/or IgA antibodies were detected in lambs as early as Day 1. Levels of IgM antibodies remained steady from D1 to D11 and increased at D22, whereas the IgG response decreased from D1 to D11 and subsequently increased. In contrast, IgA antibodies rapidly fell from D1 and all lambs were seronegative at D11 and D22. The highest levels of specific antibodies were detected in sera from ewes. In fact, all ewes were seropositives for IgM and IgA isotypes and most (16/17) showed positive levels of IgG. Four protein fractions (37-39, 42-48, 51-57 and 60-69 kDa) were the most frequently recognized by IgG and IgM from lamb sera. A low molecular weight fraction (12-14 kDa) reacting with IgG and IgA in most lamb sera was scarcely recognized by IgM and three broad bands were frequently recognized by IgA antibodies (23-25, 51-57 and 90-95 kDa). The recognition pattern of 23-25 kDa peptides by IgA from lamb sera clearly increased with the age. Peptides of 42-48, 51-57, 60-69 and 71-78 kDa were most frequently recognized by IgG and IgM from ewe sera. In relation to IgA antibodies from ewe sera, a frequent immunoreactivity was found with proteins in the intervals between 12 and 22 kDa as well as between 32 and 34 kDa and practically all sera reacted with fractions from 42 to 95 kDa.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=118126170304-4017 Journal Article11812617Departamento de Patologia Animal, Parasitologia y Enfermedades Parasitarias, Facultad de Veterinaria, Universidad de Zaragoza, C/Miguel Servet 177, 50013 Zaragoza, Spain. jquilez@posta.unizar.es~?\Castro-Hermida, J. A. Gonzalez-Losada, Y. Freire-Santos, F. Mezo-Menendez, M. Ares-Mazas, E.2001YEvaluation of beta-cyclodextrin against natural infections of cryptosporidiosis in calves85-9 Vet Parasitol1012Animals Animals, Suckling Cattle Cattle Diseases/*drug therapy/parasitology/prevention & control Coccidiostats/pharmacology/therapeutic use Cryptosporidiosis/drug therapy/parasitology/prevention & control/*veterinary Cryptosporidium parvum/*drug effects/growth & development Cyclodextrins/pharmacology/*therapeutic use Diarrhea/drug therapy/parasitology/prevention & control/*veterinary Feces/parasitology Female Male Parasitic Sensitivity Tests/veterinary Support, Non-U.S. Gov'tNov 5The effectiveness of beta-cyclodextrin, excipient used in pharmaceutical industry, in the treatment of natural infection by Cryptosporidium parvum in suckling calves, was evaluated. Administration of the drug at a dose of 500 mg/kg body weight for 3 consecutive days from birth (prophylactically) or following confirmation of the infection (therapeutically) decreased the severity of diarrhoea and shortened the duration of oocyst shedding.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11587837,0304-4017 Evaluation Studies Journal Article11587837Laboratorio de Parasitologia, Departamento de Microbiologia y Parasitologia, Facultad de Farmacia, Universidad de Santiago de Compostela, Avda. de Vigo s/n, 15782 Santiago de Compostela, La Coruna, Spain.F~?hCastro Hermida, J. A. Freire Santos, F. Oteiza Lopez, A. M. Vergara Castiblanco, C. A. Ares-Mazas, M. E.2000ZIn vitro and in vivo efficacy of lasalocid for treatment of experimental cryptosporidiosis265-70 Vet Parasitol904Animals Animals, Newborn Coccidiostats/administration & dosage/standards/*therapeutic use Cryptosporidiosis/drug therapy/*veterinary Cryptosporidium parvum/drug effects/*growth & development Female Fluorescent Dyes/chemistry Indoles/chemistry Intestine, Large/parasitology Intestine, Small/parasitology Lasalocid/administration & dosage/standards/*therapeutic use Mice Microscopy, Fluorescence/veterinary Microscopy, Phase-Contrast/veterinary Parasite Egg Count/veterinary Propidium/chemistry Statistics, Nonparametric Support, Non-U.S. Gov'tJul 4In vitro viability of purified Cryptosporidium parvum oocysts, exposed for 30, 60, 90 and 120min to 0.27mg/ml lasalocid suspension was evaluated by inclusion or exclusion of two fluorogenic vital dyes and an excystation technique. Continuously, preventive and curative efficacies at different doses (9, 6.75, 5.625 and 4.5mg/kg body weight) and regimens of lasalocid against cryptosporidial infection were evaluated on an experimental neonatal mice model. In vitro assays demonstrated a decrease in the oocyst viability related to an increase in exposure time for exposure to the lasalocid suspension. The infection was eradicated when the suspension was administered with a dose of > or = 6.75mg/kg body weight. No apparent toxic effects were observed.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=108568130304-4017 Journal Article10856813Laboratorio de Parasitologia, Departamento de Microbiologia y Parasitologia, Facultad de Farmacia, Universidad de Santiago de Compostela, La Coruna, Spain.~?RVergara-Castiblanco, C. A. Freire-Santos, F. Oteiza-Lopez, A. M. Ares-Mazas, M. E.2000lViability and infectivity of two Cryptosporidium parvum bovine isolates from different geographical location261-7 Vet Parasitol894Animals Antibodies, Protozoan/analysis Cattle Cattle Diseases/*parasitology Colombia Cryptosporidiosis/parasitology/*veterinary Cryptosporidium parvum/*pathogenicity Enzyme-Linked Immunosorbent Assay/veterinary Mice SpainMay 17The viability of two Cryptosporidium parvum bovine isolates from Spain and Colombia was evaluated by in vitro excystation, inclusion/exclusion of two fluorogenic vital dyes (DAPI and PI) and infectivity assay in a suckling murine model. Excystation percentages were similar for both Spain and Colombia isolates (83% and 87%, respectively). The total viability of the Spain isolate, measured by inclusion/exclusion of two fluorogenic vital dyes, was 71% in comparison with that detected for oocysts of the Colombia isolate, 32.3%. The bovine C. parvum oocysts of both isolates were viable and infectious for suckling Swiss CD-1 mice. However, infectivity percentage and the mean intensity of infection were consistently higher in the Spain isolate than those from Colombia isolate. It was not possible to obtain a good correlation between in vitro excystation, inclusion/exclusion of vital dyes and in vivo infectivity for the Colombia isolate, while data obtained with the Spain isolate indicated that there was an apparent strong correlation between excystation efficiency, total viability and the infectivity. Although a comparative analysis of genetic variation among these isolates from different geographical location is necessary, variations observed between the both isolates seemed to be a result of parasite adaptation to environmental stresses such as temperature which appears to have a direct effect on the permeability of the oocysts.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=107998390304-4017 Journal Article10799839Laboratorio de Parasitologia, Departamento de Microbiologia y Parasitologia, Facultad de Farmacia, Universidad de Santiago de Compostela, Spain.b~? OFreire-Santos, F. Oteiza-Lopez, A. M. Vergara-Castiblanco, C. A. Ares-Mazas, E.2000Study of the combined influence of environmental factors on viability of cryptosporidium parvum oocysts in water evaluated by fluorogenic vital dyes and excystation techniques253-9 Vet Parasitol894Animals *Cryptosporidium parvum Fluorescent Dyes Osmolar Concentration Parasitology/*methods Temperature Time Factors Water/*parasitologyMay 17iUsing a factorial experimental design, the combined effect of salinity, temperature and storage time on the viability of Cryptosporidium parvum oocysts in water was evaluated by fluorogenic vital dyes (4',6-diamidino-2-phenylindole and propidium iodide) and an excystation technique. Salinity, storage time and their interaction seemed to be the most influential factors, whereas temperature was not a significant factor. Under unfavourable conditions (salinity 35 per thousand, storage time 40 days), even more than 20% of oocysts remain viable, indicating a high risk of infection for immunocompromised individuals.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=107998380304-4017 Journal Article10799838Laboratorio de Parasitologia, Departamento de Microbiologia y Parasitologia, Facultad de Farmacia, Universidad de Santiago de Compostela, Coruna, Spain.~?!RFreire-Santos, F. Oteiza-Lopez, A. M. Vergara-Castiblanco, C. A. Ares-Mazas, M. E.1999jEffect of salinity, temperature and storage time on mouse experimental infection by Cryptosporidium parvum1-7 Vet Parasitol871Animals Animals, Newborn Animals, Suckling Cattle Cattle Diseases/*parasitology Cryptosporidiosis/parasitology/*veterinary Cryptosporidium parvum/*growth & development Intestines/parasitology Mice Multivariate Analysis Regression Analysis Sodium Chloride Temperature Time FactorsNovCryptosporidium parvum oocysts collected from a naturally infected calf were exposed to different salinity and temperature for 2, 21 and 40 days, and then inoculated intragastrically into coccidium-free neonatal mice. The intensity of infection as determined seven days later by examination of intestinal homogenates were statistically analysed. Salinity, time and salinity-time interaction were the only factors with significant effect on the infection intensity.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=106286950304-4017 Journal Article10628695}Departamento de Microbiologia y Parasitologia, Facultad de Farmacia, Universidad de Santiago de Compostela, La Coruna, Spain. ~?"Ares-Mazas, M. E. Fernandez-da Ponte, B. Vergara-Castiblanco, C. A. Freire-Santos, F. Quilez-Cinca, J. Causape-Valenzuela, A. C. Sanchez-Acedo, C.1999Oocysts, IgG levels and immunoblot patterns determined for Cryptosporidium parvum in bovine examined during a visit to a farm (northeastern Spain)185-93 Vet Parasitol813;Age Distribution Animals Antibodies, Monoclonal Antibodies, Protozoan/blood Antigens, Protozoan/chemistry Blotting, Western/veterinary Cattle Cattle Diseases/*epidemiology/immunology/parasitology Comparative Study Cryptosporidiosis/epidemiology/immunology/*veterinary Cryptosporidium parvum/*immunology Electrophoresis, Polyacrylamide Gel/veterinary Enzyme-Linked Immunosorbent Assay/veterinary Feces/parasitology Fluorescent Antibody Technique, Indirect/veterinary Immunoglobulin G/blood Incidence Parasite Egg Count/veterinary Spain/epidemiology Support, Non-U.S. Gov'tMar 1Single fecal and serum samples were individually collected from 101 bovines selected at random during a visit to a farm in northeastern Spain (Group I, 26 animals aged 2-36 days; Group II, 34 animals aged 1.5-4.5 months; Group III, 41 animals aged 20-24 months). Testing for the presence of Cryptosporidium parvum oocysts in feces (Monofluo Kit Cryptosporidium, Diagnostics Pasteur, France) indicated that 26% animals were infected (81% of Group I, 15% of Group II and 0% of Group III). Serological testing (ELISA for detection of specific anti-C. parvum IgG) indicated that 59% animals were seropositive (12% of Group I, 74% of Group II and 78% of Group III). Immunoblotting results indicate that cattle sera recognize C. parvum antigens of widely varying molecular weights and that the number of antigens recognized increases with age. Immunoblots revealed that some of the sera belonging to the Group I reacted with protein fractions between 15 and 20 kDa but none recognized the 21-23 kDa antigen. Only few sera in the Group II recognized the protein fraction between 15 and 20 kDa. The recognition of 21-23 kDa fraction was observed by four sera from uninfected and seropositive animals. Sera from all the seronegative Group II animals recognized few antigens and always with molecular weight greater than 50 kDa. Serum samples from both seropositive and seronegative animals belonging to the Group III recognized antigens with molecular weight ranging 15-20 kDa. Surprisingly, the protein fractions between 21 and 28 kDa reacted with approximately 30% of the sera from seropositive animals and only one of the nine sera from seronegative animals. The recognition of 42-46 kDa antigens increased with the age and only reacted with the sera from uninfected animals.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=101908620304-4017 Journal Article10190862Departamento de Microbiologia y Parasitologia, Facultad de Farmacia, Universidad de Santiago de Compostela, La Coruna, Spain. mpeares@usc.es~?#3Mtambo, M. M. Wright, E. Nash, A. S. Blewett, D. A.1996iInfectivity of a Cryptosporidium species isolated from a domestic cat (Felis domestica) in lambs and mice61-4 Res Vet Sci601Animals Animals, Domestic Animals, Newborn *Cat Diseases Cats Cryptosporidiosis/transmission/*veterinary Cryptosporidium/isolation & purification/*pathogenicity Feces/parasitology Mice Sheep Species Specificity Support, Non-U.S. Gov'tJanOTwo neonatal lambs were inoculated orally with purified Cryptosporidium species oocysts isolated from a farm cat. Oocysts first appeared in the faeces of the two lambs three and 10 days after infection. Two distinct sizes of oocysts were observed in the faeces of both the cat and the lambs, the smaller measuring approximately 5.0 x 4.5 microns and the larger measuring approximately 6.0 x 5.0 microns in diameter. The smaller type predominated. Histological examination of the alimentary tract of the lambs revealed endogenous stages of Cryptosporidium in the epithelial borders of the ileum. In addition, Cryptosporidium oocysts were detected in impression smears from the jejunum, ileum, caecum and colon. Suspensions of 10(3) oocysts from the faeces of the farm cat were inoculated into each of 10 newborn mice and 10(4) oocysts from the two experimentally infected lambs were inoculated into each of 20 newborn mice. Cryptosporidium oocysts were detected in gut homogenates from 19 of the 20 mice inoculated with oocysts from the lambs but in none of the mice inoculated with oocysts from the cat.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=87452580034-5288 Journal Article8745258UDepartment of Veterinary Medicine, University of Glasgow Veterinary School, Bearsden.}~?$iRobinson, P. Okhuysen, P. C. Chappell, C. L. Weinstock, J. V. Lewis, D. E. Actor, J. K. White, A. C., Jr.2003PSubstance P expression correlates with severity of diarrhea in cryptosporidiosis290-6 J Infect Dis1882.AIDS-Related Opportunistic Infections/complications/genetics Acquired Immunodeficiency Syndrome/complications/genetics Adolescent Adult Animals Chronic Disease Cryptosporidiosis/*complications/genetics Cryptosporidium parvum Diarrhea/*complications/*genetics/microbiology/physiopathology Female *Gene Expression Regulation Genetic Predisposition to Disease/genetics Human Jejunum/metabolism Male Middle Aged RNA, Messenger/analysis/genetics Substance P/*genetics/*metabolism Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.Jul 15/Cryptosporidiosis, caused by Cryptosporidium parvum, is self-limited in immunocompetent hosts but may cause chronic diarrhea in patients with acquired immunodeficiency syndrome (AIDS). Substance P (SP), a neuropeptide belonging to the tachykinin family, is expressed in gastrointestinal tract and can cause electrogenic chloride anion secretion. Therefore, we studied SP mRNA and protein expression in jejunal tissue samples of patients with AIDS with naturally occurring chronic cryptosporidiosis and healthy volunteers with mild cryptosporidiosis or asymptomatic infection after experimental C. parvum challenge. SP mRNA was associated with symptoms in cryptosporidiosis. SP protein levels were greater in symptomatic than asymptomatic volunteers. Similarly, greater expression of SP mRNA and protein were noted in patients with AIDS with chronic cryptosporidiosis versus immunocompetent volunteers with self-limited infection. This study demonstrates a direct correlation between SP levels and disease severity and may imply that SP plays a role in diarrhea mediation.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=128540860022-1899 Journal Article12854086Infectious Diseases Section, Department of Medicine, Baylor College of Medicine, One Baylor Plaza, Houston, TX 77030, USA. premar@bcm.tmc.edu~?%YOkhuysen, P. C. Rich, S. M. Chappell, C. L. Grimes, K. A. Widmer, G. Feng, X. Tzipori, S.2002wInfectivity of a Cryptosporidium parvum isolate of cervine origin for healthy adults and interferon-gamma knockout mice1320-5 J Infect Dis1859+Adult Animals Body Weight Cryptosporidiosis/*etiology Cryptosporidium parvum/classification/genetics/*pathogenicity Deer/*parasitology Diarrhea/etiology Disease Susceptibility Female Human Interferon Type II/*physiology Mice Mice, Knockout Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.May 1=The infectivity of a Cryptosporidium parvum isolate of cervine origin (type 2, Moredun) propagated in calves was investigated simultaneously in healthy adult human volunteers and in interferon-gamma knockout (GKO) mice. After exposure to 100-3000 oocysts, 16 volunteers recorded, for a duration of 6 weeks, the number and form of stools that they passed and any symptoms that they experienced. Oocyst excretion was assessed by enzyme-linked immunosorbent assay and direct immunofluorescence assay. Eleven subjects (69%) became ill, and 8 subjects (50%) shed oocysts in stool. The median duration of illness was 169 h, and the median number of unformed stools passed was 24. The duration and intensity of symptoms were more severe than were those associated with previously studied isolates. The median infectious dose was estimated to be 300 oocysts for humans and 1 oocyst for the GKO mouse model. The Moredun isolate was more pathogenic than the reference GCH-1 isolate. The GKO mouse model of cryptosporidiosis is useful for discerning isolate-specific differences in pathogenicity.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=120010500022-1899 Journal Article12001050Department of Medicine, Division of Infectious Diseases, and School of Public Health, University of Texas-Houston Health Science Center, Houston, TX 77030, USA. Pablo.C.Okhuysen@uth.tmc.edu~?&jWhite, A. C. Robinson, P. Okhuysen, P. C. Lewis, D. E. Shahab, I. Lahoti, S. DuPont, H. L. Chappell, C. L.2000Interferon-gamma expression in jejunal biopsies in experimental human cryptosporidiosis correlates with prior sensitization and control of oocyst excretion701-9 J Infect Dis1812AIDS-Related Opportunistic Infections/immunology/parasitology Animals Antibodies, Protozoan/blood Biopsy Cryptosporidiosis/*immunology/*parasitology/pathology Cryptosporidium parvum/growth & development/*immunology Cytokines/genetics/metabolism Human Immunohistochemistry In Situ Hybridization Interferon Type II/*biosynthesis/genetics Jejunum/*immunology/metabolism Plasmids/genetics RNA, Messenger/metabolism Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.FebnTo investigate the role of interferon (IFN)-gamma in human cryptosporidiosis, jejunal biopsies from experimentally infected volunteers and chronically infected AIDS patients were examined for IFN-gamma expression by in situ hybridization. IFN-gamma expression was compared with oocyst excretion, baseline serum anti-Cryptosporidium antibody, and symptoms. IFN-gamma mRNA was detected in biopsies from 13 of 26 volunteers after experimental infection but not in biopsies taken before C. parvum exposure or in biopsies from patients with AIDS-associated cryptosporidiosis. After challenge, 9 of 10 volunteers with baseline C. parvum antibody produced IFN-gamma, compared with 4 of 16 volunteers without baseline antibody (P<.01). Furthermore, IFN-gamma mRNA was detected in 9 of 13 volunteers who did not excrete oocysts, compared with 4 of 13 with organisms (P<.05). Thus, expression of IFN-gamma in the jejunum was associated with prior sensitization and absence of oocyst shedding. IFN-gamma production may explain the resistance to infection noted in sensitized persons but may not be involved in control of human primary infection.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=106693580022-1899 Journal Article10669358rInfectious Diseases Section, Dept. of Medicine, Baylor College of Medicine, Houston, TX 77030. arthurw@bcm.tmc.edu~?'JOkhuysen, P. C. Chappell, C. L. Crabb, J. H. Sterling, C. R. DuPont, H. L.1999NVirulence of three distinct Cryptosporidium parvum isolates for healthy adults1275-81 J Infect Dis1804Adult Animals Cattle Confidence Intervals Cryptosporidiosis/*physiopathology Cryptosporidium parvum/*genetics/isolation & purification/*pathogenicity Diarrhea/parasitology Enzyme-Linked Immunosorbent Assay Feces/parasitology Fluorescent Antibody Technique, Direct Genotype Human Iowa Parasite Egg Count Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S. Time Factors VirulenceOctIThe infectivity of three Cryptosporidium parvum isolates (Iowa [calf], UCP [calf], and TAMU [horse]) of the C genotype was investigated in healthy adults. After exposure, volunteers recorded the number and form of stools passed and symptoms experienced. Oocyst excretion was assessed by immunofluorescence. The ID50 differed among isolates: Iowa, 87 (SE, 19; 95% confidence interval [CI], 48.67-126); UCP, 1042 (SE, 1000; 95% CI, 0-3004); and TAMU, 9 oocysts (SE, 2.34; 95% CI, 4.46-13.65); TAMU versus Iowa, P=.002 or UCP, P=.019. Isolates also differed significantly (P=.045) in attack rate between TAMU (86%) and Iowa (52%) or UCP (59%). A trend toward a longer duration of diarrhea was seen for the TAMU (94.5 h) versus UCP (81.6 h) and Iowa (64.2 h) isolates. C. parvum isolates of the C genotype differ in their infectivity for humans.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=104791580022-1899 Journal Article10479158Division of Infectious Diseases, The University of Texas Medical School, Houston, Texas 77030, USA. Okhuysen@heart.med.uth.tmc.edu~?(hMoss, D. M. Chappell, C. L. Okhuysen, P. C. DuPont, H. L. Arrowood, M. J. Hightower, A. W. Lammie, P. J.1998qThe antibody response to 27-, 17-, and 15-kDa Cryptosporidium antigens following experimental infection in humans827-33 J Infect Dis1783Animals Antibodies, Protozoan/*immunology Antigens, Protozoan/*immunology Cryptosporidiosis/*immunology Cryptosporidium parvum/*immunology Human Immunoblotting Kinetics Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.SeplPrevious studies have suggested that persons infected with Cryptosporidium parvum develop antibody responses to 27-, 17-, and 15-kDa C. parvum antigens. Studies of volunteers infected with Cryptosporidium species provided an opportunity to evaluate the relationship between antibody reactivity to these antigens and infection outcome. As monitored by immunoblot, increases in specific antibody reactivity were more prevalent among volunteers who developed signs and symptoms of cryptosporidiosis (n = 11) than among asymptomatic infected (n = 7; P = .05) or oocyst-negative volunteers (n = 11; P = .02). Volunteers with preexisting IgG antibody to the 27-kDa antigen excreted fewer oocysts than volunteers without this antibody (P = .003). IgG reactivity to the 17-kDa antigens and IgM reactivity to the 27-kDa antigens were higher at day 0 for asymptomatic infected persons than for those who developed symptoms (P = .03 and P = .04, respectively). These results suggest that characteristic antibody responses develop following C. parvum infection and that persons with preexisting antibodies may be less likely to develop illness.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=97285530022-1899 Journal Article9728553Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Public Health Service, US Department of Health and Human Services, Atlanta, Georgia 30341, USA.d~?)=Chappell, C. L. Okhuysen, P. C. Sterling, C. R. DuPont, H. L.1996bCryptosporidium parvum: intensity of infection and oocyst excretion patterns in healthy volunteers232-6 J Infect Dis1731'Animals Cryptosporidiosis/*parasitology/transmission Cryptosporidium parvum/isolation & purification/*pathogenicity Diarrhea/parasitology Disease Susceptibility Feces/parasitology Human Parasite Egg Count Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S. TravelJaniData about human Cryptosporidium parvum infection have originated from travelers, community and day care center outbreaks, and persons infected with the human immunodeficiency virus. In addition, experimental infection in 29 antibody-negative, healthy, adult volunteers generated information on the dose-infection response of C. parvum (Iowa strain). In that report, low inocula were sufficient to cause infection in 18 and illness in 7 persons. To further define the duration and intensity of infection in this population, oocyst shedding patterns were investigated in the 18 subjects infected with C. parvum. Oocyst quantitation revealed that volunteers with diarrheal illness (n = 7) excreted more oocysts over the course of the infection than did volunteers without diarrhea (n = 11; P < .05). Symptomatic subjects were more likely to shed oocysts on consecutive days. Further, a statistical nonsignificant inverse trend (r2 = .330, P = .136) was seen between challenge dose and total excreted oocysts. This paradox may relate to receptor saturation or a toxic effect on cells, parasites, or both afforded by a high inoculum.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=85376640022-1899 Journal Article8537664`Center for Infectious Diseases, University of Texas School of Public Health, Houston 77030, USA.=~?+OSulaiman, I. M. Fayer, R. Lal, A. A. Trout, J. M. Schaefer, F. W., 3rd Xiao, L.2003Molecular characterization of microsporidia indicates that wild mammals Harbor host-adapted Enterocytozoon spp. as well as human-pathogenic Enterocytozoon bieneusi4495-501Appl Environ Microbiol698Animals Animals, Wild Base Sequence Enterocytozoon/classification/*genetics/isolation & purification Female Genotype Human Male Molecular Sequence Data Phylogeny Support, U.S. Gov't, Non-P.H.S.AugOver 13 months, 465 beavers, foxes, muskrats, otters, and raccoons were trapped in four counties in eastern Maryland and examined by molecular methods for microsporidia. A two-step nested PCR protocol was developed to amplify a 392-bp fragment of the internal transcribed spacer region of the rRNA gene of Enterocytozoon spp., with the use of primers complementary to the conserved regions of published nucleotide sequences. Fifty-nine PCR-positive samples were sequenced. Multiple alignments of these sequences identified 17 genotypes of Enterocytozoon spp. (WL1 to WL17); of these, 15 have not been reported before. Most of the genotypes were found in multiple species of wildlife and belonged to a major group consisting of all the previously described Enterocytozoon bieneusi genotypes from human and domestic animals. Some of the isolates from muskrats and raccoons formed two distinct groups. Results of this study indicate that fur-bearing mammals, especially those closely associated with surface water, can be a potential source of human-pathogenic E. bieneusi. However, there are also host-adapted Enterocytozoon genotypes in wildlife, which may represent species different from E. bieneusi and have no apparent public health significance. This is the first report of E. bieneusi in wildlife.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=129022340099-2240 Journal Article12902234Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 30341, USA.(~?,;Ryan, U. Xiao, L. Read, C. Zhou, L. Lal, A. A. Pavlasek, I.2003IIdentification of novel Cryptosporidium genotypes from the Czech Republic4302-7Appl Environ Microbiol697Adolescent Animals Animals, Domestic/parasitology Animals, Zoo/parasitology Cats Cattle Child Child, Preschool Cryptosporidiosis/*epidemiology/parasitology/veterinary Cryptosporidium/*classification/*genetics Czech Republic/epidemiology DNA, Protozoan/analysis DNA, Ribosomal/analysis Female Genotype Hamsters Heat-Shock Proteins 70/genetics Human Male Mice Molecular Sequence Data Phylogeny Polymerase Chain Reaction RNA, Ribosomal, 18S/genetics Rabbits Sequence Analysis, DNAJulkIsolates of Cryptosporidium from the Czech Republic were characterized from a variety of different hosts using sequence and phylogenetic analysis of the 18S ribosomal DNA and the heat-shock (HSP-70) gene. Analysis expanded the host range of accepted species and identified several novel genotypes, including horse, Eurasian woodcock, rabbit, and cervid genotypes.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=128398190099-2240 Journal Article12839819{Division of Health Sciences, Murdoch University, Murdoch, Western Australia 6150, Australia. unaryan@central.murdoch.edu.au6~?->Xiao, L. Singh, A. Limor, J. Graczyk, T. K. Gradus, S. Lal, A.2001dMolecular characterization of cryptosporidium oocysts in samples of raw surface water and wastewater1097-101Appl Environ Microbiol673Animals Cryptosporidiosis/parasitology Cryptosporidium/*classification/*genetics/growth & development/isolation & purification DNA, Protozoan/analysis/genetics DNA, Ribosomal/analysis/genetics Fresh Water/*parasitology Genotype Human Polymerase Chain Reaction Polymorphism, Restriction Fragment Length RNA, Ribosomal/genetics Sewage/*parasitology Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. *Water PollutionMarRecent molecular characterizations of Cryptosporidium parasites make it possible to differentiate the human-pathogenic Cryptosporidium parasites from those that do not infect humans and to track the source of Cryptosporidium oocyst contamination in the environment. In this study, we used a small-subunit rRNA-based PCR-restriction fragment length polymorphism (RFLP) technique to detect and characterize Cryptosporidium oocysts in 55 samples of raw surface water collected from several areas in the United States and 49 samples of raw wastewater collected from Milwaukee, Wis. Cryptosporidium parasites were detected in 25 surface water samples and 12 raw wastewater samples. C. parvum human and bovine genotypes were the dominant Cryptosporidium parasites in the surface water samples from sites where there was potential contamination by humans and cattle, whereas C. andersoni was the most common parasite in wastewater. There may be geographic differences in the distribution of Cryptosporidium genotypes in surface water. The PCR-RFLP technique can be a useful alternative method for detection and differentiation of Cryptosporidium parasites in water.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11229897,0099-2240 Evaluation Studies Journal Article11229897uDivision of Parasitic Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 30341, USA. lax0@cdc.gov~?.KXiao, L. Limor, J. Morgan, U. M. Sulaiman, I. M. Thompson, R. C. Lal, A. A.2000qSequence differences in the diagnostic target region of the oocyst wall protein gene of Cryptosporidium parasites5499-502Appl Environ Microbiol6612RAnimals Base Sequence Cattle Comparative Study Cryptosporidium/*genetics DNA Primers/genetics DNA, Protozoan/genetics Dogs *Genes, Protozoan Human Mice Molecular Sequence Data Polymerase Chain Reaction Protozoan Proteins/*genetics Sequence Homology, Nucleic Acid Species Specificity Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S.Dec&Nucleotide sequences of the Cryptosporidium oocyst wall protein (COWP) gene were obtained from various Cryptosporidium spp. (C. wrairi, C. felis, C. meleagridis, C. baileyi, C. andersoni, C. muris, and C. serpentis) and C. parvum genotypes (human, bovine, monkey, marsupial, ferret, mouse, pig, and dog). Significant diversity was observed among species and genotypes in the primer and target regions of a popular diagnostic PCR. These results provide useful information for COWP-based molecular differentiation of Cryptosporidium spp. and genotypes.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=110979360099-2240 Journal Article11097936National Center for Infectious Diseases, Centers for Disease Control and Prevention, Public Health Service, U.S. Department of Health and Human Services, Atlanta, Georgia 30341, USA. LAX0@CDC.GOV~?/5Xiao, L. Alderisio, K. Limor, J. Royer, M. Lal, A. A.2000Identification of species and sources of Cryptosporidium oocysts in storm waters with a small-subunit rRNA-based diagnostic and genotyping tool5492-8Appl Environ Microbiol6612Animals Base Sequence Cryptosporidium/classification/*genetics/*isolation & purification DNA Primers/genetics DNA, Protozoan/genetics DNA, Ribosomal/genetics Genotype Molecular Sequence Data New York Phylogeny Polymerase Chain Reaction/*methods Polymorphism, Restriction Fragment Length RNA, Protozoan/*genetics RNA, Ribosomal/*genetics Sequence Homology, Nucleic Acid Support, U.S. Gov't, Non-P.H.S. Variation (Genetics) Water/*parasitologyDecThe identification of Cryptosporidium oocysts in environmental samples is largely made by the use of an immunofluorescent assay. In this study, we have used a small-subunit rRNA-based PCR-restriction fragment length polymorphism technique to identify species and sources of Cryptosporidium oocysts present in 29 storm water samples collected from a stream in New York. A total of 12 genotypes were found in 27 positive samples; for 4 the species and probable origins were identified by sequence analysis, whereas the rest represent new genotypes from wildlife. Thus, this technique provides an alternative method for the detection and differentiation of Cryptosporidium parasites in environmental samples.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=110979350099-2240 Journal Article11097935vDivision of Parasitic Diseases, Centers for Disease Control and Prevention, Chamblee, Georgia 30341, USA. LAX0@CDC.GOV~?0ASulaiman, I. M. Morgan, U. M. Thompson, R. C. Lal, A. A. Xiao, L.2000rPhylogenetic relationships of Cryptosporidium parasites based on the 70-kilodalton heat shock protein (HSP70) gene2385-91Appl Environ Microbiol666RAnimals Base Sequence Cryptosporidiosis/*parasitology Cryptosporidium/classification/*genetics/isolation & purification Feces/parasitology Genes, Protozoan Heat-Shock Proteins 70/*genetics Human Molecular Sequence Data *Phylogeny Polymerase Chain Reaction Sequence Analysis, DNA Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.JunWe have characterized the nucleotide sequences of the 70-kDa heat shock protein (HSP70) genes of Cryptosporidium baileyi, C. felis, C. meleagridis, C. muris, C. serpentis, C. wrairi, and C. parvum from various animals. Results of the phylogenetic analysis revealed the presence of several genetically distinct species in the genus Cryptosporidium and eight distinct genotypes within the species C. parvum. Some of the latter may represent cryptic species. The phylogenetic tree constructed from these sequences is in agreement with our previous results based on the small-subunit rRNA genes of Cryptosporidium parasites. The Cryptosporidium species formed two major clades: isolates of C. muris and C. serpentis formed the first major group, while isolates of C. felis, C. meleagridis, C. wrairi, and eight genotypes of C. parvum formed the second major group. Sequence variations were also observed between C. muris isolates from ruminants and rodents. The HSP70 gene provides another useful locus for phylogenetic analysis of the genus Cryptosporidium.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=108314150099-2240 Journal Article10831415Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Public Health Services, U.S. Department of Health and Human Services, Atlanta, Georgia 30341, USA.~?1qMorgan, U. M. Xiao, L. Monis, P. Fall, A. Irwin, P. J. Fayer, R. Denholm, K. M. Limor, J. Lal, A. Thompson, R. C.20009Cryptosporidium spp. in domestic dogs: the "dog" genotype2220-3Appl Environ Microbiol665Animals Carbohydrate Sequence Cattle Consensus Sequence Cryptosporidium/*classification/*genetics/isolation & purification DNA, Protozoan/chemistry/genetics DNA, Ribosomal/chemistry/*genetics Dogs/*microbiology Genotype Heat-Shock Proteins 70/*genetics Human Marsupialia Mice Molecular Sequence Data *Phylogeny RNA, Ribosomal, 16S/*genetics Sequence Alignment Sequence Homology, Nucleic Acid Support, Non-U.S. Gov't SwineMayGenetic and phylogenetic characterization of Cryptosporidium isolates at two loci (18S rRNA gene and heat shock gene) from both Australian and United States dogs demonstrated that dog-derived Cryptosporidium isolates had a distinct genotype which is conserved across geographic areas. Phylogenetic analysis provided support for the idea that the "dog" genotype is, in fact, a valid species.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=107884040099-2240 Journal Article10788404World Health Organisation Collaborating Centre for the Molecular Epidemiology of Parasitic Infections and State Agricultural Biotechnology Centre, Division of Veterinary and Biomedical Sciences, Murdoch University, Murdoch, Western Australia.~?2#Sulaiman, I. M. Xiao, L. Lal, A. A.1999:Evaluation of Cryptosporidium parvum genotyping techniques4431-5Appl Environ Microbiol6510Animals Base Sequence Cryptosporidium parvum/*classification/genetics DNA, Protozoan/analysis Genotype Molecular Sequence Data Polymerase Chain Reaction Polymorphism, Restriction Fragment Length Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.OctWe evaluated the specificity and sensitivity of 11 previously described species differentiation and genotyping PCR protocols for detection of Cryptosporidium parasites. Genomic DNA from three species of Cryptosporidium parasites (genotype 1 and genotype 2 of C. parvum, C. muris, and C. serpentis), two Eimeria species (E. neischulzi and E. papillata), and Giardia duodenalis were used to evaluate the specificity of primers. Furthermore, the sensitivity of the genotyping primers was tested by using genomic DNA isolated from known numbers of oocysts obtained from a genotype 2 C. parvum isolate. PCR amplification was repeated at least three times with all of the primer pairs. Of the 11 protocols studied, 10 amplified C. parvum genotypes 1 and 2, and the expected fragment sizes were obtained. Our results indicate that two species-differentiating protocols are not Cryptosporidium specific, as the primers used in these protocols also amplified the DNA of Eimeria species. The sensitivity studies revealed that two nested PCR-restriction fragment length polymorphism (RFLP) protocols based on the small-subunit rRNA and dihydrofolate reductase genes are more sensitive than single-round PCR or PCR-RFLP protocols.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=105080710099-2240 Journal Article10508071Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 30341, USA. ~?3kXiao, L. Morgan, U. M. Limor, J. Escalante, A. Arrowood, M. Shulaw, W. Thompson, R. C. Fayer, R. Lal, A. A.1999SGenetic diversity within Cryptosporidium parvum and related Cryptosporidium species3386-91Appl Environ Microbiol658Animals Base Sequence Comparative Study Cryptosporidium/*genetics/isolation & purification Cryptosporidium parvum/*genetics/isolation & purification DNA Primers/genetics DNA, Protozoan/genetics DNA, Ribosomal/genetics Dogs Genes, Protozoan Human Mice Molecular Sequence Data Phylogeny Polymorphism, Restriction Fragment Length Sequence Homology, Nucleic Acid Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S. Variation (Genetics)AugTo assess the genetic diversity in Cryptosporidium parvum, we have sequenced the small subunit (SSU) rRNA gene of seven Cryptosporidium spp., various isolates of C. parvum from eight hosts, and a Cryptosporidium isolate from a desert monitor. Phylogenetic analysis of the SSU rRNA sequences confirmed the multispecies nature of the genus Cryptosporidium, with at least four distinct species (C. parvum, C. baileyi, C. muris, and C. serpentis). Other species previously defined by biologic characteristics, including C. wrairi, C. meleagridis, and C. felis, and the desert monitor isolate, clustered together or within C. parvum. Extensive genetic diversities were present among C. parvum isolates from humans, calves, pigs, dogs, mice, ferrets, marsupials, and a monkey. In general, specific genotypes were associated with specific host species. A PCR-restriction fragment length polymorphism technique previously developed by us could differentiate most Cryptosporidium spp. and C. parvum genotypes, but sequence analysis of the PCR product was needed to differentiate C. wrairi and C. meleagridis from some of the C. parvum genotypes. These results indicate a need for revision in the taxonomy and assessment of the zoonotic potential of some animal C. parvum isolates.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=104270230099-2240 Journal Article10427023Division of Parasitic Diseases, Centers for Disease Control and Prevention, Public Health Service, U.S. Department of Health and Human Services, Atlanta, Georgia 30341, USA. lax0@cdc.gov ~?4aXiao, L. Escalante, L. Yang, C. Sulaiman, I. Escalante, A. A. Montali, R. J. Fayer, R. Lal, A. A.1999]Phylogenetic analysis of Cryptosporidium parasites based on the small-subunit rRNA gene locus1578-83Appl Environ Microbiol654Animals Cattle Cryptosporidiosis/*parasitology Cryptosporidium/*classification/*genetics/isolation & purification Genes, Protozoan *Genes, rRNA Guinea Pigs Human Molecular Sequence Data Phylogeny Polymerase Chain Reaction/methods Polymorphism, Restriction Fragment Length RNA, Protozoan/genetics RNA, Ribosomal/genetics Sequence Analysis, DNA Species Specificity Support, U.S. Gov't, Non-P.H.S.AprBiological data support the hypothesis that there are multiple species in the genus Cryptosporidium, but a recent analysis of the available genetic data suggested that there is insufficient evidence for species differentiation. In order to resolve the controversy in the taxonomy of this parasite genus, we characterized the small-subunit rRNA genes of Cryptosporidium parvum, Cryptosporidium baileyi, Cryptosporidium muris, and Cryptosporidium serpentis and performed a phylogenetic analysis of the genus Cryptosporidium. Our study revealed that the genus Cryptosporidium contains the phylogenetically distinct species C. parvum, C. muris, C. baileyi, and C. serpentis, which is consistent with the biological characteristics and host specificity data. The Cryptosporidium species formed two clades, with C. parvum and C. baileyi belonging to one clade and C. muris and C. serpentis belonging to the other clade. Within C. parvum, human genotype isolates and guinea pig isolates (known as Cryptosporidium wrairi) each differed from bovine genotype isolates by the nucleotide sequence in four regions. A C. muris isolate from cattle was also different from parasites isolated from a rock hyrax and a Bactrian camel. Minor differences were also detected between C. serpentis isolates from snakes and lizards. Based on the genetic information, a species- and strain-specific PCR-restriction fragment length polymorphism diagnostic tool was developed.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=101032530099-2240 Journal Article10103253Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, U.S. Department of Health and Human Services, Atlanta, Georgia 30341, USA. ~?5FTate, K. W. Atwill, E. R. George, M. R. McDougald, M. K. Larsen, R. E.2000TCryptosporidium parvum transport from cattle fecal deposits on California rangelands295-299Journal of Range Management533Water-quality. Infection. Patterns. Oocysts. Giardia. Environment/Ecology in Current Contents(R)/Agricultural, Biology & Environmental Sciences. 2000 week 34 Reprint available from: Tate KW. Univ Calif Davis, Davis, CA 95616, USA, .,Cryptosporidium parvum is a fecal borne protozoan parasite that can be carried by and cause gastrointestinal illness in humans, cattle, and wildlife. The illness, cryptosporidiosis, can be fatal to persons with compromised immune systems. At question is the potential for C. parvum in cattle fecal deposits on rangeland watersheds to contaminate surface water. First, C. parvum oocysts must be released from fecal deposits during rainfall, becoming available for transport. In 1996, we examined the transport of C. parvum oocysts in overland flow from fecal deposits under natural rainfall and rangeland conditions at the San Joaquin Experimental Range in Madera County, Calif. Our null hypothesis was that C. parvum oocysts are not released from fecal pats and transported 1 m downslope as overland flow with rainfall. Paired plots were located on 10, 20, and 30% slope sites. Each plot was loaded with four, 200 g fecal pats dosed with 10(5) oocysts g(-1). Fats were placed 1.0 m above the base of each plot. Composite runoff samples from each plot were analyzed for oocyst concentration following each of 4 storm events. Oocysts were transported during each storm. Slope was a significant factor in oocyst transport, with oocyst transport increasing with slope. Although not significant, there was an apparent flushing effect of oocysts across storms, with the majority transported in the first 2 storms. A pilot rainfall simulation experiment also revealed a flushing phenomenon from pats during individual rainfall events. C. parvum oocysts in fecal pats on rangeland can be transported from fecal deposits during rainfall events, becoming available for transport to water-bodies. Future studies need to examine surface and subsurface transport of oocysts on rangeland hillslopes for distances greater than 1 m. [References: 25] 25English Article Current Contents(R)/Agriculture, Biology & Environmental Sciences. Reprint available from: Tate KW Univ Calif Davis Davis, CA 95616 USA Univ Calif Davis Davis, CA 95616 USA Univ Calif, Vet Med Teaching & Res Ctr, Sch Vet Med Tulare, CA 93274 USA Univ Calif Cooperat Extens San Luis Obispo, CA 93446 USA Univ Calif Cooperat Extens Madera, CA 93637 USA 0009 J. Range Manage-337DA-0009 337DA: Document Delivery available ~?68Tate, K. W. Atwill, E. R. McDougald, N. K. George, M. R.2003ESpatial and temporal patterns of cattle feces deposition on rangeland432-438Journal of Range Management565*Cryptosporidium-parvum oocysts. Dehydrated molasses supplement. Grazing distribution. Sheep. Herds. Environment/Ecology in Current Contents(R)/Agricultural, Biology & Environmental Sciences. 2003 week 42 Reprint available from: Tate KW. Univ Calif Davis, Agron & Range Serv, Davis, CA 95616, USA, .3The objective of this study was to identify and model environmental and management factors associated with cattle feces deposition patterns across annual rangeland watersheds in the Sierra Nevada foothills. Daily cattle fecal load accumulation rates were calculated from seasonal fecal loads measured biannually on 40 m(2) permanent transects distributed across a 150.5 ha pasture in Madera County, Calif. during the 4 year period from 1995 through 1998. Associations between daily fecal load per season, livestock management, and environmental factors measured for each transect were determined using a linear mixed effects model. Cattle feces distribution patterns were significantly associated with location of livestock attractants, slope percentage, slope aspect, hydrologic position, and season. Transects located in livestock concentration areas experienced a significantly higher daily fecal load compared to transects outside of these concentration areas (P < 0.001). Percent slope was negatively associated with daily fecal load, but this association had a significant interaction with slope aspect (P = 0.02). Daily fecal load was significantly lower during the wet season compared to the dry season (P = 0.002). Daily fecal loading rates across hydrologic positions were dependent upon season. Our results illustrate the opportunities to reduce the risk of water quality contamination by strategic placement of cattle attractants, and provide a means to predict cattle feces deposition based upon inherent watershed characteristics and management factors. [References: 27] 27rEnglish Article Current Contents(R)/Agriculture, Biology & Environmental Sciences. Reprint available from: Tate KW Univ Calif Davis, Agron & Range Serv Davis, CA 95616 USA Univ Calif Davis, Agron & Range Serv Davis, CA 95616 USA Univ Calif, Sch Vet Med, Vet Med Teaching & Res Ctr Tulare, CA 93274 USA Univ Calif Cooperat Extens Madera, CA 93637 USA 0005 J. Range Manage-724AD-0005 724AD: Document Delivery available~?7LBajer, A. Caccio, S. Bednarska, M. Behnke, J. M. Pieniazek, N. J. Sinski, E.2003iPreliminary molecular characterization of Cryptosporidium parvum isolates of wildlife rodents from Poland1053-5 J Parasitol895Animals Animals, Wild Base Sequence Cryptosporidiosis/parasitology/*veterinary Cryptosporidium parvum/classification/*genetics DNA Primers/chemistry DNA, Protozoan/chemistry Disease Reservoirs/veterinary Genotype Microtinae/*parasitology Molecular Sequence Data Muridae/*parasitology Poland Polymerase Chain Reaction/veterinary Protozoan Proteins/genetics RNA, Ribosomal, 18S/genetics Rodent Diseases/*parasitology Sequence Analysis, DNA/veterinary Support, Non-U.S. Gov'tOctIsolates of Cryptosporidium were collected from 3 species of woodland and field rodents (Clethrionomys glareolus, Microtus arvalis, and Apodemus flavicollis) and were characterized by polymerase chain reaction amplification and sequencing of fragments of the oocyst wall protein (COWP) gene and of the 18S ribosomal RNA gene. Sequence analysis of these markers revealed that the animals were infected with C. parvum, and that the genotype involved was almost identical to the mouse genotype previously described from Mus musculus. Thus, small rodents should be considered as an important reservoir of C. parvum genotypes closely related to the zoonotic genotype 2 and potentially hazardous to humans.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=146271560022-3395 Journal Article14627156rDepartment of Parasitology, Faculty of Biology, Warsaw University, Warszawa, Miecznikowa 1, 02-096 Warsaw, Poland.s~?8CChalmers, R. M. Sturdee, A. P. Bull, S. A. Miller, A. Wright, S. E.1997The prevalence of Cryptosporidium parvum and C. muris in Mus domesticus, Apodemus sylvaticus and Clethrionomys glareolus in an agricultural system478-82 Parasitol Res835#Agriculture Animals Animals, Wild/parasitology Cryptosporidiosis/epidemiology/parasitology/*veterinary Cryptosporidium parvum/*isolation & purification Female Male Mice Microtinae/parasitology Muridae/parasitology Prevalence Rodent Diseases/*epidemiology/parasitology Support, Non-U.S. Gov'tOWild mice and voles were tested for Cryptosporidium during a 2-year survey at an agricultural site in Warwickshire, United Kingdom. C. parvum and C. muris, the two cryptosporidial species known to infect mammals, were detected. Prevalence figures of 22%, 21% and 13% noted for C. parvum for Mus domesticus, Apodemus sylvaticus and Clethrionomys glareolus, respectively, were higher than those recorded for C. muris at 10%, 6% and 2%. C. parvum causes the sometimes severe diarrhoeal disease cryptosporidiosis in many hosts, but the wild rodents were asymptomatic. The discovery of C. muris in A. sylvaticus and C. glareolus confirms a wider distribution in wild rodents than has previously been reported. Rodents may represent a significant reservoir of Cryptosporidium with a high potential for infection of man and livestock due to cohabitation.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=91973960932-0113 Journal Article9197396\School of Natural and Environmental Sciences, Coventry University, UK. byk032@coventry.ac.uk ~?9)Daniels, M. J. Hutchings, M. R. Greig, A.2003lThe risk of disease transmission to livestock posed by contamination of farm stored feed by wildlife excreta561-8Epidemiol Infect1303KAnimal Feed/analysis/*microbiology Animals *Animals, Wild Birds/microbiology Cattle Cattle Diseases/epidemiology/prevention & control/transmission Cryptosporidiosis/epidemiology/prevention & control/transmission/*veterinary Feces/*microbiology Food Contamination Incidence Models, Statistical Paratuberculosis/epidemiology/*prevention & control/transmission Prevalence Risk Rodentia/microbiology Salmonella Infections, Animal/epidemiology/*prevention & control/transmission Scotland/epidemiology Sheep Sheep Diseases/epidemiology/prevention & control/transmission Support, Non-U.S. Gov'tJunLivestock feed is susceptible to contamination from wildlife excreta during on farm storage. Pathogens associated with diseases such as paratuberculosis, salmonella and cryptosporidiosis are present in wild rodent and bird excreta. Feed stores on four farms in the east of Scotland were monitored monthly over the winter of 1998/9 to quantify the levels of wildlife faecal contamination. A mean of 79.9 rodent (95% confidence interval: 37.5-165.9) and 24.9 (14.3-41.7) bird faeces were deposited per m2 of stored feed per month. It was estimated that individual cattle and sheep could encounter 1626 and 814 wildlife faeces over the winter. A model based on the numbers of infected faeces consumed per annum was used to estimate 'infectious probabilities' (Pinf) required to account for the reported prevalence of paratuberculosis, salmonella and cryptosporidiosis in sheep and cattle in the east of Scotland in 1998. Based on empirical data for input variables [the number of faeces encountered (Fe), the number ingested (Fi) and the prevalence of infection in wildlife species (Ip)], Pinf estimates ranged from 1.6 x 10(-8) for cryptosporidiosis in sheep to 8.2 x 10(-8) for paratuberculosis in cattle. The model suggested that ingestion of feed contaminated by wildlife faeces could account for the prevalence of all three diseases. Wildlife faecal contamination of stored feed should be given serious consideration as a potential source of infection to livestock.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12825742+0950-2688 Journal Article Multicenter Study12825742jAnimal Nutrition and Health Department, Scottish Agricultural College, West Mains Road, Edinburgh EH9 3JG.~?:jXiao, L. Sulaiman, I. M. Ryan, U. M. Zhou, L. Atwill, E. R. Tischler, M. L. Zhang, X. Fayer, R. Lal, A. A.2002nHost adaptation and host-parasite co-evolution in Cryptosporidium: implications for taxonomy and public health1773-85Int J Parasitol3214Actins/genetics Animals Base Sequence Cryptosporidium/classification/*genetics/physiology DNA, Protozoan/genetics *Evolution Genes, rRNA/genetics Genotype Heat-Shock Proteins 70/genetics Host-Parasite Relations/genetics Human Molecular Sequence Data Phylogeny Polymerase Chain Reaction/methods *Public Health Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S. Variation (Genetics)Dec 19To assess the genetic diversity and evolution of Cryptosporidium parasites, the partial ssrRNA, actin, and 70kDa heat shock protein (HSP70) genes of 15 new Cryptosporidium parasites were sequenced. Sequence data were analysed together with those previously obtained from other Cryptosporidium parasites (10 Cryptosporidium spp. and eight Cryptosporidium genotypes). Results of this multi-locus genetic characterisation indicate that host adaptation is a general phenomenon in the genus Cryptosporidium, because specific genotypes were usually associated with specific groups of animals. On the other hand, host-parasite co-evolution is also common in Cryptosporidium, as closely related hosts usually had related Cryptosporidium parasites. Results of phylogenetic analyses suggest that the Cryptosporidium parvum bovine genotype and Cryptosporidium meleagridis were originally parasites of rodents and mammals, respectively, but have subsequently expanded their host ranges to include humans. Understanding the evolution of Cryptosporidium species is important not only for clarification of the taxonomy of the parasites but also for assessment of the public health significance of Cryptosporidium parasites from animals.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=124644240020-7519 Journal Article12464424Division of Parasitic Diseases, Mail Stop F-12, Centers for Disease Control and Prevention, U.S. PHS/DHHS, 4770 Buford Highway, Atlanta, GA 30341, USA. lax0@cdc.gov~?;Morgan, U. M. Monis, P. T. Xiao, L. Limor, J. Sulaiman, I. Raidal, S. O'Donoghue, P. Gasser, R. Murray, A. Fayer, R. Blagburn, B. L. Lal, A. A. Thompson, R. C.2001IMolecular and phylogenetic characterisation of Cryptosporidium from birds289-96Int J Parasitol313SAnimals Bird Diseases/*parasitology Birds Cryptosporidiosis/parasitology/*veterinary Cryptosporidium/*classification/*genetics DNA, Protozoan/analysis/genetics DNA, Ribosomal/analysis/genetics Ducks Genes, rRNA Heat-Shock Proteins 70/genetics Molecular Sequence Data *Phylogeny RNA, Ribosomal, 18S/genetics Sequence Analysis, DNA SongbirdsMar3Avian isolates of Cryptosporidium species from different geographic locations were sequenced at two loci, the 18S rRNA gene and the heat shock gene (HSP-70). Phylogenetic analysis of the sequence data provided support for the existence of a new avian species of Cryptosporidium infecting finches and a second species infecting a black duck. The identity of Cryptosporidium baileyi and Cryptosporidium meleagridis as valid species was confirmed. Also, C. baileyi was identified in a number of isolates from the brown quail extending the host range of this species.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=112264560020-7519 Journal Article11226456World Health Organisation Collaborating Centre for the Molecular Epidemiology of Parasitic Infections and State Agricultural Biotechnology Centre, Murdoch University, WA 6150, Australia. morgan@numbat.murdoch.edu.au\~?<!Fayer, R. Morgan, U. Upton, S. J.2000KEpidemiology of Cryptosporidium: transmission, detection and identification1305-22Int J Parasitol3012-13Animals Cryptosporidiosis/diagnosis/*transmission Cryptosporidium/classification/genetics/*isolation & purification Genotype Human *ZoonosesNovThere are 10 valid species of Cryptosporidium and perhaps other cryptic species hidden under the umbrella of Cryptosporidium parvum. The oocyst stage is of primary importance for the dispersal, survival, and infectivity of the parasite and is of major importance for detection and identification. Because most oocysts measure 4-6 microm, appear nearly spherical, and have obscure internal structures, there are few or no morphometric features to differentiate species and in vitro cultivation does not provide differential data as for bacteria. Consequently, we rely on a combination of data from three tools: morphometrics, molecular techniques, and host specificity. Of 152 species of mammals reported to be infected with C. parvum or an indistinguishable organism, very few oocysts have ever been examined using more than one of these tools. This paper reviews the valid species of Cryptosporidium, their hosts and morphometrics; the reported hosts for the human pathogen, C. parvum; the mechanisms of transmission; the drinking water, recreational water, and food-borne outbreaks resulting from infection with C. parvum; and the microscopic, immunological, and molecular methods used to detect and identify species and genotypes.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1111325710020-7519 Journal Article Review Review, Academic11113257United States Department of Agriculture, Agricultural Research Institute, LPSI, 10300 Baltimore Avenue, Beltsville, MD 20705, USA. rfayer@lpsi.barc.usda.gov~?=;Morgan, U. M. Xiao, L. Fayer, R. Lal, A. A. Thompson, R. C.1999GVariation in Cryptosporidium: towards a taxonomic revision of the genus1733-51Int J Parasitol2911wAnimals Cryptosporidiosis/*parasitology Cryptosporidium/*classification/*genetics Human Phylogeny *Variation (Genetics)NovVCryptosporidium is an important cause of enteric disease in humans and other animals. Limitations associated with conventional diagnostic methods for cryptosporidiosis based on morphological features, coupled with the difficulty of characterising parasites isolated in the laboratory, have restricted our ability to clearly identify species. The application of sensitive molecular approaches has obviated the necessity for laboratory amplification. Such studies have found considerable evidence of genetic heterogeneity among isolates of Cryptosporidium from different species of vertebrate, and there is now mounting evidence suggesting that a series of host-adapted genotypes/strains/species of the parasite exist. In this article, studies on the molecular characterisation of Cryptosporidium during the last 5 years are reviewed and put into perspective with the past and present taxonomy of the genus. The predictive value of achieving a sound taxonomy for the genus Cryptosporidium with respect to understanding its epidemiology and transmission and controlling outbreaks of the disease is also discussed.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1061692010020-7519 Journal Article Review Review, Tutorial10616920World Health Organisation Collaborating Centre for the Molecular Epidemiology of Parasitic Infections, and State Agricultural Biotechnology Centre, Murdoch University, Australia. morgan@numbat.murdoch.edu.au ~?>GFayer, R. Gasbarre, L. Pasquali, P. Canals, A. Almeria, S. Zarlenga, D.1998iCryptosporidium parvum infection in bovine neonates: dynamic clinical, parasitic and immunologic patterns49-56Int J Parasitol281Animals Animals, Newborn CD4-Positive T-Lymphocytes/immunology CD8-Positive T-Lymphocytes/immunology Cattle *Cattle Diseases Cryptosporidiosis/immunology/physiopathology/*veterinary *Cryptosporidium parvum/isolation & purification Diarrhea/parasitology/veterinary Feces/parasitology Immunity, Cellular Interferon Type II/biosynthesis Interleukin-12/biosynthesis Lymphocyte Count Male RNA, Messenger/biosynthesis Transcription, GeneticJanTwenty-six experimentally infected calves were monitored daily for oocyst excretion. All began excreting oocysts 3-6 days p.i. Most calves (n = 23) excreted oocysts for 6-9 days, with a daily range from 4 x 10(2) to 4.15 x 10(7) oocysts g(-1) of faeces. Over half the calves excreted peak numbers of oocysts 6-8 days p.i. Diarrhoea, observed intermittently beginning as early as day 3 p.i., lasted 4-16 days and varied greatly in severity from calf to calf. In a second study, nine of 18 calves were orally inoculated with 5 x 10(6) oocysts between birth and 2 days of age and nine remained uninfected. Monoclonal antibodies for cell surface markers indicated substantial increases in CD4+ and CD8+ T cells in the intraepithelial lymphocyte population of the ilea of infected calves at 7-9 days of age. RT-PCR demonstrated increases in mRNA for interleukin-12 and interferon-gamma that correlated with increases in both CD4+ and CD8 + intraepithelial lymphocyte cells. Increased mRNA for interleukin-12 and interferon-gamma from lamina propria lymphocytes correlated with increased numbers of CD8+ cells. No changes were found in interleukin-2, interleukin-4 or interleukin-10 mRNA levels. However, interleukin-15 mRNA, possibly from epithelial cells contaminating intraepithelial lymphocytes, was decreased in infected calves and had a negative correlation with increases in CD4+ and CD8+ cells. No differences were detected in mRNA levels for cytokines from lymph node lymphocytes.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=95043340020-7519 Journal Article9504334United States Department of Agriculture, Agricultural Research Service, Immunology and Disease Resistance Laboratory, Beltsville, MD 20705, USA. rfayer@ggpl.arsusda.gov~??Fayer, R. Dubey, J. P.1987aComparative epidemiology of coccidia: clues to the etiology of equine protozoal myeloencephalitis615-20Int J Parasitol172Animals Coccidia/physiology Comparative Study Encephalomyelitis, Equine/etiology/*veterinary Female Horse Diseases/*etiology Horses Male *Protozoan Infections, AnimalFebdhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=3294672 0020-7519 Journal Article Review3294672 K~?@fHeitman, T. L. Frederick, L. M. Viste, J. R. Guselle, N. J. Morgan, U. M. Thompson, R. C. Olson, M. E.2002Prevalence of Giardia and Cryptosporidium and characterization of Cryptosporidium spp. isolated from wildlife, human, and agricultural sources in the North Saskatchewan River Basin in Alberta, Canada530-41Can J Microbiol486Animals Canada/epidemiology Cattle Cryptosporidiosis/epidemiology/*veterinary Cryptosporidium/genetics/*isolation & purification Electrophoresis, Polyacrylamide Gel Environmental Monitoring Feces/parasitology Giardia/genetics/isolation & purification Giardiasis/epidemiology/*veterinary Human Polymerase Chain Reaction Prevalence Support, Non-U.S. Gov't Water/*parasitology *Water SupplyJungThe environmental distribution of Giardia spp. and Cryptosporidium spp. is dependent upon human, agricultural, and wildlife sources. The significance of each source with regard to the presence of parasites in the environment is unknown. This 2-year study examined parasite prevalence in human sewage influent, wildlife, and agricultural sources associated with the North Saskatchewan River Basin in Alberta, Canada. Fecal samples were collected from cow-calf, dairy, and hog operations in the watershed area. Sewage-treatment facilities were sampled bimonthly during the 2-year study, and wildlife scat was collected at locations along tributaries of the North Saskatchewan River. All samples were analyzed for the presence of Giardia and Cryptosporidium, using sucrose-gradient separation followed by immunofluorescent microscopy. Giardia and Cryptosporidium were detected in all three sources. The lowest prevalence of both Giardia (3.28%) and Cryptosporidium (0.94%) was found in wildlife, with 6 of 19 species testing positive. Sewage influent had the highest prevalence of Giardia (48.80%) and Cryptosporidium parvum-like oocysts (5.42%); however, the concentration of both parasites was minimal compared with the concentration detected in cattle feces. Cow-calf sources contained the highest concentration of Giardia (mean 5800/g feces, P < 0.01), and dairy sources contained the highest concentration of C. parvum-like oocysts (mean 295/g feces, P < 0.01). Although prevalence and concentration are higher in cattle feces than in sewage, the Giardia and Cryptosporidium in animal manure do not have direct access to water draining into the North Saskatchewan River. PCR-based characterization of rDNA from isolates of Cryptosporidium collected from Alberta human, pig, calf, mature steer, dog, cat, and beaver hosts revealed distinct genetic differences that may reflect host specificity.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=121666800008-4166 Journal Article12166680ZDepartment of Microbiology and Infectious Diseases, The University of Calgary, AB, Canada.k~?A:Jellison, K. L. Distel, D. L. Hemond, H. F. Schauer, D. B.2004Phylogenetic analysis of the hypervariable region of the 18S rRNA gene of Cryptosporidium oocysts in feces of Canada geese (Branta canadensis): evidence for five novel genotypes452-8Appl Environ Microbiol701kAnimals Cryptosporidium/classification/genetics/growth & development/*isolation & purification DNA, Protozoan/analysis DNA, Ribosomal/analysis Feces/*parasitology Geese/*parasitology *Genes, rRNA Oocysts/classification/genetics/*isolation & purification *Phylogeny RNA, Ribosomal, 18S/*genetics Sequence Analysis, DNA Support, U.S. Gov't, Non-P.H.S. United StatesJanTo assess genetic diversity in Cryptosporidium oocysts from Canada geese, 161 fecal samples from Canada geese in the United States were analyzed. Eleven (6.8%) were positive for Cryptosporidium spp. following nested PCR amplification of the hypervariable region of the 18S rRNA gene. Nine PCR products from geese were cloned and sequenced, and all nine diverged from previously reported Cryptosporidium 18S rRNA gene sequences. Five sequences were very similar or identical to each other but genetically distinct from that of Cryptosporidium baileyi; two were most closely related to, but genetically distinct from, the first five; and two were distinct from any other sequence analyzed. One additional sequence in the hypervariable region of the 18S rRNA gene isolated from a cormorant was identical to that of C. baileyi. Phylogenetic analysis provided evidence for new genotypes of Cryptosporidium species in Canada geese. Results of this study suggest that the taxonomy of Cryptosporidium species in geese is complex and that a more complete understanding of genetic diversity among these parasites will facilitate our understanding of oocyst sources and species in the environment.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=147116740099-2240 Journal Article14711674Department of Civil and Environmental Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA. kjellison@lehigh.edu~?B,Jellison, K. L. Hemond, H. F. Schauer, D. B.2002SSources and species of cryptosporidium oocysts in the Wachusett Reservoir watershed569-75Appl Environ Microbiol682Animals Cattle Cryptosporidiosis/epidemiology/parasitology/prevention & control Cryptosporidium/*classification/genetics/growth & development/*isolation & purification DNA, Protozoan/analysis DNA, Ribosomal/analysis Feces/parasitology Fresh Water/*parasitology Human Massachusetts Polymerase Chain Reaction Polymorphism, Restriction Fragment Length RNA, Ribosomal, 16S/genetics Sequence Analysis, DNA Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S. *Water SupplyFebUnderstanding the behavior of Cryptosporidium oocysts in the environment is critical to developing improved watershed management practices for protection of the public from waterborne cryptosporidiosis. Analytical methods of improved specificity and sensitivity are essential to this task. We developed a nested PCR-restriction fragment length polymorphism assay that allows detection of a single oocyst in environmental samples and differentiates the human pathogen Cryptosporidium parvum from other Cryptosporidium species. We tested our method on surface water and animal fecal samples from the Wachusett Reservoir watershed in central Massachusetts. We also directly compared results from our method with those from the immunofluorescence microscopy assay recommended in the Information Collection Rule. Our results suggest that immunofluorescence microscopy may not be a reliable indicator of public health risk for waterborne cryptosporidiosis. Molecular and environmental data identify both wildlife and dairy farms as sources of oocysts in the watershed, implicate times of cold water temperatures as high-risk periods for oocyst contamination of surface waters, and suggest that not all oocysts in the environment pose a threat to public health.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11823192,0099-2240 Evaluation Studies Journal Article11823192Department of Civil Engineering. Division of Bioengineering and Environmental Health, Massachusetts Institute of Technology, 77 Massachusetts Ave., Cambridge, MA 02139, USA.9~?CAKatsumata, T. Hosea, D. Ranuh, I. G. Uga, S. Yanagi, T. Kohno, S.2000@Short report: possible Cryptosporidium muris infection in humans70-2Am J Trop Med Hyg621xAnimals Antibodies, Monoclonal Child, Preschool Cryptosporidiosis/*parasitology/pathology Cryptosporidium/chemistry/classification/*isolation & purification DNA Primers DNA, Protozoan/chemistry/isolation & purification Feces/parasitology Female Fluorescent Antibody Technique, Direct Human Indonesia Microscopy, Phase-Contrast Polymerase Chain Reaction Support, Non-U.S. Gov'tJanOocysts of cryptosporidia whose morphology resembled that of Cryptosporidium muris were found in the stool of 2 healthy girls in Surabaya, Indonesia. The oocysts were predominantly oval and measured 7.75+/-0.17 x 5.55+/-0.13 microm (mean+/-SD). The number of oocysts excreted were more than 10(5) per gram of stool. The oocysts were well stained with fluorescein-conjugated monoclonal antibody to Cryptosporidium. The specimens from both girls containing the oocysts showed a positive result by the polymerase chain reaction (PCR) using primers specific for the genus Cryptosporidium, but a negative result by the PCR using primers specific for C. parvum. The 2 girls passed oocysts for 5 and 6 days, respectively. They did not complain of any symptoms during the passage of oocysts.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=107617260002-9637 Journal Article10761726WDepartment of Parasitology, Institute of Tropical Medicine, Nagasaki University, Japan.~?DRKatsumata, T. Hosea, D. Wasito, E. B. Kohno, S. Hara, K. Soeparto, P. Ranuh, I. G.1998SCryptosporidiosis in Indonesia: a hospital-based study and a community-based survey628-32Am J Trop Med Hyg594Adolescent Adult Aged Animals Cats Child Child, Preschool Cryptosporidiosis/*epidemiology/prevention & control/transmission Diarrhea/etiology Female Human Indonesia/epidemiology Infant Infant, Newborn Male Middle Aged Prevalence Seasons Support, Non-U.S. Gov'tOctHospital-based and community-based studies were conducted to understand the prevalence and mode of transmission of Cryptosporidium parvum infection in Surabaya, Indonesia. In both studies people with and without diarrhea were examined for oocysts. A community-based survey included questionnaires to a community and stool examination of cats. Questionnaires covered demographic information, health status, and hygienic indicators. In the hospital, C. parvum oocysts were found in 26 (2.8%) of 917 patients with diarrhea and 15 (1.4%) of 1,043 control patients. The most susceptible age was less than two years old. The prevalence was higher during the rainy season. A community-based study again showed that C. parvum oocysts were frequently detected in diarrhea samples (8.2%), exclusively during rainy season. Thirteen (2.4%) of 532 cats passed C. parvum oocysts. A multiple logistic regression model indicated that contact with cats, rain, flood, and crowded living conditions are significant risk factors for Cryptosporidium infection.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=97904420002-9637 Journal Article9790442aDepartment of Parasitology, Institute of Tropical Medicine, Nagasaki University, Sakamoto, Japan.\~?E^Ong, C. S. Eisler, D. L. Alikhani, A. Fung, V. W. Tomblin, J. Bowie, W. R. Isaac-Renton, J. L.2002}Novel cryptosporidium genotypes in sporadic cryptosporidiosis cases: first report of human infections with a cervine genotype263-8Emerg Infect Dis83Animals British Columbia Cryptosporidiosis/*genetics Cryptosporidium/*genetics/isolation & purification Deer Feces/parasitology Genome Genotype Human New York Polymerase Chain Reaction Polymorphism, Restriction Fragment Length RNA, Ribosomal, 18S/genetics Support, Non-U.S. Gov'tMarCIn this study, we genotyped parasites from the fecal specimens of sporadic cryptosporidiosis cases in British Columbia from 1995 to 1999. Genotyping was conducted by polymerase chain amplification of the internal transcribed spacer region, a hypervariable region in the 18S rRNA gene and the Cryptosporidium oocyst wall protein gene. Subsequent analysis was by restriction fragment length polymorphism and DNA sequencing. We identified two new Cryptosporidium genotypes in humans. One of these genotypes has been found recently in deer in New York state. The other genotype has not been identified in humans or animals. These results have important implications for drinking water quality strategies, especially for communities that obtain drinking water supplies from surface sources located in forested regions with deer populations.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=119270231080-6040 Journal Article11927023NUniversity of British Columbia, Vancouver, BC, Canada. cong@interchange.ubc.ca~?FPeng, M. M. Xiao, L. Freeman, A. R. Arrowood, M. J. Escalante, A. A. Weltman, A. C. Ong, C. S. Mac Kenzie, W. R. Lal, A. A. Beard, C. B.1997nGenetic polymorphism among Cryptosporidium parvum isolates: evidence of two distinct human transmission cycles567-73Emerg Infect Dis34Amino Acid Sequence Animals Base Sequence Cattle Cryptosporidium parvum/classification/*genetics DNA, Protozoan/analysis/chemistry Genotype Human Mice Molecular Sequence Data Polymerase Chain Reaction Polymorphism (Genetics)Oct-DecWe report the results of molecular analysis of 39 isolates of Cryptosporidium parvum from human and bovine sources in nine human outbreaks and from bovine sources from a wide geographic distribution. All 39 isolates could be divided into either of two genotypes, on the basis of genetic polymorphism observed at the thrombospondin-related adhesion protein (TRAP-C2) locus. Genotype 1 was observed only in isolates from humans. Genotype 2, however, was seen in calf isolates and in isolates from a subset of human patients who reported direct exposure to infected cattle or consumed items thought to be contaminated with cattle faces. Furthermore, experimental infection studies showed that genotype 2 isolates were infective to mice or calves under routine laboratory conditions, whereas genotype 1 isolates were not. These results support the occurrence of two distinct transmission cycles of C. parvum in humans.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=936661111080-6040 Journal Article Review Review, Tutorial9366611'University of Michigan, Ann Arbor, USA.7~?G(Pedraza-Diaz, S. Amar, C. McLauchlin, J.2000The identification and characterisation of an unusual genotype of Cryptosporidium from human faeces as Cryptosporidium meleagridis189-94FEMS Microbiol Lett1892Amino Acid Sequence Animals Cryptosporidium/*genetics/isolation & purification Feces/*parasitology *Genes, Protozoan Genotype Human Molecular Sequence Data Support, Non-U.S. Gov'tAug 15ZAn unusual genotype of Cryptosporidium was identified in the faeces of six human patients by PCR/RFLP analysis of the Cryptosporidium oocyst wall protein (COWP) gene. Conventional microscopy showed oocysts indistinguishable in size from those of Cryptosporidium parvum, which reacted with two different commercially available anti-oocyst monoclonal antibodies. The isolates were further characterised by PCR/RFLP analysis of the thrombospondin-related adhesive protein of Cryptosporidium-1 (TRAP-C1) genes as well as by DNA sequencing of the COWP and the TRAP-C1 gene fragments and of two regions of the 18S rRNA gene. Sequence analysis of the COWP, TRAP-C1, and 18S rRNA gene fragments confirmed that this genotype is genetically distinct from C. parvum. 18S rRNA gene sequences were found to be identical to those published for Cryptosporidium meleagridis.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=109307360378-1097 Journal Article10930736Food Safety Microbiology Laboratory, Division of Gastrointestinal Infections, PHLS Central Public Health Laboratory, 61 Colindale Avenue, NW9 5HT, London, UK. g~?HFPedraza-Diaz, S. Amar, C. Iversen, A. M. Stanley, P. J. McLauchlin, J.2001Unusual cryptosporidium species recovered from human faeces: first description of Cryptosporidium felis and Cryptosporidium 'dog type' from patients in England293-6J Med Microbiol503Adult Animals Child Child, Preschool Cryptosporidium/*isolation & purification Dogs England Feces/*parasitology Female Heat-Shock Proteins 70/genetics Human Male RNA, Ribosomal, 18S/genetics Support, Non-U.S. Gov'tMar|DNA was extracted from faecal samples collected from 1680 patients in which Cryptosporidium oocysts were recognised by light microscopy. DNA from faeces from five of these patients failed to amplify by PCR three gene fragments--the Cryptosporidium oocyst wall protein (COWP) gene, the thrombospondin-related adhesive protein of Cryptosporidium-1 (TRAP-C1) gene and the thrombospondin-related adhesive protein of Cryptosporidium-2 (TRAP-C2) gene--with primers designed from C. parvum sequences. However, DNA from these five patients did amplify cryptosporidial 18S rDNA gene fragments and a heat-shock protein (HSP70) gene fragment was also amplified from four of them. The purpose of this study was to characterise further the Cryptosporidium associated with infection in these patients. DNA sequence analysis of 18S rDNA genes showed that four of these patients were infected by C. felis, and the remaining one by an as yet un-named Cryptosporidium species designated the 'dog type' (C. dt). Infection by C. felis was further confirmed in all four patients by DNA sequence analysis of the HSP70 gene. Oocysts present in all five samples reacted strongly with two anti-cryptosporidial oocyst monoclonal antibodies, except for the C. dt, which was tested with only one of the antibodies. Two of the patients infected by C. felis had underlying illness; one 8-year-old male had an undefined severe inherited underlying condition, and the second patient, a 32-year-old male, was HIV positive. Two of the remaining three patients (two females aged 1 and 2 years, respectively) were apparently immunocompetent (one infected with C. felis and one with the C. dt). No information was obtained for the fifth patient. The patient infected by C. dt had a recent history of travel to Africa. This is the first report of infection with these two Cryptosporidium species in immunocompetent patients, and in any patient in the UK.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=112327770022-2615 Journal Article11232777WDivision of Gastrointestinal Infections, PHLS Central Public Health Laboratory, London.D~?I>Eberhard, M. L. da Silva, A. J. Lilley, B. G. Pieniazek, N. J.1999Morphologic and molecular characterization of new Cyclospora species from Ethiopian monkeys: C. cercopitheci sp.n., C. colobi sp.n., and C. papionis sp.n651-8Emerg Infect Dis55Animals Base Sequence Cercopithecus aethiops/*parasitology Coccidia/*classification/*isolation & purification/pathogenicity Colobus/*parasitology Feces/parasitology Human Molecular Sequence Data Papio/*parasitology PhylogenySep-OctIn recent years, human cyclosporiasis has emerged as an important infection, with large outbreaks in the United States and Canada. Understanding the biology and epidemiology of Cyclospora has been difficult and slow and has been complicated by not knowing the pathogen s origins, animal reservoirs (if any), and relationship to other coccidian parasites. This report provides morphologic and molecular characterization of three parasites isolated from primates and names each isolate: Cyclospora cercopitheci sp.n. for a species recovered from green monkeys, C. colobi sp.n. for a parasite from colobus monkeys, and C. papionis sp.n. for a species infecting baboons. These species, plus C. cayetanensis, which infects humans, increase to four the recognized species of Cyclospora infecting primates. These four species group homogeneously as a single branch intermediate between avian and mammalian Eimeria. Results of our analysis contribute toward clarification of the taxonomic position of Cyclospora and its relationship to other coccidian parasites.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1051152111080-6040 Journal Article Review Review, Tutorial10511521zDivision of Parasitic Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 30341-3724, USA. mle1@cdc.gov:~?J~Pieniazek, N. J. Bornay-Llinares, F. J. Slemenda, S. B. da Silva, A. J. Moura, I. N. Arrowood, M. J. Ditrich, O. Addiss, D. G.19995New cryptosporidium genotypes in HIV-infected persons444-9Emerg Infect Dis53AIDS-Related Opportunistic Infections/*parasitology Animals Base Sequence Cat Diseases/parasitology Cats Cattle Cryptosporidiosis/*parasitology/veterinary Cryptosporidium/*classification/*genetics/isolation & purification Cryptosporidium parvum/genetics/isolation & purification DNA, Protozoan/analysis Dog Diseases/parasitology Dogs Feces/parasitology Genotype Human Molecular Sequence Data Phylogeny Polymerase Chain Reaction/methods RNA, Protozoan/analysis Sequence Analysis, DNAMay-JunsUsing DNA sequencing and phylogenetic analysis, we identified four distinct Cryptosporidium genotypes in HIV-infected patients: genotype 1 (human), genotype 2 (bovine) Cryptosporidium parvum, a genotype identical to C. felis, and one identical to a Cryptosporidium sp. isolate from a dog. This is the first identification of human infection with the latter two genotypes.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=103411841080-6040 Journal Article10341184UCenters for Disease Control and Prevention, Atlanta, Georgia 30333, USA. nxp3@cdc.gov~?K Pieniazek, N. J. Herwaldt, B. L.1997`Reevaluating the molecular taxonomy: is human-associated Cyclospora a mammalian Eimeria species?381-3Emerg Infect Dis33Animals Coccidiosis/*parasitology Diarrhea/parasitology Eimeria/*classification/*genetics/pathogenicity Eucoccidiida/*classification/*genetics/pathogenicity Human Mammals Molecular Sequence Data Phylogeny RNA, Protozoan/genetics RNA, Ribosomal/geneticsJul-Sep Human-associated Cyclospora is a coccidian parasite that causes diarrheal disease. A reevaluation of the parasite's molecular taxonomy that takes into account newly published data for seven Eimeria species shows that Cyclospora belongs to the Eimeria clade (Eimeriidae family). The Cyclospora branch on the phylogenetic tree is between the branches of the eight avian and two mammalian Eimeria species that have been evaluated to date. Furthermore, preliminary results indicate that Cyclospora and Isospora belli, another coccidian parasite that causes diarrheal disease in humans, belong to different families. To improve our understanding of the taxonomy of human-associated Cyclospora, molecular evaluation of isolates of additional Cyclospora and Eimeria species is needed.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=92843871080-6040 Journal Article9284387BCenters for Disease Control and Prevention, Atlanta, Georgia, USA.W~?LNPieniazek, N. J. Slemenda, S. B. da Silva, A. J. Alfano, E. M. Arrowood, M. J.1996:PCR confirmation of infection with Cyclospora cayetanensis357-9Emerg Infect Dis24Animals Coccidiosis/*diagnosis/epidemiology DNA Probes Diarrhea/*diagnosis/*parasitology Disease Outbreaks Eucoccidiida/genetics/*isolation & purification Feces/parasitology Food Parasitology Human Polymerase Chain Reaction/methods Sensitivity and SpecificityOct-Decdhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=89692551080-6040 Letter8969255 |~?M?Sturdee, A. P. Bodley-Tickell, A. T. Archer, A. Chalmers, R. M.2003ULong-term study of Cryptosporidium prevalence on a lowland farm in the United Kingdom97-113 Vet Parasitol1162SAnimals Animals, Domestic/*parasitology Animals, Wild/parasitology Cryptosporidiosis/epidemiology/*veterinary Cryptosporidium/*isolation & purification Cryptosporidium parvum/isolation & purification Feces/*parasitology Female Great Britain/epidemiology Male Oocysts Parasite Egg Count/veterinary Prevalence Seasons Support, Non-U.S. Gov'tOct 8% A longitudinal sample survey testing for Cryptosporidium in livestock and small wild mammals conducted over 6 years (1992-1997) on a lowland farm in Warwickshire, England, has shown the parasite to be endemic and persistently present in all mammalian categories. Faecal samples were taken throughout the year and oocysts concentrated by a formal ether sedimentation method for detection by immunofluorescence staining using a commercially available genus specific monoclonal antibody. Cryptosporidium parvum was identified by morphology and measurement of modified Ziehl-Neelsen stained oocysts. C. muris was rarely found in wild mammals and C. andersoni oocysts were never detected in livestock. Faecal samples from 3721 individuals gave cumulative 6-year prevalences for C. parvum as follows: bull beef, 3.6%; dairy cows, 3.5%; ewes, 6.4%; horses, 8.9%; calves (home bred), 52%; calves (bought-in) 23.2%; lambs, 12.9%; small wild mammals (rodents) living in and around farm buildings, 32.8%; small wild mammals (mainly rodents) living in areas of pasture, 29.9%. Animal categories with the highest prevalences also shed the highest average oocyst numbers per gram of faeces (ranging from 1.4 x 10(3) for bull beef to 1.1 x 10(5) for calves). Analysis of annual and seasonal data for each animal category revealed that patterns of infection were variable and sporadic; this means that short-term sampling was never likely to provide a true or representative picture. Seasonally combined data for adult livestock, young livestock and small wild mammals showed all three categories tended to have the highest Cryptosporidium prevalences in the autumn. Calves were separated from their dams within 24h of birth and yet showed high prevalence of infection in most years despite the low prevalence for the dairy herd. It is possible the coincidence of high autumn prevalence in mice with the main period for the rearing of calves contributed to the infection of the latter. The farming estate was used to teach students of agriculture and took pride in good land management and husbandry practices that produced well fed and healthy livestock. The data from this estate may represent, therefore, the baseline, the lowest possible levels to be expected, for Cryptosporidium infection and oocyst production on a lowland farm in the United Kingdom.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=145193150304-4017 Journal Article14519315Cell and Molecular Biosciences, School of Science and Environment, Coventry University, Priory Street, Coventry CV1 5FB, UK. t.sturdee@coventry.ac.uk~?N*Sturdee, A. P. Chalmers, R. M. Bull, S. A.1999HDetection of Cryptosporidium oocysts in wild mammals of mainland Britain273-80 Vet Parasitol804BAnimals Animals, Wild/*parasitology Antibodies, Monoclonal/chemistry Cryptosporidiosis/epidemiology/*veterinary Cryptosporidium/*isolation & purification Cryptosporidium parvum/*isolation & purification Feces/parasitology Great Britain/epidemiology Mammals/*parasitology Prevalence Rural Population Support, Non-U.S. Gov'tJan 28This paper combines the results from a preliminary survey of occurrence of Cryptosporidium species in faecal samples from a range of wild mammal species inhabiting mainland Britain with a tabulated literature review of world-wide reports of the parasite in those British mammals. In the literature, C. parvum was reported from 11 wild mammals found in Britain and elsewhere, mainly in rodents but also in insectivores, lagomorphs and ungulates. C. muris has been reported only in wild rodents. The sample survey detected C. parvum in seven additional British species, including carnivores. Overall, 12% of 184 faecal samples tested with a genus-specific monoclonal antibody contained oocysts of C. parvum. The results further emphasise the widespread distribution of Cryptosporidium amongst wild mammals in Britain, highlight the potential for transmission between host species and warn of the possibility of direct exposure for anybody using the countryside for professional or recreational purposes (e.g. farmers and ramblers) to previously unregarded sources of infection. It seems increasingly likely that most, if not all, mammalian species can be infected with C. parvum.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=99503330304-4017 Journal Article9950333_School of Natural and Environmental Sciences, Coventry University, UK. t.sturdee@coventry.ac.uk@~?O;Bull, S. Chalmers, R. Sturdee, A. P. Curry, A. Kennaugh, J.1998cCross-reaction of an anti-Cryptosporidium monoclonal antibody with sporocysts of Monocystis species195-7 Vet Parasitol772-3 Animals Antibodies, Monoclonal/*immunology Antibodies, Protozoan/*immunology Apicomplexa/*immunology Cross Reactions Cryptosporidiosis/complications/diagnosis/immunology Cryptosporidium/*immunology Feces/parasitology Fluorescent Antibody Technique, Indirect/veterinary Intestinal Diseases, Parasitic/diagnosis/immunology/veterinary Microtinae/parasitology Protozoan Infections/complications/diagnosis/immunology Protozoan Infections, Animal Rodent Diseases/diagnosis/immunology Shrews/parasitology Support, Non-U.S. Gov'tJun 15lThe non-specific cross-reaction of a fluorescently labelled anti-Cryptosporidium monoclonal antibody was observed microscopically when testing faecal specimens from small mammals. The reactive particles were identified as sporocysts of the Gregarine family Monocystidae, and indicate that considerable care should be taken so that false positives are not recorded.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=97462900304-4017 Journal Article9746290SBiosciences, School of Natural and Environmental Sciences, Coventry University, UK.~?PWebster, J. P. Macdonald, D. W.1995LCryptosporidiosis reservoir in wild brown rats (Rattus norvegicus) in the UK207-9Epidemiol Infect1151Animals Cryptosporidiosis/*epidemiology/parasitology Cryptosporidium parvum/growth & development/*isolation & purification *Disease Reservoirs Female Great Britain/epidemiology Male Prevalence Rats/*parasitology Seasons Support, Non-U.S. Gov'tAug~Rats (n = 73) were trapped from nine rural farms around Oxfordshire and faeces were examined using the auramine-phenol and the Modified Ziehl-Neelsen techniques. Cryptosporidium parvum oocysts were detected in the faeces from 46 (63%) rats. This suggests that wild rats represent a risk to human and livestock health through the carriage and transmission of this zoonotic protozoan.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=76418340950-2688 Journal Article7641834,Department of Zoology, University of Oxford.W~?Q2Pereira, S. J. Ramirez, N. E. Xiao, L. Ward, L. A.2002KPathogenesis of human and bovine Cryptosporidium parvum in gnotobiotic pigs715-8 J Infect Dis1865Animals Animals, Newborn Cattle Cecum/parasitology Comparative Study Cryptosporidiosis/*parasitology Cryptosporidium parvum/*pathogenicity *Germ-Free Life Human Ileum/parasitology Intestine, Small/parasitology Lymphoid Tissue/parasitology Support, U.S. Gov't, Non-P.H.S. SwineSep 1To compare the pathogenesis of human genotype 1 (HuG1) and bovine genotype 2 (BoG2) Cryptosporidium parvum, neonatal gnotobiotic pigs were given 1-10 HuG1 or BoG2 oocysts. The prepatent and patent periods were significantly longer for HuG1 than for BoG2 C. parvum (prepatent, 8.6 vs. 5.6 days; patent, 16.6 vs. 10.3 days). BoG2-infected pigs developed significantly more severe disease than did HuG1-infected pigs. BoG2 parasites were seen microscopically throughout the intestines during the prepatent and patent periods. HuG1 parasites were only detected during the patent period in the ileum and colon but colonized the mucosal surface in significantly larger numbers than did BoG2. Moderate-to-severe villus/mucosal attenuation with lymphoid hyperplasia was seen throughout the intestines of BoG2-infected pigs, whereas lesions in HuG1-infected pigs were mild to moderate and restricted to the ileum and colon. These findings provide additional support for the hypothesis that human and bovine C. parvum genotypes may be separate species.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=121953620022-1899 Journal Article12195362Food Animal Health Research Program, Ohio Agricultural Research and Development Center, Department of Veterinary Preventive Medicine, College of Veterinary Medicine, Ohio State University, Wooster, Ohio 44691-4096, USA.~?RaXiao, L. Bern, C. Arrowood, M. Sulaiman, I. Zhou, L. Kawai, V. Vivar, A. Lal, A. A. Gilman, R. H.2002EIdentification of the cryptosporidium pig genotype in a human patient1846-8 J Infect Dis18512CAdult Animals Cryptosporidiosis/*genetics Cryptosporidium/*genetics Feces/parasitology Genotype HIV Infections/*parasitology Homosexuality, Male Human Male Molecular Sequence Data Peru Polymerase Chain Reaction Polymorphism, Restriction Fragment Length Support, U.S. Gov't, P.H.S. Swine Swine Diseases/genetics/parasitologyJun 15ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12085341%0022-1899 Case Reports Comment Letter12085341X~?SfXiao, L. Bern, C. Limor, J. Sulaiman, I. Roberts, J. Checkley, W. Cabrera, L. Gilman, R. H. Lal, A. A.2001PIdentification of 5 types of Cryptosporidium parasites in children in Lima, Peru492-7 J Infect Dis1833Animals Cattle Child Child, Preschool Cryptosporidiosis/epidemiology/*parasitology Cryptosporidium parvum/*classification/*genetics/isolation & purification DNA, Protozoan/analysis/genetics Diarrhea/parasitology Dogs Feces/parasitology Genotype Human Infant Peru/epidemiology Polymerase Chain Reaction Polymorphism, Restriction Fragment Length Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.Feb 1Cryptosporidium parvum is usually considered to be the pathogen responsible for human cryptosporidiosis. We genotyped Cryptosporidium in 132 stool specimens from 80 Peruvian children, representing 85 infection episodes, using techniques that differentiate Cryptosporidium species and C. parvum genotypes. Five types of Cryptosporidium were identified: C. parvum human (67), bovine (8), and dog (2) genotypes, C. meleagridis (7), and C. felis (1). Twenty-five (29%) of the 85 infection episodes were associated with diarrhea. There was no significant difference in age, antecedent stunting, percentage with diarrhea, or duration of diarrhea for episodes with human genotype, compared with those of zoonotic Cryptosporidium. Duration of oocyst shedding was longer for human genotype than for zoonotic Cryptosporidium (mean, 13.9 days and 6.4 days, respectively; P=.004). Serum samples from 8 children with C. meleagridis, C. felis, or C. parvum dog genotype were tested for anti-human immunodeficiency virus (HIV) type 1 antibodies; all were found to be negative. Contrary to common belief, novel Cryptosporidium species and C. parvum genotypes can infect HIV-negative children.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=111333820022-1899 Journal Article11133382Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, GA 30341-3717, USA. lax0@cdc.gov~?TzQuiroz, E. S. Bern, C. MacArthur, J. R. Xiao, L. Fletcher, M. Arrowood, M. J. Shay, D. K. Levy, M. E. Glass, R. I. Lal, A.20008An outbreak of cryptosporidiosis linked to a foodhandler695-700 J Infect Dis1812{Adolescent Adult Animals Case-Control Studies Cryptosporidiosis/*epidemiology/parasitology Cryptosporidium parvum/*isolation & purification Diarrhea/parasitology *Disease Outbreaks District of Columbia/epidemiology Feces/microbiology *Food Handling *Food Microbiology Food Poisoning/epidemiology/parasitology Human Questionnaires Students Support, U.S. Gov't, P.H.S. UniversitiesFebIn September and October 1998, a cryptosporidiosis outbreak occurred on a Washington, DC, university campus. In a case-control study of 88 case patients and 67 control subjects, eating in 1 of 2 cafeterias was associated with diarrheal illness (P<.001). Morbidity was associated with eating dinner on 22 September (odds ratio, 8.1; 95% confidence interval, 3.4-19.5); weaker associations were found for 6 other meals. Cryptosporidium parvum was detected in stool specimens of 16 (70%) of 23 ill students and 2 of 4 ill employees. One ill foodhandler with laboratory-confirmed C. parvum prepared raw produce on 20-22 September. All 25 Cryptosporidium isolates submitted for DNA analysis, including 3 from the ill foodhandler, were genotype 1. This outbreak illustrates the potential for cryptosporidiosis to cause foodborne illness. Epidemiologic and molecular evidence indicate that an ill foodhandler was the likely outbreak source.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=106693570022-1899 Journal Article10669357Epidemic Intelligence Service, Epidemiology Program Office, and Respiratory and Enteric Viruses Branch, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, GA, 30341, USA.~?UXiao, L. Lal, R. B. Lal, A. A.1999Effect of immune activation induced by Cryptosporidium parvum whole antigen on in vitro human immunodeficiency virus type 1 infection559-63 J Infect Dis1802Animals Antigens, Protozoan/*immunology Cells, Cultured Cryptosporidium parvum/*immunology Cytokines/biosynthesis HIV Core Protein p24/*biosynthesis HIV-1/*physiology Human Leukocytes, Mononuclear/*immunology/*virology Lymphocyte Activation Tumor Necrosis Factor/biosynthesisAugPrevious epidemiologic investigations have suggested that persons with AIDS who are infected with Cryptosporidium parvum have a shorter survival time than those with other opportunistic infections. In this study, the effect of immune activation by a crude Cryptosporidium whole antigen on human immunodeficiency virus type 1 (HIV-1) infection was evaluated. Peripheral blood mononuclear cells from healthy persons without HIV-1 infection had increased proliferative and cytokine (interleukin-4, interferon-gamma, and tumor necrosis factor [TNF]-alpha) responses to stimulation with the crude Cryptosporidium whole antigen. This stimulation increased HIV-1 p24 antigen production in in vitro infection by>30-fold. A similar increase in p24 production was also seen when stimulation was done after cells were infected with HIV-1. Neutralization of TNF-alpha reduced Cryptosporidium antigen-induced p24 production by >50%. Results of this study suggest that Cryptosporidium-induced immune activation may be a cofactor in regulating HIV-1 production.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=103958830022-1899 Journal Article10395883wImmunology Branch, Division of Parasitic Diseases, Centers for Disease Control and Prevention, Chamblee, GA 30341, USA. ~?V9Xiao, L. Owen, S. M. Rudolph, D. L. Lal, R. B. Lal, A. A.1998Plasmodium falciparum antigen-induced human immunodeficiency virus type 1 replication is mediated through induction of tumor necrosis factor-alpha437-45 J Infect Dis1772Animals Antibodies, Blocking/immunology Antigens, CD14/immunology Antigens, Protozoan/*immunology CD8-Positive T-Lymphocytes/immunology Cell Division/immunology HIV Core Protein p24/analysis HIV Infections/complications/*immunology/metabolism HIV Long Terminal Repeat/immunology *Hiv-1 HLA-DR Antigens/immunology Human Interleukin-10/metabolism Interleukin-6/immunology/metabolism Leukocytes, Mononuclear/cytology/virology Lymphocyte Activation Plasmodium falciparum/*immunology Polymerase Chain Reaction RNA, Messenger/metabolism RNA, Viral/metabolism Receptors, CCR5/metabolism Receptors, CXCR4/metabolism Transcription, Genetic/immunology Tumor Necrosis Factor/immunology/*metabolism Virus Replication/*immunologyFebBecause malaria-stimulated cytokine production may have deleterious effects on human immunodeficiency virus type 1 (HIV-1) replication, the effects of Plasmodium falciparum antigens on HIV-1 replication were studied. Stimulation with malarial antigens significantly enhanced HIV-1 replication of HIV-1LAV and primary HIV-1 isolates (subtype A) in CD8-depleted peripheral blood mononuclear cells from naive donors. The malarial antigen-induced activation of HIV-1 was due to cellular activation as judged by the expression of cell activation markers and proliferative responses. While malarial antigen stimulation increased expression of tumor necrosis factor (TNF-alpha) and interleukin-6 (IL-6), only monoclonal antibodies (MAbs) to TNF-alpha inhibited malarial antigen-induced HIV-1 replication, whereas MAb to IL-6 had no effect. Malarial antigen increased HIV-1 replication by increasing viral mRNA expression and by activating long terminal repeat-directed viral transcription. These data suggest that P. falciparum infection can modulate HIV-1 pathogenesis by activating lymphocytes and stimulating viral replication through the production of cytokines.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=94665330022-1899 Journal Article9466533Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia, USA.~?Z<Bajer, A. Behnke, J. M. Bednarska, M. Kanicka, M. Sinski, E.2000QThe common vole (Microtus arvalis) as a competent host for Cryptosporidium parvum178Acta Parasitologica453[64002] Invertebrata: comparative, experimental morphology, physiology and pathology - Protozoa [60502] Parasitology - General [00520] General biology - Symposia, transactions and proceedings Parasitology [86310] Cricetidae [35400] Sporozoa [86310] Cricetidae, Rodentia, Mammalia, Vertebrata, Chordata, Animalia [35400] Sporozoa, Protozoa, Invertebrata, Animalia Cricetidae: Animals, Chordates, Mammals, Nonhuman Vertebrates, Nonhuman Mammals, Rodents, Vertebrates; Sporozoa: Animals, Invertebrates, Microorganisms, Protozoans Microtus arvalis: parasite host [Cricetidae]; Cryptosporidium parvum: parasite [Sporozoa] Poland: Europe, Palearctic region Meeting AbstractJuly[Reprint author] [Author] Poland Meeting. Meeting Abstract. 1230-2821 VIII European Muticolloquium of Parasitology, Poznan, Poland; September 10-14, 2000. Witold Stefanski Institute of Parasitology English'PREV200000434591 Copyright BIOSIS 2003.)University of Warszawa, Warszawa, Poland.~?[@Bajer, Anna Behnke, Jerzy M. Pawelczyk, Agnieszka Sinski, Edward1999CFirst evidence of Ehrlichia sp in wild Microtus arvalis from Poland204-205Acta Parasitologica443[07502] Ecology: environmental biology - General and methods [36001] Medical and clinical microbiology - General and methods [37057] Public health: disease vectors - General [64060] Invertebrata: comparative, experimental morphology, physiology and pathology - Arthropoda: chelicerata Infection; Epidemiology: Population Studies [75403] Acarina [86310] Cricetidae [86375] Muridae [07113] Rickettsiaceae [75403] Acarina, Chelicerata, Arthropoda, Invertebrata, Animalia [86310] Cricetidae, Rodentia, Mammalia, Vertebrata, Chordata, Animalia [86375] Muridae, Rodentia, Mammalia, Vertebrata, Chordata, Animalia [07113] Rickettsiaceae, Rickettsiales, Rickettsias and Chlamydias, Eubacteria, Bacteria, Microorganisms Acarina: Animals, Arthropods, Chelicerates, Invertebrates; Cricetidae: Animals, Chordates, Mammals, Nonhuman Vertebrates, Nonhuman Mammals, Rodents, Vertebrates; Muridae: Animals, Chordates, Mammals, Nonhuman Vertebrates, Nonhuman Mammals, Rodents, Vertebrates; Rickettsiaceae: Bacteria, Eubacteria, Microorganisms Ixodes ricinus: vector [Acarina]; Clethrionomys glareolus: host [Cricetidae]; Microtus arvalis: host [Cricetidae]; Apodemus flavicollis: host [Muridae]; Ehrlichia sp.: pathogen [Rickettsiaceae]JulyKWe examined a total of 73 Microtus arvalis, 168 Clethrionomys glareolus and 17 Apodemus flavicollis trapped in the Mazury Lakes district of North-Eastern Poland, in the spring, summer and autumn of 1998. Three M. arvalis, (2 in summer and 1 in autumn) carried Ehrlichia sp. (overall prevalence = 4.1%), whereas infection was not detected in the other rodent species. We hypothesize that Ixodes ricinus (the most common tick in the region) with which the animals were heavily infested, constitutes the likely natural vector for this pathogen and that M. arvalis are its natural reservoir.2[Author] [Reprint author] Poland 1230-2821 English'PREV200000069014 Copyright BIOSIS 2003.Department of Parasitology, Institute of Zoology, University of Warszawa, Krakowskie Przedmiescie 26/28, 00-927, Warszawa, Poland.~?\0Bajer, Anna Bednarska, Malgorzata Sinski, Edward1997RWildlife rodents from different habitats as a reservoir for Cryptosporidium parvum192-194Acta Parasitologica424[37003] Public health - Epizootiology [07508] Ecology: environmental biology - Animal [16501] Reproductive system - General and methods [37058] Public health: disease vectors - Animate [60502] Parasitology - General [64002] Invertebrata: comparative, experimental morphology, physiology and pathology - Protozoa Parasitology; Vector Biology [86295] Castoridae [86310] Cricetidae [86375] Muridae [35400] Sporozoa [86295] Castoridae, Rodentia, Mammalia, Vertebrata, Chordata, Animalia [86310] Cricetidae, Rodentia, Mammalia, Vertebrata, Chordata, Animalia [86375] Muridae, Rodentia, Mammalia, Vertebrata, Chordata, Animalia [35400] Sporozoa, Protozoa, Invertebrata, Animalia Castoridae: Animals, Chordates, Mammals, Nonhuman Vertebrates, Nonhuman Mammals, Rodents, Vertebrates; Cricetidae: Animals, Chordates, Mammals, Nonhuman Vertebrates, Nonhuman Mammals, Rodents, Vertebrates; Muridae: Animals, Chordates, Mammals, Nonhuman Vertebrates, Nonhuman Mammals, Rodents, Vertebrates; Sporozoa: Animals, Invertebrates, Microorganisms, Protozoans Castor-fiber: disease vector, parasite host [Castoridae]; Clethrionomys-glareolus: disease vector, parasite host [Cricetidae]; Microtus-arvalis: disease vector, parasite host [Cricetidae]; Microtus-oeconomus: disease vector, parasite host [Cricetidae]; Ondatra-zibethicus: disease vector, parasite host [Cricetidae]; Apodemus-agrarius: disease vector, parasite host [Muridae]; Apodemus-flavicollis: disease vector, parasite host [Muridae]; Apodemus-sylvaticus: disease vector, parasite host [Muridae]; Cryptosporidium-parvum: oocyst, parasite [Sporozoa] cryptosporidiosis: (Cryptosporidiosis (MeSH)), digestive system disease, parasitic disease Poland: Europe, Palearctic regionOct.pA variety of domestic and wild animals are considered to be potential sources of cryptosporidiosis in humans. However, despite considerable information available on Cryptosporidium spp. in humans and ruminants, a paucity of information exists for many species of wildlife rodents. The current study reports the prevalence of Cryptosporidium infection among rodent populations from different habitats in northern Poland. In autumn 1996, spring and summer 1997, a total of 287 animals (8 species of rodents from three different habitats: Clethrionomys glareolus and Apodemus flavicollis and A. sylvaticus from forest; A. agrarius, Microtus arvalis and M. oeconomus from field/meadow; Castor fiber and Ondatra zibethicus from semi-aquatic habitat) were examined for Cryptosporidium oocysts. Direct wet smears from unconcentrated faecal specimens were stained using a modified Ziehl-Neelsen method and/or direct, immunofluorescent (MerIFluor Cryptosporidium/Giardia) assay. Faecal examination revealed that 41 of 132 (31%) C. glareolus, 26 of 96 (27%) Apodemus sp., 19 of 28 (68%) Microtus sp., 1 A. agrarius, 2 of 19 (10.5% colonies of C fiber and 5 of 12 (42%) O. zibethicus were infected with Cryptosporidium oocysts. The intensity of infection was generally low, with <5 oocysts/slide film in most of the samples studied. The morphological characteristics and measurements of the oocysts (mean size 4.8 X 4.5 mum) conformed with those of C. parvum. This study indicates, for the first time in Poland, that semi-aquatic rodents, mostly O. zibethicus, and possibly rodents from other habitats may be involved as reservoirs of infection for C. parvum.2[Reprint author] [Author] Poland 1230-2821 English'PREV199800234564 Copyright BIOSIS 2003.FDep. Parasitol., Inst. Zool., Univ. Warszawa, 00-927 Warszawa, Poland. ~?`5Sinski, Edward Hlebowicz, Edyta Bednarska, Malgorzata1993fOccurrence of Cryptosporidium parvum infection in wild small mammals in District of Mazury Lake Poland59-61Acta Parasitologica3820[01056] Microscopy - Histology and histochemistry [11108] Anatomy and Histology - Microscopic and ultramicroscopic anatomy [37058] Public health: disease vectors - Animate [60502] Parasitology - General [62800] Animal distribution [64002] Invertebrata: comparative, experimental morphology, physiology and pathology - Protozoa Morphology; Parasitology; Physiology; Vector Biology [86310] Cricetidae [86375] Muridae [51300] Nematoda [00500] Organisms [35400] Sporozoa [86310] Cricetidae, Rodentia, Mammalia, Vertebrata, Chordata, Animalia [86375] Muridae, Rodentia, Mammalia, Vertebrata, Chordata, Animalia [51300] Nematoda, Aschelminthes, Helminthes, Invertebrata, Animalia [00500] Organisms, Organisms [35400] Sporozoa, Protozoa, Invertebrata, Animalia Cricetidae: Animals, Chordates, Mammals, Nonhuman Vertebrates, Nonhuman Mammals, Rodents, Vertebrates; Muridae: Animals, Chordates, Mammals, Nonhuman Vertebrates, Nonhuman Mammals, Rodents, Vertebrates; Nematoda: Animals, Aschelminths, Helminths, Invertebrates; Organisms: Organisms; Sporozoa: Animals, Invertebrates, Microorganisms, Protozoans Neotoma floridana [Cricetidae]; Muridae [Muridae]; Longistriata neotoma [Nematoda]; Trichuris muris [Nematoda]; Bohmiella wilsoni [Organisms]; Sporozoa [Sporozoa] USA: North America, Nearctic region parasitism; seasonal abundanceXThis study was conducted in population of wild small mammals in northern Poland, District of Mazury Lake, Urwitalt, near Mikolajki. Samples were collected four times, from autumn 1989 until spring 1991. Twenty percent (66 of 330) of the examined mammals were positive for Cryptosporidium, i.e., 55 of 275 Clethrionomys glareolus, 6 of 39 Apodemus flavicollis, 5 of 16 Sorex araneus were naturally infected with Cryptosporidium. There was no infection of Cryptosporidium assessed in A. agrarius and S. minutus. Our histological data clearly indicate that the population of C. glareolus was infected with C. parvum. This population of rodents appears to have maintained an infection throughout two years. These findings indicate that C. glareolus and possibly other rodents have potential to act as a reservoir for C. parvum, a pathogen of man and ruminants.![Author] Poland 1230-2821 English'PREV199396091139 Copyright BIOSIS 2003.FDep. Parasitol., Zool. Inst., Univ. Warszawa, 00-929 Warszawa, Poland. q~?a7Sinski, Edward Bednarska, Malgorzata Adamczewska, Agata1992dBiological characterization of Cryptosporidium parvum isolates in suckling and immunosuppressed mice139-143Acta Parasitologica373o[25508] Development and Embryology - Morphogenesis [34508] Immunology - Immunopathology, tissue immunology [35000] Immunology, parasitological [60502] Parasitology - General [64002] Invertebrata: comparative, experimental morphology, physiology and pathology - Protozoa Development; Immune System: Chemical Coordination and Homeostasis; Parasitology; Physiology [86375] Muridae [35400] Sporozoa [86375] Muridae, Rodentia, Mammalia, Vertebrata, Chordata, Animalia [35400] Sporozoa, Protozoa, Invertebrata, Animalia Muridae: Animals, Chordates, Mammals, Nonhuman Vertebrates, Nonhuman Mammals, Rodents, Vertebrates; Sporozoa: Animals, Invertebrates, Microorganisms, Protozoans Muridae [Muridae]; Cryptosporidium parvum [Sporozoa] prednisone: 53-03-2; cyclophosphamide: 50-18-0 age-dependent immunity; cyclophosphamide; endogenous parasite development; immunosuppressant; prednisoneSuckling mice and immunosuppressed adult mice were used as model hosts to compare the endogenous development of two isolates of C. parvum obtained from naturally infected calves in different seasons of the year. There were no differences in these two isolates as to their location, time of appearance, intensity of infection and antigenicity. Present study indicates that Cryptosporidium infection may be easily established in 5-7 day old outbred Swiss mice. The peak of infection is shown on day 8 after inoculation and shortly after that a self-limiting process of infection takes place. However, C. parvum does not develop endogenously in adult immunosuppressed mice. The lack of establishment of infection in these animals, even in latent state, indicates that age dependent mechanisms of immunity, developed in mice, are not impaired by prednisolone or cyclophosphamide. There was no difference in biological characteristics of two isolates C. parvum. Moreover, the present study illustrates that oocysts, spread by naturally infected calves in autumn and winter, are infective for suckling mice, which might be associated in transmissibility of C. parvum.![Author] Poland 1230-2821 English'PREV199395134500 Copyright BIOSIS 2003.FDep. Parasitol., Zool. Inst., Univ. Warszawa, 00-927 Warszawa, Poland.C~?bSinski, E. Czarnogrecka, M.1989&Cryptosporidium-sp infection in snakes307-310Acta Parasitologica Polonica344[07508] Ecology: environmental biology - Animal [14006] Digestive system - Pathology [60502] Parasitology - General [64002] Invertebrata: comparative, experimental morphology, physiology and pathology - Protozoa Digestive System: Ingestion and Assimilation; Ecology: Environmental Sciences; Parasitology; Physiology [35400] Sporozoa [85410] Serpentes [35400] Sporozoa, Protozoa, Invertebrata, Animalia [85410] Serpentes, Reptilia, Vertebrata, Chordata, Animalia Sporozoa: Animals, Invertebrates, Microorganisms, Protozoans; Serpentes: Animals, Chordates, Nonhuman Vertebrates, Reptiles, Vertebrates elaphe-obsoleta elaphe-quatuorlineata elaphe-schrencki elaphe-guttata cryptosporidium chronic gastritisQThis is the first record of Cryptosporidium sp. infection in snakes of genus Elaphe (E. obsoleta, E. quatuorlineata, E. shrencki, E. guttata) in Poland. Fecal smears from snakes contained spherical organisms, confirmed by specific method of Ziehl-Neelson to be oocyst of Cryptosporidium. The examined snakes had severe chronic gastritis.2[Reprint author] [Author] POLAND 0065-1478 English'PREV199090031302 Copyright BIOSIS 2003.ADep Parasitol, Zool Inst, Univ Warszawa, 00-927 Warszawa, Poland. ~?c1Guy, R. A. Payment, P. Krull, U. J. Horgen, P. A.2003iReal-time PCR for quantification of Giardia and Cryptosporidium in environmental water samples and sewage5178-85Appl Environ Microbiol699iAnimals Base Sequence Canada Cryptosporidium/genetics/growth & development/*isolation & purification DNA Primers DNA, Bacterial/genetics/isolation & purification Environment Fresh Water/microbiology Giardia/genetics/growth & development/*isolation & purification Polymerase Chain Reaction/methods Sewage/*microbiology Support, Non-U.S. Gov't *Water MicrobiologySepThe protozoan pathogens Giardia lamblia and Cryptosporidium parvum are major causes of waterborne enteric disease throughout the world. Improved detection methods that are very sensitive and rapid are urgently needed. This is especially the case for analysis of environmental water samples in which the densities of Giardia and Cryptosporidium are very low. Primers and TaqMan probes based on the beta-giardin gene of G. lamblia and the COWP gene of C. parvum were developed and used to detect DNA concentrations over a range of 7 orders of magnitude. It was possible to detect DNA to the equivalent of a single cyst of G. lamblia and one oocyst of C. parvum. A multiplex real-time PCR (qPCR) assay for simultaneous detection of G. lamblia and C. parvum resulted in comparable levels of detection. Comparison of DNA extraction methodologies to maximize DNA yield from cysts and oocysts determined that a combination of freeze-thaw, sonication, and purification using the DNeasy kit (Qiagen) provided a highly efficient method. Sampling of four environmental water bodies revealed variation in qPCR inhibitors in 2-liter concentrates. A methodology for dealing with qPCR inhibitors that involved the use of Chelex 100 and PVP 360 was developed. It was possible to detect and quantify G. lamblia in sewage using qPCR when applying the procedure for extraction of DNA from 1-liter sewage samples. Numbers obtained from the qPCR assay were comparable to those obtained with immunofluorescence microscopy. The qPCR analysis revealed both assemblage A and assemblage B genotypes of G. lamblia in the sewage. No Cryptosporidium was detected in these samples by either method.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=129578990099-2240 Journal Article12957899`University of Toronto at Mississauga, Mississauga, Ontario, Canada L5L 1C6. rguy@utm.utoronto.ca~?dKreader, C. A.1996YRelief of amplification inhibition in PCR with bovine serum albumin or T4 gene 32 protein1102-6Appl Environ Microbiol623Bacteriophage T4/*genetics Culture Media Membrane Proteins/*genetics Polymerase Chain Reaction *Serum Albumin, Bovine Support, U.S. Gov't, Non-P.H.S. Viral Proteins/*geneticsMar,The benefits of adding bovine serum albumin (BSA) or T4 gene 32 protein (gp32) to PCR were evaluated with reaction mixtures containing substances that inhibit amplification. Whereas 10- to 1,000-fold more FeCl3, hemin, fulvic acids, humic acids, tannic acids, or extracts from feces, freshwater, or marine water were accommodated in PCR when either 400 ng of BSA per microl or 150 ng of gp32 per microl was included in the reactions, neither BSA nor gp32 relieved interference significantly when minimum inhibitory levels of bile salts, bilirubin, EDTA, NaCl, sodium dodecyl sulfate, or Triton X-100 were present. Use of BSA and gp32 together offered no more relief of inhibition than either alone at its optimal level, and neither protein had any noticeable effect on amplification in the absence of inhibitors.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=89756030099-2240 Journal Article8975603Microbiology Research Division, Environmental Monitoring Systems Laboratory, U.S. Environmental Protection Agency, Cincinnati, Ohio 45268, USA.~?eRHiggins, J. A. Fayer, R. Trout, J. M. Xiao, L. Lal, A. A. Kerby, S. Jenkins, M. C.20019Real-time PCR for the detection of Cryptosporidium parvum323-37J Microbiol Methods473Animals Cattle Cloning, Molecular Cryptosporidiosis/diagnosis/parasitology/veterinary Cryptosporidium parvum/genetics/*isolation & purification DNA, Protozoan/genetics/isolation & purification Feces/parasitology Genotype Human Male Parasite Egg Count Polymerase Chain Reaction/*methods RNA, Ribosomal, 18S/chemistry/genetics Reagent Kits, Diagnostic Sensitivity and Specificity Sequence Homology, Nucleic Acid Support, Non-U.S. Gov'tDecReal time, TaqMan PCR assays were developed for the Cp11 and 18S rRNA genes of the protozoan parasite Cryptosporidium parvum. The TaqMan probes were specific for the genus Cryptosporidium, but could not hybridize exclusively with human-infectious C. parvum species and genotypes. In conjunction with development of the TaqMan assays, two commercial kits, the Mo Bio UltraClean Soil DNA kit, and the Qiagen QIAamp DNA Stool kit, were evaluated for DNA extraction from calf diarrhea and manure, and potassium dichromate and formalin preserved human feces. Real-time quantitation was achieved with the diarrhea samples, but nested PCR was necessary to detect C. parvum DNA in manure and human feces. Ileal tissues were obtained from calves at 3, 7, and 14 days post-infection, and DNA extracted and assayed. Nested PCR detected C. parvum DNA in the 7-day post-infection sample, but neither of the other time point samples were positive. These results indicate that real-time quantitation of C. parvum DNA, extracted using the commercial kits, is feasible on diarrheic feces, with large numbers of oocysts and small concentrations of PCR inhibitor(s). For samples with few oocysts and high concentrations of PCR inhibitor(s), such as manure, nested PCR is necessary for detection.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=117145230167-7012 Journal Article11714523kUSDA-ARS, Rm. 202, Bldg. 173, 10300 Baltimore Blvd., Beltsville, MD 20705, USA. jhiggins@anri.barc.usda.gov C~?fNJenkins, M. C. Trout, J. Abrahamsen, M. S. Lancto, C. A. Higgins, J. Fayer, R.2000Estimating viability of Cryptosporidium parvum oocysts using reverse transcriptase-polymerase chain reaction (RT-PCR) directed at mRNA encoding amyloglucosidase97-106J Microbiol Methods432'Animals Cell Survival Cryptosporidium parvum/*cytology Glucan 1,4-alpha-Glucosidase/genetics/isolation & purification Mice Mice, Inbred BALB C RNA, Messenger/isolation & purification RNA, Protozoan/isolation & purification *Reverse Transcriptase Polymerase Chain Reaction Support, Non-U.S. Gov'tDec 15The purpose of the present study was to determine if reverse transcriptase-polymerase chain reaction (RT-PCR) directed at mRNA encoding the enzyme amyloglucosidase (CPAG) could serve as a indicator for C. parvum oocyst viability. Oocysts were stored for 1-11 months in the refrigerator and at monthly intervals extracted for total RNA for RT-PCR analysis. An aliquot of these C. parvum oocysts was inoculated into neonatal mice which were necropsied 4 days later for ileal tissue that was analyzed by semi-quantitative PCR to determine the level of parasite replication. The CPAG RT-PCR assay detected RNA from as few as 10(3) C. parvum oocysts. An effect of storage time on both RT-PCR signal and mouse infectivity was observed. RNA from oocysts stored for 1-7 months, unlike oocysts stored for 9 or 11 months, contained CPAG mRNA that was detectable by RT-PCR. A gradual decrease in the RT-PCR signal intensity was observed between 5 and 7 months storage. The intensity of RT-PCR product from oocysts and the signal from semi-quantitative PCR of ileal tissue DNA from mice infected with these same aged oocysts were comparable. The RT-PCR assay of CPAG mRNA in cultured cells infected with viable C. parvum oocysts first detected expression at 12 h with highest expression levels observed at 48 h post-infection. These results indicate that CPAG RT-PCR may be useful for differentiating viable from non-viable C. parvum oocysts and for studying the expression of the gene for amyloglucosidase in vitro.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=111216080167-7012 Journal Article11121608mImmunology and Disease Resistance Laboratory, Agricultural Research Service, USDA, Beltsville, MD 20705, USA. R~?g)Deng, M. Q. Peterson, R. P. Cliver, D. O.2000^First findings of Cryptosporidium and Giardia in California sea lions (Zalophus californianus)490-4 J Parasitol863Animals Base Sequence California/epidemiology Cryptosporidiosis/epidemiology/parasitology/*veterinary Cryptosporidium/genetics/*isolation & purification DNA, Protozoan/chemistry Feces/parasitology Fluorescent Antibody Technique, Direct/veterinary Fluorescent Antibody Technique, Indirect/veterinary Giardia/genetics/*isolation & purification Giardiasis/epidemiology/parasitology/*veterinary Immunomagnetic Separation/veterinary Molecular Sequence Data Polymerase Chain Reaction/veterinary Seals/*parasitology Sequence Alignment/veterinaryJunYWe report the detection and identification of Cryptosporidium and Giardia from 1 of 3 species of pinnipeds. Fecal samples were collected from Pacific harbor seal (Phoca vitulina richardsi), northern elephant seal (Mirounga angustirostris), and California sea lion (Zalophus californianus) in the northern California coastal area. By means of fluorescently labeled monoclonal antibodies, Cryptosporidium oocysts were detected in 3 samples from California sea lions, 1 of which also contained Giardia cysts. Oocysts of Cryptosporidium and cysts of Giardia were morphologically indistinguishable from oocysts of C. parvum and cysts of G. duodenalis from other animal origins. Oocysts and cysts were then purified using immunomagnetic separation techniques and identified by polymerase chain reaction (PCR), from which species-specific products were obtained. Sequence analysis revealed that the 452-bp and 358-bp PCR products of Cryptosporidium isolated from California sea lion had identities of 98% with sequences of their template fragments of C. parvum obtained from infected calves. Based on morphological, immunological, and genetic characterization, the isolates were identified as C. parvum and G. duodenalis, respectively. The findings suggested that California sea lions could serve as reservoirs in the environmental transmission of Cryptosporidium and Giardia.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=108642440022-3395 Journal Article10864244Department of Population Health and Reproduction, School of Veterinary Medicine, Univerity of California at Davis, 95616-8743, USA.n~?hDeng, M. Q. Cliver, D. O.1998:Cryptosporidium parvum development in the BS-C-1 cell line8-15 J Parasitol841Animals Cell Line Cercopithecus aethiops Cryptosporidium parvum/genetics/*growth & development/immunology DNA, Protozoan/analysis Fluorescent Antibody Technique, Indirect Immune Sera/immunology Polymerase Chain Reaction Species Specificity Support, Non-U.S. Gov'tFebCryptosporidium parvum is a worldwide parasitic protozoon capable of causing life-threatening disease in immunocompromised patients. In vitro cultivation of C. parvum has been under investigation for development of a well-defined in vitro model for C. parvum infectivity assay. This is the first report of C. parvum completing its life cycle in BS-C-1, an African green monkey kidney cell line. Both sodium hypochlorite-stimulated oocysts and purified sporozoites were able to initiate infection that led to completion of the whole life cycle, although inoculating purified sporozoites was less efficient. Beside Giemsa staining and normal light microscopy, an indirect fluorescent antibody staining method was developed to facilitate the detection and interpretation of C. parvum life stages developed in vitro. A C. parvum-specific polymerase chain reaction was also applied to detect and confirm the presence of C. parvum life stages in liquid from oocyst-infected cell cultures. In addition to the 452-base-pair (bp) product that can be specifically amplified from C. parvum oocysts and sporozoites, a fragment near 280 bp was also obtained. Various applications of this in vitro culture system are envisioned.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=94883300022-3395 Journal Article9488330Department of Population Health and Reproduction, School of Veterinary Medicine, University of California at Davis, 95616-8743, USA.~?i^Enemark, H. L. Ahrens, P. Bille-Hansen, V. Heegaard, P. M. Vigre, H. Thamsborg, S. M. Lind, P.2003OCryptosporidium parvum: infectivity and pathogenicity of the 'porcine' genotype407-16 Parasitology126Pt 5Animals Autopsy Cattle Cattle Diseases/parasitology Cryptosporidiosis/complications/*parasitology/*veterinary Cryptosporidium parvum/classification/*genetics/*pathogenicity/physiology Diarrhea/complications/microbiology Eating Feces/microbiology/virology Genotype Haptoglobins/analysis Human Molecular Sequence Data Species Specificity Support, Non-U.S. Gov't Swine/microbiology/*parasitology/virology Swine Diseases/microbiology/parasitology/physiopathology/virology Virulence Vomiting/complications/microbiologyMayGenetic studies have demonstrated profound differences between the 'porcine' genotype of Cryptosporidium parvum, versus 'human' and 'bovine' genotypes. The study analysed infectivity and pathogenicity of the 'porcine' genotype (CPP-13 isolate) of C. parvum, and compared the results with published data on the 'bovine' genotype (CPB-0 isolate). This was investigated in calves and piglets from commercial herds. Piglets were mildly affected by the CPP-13 isolate, contrary to piglets infected with the CPB-0 isolate, which caused diarrhoea of a mean duration of 3.5 days. CPP-13 produced no or very mild clinical signs in piglets despite the excretion of high numbers of oocysts. Concomitant infection with rotavirus, however, caused a dramatic aggravation of the clinical signs, and 5 of 6 experimentally infected piglets died. CPP-13 appeared to be adapted to porcine hosts as illustrated by the lack of infectivity to 1 experimentally inoculated calf, and the absence of clinical signs, the long pre-patent period (15 days), and the excretion of very low numbers of oocysts following experimental infection of another calf. Thus, in accordance with other molecular studies, our results support the genetic evidence for the existence of a new species of Cryptosporidium adapted to pigs.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=127936440031-1820 Journal Article12793644XDanish Veterinary Institute, Bulowsvej 27, DK-1790 Copenhagen V, Denmark. hle@vetinst.dk R~?jlEnemark, H. L. Ahrens, P. Juel, C. D. Petersen, E. Petersen, R. F. Andersen, J. S. Lind, P. Thamsborg, S. M.2002DMolecular characterization of Danish Cryptosporidium parvum isolates331-41 Parasitology125Pt 4qAnimals Base Sequence Cattle Cryptosporidium parvum/*classification/*genetics/isolation & purification Denmark Genes, Protozoan/genetics Genotype Human Microsatellite Repeats/genetics Molecular Sequence Data Polymerase Chain Reaction Polymorphism (Genetics) Protozoan Proteins/genetics RNA, Ribosomal, 18S/genetics Support, Non-U.S. Gov't Variation (Genetics)/*geneticsOctThe genetic polymorphism among 271 Danish Cryptosporidium isolates of human and animal origin was studied by partial amplification and sequencing of the Cryptosporidium oocyst wall protein (COWP) gene, the 1 8S rDNA, and a microsatellite locus. Furthermore, the microsatellite locus was studied directly using fragment analysis. A comparative analysis of DNA sequences showed the presence of 3 different subgenotypes (Cl, C2 and C3) in C. parvum isolates from Danish cattle, with prevalences of 16.7, 17.2 and 73.1% including 13 (7.0%) mixed infections. Subgenotype Cl was significantly more prevalent (P < 0.001) in the southern part of Denmark. In Cryptosporidium isolates of human origin the anthroponotic subgenotype H1 was identified, in addition to the zoonotic subgenotypes C1, C2, and C3. Of 44 human samples, 56.8% were anthroponotic, whereas 40.9% were zoonotic genotypes. One human isolate was characterized as C. meleagridis. The porcine Cryptosporidium isolates (N = 4) revealed a pattern which was genetically distinct from human and bovine isolates. Cryptosporidium in a hedgehog (Erinaceus europaeus L.) was identified for the first time. By microsatellite sequencing the hedgehog isolate showed a subgenotype distinct from the previously detected types. The assignment to subgenotype by microsatellite sequencing and fragment typing was 100% identical in samples where results were achieved by both methods. In addition, the fragment analysis proved more sensitive, easier, faster, and less expensive compared to sequencing.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=124033210031-1820 Journal Article12403321QSection for Parasitology, Danish Veterinary Institute, Copenhagen. hle@vetinst.dk~?l]Wallis, P. M. Erlandsen, S. L. Isaacrenton, J. L. Olson, M. E. Robertson, W. J. Vankeulen, H.1996Prevalence of Giardia Cysts and Cryptosporidium Oocysts and Characterization of Giardia Spp Isolated from Drinking Water in Canada 2789-2797$Applied & Environmental Microbiology628Animal infectivity. Excystation. Duodenalis. Supplies. Sequence. Gerbils. Microbiology Biology. Reprint available from: Wallis PM. HYPER RES LTD, 350-12 ST SW, MEDICINE HAT, AB T1A 4T7.A This study was carried out to estimate the prevalence and potential for human infectivity of Giardia cysts in Canadian drinking water supplies. The presence of Cryptosporidium oocysts was also noted, but isolates were not collected for further study, A total of 1,760 raw water samples, treated water samples, and raw sewage samples were collected from 72 municipalities across Canada for analysis, 58 of which treat their water by chlorination alone, Giardia cysts were found in 73% of raw sewage samples, 21% of raw water samples, and 18.2% of treated water samples, There was a trend to higher concentration and more frequent incidence of Giardia cysts in the spring and fall, but positive samples were found in all seasons, Cryptosporidium oocysts were found in 6.1% of raw sewage samples, 4.5% of raw water samples, and 3.5% of treated water samples, Giardia cyst viability was assessed by infecting Mongolian gerbils (Meriones unguiculatus) and by use of a modified propidium iodide dye exclusion test, and the results were not always in agreement, No Cryptosporidium isolates were recovered from gerbils, but 8 of 276 (3%) water samples and 19 of 113 (17%) sewage samples resulted in positive Giardia infections, Most of the water samples contained a low number of cysts, and 12 Giardia isolates were successfully recovered from gerbils and cultured, Biotyping of these isolates by isoenzyme analysis and karyotyping by pulsed-field gel electrophoresis separated the isolates into the same three discrete groups, Karyotyping revealed four or five chromosomal bands ranging in size from 0.9 to 2 Mb, and four of the isolates had the same banding pattern as that of the WB strain, Analysis of the nucleotide sequences of the 16S DNA coding for rRNA divided the isolates into two distinct groups corresponding to the Polish and Belgian designations found by other investigators, The occurrence of these biotypes and karyotypes appeared to be random and was not related to geographic or other factors (e.g., different types were found in both drinking water and sewage from the same community), Biotyping and karyotyping showed that isolates from this study were genetically and biochemically similar to those found elsewhere, including well-described human source strains such as WB. We conclude that potentially human-infective Giardia cysts are commonly found in raw surface waters and sewage in Canada, although cyst viability is frequently low. Cryptosporidium oocysts are less common in Canada, An action level of three to five Giardia cysts per 100 liters in treated drinking water is proposed on the basis of the monitoring data from outbreak situations, This action level is lower than that proposed by Haas and Rose (C. N. Haas and J. B. Rose, J. Am. Water Works Assoc, 87(9):81-84, 1995) for Clyptosporidium spp, (10 to 30 oocysts per 100 liters). [References: 25] 25"English Article Current Contents/Life Sciences Current Contents/Agriculture, Biology & Environmental Sciences. Reprint available from: Wallis PM HYPER RES LTD 350-12 ST SW MEDICINE HAT AB T1A 4T7 CANADA UNIV MINNESOTA SCH MED DEPT CELL BIOL & NEUROANAT MINNEAPOLIS, MN 55455 USA UNIV BRITISH COLUMBIA DEPT PATHOL DIV MED MICROBIOL VANCOUVER BC V5Z 1M9 CANADA UNIV CALGARY FAC MED ANIM RESOURCE CTR CALGARY AB T2N 1N4 CANADA HLTH CANADA ENVIRONM HLTH DIRECTORATE OTTAWA ON K1A 0L2 CANADA CLEVELAND STATE UNIV DEPT BIOL CLEVELAND, OH 44115 USA 0017-UZ996-0017 UZ996: Document Delivery available~?m]Olson, M. E. McAllister, T. A. Deselliers, L. Morck, D. W. Cheng, K. J. Buret, A. G. Ceri, H.1995GEffects of Giardiasis on Production in a Domestic Ruminant (Lamb) Model 1470-1474'American Journal of Veterinary Research5611Small-intestine. Cryptosporidium. Infection. Animals. Growth. Veterinary medicine/animal health. Reprint available from: Olson ME. UNIV CALGARY, GASTROINTESTINAL RES GRP, CALGARY, AB T2N 4N1.Objective-To examine the effects of giardiasis on production and carcass quality, using growing lambs as a domestic ruminant model. Design-Randomized block. Animals-Giardia-free lambs: 23 in infected group, 24 in control group. Procedure-Six-week-old, specific-pathogen-free lambs were infected with Giardia trophozoites; control lambs received saline solution. Clinical signs of infection, body weight, and feed intake were determined for 10 weeks. Carcass weight and quality were determined at slaughter weight of 45 kg. Results-Giardia infection persisted from weeks 7 to 16. For 5 weeks after challenge exposure, abnormal feces were more frequently observed in infected lambs. Giardia infection was associated with a decrease in rate of weight gain and impairment in feed efficiency. Time to reach slaughter weight was extended in infected lambs, and the carcass weight of Giardia-infected lambs was lower than that of control lambs. Conclusion-Giardiasis has a negative effect on domestic ruminant production. Clinical Relevance-Giardiasis in domestic ruminants is an economically important disease, thus necessitating control or elimination of the infection. [References: 31] 31sEnglish Article Current Contents/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME UNIV CALGARY GASTROINTESTINAL RES GRP CALGARY AB T2N 4N1 CANADA UNIV CALGARY DEPT MICROBIOL & INFECT DIS CALGARY AB T2N 4N1 CANADA UNIV CALGARY DEPT SCI BIOL CALGARY AB T2N 4N1 CANADA AGR & AGRIFOOD CANADA LETHBRIDGE RES STN LETHBRIDGE AB T1J 4B1 CANADA 0014-TC132-0014 TC132: Document Delivery available ~?n"Olson, M. E. Morck, D. W. Ceri, H.19964The Efficacy of a Giardia Lamblia Vaccine in Kittens249-256SCanadian Journal of Veterinary Research - Revue Canadienne de Recherche Veterinaire604Muris trophozoites. Experimental infections. Feline giardiasis. Cats. Dogs. Cysts. Oocysts. Veterinary medicine/animal health. Reprint available from: Olson ME. UNIV CALGARY, GASTROINTESTINAL RES GRP, CALGARY, AB T2N 4N1.Twenty kittens were vaccinated with a Giardia lamblia vaccine prepared on a commercial scale on day 0 and boosted on day 21 (group 1); while 10 kittens received only saline (group 2). These kittens were challenged on day 35 with 10(6) Giardia lamblia trophozoites by a surgical intraduodenal injection. Three control kittens were not vaccinated and not challenged (group 3). Following challenge, Giardia vaccinated kittens had significantly fewer days in which abnormal stools were observed and reduced food intake occurred compared to saline injected animals. The rate of weight gain between group 1 and group 2 animals was not different in the prechallenge period (day 0 to day 35), but vaccinated animals had a significantly higher weight gain in the postchallenge period (P < 0.05). On day 56, all vaccinated animals were not passing cysts in their feces, while 40% of saline injected kittens had Giardia cysts in their feces. In vaccinated kittens, cysts were never demonstrated in 45% of the animals, while cysts were detected in 90% of the saline injected kittens. Viability of the cysts in vaccinated kittens was 38% while the cysts viability in saline injected kittens was 99%. On postmortem examination, trophozoites could be detected in 5% of vaccinated kittens and 60% of saline injected kittens. Vaccination produced an elevated Giardia specific serum IgG and IgA response prior to challenge and throughout the postinfection period. The Giardia infection in the saline injected group did not induce an elevated specific serum response. Giardia vaccination of kittens provides protection in kittens from an experimental challenge by reducing or eliminating intestinal trophozoites and fecal cyst excretion. [References: 37] 37 English Article Current Contents/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME UNIV CALGARY GASTROINTESTINAL RES GRP CALGARY AB T2N 4N1 CANADA UNIV CALGARY ANIM HLTH UNIT CALGARY AB T2N 4N1 CANADA UNIV CALGARY DEPT BIOL SCI CALGARY AB T2N 4N1 CANADA 0002-VN250-0002 VN250: Document Delivery available ~?oFMcAllister, T. A. Annett, C. B. Olson, M. E. Morck, D. W. Cheng, K. J.1996DEffect of Salinomycin on Giardiasis and Coccidiosis in Growing Lambs 2896-2903Journal of Animal Science7412Feedlot performance. Cattle. Cryptosporidium. Supplementation. Infections. Monensin. Invitro. Animals. Calves. Level. Animal sciences. Reprint available from: Mcallister TA. AGR & AGRI FOOD CANADA, RES CTR, LETHBRIDGE, AB T1J 4B1.An experiment was undertaken to determine the effect of salinomycin on Giardia in vitro and on Giardia and coccidia in growing lambs. Concentrations of salinomycin above (3.9 mu g/mL) reduced the adherence (index of viability) of Giardia S2 trophozoites by more than 50%. This strain did not develop resistance after repeated exposure to sublethal concentrations of salinomycin. Five of 40 lambs escaped natural infection by Giardia, and these were inoculated with greater than or equal to 500,000 cysts. Giardiasis (presence of cysts in feces) was confirmed in all lambs before commencement of the experiment. Coccidiosis (presence of oocysts) developed by natural exposure. Lambs were assigned randomly to diets containing 0, 4, 10, or 16 ppm of salinomycin. Giardia cyst and coccidia oocyst excretions were determined on 6 d during the first week and weekly thereafter. Giardia cysts were detected at each sampling date in all treatments (highest release, 2.3 x 10(6) cysts/g feces). The number of Giardia cysts shed in feces was not affected(P >.05) by salinomycin but did decline (P <.05) with time. Average infection rates remained above 50% until d 41 of the experiment and declined linearly(P <.05) with salinomycin concentration and time. The number of coccidia oocysts in feces was low !highest release, 6.8 x 10(4) oocysts/g feces), but shedding occurred in 38 of the 40 lambs. Treatment with salinomycin had a cubic effect (P <.05) on coccidia oocyst excretion, and no oocysts were detected beyond d 28. Treatment effect on average daily gain (P <.002), dry matter intake (P <.02), and final live weight(P <.07) was cubic, whereas carcass weight (P <.003) and dressing percentage (P <.08) were linearly affected by salinomycin concentration. Although a beneficial effect of 10 ppm salinomycin on lamb performance was apparent, the development of natural resistance makes it difficult to attribute this response to the control of coccidiosis or giardiasis. [References: 39] 39.English Article Current Contents/Agriculture, Biology & Environmental Sciences. Reprint available from: Mcallister TA AGR & AGRI FOOD CANADA RES CTR LETHBRIDGE AB T1J 4B1 CANADA UNIV CALGARY DEPT MICROBIOL & INFECT DIS CALGARY AB T2N 1N4 CANADA UNIV CALGARY DEPT BIOL SCI CALGARY AB T2N 1N4 CANADA 0005-WE449-0005 WE449: Document Delivery available `~?p_Ohandley, R. M. Olson, M. E. McAllister, T. A. Morck, D. W. Jelinski, M. Royan, G. Cheng, K. J.1997>Efficacy of Fenbendazole for Treatment of Giardiasis in Calves384-388'American Journal of Veterinary Research584Albendazole. Metronidazole. Dogs. Cryptosporidium. Infections. Benzimidazoles. Duodenalis. Resistance. Animals. Invitro. Veterinary medicine/animal health. Reprint available from: Olson ME. UNIV CALGARY, GASTROINTESTINAL RES GRP, CALGARY, AB T2N 4N1.Objective-To determine efficacy of fenbendazole for treatment of giardiasis in calves. Animals-Twenty male and 15 female Holstein calves (100 to 180 kg), naturally infected with Giardia sp. Procedure-In vitro fenbendazole susceptibility and resistance development was determined for a ruminant Giardia isolate by use of an adherence assay. Carves were treated as follows. group 1, a single administration of 5 mg of fenbendazole/kg of body weight; group 2, a single administration of 10 mg of fenbendazole/kg; group 3, 5 mg of fenbendazole/kg, every 24 hours for 3 days; group 4, 10 mg of fenbendazole/kg, every 24 hours for 3 days; group 5, 20 mg of fenbendazole/kg, every 24 hours for 3 days; group 6, 0.833 mg of fenbendazole/kg, even/ 24 hours for 6 days; and group 7, saline solution. Fecal Giardia cysts were counted on days -3 through -1 and 1 through 7, 9, 11, 13, 21, and 28 by use of sucrose gradient concentration and staining with a fluorescent monoclonal antibody. Results-The 50% adherence inhibition concentration was 0.024 +/- 0.002 mu g/ml, and resistance could not be detected after 5 weeks of continuous culture at sublethal concentration of fenbendazole (0.01 mu g/ kg). Fenbendazole was 100% effective in eliminating cysts from the feces within 6 days for calves in treatment groups 2-6. Reinfection was observed in some calves within the 28-day study period. Conclusions- Fenbendazole is effective in the elimination of Giardia infections in calves, but repeat treatments may be required in reinfected animals. Clinical Relevance-Fenbendazole is an effective and economical treatment for Giardia-associated diarrhea and growth rate reduction in calves. [References: 38] 38English Article Current Contents/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME UNIV CALGARY GASTROINTESTINAL RES GRP CALGARY AB T2N 4N1 CANADA UNIV CALGARY GASTROINTESTINAL RES GRP CALGARY AB T2N 4N1 CANADA UNIV CALGARY ANIM HLTH UNIT CALGARY AB T2N 4N1 CANADA AGR & AGRI FOOD CANADA LETHBRIDGE RES CTR LETHBRIDGE AB T1J 4B1 CANADA HOECHST ANIM HLTH REGINA SK S7N 5B4 CANADA 0024-WT576-0024 WT576: Document Delivery availableE~?qLOlson, M. E. Thorlakson, C. L. Deselliers, L. Morck, D. W. McAllister, T. A.19974Giardia and Cryptosporidium in Canadian Farm Animals375-381Veterinary Parasitology684Prevalence. Infection. Pigs. Veterinary medicine/animal health. Reprint available from: Olson ME. UNIV CALGARY, FAC MED, UNIV VET, GASTROINTESTINAL RES GRP.Giardia intestinalis and Cryptosporidium spp, are commonly identified intestinal pathogens in humans and animals. In light of the clinical disease, production losses and zoonotic potential of both Giardia and Cryptosporidium infections, a study was undertaken to investigate the prevalence of these parasites in cattle, sheep, pigs and horses in Canadian farms at different geographical locations. A total of 104 cattle, 89 sheep, 236 pigs and 35 horses were sampled from 15 different Canadian geographical locations. Fecal samples were examined after concentration and immunofluorescent staining. Giardia and Cryptosporidium were present in cattle and sheep in six out of six sites sampled. In cattle the overall prevalence was 29% for Giardia and 20% for Cryptosporidium. Giardia was identified in 38% of sheep while 23% of sheep were positive for Cryptosporidium. Giardia and Cryptosporidium were identified in four out of six hog operations with an overall prevalence of 9% for Giardia and 11% for Cryptosporidium. All horse sampling locations (4/4) were positive for Giardia with 20% of animals infected. Cryptosporidium was identified in three out of four sampling sites with a prevalence of 17%. The prevalence of Giardia and Cryptosporidium was greater in calves and lambs compared to adults. This study demonstrates that both Giardia and Cryptosporidium appear to be prevalent in farm livestock. [References: 16] 16English Article Current Contents/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME UNIV CALGARY FAC MED UNIV VET GASTROINTESTINAL RES GRP HLTH SCI BLDG CALGARY AB T2N 4N1 CANADA AGR & AGRI FOOD CANADA LETHBRIDGE RES CTR LETHBRIDGE AB T1J 4B1 CANADA 0010-WQ534-0010 WQ534: Document Delivery availableP~?r.Olson, M. E. Roach, P. D. Stabler, M. Chan, W.19972Giardiasis in Ringed Seals from the Western Arctic646-648Journal of Wildlife Diseases333Canada. Veterinary medicine/animal health. Reprint available from: Olson ME. UNIV CALGARY, FAC MED, GASTROINTESTINAL RES GRP, CALGARY.Sixteen beluga whales (Delphinapterus leucas) and fifteen ringed seals (Phoca hispida) from the western arctic region of Canada were examined for giardiasis and cryptosporidiosis. Intestinal contents from the rectum and colon were collected from animals slaughtered by Inuit hunters. A fluorescent monoclonal antibody identified Giardia sp. cysts in three of 15 (20%) seals. Thus, ringed seals are implicated as a potential reservoir for this zoonosis in the arctic. [References: 11] 11English Article Current Contents/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME UNIV CALGARY FAC MED GASTROINTESTINAL RES GRP CALGARY AB CANADA INDIAN & NO AFFAIRS WHITEHORSE YT CANADA JOINT SECRETARIAT INUVIK NT CANADA 0034-XM004-0034 XM004: Document Delivery available m~?sgOlson, M. E. Guselle, N. J. Ohandley, R. M. Swift, M. L. McAllister, T. A. Jelinski, M. D. Morck, D. W.1997?Giardia and Cryptosporidium in Dairy Calves in British Columbia703-706:Canadian Veterinary Journal - Revue Veterinaire Canadienne3811Prevalence. Infections. Outbreak. Farms. Veterinary medicine/animal health. Reprint available from: Olson ME. UNIV CALGARY, GASTROINTESTINAL SCI RES GRP, CALGARY, AB T2N 4N1.A study was undertaken to determine the prevalence of Giardia infections in dairy calves and to compare Giardia and Cryptosporidium infections in calves of different ages. Fresh fecal samples were collected from 386 male and female Holstein calves (newborn to 24 wk) in 20 dairies located in the lower Fraser river valley area of British Columbia. Giardia intestinalis, Cryptosporidium parvum, and Cryptosporidium muris were enumerated in each sample after concentration by sucrose gradient centrifugation and immunofluorescent staining. Giardia was identified at all farm locations. The overall prevalence of Giardia in calves was 73% with a geometric mean cyst count of 1180 cysts per gram of feces (CI, 41 to 5014). Cryptosporidium parvum and C. muris were identified in 80% and 40% of the farms, respectively. The prevalence of C. parvum was 59%, and the geometric mean for oocysts was 457 oocysts per gram of feces (CI, 18 to 160). The prevalence of C. muris was only 2% and the mean oocyst counts were 54 oocysts per gram of feces. Giardiasis was not age dependent, and approximately 80% of the calves from 2 to 24 wk were infected. In contrast, C. parvum infections were predominant in calves 2 to 4 wk, while C. muris was demonstrated in calves older than 4 wk. Fourty-seven percent of calves with diarrhea had high numbers of Giardia cysts in their feces. Giardia infections are highly prevalent in dairy calves and should be considered in animals with diarrhea or failure to thrive. [References: 22] 22English Article Current Contents/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME UNIV CALGARY GASTROINTESTINAL SCI RES GRP CALGARY AB T2N 4N1 CANADA UNIV CALGARY ANIM HLTH UNIT CALGARY AB T2N 4N1 CANADA PRO FORM FEEDS CHILLIWACK BC V2P 6J6 CANADA AGR & AGRI FOOD CANADA LETHBRIDGE RES CTR LETHBRIDGE AB T1J 4B1 CANADA HOECHST ROUSSEL VET REGINA SK S4N 6C2 CANADA 0015-YD538-0015 YD538: Document Delivery available~?t"Olson, M. E. Morck, A. W. Ceri, H.1997@Preliminary Data on the Efficacy of a Giardia Vaccine in Puppies777-779:Canadian Veterinary Journal - Revue Veterinaire Canadienne3812Cysts. Oocysts. Dogs. Veterinary medicine/animal health. Reprint available from: Olson ME. UNIV CALGARY, FAC MED, HLTH SCI ANIM RESOURCE CTR, CALGARY.|Twenty puppies were vaccinated with a trophozoite-derived Giardia vaccine on day 0 and boosted on day 21 (Group 1); 10 control puppies received only saline (Group 2). Both groups were experimentally infected on day 35 with 1 X 10(6) Giardia duodenalis trophozoites by intraduodenal injection. Immunization provided protection to experimental Giardia infection. [References: 11] 11oEnglish Article Current Contents/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME UNIV CALGARY FAC MED HLTH SCI ANIM RESOURCE CTR CALGARY AB T2N 4N1 CANADA UNIV CALGARY GASTROINTESTINAL SCI RES UNIT CALGARY AB T2N 4N1 CANADA UNIV CALGARY ANIM HLTH UNIT CALGARY AB T2N 4N1 CANADA UNIV CALGARY DEPT BIOL SCI CALGARY AB T2N 4N1 CANADA 0011-YL377-0011 YL377: Document Delivery available[~?u9Deselliers, L. P. Tan, D. T. M. Scott, R. B. Olson, M. E.1997gEffects of Giardia Lamblia Infection on Gastrointestinal Transit and Contractility in Mongolian Gerbils 2411-2419Digestive Diseases & Sciences4212Rat small-intestine. Smooth-muscle. Growth. Malabsorption. Motility. Medical research, organs & systems Gastroenterology and hepatology. Reprint available from: Olson ME. UNIV CALGARY, HLTH SCI CTR, GASTROINTESTINAL RES UNIT, GASTROINTESTINAL RES GRP.To determine if Giardia lamblia infection is associated with altered gastrointestinal transit and smooth muscle contractile function, Mongolian gerbils were infected orogastrically with 2 x 10(5) trophozoites (infected) or vehicle (uninfected controls). At the time of peak colonization, control and infected animals were infused either orogastrically or intraduodenally with Cr-51. Gastric emptying of isotope and intestinal transit (measured by the geometric center of distribution of intestinal Cr-51 transit) were significantly (P < 0.05) greater in the infected compared to control animals in both the fasted and the fed states. Then, to determine whether Giardia lamblia has an effect on the contractility of longitudinal and circular smooth muscle, isometric tension of jejunal segments was recorded. The development of active tension with stretch and the dose-response curve to bethanechol were significantly increased in the longitudinal muscle of infected animals compared to controls. However, the circular smooth muscle did not show a similar increase in contractility. These findings suggest that an altered gastrointestinal transit and smooth muscle contractility may be involved in the pathophysiology of giardiasis. [References: 31] 31VEnglish Article Current Contents/Life Sciences Current Contents/Clinical Medicine. Reprint available from: Olson ME UNIV CALGARY HLTH SCI CTR GASTROINTESTINAL RES UNIT GASTROINTESTINAL RES GRP 3330 HOSP DR NW CALGARY AB T2N 4N1 CANADA UNIV CALGARY HLTH SCI CTR GASTROINTESTINAL RES UNIT GASTROINTESTINAL RES GRP CALGARY AB T2N 4N1 CANADA 0005-YR203-0005 YR203: Document Delivery available~?vAYanke, S. J. Ceri, H. McAllister, T. A. Morck, D. W. Olson, M. E.1998LSerum Immune Response to Giardia Duodenalis in Experimentally Infected Lambs9-19Veterinary Parasitology751Antibody-responses. Antigenic analysis. Proteins. Muris. Mice. Veterinary medicine/animal health. Reprint available from: Olson ME. UNIV CALGARY, GASTROINTESTINAL SCI RES GRP, CALGARY, AB T2N 4N1.~Twenty-three protozoan-free lambs were experimentally infected with Giardia duodenalis trophozoites at 6 weeks of age, while 24 control lambs were not challenged. Weekly blood samples were taken and faecal cyst counts monitored for 11 weeks following infection. All experimentally infected lambs remained infected throughout the 11 week study period and control animals remained free of the parasite. Giardia-specific serum IgM, IgG and IgA antibody titers were determined weekly in 10 infected and 10 control lambs by enzyme-linked-immunosorbent assay (ELISA). Western blot analysis of serum immunoglobulins to proteins derived from four different Giardia isolates (S2, WB, D3 and NF) was performed. Weekly mean control IgM, IgG and IgA titers did not change throughout the study. Infected lambs showed no difference in IgM titers between weeks 0 and 11, but IgG and IgA titers of infected lambs differed from the preimmune (week 0) serum titer at weeks 5, 7, 9, 11 and weeks 5, 9 and 11 respectively. SDS polyacrylamide gel electrophoresis analysis showed homogeneity in the proteins of the four Giardia isolates. Antigenic proteins were also similar for all four isolates; however, the proteins recognized by IgM, IgG and IgA antibodies differed. The weak immune response of lambs to Giardia may account for the chronic nature of this disease in sheep. (C) 1998 Elsevier Science B.V. [References: 32] 32mEnglish Article Current Contents/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME UNIV CALGARY GASTROINTESTINAL SCI RES GRP CALGARY AB T2N 4N1 CANADA UNIV CALGARY GASTROINTESTINAL SCI RES GRP CALGARY AB T2N 4N1 CANADA UNIV CALGARY DEPT BIOL SCI CALGARY AB T2N 4N1 CANADA AGR CANADA LETHBRIDGE RES CTR LETHBRIDGE AB T1J 4B1 CANADA 0002-ZE199-0002 ZE199: Document Delivery available ~?w?Olson, M. E. Goh, J. Phillips, M. Guselle, N. McAllister, T. A.1999QGiardia cyst and Cryptosporidium oocyst survival in water, soil, and cattle feces 1991-1996 Journal of Environmental Quality286Parvum oocysts. Viability. Assay. Environment/Ecology in Current Contents(R)/Agricultural, Biology & Environmental Sciences. 2000 week 03 Reprint available from: Olson ME. Univ Calgary, Fac Med, 3330 Hosp Dr NW, Calgary, AB T2N 4N1.Giardiasis and cryptosporidiosis are gastrointestinal diseases caused by protozoan parasites that mag infect domestic animals, wildlife and human beings. The ability of cysts and oocysts of these parasites to persist in the environment was determined because agricultural fecal waste has the potential to contaminate municipal water supplies. The degradation rate and viability of Giardia cysts and Cryptosporidium oocysts in water, cattle (Bos taurus) feces, and soil was evaluated at temperatures of -4, 4, and 25 degrees C for up to 12 wk. Cysts and oocysts were enumerated after staining samples with a specific fluorescent monoclonal antibody and the viability was determined using propidium iodide dye exclusion and mouse infectivity assays. Giardia cysts were noninfective in water, feces, and soil following 1 wk of freezing to -4 degrees C and within 2 wk at 25 degrees C. At 4 degrees C Giardia cysts were infective for 11 wk in water, 7 wk in soil, and 1 wk in cattle feces. Cryptosporidium oocysts were more environmentally resistant. At -4 and 4 degrees C, the oocysts could survive in water and soil for >12 wk but degradation was accelerated at 25 degrees C. Cryptosporidium oocysts also were degraded more rapidly in feces and in soil containing natural microorganisms. Contaminated cattle feces should be distributed on fields during warmer weather and after 12 wk of storage to reduce potential waterborne transmission following heavy runoffs. [References: 16] 16AEnglish Article Current Contents(R)/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME Univ Calgary, Fac Med 3330 Hosp Dr NW Calgary AB T2N 4N1 Canada Univ Calgary, Fac Med Calgary AB T2N 4N1 Canada Agr & Agri Food Canada, Lethbridge Res Ctr Lethbridge AB T1J 4B1 Canada 0040 J. Environ. Qual-266EU-0040 266EU: Document Delivery available~?x"Olson, M. E. Ceri, H. Morck, D. W.2000Giardia vaccination213-217Parasitology Today165Immune-response. Lamblia. Duodenalis. Antigens. Humans. Microbiology in Current Contents(R)/Life Sciences. 2000 week 22 Reprint available from: Olson ME. Univ Calgary, Dept Microbiol & Infect Dis, 3330 Hosp Dr NW, Calgary, AB T2N 4N1.Recently, a Giardia vaccine has become commercially available in the USA for prevention of clinical signs of giardiasis and reduction of cyst shedding in dogs and cats. The vaccine is based upon the current state of knowledge of Giardia antigenicity and immunology. Here, Merle Olson, Howard Ceri and Douglas Morck describe studies that led to the development of this vaccine and subsequent efficacy studies. Immunoprophylaxis and immunotherapeutic application of the vaccine are discussed. [References: 27] 277English Article Current Contents(R)/Life Sciences. Reprint available from: Olson ME Univ Calgary, Dept Microbiol & Infect Dis 3330 Hosp Dr NW Calgary AB T2N 4N1 Canada Univ Calgary, Dept Microbiol & Infect Dis Calgary AB T2N 4N1 Canada Univ Calgary, Dept Biol Sci Calgary AB T2N 4N1 Canada 0012 Parasitol. Today-310BX-0012 310BX: Document Delivery available N~?yEO'Handley, R. M. Olson, M. E. Fraser, D. Adams, P. Thompson, R. C. A.2000nPrevalence and genotypic characterisation of Giardia in dairy calves from Western Australia and Western Canada193-200Veterinary Parasitology903<Farm-animals. Cryptosporidium. Infection. Fenbendazole. Zoonosis. Outbreak. Diarrhea. Efficacy. Veterinary Medicine/Animal Health in Current Contents(R)/Agricultural, Biology & Environmental Sciences. 2000 week 29 Reprint available from: Olson ME. Univ Calgary, Gastrointestinal Res Grp, Calgary, AB T2N 4N1, Canada.In this study, the prevalence of Giardia duodenalis infections was determined in Western Canadian and Western Australian dairy calves. Faecal samples were collected from Holstein calves located on a commercial dairy near Lethbridge, Alta., Canada (N=28) and from calves located on two commercial dairies located near Perth, WA, Australia (N=36). Faecal samples were examined for the presence of Giardia cysts using sucrose gradient centrifugation, followed by immunofluoresence microscopy. DNA was then extracted from Giardia isolates obtained from positive samples. A PCR based method was employed to amplify and sequence a 292 bp region of the 16S-rRNA gene. Genetic sequences obtained from Giardia isolates were compared to each other and to previously sequenced isolates. Following a single faecal sample, 58% of Western Australian calves and 57% of Western Canadian calves were positive for Giardia. Geometric mean cyst counts/g of faeces were 839 for Western Australian calves and 3475 for Western Canadian calves, but these values did not differ significantly. Genetic sequences were obtained from 10 calves from Western Canada, while six sequences were obtained from Western Australian calves. Of the Western Canadian isolates, eight aligned with the proposed 'Hoofed livestock' genotype. Of the five isolates obtained from Western Australian calves, four sequences were identical to the 'Hoofed-livestock' genotype. Two isolates from the Western Canadian calves and one isolate from the Western Australian calves had the identical genetic sequence to the Genotype (Assemblage) A sequence, a common human genotype. The results of this study demonstrate, for the first time, that Giardia infections occur in Western Australian calves. Also, calves from different geographical locations appear to be primarily infected with a Giardia genotype unique to hoofed livestock. However, calves can shed Giardia cysts potentially infective for humans. Thus, Giardia infections should be considered important to Australian dairy producers, and infections in calves may pose a risk to public health regardless of geographical location. (C) 2000 Elsevier Science B.V. All rights reserved. [References: 29] 29FEnglish Article Current Contents(R)/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME Univ Calgary, Gastrointestinal Res Grp Calgary AB T2N 4N1 Canada Univ Calgary, Gastrointestinal Res Grp Calgary AB T2N 4N1 Canada Murdoch Univ, Div Vet & Biomed Sci Murdoch WA 6150 Australia 0004 Vet. Parasitol-327AJ-0004 327AJ: Document Delivery available }~?zIO'Handley, R. M. Buret, A. G. McAllister, T. A. Jelinski, M. Olson, M. E.2001bGiardiasis in dairy calves: effects of fenbendazole treatment on intestinal structure and function73-79&International Journal for Parasitology311Disaccharidase activity. T-lymphocytes. Cryptosporidium. Infection. Efficacy. Gerbils. Lamblia. Model. Benzimidazoles. Giardiosis. Biology in Current Contents(R)/Agricultural, Biology & Environmental Sciences Microbiology in Current Contents(R)/Life Sciences. 2001 week 12 Reprint available from: Olson ME. Univ Calgary, Fac Med, Anim Resources Ctr, Dept Gastrointestinal Sci, 3330 Hosp Dr NW, Calgary, AB T2N 4N1.Twelve Giardia duodenalis-infected Holstein dairy calves were allocated into a treatment (n = 6) and placebo group (n = 6) according to pre-study faecal cyst counts. Calves in the treatment group received an oral dose of 5 mg/kg fenbendazole once daily for 3 days, while placebo calves received a sterile saline solution. Calves were euthanised 7 days following the initiation of treatment and intestinal were collected and prepared for trophozoite quantitation, histology, electron microscopy, and disaccharidase assays. In all calves treated with fenbendazole, intestinal trophozoites were below detection limits, while in saline-treated calves, trophozoites were observed in all intestinal segments. Histologically, no significant difference was observed between treatment groups with respect to intestinal villus height or crypt depth. However, a significant decline in the number of intraepithelial lymphocytes (IEL) was observed in fenbendazole-treated calves when compared with placebo-treated calves in the duodenum (13.9 +/- 1.2 vs. 17.0 +/- 1.1 IEL/100 enterocytes) and jejunum (21.6 +/- 0.8 vs. 30.7 +/- 1.0 IEL/100 enterocytes). In addition, measurements from TEM micrographs demonstrated a significant increase in microvillus surface area in the jejunum of fenbendazole-treated calves compared with saline-treated calves (31.2 +/- 10.2 vs. 22.8 +/- 7.6 mum(2)). This increase in microvillus surface area was also associated with an increase in jejunal maltase activity in fenbendazole-treated calves compared with calves treated with saline. These results demonstrate that fenbendazole is an effective treatment for giardiasis in calves. fenbendazole treatment eliminated Giardia trophozoites from the small intestine of calves resulting in increased microvillus surface area and greater intestinal enzyme activity. This study also demonstrates that the pathogenesis of giardiasis in calves is similar to that observed in humans and laboratory animals, and provides further evidence that Giardia is a pathogen of cattle with potential economic importance. (C) 2001 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved. [References: 32] 32English Article Current Contents(R)/Agriculture, Biology & Environmental Sciences Current Contents(R)/Life Sciences. Reprint available from: Olson ME Univ Calgary, Fac Med, Anim Resources Ctr, Dept Gastrointestinal Sci 3330 Hosp Dr NW Calgary AB T2N 4N1 Canada Univ Calgary, Fac Med, Anim Resources Ctr, Dept Gastrointestinal Sci Calgary AB T2N 4N1 Canada Univ Calgary, Dept Biol Sci Calgary AB T2N 1N4 Canada Agr Canada, Lethbridge Res Ctr Lethbridge AB T1J 4B1 Canada Hoechst Roussel Vet Regina SK S7N 5B4 Canada 0011 Int. J. Parasit-404GT-0011 404GT: Document Delivery available ~?{SMcAllister, T. A. Annett, C. B. Cockwill, C. L. Olson, M. E. Wang, Y. Cheeke, P. R.2001<Studies on the use of Yucca schidigera to control giardiosis85-99Veterinary Parasitology972wHost-parasite interactions. Steroidal saponins. Farm-animals. Ruminal fermentation. Giardiasis. Extract. Water. Cryptosporidium. Lamblia. Fenbendazole. Veterinary Medicine/Animal Health in Current Contents(R)/Agricultural, Biology & Environmental Sciences. 2001 week 28 Reprint available from: McAllister TA. Agr & Agri Food Canada, Res Ctr, POB 3000, Lethbridge, AB T1J 4B1. The potential anti-giardial activity of a powdered preparation of Yucca schidigera (yucca powder) was investigated in vitro, in a modified adherence inhibition assay, and in vivo, by enumeration of trophozoites (intestinal) or cysts (fecal) in experimentally infected gerbils and lambs receiving oral doses of whole or butanol-extracted yucca powder. Yucca powder, butanol-, acetone- and chloroform-extracted powder, and the butanol-insoluble fraction of the powder were required in concentrations of 22, 15, 62, 135 and 250 mug/ml, respectively, to reduce in vitro trophozoite adherence by 50%. Ethyl ether extract exhibited no anti-giardial activity. Virtually no trophozoites were tolerant of butanol extract at greater than or equal to 90 mug/ml. Butanol extract at 1500 mug/ml exerted effects on trophozoites similar to the nitroimidazole, metronidazole, at 40 mug/ml during a 27-h incubation. Reducing trophozoite adherence to 50% of the controls required 5-10 h of exposure to butanol extract or metronidazole. Oral administration of butanol extract (6.1 mg) or metronidazole (1 mg) once daily for 3 days reduced the number of trophozoites in the small intestine of infected gerbils, significantly (P < 0.05) in the jejunum and ileum, and numerically (P > 0.05) in the duodenum (n = 8). Oral dosing of 50 mg of butanol extract in eight doses over 3 days reduced (P < 0.05) trophozoites in the duodenum and jejunum, and eliminated them from the ileum. Including 4.5% (w/w) yucca powder in the diet did not alter Giardia trophozoite recovery from the duodenum or jejunum of infected gerbils, but trophozoite reduction (P = 0.051) was observed in the ileum (n = 9), Jejunum gut loop data were inconclusive, possibly due to insufficient duration of exposure of trophozoites to butanol extract. Compared to controls (0 g yucca powder per day) lambs receiving 10 g of yucca powder per day in their diet shed fewer (P < 0.05) Giardia cysts in their feces after 5, 9, 12 and 19 days of treatment, but a corresponding decline in the prevalence of infection was not observed (n = 10). After 26 days, Giardia infections persisted in 90% of the lambs in both treatment groups. At the dosing levels studied in vivo, yucca powder did not affect the extent of cyst shedding by experimentally infected lambs, but Further purification and concentration of the saponin fraction from Y. schidigera may provide the anti-giardial effects observed in vitro and in dosing trials. Published by Elsevier Science B.V. [References: 51] 51xEnglish Article Current Contents(R)/Agriculture, Biology & Environmental Sciences. Reprint available from: McAllister TA Agr & Agri Food Canada, Res Ctr POB 3000 Lethbridge AB T1J 4B1 Canada Agr & Agri Food Canada, Res Ctr Lethbridge AB T1J 4B1 Canada Univ Calgary, Gastrointestinal Res Grp Calgary AB T2N 4N1 Canada Desert King Int San Diego, CA 92154 USA 0001 Vet. Parasitol-439GT-0001 439GT: Document Delivery available~?|:Olson, M. E. Hannigan, C. J. Gaviller, P. F. Fulton, L. A.2001BThe use of a Giardia vaccine as an immunotherapeutic agent in dogs865-868:Canadian Veterinary Journal - Revue Veterinaire Canadienne4211Cell. Veterinary Medicine/Animal Health in Current Contents(R)/Agricultural, Biology & Environmental Sciences. 2001 week 49 Reprint available from: Olson ME. Univ Calgary, Fac Med, 3330 Hosp Dr NW, Calgary, AB T2N 4N1.jDogs (n = 13), which had failed to be cured of giardiosis following chemotherapeutic measures, were treated with a Giardia vaccine (2-3 injections). Clinical signs resolved between 16 and 42 days postvaccination and cessation of fecal cyst shedding was between 21 and 70 days. Vaccination is a potential method of treating giardiosis in dogs. [References: 13] 13English Article Current Contents(R)/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME Univ Calgary, Fac Med 3330 Hosp Dr NW Calgary AB T2N 4N1 Canada Univ Calgary, Fac Med Calgary AB T2N 4N1 Canada Forest Lawn Vet Hosp Calgary AB T2B 0X8 Canada Country Hills Vet Clin Calgary AB T2E 7A2 Canada Big Hills Vet Serv Cochrane AB T0L 0W0 Canada 0003 Can. Vet. J.-Rev. Vet. Can-489WW-0003 489WW: Document Delivery available ~?}iHeitman, T. L. Frederick, L. M. Viste, J. R. Guselle, N. J. Morgan, U. M. Thompson, R. C. A. Olson, M. E.2002Prevalence of Giardia and Cryptosporidium and characterization of Cryptosporidium spp. isolated from wildlife, human, and agricultural sources in the North Saskatchewan River Basin in Alberta, Canada530-541 Canadian Journal of Microbiology486Surface-water supplies. Molecular characterization. United-states. Human feces. Parvum. Transmission. Oocysts. Hosts. Identification. Genotypes. Biotechnology & Applied Microbiology in Current Contents(R)/Agricultural, Biology & Environmental Sciences Microbiology in Current Contents(R)/Life Sciences. 2002 week 35 Reprint available from: Olson ME. Univ Calgary, Dept Microbiol & Infect Dis, Calgary, AB T2N 4N1, Canada.zThe environmental distribution of Giardia spp. and Cryptosporidium spp. is dependent upon human, agricultural, and wildlife sources. The significance of each source with regard to the presence of parasites in the environment is unknown. This 2-year study examined parasite prevalence in human sewage influent, wildlife, and agricultural sources associated with the North Saskatchewan River Basin in Alberta, Canada. Fecal samples were collected from cow-calf, dairy, and hog operations in the watershed area. Sewage-treatment facilities were sampled bimonthly during the 2-year study, and wildlife scat was collected at locations along tributaries of the North Saskatchewan River. All samples were analyzed for the presence of Giardia and Crytosporidium, using sucrose-gradient separation followed by immunofluorescent microscopy. Giardia and Cryptosporidium were detected in all three sources. The lowest prevalence of both Giardia (3.28%) and Cryptosporidium (0.94%) was found in wildlife, with 6 of 19 species testing positive. Sewage influent had the highest prevalence of Giardia (48.80%) and Cryptosporidium parvum-like oocysts (5.42%); however, the concentration of both parasites was minimal compared with the concentration detected in cattle feces. Cow-calf sources contained the highest concentration of Giardia (mean 5800/g feces, P < 0.01), and dairy sources contained the highest concentration of C. parvum-like oocysts (mean 295/g feces, P < 0.01). Although prevalence and concentration are higher in cattle feces than in sewage, the Giardia and Cryptosporidium in animal manure do not have direct access to water draining into the North Saskatchewan River. PCR-based characterization of rDNA from isolates of Cryptosporidium collected from Alberta human, pig, calf, mature steer, dog, cat, and beaver hosts revealed distinct genetic differences that may reflect host specificity. [References: 39] 39English Article Current Contents(R)/Agriculture, Biology & Environmental Sciences Current Contents(R)/Life Sciences. Reprint available from: Olson ME Univ Calgary, Dept Microbiol & Infect Dis Calgary AB T2N 4N1 Canada Univ Calgary, Dept Microbiol & Infect Dis Calgary AB T2N 4N1 Canada Murdoch Univ, Div Vet & Biomed Sci, State Agr Biotechnol Ctr Murdoch WA 6150 Australia WHO, Collaborating Ctr Mol Epidemiol Parasit Infect Murdoch WA 6150 Australia Univ Calgary, Dept Biol Sci Calgary AB T2N 4N1 Canada 0005 Can. J. Microbiol-579RT-0005 579RT: Document Delivery available ~?~HHijjawi, N. S. Meloni, B. P. Ryan, U. M. Olson, M. E. Thompson, R. C. A.2002Successful in vitro cultivation of Cryptosporidium andersoni: evidence for the existence of novel extracellular stages in the life cycle and implications for the classification of Cryptosporidium 1719-1726&International Journal for Parasitology3214Phylogenetic analysis. Molecular characterization. Parvum. Apicomplexa. Cattle. Microscopy. Cyclospora. Coccidia. Oocysts. Muris. Biology in Current Contents(R)/Agricultural, Biology & Environmental Sciences Microbiology in Current Contents(R)/Life Sciences. 2003 week 04 Reprint available from: Thompson RCA. Murdoch Univ, Div Vet & Biomed Sci, Western Australian Biomed Res Inst, South St, Murdoch, WA 6150.The present study describes the complete development of all life cycle stages of Cryptosporidium andersoni in the HCT-8 cell line. The in vitro cultivation protocols were the same as those used for the successful growth of all life cycle stages of Cryptosporidium parvum (Int. J. Parasitol. 31 (2001) 1048). Under these culture conditions, C. andersoni grew and proliferated rapidly with the completion of the entire life cycle within 72 h post-infection. The developmental stages of C. andersoni are larger than those of C. parvum enabling easier identification of life cycle stages including a previously unrecognised extracellular stage. The presence of this extracellular stage was further confirmed following its isolation from the faeces of infected cattle using a laser microdissection technique. This stage was present in large numbers and some of them were seen undergoing syzgy. Extraction of DNA from the extracellular stage, followed by polymerase chain reaction-restriction fragment length polymorphism and sequencing of the 18S rDNA confirmed that this is a stage in the life cycle of C. andersoni. In vitro, extracellular stages were always observed moving over the HCT-8 cells infected with C. andersoni. Comparative observations with C. parvum also confirmed the presence of extracellular stages. Extracellular stages were recovered from in vitro culture after 5 days post-infection with the cattle genotype of C parvum and from infected mice. At least two morphologically different stages (stages one and two) were purified from mice after 72 h of infection. The presence and morphological characterisation of extracellular developmental stages in the life cycle of Cryptosporidium confirms its relationship to gregarines and provides important implications for our understanding of the taxonomic and phylogenetic affinities of the genus Cryptosporidium. The growth of C. andersoni in cell culture now provides a means of studying its development, metabolism, and behaviour as well as testing its response to different therapeutic agents. (C) 2002 Australian Society for Parasitology Inc. Published by Elsevier Science Ltd. All rights reserved. [References: 30] 30=English Article Current Contents(R)/Agriculture, Biology & Environmental Sciences Current Contents(R)/Life Sciences. Reprint available from: Thompson RCA Murdoch Univ, Div Vet & Biomed Sci, Western Australian Biomed Res Inst South St Murdoch WA 6150 Australia Murdoch Univ, Div Vet & Biomed Sci, Western Australian Biomed Res Inst Murdoch WA 6150 Australia Univ Western Australia, Sir Charles Gardiner Hosp Nedlands WA 6009 Australia Univ Western Australia, Ctr Neuromuscular & Neurol Disorders Nedlands WA 6009 Australia Univ Calgary Calgary AB Canada 0006 Int. J. Parasit-626XG-0006 626XG: Document Delivery available ~?+Guselle, N. J. Appelbee, A. J. Olson, M. E.2003ABiology of Cryptosporidium parvum in pigs: from weaning to market7-18Veterinary Parasitology1131fFarm-animals. Molecular characterization. Fecal transmission. Weaned piglets. Sri-lanka. Oocysts. Infection. Prevalence. Giardia. Calves. Veterinary Medicine/Animal Health in Current Contents(R)/Agricultural, Biology & Environmental Sciences. 2003 week 18 Reprint available from: Olson ME. Univ Calgary, Gastrointestinal Res Grp, Calgary, AB T2N 4N1, Canada.4Cryptosporidium parvum is commonly identified as infecting domestic livestock and humans. Prevalence of C. parvum in pigs has been reported, however, the duration and infection pattern of naturally acquired Cryptosporidium infections in pigs has not been reported. This study was undertaken to investigate the age of oocyst shedding and duration of natural Cryptosporidium parvum infections in pigs from weaning to market weight. Fecal samples were collected from weaned Yorkshire-Landrace piglets (n = 33) twice per week until Cryptosporidium oocysts were detected. Upon oocyst detection, fecal samples were collected three times per week and pigs were monitored throughout the study for diarrhea and examined after concentration and immunofluroescent staining. Cryptosporidium isolates were genotyped by polymerase chain reaction to amplify the HSP70 gene which was subsequently sequence analyzed. All 33 pigs shed oocysts some time during the study. The mean age of initial oocyst detection was 45.2 days post-weaning with the mean duration of infection 28.7 days. Mean number of Cryptosporidium oocysts was low and declined to zero prior to study completion. Episodes of diarrhea were not associated with oocyst excretion. Genetic sequences were obtained for 10 of the pigs. All of the 10 isolates aligned as the Cryptosporidium parvum 'pig' genotype. This study demonstrates that the age and duration of oocyst shedding in pigs infected with C. parvum porcine genotype is different from other livestock species. (C) 2003 Elsevier Science B.V. All rights reserved. [References: 53] 53 English Article Current Contents(R)/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME Univ Calgary, Gastrointestinal Res Grp Calgary AB T2N 4N1 Canada Univ Calgary, Gastrointestinal Res Grp Calgary AB T2N 4N1 Canada 0002 Vet. Parasitol-664FD-0002 664FD: Document Delivery availablei~?<Appelbee, A. J. Frederick, L. M. Heitman, T. L. Olson, M. E.2003SPrevalence and genotyping of Giardia duodenalis from beef calves in Alberta, Canada289-294Veterinary Parasitology11245Dairy calves. Cryptosporidium. Australia. Animals. Humans. Veterinary Medicine/Animal Health in Current Contents(R)/Agricultural, Biology & Environmental Sciences. 2003 week 16 Reprint available from: Olson ME. Univ Calgary, Fac Med, Dept Med Infect Dis, Gastrointestinal Res Grp, Calgary, AB T2N 4N1, Canada.Giardia infections in domestic cattle has come under increasing scrutiny owing to the potential contamination of surface and ground waters through manure distribution on fields and pasture runoff. The objective of the study was to determine the prevalence and genotypes of Giardia duodenalis in beef calves in major beef cow calf farms in Alberta, Canada. Fecal samples were collected from beef calves aged 2-10 weeks at nine farms in Alberta. Samples were examined for the presence of G. duodenalis cysts by immunofluorescent staining. Giardia cysts were found in 168 of the 495 fecal samples examined, with prevalence ranging from 7 to 60% among farms. Genotypic analysis of positive isolates utilizing PCR and sequencing of a 292 bp fragment of the 16S-rRNA locus, revealed the hoofed livestock genotype in 41 of the 42 isolates. One isolate was identical to the Assemblage A genotype. The results of this study demonstrate that beef calves in this area are primarily infected with the livestock genotype which is thought to be specific to artiodactyl hosts and non-infective to humans. This suggests that the Giardia carried by beef cattle may be a minimal zoonotic threat. (C) 2003 Elsevier Science B.V. All rights reserved. [References: 15] 15EEnglish Article Current Contents(R)/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME Univ Calgary, Fac Med, Dept Med Infect Dis, Gastrointestinal Res Grp Calgary AB T2N 4N1 Canada Univ Calgary, Fac Med, Dept Med Infect Dis, Gastrointestinal Res Grp Calgary AB T2N 4N1 Canada 0004 Vet. Parasitol-658TR-0004 658TR: Document Delivery available ~?aMcDonnell, P. A. Scott, K. G. E. Teoh, D. A. Olson, M. E. Upcroft, J. A. Upcroft, P. Buret, A. G.2003Giardia duodenalis trophozoites isolated from a parrot (Cacatua galerita) colonize the small intestinal tracts of domestic kittens and lambs31-46Veterinary Parasitology1111Cryptosporidium-parvum oocysts. Ibis threskiornis-spinicollis. Mongolian gerbils. Western-australia. Farm-animals. Infection. Mice. Cysts. Ultrastructure. Growth. Veterinary Medicine/Animal Health in Current Contents(R)/Agricultural, Biology & Environmental Sciences. 2003 week 09 Reprint available from: McDonnell PA. Griffith Univ, Fac Sci, Sch Biomol & Biomed Sci, Kessels Rd, Nathan, Qld 4111.mThis study examines the ability of Giardia duodenalis trophozoites, isolated from a wild bird, to colonize the intestinal tracts of companion animals (kittens) and domestic ruminants (lambs). Trophozoites colonized the intestinal tracts of intraduodenally inoculated animals as demonstrated by increasing parasite burdens within the duodenum and jejunum and by fecal passage of cysts within 4 days post-inoculation. The pathogenesis of the trophozoites was further investigated in kittens. In these animals, infection significantly reduced jejunal brush border microvillous length and density, which resulted in a loss of overall epithelial brush border surface area. This injury was associated with the production of diarrhea in four of five infected kittens. These findings indicate that some bird species may carry G. duodenalis that represent a possible health threat to companion animals and livestock. Our results describe the first successful colonization of avian-derived G. duodenalis trophozoites in the small intestines of domestic kittens and lambs. (C) 2002 Elsevier Science B.V. All rights reserved. [References: 62] 62English Article Current Contents(R)/Agriculture, Biology & Environmental Sciences. Reprint available from: McDonnell PA Griffith Univ, Fac Sci, Sch Biomol & Biomed Sci Kessels Rd Nathan Qld 4111 Australia Griffith Univ, Fac Sci, Sch Biomol & Biomed Sci Nathan Qld 4111 Australia Univ Calgary, Dept Biol Sci Calgary AB T2N 1N4 Canada Queensland Inst Med Res, Bancroft Ctr Herston Qld 4029 Australia 0004 Vet. Parasitol-639DL-0004 639DL: Document Delivery available s~?1O'Handley, R. M. Ceri, H. Anette, C. Olson, M. E.2003vPassive immunity and serological immune response in dairy calves associated with natural Giardia duodenalis infections89-98Veterinary Parasitology1132Serum antibody-responses. Muris trophozoites. Immunoglobulin-g. Lamblia. Cryptosporidium. Diarrhea. Efficacy. Mice. Fenbendazole. Vaccine. Veterinary Medicine/Animal Health in Current Contents(R)/Agricultural, Biology & Environmental Sciences. 2003 week 21 Reprint available from: O'Handley RM. Univ Prince Edward Isl, Atlantic Vet Coll, Dept Pathol & Microbiol, Charlottetown, PE C1A 4P3, Canada.SIn a previous study, Giardia infection patterns were studied in newborn dairy calves over a 4-month period. Chronic Giardia infections were observed in all calves with initial cyst excretion occurring at approximately 1 month of age. In the work presented here, the passive immunity and serological immune response associated with these Giardia infections were examined. Colostrum and milk samples were collected from the dams of these calves, and monthly serum samples were collected from each calf. The colostrum, milk and sera samples were analyzed by ELISA and Western blot for the presence of anti-Giardia IgG antibodies. In addition, the in vitro anti-Giardia activity of milk and colostrum was examined using a miniculture adherence assay. When examined by ELISA, mean anti-Giardia antibody titres were found to be significantly higher in colostrum compared to milk. The monthly mean serum antibody titres in the calves were not found to differ significantly at any time point during the study. Western blot analysis revealed that colostrum from the dams reacted strongly with many different Giardia antigens between 205 and 7.5 kDa, while milk reacted with few antigens in the same size range. Sera collected from the calves when 30 and 60 days of age reacted with few Giardia antigens, but as the calves aged, IgG antibodies in their sera began to react with antigens of 21, 50, 65, 73 and 79 kDa. The miniculture adherence assay demonstrated that colostrum had significantly more anti-Giardia activity in vitro compared to milk. These results suggest that the calves in this dairy did not mount a significant humoral immune response against Giardia following infection. However, colostrum contained a high level of anti-Giardia antibodies and exhibited anti-Giardia activity in vitro. Therefore, colostrum may have the potential to provide initial protection against Giardia infections in calves, but the lack of a strong, specific humoral immune response by these calves could account for the high prevalence and chronic duration of the infections. (C) 2003 Elsevier Science B.V. All rights reserved. [References: 31] 31oEnglish Article Current Contents(R)/Agriculture, Biology & Environmental Sciences. Reprint available from: O'Handley RM Univ Prince Edward Isl, Atlantic Vet Coll, Dept Pathol & Microbiol Charlottetown PE C1A 4P3 Canada Univ Calgary, Fac Med, Dept Gastrointestinal Sci Calgary AB T2N 4N1 Canada Univ Calgary, Dept Biol Sci Calgary AB T2N 4N1 Canada 0001 Vet. Parasitol-671VF-0001 671VF: Document Delivery available 9~?\Ralston, B. J. Cockwill, C. L. Guselle, N. J. Van Herk, F. H. McAllister, T. A. Olson, M. E.2003kPrevalence of Giardia and Cryptosporidium andersoni and their effects on performance in feedlot beef cattle153-159"Canadian Journal of Animal Science831Dairy calves. Parvum oocysts. Farm-animals. Infection. Excretion. Muris. Animal Sciences in Current Contents(R)/Agricultural, Biology & Environmental Sciences. 2003 week 23 Reprint available from: Olson ME. Alberta Dept Agr, Food & Rural Dev, 900 Irricana Rd NE, Airdrie, AB T4A 2G6.8Sixty individually housed Charolais crossbred steers originating from one ranch source had a 12-d (days 0-12) adaptation period in their pens to adjust to their ration and surroundings, followed by two consecutive backgrounding periods (85.5% roughage, 12% concentrate rations) with durations of 84 d (days 13-97) and 63 d (days 98-153), respectively. Steers had a 21-d adaptation period (days 154-174), followed by a 77-d (days 175-257) finishing period (20% roughage, 75% concentrate ration). Fecal samples and animal weights were collected from each steer every 28 d initially, then every 21 d during a test duration of 257 d. Feed weigh-backs were performed weekly for each steer. Fecal samples were processed, and Giardia duodenalis cysts and Cryptosporidium andersoni oocysts were counted. ADG, DMI and FE were calculated for each of the periods (Backgrounding Period 1, Backgrounding Period 2, Finishing Period 3 and Overall). Overall prevalence of C. andersoni and G. duodenalis was 85 and 82%, respectively. There was a decrease (P < 0.05) in the percentage of G. duodenalis infected steers from day 132 to the completion of the trial. The percentage of C. andersoni infected steers decreased (P < 0.05) from day 97 to the completion of the trial (day 257). Shedding of G. duodenalis cysts and C. andersoni oocysts in the feces was intermittent throughout the trial period. A comparison between the ADG, DMI and FE of G. duodenalis infected and non-infected steers demonstrated no overall differences (P > 0.05). A similar comparison between C andersoni infected and non-infected steers showed no overall difference (P > 0.05) with the exception of a lower (P < 0.05) DMI for infected steers. The degree of Giardia or C. andersoni infection observed in the present study did not effect DMI, ADG or FE of feedlot steers. [References: 28] 28English Article Current Contents(R)/Agriculture, Biology & Environmental Sciences. Reprint available from: Olson ME Alberta Dept Agr, Food & Rural Dev 900 Irricana Rd NE Airdrie AB T4A 2G6 Canada Alberta Dept Agr, Food & Rural Dev Airdrie AB T4A 2G6 Canada Univ Calgary, Dept Gastrointestinal Sci Calgary AB T2N 4N1 Canada Agr & Agri Food Canada, Res Ctr Lethbridge AB T1J 4B1 Canada 0022 Can. J. Anim. Sci-675XW-0022 675XW: Document Delivery available~?QOlson, M. E. O'Handley, R. M. Ralston, B. J. McAllister, T. A. Thompson, R. C. A.2004CUpdate on Cryptosporidium and Giardia infections in cattle [Review]185-191Trends in Parasitology204cN. sp apicomplexa. New-york-state. Dairy calves. Molecular characterization. Parvum oocysts. Fenbendazole treatment. Immune-response. United-kingdom. Farm-animals. Beef-calves. Microbiology in Current Contents(R)/Life Sciences. 2004 week 20 Reprint available from: Olson ME. Univ Calgary, Dept Microbiol & Infect Dis, 3900 Hosp Dr NW, Calgary, AB T2N 4N1. Cattle are frequently parasitized with Giardia duodenalis, Cryptosporidium parvum and Cryptosporidium andersoni. These parasites cause diarrhoea and impair gain of body weight. Giardia and Cryptosporidium from cattle are potential zoonotic pathogens, and contact with animals, manure or contaminated water is believed to lead to infections in humans. Molecular epidemiology has suggested that cattle are not as significant a reservoir for human infections as was once believed. Most G. duodenalis from cattle (Assemblage E) are different from those in humans (Assemblages A and B), and C. andersoni does not infect humans. However, molecular tools have shown that humans can be infected with zoonotic C. parvum, as well as anthroponotic Cryptosporidium hominis. [References: 73] 73lEnglish Review Current Contents(R)/Life Sciences. Reprint available from: Olson ME Univ Calgary, Dept Microbiol & Infect Dis 3900 Hosp Dr NW Calgary AB T2N 4N1 Canada Univ Calgary, Dept Microbiol & Infect Dis Calgary AB T2N 4N1 Canada Univ Prince Edward Isl, Atlantic Vet Coll Charlottetown PE C1A 4P3 Canada Agr & Agri Food Canada, Lethbridge Res Ctr Lethbridge AB T1J 4B1 Canada Alberta Agr Food & Rural Dev Airdrie AB T4B 2C1 Canada Murdoch Univ, WHO, Collaborating Ctr Mol Epidemiol Parasit Infect Perth WA 6150 Australia Murdoch Univ, Western Australian Biomed Res Inst Perth WA 6150 Australia 0011 Trends Parasitol-812LW-0011 812LW: Document Delivery available~?zPalmer, C. J. Xiao, L. Terashima, A. Guerra, H. Gotuzzo, E. Saldias, G. Bonilla, J. A. Zhou, L. Lindquist, A. Upton, S. J.2003HCryptosporidium muris, a rodent pathogen, recovered from a human in Peru1174-6Emerg Infect Dis99eAdult Animals Communicable Diseases, Emerging/epidemiology/*parasitology Cryptosporidiosis/diagnosis/*epidemiology Cryptosporidium/genetics/*isolation & purification/pathogenicity Feces/parasitology Female Human Peru/epidemiology Polymerase Chain Reaction Polymorphism, Restriction Fragment Length Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.SeptCryptosporidium muris, predominantly a rodent species of Cryptosporidium, is not normally considered a human pathogen. Recently, isolated human infections have been reported from Indonesia, Thailand, France, and Kenya. We report the first case of C. muris in a human in the Western Hemisphere. This species may be an emerging zoonotic pathogen capable of infecting humans.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14519260&1080-6040 Case Reports Journal Article145192601University of Florida, Gainesville, Florida, USA.~?Ryan, U. O'Hara, A. Xiao, L.2004vMolecular and Biological Characterization of a Cryptosporidium molnari-Like Isolate from a Guppy (Poecilia reticulata)3761-5Appl Environ Microbiol706JunoHistological, morphological, genetic, and phylogenetic analyses of a Cryptosporidium molnari-like isolate from a guppy (Poecilia reticulata) identified stages consistent with those of C. molnari and revealed that C. molnari is genetically very distinct from all other species of Cryptosporidium. This study represents the first genetic characterization of C. molnari.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=151841870099-2240 Journal Article15184187Division of Health Sciences, School of Veterinary and Biomedical Sciences, Murdoch University, Murdoch, Western Australia 6150, Australia. unaryan@central.murdoch.edu.au~?Xiao, L. Ryan, U. M. Graczyk, T. K. Limor, J. Li, L. Kombert, M. Junge, R. Sulaiman, I. M. Zhou, L. Arrowood, M. J. Koudela, B. Modry, D. Lal, A. A.2004=Genetic diversity of Cryptosporidium spp. in captive reptiles891-9Appl Environ Microbiol702Animals Base Sequence Cattle Cryptosporidiosis/microbiology/*veterinary Cryptosporidium/*classification/*genetics/growth & development DNA, Ribosomal/analysis Genotype Lizards/microbiology Molecular Sequence Data Polymerase Chain Reaction Polymorphism, Restriction Fragment Length RNA, Ribosomal/genetics Reptiles/*microbiology Sequence Analysis, DNA Snakes/microbiology Support, Non-U.S. Gov't *Variation (Genetics)Feb)The genetic diversity of Cryptosporidium in reptiles was analyzed by PCR-restriction fragment length polymorphism and sequence analysis of the small subunit rRNA gene. A total of 123 samples were analyzed, of which 48 snake samples, 24 lizard samples, and 3 tortoise samples were positive for Cryptosporidium: Nine different types of Cryptosporidium were found, including Cryptosporidium serpentis, Cryptosporidium desert monitor genotype, Cryptosporidium muris, Cryptosporidium parvum bovine and mouse genotypes, one C. serpentis-like parasite in a lizard, two new Cryptosporidium spp. in snakes, and one new Cryptosporidium sp. in tortoises. C. serpentis and the desert monitor genotype were the most common parasites and were found in both snakes and lizards, whereas the C. muris and C. parvum parasites detected were probably the result of ingestion of infected rodents. Sequence and biologic characterizations indicated that the desert monitor genotype was Cryptosporidium saurophilum. Two host-adapted C. serpentis genotypes were found in snakes and lizards.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=147665690099-2240 Journal Article14766569vDivision of Parasitic Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 30341, USA. lxiao@cdc.gov8~?TRyan, U. M. Samarasinghe, B. Read, C. Buddle, J. R. Robertson, I. D. Thompson, R. C.2003:Identification of a novel Cryptosporidium genotype in pigs3970-4Appl Environ Microbiol697Animals Base Sequence Cryptosporidiosis/epidemiology/parasitology/*veterinary Cryptosporidium/*classification/*genetics/isolation & purification DNA, Protozoan/analysis DNA, Ribosomal/analysis Feces/parasitology Genotype Molecular Sequence Data Parasite Egg Count Phylogeny Prevalence RNA, Ribosomal, 18S/genetics Sequence Analysis, DNA Support, Non-U.S. Gov't Swine/*parasitology Swine Diseases/*epidemiology/parasitologyJulOver a 3-year period, a total of 646 fecal samples from pigs in 22 indoor and outdoor herds from Western Australia were screened for Cryptosporidium spp. by microscopy. Results revealed that 39 of 646 samples (6.03%) were positive for Cryptosporidium. Cryptosporidium was much more common in outdoor herds (17.2%) than in indoor herds (0.5%) and was more common in animals between the ages of 5 and 8 weeks (69.2%) than in younger animals (P < 0.0001). Molecular characterization of the positive samples at the 18S ribosomal DNA locus identified two distinct genotypes of Cryptosporidium: the previously identified pig genotype I and a novel pig genotype (pig genotype II), both of which warrant species status.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=128397690099-2240 Journal Article12839769\Division of Health Sciences, Murdoch University, Murdoch, Western Australia 6150, Australia.~?YRyan, U. M. Xiao, L. Read, C. Sulaiman, I. M. Monis, P. Lal, A. A. Fayer, R. Pavlasek, I.2003cA redescription of Cryptosporidium galli Pavlasek, 1999 (Apicomplexa: Cryptosporidiidae) from birds809-13 J Parasitol894Actins/genetics Animals Base Sequence Bird Diseases/*parasitology Birds Chickens Cluster Analysis Cryptosporidiosis/parasitology/*veterinary Cryptosporidium/*classification/genetics/ultrastructure DNA, Protozoan/chemistry/isolation & purification DNA, Ribosomal/chemistry/isolation & purification Heat-Shock Proteins 70/genetics Molecular Sequence Data Phylogeny RNA, Ribosomal, 18S/genetics Sequence Alignment/veterinary Sequence Analysis, DNA/veterinary SongbirdsAugCryptosporidium galli Pavlasek, 1999, described from the feces of birds, is redescribed with additional molecular and biological data. Oocysts are ellipsoidal, are passed fully sporulated, lack sporocysts, and measure 8.25 x 6.3 microm (range 8.0-8.5 x 6.2-6.4 microm) with a length-width ratio of 1.30 (n = 50). Oocysts are structurally similar to those of Cryptosporidium baileyi described from chickens, but in addition to being considerably larger than oocysts of C. baileyi, these oocysts infect the proventriculus in a variety of birds and not the respiratory tract. Oocysts were successfully transmitted from chickens to chickens, and morphologically similar oocysts also were observed in a variety of exotic and wild birds (Order Passeriformes, Phasianidae, Fringillidae, and Icteridae). Molecular and phylogenetic analyses at the 18S rRNA, HSP70, and actin gene loci demonstrate that this species is genetically distinct from all known species and genotypes of Cryptosporidium and, thus, was named C. galli.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=145336940022-3395 Journal Article14533694Division of Veterinary and Biomedical Sciences, Murdoch University, Murdoch, Western Australia, 6150. unaryan@central.murdoch.edu.auG~?#Sulaiman, I. M. Lal, A. A. Xiao, L.2002bMolecular phylogeny and evolutionary relationships of Cryptosporidium parasites at the actin locus388-94 J Parasitol882TActins/chemistry/*genetics Animals Base Sequence Cryptosporidium/chemistry/*genetics DNA, Protozoan/chemistry/genetics *Evolution, Molecular Human Molecular Sequence Data Oocytes/chemistry *Phylogeny Polymerase Chain Reaction Sequence Analysis, DNA Sequence Homology, Nucleic Acid Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.AprTo further validate previous observations in the taxonomy of Cryptosporidium parasites, the phylogenetic relationship was analyzed among various Cryptosporidium parasites at the actin locus. Nucleotide sequences of the actin gene were obtained from 9 putative Cryptosporidium species (C. parvum, C. andersoni, C. baileyi, C. felis, C. meleagridis, C. muris, C. saurophilum, C. serpentis, and C. wrairi) and various C. parvum genotypes. After multiple alignment of the obtained actin sequences, genetic distances were measured, and phylogenetic trees were constructed. Results of the analysis confirmed the presence of genetically distinct species within Cryptosporidium and various distinct genotypes within C. parvum. The phylogenetic tree constructed on the basis of the actin sequences was largely in agreement with previous results based on small subunit rRNA, 70-kDa heat shock protein, and Cryptosporidium oocyst wall protein genes. The Cryptosporidium species formed 2 major clades; isolates of C. andersoni, C. muris, and C. serpentis formed the first major group, whereas isolates of all other species, as well as various C. parvum genotypes, formed the second major group. Intragenotype variations were low or absent at this locus.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=120540170022-3395 Journal Article12054017Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, U.S. Department of Health and Human Services, Atlanta, Georgia 30341, USA.&~?>Xiao, L. Limor, J. R. Sulaiman, I. M. Duncan, R. B. Lal, A. A.2000IMolecular characterization of a Cryptosporidium isolate from a black bear1166-70 J Parasitol865Animals Base Sequence Bears/*parasitology Cattle Cryptosporidiosis/parasitology/*veterinary Cryptosporidium parvum/*classification/*genetics Dogs Genes, rRNA/genetics Heat-Shock Proteins 70/genetics Human Molecular Sequence Data Polymerase Chain Reaction Polymorphism, Restriction Fragment Length RNA, Ribosomal/genetics Sequence Analysis, DNA Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.OctTo further validate the observation of the existence of host-adapted strains of Cryptosporidium parvum, we genetically characterized an isolate of Cryptosporidium parasite from a black bear. Sequence analysis of the ribosomal RNA small subunit and the 70-kDa heat shock protein (HSP70) showed that this parasite represents a new genotype of C. parvum and is related to the C. parvum dog genotype. This finding is helpful for clarifying Cryptosporidium taxonomy.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=111285050022-3395 Journal Article11128505hDivision of Parasitic Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia 30341, USA.G~?9Jongwutiwes, S. Tiangtip, R. Yentakarm, S. Chantachum, N.2002qSimple method for long-term copro-preservation of Cryptosporidium oocysts for morphometric and molecular analysis257-64Trop Med Int Health73Animals Cryptosporidium/*genetics Ethanol Feces/*parasitology Nucleic Acid Amplification Techniques *Organ Preservation Solutions Polymerase Chain Reaction RNA, Ribosomal/*genetics Sensitivity and Specificity Support, Non-U.S. Gov't Time FactorsMarSPreservation of Cryptosporidium oocysts in faecal specimens containing 75% ethanol is suitable for subsequent morphometric and molecular analysis. No significant morphologic alteration occurred after storage at ambient temperatures, ranging from 22 to 38 degrees C, for more than 2 years. After washing, sugar floatation and DNA extraction, a nested polymerase chain reaction targeting the small subunit ribosomal RNA gene successfully amplified Cryptosporidium DNA in all 15 isolates examined. The sensitivity of detection by polymerase chain reaction (PCR) was found to be as high as 1.25 oocysts per reaction (mean=3.01, SD=1.14). Importantly, a 2.2-kb of the complete DNA sequence of a gene encoding Cryptosporidium thrombospondin-related adhesive protein (TRAP-C1) was also consistently amplified by PCR in all isolates. The PCR-amplified product can be used as a good template for sequencing. Therefore, this simple procedure should be useful for epidemiological analysis of clinical samples from outbreaks, endemic or sporadic cases of cryptosporidiosis when long-term storage of oocysts is required.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=119039881360-2276 Journal Article11903988xDepartment of Parasitology, Faculty of Medicine, Chulalongkorn University, Bangkok, Thailand. fmedsjw@md2.md.chula.ac.th~?(Xiao, L. Fayer, R. Ryan, U. Upton, S. J.2004LCryptosporidium taxonomy: recent advances and implications for public health72-97Clin Microbiol Rev171Animals Cryptosporidium/*classification/genetics/isolation & purification Evolution *Genes, Protozoan Host-Parasite Relations Human Public Health/standards/trends Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S. TerminologyJanThere has been an explosion of descriptions of new species of Cryptosporidium during the last two decades. This has been accompanied by confusion regarding the criteria for species designation, largely because of the lack of distinct morphologic differences and strict host specificity among Cryptosporidium spp. A review of the biologic species concept, the International Code of Zoological Nomenclature (ICZN), and current practices for Cryptosporidium species designation calls for the establishment of guidelines for naming Cryptosporidium species. All reports of new Cryptosporidium species should include at least four basic components: oocyst morphology, natural host specificity, genetic characterizations, and compliance with the ICZN. Altogether, 13 Cryptosporidium spp. are currently recognized: C. muris, C. andersoni, C. parvum, C. hominis, C. wrairi, C. felis, and C. cannis in mammals; C. baileyi, C. meleagridis, and C. galli in birds; C. serpentis and C. saurophilum in reptiles; and C. molnari in fish. With the establishment of a framework for naming Cryptosporidium species and the availability of new taxonomic tools, there should be less confusion associated with the taxonomy of the genus Cryptosporidium. The clarification of Cryptosporidium taxonomy is also useful for understanding the biology of Cryptosporidium spp., assessing the public health significance of Cryptosporidium spp. in animals and the environment, characterizing transmission dynamics, and tracking infection and contamination sources.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1472645610893-8512 Journal Article Review Review, Academic14726456Division of Parasitic Diseases, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Chamblee, Georgia 30341, USA. lxiao@cdc.gov ~?rLeChevallier, M. W. Di Giovanni, G. D. Clancy, J. L. Bukhari, Z. Bukhari, S. Rosen, J. S. Sobrinho, J. Frey, M. M.2003eComparison of method 1623 and cell culture-PCR for detection of Cryptosporidium spp. in source waters971-9Appl Environ Microbiol692Animals Cells, Cultured Comparative Study Cryptosporidium/genetics/*growth & development/*isolation & purification/pathogenicity DNA, Protozoan/analysis/genetics Fluorescent Antibody Technique Fresh Water/*parasitology Human Oocysts/growth & development/isolation & purification Polymerase Chain Reaction/*methods Risk Assessment Seasons Sequence Analysis, DNA Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. *Water SupplyFebAnalysis of Cryptosporidium occurrence in six watersheds by method 1623 and the integrated cell culture-PCR (CC-PCR) technique provided an opportunity to evaluate these two methods. The average recovery efficiencies were 58.5% for the CC-PCR technique and 72% for method 1623, but the values were not significantly different (P = 0.06). Cryptosporidium oocysts were detected in 60 of 593 samples (10.1%) by method 1623. Infectious oocysts were detected in 22 of 560 samples (3.9%) by the CC-PCR technique. There was 87% agreement between the total numbers of samples positive as determined by method 1623 and CC-PCR for four of the sites. The other two sites had 16.3 and 24% correspondence between the methods. Infectious oocysts were detected in all of the watersheds. Overall, approximately 37% of the Cryptosporidium oocysts detected by the immunofluorescence method were viable and infectious. DNA sequence analysis of the Cryptosporidium parvum isolates detected by CC-PCR showed the presence of both the bovine and human genotypes. More than 90% of the C. parvum isolates were identified as having the bovine or bovine-like genotype. The estimates of the concentrations of infectious Cryptosporidium and the resulting daily and annual risks of infection compared well for the two methods. The results suggest that most surface water systems would require, on average, a 3-log reduction in source water Cryptosporidium levels to meet potable water goals.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12571019,0099-2240 Evaluation Studies Journal Article12571019aAmerican Water Works Service Company, Inc., Voorhees, New Jersey 08043, USA. mlecheva@amwater.com ~?]Bukhari, Z. Marshall, M. M. Korich, D. G. Fricker, C. R. Smith, H. V. Rosen, J. Clancy, J. L.2000jComparison of Cryptosporidium parvum viability and infectivity assays following ozone treatment of oocysts2972-80Appl Environ Microbiol667:Animals Animals, Newborn Comparative Study Cryptosporidiosis/parasitology Cryptosporidium parvum/*growth & development/*pathogenicity *Disinfection Dyes/metabolism Fluorescent Dyes/metabolism Great Britain Indoles/metabolism Mice Ozone/*pharmacology Reproducibility of Results Support, Non-U.S. Gov't United StatesJulSeveral in vitro surrogates have been developed as convenient, user-friendly alternatives to mouse infectivity assays for determining the viability of Cryptosporidium parvum oocysts. Such viability assays have been used increasingly to determine oocyst inactivation following treatment with chemical, physical, or environmental stresses. Defining the relationship between in vitro viability assays and oocyst infectivity in susceptible hosts is critical for determining the significance of existing oocyst inactivation data for these in vitro assays and their suitability in future studies. In this study, four viability assays were compared with mouse infectivity assays, using neonatal CD-1 mice. Studies were conducted in the United States and United Kingdom using fresh (<1 month) or environmentally aged (3 months at 4 degrees C) oocysts, which were partially inactivated by ozonation before viability and/or infectivity analyses. High levels of variability were noted within and between the viability and infectivity assays in the U.S. and United Kingdom studies despite rigorous control over oocyst conditions and disinfection experiments. Based on the viability analysis of oocyst subsamples from each ozonation experiment, SYTO-59 assays demonstrated minimal change in oocyst viability, whereas 4',6'-diamidino-2-phenylindole-propidium iodide assays, in vitro excystation, and SYTO-9 assays showed a marginal reduction in oocyst viability. In contrast, the neonatal mouse infectivity assay demonstrated significantly higher levels of oocyst inactivation in the U.S. and United Kingdom experiments. These comparisons illustrate that four in vitro viability assays cannot be used to reliably predict oocyst inactivation following treatment with low levels of ozone. Neonatal mouse infectivity assays should continue to be regarded as a "gold standard" until suitable alternative viability surrogates are identified for disinfection studies.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=108777940099-2240 Journal Article10877794]Clancy Environmental Consultants, Inc., St. Albans, Vermont 05478, USA. zbukhari@together.net ~?6Bukhari, Z. McCuin, R. M. Fricker, C. R. Clancy, J. L.1998dImmunomagnetic separation of Cryptosporidium parvum from source water samples of various turbidities4495-9Appl Environ Microbiol6411Animals Comparative Study Cryptosporidium parvum/*isolation & purification/physiology Immunomagnetic Separation/methods Nephelometry and Turbidimetry Reagent Kits, Diagnostic Support, Non-U.S. Gov't Water/analysis/*parasitology ZygoteNov`Immunomagnetic separation (IMS) procedures which specifically capture Cryptosporidium oocysts and have the potential to isolate oocysts from debris have become commercially available. We compared two IMS kits (kit DB [Dynabeads anti-Cryptosporidium; product no. 730.01; Dynal A.S., Oslo, Norway] and kit IC1 [Crypto Scan IMS; product no. R10; Clearwater Diagnostics Company, LLC, Portland, Maine]) and a modification of kit IC1 (kit IC2 [Crypto Scan IMS; product no. R10; Clearwater Diagnostics Company, LLC]) at three turbidity levels (50, 500, and 5,000 nephelometric turbidity units [ntu]) by using water matrices obtained from different geographical locations. In deionized water, kit DB yielded recoveries between 68 and 83%, whereas the recoveries obtained with kits IC1 and IC2 were more variable and ranged from 0.2 to 74.5%. In water matrices with turbidity levels up to 500 ntu, the oocyst recoveries were more variable with kit DB; however, the recoveries were similar to those obtained in deionized water. In contrast, there were notable reductions in oocyst recoveries in the turbid matrices with kits IC1 and IC2, and the highest recovery (8.3%) was obtained with a 50-ntu sample. An examination of the effects of age on oocyst recovery with kit DB revealed that oocysts up to 16 weeks old yielded recoveries similar to the recoveries observed with fresh oocysts. These data indicate that all IMS kits do not perform equally well, and it is important to conduct in-house quality assurance work before a commercially available IMS kit is selected to replace flotation procedures for recovery of Cryptosporidium oocysts.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=97973130099-2240 Journal Article9797313WClancy Environmental Consultants, St. Albans, Vermont 05478, USA. zbukhari@together.net ~?AFeng, Y. Y. Ong, S. L. Hu, J. Y. Song, L. F. Tan, X. L. Ng, W. J.2003oEffect of particles on the recovery of cryptosporidium oocysts from source water samples of various turbidities1898-903Appl Environ Microbiol694Animals Cryptosporidium parvum/growth & development/*isolation & purification Filtration Fluorescent Antibody Technique Immunomagnetic Separation Laboratory Techniques and Procedures Microscopy, Fluorescence Nephelometry and Turbidimetry Oocysts/*isolation & purification Particle Size United States United States Environmental Protection Agency Water/*parasitology Water Purification/*methods *Water SupplyAprCryptosporidium parvum can be found in both source and drinking water and has been reported to cause serious waterborne outbreaks which threaten public health safety. The U.S. Environmental Protection Agency has developed method 1622 for detection of Cryptosporidium oocysts present in water. Method 1622 involves four key processing steps: filtration, immunomagnetic separation (IMS), fluorescent-antibody (FA) staining, and microscopic evaluation. The individual performance of each of these four steps was evaluated in this study. We found that the levels of recovery of C. parvum oocysts at the IMS-FA and FA staining stages were high, averaging more than 95%. In contrast, the level of recovery declined significantly, to 14.4%, when the filtration step was incorporated with tap water as a spiking medium. This observation suggested that a significant fraction of C. parvum oocysts was lost during the filtration step. When C. parvum oocysts were spiked into reclaimed water, tap water, microfiltration filtrate, and reservoir water, the highest mean level of recovery of (85.0% +/- 5.2% [mean +/- standard deviation]) was obtained for the relatively turbid reservoir water. Further studies indicated that it was the suspended particles present in the reservoir water that contributed to the enhanced C. parvum oocyst recovery. The levels of C. parvum oocyst recovery from spiked reservoir water with different turbidities indicated that particle size and concentration could affect oocyst recovery. Similar observations were also made when silica particles of different sizes and masses were added to seeded tap water. The optimal particle size was determined to be in the range from 5 to 40 micro m, and the corresponding optimal concentration of suspended particles was 1.42 g for 10 liters of tap water.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12676662,0099-2240 Evaluation Studies Journal Article12676662hCenter for Water Research, Department of Civil Engineering, National University of Singapore, Singapore.~?IJohnson, C. H. Marshall, M. M. DeMaria, L. A. Moffet, J. M. Korich, D. G.20036Chlorine inactivation of spores of Encephalitozoon spp1325-6Appl Environ Microbiol692?Animals Cells, Cultured Chlorine/*pharmacology Colony Count, Microbial Disinfection/*methods Encephalitozoon/classification/drug effects/*pathogenicity/*physiology Encephalitozoon cuniculi/drug effects/pathogenicity/physiology Human Kidney/cytology Parasitology/methods Rabbits Spores, Protozoan/drug effects/physiologyFebThis report is an extension of a preliminary investigation on the use of chlorine to inactivate spores of Encephalitozoon intestinalis and to investigate the effect of chlorine on two other species, E cuniculi and E. hellem, associated with human infection. The 50% tissue culture infective doses of these three species were also determined. On the basis of the results obtained, it appears that chlorination of water is an effective means of controlling spores of these organisms in the aquatic environment.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=125710670099-2240 Journal Article12571067XU.S. Environmental Protection Agency, Cincinnati, Ohio 45268, USA. johnson.cliff@epa.gov ~?aRochelle, P. A. Marshall, M. M. Mead, J. R. Johnson, A. M. Korich, D. G. Rosen, J. S. De Leon, R.2002iComparison of in vitro cell culture and a mouse assay for measuring infectivity of Cryptosporidium parvum3809-17Appl Environ Microbiol688tAnimals Caco-2 Cells Cattle Cell Line Comparative Study Cryptosporidiosis/parasitology/*physiopathology Cryptosporidium parvum/*classification/genetics/growth & development/*pathogenicity Disease Models, Animal Dogs Genotype Human Mice Ozone/pharmacology Parasitology/methods Reverse Transcriptase Polymerase Chain Reaction Support, U.S. Gov't, Non-P.H.S. Ultraviolet RaysAugGIn vitro cell cultures were compared to neonatal mice for measuring the infectivity of five genotype 2 isolates of Cryptosporidium parvum. Oocyst doses were enumerated by flow cytometry and delivered to animals and cell monolayers by using standardized procedures. Each dose of oocysts was inoculated into up to nine replicates of 9 to 12 mice or 6 to 10 cell culture wells. Infections were detected by hematoxylin and eosin staining in CD-1 mice, by reverse transcriptase PCR in HCT-8 and Caco-2 cells, and by immunofluorescence microscopy in Madin-Darby canine kidney (MDCK) cells. Infectivity was expressed as a logistic transformation of the proportion of animals or cell culture wells that developed infection at each dose. In most instances, the slopes of the dose-response curves were not significantly different when we compared the infectivity models for each isolate. The 50% infective doses for the different isolates varied depending on the method of calculation but were in the range from 16 to 347 oocysts for CD-1 mice and in the ranges from 27 to 106, 31 to 629, and 13 to 18 oocysts for HCT-8, Caco-2, and MDCK cells, respectively. The average standard deviations for the percentages of infectivity for all replicates of all isolates were 13.9, 11.5, 13.2, and 10.7% for CD-1 mice, HCT-8 cells, Caco-2 cells, and MDCK cells, respectively, demonstrating that the levels of variability were similar in all assays. There was a good correlation between the average infectivity for HCT-8 cells and the results for CD-1 mice across all isolates for untreated oocysts (r = 0.85, n = 25) and for oocysts exposed to ozone and UV light (r = 0.89, n = 29). This study demonstrated that in vitro cell culture was equivalent to the "gold standard," mouse infectivity, for measuring the infectivity of C. parvum and should therefore be considered a practical and accurate alternative for assessing oocyst infectivity and inactivation. However, the high levels of variability displayed by all assays indicated that infectivity and disinfection experiments should be limited to discerning relatively large differences.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12147476,0099-2240 Evaluation Studies Journal Article12147476Water Quality Laboratory, Metropolitan Water District of Southern California, La Verne, California 91750, USA. prochelle@mwdh2o.com~?GKorich, D. G. Mead, J. R. Madore, M. S. Sinclair, N. A. Sterling, C. R.1990kEffects of ozone, chlorine dioxide, chlorine, and monochloramine on Cryptosporidium parvum oocyst viability1423-8Appl Environ Microbiol565Animals Chloramines/*pharmacology Chlorine/*pharmacology *Chlorine Compounds Coccidia/*drug effects Cryptosporidium/*drug effects/physiology Disinfectants/*pharmacology Oxides/*pharmacology Ozone/*pharmacology Support, U.S. Gov't, Non-P.H.S.MayPurified Cryptosporidium parvum oocysts were exposed to ozone, chlorine dioxide, chlorine, and monochloramine. Excystation and mouse infectivity were comparatively evaluated to assess oocyst viability. Ozone and chlorine dioxide more effectively inactivated oocysts than chlorine and monochloramine did. Greater than 90% inactivation as measured by infectivity was achieved by treating oocysts with 1 ppm of ozone (1 mg/liter) for 5 min. Exposure to 1.3 ppm of chlorine dioxide yielded 90% inactivation after 1 h, while 80 ppm of chlorine and 80 ppm of monochloramine required approximately 90 min for 90% inactivation. The data indicate that C. parvum oocysts are 30 times more resistant to ozone and 14 times more resistant to chlorine dioxide than Giardia cysts exposed to these disinfectants under the same conditions. With the possible exception of ozone, the use of disinfectants alone should not be expected to inactivate C. parvum oocysts in drinking water.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=23398940099-2240 Journal Article2339894ODepartment of Microbiology and Immunology, University of Arizona, Tucson 85721.~?FJohnson, D. W. Pieniazek, N. J. Griffin, D. W. Misener, L. Rose, J. B.1995aDevelopment of a PCR protocol for sensitive detection of Cryptosporidium oocysts in water samples3849-55Appl Environ Microbiol6111qAnimals Base Sequence Cryptosporidium/*genetics/*isolation & purification DNA Probes/genetics DNA, Protozoan/genetics Evaluation Studies Molecular Sequence Data Polymerase Chain Reaction/*methods/statistics & numerical data RNA, Ribosomal, 18S/genetics Sensitivity and Specificity Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Water/*parasitology Water SupplyNovThe development of a reliable method of using PCR for detection of Cryptosporidium oocysts in environmental samples with oligonucleotide primers which amplify a portion of the sequence encoding the small (18S) subunit of rRNA producing a 435-bp product was demonstrated. The PCR assay was found to provide highly genus-specific detection of Cryptosporidium spp. after release of nucleic acids from oocysts by a simple freeze-thaw procedure. The assay routinely detected 1 to 10 oocysts in purified oocyst preparations, as shown by direct microscopic counts and by an immunofluorescence assay. The sensitivity of the PCR assay in some seeded environmental water samples was up to 1,000-fold lower. However, this interference was eliminated by either flow cytometry or magnetic-antibody capture. Sensitivity was also improved 10- to 1,000-fold by probing of the PCR product on dot blots with an oligonucleotide probe detected by chemiluminescence. Confirmation of the presence of Cryptosporidium oocysts in water samples from the outbreak in Milwaukee, Wis., was obtained with this technique, and PCR was found to be as sensitive as immunofluorescence for detection of oocysts in wastewater concentrates.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=85264960099-2240 Journal Article8526496ZDepartment of Marine Science, University of South Florida, St. Petersburg 33701-5016, USA. ~?WSturbaum, G. D. Klonicki, P. T. Marshall, M. M. Jost, B. H. Clay, B. L. Sterling, C. R.2002Immunomagnetic separation (IMS)-fluorescent antibody detection and IMS-PCR detection of seeded Cryptosporidium parvum oocysts in natural waters and their limitations2991-6Appl Environ Microbiol686Animals Cryptosporidium parvum/*isolation & purification DNA, Protozoan/*analysis Fluorescent Antibody Technique Immunomagnetic Separation/*methods Polymerase Chain Reaction Support, Non-U.S. Gov'tJun{Detection and enumeration of Cryptosporidium parvum in both treated and untreated waters are important to facilitate prevention of future cryptosporidiosis incidents. Immunomagnetic separation (IMS)-fluorescent antibody (FA) detection and IMS-PCR detection efficiencies were evaluated in two natural waters seeded with nominal seed doses of 5, 10, and 15 oocysts. IMS-FA detected oocysts at concentrations at or below the three nominal oocyst seed doses, illustrating that IMS-FA is sensitive enough to detect low oocyst numbers. However, the species of the oocysts could not be determined with this technique. IMS-PCR, targeting the 18S rRNA gene in this study, yielded positive amplification for 17 of the 18 seeded water samples, and the amplicons were subjected to restriction fragment length polymorphism digestion and DNA sequencing for species identification. Interestingly, the two unseeded, natural water samples were also PCR positive; one amplicon was the same base pair size as the C. parvum amplicon, and the other amplicon was larger. These two amplified products were determined to be derived from DNA of Cryptosporidium muris and a dinoflagellate. These IMS-PCR results illustrate that (i) IMS-PCR is able to detect low oocyst numbers in natural waters, (ii) PCR amplification alone is not confirmatory for detection of target DNA when environmental samples are used, (ii) PCR primers, especially those designed against the rRNA gene region, need to be evaluated for specificity with organisms closely related to the target organism, and (iv) environmental amplicons should be subjected to appropriate species-specific confirmatory techniques.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=120397590099-2240 Journal Article12039759eDepartment of Veterinary Science and Microbiology, University of Arizona, Tucson, Arizona 85721, USA.~?6Reed, C. Sturbaum, G. D. Hoover, P. J. Sterling, C. R.2002hCryptosporidium parvum mixed genotypes detected by PCR-restriction fragment length polymorphism analysis427-9Appl Environ Microbiol681Animals Cryptosporidium parvum/*classification/*genetics/growth & development DNA, Ribosomal/analysis Genotype *Polymerase Chain Reaction *Polymorphism, Restriction Fragment Length RNA, Ribosomal, 18S/genetics Sensitivity and SpecificityJanCombinations of 10 Cryptosporidium parvum oocysts, with various ratios of genotype I to genotype II, were isolated and subjected to PCR-restriction fragment length polymorphism analysis. Amplification of both genotypes in these samples ranged from 31 to 74% and yielded no information about the genotype proportions. In addition, since both genotypes were not always detected, amplification of a single genotype is not conclusive evidence that the sample contains only a single genotype.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11772657,0099-2240 Evaluation Studies Journal Article11772657eDepartment of Veterinary Science and Microbiology, University of Arizona, Tucson, Arizona 85721, USA. 9~?RSturbaum, G. D. Reed, C. Hoover, P. J. Jost, B. H. Marshall, M. M. Sterling, C. R.2001xSpecies-specific, nested PCR-restriction fragment length polymorphism detection of single Cryptosporidium parvum oocysts2665-8Appl Environ Microbiol676$Animals Cryptosporidium parvum/genetics/*isolation & purification Micromanipulation Polymerase Chain Reaction/*methods *Polymorphism, Restriction Fragment Length RNA, Protozoan/genetics/isolation & purification RNA, Ribosomal, 18S/genetics/isolation & purification Sensitivity and SpecificityJunConcurrent with recent advances seen with Cryptosporidium parvum detection in both treated and untreated water is the need to properly evaluate these advances. A micromanipulation method by which known numbers of C. parvum oocysts, even a single oocyst, can be delivered to a test matrix for detection sensitivity is presented. Using newly developed nested PCR-restriction fragment length polymorphism primers, PCR sensitivity was evaluated with 1, 2, 3, 4, 5, 7, or 10 oocysts. PCR detection rates (50 samples for each number of oocysts) ranged from 38% for single oocysts to 92% for 5 oocysts, while 10 oocysts were needed to achieve 100% detection. The nested PCR conditions amplified products from C. parvum, Cryptosporidium baileyi, and Cryptosporidium serpentis but no other Cryptosporidium sp. or protozoan tested. Restriction enzyme digestion with VspI distinguished between C. parvum genotypes 1 and 2. Restriction enzyme digestion with DraII distinguished C. parvum from C. baileyi and C. serpentis. Use of known numbers of whole oocysts encompasses the difficulty of liberating DNA from the oocyst and eliminates the standard deviation inherent within a dilution series. To our knowledge this is the first report in which singly isolated C. parvum oocysts were used to evaluate PCR sensitivity. This achievement illustrates that PCR amplification of a single oocyst is feasible, yet sensitivity remains an issue, thereby illustrating the difficulty of dealing with low oocyst numbers when working with environmental water samples.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=113751780099-2240 Journal Article11375178eDepartment of Veterinary Science and Microbiology, University of Arizona, Tucson, Arizona 85721, USA.7~?,Deng, M. Rutherford, M. S. Abrahamsen, M. S.2004=Host intestinal epithelial response to Cryptosporidium parvum869-84Adv Drug Deliv Rev566Apr 19fCryptosporidium parvum is an obligate intracellular protozoan parasite that is a well-recognized cause of diarrhea in humans and animals throughout the world, and is associated with a substantial degree of morbidity and mortality in patients with the acquired immunodeficiency syndrome (AIDS). C. parvum primarily infects epithelial cells of the gastrointestinal tract, resulting in acute watery diarrhea for which there is no effective therapy. During infection, all parasite development, sexual or asexual, occurs within epithelial cells of the host. This requires a unique and complex association between two distinct eukaryotic organisms. Conversely, due to the intracellular nature of C. parvum, epithelial cells appear to play a key role in activating and communicating with the host immune system. Delineation of the biochemical processes that are regulated within infected epithelial cells is crucial for understanding the pathology of C. parvum infection, the process by which the host clears and ultimately develops resistance to infection, and the development of chemotherapeutic strategies to intercede infections.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=150635950169-409x Journal Article15063595Department of Veterinary PathoBiology, College of Veterinary Medicine, University of Minnesota, 1988 Fitch Avenue, St. Paul, MN 55108, USA.~?uMorgan-Ryan, U. M. Fall, A. Ward, L. A. Hijjawi, N. Sulaiman, I. Fayer, R. Thompson, R. C. Olson, M. Lal, A. Xiao, L.2002QCryptosporidium hominis n. sp. (Apicomplexa: Cryptosporidiidae) from Homo sapiens433-40J Eukaryot Microbiol496KAnimals Cats Cattle Cryptosporidiosis/*parasitology/physiopathology/transmission Cryptosporidium/*classification/genetics/growth & development/*pathogenicity Dogs Female Genotype Germ-Free Life Human Male Mice Oocysts/genetics/pathogenicity Protozoan Proteins/genetics Rats Species Specificity Support, U.S. Gov't, Non-P.H.S. SwineNov-Dec2The structure and infectivity of the oocysts of a new species of Cryptosporidium from the feces of humans are described. Oocysts are structurally indistinguishable from those of Cryptosporidium parvum. Oocysts of the new species are passed fully sporulated, lack sporocysts. and measure 4.4-5.4 microm (mean = 4.86) x 4.4-5.9 microm (mean = 5.2 microm) with a length to width ratio 1.0-1.09 (mean 1.07) (n = 100). Oocysts were not infectious for ARC Swiss mice, nude mice. Wistar rat pups, puppies, kittens or calves, but were infectious to neonatal gnotobiotic pigs. Pathogenicity studies in the gnotobiotic pig model revealed significant differences in parasite-associated lesion distribution (P = 0.005 to P = 0.02) and intensity of infection (P = 0.04) between C. parvum and this newly described species from humans. In vitro cultivation studies have also revealed growth differences between the two species. Multi-locus analysis of numerous unlinked loci, including a preliminary sequence scan of the entire genome demonstrated this species to be distinct from C. parvum and also demonstrated a lack of recombination, providing further support for its species status. Based on biological and molecular data, this Cryptosporidium infecting the intestine of humans is proposed to be a new species Cryptosporidium hominis n. sp.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=125036761066-5234 Journal Article12503676Division of Veterinary and Biomedical Sciences, Murdoch University, Murdoch, Western Australia 6150. unaryan@central.murdoch.edu.au ~?XDuPont, H. L. Chappell, C. L. Sterling, C. R. Okhuysen, P. C. Rose, J. B. Jakubowski, W.1995?The infectivity of Cryptosporidium parvum in healthy volunteers855-9 N Engl J Med33213WAdult Animals Cryptosporidiosis/*parasitology/transmission Cryptosporidium parvum/isolation & purification/*pathogenicity Disease Susceptibility Feces/parasitology Female Human Linear Models Male Middle Aged Parasite Egg Count Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S. Water/*parasitology Water SupplyMar 30-BACKGROUND. Small numbers of Cryptosporidium parvum oocysts can contaminate even treated drinking water, and ingestion of oocysts can cause diarrheal disease in normal as well as immunocompromised hosts. Since the number of organisms necessary to cause infection in humans is unknown, we performed a study to determine the infective dose of the parasite in healthy adults. METHODS. After providing informed consent, 29 healthy volunteers without evidence of previous C. parvum infection, as determined by the absence of anti-cryptosporidium-specific antibodies, were given a single dose of 30 to 1 million C. parvum oocysts obtained from a calf. They were then monitored for oocyst excretion and clinical illness for eight weeks. Household contacts were monitored for secondary spread. RESULTS. Of the 16 subjects who received an intended dose of 300 or more oocysts, 14 (88 percent) became infected. After a dose of 30 oocysts, one of five subjects (20 percent) became infected, whereas at a dose of 1000 or more oocysts, seven of seven became infected. The median infective dose, calculated by linear regression, was 132 oocysts. Of the 18 subjects who excreted oocysts after the challenge dose, 11 had enteric symptoms and 7 (39 percent) had clinical cryptosporidiosis, consisting of diarrhea plus at least one other enteric symptom. All recovered, and there were no secondary cases of diarrhea among household contacts. CONCLUSIONS. In healthy adults with no serologic evidence of past infection with C. parvum, a low dose of C. parvum oocysts is sufficient to cause infection.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=78701400028-4793 Journal Article7870140,University of Texas Medical School, Houston.@~? DuPont, H. L.1985&Cryptosporidiosis and the healthy host1319-20 N Engl J Med31220>Animals *Cryptosporidiosis/transmission Human ImmunocompetenceMay 16dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=39907290028-4793 Editorial3990729~?_Lowery, C. J. Moore, J. E. Millar, B. C. Burke, D. P. McCorry, K. A. Crothers, E. Dooley, J. S.2000Detection and speciation of Cryptosporidium spp. in environmental water samples by immunomagnetic separation, PCR and endonuclease restriction779-85J Med Microbiol499Animals Cryptosporidiosis/parasitology Cryptosporidium/classification/genetics/*isolation & purification Cryptosporidium parvum/classification/genetics/isolation & purification DNA Restriction Enzymes/metabolism DNA, Ribosomal/isolation & purification Human Immunomagnetic Separation/*methods Polymerase Chain Reaction/*methods RNA, Ribosomal, 18S/genetics Sensitivity and Specificity Support, Non-U.S. Gov't Water/*parasitologySepCurrent methods for the detection of Cryptosporidium oocysts in water samples are both time-consuming and subject to variation in sensitivity. A genus-specific PCR assay was designed for the specific amplification of a 552-bp region of the 18S rRNA gene. Postamplification endonuclease restriction generated unique digest patterns that enabled differentiation between the three species, C. muris, C. baileyi and C. parvum, the major human pathogen. Theoretical restriction profiles for other Cryptosporidium species were also predicted. The assay routinely detected 10 oocysts in 10-ml purified oocyst preparations, but sensitivity was found to be 10(3)-10(4) -fold lower in environmental water samples. The use of Chelex resin and an immunomagnetic separation procedure overcame this inhibition. This provided detection levels of 10(1)-10(3) oocysts, depending on water turbidity. Rapid and sensitive pathogen detection methods are essential for the water industry. The results of this study demonstrate that PCR has the potential to improve current detection capabilities greatly by differentiating the major human pathogens from non-pathogenic species. This will greatly facilitate a closer examination of the epidemiology of this important pathogen.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=109662250022-2615 Journal Article10966225jDepartment of Applied Biological and Chemical Sciences, University of Ulster, Coleraine, Northern Ireland. ~?Mac Kenzie, W. R. Hoxie, N. J. Proctor, M. E. Gradus, M. S. Blair, K. A. Peterson, D. E. Kazmierczak, J. J. Addiss, D. G. Fox, K. R. Rose, J. B. et al.,1994hA massive outbreak in Milwaukee of cryptosporidium infection transmitted through the public water supply161-7 N Engl J Med3313<Adolescent Adult Aged Animals Child Cryptosporidiosis/*epidemiology/transmission Cryptosporidium/isolation & purification Diarrhea/epidemiology/parasitology *Disease Outbreaks Female Gastrointestinal Diseases/*epidemiology/parasitology Human Male Middle Aged Seasons Urban Health *Water Supply Wisconsin/epidemiologyJul 21<BACKGROUND. Early in the spring of 1993 there was a widespread outbreak of acute watery diarrhea among the residents of Milwaukee. METHODS. We investigated the two Milwaukee water-treatment plants, gathered data from clinical laboratories on the results of tests for enteric pathogens, and examined ice made during the time of the outbreak for cryptosporidium oocysts. We surveyed residents with confirmed cryptosporidium infection and a sample of those with acute watery diarrhea consistent with cryptosporidium infection. To estimate the magnitude of the outbreak, we also conducted a survey using randomly selected telephone numbers in Milwaukee and four surrounding counties. RESULTS. There were marked increases in the turbidity of treated water at the city's southern water-treatment plant from March 23 until April 9, when the plant was shut down. Cryptosporidium oocysts were identified in water from ice made in southern Milwaukee during these weeks. The rates of isolation of other enteric pathogens remained stable, but there was more than a 100-fold increase in the rate of isolation of cryptosporidium. The median duration of illness was 9 days (range, 1 to 55). The median maximal number of stools per day was 12 (range, 1 to 90). Among 285 people surveyed who had laboratory-confirmed cryptosporidiosis, the clinical manifestations included watery diarrhea (in 93 percent), abdominal cramps (in 84 percent), fever (in 57 percent), and vomiting (in 48 percent). We estimate that 403,000 people had watery diarrhea attributable to this outbreak. CONCLUSIONS. This massive outbreak of watery diarrhea was caused by cryptosporidium oocysts that passed through the filtration system of one of the city's water-treatment plants. Water-quality standards and the testing of patients for cryptosporidium were not adequate to detect this outbreak.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=78186400028-4793 Journal Article7818640EWisconsin Division of Health, Bureau of Public Health, Madison 53703. ~?4McCuin, R. M. Bukhari, Z. Sobrinho, J. Clancy, J. L.2001tRecovery of Cryptosporidium oocysts and Giardia cysts from source water concentrates using immunomagnetic separation69-76J Microbiol Methods452qAge Factors Animals Antigens, Protozoan/analysis California Cryptosporidiosis/prevention & control Cryptosporidium parvum/*isolation & purification Giardia lamblia/*isolation & purification Giardiasis/prevention & control Heat *Immunomagnetic Separation Microscopy, Fluorescence Nebraska Pennsylvania Reagent Kits, Diagnostic Support, Non-U.S. Gov't Water/*parasitologyJunImmunomagnetic separation (IMS) procedures for the simultaneous isolation of Cryptosporidium oocysts and Giardia cysts have recently become available. We validated Dynal's GC-Combo IMS kit using source water at three turbidity levels (5000, 500 and 50 nephelometric turbidity units [ntu]) obtained from different geographical locations and spiked with approximately 9--11 (oo)cysts per ml. Mean recoveries of Cryptosporidium oocysts and Giardia cysts in deionized water were 62% and 69%, respectively. In turbid water matrices, mean recoveries of Cryptosporidium oocysts were between 55.9% and 83.1% while mean recoveries of cysts were between 61.1% and 89.6%. Marginally higher recoveries of the heat inactivated (oo)cysts were observed (119.4% Cryptosporidium oocysts and 90.9% Giardia cysts) in deionized water when compared with recoveries of viable (oo)cysts (69.7% Cryptosporidium oocysts and 79% Giardia cysts). Age of (oo)cysts on recoveries using the GC-Combo IMS kit demonstrated no effects up to 20 months old. Recovery of Giardia cysts was consistent for isolates aged up to 8 months (81.4%), however, a significant reduction in recoveries was noted at 20 months age. Recoveries of low levels (5 and 10 (oo)cysts) of Cryptosporidium oocysts and Giardia cysts in deionized water using IMS ranged from 51.3% to 78% and from 47.6% to 90.0%, respectively. Results of this study indicate that Dynal's GC-Combo IMS kit is an efficient technique to separate Cryptosporidium/Giardia from turbid matrices and yields consistent, reproducible recoveries. The use of fresh (recently voided and purified) (oo)cysts, aged (oo)cysts, viable and heat-inactivated (oo)cysts indicated that these parameters do not influence IMS performance.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=113113910167-7012 Journal Article11311391NClancy Environmental Consultants, Inc., PO Box 314, St. Albans, VT 05478, USA.~?eMorgan, U. Weber, R. Xiao, L. Sulaiman, I. Thompson, R. C. Ndiritu, W. Lal, A. Moore, A. Deplazes, P.2000Molecular characterization of Cryptosporidium isolates obtained from human immunodeficiency virus-infected individuals living in Switzerland, Kenya, and the United States1180-3J Clin Microbiol383AIDS-Related Opportunistic Infections/*diagnosis/parasitology Acetate-CoA Ligase/genetics Animals Cryptosporidiosis/*diagnosis Cryptosporidium/classification/*genetics/isolation & purification DNA, Protozoan/genetics DNA, Ribosomal/genetics HIV Infections/complications/transmission Heat-Shock Proteins 70/genetics Homosexuality, Male Human Kenya Male RNA, Protozoan/genetics RNA, Ribosomal, 18S/genetics Substance Abuse, Intravenous Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S. Switzerland United StatesMar/A total of 22 Cryptosporidium isolates from human immunodeficiency virus-infected patients from Kenya, Switzerland, and the United States were examined at three genetic loci: the 18S ribosomal DNA, HSP-70, and acetyl coenzyme A synthetase genes. Four distinct Cryptosporidium genotypes were identified: (i) the Cryptosporidium parvum "human" genotype, (ii) the C. parvum "cattle" genotype, (iii) Cryptosporidium felis, and (iv) Cryptosporidium meleagridis. This is the first report of C. meleagridis in a human host. These results and those of others indicate that immunocompromised individuals are susceptible to a wide range of Cryptosporidium species and genotypes. Future studies are required to understand the full public health significance of Cryptosporidium genotypes and species in immunocompromised hosts.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=106990170095-1137 Journal Article10699017World Health Organisation Collaborating Centre for the Molecular Epidemiology of Parasitic Infections and State Agricultural Biotechnology Centre, Murdoch University, Murdoch, WA 6150, Australia. morgan@numbat.murdoch.edu.au~?Morgan, U. M. Sturdee, A. P. Singleton, G. Gomez, M. S. Gracenea, M. Torres, J. Hamilton, S. G. Woodside, D. P. Thompson, R. C.1999IThe Cryptosporidium "mouse" genotype is conserved across geographic areas1302-5J Clin Microbiol375Acetate-CoA Ligase/genetics Animals Cattle/parasitology Cryptosporidium parvum/classification/*genetics DNA, Ribosomal/chemistry Genotype Human Mice/*parasitology Phylogeny RNA, Ribosomal, 18S/genetics Support, Non-U.S. Gov'tMaysA 298-bp region of the Cryptosporidium parvum 18S rRNA gene and a 390-bp region of the acetyl coenzyme A synthetase gene were sequenced for a range of Cryptosporidium isolates from wild house mice (Mus domesticus), a bat (Myotus adversus), and cattle from different geographical areas. Previous research has identified a distinct genotype, referred to as the "mouse"-derived Cryptosporidium genotype, common to isolates from Australian mice. Comparison of a wider range of Australian mouse isolates with United Kingdom and Spanish isolates from mice and cattle and also an Australian bat-derived Cryptosporidium isolate revealed that the "mouse" genotype is conserved across geographic areas. Mice are also susceptible to infection with the "cattle" Cryptosporidium genotype, which has important implications for their role as reservoirs of infection for humans and domestic animals.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=102034750095-1137 Journal Article10203475World Health Organisation Collaborating Centre for the Molecular Epidemiology of Parasitic Infections, School of Veterinary and Biomedical Sciences, Murdoch University, Murdoch, Western Australia, Australia. morgan@numbat.murdoch.edu.au~?uSpano, F. Putignani, L. Crisanti, A. Sallicandro, P. Morgan, U. M. Le Blancq, S. M. Tchack, L. Tzipori, S. Widmer, G.1998nMultilocus genotypic analysis of Cryptosporidium parvum isolates from different hosts and geographical origins3255-9J Clin Microbiol3611+Animals Australia/epidemiology Base Sequence Cryptosporidiosis/epidemiology/*parasitology/transmission Cryptosporidium parvum/classification/*genetics/*isolation & purification DNA Primers/genetics DNA, Protozoan/genetics/isolation & purification Epidemiology, Molecular Europe/epidemiology Feces/parasitology Genes, Protozoan Genetic Markers Genotype Human North America/epidemiology Polymerase Chain Reaction Polymorphism (Genetics) Polymorphism, Restriction Fragment Length South America/epidemiology Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S.NovOThe genetic analysis of oocysts recovered from the stools of humans and animals infected with Cryptosporidium parvum has consistently shown the existence of two distinct genotypes. One of the genotypes is found exclusively in some human infections, whereas the other genotype is found in human as well as in animal infections. On the basis of these observations and the results of published epidemiological studies with single polymorphic markers, the existence of two separate transmission cycles has been postulated, one exclusively anthroponotic and the other involving both animals and humans. To test this hypothesis, C. parvum isolates of different geographic and host origins were analyzed by using unlinked genetic polymorphisms. A total of 28 isolates originating from Europe, North and South America, and Australia were examined. Isolates clustered into two groups, one comprising both human and animal isolates and the other comprising isolates only of human origin. The absence of recombinant genotypes is consistent with two reproductively isolated populations within the species C. parvum.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=97745750095-1137 Journal Article9774575PIstituto di Parassitologia, Universita di Roma "La Sapienza," 00185 Rome, Italy. ~?MMorgan, U. M. Pallant, L. Dwyer, B. W. Forbes, D. A. Rich, G. Thompson, R. C.1998qComparison of PCR and microscopy for detection of Cryptosporidium parvum in human fecal specimens: clinical trial995-8J Clin Microbiol364Animals Comparative Study Cryptosporidium parvum/*isolation & purification Feces/*parasitology Human Microscopy *Polymerase Chain Reaction Sensitivity and Specificity Support, Non-U.S. Gov'tAprPCR technology offers alternatives to conventional diagnosis of Cryptosporidium for both clinical and environmental samples. We compared microscopic examination by a conventional acid-fast staining procedure with a recently developed PCR test that can not only detect Cryptosporidium but is also able to differentiate between what appear to be host-adapted genotypes of the parasite. Examinations were performed on 511 stool specimens referred for screening on the basis of diarrhea. PCR detected a total of 36 positives out of the 511 samples, while routine microscopy detected 29 positives. Additional positives detected by PCR were eventually confirmed to be positive by microscopy. A total of five samples that were positive by routine microscopy at Western Diagnostic Pathology but negative by PCR and by microscopy in our laboratory were treated as false positives. Microscopy therefore exhibited 83.7% sensitivity and 98.9% specificity compared to PCR. PCR was more sensitive and easier to interpret but required more hands-on time to perform and was more expensive than microscopy. PCR, however, was very adaptable to batch analysis, reducing the costs considerably. Bulk buying of reagents and modifications to the procedure would decrease the cost of the PCR test even more. An important advantage of the PCR test, its ability to directly differentiate between different Cryptosporidium genotypes, will assist in determining the source of cryptosporidial outbreaks. Sensitivity, specificity, ability to genotype, ease of use, and adaptability to batch testing make PCR a useful tool for future diagnosis and studies on the molecular epidemiology of Cryptosporidium infections.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9542924(0095-1137 Clinical Trial Journal Article9542924World Health Organisation Collaborating Center for the Molecular Epidemiology of Infectious Diseases, Division of Veterinary and Biomedical Sciences, Murdoch University, WA, Australia. morgan@numbat.murdoch.edu.au ~?#Strong, W. B. Gut, J. Nelson, R. G.2000Cloning and sequence analysis of a highly polymorphic Cryptosporidium parvum gene encoding a 60-kilodalton glycoprotein and characterization of its 15- and 45-kilodalton zoite surface antigen products4117-34 Infect Immun687fAlleles Amino Acid Sequence Animals Antibodies, Monoclonal Antigens, Protozoan/chemistry/*genetics/metabolism Antigens, Surface/chemistry/*genetics/metabolism Base Sequence Cloning, Molecular Cryptosporidiosis/parasitology Cryptosporidium parvum/*genetics/*immunology/pathogenicity DNA Primers/genetics DNA, Protozoan/genetics Gene Expression Regulation, Developmental *Genes, Protozoan Genotype Human Molecular Sequence Data Molecular Weight Polymorphism (Genetics) Protein Processing, Post-Translational Protozoan Proteins/chemistry/*genetics/*immunology Sequence Homology, Amino Acid Support, U.S. Gov't, P.H.S.JulThe apicomplexan parasite Cryptosporidium parvum is a major cause of serious diarrheal disease in both humans and animals. No efficacious chemo- or immunotherapies have been identified for cryptosporidiosis, but certain antibodies directed against zoite surface antigens and/or proteins shed by gliding zoites have been shown to neutralize infectivity in vitro and/or to passively protect against, or ameliorate, disease in vivo. We previously used monoclonal antibody 11A5 to identify a 15-kDa surface glycoprotein that was shed behind motile sporozoites and was recognized by several lectins that neutralized parasite infectivity for cultured epithelial cells. Here we report the cloning and sequence analysis of the gene encoding this 11A5 antigen. Surprisingly, the gene encoded a 330-amino-acid, mucin-like glycoprotein that was predicted to contain an N-terminal signal peptide, a homopolymeric tract of serine residues, 36 sites of O-linked glycosylation, and a hydrophobic C-terminal peptide specifying attachment of a glycosylphosphatidylinositol anchor. The single-copy gene lacked introns and was expressed during merogony to produce a 60-kDa precursor which was proteolytically cleaved to 15- and 45-kDa glycoprotein products that both localized to the surface of sporozoites and merozoites. The gp15/45/60 gene displayed a very high degree of sequence diversity among C. parvum isolates, and the numerous single-nucleotide and single-amino-acid polymorphisms defined five to six allelic classes, each characterized by additional intra-allelic sequence variation. The gp15/45/60 single-nucleotide polymorphisms will prove useful for haplotyping and fingerprinting isolates and for establishing meaningful relationships between C. parvum genotype and phenotype.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=108582290019-9567 Journal Article10858229Division of Infectious Diseases, San Francisco General Hospital, University of California, San Francisco, San Francisco, California 94143-0811, USA.~?Lisle, J. T. Rose, J. B.1995GCryptosporidium contamination of water in the USA and UK: A mini-review103-117 AQUA, vol. 443:USA; drinking water; public health; water treatment; water quality control; bacteria; pathogens; disinfection; resistance; parasites; parasitic diseases; human diseases; disease transmission; hazard assessment; water supply; microbial contamination; water purification; USA; British Isles; Cryptosporidium Cryptosporidium SW 3020 Sources and fate of pollution; Q5 01524 Public health, medicines, dangerous organisms; H SE3.21 WATER POLLUTION/WATER QUALITY; P 2000 FRESHWATER POLLUTION Environmental Quality; Health & Safety Science Abstracts; Pollution Abstracts+During the past 10 years the protozoan parasite Cryptosporidium has been recognised as a public health threat in drinking waters. Recently, the largest outbreak to date occurred in Milwaukee, Wisconsin, USA. Over 1.5 million consumers were exposed to this intestinal pathogen, of which 403 000 became ill. Many of those who were immunocompromised died. The probability of an outbreak of cryptosporidiosis occurring in drinking water systems, relative to that of bacterial and viral pathogens, is increased due to the resistant nature of oocysts to concentrations of disinfectants routinely used in drinking-water treatment. Surveys of surface and drinking waters in the USA and UK have shown Cryptosporidium oocysts to be present in polluted, pristine and drinking waters at concentrations that may put the consumer at risk of infection, based upon current risk assessment models. This mini-review is an attempt to present the most recent literature concerning Cryptosporidium in regard to outbreaks, occurrence, monitoring and detection, and regulatory implications.CISSN: 0003-7214 Language: English Summary Language: English; French3848674<Univ. South Florida, Dep. Mar. Sci., St. Petersburg, FL, USA>?Gale, P. Stanfield, G.2000+Cryptosporidium during a simulated outbreak105-116/Journal of the American Water Works Association929Cryptosporidium; Protozoa; Viruses; Pathogens; Diseases; Oocysts; Treated Water; Intestinal protozoa; Viruses (-general-) (see also Individ Groups); Pathogenic organism; Diseases (see also Individual groups); Oocytes; Water supplies; Outbreaks; Water treatment; Monitoring; Risk assessment; Cryptosporidium SW 3060 Water treatment and distribution; AQ 00004 Water Treatment; K 03099 PollutionSep[Protozoa and viruses have in recent years replaced bacterial pathogens as the agents of primary concern in waterborne disease. Unfortunately, so-called "spot" sampling for Cryptosporidium may underestimate the risk of infection. Continuous large-volume sampling of treated water may help overcome the variation in oocyst counts in treated water. Gale and Stanfield provide a model to help utility managers interpret data during outbreaks and to design strategies to monitor for Cryptosporidium. The authors' model also helps explain why some, and often all, samples collected during an outbreak contain zero oocysts. They show that there is no clear association between measured oocyst densities in the drinking water supply and the outbreak of illness. Instead they emphasize the need to accommodate the spatial and temporal variation in oocyst counts in treated water in order to fully assess the net oocyst removal by treatment processes. The model they present promises to enhance water professionals' understanding of the association between measured oocyst densities and the risk of illness in the population.;ISSN: 0003-150X Language: English Summary Language: EnglishJ. Am. Water Works Assoc.4858548WRc-NSF Ltd., Bucks, England 5D|?Stanfield, G. Carrington, E. Albinet, F. Compagnon, B. Dumoutier, N. Hambsch, B. Lorthioy, A. Medema, G. Pezoldt, H. De Roubin, M. R. De Lohman, A. Whitmore, T.2000qAn optimised and standardised test to determine the presence of the protozoa Cryptosporidium and Giardia in waterInternational Conference on Minimising Risk from Cryptosporidium and Other Waterborne Particles, Paris (France), 19-23 Apr 1999Rapinat, M. Weitz, E.Water sampling; Protozoa; Research programs; Filters; Flocculation; Purification; Microbial contamination; Sampling methods; Cryptosporidium; Giardia; Europe; Water pollution; Water Analysis; Microbiological Studies; Analytical Methods; Testing Procedures; Intestinal protozoa; Flagellates (Intestinal); Analytical techniques; Water filtration; River water; Fresh water; Pollution detection; Microbiological analysis; Cysts; Microorganisms; Protozoan diseases; Human diseases; Public health; Risks; Cryptosporidium; Giardia; Europe standardization; purification; optimization; Ferric sulphate P 2000 FRESHWATER POLLUTION; K 03099 Pollution; SW 3010 Identification of pollutants; AQ 00002 Water Quality; Q5 01502 Methods and instrumentsWith funding from the European Commission, a consortium of members of the European Water Research Institutes is carrying out a programme of work with the objective of optimising and standardising a method for determining the presence in water of (oo)cysts of Cryptosporidium and Giardia. Each of the stages of the conventional analysis procedure (initial concentration, recovery and identification and enumeration) are being investigated and the relative merits of existing and new methods are being assessed. Newly developed filters (Envirochek and Filta-Max) have been shown to be more efficient for initial recovery of (oo)cysts from water than the previously used Cuno cartridge filters. In addition, for the analysis of raw waters, flocculation with ferric sulphate has been shown to give recoveries similar to the Envirochek and Filta Max. Modern purification systems such as immunomagnetic separation have also been assessed and found to offer some advantages over flotation although optimisation of the latter has brought improved efficiency. Preliminary assessment of solid phase cytometry has indicated that this technique could offer significant time savings compared to conventional microscopic counting. The results of the study will be used to propose a revised standard method to CEN.MINIMISING RISK FROM CRYPTOSPORIDIUM AND OTHER WATERBORNE PARTICLES. pp. 103-110. Water Science & Technology [Water Sci. Technol.]. Vol. 41, no. 7. Language: English Summary Language: English Book Monograph; Conference ASFA Input Center Number: CS0206546 ISSN: 0273-1223 ISBN: 19002223614800631 PARTICLES3WRc plc, Henley Road, Medmenham, Bucks, SL7 2HD, UK>?Stanfield, G. Carrington, E. Albinet, F. Compagnon, B. Dumoutier, N. Hambsch, B. Lorthioy, A. Medema, G. Pezoldt, H. de Roubin, M. R. de Lohman, A. Whitmore, T.2000nOptimized and standardized test to determine the presence of the protozoa Cryptosporidium and Giardia in water103-110Water Science & Technology417VWater analysis; Filters (for fluids); Flocculation; Iron compounds; Purification; Magnetic separation; Flotation; Microscopic examination Cytometry; Iron sulfate; Immunomagnetic separation (IMS) EE 445.2 Water Analysis; EE 801.2 Biochemistry; EE 445.1 Water Treatment Techniques; EE 802.3 Chemical Operations; EE 804.2 Inorganic CompoundsWith funding from the European Commission, a consortium of members of the European Water Research Institutes is carrying out a programme of work with the objective of optimizing and standardizing a method for determining the presence in water of (oo)cysts of Cryptosporidium and Giardia. Each of the stages of the conventional analysis procedure (initial concentration, recovery and identification and enumeration) are being investigated and the relative merits of existing and new methods are being assessed. Newly developed filters (Envirochek and Filta-Max) have been shown to be more efficient for initial recovery of (oo)cysts from water than the previously used Cuno cartridge filters. In addition, for the analysis of raw waters, flocculation with ferric sulphate has been shown to give recoveries similar to the Envirochek and Filta Max. Modern purification systems such as immunomagnetic separation have also been assessed and found to offer some advantages over flotation although optimization of the latter has brought improved efficiency. Preliminary assessment of solid phase cytometry has indicated that this technique could offer significant time savings compared to conventional microscopic counting. The results of the study will be used to propose a revised standard method to CEN.!ISSN: 0273-1223 Language: EnglishWATER SCI TECHNOL491866WRc plc, Bucks, UK ~?)Gale, P. Van Dijk, P. A. H. Stanfield, G.1997rDrinking water treatment increases micro-organism clustering; the implications for microbiological risk assessment117-126,AQUA: J. WATER SUPPLY RES. TECHNOL., vol. 463microbiological studies; risk; drinking water; water treatment; microorganisms; public health; pathogens; spatial distribution; spores; model studies; Cryptosporidium; density; microbial contamination; risk assessment; Cryptosporidium parvum SW 3060 Water treatment and distribution; P 2000 FRESHWATER POLLUTION; H SE3.21 WATER POLLUTION/WATER QUALITY; R2 23060 Medical and environmental healthJunCurrent models to assess the risks to public health from waterborne pathogens assume that micro-organisms are randomly dispersed within drinking water samples. One manifestation of such an assumption is that, under non-outbreak conditions, models predict that consumers are only exposed to daily doses of either zero pathogens or just one pathogen. This paper presents evidence against this assumption and demonstrates that in drinking water bacteria spores are spatially associated or clustered to some degree. The variability between spore counts in most 10-mL or 100-mL sub-samples within 100 L treated water volumes was accommodated by a negative binomial distribution. However, a few sub-samples contained extremely high counts which could not be so accommodated. A comparison of spore counts within 100 L volumes before and after alum coagulation and rapid gravity filtration demonstrated that drinking water treatment not only removed 95-99% of spores but also appeared to promote their clustering. More extreme clustering could occur from tight association, where micro-organisms are bound to particles such as metal hydroxides from coagulation. This was not investigated. Through clustering, a few drinking water consumers could be exposed to higher doses of pathogen than just one a day. This is an important consideration in risk models for waterborne pathogens such as Cryptosporidium parvum with an ID sub(50) for healthy human volunteers of about 200 oocysts. The data presented here contribute to modelling the effect of treatment on raw water pathogen density data (measured in 100 L volumes) and simulating pathogen counts in the finished waters at the resolution of unboiled volumes consumed daily by consumers (0.1-1 L).CISSN: 0003-7214 Language: English Summary Language: English; French40925739WRc plc, Henley Rd., Medmenham, Marlow, Bucks SL7 2HD, UK~?HTaghi-Kilani, R. Gyurek, L. L. Millard, P. J. Finch, G. R. Belosevic, M.1996XNucleic acid stains as indicators of Giardia muris viability following cyst inactivation637-46Int J Parasitol266cAnimals Anti-Infective Agents/pharmacology Chlorine/pharmacology Comparative Study Disinfection/*methods Dyes Female Giardia/drug effects/*isolation & purification/pathogenicity Hamsters Mice *Microbiological Techniques Nucleic Acids/*isolation & purification Oxidants/pharmacology Ozone/pharmacology Staining and Labeling/*methods Support, Non-U.S. Gov'tJunA reliable viability assay for Giardia is required for the development of disinfection process design criteria and pathogen monitoring by water treatment utilities. Surveys of single-staining nucleic acid dyes (stain dead parasites only), and double-staining vital dye kits from Molecular Probes (stain live and dead parasites) were conducted to assess the viability of untreated, heat-killed, and chemically inactivated Giardia muris cysts. Nucleic acid staining results were compared to those of in vitro excystation and animal infectivity. Nucleic acid stain, designated as SYTO-9, was considered the best among the single-staining dyes for its ability to stain dead cysts brightly and its relatively slow decay rate of visible light emission following DNA binding. SYTO-9 staining was correlated to animal infectivity. A Live/Dead BacLight was found to be the better of 2 double-staining viability kits tested. Logarithmic survival ratios based on SYTO-9 and Live/Dead BacLight were compared to excystation and infectivity results for G. muris cysts exposed to ozone or free chlorine. The results indicate that SYTO-9 and Live/Dead BacLight staining is stable following treatment of cysts with chemical disinfectants.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=88753090020-7519 Journal Article8875309KDepartment of Biological Sciences, University of Alberta, Edmonton, Canada.~?8Black, E. K. Finch, G. R. Taghi-Kilani, R. Belosevic, M.1996]Comparison of assays for Cryptosporidium parvum oocysts viability after chemical disinfection187-9FEMS Microbiol Lett1352-3Animals Animals, Newborn Comparative Study Cryptosporidium parvum/isolation & purification/pathogenicity/*physiology *Disinfection Fluorescent Dyes Indoles Mice Propidium Support, Non-U.S. Gov'tJan 15=In vitro excystation, vital dyes (4', 6-diamidino-2-phenylindole (DAPI) and propidium iodide (PI), and infectivity in neonatal CD-1 mice were used to assess the viability of Cryptosporidium parvum oocysts after chemical disinfection. In vitro excystation and DAPI/PI staining provided similar estimates of viability in bench-scale experiments, but both of these methods significantly overestimated the viability when compared with infectivity (Pr < or = 0.01). Infectivity was the most reliable measure of the viability of C. parvum oocysts following chemical disinfection.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=85958560378-1097 Journal Article8595856IDepartment of Civil Engineering, University of Alberta, Edmonton, Canada. ~?:Chauret, C. Nolan, K. Chen, P. Springthorpe, S. Sattar, S.1998~Aging of Cryptosporidium parvum oocysts in river water and their susceptibility to disinfection by chlorine and monochloramine1154-60Can J Microbiol4412Age Factors Animals Anti-Infective Agents/pharmacology Cattle Chloramines/*pharmacology Chlorine/*pharmacology Cryptosporidium parvum/*drug effects/*physiology Disinfection/*methods Feces/parasitology Fresh Water/*parasitology Support, Non-U.S. Gov't Water SupplyDecCryptosporidium parvum oocysts were aged in waters from both the St. Lawrence River and the Ottawa River. In situ survival experiments were carried out by incubating the oocysts in either dialysis cassettes or microtubes floated into an overflow tank. A significant portion of the oocysts survived in the test waters for several weeks. Oocyst survival in the St. Lawrence River was better in membrane-filtered (0.2 microm-pore diameter) water than in unfiltered water, suggesting that biological antagonism may play a role in the environmental fate of the parasite. Oocysts aged in river waters under in situ conditions and control oocysts kept refrigerated in synthetic water (100 ppm as CaCO3); pH 7.0) were subjected to the same disinfection protocol. Aged oocysts were at least as resistant as, if not more resistant than, the control oocysts to disinfection. This indicates that the oocysts surviving in the water environment may be just as difficult to inactivate by potable water disinfection as freshly shed oocysts. Therefore, water treatment should not be based on the assumption that environmental oocysts may be more easily inactivated than freshly shed oocysts. First-order kinetics die-off rates varied from one river to another (from 0.013 to 0.039 log(10).day(-1)) and from one experiment to another with water from the same river collected at different times. Calculation of the die-off rates based on either in vitro excystation or in vitro excystation in combination with total counts (overall die-off rates) showed that the assessment of oocyst viability by microscopic methods must account for the total oocyst loss observed during long-term inactivation assays of river waters.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=103832270008-4166 Journal Article10383227fBiological and Physical Sciences Unit, Indiana University Kokomo, IN 46904-9003, USA. cchauret@iuk.edu~?Current, W. L. Long, P. L.1983@Development of human and calf Cryptosporidium in chicken embryos1108-13 J Infect Dis1486Animals Apicomplexa/cytology/*growth & development/physiology Cattle/parasitology Chick Embryo/*parasitology Comparative Study Human Ileum/parasitology Mice Microvilli/parasitology Movement Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Temperature Time FactorsDecCryptosporidium is a newly recognized, zoonotic protozoan that produces short-term, flu-like, gastrointestinal illness in immunocompetent humans and prolonged, severe, diarrhea in immunocompromised individuals. Successful completion of the life cycle, from sporozoite to infective oocyst, of isolates of Cryptosporidium from humans and calves was demonstrated in endoderm cells of the chorioallantoic membrane (CAM) of chicken embryos maintained at 37 C. The human and calf isolates of Cryptosporidium were morphologically and developmentally indistinguishable when grown in chicken embryos. The human isolate also completed its entire life cycle in the CAMs of chicken embryos maintained at 35 C and 41 C. Oocysts recovered from endoderm cells of infected CAMs produced heavy infections in suckling mice. The timing, presence, and morphology of developmental stages in CAM cells during the first eight days after inoculation of sporozoites were similar to those in enterocytes of mice inoculated with oocysts. The method described is safe and convenient for cultivating and studying Cryptosporidium in a bacteria-free environment; the system also lends itself to well-established procedures for evaluating antiprotozoan drugs.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=64188300022-1899 Journal Article6418830~?Blagburn, B. L. Current, W. L.1983?Accidental infection of a researcher with human Cryptosporidium772-3 J Infect Dis1484eAcquired Immunodeficiency Syndrome/complications Adult Coccidiosis/pathology/*transmission Human MaleOctdhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=66310720022-1899 Case Reports Letter6631072?~?^Koch, K. L. Shankey, T. V. Weinstein, G. S. Dye, R. E. Abt, A. B. Current, W. L. Eyster, M. E.1983Cryptosporidiosis in a patient with hemophilia, common variable hypogammaglobulinemia, and the acquired immunodeficiency syndrome337-40Ann Intern Med993Adult Agammaglobulinemia/*complications/immunology Animals Cat Diseases/parasitology/transmission Cats Diarrhea/*etiology Feces/parasitology Hemophilia A/*complications Human Leukocyte Count Lymphocyte Activation Lymphopenia/etiology Male Protozoan Infections/complications/*transmission Protozoan Infections, Animal T-Lymphocytes, Helper-Inducer/immunology T-Lymphocytes, Suppressor-Effector/immunologySepA 36-year-old man had chronic, debilitating diarrhea due to cryptosporidiosis. This patient had longstanding common variable hypogammaglobulinemia and recurrent bacterial infections. Immunologic evaluation after discovery of Cryptosporidium showed lymphopenia with persistently reduced numbers of helper/inducer cells (OKT-4), variable numbers of suppressor/cytotoxic cells (OKT-8), OKT-4/OKT-8 ratio of 0.09, and increased levels of serum alpha-interferon, all of which describe the acquired immunodeficiency syndrome. Oocysts of Cryptosporidium were found in feces from the patient's cat, thus identifying a possible source of his infection. The patient had disseminated candidiasis, cytomegalovirus pneumonia, and cryptosporidiosis when he died.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=6225362&0003-4819 Case Reports Journal Article6225362~?*Lindsay, D. S. Current, W. L. Ernst, J. V.1983!Motility of Isospora suis meronts783-4 J Parasitol694pAnimals Isospora/*physiology Movement Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Swine/parasitologyAugdhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=66316450022-3395 Journal Article6631645~?Campbell, P. N. Current, W. L.1983wDemonstration of serum antibodies to Cryptosporidium sp. in normal and immunodeficient humans with confirmed infections165-9J Clin Microbiol181Acquired Immunodeficiency Syndrome/*immunology Agammaglobulinemia/*immunology Antibodies/*analysis Coccidiosis/*immunology Fluorescent Antibody Technique Human Species Specificity Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S.Jul6Antibodies to Cryptosporidium sp. were detected in sera from 12 immunocompetent individuals recovered from cryptosporidiosis and from 5 subjects with an acquired immunodeficiency syndrome and persistent cryptosporidiosis by an indirect immunofluorescent (IIF) test. Marked seroconversion accompanied recovery from infection in immunocompetent individuals, and their IIF titers remained high (1:40 to 1:640) for at least 1 year. No antibodies to Cryptosporidium sp. were detected in sera from two subjects with hypogammaglobulinemia, normal T-cell function, and persistent cryptosporidiosis or in sera from individuals not previously exposed to Cryptosporidium sp. Very little or no cross-reactivity with the other coccidia--Toxoplasma, Sarcocystis, and Isospora spp.--occurred in the IIF test procedure. The application of this IIF procedure, along with recently developed techniques to detect oocysts in the feces, should provide the basis for a more accurate assessment of the number of individuals within any subject group with previous and active Cryptosporidium infections.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=63503450095-1137 Journal Article6350345~?UCurrent, W. L. Reese, N. C. Ernst, J. V. Bailey, W. S. Heyman, M. B. Weinstein, W. M.1983|Human cryptosporidiosis in immunocompetent and immunodeficient persons. Studies of an outbreak and experimental transmission1252-7 N Engl J Med30821Acquired Immunodeficiency Syndrome/complications Adolescent Adult Agammaglobulinemia/complications Animals Cats Cattle Cattle Diseases/transmission Coccidia/isolation & purification Coccidiosis/diagnosis/*transmission/veterinary Disease Outbreaks/epidemiology Dogs Female Goats Human Immune Tolerance Immunologic Deficiency Syndromes/*complications Male Mice Middle Aged Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. ZoonosesMay 26Infection with cryptosporidium occurred in 12 immunocompetent persons who had direct contact with the feces of infected calves during three unrelated outbreaks of calf cryptosporidiosis. Nine of the twelve subjects had diarrhea and abdominal cramps that lasted 1 to 10 days. Infections were diagnosed and monitored by detection of oocysts in feces, with a modified Sheather's flotation technique and phase-contrast microscopy. Oocysts of cryptosporidium were isolated from calves but not from other animals with which these subjects had been in contact. Oocysts of cryptosporidium were also detected during repeated examinations of feces from two immunodeficient patients with persistent cryptosporidiosis. An apparently identical infection was transmitted to calves and mice, using oocysts from infected calves and human beings. Oocysts from an immunodeficient person also produced infections in kittens, puppies, and goats. This study shows that cryptosporidium may produce a moderate self-limited illness in immunocompetent persons, which contrasts sharply with the prolonged severe diarrhea in immunocompromised patients who contract cryptosporidiosis. Calves with diarrhea should be considered a potential source of human infection, and immunocompromised persons should avoid contact with such animals.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=6843609&0028-4793 Case Reports Journal Article6843609s~?*Lindsay, D. S. Current, W. L. Ernst, J. V.19833Excystation of Isospora suis Biester, 1934 of swine27-34 Z Parasitenkd691hAnimals Isospora/*physiology Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Swine/*parasitologyThe in vitro excystation of sporozoites of Isospora suis Biester 1934 is described. Sporocysts of I. suis lack a Stieda body. Upon incubation in 0.75% sodium taurocholate or in 0.25% trypsin + 0.75% sodium taurocholate excystation solutions, sporozoites were released by separation of the sporocyst wall into four plates. Occasionally, the sporocyst wall did not separate completely but opened partially and released the sporozoite. At the time of excystation, sporozoites were short and broad but became elongated after 5 to 10 min in the excystation fluids. Elongate sporozoites measuring 11.7 x 3.8 micrometers, had a pointed anterior end and a nucleus located in the posterior half of the cell. Living sporozoites exhibited gliding movements, side-to-side flexion, and probed with their anterior ends. Incubation in 5.25% sodium hypochlorite removed the oocyst walls from most oocysts. Sporozoites did not excyst from sporocysts that were released during treatment with sodium hypochlorite.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=68371000044-3255 Journal Article6837100~?Entrala, E. Mascaro, C.1997>Glycolytic enzyme activities in Cryptosporidium parvum oocysts51-7FEMS Microbiol Lett1511Animals Cell Compartmentation Cryptosporidiosis/veterinary Cryptosporidium parvum/*enzymology *Glycolysis Goat Diseases/parasitology Goats Isoelectric Focusing Support, Non-U.S. Gov'tJun 1JOocysts of Cryptosporidium parvum were obtained from an experimentally infected newborn goat. After purification, the oocysts were homogenised and the activities of the glycolytic enzymes measured in the different subcellular fractions. All of the activities of the Embden-Meyerhoff pathway were located in the non-sedimentable, cytoplasmic fraction. Under the conditions used, hexokinase activity was below the limits of detection. The pathway is also characterised by the presence of a pyrophosphate-dependent phosphofructokinase and a carbon dioxide-fixing cycle comprising phosphoenolpyruvate carboxylase, malate dehydrogenase and malate dehydrogenase (decarboxylating) activities. The data presented in this paper suggest that the infective stage of this parasite probably relies on substrate-level phosphorylation for energy generation.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=91982810378-1097 Journal Article9198281WDept. Parasitologia, Facultad de Ciencias, Instituto de Biotechnologia, Granada, Spain.2~?#Entrala, E. Mascaro, C. Barrett, J.19976Anti-oxidant enzymes in Cryptosporidium parvum oocysts13-7 Parasitology 114 ( Pt 1)sAnimals Catalase/analysis Cattle Cryptosporidiosis/parasitology Cryptosporidium parvum/*enzymology Feces/parasitology Glutathione Peroxidase/analysis Glutathione Reductase/analysis Glutathione Transferase/analysis Isoelectric Point Molecular Weight NADH, NADPH Oxidoreductases/analysis Peroxidases/analysis Superoxide Dismutase/*analysis/chemistry Support, Non-U.S. Gov'tJanNOocysts of Cryptosporidium parvum showed relatively low levels of SOD activity. The SOD which had a pI of 4.8 and an approximate molecular weight of 35 kDa appeared to be iron dependent. Catalase, glutathione transferase, glutathione reductase and glutathione peroxidase activity could not be detected, nor could trypanothione reductase. No NADH or NADPH oxidase activity could be detected, nor could peroxidase activity be demonstrated using o-dianisidine, guaiacol, NADPH or NADH as co-substrates. However, an NADPH-dependent H2O2 scavenging system was detected in the insoluble fraction.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=90110700031-1820 Journal Article9011070@Departmento Parasitologia, Facultad de Ciencias, Granada, Spain.~?)Graczyk, T. K. Fayer, R. Cranfield, M. R.1997_Zoonotic transmission of Cryptosporidium parvum: Implications for water-borne cryptosporidiosis348-51Parasitol Today139SepThe emergence of Cryptosporidium parvum-associated cryptosporidiosis as a worldwide zoonosis has stimulated interest in the modes of pathogen transmission. Here, Thaddeus Graczyk, Ronald Fayer and Michael Cranfield discuss the complex epidemiology of C. parvum, emphasizing the crosstransmission potential of the pathogen, mechanical vectors involved in water-borne transmission of the oocysts, and factors contributing to contamination of pristine waters with Cryptosporidium. They also outline the public health importance of proper interpretation of positive detection of Cryptosporidium oocysts at water-treatment facilities and identify means by which watersheds can be protected from Cryptosporidium contamination.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=152750480169-4758 Journal Article15275048Department of Molecular Microbiology and Immunology, Johns Hopkins University, School of Hygiene and Public Health, 615 North Wolfe Street, Baltimore, MD 21205-2179, USA.~?@Graczyk, T. K. Fayer, R. Lewis, E. J. Farley, C. A. Trout, J. M.1997In vitro interactions between hemocytes of the eastern oyster, Crassostrea virginica Gmelin, 1791 and Cryptosporidium parvum oocysts949-52 J Parasitol835Analysis of Variance Animals Cells, Cultured Cryptosporidium parvum/*immunology Fluorescent Antibody Technique, Indirect Hemocytes/immunology/*parasitology Mice Oysters/immunology/*parasitology *PhagocytosisOctIt was demonstrated by an in vitro slide phagocytosis assay that hemocytes of the Eastern oyster, Crassostrea virginica Gmelin, 1791 are capable of rapid recognition and internalization of infectious Cryptosporidium parvum (AUCP-1 strain) oocysts. The incubation of hemocyte monolayers (8.5 x 10(4) cells) that had received 6.8 x 10(5) or 3.4 x 10(5) oocysts was arrested at 5, 15, 30, 60, 90, and 120 min and the oocytes detected by acid-fast stain and immunofluorescent antibody (IFAT). An average of 20.5, 38.3, 50.2, 58.9, 69.0, and 75.0% oocysts were phagocytosed after 5, 15, 30, 60, 90, and 120 min, respectively. The intensity of fluorescence of phagocytosed oocysts significantly decreased over time (P < 0.01), and their round shape was altered. The number of cells containing oocysts and the mean number of ingested oocysts (range: 1.2-4.5 per cell) increased significantly over time (P < 0.01), whereas the numbers of nonphagocytosed oocysts that were adherent to the glass slides significantly decreased (P < 0.05). By extrapolation, the results indicate that Cr. virginica is capable of internalizing up to 6.4 x 10(6) Cryptosporidium oocysts per ml of its hemolymph.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=93793060022-3395 Journal Article9379306Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA.)~?@Schrum, D. P. Alugupalli, S. Kelly, S. T. White, D. C. Fayer, R.1997Structural characterization of a "signature" phosphatidylethanolamine as the major 10-hydroxy stearic acid-containing lipid of Cryptosporidium parvum oocysts789-93Lipids327 Animals Chromatography, High Pressure Liquid Cryptosporidium parvum/*chemistry Magnetic Resonance Spectroscopy Phosphatidylethanolamines/*chemistry Spectrometry, Mass, Fast Atom Bombardment Spectrophotometry, Infrared Stearic Acids/*analysis Support, Non-U.S. Gov'tJul%A 10-hydroxy stearic acid-containing lipid from Cryptosporidium parvum was purified by thin-layer chromatography and analyzed by infrared spectroscopy, fast-atom bombardment mass spectrometry, 1H and 31P nuclear magnetic resonance spectroscopy, and was identified as phosphatidyl-ethanolamine.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=92529700024-4201 Journal Article9252970OMicrobial Insights, Inc., Rockford, Tennessee 37853-3044, USA. milipids@sol.com ~?BGraczyk, T. K. Cranfield, M. R. Fayer, R. Trout, J. Goodale, H. J.1997Infectivity of Cryptosporidium parvum oocysts is retained upon intestinal passage through a migratory water-fowl species (Canada goose, Branta canadensis)341-7Trop Med Int Health24 Animals Cryptosporidiosis/epidemiology/*transmission *Cryptosporidium parvum Disease Reservoirs Disease Transmission, Horizontal Environmental Microbiology Feces/parasitology Geese Mice Mice, Inbred BALB C Parasite Egg Count Support, Non-U.S. Gov't Water MicrobiologyAprFive Cryptosporidium-free Canada geese (Branta canadensis) were individually orally dosed with 3.5 x 10(6) Cryptosporidium parvum oocysts infectious to neonatal BALB/c mice. After intestinal passage, inoculum-derived oocysts extracted from goose faeces established severe infection in 14 neonatal BALB/c mice (inoculum dose 2.5 x 10(5)/mouse). The inoculum-derived oocysts were detected in goose faeces up to 9 days post-inoculation (PI); the number of intact oocysts and oocyst shells shed during the first 3 days PI was significantly higher than for the remaining 6 days PI (P < 0.01). Based on acid-fast stained air-dried direct wet smears, 62% of the oocysts in goose faeces were intact (oocyst shells) constituted 38%) and conformed to morphological features of viable and infectious inoculum oocysts. The fluorescence scores of the inoculated oocysts, obtained by use of the MERIFLUOR test, were identical to those obtained for the faeces-recovered oocysts (majority 3+ to 4+). The dynamics of oocyst shedding showed that overall, the birds released a significantly higher number of intact oocysts than oocyst (P < 0.01). Retention of the viability and infectivity of C. parvum oocysts following intestinal passage through a migratory water-fowl species has serious epidemiological implications. Water-fowl can serve as mechanical vectors for the water-borne oocysts and can contaminate surface waters with C. parvum. As the concentration of Cryptosporidium oocysts in source waters is attributable to water-shed management practices, water-shed protection programme officials should consider water-fowl as a potential factor enhancing contamination of the source water with Cryptosporidium.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=91718421360-2276 Journal Article9171842}Department of Molecular Microbiology and Immunology, Johns Hopkins University, Baltimore, MD, USA. tgraczyk@phnet.sph.jhu.edu~?LPasquali, P. Fayer, R. Almeria, S. Trout, J. Polidori, G. A. Gasbarre, L. C.1997\Lymphocyte dynamic patterns in cattle during a primary infection with Cryptosporidium parvum247-50 J Parasitol832Animals Animals, Newborn Antibodies, Monoclonal/immunology Antibody Specificity Cattle Cattle Diseases/*immunology/pathology Cryptosporidiosis/*immunology/pathology Fluorescent Antibody Technique, Indirect/veterinary Ileum/immunology/pathology Immunity, Cellular Intestinal Mucosa/immunology/pathology Lymph Nodes/immunology/pathology Lymphocyte Count/veterinary Male T-Lymphocyte Subsets/*pathologyApr+Changes in intraepithelial (IEL), lamina propria (LPL), and draining lymph node (LNL) lymphocytes were assessed in 9-day-old calves during primary infection with Cryptosporidium parvum and in similarly aged noninfected calves. A very low percentage of both CD4+ and CD8+ T cells were found in IEL and LPL of noninfected calves. In infected compared to controls, percentages of CD2+, CD3+, CD4+, and CD8+ T cells in IEL exhibited a significant increase (P < 0.05), whereas the percentage of IL2R+ increased and the percentage of IgG+ cells decreased, but neither of these changes were statistically significant. In LPL, percentages of CD2+, CD3+, CD8+, and IL2R+ T cells were increased in infected compared to noninfected calves, whereas the percentage of IgG-bearing cells decreased; but only the increase in CD3+, CD8+, and IL2R+ cells was significant (P < or = 0.05). In LNL only minimal changes were seen. In fact, the percentage of CD2+ T cells increased whereas the percentage of CD8+ T cells decreased, but neither of these differences was statistically significant. These findings indicate that T cells subsets in the ileal mucosa of naive neonatal calves are different than those of adult cattle, and that the immune response to C. parvum infection differs in ileal mucosa when compared to the regional lymph nodes.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=91053050022-3395 Journal Article9105305_Immunology and Disease Resistance Laboratory, LPSI, ARS, USDA, Beltsville, Maryland 20705, USA.~?3Graczyk, T. K. Fayer, R. Cranfield, M. R. Owens, R.1997vCryptosporidium parvum oocysts recovered from water by the membrane filter dissolution method retain their infectivity111-4 J Parasitol831!Acetone Animals Animals, Newborn Biological Assay Cattle Cellulose/analogs & derivatives Cryptosporidium parvum/drug effects/isolation & purification/*physiology Ethanol Filtration/methods *Membranes, Artificial Mice Mice, Inbred BALB C Solvents Support, Non-U.S. Gov't Water/*parasitologyFebCryptosporidium parvum oocysts infectious to neonatal BALB/c mice were processed by the cellulose-acetate membrane (CAM) filter dissolution method to determine if the procedure that utilizes acetone incubation and alcohol centrifugations alters their viability (determined by in vitro excystation) or infectivity (determined by infectivity bioassay). In addition, most oocysts with altered viability by desiccation, heat inactivation, and snap freezing that were processed by the CAM filter dissolution method were nonrefractile, unstained oocyst ghosts. The remaining organisms, oocyst shells, were lightly stained with the acid-fast stain. Infectious oocysts retained their infectivity and nonviable oocysts (oocyst shells) retained their morphology when processed by the CAM dissolution method. Infectious oocysts, oocyst shells, and oocyst ghosts produced positive reactions of similar intensity in direct immunofluorescence antibody staining, utilizing the MERIFLUOR Cryptosporidium/Giardia test kit. Cryptosporidium oocysts recovered from finished drinking water by the CAM dissolution method can be subjected to testing for their viability and infectivity.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=90577050022-3395 Journal Article9057705Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA.~?)Graczyk, T. K. Cranfield, M. R. Fayer, R.1997nRecovery of waterborne oocysts of Cryptosporidium from water samples by the membrane-filter dissolution method121-5 Parasitol Res832Animals Cattle Cryptosporidiosis/transmission Cryptosporidium parvum/*isolation & purification Filtration Human Support, Non-U.S. Gov't Water/*parasitology *Water Supply ZygotepThe cellulose-acetate membrane (CAM)-filter dissolution method implemented into a Millipore Glass Microanalysis system was used for recovery of Cryptosporidium parvum oocysts seeded into 25 l of drinking water in polyethylene carboy aspirator bottles. CAM-entrapped oocysts were detected by immunofluorescence microscopy. From 65 to 94 oocysts/l (mean 75 oocysts/l), 34.7% overall of the inoculated oocysts, were unrecovered as determined after the water had been drained from the bottle, rinsed with 1 l of eluting fluid (EF), and CAM-filtered. Efficiency rates of oocyst recovery ranged from 24.0% to 64.0% (mean 44.1%), without the use of EF and from 72.1% to 82.3% (mean 78.8%) when EF was used. To ensure a high recovery efficiency of Cryptosporidium oocysts from sampled water by the CAM-filter dissolution method, it is recommended that 1 l of EF per 25 l of water be used.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=90396930932-0113 Journal Article9039693Department of Molecular Microbiology and Immunology, Johns Hopkins University, School of Hygiene and Public Health, Baltimore, MD 21205, USA.T~?EFayer, R. Fischer, J. R. Sewell, C. T. Kavanaugh, D. M. Osborn, D. A.1996SSpontaneous cryptosporidiosis in captive white-tailed deer (Odocoileus virginianus)619-22 J Wildl Dis324Animals Animals, Newborn Animals, Zoo Cattle *Cryptosporidiosis/parasitology Deer/*parasitology Feces/parasitology Female Male Mice Parasite Egg Count/veterinary PregnancyOctIn August 1994, cryptosporidiosis was diagnosed in a diarrheic fawn from a captive white-tailed deer (Odocoileus virginianus) herd maintained for research purposes at The University of Georgia's Warnell School of Forest Resources in Athens, Georgia (USA). From June through August 1995, 11 captive female white-tailed deer were housed in individual barn stalls where they gave birth to 18 fawns. Feces collected at 2 or 3 day intervals from the 18 neonatal fawns for at least 21 days and from 11 adult females once from 1 to 30 days before fawns were born and on three to 12 occasions after their birth were examined for oocysts of Cryptosporidium spp. Feces from all animals appeared normal throughout the period of examination. Oocysts morphologically indistinguishable from those of Cryptosporidium parvum were detected intermittently in the feces of one adult female from 1 to 25 days after parturition and in the feces of her fawn from 11 to 22 days of age. Oocysts also were detected intermittently in feces from twin fawns from 9 to 20 days of age, but not from their mother. Oocysts from deer were infectious for neonatal mice as determined histologically, and for calves as determined by clinical signs and excretion of oocysts.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=93590610090-3558 Journal Article9359061hUnited States Department of Agriculture, Agricultural Research Service, Beltsville, Maryland 20705, USA. W~?Urban, J. F., Jr. Fayer, R. Sullivan, C. Goldhill, J. Shea-Donohue, T. Madden, K. Morris, S. C. Katona, I. Gause, W. Ruff, M. Mansfield, L. S. Finkelman, F. D.1996eLocal TH1 and TH2 responses to parasitic infection in the intestine: regulation by IFN-gamma and IL-4337-44Vet Immunol Immunopathol541-4nAnimals Cryptosporidiosis/*immunology Cryptosporidium parvum/immunology Interferon Type II/*pharmacology Interleukin-4/*pharmacology Intestines/drug effects/*immunology/*parasitology Mice Mice, SCID Nematospiroides dubius/immunology Nippostrongylus/immunology Strongylida Infections/*immunology Th1 Cells/*drug effects/*immunology Th2 Cells/*drug effects/*immunologyNovControl of parasitic infections is dependent on the production of cytokines that activate mechanisms which limit invasion, reproduction or survival of the parasite. In contrast, conditions that induce inappropriate cytokine responses facilitate the spread of infection and ultimately exacerbate the level of disease. Measurement of local cytokine responses to different gastrointestinal parasites, such as the intracellular protozoan, Cryptosporidium parvum, and luminal dwelling nematodes like Nippostrongylus brasiliensis and Heligmosomoides polygyrus, reveal stereotype response patterns. In general, intracellular parasites stimulate type 1 responses where IFN-gamma is the predominant immune activator, while extracellular parasites stimulate type 2 responses where IL-4 plays a prominent role in elevating humoral immune mechanisms. Cytokines alter cellular function and the milieu of the intestinal lumen to affect the outcome of an infection. The importance of a particular response during the course of an infection can be studied by selective enhancement with an excess of exogenous recombinant cytokine or cytokine antagonists. For example, exogenous IL-12 enhances resistance to C.parvum, but suppresses the normally rapid cure of an infection with N. brasiliensis. Both mechanisms are dependent on expression of IFN-gamma. At the molecular level, exogenous IL-12 stimulates IFN-gamma production which elevates a protective type 1 response to C. parvum but converts the normally anti-worm type 2 response to a type 1 response that inappropriately regulates the infection. Alternatively, excess IL-4 plays a prominent role in modulating effector elements that change intestinal physiology to create a hostile environment for worm parasites. Exogenous IL-4 can cure chronic worm infection, while IL-4 antagonists interfere with protective responses to infection. These observations provide a paradigm for analysis of stereotype responses to different gastrointestinal parasites, and demonstrate how cytokine-induced immune system-dependent and independent effector mechanisms can limit parasitic infection, while inappropriate cytokine responses can exacerbate the state of disease.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=89888790165-2427 Journal Article8988879rImmunology and Disease Resistance Laboratory, Livestock and Poultry Sciences Institute, Beltsville, MD 20705, USA.~?)Graczyk, T. K. Fayer, R. Cranfield, M. R.1996LCryptosporidium parvum is not transmissible to fish, amphibians, or reptiles748-51 J Parasitol825zAnimals Anura/*parasitology Cloaca/parasitology Cryptosporidiosis/parasitology/transmission/*veterinary Cryptosporidium parvum/*physiology Elapidae/*parasitology Fluorescent Antibody Technique Ileum/parasitology Jejunum/parasitology Lizards/*parasitology Mice Mice, Inbred BALB C Perciformes/*parasitology Stomach/parasitology Support, Non-U.S. Gov't Xenopus laevis/parasitologyOctOA recent report suggested that an isolate of Cryptosporidium parvum had established infections in fish, amphibians, and reptiles and raises concern that animals other than mammals might be a potential source of waterborne Cryptosporidium oocysts. To test this possibility, viable C. parvum oocysts, infectious for neonatal BALB/c mice, were delivered by gastric intubation to bluegill sunfish, poison-dart frogs, African clawed frogs, bearded dragon lizards, and corn snakes. Histological sections of the stomach, jejunum, ileum, and cloaca prepared from tissues collected on days 7 and 14 postinoculation (PI) were negative for Cryptosporidium developmental stages. However, inoculum-derived oocysts were detectable by fluorescein-labeled monoclonal antibody in feces of inoculated animals from day 1 to day 12 PI in fish and frogs, and up to day 14 PI in lizards. Snakes did not defecate for 14 days PI. Impression smears taken at necropsy on days 7 and 14 PI revealed C. parvum oocysts in the lumen of the cloaca of 2 fish and 1 lizard on day 7 PI only. Because tissue stages of the pathogen were not found, it appears that C. parvum was not heterologously transmitted to lower vertebrates. Under certain circumstances, however, such as after the ingestion of C. parvum-infected prey, lower vertebrates may disseminate C. parvum oocysts in the environment.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=88858830022-3395 Journal Article8885883Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA.~?6Fayer, R. Graczyk, T. K. Cranfield, M. R. Trout, J. M.19966Gaseous disinfection of Cryptosporidium parvum oocysts3908-9Appl Environ Microbiol6210Ammonia/pharmacology Animals Animals, Newborn Carbon Monoxide/pharmacology Cecum/parasitology Colon/parasitology Cryptosporidiosis/parasitology/prevention & control Cryptosporidium parvum/*drug effects/growth & development/pathogenicity Disinfectants/pharmacology Disinfection/*methods Ethylene Oxide/pharmacology Formaldehyde/pharmacology *Gases Hydrocarbons, Brominated/pharmacology Ileum/parasitology Mice Mice, Inbred BALB COctIPurified oocysts of Cryptosporidium parvum suspended in approximately 400 microliters of phosphate-buffered saline or deionized water in microcentrifuge tubes were exposed at 21 to 23 degrees C for 24 h to a saturated atmosphere of ammonia, carbon monoxide, ethylene oxide, formaldehyde, or methyl bromide gas. Controls were exposed to air. Oocysts in each tube were then rinsed and resuspended in fresh, deionized water, and 1 million oocysts exposed to each gas were orally administered to each of three to six neonatal BALB/c mice in replicate groups. Histologic sections of ileum, cecum, and colon tissues taken from each mouse 72 h after oral administration of oocysts were examined microscopically to determine if infection had been established. All 15 mice given oocysts exposed to carbon monoxide had numerous developmental stages of cryptosporidium in all three intestinal segments. Of 10 mice given oocysts exposed to formaldehyde, 6 had a few developmental stages of cryptosporidium in the ileum. No mice given oocysts exposed to ammonia, ethylene oxide, or methyl bromide were found to be infected. These findings indicate the efficacy of these low-molecular-weight gases (ammonia, ethylene oxide, and methyl bromide) as potential disinfectants for C. parvum oocysts where soil, rooms, buildings, tools, or instruments might be contaminated.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=88374510099-2240 Journal Article8837451sImmunology and Disease Resistance Laboratory, U.S. Department of Agriculture, Beltsville, Maryland 20705-2350, USA.~?Fayer, R. Trout, J. Nerad, T.1996XEffects of a wide range of temperatures on infectivity of Cryptosporidium parvum oocysts64SJ Eukaryot Microbiol435pAnimals Cattle Cryptosporidium parvum/*pathogenicity Freezing Heating Male Mice Mice, Inbred BALB C *TemperatureSep-Octdhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=88228611066-5234 Journal Article8822861USDA, ARS, Beltsville, MD, USA. ~?9Graczyk, T. K. Cranfield, M. R. Fayer, R. Anderson, M. S.1996Viability and infectivity of Cryptosporidium parvum oocysts are retained upon intestinal passage through a refractory avian host3234-7Appl Environ Microbiol629Animals Cryptosporidium parvum/*physiology Ducks/*parasitology Feces/parasitology Intestines/*parasitology Mice Mice, Inbred BALB C Support, Non-U.S. Gov't Water/parasitologySepSix Cryptosporidium-free Peking ducks (Anas platyrhynchos) were each orally inoculated with 2.0 x 10(6) Cryptosporidium parvum oocysts infectious to neonatal BALB/c mice. Histological examination of the stomachs jejunums, ilea, ceca, cloacae, larynges, tracheae, and lungs of the ducks euthanized on day 7 postinoculation (p.i.) revealed no life-cycle stages of C. parvum. However, inoculum-derived oocysts extracted from duck feces established severe infection in eight neonatal BALB/c mice (inoculum dose, 2.5 x 10(5) per mouse). On the basis of acid-fast stained direct wet smears, 73% of the oocysts in duck feces were intact (27% were oocyst shells), and their morphological features conformed to those of viable and infectious oocysts of the original inoculum. The fluorescence scores of the inoculated oocysts, obtained by use of the MERIFLUOR test, were identical to those obtained for the feces-recovered oocysts (the majority were 3+ to 4+). The dynamics of oocyst shedding showed that the birds released a significantly higher number of intact oocysts than the oocyst shells (P < 0.01). The number of intact oocysts shed (87%) during the first 2 days p.i. was significantly higher than the number shed during the remaining 5 days p.i. (P < 0.01) and significantly decreased from day 1 to day 2 p.i. (P < 0.01). The number of oocyst shells shed during 7 days p.i. did not vary significantly (P > 0.05). The retention of infectivity of C. parvum oocysts after intestinal passage through an aquatic bird has serious epidemiological and epizootiological implications. Waterfowl may serve as mechanical vectors for the waterborne oocysts and may enhance contamination of surface waters with C. parvum. As the concentration of Cryptosporidium oocysts in source waters is attributable to watershed management practices, the watershed protection program should consider waterfowl as a potential factor enhancing contamination of the source water with C. parvum.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=87952130099-2240 Journal Article8795213Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA. tgraczyk@phnet.sph.jhu.edu~?1Harp, J. A. Fayer, R. Pesch, B. A. Jackson, G. J.1996[Effect of pasteurization on infectivity of Cryptosporidium parvum oocysts in water and milk2866-8Appl Environ Microbiol628Animals Cattle Cryptosporidium parvum/*pathogenicity Heat Mice Mice, Inbred BALB C Milk/*parasitology *Sterilization Time Factors Water/*parasitologyAugCryptosporidium parvum is a major cause of diarrheal disease in humans and has been identified in 78 other species of mammals. The oocyst stage, excreted in feces of infected humans and animals, has been responsible for recent waterborne outbreaks of human cryptosporidiosis. High temperature and long exposure time have been shown to render oocysts (suspended in water) noninfectious, but for practical purposes, it is important to know if high-temperature--short-time conditions (71.7 degrees C for 15 s) used in commercial pasteurization are sufficient to destroy infectivity of oocysts. In this study, oocysts were suspended in either water or whole milk and heated to 71.7 degrees C for 15, 10, or 5 s in a laboratory-scale pasteurizer. Pasteurized and nonpasteurized (control) oocysts were then tested for the ability to infect infant mice. No mice (0 of 177) given 10(5) oocysts pasteurized for 15, 10, or 5 s in either water or milk were found to be infected with C. parvum on the basis of histologic examination of the terminal ileum. In contrast, all (80 of 80) control mice given nonpasteurized oocysts were heavily infected. These data indicate that high-temperature--short-time pasteurization is sufficient to destroy the infectivity of C. parvum oocysts in water and milk.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=87022790099-2240 Journal Article8702279Metabolic Diseases and Immunology Research Unit, USDA Agricultural Research Service, National Animal Disease Center, Ames, Iowa 50010-0070, USA. !A03RLMDIR@ATTMAIL.COM ~?Fayer, R. Nerad, T.1996JEffects of low temperatures on viability of Cryptosporidium parvum oocysts1431-3Appl Environ Microbiol624Animals Animals, Newborn Cold Comparative Study Cryopreservation Cryptosporidium parvum/cytology/growth & development/*pathogenicity Freezing Mice Mice, Inbred BALB C Time FactorsAprMicrocentrifuge tubes containing 8 x 10(6) purified oocysts of Cryptosporidium parvum suspended in 400 microliters of deionized water were stored at 5 degrees C for 168 h or frozen at -10, -15, -20, and -70 degrees C for 1 h to 168 h and then thawed at room temperature (21 degrees C). Fifty microliters containing 10(6) oocysts was administered to each of five to seven neonatal BALB/c mice by gastric intubation. Segments of ileum, cecum, and colon were taken for histology from each mouse 72 or 96 h later. Freeze-thawed oocysts were considered viable and infectious only when developmental-stage C. parvum organisms were found microscopically in the tissue sections. Developmental-stage parasites were not found in tissues from any mice that received oocysts frozen at -70 degrees C for 1, 8, or 24 h. All mice that received oocysts frozen at -20 degrees C for 1, 3, and 5 h had developmental-stage C. parvum; one of 6 mice that received oocysts frozen at -20 degrees C for 8 h had a few developmental-stage parasites; mice that received oocysts frozen at -20 degrees C for 24 and 168 h had no parasites. All mice that received oocysts frozen at -15 degrees C for 8 and 24 h had developmental-stage parasites; mice that received oocysts frozen at -15 degrees C for 168 h had no parasites. All mice that received oocysts frozen at -10 degrees C for 8, 24, and 168 h and those that received oocysts stored at 5 degrees C for 168 h had developmental-stage parasites. These findings demonstrate for the first time that oocysts of C. parvum in water can retain viability and infectivity after freezing and that oocysts survive longer at higher freezing temperatures.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=89198060099-2240 Journal Article8919806Livestock and Poultry Sciences Institute, Agricultural Research Service, United States Department of Agriculture, Beltsville, Maryland 20705, USA. x~?)Graczyk, T. K. Cranfield, M. R. Fayer, R.1996Evaluation of commercial enzyme immunoassay (EIA) and immunofluorescent antibody (FA) test kits for detection of Cryptosporidium oocysts of species other than Cryptosporidium parvum274-9Am J Trop Med Hyg543IAnimals Camels Cattle Chickens Comparative Study Cross Reactions Cryptosporidiosis/*diagnosis Cryptosporidium/*isolation & purification Feces/*parasitology *Fluorescent Antibody Technique, Direct Guinea Pigs Human Hyraxes *Immunoenzyme Techniques Lizards Sensitivity and Specificity Snakes Support, Non-U.S. Gov't Turkeys TurtlesMarA commercial enzyme immunoassay (EIA) (ProSpect Rapid Assay), a direct immunofluorescence antibody (IFA) test for stool testing (MERIFLUOR Cryptosporidium/Giardia), and an indirect IFA test for environmental testing (Hydrofluor-Combo Cryptosporidium/Giardia) were evaluated for detection of low public health risk Cryptosporidium oocyst isolates, and for C. parvum oocyst isolates from human and bovine feces that represent a high public health risk. There was no cross-reactivity of EIA with ova of eight medically important helminths, three Eimeria species oocysts, Sarcocystis cruzi sporocysts, and two Candida sp. isolates. All nine snake oocyst isolates (C. serpentis), two of seven lizard oocyst isolates, one turtle oocyst isolate, two avian oocyst isolates (turkey, C. meleagridis), one C. wrairi oocyst isolate from guinea pigs, one C. muris oocyst isolate from hyrax, one heifer C. muris isolate, and two C. muris-like oocyst isolates from a camel were positive by both IFA tests; six of these 19 oocyst isolates were EIA-positive. There was no difference in the sensitivity and specificity between direct and indirect IFA tests. The sensitivity of the EIA and both IFA tests to the C. parvum oocysts was 100%. The EIA showed less cross-reactivity with the non-C. parvum oocysts (24%) than direct or indirect IFA (76%), and was less sensitive to those isolates (20%) than both IFA tests (63%). A simulated sampling model for high and low public health risk Cryptosporidium oocysts showed that the low risk oocyst isolates may constitute up to 35% of all positive environmental samples by direct or indirect IFA determination, and up to 12% of all EIA positive samples. This study indicates a superiority of direct and indirect IFA and EIA for screening of human-or-bovine-origin fecal specimens, whereas testing of environmental samples may lead to misidentification of medically important isolates. The results demonstrated that the EIA kit can more accurately identify environmental samples containing oocytes pathogenic for humans than both IFA tests. The specificity of commercially available diagnostic kits to C. parvum should be critically examined for cross-species identification before they are recommended or adopted for use in testing environmental samples.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=86007650002-9637 Journal Article8600765Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, The Johns Hopkins University, Baltimore, Maryland, USA. (~?SUrban, J. F., Jr. Fayer, R. Chen, S. J. Gause, W. C. Gately, M. K. Finkelman, F. D.1996nIL-12 protects immunocompetent and immunodeficient neonatal mice against infection with Cryptosporidium parvum263-8 J Immunol1561Adjuvants, Immunologic/biosynthesis/pharmacology Animals Animals, Newborn/*immunology Coccidiostats/*therapeutic use Cryptosporidiosis/immunology/parasitology/*prevention & control Cryptosporidium parvum/*drug effects/*immunology Interferon Type II/physiology Interleukin-12/biosynthesis/*therapeutic use Mice Mice, Inbred BALB C Mice, SCID Severe Combined Immunodeficiency/immunology/parasitology/prevention & control Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.Jan 17The protozoan parasite Cryptosporidium parvum (Cp) causes diarrhea that can be acutely severe in immunocompetent persons and can become chronic and life-threatening in immunocompromised individuals. Because studies in mice have implicated IFN-gamma in protection against this parasite, and IL-12 can induce IFN-gamma production, we examined the ability of IL-12 to prevent or cure Cp infection in neonatal BALB/c and SCID mice. Treatment of both immunocompetent and immunodeficient mice with IL-12 before experimental inoculation with Cp oocysts prevented or greatly reduced the severity of infection. Intestinal epithelial cell invasion and/or early intracellular development of Cp was inhibited by exogenous IL-12. The protective effect of IL-12 was completely blocked by anti-IFN-gamma mAb. Established infections were associated with elevated IFN-gamma gene expression and were not ameliorated by IL-12 treatment, even though such treatment further enhanced IFN-gamma gene expression. The severity of Cp infection was, however, exacerbated by treatment with anti-IL-12 Ab. These observations provide the first evidence that treatment with exogenous IL-12 can prevent Cp infection through an IFN-gamma-dependent, specific immune system-independent mechanism, and that endogenous IL-12 production has a role in limiting Cp infection.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=85984710022-1767 Journal Article8598471fParasite Immunobiology Laboratory, United States Department of Agriculture, Beltsville, MD 20705, USA. ~?'Jenkins, M. Kerr, D. Fayer, R. Wall, R.1995{Serum and colostrum antibody responses induced by jet-injection of sheep with DNA encoding a Cryptosporidium parvum antigen1658-64Vaccine1317Animals Antibodies, Protozoan/*biosynthesis/*blood Antigens, Protozoan/administration & dosage/*genetics/*immunology Colostrum/*immunology Cryptosporidiosis/immunology/prevention & control/veterinary Cryptosporidium parvum/*immunology DNA, Protozoan/administration & dosage/*immunology Female Hindlimb Injections, Jet Mammary Glands, Animal Protozoan Vaccines/immunology Sheep Sheep Diseases/immunology/prevention & control Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Vaccines, Synthetic/immunologyDecIn an effort to generate high titer colostrum for immunotherapy of cryptosporidiosis, a study was conducted to test the efficacy of immunizing sheep with recombinant plasmid DNA (pCMV-CP15/60) encoding epitopes of 15 and 60 kDa surface antigens of Cryptosporidium parvum sporozoites. The plasmid DNA was used to immunize preparturient ewes at three dose levels by jet-injection into either hind limb muscle (IM) or mammary tissue (IMAM). Regardless of route of injection, a dose-dependent anti-CP15/60 immunoglobulin response was observed in sera and colostrum from sheep immunized with pCMV-CP15/60 plasmid DNA. High titer antibody responses were observed in one of three animals per group receiving an IM injection of 100 or 1000 micrograms pCMV-CP15/60. IMAM immunization with 100 or 1000 micrograms pCMV-CP15/60 plasmid DNA elicited higher titer colostrum responses and more consistent serum responses compared to IM injections. A negligible serum and colostrum anti-CP15/60 response was observed in ewes injected IM with 10 micrograms pCMV-CP15/60 or 1000 micrograms control plasmid DNA. Immunoblotting of native C. parvum sporozoite/oocyst protein with hyperimmune serum and colostrum corroborated the increased titers against CP15/60 antigen. Serum and colostrum antibodies from pCMV-CP15/60-immunized sheep were eluted from native CP15 protein and bound a surface antigen of C. parvum sporozoites as indicated by indirect immunofluorescence staining.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=87195160264-410x Journal Article8719516bParasite Immunobiology Laboratory, Agricultural Research Service, USDA, Beltsville, MD 20705, USA.~?Fayer, R. Fetterer, R.1995LActivity of benzimidazoles against cryptosporidiosis in neonatal BALB/c mice794-5 J Parasitol815Animals Animals, Newborn Benzimidazoles/*therapeutic use Coccidiostats/*therapeutic use Cryptosporidiosis/*drug therapy/parasitology Cryptosporidium parvum/drug effects/isolation & purification Intestines/parasitology Mice Mice, Inbred BALB COctThe need for an effective compound for the prevention and treatment of cryptosporidiosis in humans and animals has led to the testing of benzimidazoles based on reports that albendazole was clinically effective against related protozoan parasites causing microsporidiosis in humans. Albendazole and other benzimidazole derivatives were tested for prophylactic efficacy against cryptosporidiosis at dosage levels 1-3x the levels found effective for treatment of cattle or sheep for helminth infections. Daily dosage levels of thiabendazole, parbendazole, oxibendazole, mebendazole, and albendazole, as high as 200, 30, 10, 15, and 15 mg/kg of body weight, respectively, were not efficacious in neonatal mice. Although the number of parasites in histologic sections of intestine from mice mediated with 15 mg albendazole/kg of body weight was significantly lower than in unmedicated control mice, suggesting activity against the parasite, a high percentage of epithelial cells in the medicated mice were infected.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=74728800022-3395 Journal Article7472880Parasite Immunobiology Laboratory, Livestock and Poultry Sciences Institute, United States Department of Agriculture, Beltsville, Maryland 20705, USA. v~?Weigel, R. M. Dubey, J. P. Siegel, A. M. Hoefling, D. Reynolds, D. Herr, L. Kitron, U. D. Shen, S. K. Thulliez, P. Fayer, R. et al.,1995JPrevalence of antibodies to Toxoplasma gondii in swine in Illinois in 19921747-51J Am Vet Med Assoc20611Age Factors Animals Antibodies, Protozoan/*isolation & purification Female Illinois/epidemiology Prevalence Seroepidemiologic Studies Support, U.S. Gov't, Non-P.H.S. Swine Swine Diseases/*epidemiology/immunology Toxoplasma/*immunology Toxoplasmosis, Animal/*epidemiology/immunologyJun 1>A serologic survey that tested for antibodies to Toxoplasma gondii was conducted, using the modified direct agglutination test, on 6,965 serum samples collected from swine in 179 herds in Illinois in 1992. In breeding swine, results for 1,057 of 5,080 (20.8%) sera tested were positive. In growing/finishing swine, results for 59 of 1,885 (3.1%) sera tested were positive, which was substantially lower than the seroprevalence rate estimated in a serosurvey of pigs from abattoirs in Illinois in 1983 and 1984. Data in the survey reported here were summarized for herds having at least 28 samples/herd. Among all herds, the median, mean, and maximum seroprevalence rates were 6.7, 16.1, and 96.8%, respectively, for breeding swine in 172 herds, and 0.0, 2.8, and 20.0%, respectively, for growing/finishing pigs in 44 herds. Among the 172 herds with breeding swine, 61 (35.5%) had no seropositive pigs. Among the 44 herds with growing/finishing swine, 28 (63.6%) had no seropositive pigs. A logistic regression model was used to estimate that the cumulative risk of T gondii infection for swine in herds containing seropositive pigs was 9.0% by 6 months of age for a herd that had the median seroprevalence rate. In contrast, for pigs in herds in the upper quartile of seroprevalence rates, risk of infection by 6 months of age was estimated to be greater than 20%. Analysis of these data would suggest that overall prevalence of T gondii infection in pigs from Illinois is low; nevertheless, there is a small proportion of farms for which the rate of T gondii infection in swine is moderately high.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=77822490003-1488 Journal Article7782249qDepartment of Veterinary Pathobiology, College of Veterinary Medicine, University of Illinois, Urbana 61801, USA. ~?)Fayer, R. Graczyk, T. K. Cranfield, M. R.1995Multiple heterogenous isolates of Cryptosporidium serpentis from captive snakes are not transmissible to neonatal BALB/c mice (Mus musculus)482-4 J Parasitol813Animal Feed/parasitology Animals Animals, Newborn Animals, Zoo/*parasitology Boidae/*parasitology Cryptosporidiosis/*transmission Feces/parasitology Food Parasitology Mice Mice, Inbred BALB C Support, Non-U.S. Gov'tJunHOral inoculations of 9 litter-groups of 3 5-day-old suckling BALB/c mouse pups (Mus musculus) with 6.7 x 10(3) to 1.2 x 10(5) per pup of viable, Cryptosporidium serpentis oocysts from snakes resulted in no transmission. Mice showed normal development; the litter-group weight gain was not altered significantly (P > 0.05) relative to the total number of C. serpentis oocysts inoculated or to the initial group weight (P > 0.05). Histological sections of stomach, duodenum, jejunum, ileum, cecum, and colon 4 days postinoculation did not contain life-cycle stages of Cryptosporidium in any inoculated mice. Because these neonatal, C. parvum-susceptible BALB/c mice were resistant to infection it is unlikely that C. serpentis transmission to the snakes "via infected prey" results when captive snakes are maintained on a diet of BALB/c mice.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=77761380022-3395 Journal Article7776138bParasite Immunobiology Laboratory, Agricultural Research Service, Beltsville, Maryland 20705, USA.`~?Jenkins, M. C. Fayer, R.1995SCloning and expression of cDNA encoding an antigenic Cryptosporidium parvum protein149-52Mol Biochem Parasitol711Amino Acid Sequence Animals Antigens, Protozoan/genetics Base Sequence Cloning, Molecular Cryptosporidium parvum/*genetics/immunology DNA, Complementary/*genetics Epitopes/analysis Escherichia coli/genetics Genes, Structural, Protozoan/*genetics Molecular Sequence Data Molecular Weight Open Reading Frames/genetics Pichia/genetics Protozoan Proteins/chemistry/*genetics/immunology Recombinant Proteins/immunology Sequence Analysis, DNAAprdhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=75431820166-6851 Journal Article7543182fParasite Immunobiology Laboratory Agricultural Research Service LPSI, USDA, Beltsville, MD 20705, USA.@~? Fayer, R.1995pEffect of sodium hypochlorite exposure on infectivity of Cryptosporidium parvum oocysts for neonatal BALB/c mice844-6Appl Environ Microbiol612.Animals Animals, Newborn Cryptosporidiosis/etiology/parasitology/prevention & control Cryptosporidium parvum/*drug effects/growth & development/pathogenicity Female Human Intestines/parasitology Mice Mice, Inbred BALB C Sodium Hypochlorite/administration & dosage/*pharmacology Time Factors Trypan BlueFebXOocysts of Cryptosporidium parvum suspended in 5.25, 2.63, or 1.31% aqueous sodium hypochlorite (Clorox laundry bleach) for 10, 30, 60, or 120 min at 21 degrees C were administered by gastric intubation to neonatal BALB/c mice. Microscopic examination of intestinal tissue sections revealed developmental stages of C. parvum in all of the mice.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=75746260099-2240 Journal Article7574626cParasite Immunobiology Laboratory, U.S. Department of Agriculture, Beltsville, Maryland 20705, USA.~?DBarr, S. C. Jamrosz, G. F. Hornbuckle, W. E. Bowman, D. D. Fayer, R.1994>Use of paromomycin for treatment of cryptosporidiosis in a cat1742-3J Am Vet Med Assoc20512Animals Cat Diseases/*drug therapy Cats Cryptosporidiosis/*drug therapy Cryptosporidium/isolation & purification Diarrhea/drug therapy/*veterinary Feces/parasitology Female Paromomycin/*therapeutic useDec 15 Cryptosporidium oocysts were found in the feces of a 6-month-old female cat with persistent diarrhea. The oocysts disappeared from the feces immediately after treatment with paromomycin (165 mg/kg of body weight, PO, for 5 days), and the diarrhea eventually resolved.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=7744647&0003-1488 Case Reports Journal Article7744647fDepartment of Clinical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, NY 14853.i~?Fayer, R. Ellis, W.1994zQinghaosu (artemisinin) and derivatives fail to protect neonatal BALB/c mice against Cryptosporidium parvum (Cp) infection41SJ Eukaryot Microbiol415Animals Animals, Newborn Antiprotozoal Agents/*pharmacology *Artemisinins Cryptosporidiosis/*prevention & control *Cryptosporidium parvum Drug Evaluation, Preclinical Mice Mice, Inbred BALB C Sesquiterpenes/*pharmacologySep-Octdhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=78042411066-5234 Journal Article7804241%USDA, ARS, LPSI Beltsville, MD 20705.O~? Fayer, R.1994ODevelopment of a precocious strain of Cryptosporidium parvum in neonatal calves40SJ Eukaryot Microbiol415Animals Animals, Newborn Cattle Cryptosporidiosis/parasitology Cryptosporidium parvum/*growth & development/isolation & purification/pathogenicity Disease Models, Animal Feces/parasitology Intestinal Diseases, Parasitic/parasitology Male VirulenceSep-Octdhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=78042401066-5234 Journal Article7804240%USDA, ARS, LPSI Beltsville, MD 20705.~? Fayer, R.1994TEffect of high temperature on infectivity of Cryptosporidium parvum oocysts in water2732-5Appl Environ Microbiol608wAnimals Cattle Cryptosporidium parvum/*pathogenicity *Heat Intestines/parasitology Male Mice Mice, Inbred BALB C *WaterAugCryptosporidium parvum oocysts suspended in 0.5 ml of distilled water were pipetted into plastic vials which were inserted into wells in the heated metal block of a thermal DNA cycler. Block temperatures were set at 5 degrees C incremental temperatures from 60 to 100 degrees C. At each temperature setting four vials containing C. parvum oocysts were placed into wells and held for 15 s before time was recorded as zero, and then pairs of vials were removed 1 and 5 min later. Upon removal, all vials were immediately cooled on crushed ice. Also, at each temperature interval one vial containing 0.5 ml of distilled water was placed in a well and a digital thermometer was used to record the actual water temperature at 30-s intervals. Heated oocyst suspensions as well as unheated control suspensions were orally inoculated by gavage into 7- to 10-day-old BALB/c mouse pups to test for infectivity. At 96 h after inoculation the ileum, cecum, and colon from each mouse were removed and prepared for histology. Tissue sections were examined microscopically. Developmental-stage C. parvum was found in all three gut segments from all mice that received oocysts in unheated water and in water that reached temperatures of 54.4, 59.9, and 67.5 degrees C at 1 min when vials were removed from the heat source. C. parvum was also found in the ileum of one of six mice that received oocysts in water that reached a temperature of 59.7 degrees C at 5 min.(ABSTRACT TRUNCATED AT 250 WORDS)dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=80858160099-2240 Journal Article8085816]Zoonotic Diseases Laboratory, USDA Agricultural Research Service, Beltsville, Maryland 20705. ~?eZu, S. X. Li, J. F. Barrett, L. J. Fayer, R. Shu, S. Y. McAuliffe, J. F. Roche, J. K. Guerrant, R. L.1994}Seroepidemiologic study of Cryptosporidium infection in children from rural communities of Anhui, China and Fortaleza, Brazil1-10Am J Trop Med Hyg511Adolescent Adult Age Factors Animals Antibodies, Protozoan/*blood Brazil/epidemiology Child Child, Preschool China/epidemiology Comparative Study Cross-Sectional Studies Cryptosporidiosis/*epidemiology Cryptosporidium/*immunology Enzyme-Linked Immunosorbent Assay Feces/parasitology Female Human Immunoglobulin G/blood Immunoglobulin M/blood Infant Male Prevalence Reproducibility of Results Rural Population Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S.Jul A cluster-sampling, cross-sectional study was conducted for assessing the prevalence of Cryptosporidium infection in children less than 16 years of age from three villages, Dondian, Linshan, and Fuziyin, in rural Anhui in eastern China. Among 320 apparently healthy children less than 10 years of age from Dondian who had stool specimens collected, cryptosporidial oocysts were found in stools of three children from Dondian, and no positive specimens were found in 239 children studied from Linshan. In addition, a total of 610 serum samples from children in these three villages were tested for specific IgG antibody to Cryptosporidium with an enzyme-linked immunosorbent assay (ELISA) and the prevalence rates were 42.3%, 51.7%, and 57.5%, respectively, in Dondian, Linshan, and Fuziyin. Seroprevalence increased progressively with age. No detectable antibody was found in infants between two and six months of age, and seropositivity steadily increased after one year of age. Among 36 sera from adults 15-60 years of age without diarrheal illness in Huanglu villages of rural Chaohu, 50% (18 of 36) were positive. As expected, a good correlation was found in the specific IgG antibody between the paired serum specimens from 30 matched mother-neonates who showed transplacental transfer of IgG. However, little or no IgM antibody was seen in the neonates even though several mothers had a positive anticryptosporidial IgM enzyme-linked immunoassay result. Forty randomly selected serum samples from children less than four years of age in a similarly impoverished semiurban community in Fortaleza, Brazil, where the majority of households also have pit toilets and shared community water supplies and 172 serum samples from patients one month to 29 years of age admitted to the University of Virginia Hospital without diarrhea were also examined. In Fortaleza, almost all children acquired antibody by their second year of life, demonstrating the high prevalence of this infection. In rural Anhui, only about half the children were infected by 5-7 years of age. The overall prevalence rate (16.9%) of seropositivity among children and young adults in Virginia was much lower than in China and Brazil. These results indicate that cryptosporidial infection is ubiquitous, and is highly endemic in these impoverished communities. The difference between China and Brazil may reflect earlier weaning, hygiene practices, poorer water or sanitation, multiple siblings in family and geographic environment in Brazil.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=80599060002-9637 Journal Article8059906xDepartment of Clinical Epidemiology and Community Medicine, Anhui Medical University, Hefei, People's Republic of China.~?Fayer, R. Ellis, W.1993MParomomycin is effective as prophylaxis for cryptosporidiosis in dairy calves771-4 J Parasitol795Animals Animals, Newborn Cattle Cattle Diseases/*prevention & control Cryptosporidiosis/*prevention & control Diarrhea/prevention & control/*veterinary Feces/parasitology Paromomycin/*therapeutic use Weight GainOctMOf 16 experimentally infected neonatal dairy calves, 12 were fed paromomycin twice daily in their milk for 11 consecutive days beginning 1 day before oral inoculation with 1.5-2.0 x 10(6) oocysts of Cryptosporidium parvum. Four calves each in groups A, B, C, and D received total daily doses of 100, 50, 25, and 0 mg of paromomycin per kilogram of body weight, respectively. From birth until 28 days of age feces from each calf were examined for diarrhea, and oocysts were enumerated, rectal temperature was recorded, and weight gain was determined. Total days of diarrhea, severity of diarrhea, the total number of days oocysts were shed, and the number of oocysts shed were significantly less in group A than in the unmedicated group D. The severity of diarrhea was also significantly less in groups B and C than in group D. Oocysts were not detected in feces from calves in group A. Except for 1 calf, oocysts were not detected from calves in groups B and C during the first week the drug was administered and those calves that shed oocysts began shedding at or near the end of paromomycin administration or more than 1 wk after treatment ended. Frequency of fever and weight gains did not vary significantly between the unmedicated and medicated groups except for group C, calves of which gained significantly less weight than those in all other groups.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=84105520022-3395 Journal Article8410552^Zoonotic Diseases Laboratory, U.S. Department of Agriculture, Beltsville, Maryland 20705-2350._~?Fayer, R. Ellis, W.1993Glycoside antibiotics alone and combined with tetracyclines for prophylaxis of experimental cryptosporidiosis in neonatal BALB/c mice553-8 J Parasitol794Animals Animals, Newborn Anti-Bacterial Agents/*therapeutic use Azithromycin Clarithromycin/therapeutic use Cryptosporidiosis/*prevention & control *Cryptosporidium parvum Disease Models, Animal Doxycycline/therapeutic use Drug Synergism Drug Therapy, Combination Erythromycin/analogs & derivatives/therapeutic use Mice Mice, Inbred BALB C Minocycline/therapeutic use Paromomycin/therapeutic use Tetracyclines/*therapeutic useAugGlycoside antibiotics including the macrolide antibiotics azithromycin, clarithromycin, and erythromycin and the aminoglycoside paromomycin were administered alone or combined with doxycycline, minocycline, or tetracycline to neonatal BALB/c mice experimentally infected with Cryptosporidium parvum. Glycosides at 100 or 200 mg/kg of body weight and tetracyclines at 50 mg/kg of body weight were dissolved in dimethylsulfoxide (DMSO), which was then diluted with phosphate-buffered saline (PBS) and given orally by gavage. Drugs were administered at 0, 24, 48, and 72 hr postinfection (PI) for prophylaxis. Histologic sections of ileum, cecum, and colon from tissues fixed at 96 hr PI were examined microscopically to determine the number of developing parasites and assign a quantitative score based on infectivity. All groups that received glycosides had significantly (P < 0.01) lower scores than controls that received only DMSO/PBS. A range in efficacy was apparent. None or extremely few parasites were found in paromomycin- and azithromycin-treated groups, whereas few to moderate numbers of parasites were found in erythromycin- and clarithromycin-treated groups. The addition of tetracyclines did not consistently result in significantly lower scores.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=83925410022-3395 Journal Article8392541^Zoonotic Diseases Laboratory, U.S. Department of Agriculture, Beltsville, Maryland 20705-2350.~?0Jenkins, M. C. Fayer, R. Tilley, M. Upton, S. J.1993Cloning and expression of a cDNA encoding epitopes shared by 15- and 60-kilodalton proteins of Cryptosporidium parvum sporozoites2377-82 Infect Immun616Amino Acid Sequence Animals Antibodies, Monoclonal/immunology Base Sequence Cattle Cloning, Molecular Cryptosporidium parvum/genetics/*immunology DNA, Protozoan Epitopes/*genetics Fluorescent Antibody Technique Immune Sera/immunology Molecular Sequence Data Nucleic Acid Hybridization Protozoan Proteins/genetics/*immunology/isolation & purification Rats Recombinant Proteins/genetics/*immunology/isolation & purificationJun%A cDNA (CP15/60) encoding epitopes of Cryptosporidium parvum 15- and 60-kDa sporozoite proteins was isolated and expressed in Escherichia coli toward the goal of developing an immunogen for producing high-titer anticryptosporidial colostrum. Antisera prepared in rats to native C. parvum 15-kDa protein and used to identify the CP15/60 bacteriophage clone recognized both 15- and 60-kDa in vitro translation products derived from sporozoite RNA. Antisera specific for recombinant CP15/60 antigen recognized native 15- and 60-kDa C. parvum sporozoite proteins by immunoblotting and identified both surface and internal antigens on C. parvum sporozoites by immunofluorescence staining. Northern (RNA) and Southern blot hybridization experiments using sporozoite RNA and DNA indicated that CP15/60 DNA is transcribed as a single 1.4-kb RNA species from a single-copy gene. Recombinant CP15/60 antigen was recognized by hyperimmune colostrum from cows immunized with C. parvum oocyst-sporozoite protein and by convalescent-phase sera from C. parvum-infected calves.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=76847260019-9567 Journal Article7684726ZProtozoan Diseases Laboratory, U.S. Department of Agriculture, Beltsville, Maryland 20705.l~?iGranstrom, D. E. Dubey, J. P. Davis, S. W. Fayer, R. Fox, J. C. Poonacha, K. B. Giles, R. C. Comer, P. F.1993_Equine protozoal myeloencephalitis: antigen analysis of cultured Sarcocystis neurona merozoites88-90J Vet Diagn Invest51Animals Antigens, Protozoan/*analysis/isolation & purification Cattle Cells, Cultured Electrophoresis, Polyacrylamide Gel *Horse Diseases Horses Immunoblotting Molecular Weight Sarcocystis/*immunology/isolation & purification Sarcocystosis/*veterinary Support, Non-U.S. Gov'tJaniAntigens of cultured Sarcocystis neurona merozoites were examined using immunoblot analysis. Blotted proteins were probed with S. cruzi, S. muris, and S. neurona antisera produced in rabbits, S. fayeri (pre- and post-infection) and S. neurona (pre- and post-inoculation) sera produced in horses, immune sera from 7 histologically confirmed cases of equine protozoal myeloencephalitis (EPM), and pre-suckle serum from a newborn foal. Eight proteins, 70, 24, 23.5, 22.5, 13, 11, 10.5, and 10 Kd, were detected only by S. neurona antiserum and/or immune serum from EPM-affected horses. Equine sera were titered by the indirect immunofluorescent antibody (IFA) method using air-dried, cultured S. neurona merozoites. Anti-Sarcocystis IFA titers were found in horses with or without EPM. Serum titers did not correspond to the number of specific bands recognized on immunoblots.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=84669881040-6387 Journal Article8466988ODepartment of Veterinary Science, University of Kentucky, Lexington 40546-0099.~?/Zu, S. X. Fang, G. D. Fayer, R. Guerrant, R. L.1992.Cryptosporidiosis: Pathogenesis and immunology24-27Parasitol Today81JanxCryptosporidium parvum is an increasingly recognized agent of intestinal infection in normal and immunocompromised humans, and in many other animals. The intraepithelial cell infection results in villous atrophy, mild submucosal in flammation, reduction of brush-border enzymes and a characteristic persistent watery diarrhea. The infection is self-limiting in immunocompetent hosts, probably because of specific acquired immunity; specific serum and secretory antibody responses develop that may be required for clearance and protection against reinfection. Passive milk antibody, especially i f in high titers, may be partially protective but severe, persistent infection in athymic rodents and humans with AIDS demonstrate that T cells are essential for controlling the infection. Specific anti-bodies and lymphocyte extracts have been tested in cases of cryptosporidiosis but the interpretation of the results remains controversial. Here, Shu-Xian Zu, Guo-Dong Fang, Ronald Foyer and Richard Guerrant emphasize that effective treatment and prevention remain dependent on advances in our understanding of the host cell-parasite relationship.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=154635220169-4758 Journal article15463522YShu-Xian Zu, Guo-Dong Fang and Richard Guerrant are at the Division of Geographic Medicine, Department of Medicine, University of Virginia School of Medicine, Charlottesville, VA 22908, USA; Shu-Xian Zu is also at the Department of Clinical Epidemiology and Community Medicine, Anhui Medical University, Hefei 230032, People's Republic of China. E~?Fayer, R. Jenkins, M. C.1992pColostrum from cows immunized with Eimeria acervulina antigens reduces parasite development in vivo and in vitro1637-45 Poult Sci7110iAnimals Antibodies, Protozoan/analysis Antigens, Protozoan/*immunology Cattle/*immunology Chickens/*immunology/parasitology Coccidiosis/parasitology/prevention & control/*veterinary Colostrum/*immunology Duodenum/parasitology Eimeria/*immunology Enzyme-Linked Immunosorbent Assay Female *Immunization, Passive Poultry Diseases/parasitology/*prevention & controlOct%Experiments were undertaken to determine whether passive immunization utilizing hyperimmune bovine colostrum (HBC) specific for Eimeria acervulina (EA) antigens conferred protection against coccidiosis in chickens. The HBC was produced by immunizing three pregnant, nonmilking Jersey cows with EA antigens administered via one intramuscular injection followed by three intramammary infusions at approximately 10, 8, 6, and 4 wk before parturition. One cow was immunized with sporozoites (SZ), the second with merozoites (MZ), and the third with recombinant merozoite antigen (rMZ). A fourth cow, unimmunized, provided normal colostrum (NC) for control purposes. Colostral whey from each cow was tested by ELISA for antibody against SZ, MZ, and rMZ antigens. In all immunized cows, antiparasite titers were elevated above those of the control. Antibodies from MZ- and rMZ-immunized cows recognized both MZ and rMZ antigen. Separate groups of 2-wk-old chickens received two oral doses of anti-SZ, -MZ, or -rMZ HBC or NC or PBS daily from 1 day before through 6 days after oral inoculation (DAI) with EA oocysts. Feces from each group were examined for oocysts. Intestines were examined for lesions 6 DAI. Histologic sections of duodenum were examined for asexual stages and gametocytes utilizing monoclonal antibody and fluorescence microscopy. In Experiments 1 and 2, oocyst production was reduced in all HBC-treated groups, except one treated with rMZ HBC, compared with PBS- or NC-treated groups. In Experiment 2, the severity of lesions was significantly reduced in all HBC-treated groups compared with those that received NC or PBS. Significantly fewer developmental stages were found in histological sections from all chickens treated with anti-SZ and anti-rMZ HBC than from controls. Anti-SZ HBC significantly reduced the number of intracellular SZ found 24 h after their inoculation into cultures of primary chicken kidney cells. These results suggest that HBC specific for certain EA antigens can inhibit parasite development and reduce severity of parasite-related gut lesions.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14546820032-5791 Journal Article1454682[Zoonotic Diseases Laboratory, Beltsville Agricultural Research Center, Maryland 20705-2350.~? Fayer, R.1992FActivity of sulfadimethoxine against cryptosporidiosis in dairy calves534-7 J Parasitol783Administration, Oral Animals Cattle Cattle Diseases/*drug therapy Cryptosporidiosis/*drug therapy Diarrhea/drug therapy/*veterinary Feces/parasitology Male Sulfadimethoxine/administration & dosage/*therapeutic use Weight GainJunOf 13 neonatal calves inoculated orally with 1.5 x 10(6) oocysts of Cryptosporidium parvum, 7 in group A were fed 5-g boluses of sulfadimethoxine for 21 consecutive days beginning 1 day before infection, and 6 calves in group B were untreated controls. Calves in group A had diarrhea for 6-18 days (mean = 11 days); those in group B had diarrhea for 4-14 days (mean = 8.7 days). The severity of diarrhea, based on a daily numerical scoring system, was similar for both groups. Calves in group A shed an average of 18 x 10(6) oocysts/ml of feces for 3.9 days; those in group B shed an average of 2.4 x 10(6) oocysts/ml of feces for 5.3 days. By 28 days of age, calves in group A vs. group B gained an average of 8.9 kg vs. 15.7 kg. These findings indicate that sulfadimethoxine did not significantly reduce the number of days or severity of diarrhea, or the number of oocysts or patent period, nor did it improve weight gains.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=15978030022-3395 Journal Article1597803hUnited States Department of Agriculture, Agricultural Research Service, Beltsville, Maryland 20705-2350.~?SChrisp, C. E. Suckow, M. A. Fayer, R. Arrowood, M. J. Healey, M. C. Sterling, C. R.1992tComparison of the host ranges and antigenicity of Cryptosporidium parvum and Cryptosporidium wrairi from guinea pigs406-9 J Protozool393Animals Antibodies, Monoclonal/immunology Antibodies, Protozoan/*immunology Cattle Comparative Study Cross Reactions Cryptosporidiosis/immunology/parasitology Cryptosporidium/immunology/*physiology Cryptosporidium parvum/immunology/*physiology Female Guinea Pigs Host-Parasite Relations Mice Mice, Inbred C57BL Specific Pathogen-Free Organisms Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.May-JunvOocysts of a Cryptosporidium isolate from guinea pigs were not infectious for adult mice, but were infectious for two of three newborn calves and for suckling mice. However, oocysts isolated from calves or mice infected with guinea pig Cryptosporidium were not infectious for guinea pigs. Four isolates of C. parvum from calves were incapable of infecting weanling guinea pigs. Microscopic examination of tissue from the colon and cecum of suckling guinea pigs inoculated with C. parvum revealed sparse infection of some pups. These host range studies and previously described differences in 125I-labeled oocyst surface protein profiles between Cryptosporidium sp. from guinea pigs and C. parvum suggest they are distinct species. We propose the name Cryptosporidium wrairi be retained. Studies with monoclonal antibodies indicate that C. wrairi and C. parvum are antigenically related.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=13863850022-3921 Journal Article1386385\Unit for Laboratory Animal Medicine, University of Michigan Medical School, Ann Arbor 48109.@~?Fayer, R. Elsasser, T. H.1991<Bovine sarcocystosis: How parasites negatively affect growth250-5Parasitol Today79Growth, though genetically encoded, is markedly influenced in healthy animals by the interaction of hormonal and nutritional factors. The uptake and use of nutrients by specific tissues is regulated by a priority system that modulates physiological processes. Nutritional, hormonal and immunological consequences of parasitism often lead to partitioning of nutrients away from growth. In this article, Ron Foyer and Ted Elsosser use a bovine sarcocystosis model to show that changes in plasma concentrations of insulin-like growth factor-I (IGF-I), growth hormone (GH) and somotostotin (SSN), as well as the host's immunological response to the parasite via cytokine interactions with the endocrine system, are modulators of perturbed growth.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=154635110169-4758 Journal Article154635116Ron Foyer is at the Zoonotic Diseases Laboratory, USA.~?VFayer, R. Tilley, M. Upton, S. J. Guidry, A. J. Thayer, D. W. Hildreth, M. Thomson, J.1991iProduction and preparation of hyperimmune bovine colostrum for passive immunotherapy of cryptosporidiosis38S-39S J Protozool386Animals Blotting, Western Cattle Colostrum/*immunology/microbiology/radiation effects Cryptosporidiosis/*therapy Enzyme-Linked Immunosorbent Assay Female Freeze Drying Immunoglobulin Isotypes/immunology *Immunotherapy, Adoptive VaccinationNov-Dec$Pregnant cows were immunized to produce hyperimmune bovine colostrum (HBC) by intramuscular injection or intramammary infusion (TI) followed by 3 successive TI boosters with Cryptosporidium parvum (Cp) oocyst antigen mixed with Freund's (F) or Ribi (R) adjuvant. Control cows received no Cp. Colostrum from all cows was skimmed of butterfat and tested for specific anti-Cp immunoglobulin isotypes by ELISA. The HBC from Cp-F and Cp-R immunized cows had IgG1 titers exceeding 1:400,000 and 1:800,000, respectively. Some HBC from Cp-F immunized cows was freeze-dried to facilitate storage and some were irradiated at 42.5 kGy to kill potentially contaminating pathogens. Freeze-drying, but not irradiation, reduced IgG1 titers by only one dilution. Neither treatment affected Western blot banding patterns.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18181910022-3921 Journal Article1818191&USDA, ARS, LPSI, Beltsville, MD 20705.z~?1Hoskins, D. Chrisp, C. E. Suckow, M. A. Fayer, R.1991Effect of hyperimmune bovine colostrum raised against Cryptosporidium parvum on infection of guinea pigs by Cryptosporidium wrairi 185S-186S J Protozool386Animals Cattle Colostrum/*immunology Cryptosporidiosis/*prevention & control Cryptosporidium parvum/*immunology Guinea Pigs Immunotherapy Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.Nov-DecOocysts shedding was markedly reduced in guinea pigs inoculated intraintestinally with Cryptosporidium wrairi sporozoites that had been incubated with hyperimmune bovine colostrum raised to C. parvum when compared with shedding in guinea pigs inoculated with sporozoites incubated in either non-immune bovine colostrum or buffered saline. However oocyst shedding was apparently not reduced in guinea pigs inoculated by gavage with oocysts of C. wrairi and subsequently treated twice daily per os with hyperimmune bovine colostrum. Similarly, oocyst shedding was apparently not reduced by oral treatment with hyperimmune bovine colostrum when treatment was begun simultaneously with inoculation of C. wrairi oocysts.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=18181600022-3921 Journal Article1818160\Unit for Laboratory Animal Medicine, University of Michigan Medical School, Ann Arbor 48109.~?4Fayer, R. Barta, J. R. Guidry, A. J. Blagburn, B. L.1991uImmunogold labeling of stages of Cryptosporidium parvum recognized by immunoglobulins in hyperimmune bovine colostrum487-90 J Parasitol773Animals Cattle Colostrum/*immunology Cryptosporidium/*immunology/ultrastructure Ileum/*parasitology/ultrastructure Immunoglobulins/*immunology Immunohistochemistry Mice Mice, Inbred BALB C Microscopy, ElectronJun)Ultrathin sections of mouse ileum infected with Cryptosporidium parvum were stained by immunogold techniques. Sections first were stained with polyvalent antibodies in whey from hyperimmune bovine colostrum (HBC), then stained by secondary antibodies in rabbit antibovine IgA, IgM, IgG1, and IgG2, and lastly labeled by goat anti-rabbit gold conjugate. Examination of the immunostained specimens by electron microscopy revealed that each bovine immunoglobulin isotype in the whey recognized antigens in meronts, merozoites, microgametocytes, microgametes, and macrogamonts. Based on these findings it is hypothesized that antigens in all stages of C. parvum provide targets of opportunity for the antiparasitic activity of HBC whey antibodies thereby accounting for its efficacy as an immunotherapeutic agent.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20409610022-3395 Journal Article2040961&USDA, ARS, Beltsville, Maryland 20705.~?;Fayer, R. Nerad, T. Rall, W. Lindsay, D. S. Blagburn, B. L.19915Studies on cryopreservation of Cryptosporidium parvum357-61 J Parasitol773Animals Animals, Newborn *Cryopreservation *Cryoprotective Agents Cryptosporidiosis/*parasitology Cryptosporidium/*growth & development Dimethyl Sulfoxide Glycerol Mice Mice, Inbred BALB C Support, U.S. Gov't, Non-P.H.S.JunbNeonatal BALB/c mice received oocysts or sporozoites of Cryptosporidium parvum pretreated by a variety of cryopreservation protocols. Histologic sections of infected and control mice were examined to determine if pretreated organisms established infection in the intestine. Sporozoites were inoculated rectally, oocysts orally. Freshly excysted sporozoites were frozen in Hanks' balanced salt solution (HBSS) containing dimethylsulfoxide (DMSO) or glycerol at concentrations of 5%, 10%, or 15% at cooling rates of -1 C and -10 C per min. Other sporozoites were frozen to -70 C in the absence of cryoprotectant without controlled reduction of temperature, others placed in HBSS with 10% DMSO but not subjected to freezing, whereas others were placed in vitrification media containing 5.5 M propylene glycol, 6.5 M glycerol, or 8 M ethylene glycol for 1 min before resuspension in fresh HBSS and inoculation into mice. Intact oocysts were frozen without controlled reduction of temperature directly to -70 C in HBSS containing no cryoprotectant or in HBSS that contained 10% DMSO. Others were cooled at -0.3 C per min from 4 C to -70 C in HBSS with 5% or 10% DMSO. Still others were cooled at a rate of -1 C per min until reaching -40 C and then cooled at -10 C per min until reaching -70 C in HBSS with 7.5% DMSO. Oocysts and sporozoites not exposed to cryoprotectants were inoculated into mice orally and rectally, respectively, for control purposes. Only unfrozen oocysts and sporozoites not exposed to cryoprotectant, and some of the unfrozen oocysts and sporozoites exposed to 10% DMSO, successfully established infections in mice.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20409480022-3395 Journal Article2040948eLivestock and Poultry Sciences Institute, U.S. Department of Agriculture, Beltsville, Maryland 20705. ~?xTilley, M. Upton, S. J. Fayer, R. Barta, J. R. Chrisp, C. E. Freed, P. S. Blagburn, B. L. Anderson, B. C. Barnard, S. M.1991_Identification of a 15-kilodalton surface glycoprotein on sporozoites of Cryptosporidium parvum1002-7 Infect Immun593Animals Antibodies, Monoclonal/*immunology/therapeutic use Antibodies, Protozoan/*immunology/therapeutic use Antigens, Protozoan/*immunology Blotting, Western Cross Reactions Cryptosporidiosis/therapy Cryptosporidium/analysis/*immunology Electrophoresis, Polyacrylamide Gel Epitopes/immunology Fluorescent Antibody Technique Immunohistochemistry Membrane Glycoproteins/*analysis/immunology Mice Mice, Inbred BALB C Molecular Weight Parasite Egg Count Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.MarAn immunoglobulin A monoclonal antibody (MAb5C3) was developed against a 15-kDa surface glycoprotein (GP15) of Cryptosporidium parvum sporozoites. Indirect immunofluorescence and colloidal gold immunoelectron microscopy revealed that the antibody reacted with both the sporozoite and merozoite surface plasma membranes. On Western immunoblots, MAb5C3 binding was found to be strongly inhibited when 200 mM N-acetylglucosamine was used as a competing sugar. N-Acetylgalactosamine inhibited binding of the antibody only slightly, whereas glucose, mannose, and galactose failed to inhibit binding. MAb5C3 was found to recognize a similar 15-kDa epitope associated with a Cryptosporidium sp. isolated from guinea pigs. However, MAb5C3 failed to react with any proteins or glycoproteins associated with C. baileyi from chickens, Cryptosporidium sp. (= bovine C. muris) from cattle, C. serpentis from a rat snake, bradyzoites of Besnoitia darlingi from an opossum, sporozoite/oocyst extracts of Caryospora bigenetica from an eastern diamondback rattlesnake, sporozoites of Eimeria nieschulzi and E. papillata from rats and mice, or tachyzoites of Toxoplasma gondii (RH strain). When hybridoma supernatants containing MAb5C3 were administered orally to suckling mice experimentally infected with C. parvum, a 75% reduction in developmental stages was seen histologically at 72 h postinfection and a 67.5% reduction in mean oocyst output was found at 6 days postinfection.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=17052380019-9567 Journal Article1705238>Division of Biology, Kansas State University, Manhattan 66506.Q~?%Lindsay, D. S. Dubey, J. P. Fayer, R.1991Immunohistologic examination of monoclonal antibodies generated against Eimeria bovis sporozoites for reactivity to meronts and sexual stages of E bovis and other eimerian parasites239-42 Am J Vet Res522Animals Antibodies, Monoclonal/*immunology Antibodies, Protozoan/*immunology Cattle Coccidiosis/metabolism/parasitology Comparative Study Eimeria/growth & development/*immunology ImmunohistochemistryFebSeven monoclonal antibodies (MAB) generated against sporozoites of Eimeria bovis were tested for reactivity against immature and mature first-generation meronts, sexual stages, and oocysts in tissues from experimentally infected calves by use of an avidin-biotin peroxidase complex (ABPC) immunohistologic test. Three of the 7 MAB reacted in the ABPC test. One of these, MAB-4FB4, reacted only with mature E bovis meronts. The other 2 MAB, MAB-2AE7 and MAB-4AD7, reacted with all the developmental stages of E bovis tested. Asexual stages and sexual stages of E tenella from chickens and E papillata from mice also were examined in the ABPC test. Monoclonal antibodies MAB-2AE7 and MAB-4AD7 reacted with all stages of these eimerian protozoa. None of the other 5 MAB reacted with these parasites. Results of this study suggested that antigens are shared among the asexual and sexual stages of several diverse Eimeria species.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=20123350002-9645 Journal Article2012335>Zoonotic Diseases Laboratory, USDA, ARS, Beltsville, MD 20705.w~?<Tilley, M. Fayer, R. Guidry, A. Upton, S. J. Blagburn, B. L.1990Cryptosporidium parvum (Apicomplexa: Cryptosporidiidae) oocyst and sporozoite antigens recognized by bovine colostral antibodies2966-71 Infect Immun589 Animals Antibodies, Protozoan/*immunology Antigens, Protozoan/*immunology Blotting, Western Cattle/*immunology Coccidia/*immunology Cryptosporidium/*immunology Immunization Immunoglobulin A, Secretory/*immunology Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S.SepColostral whey from seven hyperimmunized and two control cows (hyperimmune bovine colostrum) was examined by Western immunoblotting for the presence of antibody against oocysts and sporozoites of Cryptosporidium parvum, using rabbit anti-bovine immunoglobulin A (IgA), IgG1, IgG2, and IgM antibodies, followed by a horseradish peroxidase goat anti-rabbit polyvalent antibody. Although considerable variation was found in binding activity between cows on different immunization protocols, IgA and IgG1 in whey recognized a greater variety of C. parvum antigens than did IgG2 and IgM. A band at 9 to 10 kilodaltons appeared unique in that it was recognized only by IgA.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=23876310019-9567 Journal Article2387631>Division of Biology, Kansas State University, Manhattan 66506.$~?$Fayer, R. Guidry, A. Blagburn, B. L.1990Immunotherapeutic efficacy of bovine colostral immunoglobulins from a hyperimmunized cow against cryptosporidiosis in neonatal mice2962-5 Infect Immun589.Animals Animals, Newborn Cattle Coccidiostats/*therapeutic use Cryptosporidiosis/pathology/*therapy Female *Immunization, Passive Immunoglobulin A, Secretory/*immunology Intestinal Diseases, Parasitic/pathology/*therapy Mice Mice, Inbred BALB C Pregnancy Rodent Diseases/parasitology/pathology/*therapySepInfection with Cryptosporidium parvum, a ubiquitous protozoan parasite of virtually all mammals, can cause mild to severe diarrhea in immunocompetent hosts and life-threatening diarrhea in immunocompromised hosts. Passive immunotherapy of experimentally infected animals and naturally infected humans with hyperimmune bovine colostrum has been reported to be efficacious, whereas chemotherapy has not. In this study, the efficacy of specific immunoglobulin isotypes purified from bovine colostrum from a cow hyperimmunized with Cryptosporidium parvum was assessed in neonatal BALB/c mice. Mice were orally infected with oocysts and treated with whole whey immunoglobulin G1 (IgG1), IgG2, IgA, or IgM at six intervals from 22 to 66 h postinfection. In histologic sections of intestine examined at 72 h postinfection, the reduction in number of intestinal stages in treated mice versus untreated controls was very highly significant (P less than 0.0001). The greatest reduction in parasite number was found in mice treated with IgG1, IgA, or whey.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=23876300019-9567 Journal Article2387630eLivestock and Poultry Sciences Institute, U.S. Department of Agriculture, Beltsville, Maryland 20705.K~?%Lindsay, D. S. Dubey, J. P. Fayer, R.1990dExtraintestinal stages of Eimeria bovis in calves and attempts to induce relapse of clinical disease1-9 Vet Parasitol361-2Adrenal Cortex Hormones Animals Antibodies, Monoclonal/diagnostic use Cattle Cattle Diseases/*parasitology Coccidiosis/complications/parasitology/*veterinary Eimeria/immunology/*isolation & purification Immunoenzyme Techniques Immunohistochemistry Intestinal Diseases, Parasitic/complications/parasitology/*veterinary Lymph Nodes/*parasitology Male Mesentery Stress/complications/veterinaryMayMonoclonal antibodies to all life cycle stages of Eimeria bovis were used in an avidin-biotin immunoperoxidase test to examine extraintestinal tissues from experimentally infected calves for developmental stages of the parasite. First-generation meronts of Eimeria bovis were found in the mesenteric lymph nodes (MLNs) of three of four calves examined during the first 18 days of primary experimental infection. No extraintestinal stages were found in the MLNs of two calves examined 69 days after a primary infection or in five calves examined 6 months after a challenge infection. Tissue homogenates of liver, spleen, and MLNs from immune calves were not infectious for nonimmune recipient calves following oral or intraperitoneal inoculation indicating that infectious stages were not present in these extraintestinal tissues at the time of inoculations. Relapse of clinical coccidiosis and reoccurrence of oocyst shedding were not seen in E. bovis immune calves that were stressed by being put on a 16-week corticosteroid treatment program.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=22001980304-4017 Journal Article2200198SU.S. Department of Agriculture, Zoonotic Diseases Laboratory, Beltsville, MD 20705.~?%Andrews, C. D. Fayer, R. Dubey, J. P.19904Continuous in vitro cultivation of Sarcocystis cruzi254-5 J Parasitol7623Animals Cell Line Sarcocystis/*growth & developmentApr1Sporozoites of Sarcocystis cruzi were inoculated onto monolayer cultures of bovine pulmonary artery endothelial (CPA) cells. Sporozoites entered the cells, formed large and small multinucleate schizonts, and produced large numbers of merozoites. Continuous cultivation from the original sporozoite inoculum has been maintained for more than 1,320 days by subinoculating merozoites onto new cultures of CPA cells. During this time, the capacity to produce both types of schizonts was preserved, and schizogony was the only form of reproduction that was observed.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=21082360022-3395 Journal Article2108236?Zoonotic Diseases Laboratory, USDA, Beltsville, Maryland 20705.~?Fayer, R. Mayhew, I. G. Baird, J. D. Dill, S. G. Foreman, J. H. Fox, J. C. Higgins, R. J. Reed, S. M. Ruoff, W. W. Sweeney, R. W. et al.,1990uEpidemiology of equine protozoal myeloencephalitis in North America based on histologically confirmed cases. A report54-7J Vet Intern Med42Animals Encephalomyelitis/epidemiology/*veterinary Female Horse Diseases/*epidemiology Horses Male Ontario/epidemiology Prevalence Protozoan Infections/epidemiology *Protozoan Infections, Animal Retrospective Studies United States/epidemiologyMar-AprFollowing a workshop on equine protozoal myeloencephalitis (EPM) convened at the Veterinary Medical Forum of the American College of Veterinary Internal Medicine in 1988, this survey of EPM in North America was developed. It is based upon 364 histologically confirmed case records from California, Florida, Illinois, Kentucky, New York, Ohio, Oklahoma, Ontario, Pennsylvania, and Texas up to 1988. The highest rate of infection was found in young Thoroughbred, Standardbred, and quarter horses. Differences in geographic location, sex, and month (season) of infection were not discernible. This report, the first comprehensive survey of EPM in North America, is intended to serve as a basis for evaluating future changes in prevalence and spread of EPM.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=23420220891-6640 Journal Article2342022SZoonotic Diseases Laboratory, U.S. Department of Agriculture, Beltsville, MD 20705.~?/Ungar, B. L. Ward, D. J. Fayer, R. Quinn, C. A.1990Cessation of Cryptosporidium-associated diarrhea in an acquired immunodeficiency syndrome patient after treatment with hyperimmune bovine colostrum486-9Gastroenterology982Acquired Immunodeficiency Syndrome/*complications Adult Animals Cattle *Colostrum Cryptosporidiosis/complications/*therapy Diarrhea/*parasitology/therapy Human Immunotherapy/*methods Male Support, U.S. Gov't, Non-P.H.S.FebCryptosporidium is a parasite of the human gastrointestinal tract that can cause life-threatening diarrhea in immunodeficient patients. Although more than 80 agents have been tried with occasional anecdotal success, treatment remains primarily limited to hydration. A 38-yr-old homosexual man with antibody to human immunodeficiency virus and Cryptosporidium-related diarrhea is described. The patient excreted 6-12 L of stool per day for at least 3 mo, 2 of them spent in the hospital. Trials with more than 6 antidiarrheal medications were ineffective. The patient received bovine colostrum hyperimmune to Cryptosporidium by direct duodenal infusion. During infusion, the patient's fecal output decreased to less than 2 L per day, and 48 h after treatment, stools were formed and oocysts to Cryptosporidium were absent. The patient remained asymptomatic for 3 mo. Hyperimmune bovine colostrum offers an exciting new therapy for cryptosporidiosis; controlled trials to establish efficacy should be undertaken and the active factor(s) characterized.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=2295405&0016-5085 Case Reports Journal Article2295405mDivision of Tropical Public Health, Uniformed Services University of the Health Sciences, Bethesda, Maryland.~?3Perryman, L. E. Riggs, M. W. Mason, P. H. Fayer, R.1990Kinetics of Cryptosporidium parvum sporozoite neutralization by monoclonal antibodies, immune bovine serum, and immune bovine colostrum257-9 Infect Immun581%Animals Antibodies, Monoclonal/immunology Antibodies, Protozoan/*immunology Antigen-Antibody Reactions Cattle Coccidia/*immunology Colostrum/immunology Cryptosporidium/*immunology Kinetics Neutralization Tests Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S.JanMonoclonal antibodies, immune bovine serum, and immune bovine colostral whey neutralized infectivity of Cryptosporidium parvum sporozoites for mice in a time-dependent manner. Immune colostral whey neutralized sporozoites more rapidly and completely than immune serum, monoclonal antibody (MAb) 18.44, or a combination of MAb 18.44 and MAb 17.41. Mice were partially protected against oral challenge with C. parvum oocytes when treated with immune colostral whey, MAb 17.41, or a combination of MAb 17.41 and MAb 18.44.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=22940540019-9567 Journal Article2294054eDepartment of Veterinary Microbiology and Pathology, Washington State University, Pullman 99164-7040. ~?%Fayer, R. Trout, J. M. Jenkins, M. C.1998[Infectivity of Cryptosporidium parvum oocysts stored in water at environmental temperatures1165-9 J Parasitol846Amylopectin/analysis Animals Animals, Newborn Cattle Cryptosporidium parvum/chemistry/genetics/*physiology DNA, Protozoan/analysis Mice Mice, Inbred BALB C Polymerase Chain Reaction/veterinary Temperature Water/parasitologyDecMOocysts of Cryptosporidium parvum obtained from calves were cleaned of fecal debris by density gradient centrifugation and suspended in deionized water in microcentrifuge tubes. The tubes were placed in circulating water baths at temperatures of -10, -5, 0, 5, 10, 15, 20, 25, 30, or 35 C, and 2 tubes were removed from each water bath 1, 2, 4, 8, 12, 16, 20, and 24 wk later. Oocysts from 1 tube were administered at the rate of 1.5 x 10(5) oocysts per mouse to 2 litters of neonatal BALB/c mice and were considered infective when developmental stages were found in histologic sections of mouse gut and/or a positive polymerase chain reaction (PCR) was obtained for C. parvum DNA in mouse ileum. The second tube was held at -70 C until tubes from all time periods were available, then oocysts within the tubes were assayed for amylopectin concentration. Oocysts held at -10 C were infectious up to 1 wk of storage, and those held at -5 C were infectious up to 8 wk of storage, as determined by PCR but not histology. Oocysts held at 0, 5, 10, 15, and 20 C were still infectious after 24 wk of storage. By microscopic examination of mouse tissue, oocysts held at 20 C infected only 1 of 10 mice after 24 wk of storage, and the number of developmental stages began declining after 4 wk of storage; those held at 25 and 30 C each produced infections up to 12 wk after storage in 1 of 10 mice with reduced numbers of developmental stages beginning 4 wk after storage. Those held at 35 C produced light infections in 2 of 10 mice only up to 1 wk of storage. Amylopectin concentration decreased with increasing length of storage time or temperature. These findings provide a guide for estimating the potential duration of oocyst infectivity within a wide range of environmental temperatures and demonstrate the relationship between amylopectin concentration and infectivity.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=99203070022-3395 Journal Article9920307Immunology and Disease Resistance Laboratory, United States Department of Agriculture, Agricultural Research Service, Beltsville, Maryland 20705-2350, USA.~?*Xiao, L. Sulaiman, I. Fayer, R. Lal, A. A.1998eSpecies and strain-specific typing of Cryptosporidium parasites in clinical and environmental samples687-91Mem Inst Oswaldo Cruz935Animals Cryptosporidium/classification/*genetics/isolation & purification Genes, rRNA/genetics Genotype Human *Phylogeny Polymorphism (Genetics)/genetics RNA, Ribosomal/*analysis Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S. Water/*parasitologySep-OctCryptosporidiosis has recently attracted attention as an emerging waterborne and foodborne disease as well as an opportunistic infection in HIV infected individuals. The lack of genetic information, however, has resulted in confusion in the taxonomy of Cryptosporidium parasites and in the development of molecular tools for the identification and typing of oocysts in environmental samples. Phylogenetic analysis of the small subunit ribosomal RNA (SSU rRNA) gene has shown that the genus Cryptosporidium comprises several distinct species. Our data show the presence of at least four species: C. parvum, C. muris, C. baileyi and C. serpentis (C. meleagridis, C. nasorum and C. felis were not studied). Within each species, there is some sequence variation. Thus, various genotypes (genotype 1, genotype 2, guinea pig genotype, monkey genotype and koala genotype, etc.) of C. parvum differ from each other in six regions of the SSU rRNA gene. Information on polymorphism in Cryptosporidium parasites has been used in the development of species and strain-specific diagnostic tools. Use of these tools in the characterization of oocysts in various samples indicates that C. parvum genotype 1 is the strain responsible for most human Cryptosporidium infections. In contrast, genotype 2 is probably one of the major sources for environmental contamination, and has been found in most oysters examined from Chesapeake Bay that may serve as biologic monitors of estuarine waters.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=98305390074-0276 Journal Article9830539Division of Parasitic Diseases, Centers for Disease Control and Prevention, US Department of Health and Human Services, Atlanta, GA 30341, USA.~?@Graczyk, T. K. Farley, C. A. Fayer, R. Lewis, E. J. Trout, J. M.1998Detection of Cryptosporidium oocysts and Giardia cysts in the tissues of eastern oysters (Crassostrea virginica) carrying principal oyster infectious diseases1039-42 J Parasitol845Animals Cryptosporidiosis/transmission Cryptosporidium/*isolation & purification Fluorescent Antibody Technique Giardia/*isolation & purification Giardiasis/transmission Human Oysters/*parasitology Reagent Kits, Diagnostic Shellfish/*parasitology Support, Non-U.S. Gov'tOctThe potential cross-reactivity of the combined Cryptosporidium/Giardia direct immunofluorescence antibodies (IFA) of MERIFLUOR and HYDROFLUOR-COMBO tests was examined against tissues containing known developmental stages of 12 pathogens causing the principal infectious diseases in oysters. Spores of Haplosporidium nelsoni and Haplosporidium costale produced positive acid-fast stain (AFS) reactions similar in intensity to Cryptosporidium parvum oocysts. Hexamia nelsoni trophozoites produced positive IFA reactions in both IFA tests; however, the intensity of fluorescence was considerably lower and the fluorescein-staining pattern different than those of Giardia cysts. The applicability of AFS for screening oysters for Cryptosporidium oocysts is low, and positive identification of Cryptosporidium oocysts cannot be accomplished based on the AFS. Presumptive IFA identification of the Cryptosporidium oocysts or Giardia cysts in the oyster tissue should fulfill 3 criteria, i.e., bright-green fluorescence of the same intensity as C. parvum oocysts and Giardia cysts in the positive control, correct size and shape of the fluorescein-stained objects, and oocyst or cyst shell clearly visible.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=97946510022-3395 Journal Article9794651Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA.~?BBurkhalter, R. S. Smith, C. A. White, D. C. Fayer, R. White, A. B.1998The signature 10-hydroxy stearic acid thought to correlate with infectivity in oocysts of Cryptosporidium species is an artifact829-33Lipids338Animals *Artifacts Cryptosporidiosis/*metabolism/parasitology Cryptosporidium/chemistry/*pathogenicity Mice Mice, Inbred BALB C Stearic Acids/*chemistry Support, Non-U.S. Gov'tAugHeating or freezing leads to loss in infectivity of oocysts of Cryptosporidium parvum toward neonatal BALB/c mice and is reflected in the profile of the polar lipid fatty acids. Upon loss of infectivity, the ratio of polar lipid to neutral lipid fatty acid decreased and the relative proportions of 18:1n-9 also decreased; proportions of 18:2n-6 and 20:5n-6 increased, whereas the proportions of 16:0 remained constant with freezing. During these investigations, a novel fatty acid, 10-OH 18:0, was discovered in the glycolipid fraction. The identification of a fatty acid unique to species of Cryptosporidium was thought to provide a specific biomarker for this organism. Cryptosporidium also demonstrated fluctuations in absolute quantities of 10-OH 18:0 with events that lead to loss of infectivity. This led to the presumed correlation of this biomarker with infectious Cryptosporidium. The 10-OH 18:0 was putatively localized at the sn-2 position of phosphatidylethanolamine. High-performance liquid chromatography/electrospray ionization mass spectrometry revealed that the 10-OH 18:0 existed principally in the free fatty acid form. Herein, we establish that the free fatty acid 10-OH 18:0 was, in actuality, an artifact of the procedures for sample preparation.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=97276160024-4201 Journal Article9727616[Center for Environmental Biotechnology, University of Tennessee, Knoxville 37932-2575, USA.T~?RElsasser, T. H. Sartin, J. L. McMahon, C. Romo, G. Fayer, R. Kahl, S. Blagburn, B.1998Changes in somatotropic axis response and body composition during growth hormone administration in progressive cachectic parasitism239-55Domest Anim Endocrinol154Acute-Phase Reaction/blood/physiopathology/veterinary Animals Blood Urea Nitrogen Body Composition/*drug effects/physiology Body Temperature/drug effects Cachexia/etiology/prevention & control/*veterinary Cattle Cattle Diseases/etiology/*physiopathology/prevention & control Growth Hormone/blood/pharmacology/*therapeutic use Insulin-Like Growth Factor I/analysis/genetics Male Microsomes, Liver/metabolism Muscle Proteins/drug effects/metabolism RNA, Messenger/analysis Receptors, Somatotropin/analysis/genetics/metabolism Sarcocystosis/complications/physiopathology/*veterinary Somatotropin-Releasing Hormone/pharmacology Tumor Necrosis Factor/analysis Weight Gain/drug effectsJul A multistage protozoan parasitic disease was used as a cachexia model to study the effects of daily administration of bovine growth hormone (GH) on endocrine and body composition changes of young calves from the onset of the acute phase response (APR). Male calves averaging 127.5 +/- 2.0 kg body weight were assigned to control, ad libitum fed, noninfected (C); ad libitum fed, infected (250,000 oocysts Sarcocystis cruzi, per os, I); noninfected, pair-fed (PF) to matched I-treatment calves and these respective same treatments in calves injected daily with GH (USDA-bGH-B1), 12.5 mg/calf/day, im) designated as CGH, IGH and PFGH. GH injections were initiated on Day 20 postinfection (PI), 3 to 4 d before the onset of clinical signs of APR, and continued to Day 56 PI, at which time animals were euthanized for tissue collections. Abrupt increases in rectal temperature commensurate with up to 70% reduction in voluntary feed intake were observed in I and IGH beginning 23-25 d PI. For the trial period between Days 20 and 56 PI, average daily carcass protein gains were 123, 52, 109, 124, 48, and 67 g/d and average daily carcass fat gains were 85, 11, 43, 71, -23, and 29 g/d for C, I, PF, CGH, IGH, and PFGH, respectively. Effects of GH were significant for fat accretion and plasma urea depression. Rectus femoris was highly refractory to catabolic effects of infection while psoas major was significantly catabolized during infection. Plasma concentrations of insulin-like growth factor-I (IGF-I) increased significantly in all GH-treated calves between Day 20 and 23 PI. Plasma IGF-I declined well below Day 20 values in all infected calves from the onset of the APR through the end of the study. The decrease in plasma IGF-I concentrations in I and IG was highly correlated with the magnitude of the fever response. Hepatic mRNA for GH receptor and IGF-I was decreased in infected calves. Hepatic microsomal membrane binding of 125I-GH did not differ between groups. The data suggest that effects of GH and parasitism on tissue metabolism during disease may vary among different specific tissue pools. The data demonstrate that daily GH administration in young calves does not prevent lean tissue losses and may accelerate fat depletion associated with cachectic parasitism. Furthermore, the onset of APR overrode the capacity for GH to maintain elevated plasma concentrations of IGF-I, an effect not readily explained through changes of GH-receptor binding.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=96734560739-7240 Journal Article9673456UUS Department of Agriculture, Agriculture Research Service, Beltsville, MD 20705 USA.~?)Graczyk, T. K. Cranfield, M. R. Fayer, R.1998Oocysts of Cryptosporidium from snakes are not infectious to ducklings but retain viability after intestinal passage through a refractory host33-40 Vet Parasitol771Animals Bird Diseases/*parasitology/transmission Cryptosporidiosis/*parasitology/transmission Cryptosporidium/growth & development/*pathogenicity Ducks/*parasitology Feces/parasitology Snakes/*parasitology Support, Non-U.S. Gov'tMaySix 2-week-old Cryptosporidium-free Peking ducklings (Anas platyrhynchos) each received 2.0 x 10(6) viable Cryptosporidium serpentis oocysts from 6 naturally infected captive snakes. Histological sections of digestive (stomach, jejunum, ileum, cloaca, and cecum) and respiratory tract tissues (larynx, trachea, and lungs) did not contain life-cycle stages of Cryptosporidium in any of the inoculated ducklings. Because ducklings were refractory to infection, C. serpentis transmission via a diet of Peking ducklings is improbable. Viable (per in vitro excystation assay) inoculum-derived oocysts were detected in duckling feces up to 7 days post-inoculation (PI); the number of intact oocysts excreted during the first 2 days PI was significantly higher than for the remaining 5 days PI (P < 0.01). The dynamics of oocyst shedding showed that overall the birds released a significantly higher number of intact oocysts than oocyst shells (P < 0.01). Retention of the viability of C. serpentis oocysts following intestinal passage through a refractory avian species may have epizootiological implications. Under certain circumstances such as after the ingestion of C. serpentis-infected prey, herpetivorous birds may disseminate C. serpentis oocysts in the environment.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=96523810304-4017 Journal Article9652381Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, MD 21205, USA. tgraczyk@jhsph.edu~?XGraczyk, T. K. Fayer, R. Trout, J. M. Lewis, E. J. Farley, C. A. Sulaiman, I. Lal, A. A.1998zGiardia sp. cysts and infectious Cryptosporidium parvum oocysts in the feces of migratory Canada geese (Branta canadensis)2736-8Appl Environ Microbiol647Animals Cryptosporidium parvum/*isolation & purification Feces/*parasitology Geese/*parasitology Giardia/*isolation & purification Mice Support, Non-U.S. Gov'tJul Fecal droppings of migratory Canada geese, Branta canadensis, collected from nine sites near the Chesapeake Bay (Maryland), were examined for the presence of Cryptosporidium parvum and Giardia spp. Cryptosporidium sp. oocysts were found in feces at seven of nine sites, and Giardia cysts were found at all nine sites. The oocysts from three sites were infectious for mice and molecularly identified as the zoonotic genotype of Cryptosporidium parvum. Waterfowl can disseminate infectious C. parvum oocysts in the environment.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=96478600099-2240 Journal Article9647860Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA. tgraczyk@jhsph.edu ~?DGraczyk, T. K. Cranfield, M. R. Helmer, P. Fayer, R. Bostwick, E. F.1998Therapeutic efficacy of hyperimmune bovine colostrum treatment against clinical and subclinical Cryptosporidium serpentis infections in captive snakes123-32 Vet Parasitol742-4Animals Animals, Zoo/parasitology Cattle Colostrum/*immunology Cryptosporidiosis/immunology/therapy/*veterinary Cryptosporidium/*immunology Cryptosporidium parvum/immunology Feces/chemistry Female Fluorescent Antibody Technique, Indirect/veterinary Gastric Lavage/veterinary Immunization/veterinary Immunization, Passive/*veterinary Intestines/chemistry Random Allocation Snakes/*parasitology Staining and Labeling/veterinary Stomach/chemistry Support, Non-U.S. Gov'tJan 31Therapy based on the protective passive immunity of Hyperimmune Bovine Colostrum (HBC) (raised against Cryptosporidium parvum in dairy cows immunized during gestation) was tested for heterologous efficacy in subclinical and clinical infections of 12 captive snakes with C. serpentis. Six gastric HBC treatments of 1% snake weight at 1-week intervals each, have histologically cleared C. serpentis in three subclinically infected snakes, and regressed gastric histopathological changes in one of these snakes. In all snakes, each subsequent HBC treatment significantly decreased the number of oocysts recovered in gastric lavage eluants (P < 0.03). The treatments induced oocyst-negative gastric eluants and stools in all snakes, and improved clinical signs of infection. Clinically infected snakes displayed severe histopathological changes in the gastric region; however, the numbers of developmental stages of C. serpentis were moderate. Considering the severity of pathology, much lower than expected pathogen numbers were observed, and it is believed that clinically infected snakes did not have enough time to repair tissue damage that had occurred over the years of infection. As the HBC treatment was safe and highly efficacious, it is recommended to gastrically administer the HBC therapeutically to snakes that are clinically or subclinically infected with C. serpentis. Hyperimmune bovine colostrum can also be used in snake supportive therapy or prophylaxis.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=95617000304-4017 Journal Article9561700Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, MD 21205, USA. tgraczyk@phnet.sph.jhu.edu~?COlson, E. J. Epperson, W. B. Zeman, D. H. Fayer, R. Hildreth, M. B.1998KEffects of an allicin-based product on cryptosporidiosis in neonatal calves987-90J Am Vet Med Assoc2127Animals Animals, Newborn Anti-Infective Agents/*therapeutic use Cattle Cattle Diseases/*drug therapy Cryptosporidiosis/drug therapy/*veterinary *Cryptosporidium parvum Diarrhea/drug therapy/veterinary Male Sulfinic Acids/*therapeutic use Support, Non-U.S. Gov'tApr 1OBJECTIVE: To evaluate effectiveness of an allicin-based product in neonatal calves inoculated with Cryptosporidium parvum. DESIGN: Randomized controlled study. ANIMALS: 43 neonatal calves. PROCEDURE: Calves were inoculated with 1.5 x 10(8) or 7.5 x 10(5) C parvum oocysts within 2 days after birth. Calves were given an allicin-based product once after inoculation or daily for 7 days after inoculation or were not treated. Calves that developed diarrhea were treated by administration of the product. Fecal consistency scores and weight gains were statistically evaluated. RESULTS: Mean daily weight gain and severity of diarrhea in calves 4 to 21 days old were unaffected by prophylactic use of the product. However, intensive prophylactic administration may have delayed onset of C parvum-induced diarrhea in calves inoculated with the lower dose of oocysts. CLINICAL IMPLICATIONS: Administration of an allicin-based product did not alter duration of C parvum-induced diarrhea or enhance weight gain in neonatal calves. However, intensive prophylactic administration of an allicin-based product may delay onset of diarrhea in calves exposed to C parvum oocysts.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=9540869D0003-1488 Clinical Trial Journal Article Randomized Controlled Trial9540869Department of Biology and Microbiology, College of Agriculture and Biological Sciences, South Dakota State University, Brookings 57007, USA. (~?@Fayer, R. Graczyk, T. K. Lewis, E. J. Trout, J. M. Farley, C. A.1998Survival of infectious Cryptosporidium parvum oocysts in seawater and eastern oysters (Crassostrea virginica) in the Chesapeake Bay1070-4Appl Environ Microbiol643Animals Cattle Cryptosporidium parvum/*growth & development/isolation & purification Giardia/isolation & purification Mice Mice, Inbred BALB C Oysters/*parasitology Seawater/*parasitologyMar!Oocysts of Cryptosporidium parvum placed in artificial seawater at salinities of 10, 20, and 30 ppt at 10 degrees C and at 10 ppt at 20 degrees C were infectious after 12 weeks. Those placed in seawater at 20 ppt and 30 ppt at 20 degrees C were infectious for 8 and 4 weeks, respectively. These findings suggested that oocysts could survive in estuarine waters long enough to be removed by filter feeders such as oysters. Thereafter, 30 Eastern oysters, Crassostrea virginica, were collected with a dredge or with hand tongs at each of six sites within Maryland tributaries of the Chesapeake Bay in May and June and in August and September of 1997. Hemocytes and gill washings from all oysters were examined for the presence of Cryptosporidium oocysts and Giardia cysts by immunofluorescence microscopy utilizing a commercially available kit containing fluorescein isothiocyanate-conjugated monoclonal antibodies. Giardia was not detected by this method from any of the 360 oysters examined. Presumptive identification of Cryptosporidium oocysts was made in either hemocytes or gill washings of oysters from all six sites both times that surveys were conducted. In addition, during August and September, for each of the six sites, hemocytes from the 30 oysters were pooled and gill washings from the oysters were pooled. Each pool was delivered by gastric intubation to a litter of neonatal mice to produce a bioassay for oocyst infectivity. Intestinal tissue from two of three mice that received gill washings from oysters collected at a site near a large cattle farm and shoreline homes with septic tanks was positive for developmental stages of C. parvum. These findings demonstrate for the first time that oysters in natural waters harbor infectious C. parvum oocysts and can serve as mechanical vectors of this pathogen.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=95014460099-2240 Journal Article9501446Immunology and Disease Resistance Laboratory, Agricultural Research Service, U.S. Department of Agriculture, Beltsville, Maryland 20705-2350, USA. rfayer@ggpl.arsusda.gov~?"Jenkins, M. C. Trout, J. Fayer, R.1998Development and application of an improved semiquantitative technique for detecting low-level Cryptosporidium parvum infections in mouse tissue using polymerase chain reaction182-6 J Parasitol841 Animals Animals, Newborn Cryptosporidiosis/*diagnosis/parasitology Cryptosporidium parvum/genetics/*isolation & purification DNA, Protozoan/*analysis Densitometry Disease Models, Animal Mice Mice, Inbred BALB C *Polymerase Chain Reaction Sensitivity and SpecificityFebAn improved semiquantitative technique was developed for measuring low infectious doses of Cryptosporidium parvum in neonatal mice using polymerase chain reaction (PCR). Separate litters of neonatal mice were inoculated with 0, 10(2), 10(3), or 10(4) C. parvum oocysts and killed 96 hr postinfection. A segment of the ileum or the entire whole intestine was then removed from subgroups of mice in each litter and total DNA was extracted using standard procedures. By employing a CP15/60-based semiquantitative PCR technique, C. parvum DNA was detected in mice infected with as few as 10(2) oocysts. DNA isolated from the ileum of infected mice produced a more intense PCR signal than DNA isolated from the whole intestine. This technique was used to study the intracellular development of C. parvum sporozoites that had been exposed in the oocyst stage to either 0, 15, 20, 25, or 30 kRad gamma-irradiation. A CP15/60 PCR signal was observed in ileum tissue from mice infected with 0-kRad- or 15-kRad-irradiated C. parvum oocysts. A very slight PCR signal was generated by PCR on ileum tissue DNA from mice infected with 20-kRad-irradiated oocysts, whereas no signal was observed in PCR on intestinal DNA from mice infected with oocysts exposed to higher radiation doses.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=94883640022-3395 Journal Article9488364sImmunology and Disease Resistance Laboratory, Agricultural Research Service, USDA, Beltsville, Maryland 20705, USA.(~?MCanals, A. Pasquali, P. Zarlenga, D. S. Fayer, R. Almeria, S. Gasbarre, L. C.1998_Local ileal cytokine responses in cattle during a primary infection with Cryptosporidium parvum125-30 J Parasitol8411Animals Animals, Newborn Cattle Cattle Diseases/*immunology Cryptosporidiosis/immunology/*veterinary Cryptosporidium parvum/genetics/*immunology Cytokines/*biosynthesis/genetics Ileum/*immunology Lymph Nodes/cytology/immunology Lymphocytes/immunology Male Polymerase Chain Reaction RNA, Messenger/analysisFebIn the present study, localized changes in cytokine transcription profiles were examined in neonatal calves following a primary infection with Cryptosporidium parvum, using competitive reverse transcriptase polymerase chain reaction (RT-PCR). Total RNA was prepared from ileocecal lymph nodes (LN), lamina propria lymphocytes (LPL), and intraepithelial lymphocytes (IEL) isolated from neonatal calves 7 days after C. parvum infection. Competitive RT-PCR performed on cDNA samples containing internal cytokine gene competitor molecules showed increases in the levels of interferon-gamma and interleukin-12 (IL-12) (P40) mRNA in both LPL and IEL populations but not in the draining LN. In addition, the levels of mRNA of the newly identified growth factor IL-15 decreased in the IEL of the infected animals. No consistent differences were seen in any of the cell populations when the samples were analyzed for IL-10 and levels of mRNA for IL-2 and IL-4 were low and highly variable in both infected and control groups in all 3 lymphocyte populations.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=94883490022-3395 Journal Article9488349_Immunology and Disease Resistance Laboratory, LPSI, ARS, USDA, Beltsville, Maryland 20705, USA.~?5Graczyk, T. K. Fayer, R. Cranfield, M. R. Conn, D. B.1998fRecovery of waterborne Cryptosporidium parvum oocysts by freshwater benthic clams (Corbicula fluminea)427-30Appl Environ Microbiol642Animals Clams/*parasitology Cryptosporidium parvum/*isolation & purification Mice Mice, Inbred BALB C Support, Non-U.S. Gov't Water/*parasitologyFeb4Asian freshwater clams, Corbicula fluminea, exposed for 24 h to 38 liters of water contaminated with infectious Cryptosporidium parvum oocysts (1.00 x 10(6) oocysts/liter; approximately 1.9 x 10(5) oocysts/clam) were examined (hemolymph, gills, gastrointestinal [GI] tract, and feces) on days 1, 2, 3, 7, and 14 postexposure (PE). No oocysts were detected in the water 24 h after the contamination event. The percentage of oocyst-containing clams varied from 20 to 100%, depending on the type of tissue examined and the technique used--acid-fast stain (AFS) or immunofluorescent antibody (IFA). The oocysts were found in clam tissues and feces on days 1 through 14 PE; the oocysts extracted from the tissues on day 7 PE were infectious for neonatal BALB/c mice. Overall, the highest number of positive samples was obtained when gills and GI tracts were processed with IFA (prevalence, 97.5%). A comparison of the relative oocyst numbers indicated that overall, 58.3% of the oocysts were found in clam tissues and 41.7% were found in feces when IFA was used; when AFS was used, the values were 51.9 and 48.1%, respectively. Clam-released oocysts were always surrounded by feces; no free oocysts or oocysts disassociated from fecal matter were observed. The results indicate that these benthic freshwater clams are capable of recovery and sedimentation of waterborne C. parvum oocysts. To optimize the detection of C. parvum oocysts in C. fluminea tissue, it is recommended that gill and GI tract samples be screened with IFA (such as that in the commercially available MERIFLUOR test kit).dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=94643760099-2240 Journal Article9464376Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA. tgraczyk@jhsph.eduh~?AMorgan, U. M. Monis, P. T. Fayer, R. Deplazes, P. Thompson, R. C.1999^Phylogenetic relationships among isolates of Cryptosporidium: evidence for several new species1126-33 J Parasitol856^Animals Cats Cattle Cryptosporidium/*classification/genetics/isolation & purification Genotype Human Marsupialia Mice Molecular Sequence Data Parasite Egg Count Phylogeny Protozoan Proteins/chemistry/genetics RNA, Ribosomal, 18S/genetics Random Amplified Polymorphic DNA Technique Support, Non-U.S. Gov't Swine Tetrahydrofolate Dehydrogenase/geneticsDecIsolates of Cryptosporidium were characterized using nucleotide sequence analysis of the 18S rRNA and dihydrofolate reductase genes and also random-amplified polymorphic DNA analysis. Phylogenetic analysis confirmed the validity of the species of Cryptosporidium examined in this study such as Cryptospordium muris and Cryptosporidium baileyi, and also reinforced evidence from numerous researchers worldwide suggesting that Cryptosporidium parvum is not a single uniform species. The data obtained provided strong support for the validity of Cryptosporidium felis. Evidence suggests that the newly identified marsupial and pig genotypes may also be distinct and valid species, but biological studies are required for confirmation.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=106470470022-3395 Journal Article10647047World Health Organisation Collaborating Centre for the Molecular Epidemiology of Parasitic Infections, Division of Veterinary and Biomedical Sciences, Murdoch University, WA, Australia. !~?PJenkins, M. C. Trout, J. Murphy, C. Harp, J. A. Higgins, J. Wergin, W. Fayer, R.1999lCloning and expression of a DNA sequence encoding a 41-kilodalton Cryptosporidium parvum oocyst wall protein912-20Clin Diagn Lab Immunol66HAmino Acid Sequence Animals Antibodies, Protozoan Antigens, Protozoan/*genetics/immunology Base Sequence Blotting, Northern Cattle Cell Wall/chemistry/ultrastructure Cloning, Molecular Cryptosporidiosis/*diagnosis Cryptosporidium parvum/*genetics/growth & development/immunology DNA Primers DNA, Protozoan/analysis Enzyme-Linked Immunosorbent Assay Gene Expression Human Microscopy, Immunoelectron Molecular Sequence Data Protozoan Proteins/*genetics/immunology RNA, Messenger/analysis Rabbits Reverse Transcriptase Polymerase Chain Reaction Sequence Analysis, DNA Species SpecificityNovkThis study was conducted to produce a recombinant species-specific oocyst wall protein of Cryptosporidium parvum. Antigens unique to C. parvum were identified by gradient sodium dodecyl sulfate-polyacrylamide gel electrophoresis and immunoblotting of oocyst proteins from several different Cryptosporidium species. Antiserum was then prepared against a 41-kDa antigen unique to C. parvum and used to identify a recombinant DNA clone, designated rCP41. Expression of CP41 mRNA in C. parvum oocysts was confirmed by reverse transcriptase PCR (RT-PCR). Although the CP41 sequence was shown by PCR to be present in the genome of C. baileyi, CP41 mRNA was not detected in this species by RT-PCR. Immunofluorescence staining with antiserum against recombinant CP41 detected native CP41 antigen on the surface of C. parvum oocysts but failed to detect CP41 on C. baileyi oocysts. Immunoelectron microscopy demonstrated that native CP41 was distributed unevenly on the C. parvum oocyst surface and was associated with amorphous oocyst wall material. In an enzyme-linked immunosorbent assay, purified rCP41 performed as well as native C. parvum oocyst protein in measuring the serological responses of young calves and adult cows to experimental and natural C. parvum infections. These results indicate that recombinant CP41 antigen may have potential in the immunodiagnosis of cryptosporidiosis.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=105485851071-412x Journal Article10548585Immunology and Disease Resistance Laboratory, Beltsville Agricultural Research Center, Agricultural Research Service, U.S. Department of Agriculture, Beltsville, Maryland 20705, USA. mjenkins@lpsi.barc.usda.gov~?MGraczyk, T. K. Thompson, R. C. Fayer, R. Adams, P. Morgan, U. M. Lewis, E. J.1999iGiardia duodenalis cysts of genotype A recovered from clams in the Chesapeake Bay subestuary, Rhode River526-9Am J Trop Med Hyg614kAnimals Clams/*parasitology DNA Primers/chemistry DNA, Protozoan/chemistry Fluorescent Antibody Technique, Indirect/veterinary Genotype Giardia lamblia/genetics/immunology/*isolation & purification Giardiasis/*transmission Hemolymph/parasitology Maryland Polymerase Chain Reaction/veterinary Seawater Sequence Analysis, DNA Support, Non-U.S. Gov't Water PollutionOctHFilter-feeding molluscan shellfish can concentrate zoonotic and anthroponotic waterborne pathogens. Cysts of Giardia sp. were detected by immunofluorescent antibodies in tissues of the clams Macoma balthica and M. mitchelli from Rhode River, a Chesapeake Bay (Maryland) subestuary. Molecular tests identified the cysts as Giardia duodenalis Genotype A, the most common genotype recovered from humans. Macoma clams are burrowers in mud or sandy-mud substrata and preferentially feed on the surface sediment layer. Waterborne Giardia cysts settle rapidly to the bottom in slow-moving waters and contaminate the sediment. Macoma clams do not have economic value, but can serve as biologic indicators of sediment contamination with Giardia sp. cysts of public health importance. These clams can be used for sanitary assessment of water quality.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=105482840002-9637 Journal Article10548284Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA.~?aGraczyk, T. K. Fayer, R. Cranfield, M. R. Mhangami-Ruwende, B. Knight, R. Trout, J. M. Bixler, H.19999Filth flies are transport hosts of Cryptosporidium parvum726-7Emerg Infect Dis55Animals *Arthropod Vectors Cattle Cryptosporidiosis/*transmission Cryptosporidium parvum/*isolation & purification Feces/*parasitology Houseflies/*parasitology Human MaleSep-Octehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=105115341080-6040 Letter10511534~?aFayer, R. Lewis, E. J. Trout, J. M. Graczyk, T. K. Jenkins, M. C. Higgins, J. Xiao, L. Lal, A. A.1999XCryptosporidium parvum in oysters from commercial harvesting sites in the Chesapeake Bay706-10Emerg Infect Dis55.Animals Base Sequence Cryptosporidium parvum/genetics/*isolation & purification/pathogenicity Food-Processing Industry Genotype Human Maryland Mice Molecular Sequence Data Oysters/*parasitology Polymerase Chain Reaction Polymorphism, Restriction Fragment Length Support, Non-U.S. Gov't *Water PollutionSep-OctsOocysts of Cryptosporidium parvum, a zoonotic waterborne pathogen, can be removed by bivalve molluscs from contaminated water and retained on gills and in hemolymph. We identified oocysts of C. parvum in oysters from seven sites in the Chesapeake Bay area. These findings document the presence of C. parvum infectious for humans in oysters intended for human consumption.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=105115281080-6040 Journal Article10511528ZU.S. Department of Agriculture, Beltsville, Maryland 20705, USA. rfayer@lpsi.barc.usda.gov~?4Graczyk, T. K. Cranfield, M. R. Fayer, R. Bixler, H.1999JHouse flies (Musca domestica) as transport hosts of Cryptosporidium parvum500-4Am J Trop Med Hyg613FAnimals Cattle Cattle Diseases/parasitology/*transmission Cryptosporidiosis/parasitology/transmission/*veterinary Cryptosporidium parvum/growth & development/*isolation & purification Feces/*parasitology Host-Parasite Relations Houseflies/growth & development/*parasitology Insect Vectors/*parasitology Support, Non-U.S. Gov'tSepRefuse and promiscuous-landing synanthropic filth flies, such as house flies (Musca domestica), are recognized as transport hosts for a variety of protozoan and metazoan parasites in addition to viral and bacterial pathogens of public health importance. Exposure of adult M. domestica to 20 ml of bovine diarrheal feces containing Cryptosporidium parvum oocysts (2.0 x 10(5) oocysts/ml) resulted in intense deposition of the oocysts through fly feces on the surfaces visited by the flies (mean = 108 oocysts/cm2). Cryptosporidium parvum oocysts were detected by immunofluorescent antibodies on the exoskeleton of adult flies and in their digestive tracts. An average of 267, 131, 32, 19, and 14 oocysts per adult fly were eluted from its exoskeleton on days 3, 5, 7, 9, and 11 after they emerged, respectively. Approximately 320 C. parvum oocysts per pupa were eluted from the external surface of the pupae derived from maggots that breed in a substrate contaminated with the bovine feces; the oocysts were numerous on maggots (approximately 150 oocysts/maggot). Adult and larval stages of house flies breeding or having access to C. parvum-contaminated substrate will mechanically carry the oocysts in their digestive tracts and on their external surfaces.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=104979980002-9637 Journal Article10497998hDepartment of Molecular Microbiology and Immunology, Johns Hopkins University, Baltimore, Maryland, USA. ~?.Carpenter, C. Fayer, R. Trout, J. Beach, M. J.1999FChlorine disinfection of recreational water for Cryptosporidium parvum579-84Emerg Infect Dis54Animals Antiprotozoal Agents/*pharmacology Cattle Chlorine/*pharmacology Cryptosporidium parvum/*drug effects/growth & development Disinfectants/*pharmacology Disinfection Mice Mice, Inbred BALB C Recreation Temperature Time Factors Water/*parasitologyJul-Aug~We examined the effects of chlorine on oocyst viability, under the conditions of controlled pH and elevated calcium concentrations required for most community swimming pools. We found that fecal material may alter the Ct values (chlorine concentration in mg/L, multiplied by time in minutes) needed to disinfect swimming pools or other recreational water for Cryptosporidium parvum.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=104589691080-6040 Journal Article10458969@U.S. Department of Agriculture, Beltsville, Maryland 20705, USA. ~?9Jenkins, M. C. O'Brien, C. Trout, J. Guidry, A. Fayer, R.1999Hyperimmune bovine colostrum specific for recombinant Cryptosporidium parvum antigen confers partial protection against cryptosporidiosis in immunosuppressed adult mice2453-60Vaccine1719Animals Antibodies, Protozoan/immunology Antigens, Protozoan/*immunology Cattle Colostrum/*chemistry/*immunology Cryptosporidiosis/immunology/*prevention & control Cryptosporidium parvum/*immunology/isolation & purification DNA, Protozoan/analysis/genetics Dexamethasone/metabolism Female Immunization, Passive Immunocompromised Host/immunology Mice Mice, Inbred C57BL Plasmids/genetics Polymerase Chain Reaction Pregnancy Protozoan Proteins/biosynthesis/genetics Recombinant Proteins/immunologyMay 14UPreparturient cows were immunized three times over a six-week period with recombinant plasmid DNA encoding the Cryptosporidium parvum CP15/60 antigen by injecting the DNA in the mammary gland. Serum was collected at each immunization and first colostrum was collected after parturition; all were assayed for Cryptosporidium-specific antibodies (Ab). A serological response to C. parvum sporozoite and oocyst antigen was detected in cows immunized with pCP15/60 plasmid DNA. Colostrum from these cows, unlike colostrum from normal controls, contained Ab specific for C. parvum sporozoites and oocysts as indicated by immunofluorescence Ab (IFA) staining. Colostrum was also tested for conferring passive immunity against C. parvum infection by oral administration to immunosuppressed adult inbred mice. Immune colostrum and control colostrum were administered to separate groups of dexamethasone (DEX)-treated adult C57BL/6NCr mice beginning 12 h before and at 12 h intervals for 3 days after oral C. parvum oocyst infection. Cryptosporidium development was assayed in ilea of immune- and control-colostrum-treated mice 96 h postinfection by semiquantitative PCR. Mice receiving immune colostrum showed partial protection (about 50% reduction) against intestinal C. parvum development compared to mice receiving control colostrum. This protection was evident at a challenge dose of 10(3) C. parvum oocysts per mouse; no differences were noted in parasite development between groups receiving immune or control colostrum and infected with 10(4) oocysts. This study showed that serum and colostrum Ab response to C. parvum can be elicited in preparturient cows by direct injection of recombinant pCP15/60 plasmid DNA and that passive protection against cryptosporidiosis can be obtained by treating immunosuppressed mice with immune colostrum before and after C. parvum infection.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=103926280264-410x Journal Article10392628Immunology and Disease Resistance Laboratory, Agricultural Research Service, USDA, Beltsville, MD 20705, USA. mjenkins@lpsi.barc.usda.gov~?WMorgan, U. M. Xiao, L. Fayer, R. Graczyk, T. K. Lal, A. A. Deplazes, P. Thompson, R. C.1999Phylogenetic analysis of Cryptosporidium isolates from captive reptiles using 18S rDNA sequence data and random amplified polymorphic DNA analysis525-30 J Parasitol853Animals Base Sequence Cattle Cryptosporidiosis/parasitology/*veterinary Cryptosporidium/*classification/genetics DNA, Protozoan/chemistry DNA, Ribosomal/*chemistry Human Lizards/*parasitology Mice Molecular Sequence Data *Phylogeny RNA, Protozoan/genetics RNA, Ribosomal, 18S/*genetics Random Amplified Polymorphic DNA Technique/veterinary Sequence Alignment/veterinary Snakes/*parasitology Support, Non-U.S. Gov'tJunSequence alignment of a polymerase chain reaction-amplified 713-base pair region of the Cryptosporidium 18S rDNA gene was carried out on 15 captive reptile isolates from different geographic locations and compared to both Cryptosporidium parvum and Cryptosporidium muris isolates. Random amplified polymorphic DNA (RAPD) analysis was also performed on a smaller number of these samples. The data generated by both techniques were significantly correlated (P < 0.002), providing additional evidence to support the clonal population structure hypothesis for Cryptosporidium. Phylogenetic analysis of both 18S sequence information and RAPD analysis grouped the majority of reptile isolates together into 1 main group attributed to Cryptosporidium serpentis, which was genetically distinct but closely related to C. muris. A second genotype exhibited by 1 reptile isolate (S6) appeared to be intermediate between C. serpentis and C. muris but grouped most closely with C. muris, as it exhibited 99.15% similarity with C. muris and only 97.13% similarity with C. serpentis. The third genotype identified in 2 reptile isolates was a previously characterized 'mouse' genotype that grouped closely with bovine and human C. parvum isolates.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=103864470022-3395 Journal Article10386447World Health Organisation Collaborating Centre for the Molecular Epidemiology of Parasitic Infections and State Agricultural Biotechnology Centre, Murdoch University, Western Australia, Australia.~? @Graczyk, T. K. Fayer, R. Lewis, E. J. Trout, J. M. Farley, C. A.1999RCryptosporidium oocysts in Bent mussels (Ischadium recurvum) in the Chesapeake Bay518-21 Parasitol Res857Animals Cryptosporidium/*growth & development/*isolation & purification Environmental Monitoring Maryland Mussels/*parasitology Parasite Egg Count Sensitivity and Specificity Support, Non-U.S. Gov't Water/parasitologyJulFilter-feeding molluscan shellfish can concentrate environmentally derived waterborne pathogens of humans, which can be utilized in the sanitary assessment of water quality. In the present study, oocysts of Cryptosporidium were detected in Bent mussels (Ischadium recurvum) at two Chesapeake Bay sites from which C. parvum-contaminated oysters had previously been collected. Spiking of Cryptosporidium-free blue mussel (Mytilus edulis) tissue with C. parvum oocysts showed a 51.1% recovery rate of oocysts, giving an oocyst detection limit of 19 oocysts/0.7 ml of mussel tissue homogenate. The results indicate that Bent mussels, which are common throughout the Chesapeake Bay region, may prove to be useful as biological indicators of water contamination with Cryptosporidium oocysts.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=103826000932-0113 Journal Article10382600Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, MD 21205, USA. tgraczyk@jhsph.edu~? 1Graczyk, T. K. Fayer, R. Conn, D. B. Lewis, E. J.1999lEvaluation of the recovery of waterborne Giardia cysts by freshwater clams and cyst detection in clam tissue30-4 Parasitol Res851Analysis of Variance Animals Clams/*parasitology Fresh Water Gerbillinae Giardia/*isolation & purification Giardiasis/parasitology/veterinary Human Regression Analysis Sensitivity and Specificity Support, Non-U.S. Gov'tJanThe Asian freshwater clam, Corbicula fluminea, inhabits environments recognized to be contaminated with waterborne Giardia cysts. Sixty-four tissue samples of Giardia-free clams were spiked with various numbers of Giardia duodenalis cysts within the range of 50-700 cysts. Regression analysis showed that paired numbers of spiked (x) versus recovered (y) cysts regressed significantly (P < 0.01) according to the equation y = 42.57 +/- 1.81x (+/- 64.3). The cyst detection threshold was 43 cysts/clam, the coefficient of determination was 77%, and the overall sensitivity of cyst detection was 42.9%. All 20 values of cyst numbers in clam tissue samples that were processed blind were located within the 95% prediction limits of the linear regression equation. The cyst retention rate of 160 clams kept in an aquarium with 38 l of water spiked with 1.00 x 10(5) G. duodenalis cysts was approximately 1.3 x 10(3) cysts/clam. No waterborne cysts were detected by the membrane filtration method 90 min after spiking the aquarium water. G. duodenalis cysts were detected in clam tissue up to 3 weeks post-exposure. Filtration of water by clams substantially depleted the aquarium water of its particulate matter. The sampling program demonstrated that the population of 160 clams examined during the study could be accurately assessed for exposure to waterborne Giardia cysts by random sampling of 86 (54%) clams. The results indicate that C. fluminea clams can be used for biological monitoring of contamination with Giardia.dhttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=99502250932-0113 Journal Article9950225Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, MD 21205, USA. tgraczyk@jhsph.eduZ~? fGraczyk, T. K. Fayer, R. Knight, R. Mhangami-Ruwende, B. Trout, J. M. Da Silva, A. J. Pieniazek, N. J.2000[Mechanical transport and transmission of Cryptosporidium parvum oocysts by wild filth flies178-83Am J Trop Med Hyg633-4\Animals Cattle Cattle Diseases/*transmission Cryptosporidiosis/transmission/*veterinary Cryptosporidium parvum/genetics/*growth & development DNA Primers DNA, Protozoan/isolation & purification Diptera/*parasitology Human Insect Vectors/*parasitology Male Polymerase Chain Reaction/veterinary Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S.Sep-OctOver the course of six months wild filth flies were collected from traps left for 7-10 days in a barn with or without a calf shedding Cryptosporidium parvum Genotype 2 oocysts in diarrheic feces. The oocysts of C. parvum transported on the flies' exoskeletons and eluted from their droplets left on visited surfaces were infectious for mice. The mean number of oocysts carried by a fly varied from 4 to 131, and the total oocyst number per collection varied from 56 to approximately 4.56 x 10(3). Fly abundance and intensity of mechanical transmission of infectious C. parvum oocysts were positively correlated, and both increased significantly when an infected calf was in the barn. Molecular data showed that the oocysts shed by infected calves were carried by flies for at least 3 weeks. Filth flies can acquire infectious C. parvum oocysts from unsanitary sites, deposit them on visited surfaces, and therefore may be involved in human or animal cryptosporidiosis.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=113885110002-9637 Journal Article11388511Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA. 6~?2Fayer, R. Trout, J. M. Graczyk, T. K. Lewis, E. J.2000uPrevalence of Cryptosporidium, Giardia and Eimeria infections in post-weaned and adult cattle on three Maryland farms103-12 Vet Parasitol932\Animals Cattle Cattle Diseases/*epidemiology Coccidiosis/epidemiology/*veterinary Cryptosporidiosis/epidemiology/*veterinary Cryptosporidium/isolation & purification Dairying Eimeria/isolation & purification Feces/parasitology Female Giardia/isolation & purification Giardiasis/epidemiology/*veterinary Male Maryland/epidemiology Prevalence WeaningNov 10The prevalence of Cryptosporidium, Giardia and Eimeria, in healthy, asymptomatic, post-weaned and mature cattle was investigated on three Maryland farms. One farm, a dairy research facility, had 150 multiparous Holstein milking cows; 24 were examined and Cryptosporidium andersoni was detected in three (12.5%) but neither Giardia nor Eimeria was detected. The second farm, a commercial dairy, had 57 multiparous Holstein milking cows and an equal number of heifers. Of 19 cows examined, C. parvum, Giardia duodenalis, and Eimeria bovis and/or E. ellipsoidalis were detected in two (10.5%), two (10.5%) and one (5.26%) cow, respectively. Of 23 heifers examined, C. parvum, Giardia, and E. bovis and E. ellipsoidalis, was detected in two (8.7%), four (17.4%), and five (21.7%), heifers, respectively. The third farm, a beef cattle breeding and genetics research facility, had 180 7- to 9-month old purebred black Angus. Of 118 examined for C. parvum and Giardia, 34 (28.8%) and 44 (37.3%) were positive, respectively, of 97 examined for E. bovis and/or E. ellipsoidalis 32 (33.0%) were positive. These findings, based on a method with a minimum detection level of 100 oocysts of C. parvum/g of feces, which underestimates the number of infected cattle, clearly demonstrate the presence of low level, asymptomatic infections in post-weaned and adult cattle in the United States and indicate the potential role of such cattle as reservoirs of infectious parasites.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=110352280304-4017 Journal Article11035228United States Department of Agriculture, Agricultural Research Service, 10300 Baltimore Avenue, Building 1040, Beltsville, MD 20705, USA. rfayer@lpsi.barc.usda.gov~??Roche, J. K. Martins, C. A. Cosme, R. Fayer, R. Guerrant, R. L.2000Transforming growth factor beta1 ameliorates intestinal epithelial barrier disruption by Cryptosporidium parvum in vitro in the absence of mucosal T lymphocytes5635-44 Infect Immun6810Animals Cattle Cell Line Cell Membrane/metabolism Cell Membrane Permeability/*drug effects Cell Polarity Colon/*immunology/parasitology Cryptosporidium parvum/growth & development/*pathogenicity Enzyme Activation Epithelial Cells/enzymology/parasitology/physiology Human Intestinal Mucosa/*immunology/parasitology/physiology Necrosis Protein Kinase C/metabolism Receptors, Transforming Growth Factor beta/metabolism Support, Non-U.S. Gov't Support, U.S. Gov't, P.H.S. T-Lymphocytes/immunology Transforming Growth Factor beta/*pharmacologyOctn Exposure to oocysts of the protozoan Cryptosporidium parvum causes intestinal epithelial cell dysfunction in vivo and in vitro, but effective means by which mucosal injury might be prevented remain unclear. We examined the ability of transforming growth factor beta1 (TGF-beta1)-a cytokine synthesized and released by cells in the intestine-to preserve the barrier function of human colonic epithelia when challenged with C. parvum oocysts and then studied the mechanisms involved. Epithelial barrier function was monitored electrophysiologically, receptors for TGF-beta1 were localized by confocal microscopy, and TGF-beta1-induced protein kinase C activation was detected intracellularly by translocation of its alpha isozyme. TGF-beta1 alone enhanced intestinal epithelial barrier function, while exposure to C. parvum oocysts (> or =10(5)/monolayer) markedly reduced barrier function to < or =40% of that of the control. When epithelial monolayers were pretreated with TGF-beta1 at 5.0 ng/ml, the barrier-disrupting effect of C. parvum oocysts was almost completely abrogated for 96 h. Further investigation showed that (i) the RI and RII receptors for TGF-beta1 were present on 55 and 65% of human epithelial cell line cells, respectively, over a 1-log-unit range of receptor protein expression, as shown by flow cytometry and confirmed by confocal microscopy; (ii) only basolateral and not apical TGF-beta1 exposure of the polarized epithelial monolayer resulted in a protective effect; and (iii) TGF-beta1 had no direct effect on the organism in reducing its tissue-disruptive effects. In exploring mechanisms to account for the barrier-preserving effects of TGF-beta1 on epithelium, we found that the protein kinase C pathway was activated, as shown by translocation of its 80-kDa alpha isozyme within 30 s of epithelial exposure to TGF-beta1; the permeability of epithelial monolayers to passage of macromolecules was reduced by 42% with TGF-beta1, even in the face of active protozoal infection; and epithelial cell necrosis monitored by lactate dehydrogenase release was decreased by 50% 70 h after oocyst exposure. Changes in epithelial function, initiated through an established set of surface receptors, likely accounts for the remarkable barrier-sparing effect of nanogram-per-milliliter concentrations of TGF-beta1 when human colonic epithelium is exposed to an important human pathogen, C. parvum.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=109924640019-9567 Journal Article10992464Divisions of Gastroenterology and of Geographic and International Medicine, Department of Medicine, University of Virginia Health System, Charlottesville, Virginia 22908, USA. jkr7m@virginia.edu~?;Morgan, U. M. Xiao, L. Fayer, R. Lal, A. A. Thompson, R. C.2000;Epidemiology and strain variation of Cryptosporidium parvum116-39Contrib Microbiol6Animals Cryptosporidiosis/*epidemiology/*parasitology Cryptosporidium/classification/*genetics/isolation & purification Epidemiology, Molecular Human *Variation (Genetics)ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1094351011420-9519 Journal Article Review Review, Tutorial10943510World Health Organisation Collaborating Centre for the Molecular Epidemiology of Parasitic Infections, Murdoch University, Australia. morgan@numbat.murdoch.edu.au~?eMorgan, U. M. Xiao, L. Limor, J. Gelis, S. Raidal, S. R. Fayer, R. Lal, A. Elliot, A. Thompson, R. C.2000PCryptosporidium meleagridis in an Indian ring-necked parrot (Psittacula krameri)182-3 Aust Vet J783~Animals Bird Diseases/*parasitology/pathology Comparative Study Cryptosporidiosis/parasitology/pathology/*veterinary Cryptosporidium/genetics/*isolation & purification DNA, Protozoan/isolation & purification Parrots/*parasitology Polymerase Chain Reaction/veterinary Poultry Diseases/*parasitology/pathology RNA, Ribosomal, 18S/genetics Support, Non-U.S. Gov't Turkeys/*parasitologyMarXOBJECTIVE: To perform a morphological and genetic characterisation of a Cryptosporidium infection in an Indian ring-necked parrot (Psittacula krameri) and to compare this with C meleagridis from a turkey. DESIGN: Tissue and intestinal sections from an Indian ring-necked parrot were examined microscopically for Cryptosporidium. The organism was also purified from the crop and intestine, the DNA extracted and a portion of the 18S rDNA gene amplified, sequenced and compared with sequence and biological information obtained for C meleagridis from a turkey as well as sequence information for other species of Cryptosporidium. RESULTS: Morphological examination of tissue sections from an Indian ring-necked parrot revealed large numbers of Cryptosporidium oocysts attached to the apical border of enterocytes lining the intestinal tract. Purified Cryptosporidium oocysts measured about 5.1 x 4.5 microns, which conformed morphologically to C meleagridis. The sequence obtained from this isolate was identical to sequence information obtained from a C meleagridis isolate from a turkey. CONCLUSION: Cryptosporidium meleagridis was detected in an Indian ring-necked parrot using morphological and molecular methods. This is the first time that this species of Cryptosporidium has been reported in a non-galliform host and extends the known host range of C meleagridis.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=10860158&0005-0423 Case Reports Journal Article10860158yWorld Health Organisation Collaborating Centre for the Molecular Epidemiology of Parasitic Infections, Western Australia.&~?;Xiao, L. Morgan, U. M. Fayer, R. Thompson, R. C. Lal, A. A.2000>Cryptosporidium systematics and implications for public health287-92Parasitol Today167Animals Cryptosporidiosis/*parasitology/veterinary Cryptosporidium/*classification/genetics Human *Public Health Support, Non-U.S. Gov't Support, U.S. Gov't, Non-P.H.S. Support, U.S. Gov't, P.H.S. *ZoonosesJul}There is controversy in the taxonomy of Cryptosporidium parasites and the public health significance of Cryptosporidium isolates from various animals. Recent advances in molecular characterization of Cryptosporidium parasites have allowed the re-examination of species structure of the genus Cryptosporidium. Non-parvum Cryptosporidium spp and new C. parvum genotypes in immunocompromised humans can now be clearly detected. In this article, Lihua Xiao and colleagues summarize the current biological and molecular evidence for different Cryptosporidium spp, and the public health importance of these species and new C. parvum genotypes.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=1085864710169-4758 Journal Article Review Review, Tutorial10858647pDivision of Parasitic Diseases, Centers for Disease Control and Prevention, Atlanta, GA 30341, USA. lax0@cdc.gov~?)Fayer, R. Trout, J. M. Walsh, E. Cole, R.20001Rotifers ingest oocysts of Cryptosporidium parvum161-3J Eukaryot Microbiol472_Animals *Cryptosporidium parvum Diet *Feeding Behavior Rotifera/*physiology Species SpecificityMar-AprSix genera of rotifers including Philodina, Monostyla, Epiphanes, Euchlanis, Brachionus, and Asplanchna were exposed to oocysts of Cryptosporidium parvum cleaned of fecal debris. Unstained oocysts and those stained with fluorescein-conjugated monoclonal antibody were added to suspensions of viable rotifers and were examined by phase-contrast, differential interference contrast, and fluorescence microscopy. Rotifers of all six genera were observed ingesting oocysts. A maximum of 25 oocysts was observed in the stomachs of Eauchlanis and Brachionus. Euchlanis and Epiphanes were observed excreting boluses containing up to eight oocysts. It was not determined whether rotifers digested or otherwise rendered oocysts nonviable.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=107508441066-5234 Journal Article10750844United States Department of Agriculture, Agricultural Research Service, Beltsville, Maryland 20705, USA. rfayer@lpsi.barc.usda.govW~?[Graczyk, T. K. Fayer, R. Trout, J. M. Jenkins, M. C. Higgins, J. Lewis, E. J. Farley, C. A.2000_Susceptibility of the Chesapeake Bay to environmental contamination with Cryptosporidium parvum106-12 Environ Res822IAnimals Biological Assay Cryptosporidiosis/*epidemiology Cryptosporidium parvum/*isolation & purification Disease Outbreaks Environmental Monitoring/methods Feces/parasitology Geese Human Maryland/epidemiology Mice Oysters/*parasitology *Public Health Risk Assessment Seawater/parasitology Support, Non-U.S. Gov't Water PollutionFebehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=106625250013-9351 Journal Article10662525Department of Molecular Microbiology and Immunology, Johns Hopkins University, Baltimore, Maryland 21205, USA. tgraczyk@jhsph.eduK~?1Li, X. Fayer, R. Trout, J. Jenkins, M. Palmer, R.2001SEffects of gamma irradiation on the survival of Encephalitozoon intestinalis spores91SJ Eukaryot MicrobiolSupplAnimals Cell Line Dose-Response Relationship, Radiation Encephalitozoon/pathogenicity/*physiology/*radiation effects *Gamma Rays Spores/growth & development/radiation effectsehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=119060941066-5234 Journal Article11906094GUSDA, ARS, Animal Waste Pathogen Laboratory, Beltsville, MD 20705, USA.[~?4Phelps, K. K. Lindsay, D. S. Sumner, S. S. Fayer, R.2001kImmunohistochemistry based assay to determine the effects of treatments on Cryptosporidium parvum viability40S-41SJ Eukaryot MicrobiolSupplAnimals Anti-Bacterial Agents/*pharmacology Cryptosporidium parvum/*drug effects/*growth & development/pathogenicity Immunohistochemistry/methods Parasitic Sensitivity Tests/methods Paromomycin/*pharmacology Support, U.S. Gov't, Non-P.H.S. Tumor Cells, CulturedCell culture infectivity assays can provide an accurate means of detecting viable Cryptosporidium parvum oocysts from environmental samples or to test the effects of various treatments on oocyst infectivity. Cell culture assays can also be used to test candidate chemotherapeutic agents. The use of a human cell line provides a situation close to human infection. The present assay uses an anti-Cryptospordium primary antibody, combined with a biotinylated secondary antibody, and an immunoperoxidase detection system. Cryptosporidium parvum oocysts excysted in vitro when placed on monolayers of HCT-8 cells and developmental stages including schizonts and merozoites were visualized using light microscopy of the immunoperoxidase stained slides and by transmission electron microscopy of infected HCT-8 cell cultures. Because the immunoperoxidase system used gives a permanent preparation, the cell cultures can be retained and examined later. Dose titration of oocysts indicated that as few as 50 inoculated oocysts could be detected. The activity of paromomycin was evaluated in this system and 500 microg/ml produced a 97.8% reduction in infection.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11906073,1066-5234 Evaluation Studies Journal Article11906073JDepartment of Food Science and Technology, Virginia Tech, Blacksburg, USA.~?EFayer, R. Trout, J. M. Xiao, L. Morgan, U. M. Lai, A. A. Dubey, J. P.2001/Cryptosporidium canis n. sp. from domestic dogs1415-22 J Parasitol876Animals Base Sequence Cryptosporidiosis/epidemiology/*veterinary Cryptosporidium/*classification/genetics Dogs/*parasitology Genotype HIV Infections/complications Human Molecular Sequence Data Prevalence RNA, Ribosomal/genetics Sequence Homology, Nucleic AcidDecOocysts of Cryptosporidium, from the feces of a naturally infected dog and from an HIV-infected human, were identified as the previously reported canine genotype of Cryptosporidium parvum, hereafter referred to as Cryptosporidium canis n. sp. Also among the oocysts from the dog, a trace amount of C. parvum bovine genotype was detected. Cryptosporidium canis oocysts from both the dog and human were infectious for calves. Oocysts excreted by calf 1 (dog source) were approximately 90% C. canis and 10% C. parvum, whereas those excreted by calf 3 (human source) were 100% C. canis. Oocysts from calf 1 infected calf 2 resulting in excretion by calf 2 of oocysts approximately 90% C. parvum and 10% C. canis. Oocysts of C. canis were not infectious for BALB/c neonatal mice or immunosuppressed C57 juvenile mice, although all control mice became infected with the C. parvum Beltsville isolate. Oocysts of C. canis from calf 1 and the human were structurally indistinguishable from oocysts of the C. parvum Beltsville isolate (bovine). However, C. canis oocysts differed markedly at the molecular level from all known species of Cryptosporidium based on sequence data for the 18S rDNA and the HSP 70 gene. The differences in genetics and host specificity clearly differentiate C. canis as a new species.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=117808310022-3395 Journal Article11780831United States Department of Agriculture, Agricultural Research Service, Animal and Natural Resources Institute, Beltsville, Maryland 20705-2350, USA.~?CHiggins, J. A. Jenkins, M. C. Shelton, D. R. Fayer, R. Karns, J. S.2001WRapid extraction of DNA From Escherichia coli and Cryptosporidium parvum for use in PCR5321-4Appl Environ Microbiol6711-Animals Cryptosporidium parvum/*genetics/growth & development DNA, Bacterial/*isolation & purification DNA, Protozoan/*isolation & purification Escherichia coli O157/*genetics/growth & development Nucleic Acid Amplification Techniques/methods Polymerase Chain Reaction/*methods Support, Non-U.S. Gov'tNovSThe Xtra Amp tube, Isocode paper, Instagene matrix, and PrepMan matrix methods were evaluated for their ability to rapidly extract PCR-quality DNAs from Escherichia coli O157:H7 and Cryptosporidium parvum. All methods provided satisfactory DNA from E. coli, and the Xtra Amp and Instagene reagents provided satisfactory DNA from C. parvum.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=11679362,0099-2240 Evaluation Studies Journal Article11679362zU.S. Department of Agriculture-Agricultural Research Service, Beltsville, Maryland 20705, USA. jhiggins@anri.barc.usda.gov ~?KFayer, R. Trout, J. M. Lewis, E. J. Santin, M. Zhou, L. Lal, A. A. Xiao, L.2003IContamination of Atlantic coast commercial shellfish with Cryptosporidium141-5 Parasitol Res892Animals Atlantic Ocean Base Sequence Cattle Cryptosporidiosis/parasitology Cryptosporidium/genetics/growth & development/*isolation & purification *Fisheries Food Microbiology Mice Mice, Inbred BALB C Microscopy, Fluorescence Polymerase Chain Reaction Shellfish/*parasitologyJanShellfish (oysters and/or clams) were obtained from 37 commercial harvesting sites in 13 Atlantic coast states from Maine to Florida and one site in New Brunswick, Canada. Gill washings from each of 25 shellfish at each site were examined by immunofluorescence microscopy (IFA) for oocysts of Cryptosporidium. Gill washings from another 25 shellfish at each site were grouped into five pools of five shellfish each. DNA from each pool was utilized for PCR and genotyping. Oocysts were found in 3.7% of 925 oysters and clams examined by IFA in shellfish from New Brunswick and 11 of 13 states. Cryptosporidium DNA was detected by PCR in 35.2% of 185 pools. Cryptosporidium parvum genotypes 1 and 2, and Cryptosporidium meleagridis,all of which have been identified in infected humans, were identified at 37.8% of the sites. Gill washings from every site were tested for the presence of infectious oocysts by biological assay in neonatal BALB/c mice but no mice were found infected, suggesting that either the oocysts were no longer infectious or infections in mice were below the level of detection. Collectively, these findings indicate that Cryptosporidium species, indicative of pollution from human and animal feces and potentially infectious for humans, were found in commercial shellfish from 64.9% of sites examined along the Atlantic coast by either microscopy or molecular testing. Previous reports link periods of high rainfall with the elevated numbers of pathogen contaminated shellfish. Because shellfish in the present study were examined during a period of exceptionally low precipitation, the data are thought to underestimate the number of Cryptosporidium contaminated shellfish likely to be found during periods of normal or above normal precipitation.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=124890140932-0113 Journal Article12489014{Animal Waste Pathogen Laboratory, Agricultural Research Service, USDA, Beltsville, MD 20705, USA. rfayer@anri.barc.usda.gov~?BJenkins, M. Trout, J. M. Higgins, J. Dorsch, M. Veal, D. Fayer, R.2003LComparison of tests for viable and infectious Cryptosporidium parvum oocysts1-5 Parasitol Res891qAmylopectin/analysis Animals Cattle Cells, Cultured Comparative Study Cryptosporidium parvum/*growth & development/*pathogenicity DNA Probes *In Situ Hybridization, Fluorescence/methods Mice Mice, Inbred BALB C Oocysts/*growth & development/isolation & purification *Reverse Transcriptase Polymerase Chain Reaction Support, Non-U.S. Gov't Temperature Time Factors WaterJanThe purpose of this study was to compare different assays for viable Cryptosporidium parvum incubated in water at a temperature commonly found in the environment. C. parvum oocysts were stored in sterile water for 9 months at 15 degrees C. A sample was removed monthly and analyzed by five different assays to determine oocyst viability. Mouse infection and cell culture showed that C. parvum oocysts remained viable and infectious when stored for 7 months at this temperature. Fluorescence in situ hybridization (FISH) using probes directed to ribosomal RNA was also applied to these oocysts. The proportion of FISH-positive oocysts was 70-80% for the first 2 months of storage, decreased and remained nearly constant at 40-50% for 3-7 months, then decreased to 20% by 8 months, and to 0% by 9 months. Amylopectin content and mRNA for amyloglucosidase (CPAG), as measured by RT-PCR, decreased much more rapidly. By 3 months and for the remainder of the incubation period, amylopectin content was 20% of the original amount present in the oocysts. The CPAG RT-PCR signal at 3 months was 50% of that observed after 1 month storage, 20% at 4 months, and was not detected thereafter. Thus, results from cell culture and mouse infection assay exhibited the best agreement, the FISH assay showed modest agreement with these assays, and CPAG RT-PCR and the amylopectin assay displayed marginal agreement with the other three assays.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=12474036,0932-0113 Evaluation Studies Journal Article12474036}Animal Waste Pathogen Laboratory, Agricultural Research Service, USDA, Beltsville, MD 20705, USA. mjenkins@anri.barc.usda.gov~?!#Trout, J. M. Walsh, E. J. Fayer, R.2002Rotifers ingest Giardia cysts1038-40 J Parasitol885Animals Giardia/*growth & development Image Processing, Computer-Assisted Microscopy, Fluorescence Microscopy, Interference Microscopy, Phase-Contrast Rotifera/*parasitology Sewage/parasitology Water/parasitologyOctSSeven species of rotifers representing 6 genera, Epiphanes, Plationus, Asplanchna, Philodina species A, Philodina species B. Platyias, and Brachionus, were exposed to Giardia cysts isolated from the feces of experimentally infected holstein calves. Giardia cysts were prestained with a fluorescein isothiocyanate-conjugated monoclonal antibody and mixed with viable rotifers on 3-well Teflon-coated microscope slides. Organisms were observed with phase-contrast, differential interference contrast, and fluorescence microscopy. Five rotifer species, Epiphanes brachionus, Plationus patulus, Philodina (both A and B), and Platyias quadricornis, ingested varying numbers of cysts, which were retained within the rotifers' bodies throughout the observation period. Rotifer ingestion of Giardia cysts may represent a means of reducing water contamination.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=124351560022-3395 Journal Article12435156Animal and Natural Resources Institute, Agricultural Research Service, US Department of Agriculture, Beltsville, Maryland 20705, USA. jtrout@anri.barc.usda.gov{~?"RFayer, R. Trout, J. M. Lewis, E. J. Xiao, L. Lal, A. Jenkins, M. C. Graczyk, T. K.2002=Temporal variability of Cryptosporidium in the Chesapeake Bay998-1003 Parasitol Res8811^Animals Cryptosporidiosis/parasitology/physiopathology Cryptosporidium/*classification/genetics/*isolation & purification/pathogenicity Cryptosporidium parvum/classification/genetics/isolation & purification/pathogenicity Mice Mice, Inbred BALB C Oocysts/isolation & purification Oysters/*parasitology Rain *Seasons Seawater/*parasitology TemperatureNov8Although Cryptosporidium has been found worldwide in molluscan shellfish from waters contaminated with human and animal feces, little or no related environmental data have been obtained. In the present study, oysters ( Crassostrea virginica) were collected eight times over 3 years from seven sites in the Chesapeake Bay or its tributaries, with accompanying data on water temperature, salinity, rainfall, and streamflow. Oyster gill washings were examined by immunofluorescence microscopy for Cryptosporidium oocysts. Of 1,590 oysters collected, 19.6% had detectable oocysts. Of 53 collections, oocysts were detected 81% of the time. The time when the greatest percentage of oysters at most sites had detectable oocysts coincided with the time of greatest weekly and monthly rainfall, greatest streamflow into the Bay, and lowest water temperatures. In 28% of 53 collections, C. parvum genotypes 1 and 2 and C. baileyi were identified by PCR and gene sequencing. Oocyst infectivity was confirmed from 37.5% of 40 collections by initiating C. parvum genotype 2 infections in mice.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=123751660932-0113 Journal Article12375166{Animal Waste Pathogen Laboratory, Agricultural Research Service, USDA, Beltsville, MD 20705, USA. rfayer@anri.barc.usda.gov3~?#!Trout, J. M. Santin, M. Fayer, R.2003;Identification of assemblage A Giardia in white-tailed deer1254-5 J Parasitol896Animals Centrifugation, Density Gradient/veterinary Cytoskeletal Proteins/genetics Deer/*parasitology Feces/parasitology Female Fluorescent Antibody Technique/veterinary Genotype Giardia/classification/genetics/*isolation & purification Giardiasis/epidemiology/parasitology/*veterinary Maryland/epidemiology Molecular Sequence Data Polymerase Chain Reaction/veterinary Prevalence Protozoan Proteins/geneticsDecFecal samples were collected from hunter-killed white-tailed deer (Odocoileus virginianus) during a managed hunt in a central Maryland county. Fecal samples were cleaned of debris and concentrated by CsCl density gradient centrifugation and stained with MerIFluor reagents. Stained samples were examined by fluorescent microscopy for the presence of Giardia sp. cysts. One of 26 samples was found to be positive for Giardia sp. Polymerase chain reaction amplification using primers directed to the beta-giardin and TPI genes identified the same sample as the only positive one. Sequencing of the beta-giardin and TPI genes revealed that the Giardia sp. belonged to assemblage A, a genotype infectious for humans and also reported in a small percentage of cattle. This is the first report of assemblage A Giardia sp. in deer and suggests that deer could be a potential source of infectious cysts for humans and cattle.ehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=147409230022-3395 Journal Article14740923Environmental Microbial Safety Laboratory, Animal and Natural Resources Institute, Agricultural Research Service, United States Department of Agriculture, Beltsville, Maryland 20705, USA. jtrout@anri.barc.usda.gov~?$,Santin, M. Ludwig, K. Fayer, R. Trout, J. M.20032First report of Giardia in coyotes (Canis latrans)709J Eukaryot Microbiol50 SupplAnimals Animals, Wild Base Sequence Carnivora/*parasitology Cytoskeletal Proteins/genetics DNA Primers Giardia lamblia/genetics/*isolation & purification Giardiasis/diagnosis/*veterinary Molecular Sequence Data Polymerase Chain Reaction Protozoan Proteins/geneticsehttp://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=PubMed&dopt=Citation&list_uids=14736229&1066-5234 Case Reports Journal Article14736229PUSDA, ARS, Environmental Microbial Safety Laboratory, Beltsville, MD 20705, USA.e~?%&Fayer, R.