title: Detection of Cryptosporidium-specific serum immunoglobulins in captive snakes by a polyclonal antibody in the indirect ELISA.
abstract: An indirect enzyme-linked immunosorbent assay (ELISA) was developed for the detection of systemic Cryptosporidium-specific antibodies (Ab) in the serum of captive snakes. An optimal concentration of Cryptosporidium soluble protein-cocktail antigen was 20 micrograms/mL. In a direct ELISA, alkaline phosphatase-labeled rabbit-anti-snake polyvalent Ab detected snake immunoglobulins at a dilution of 1:12800. It was possible to detect Cryptosporidium-specific Ab at a serum dilution of 1:1600. Nineteen of 26 (73%) captive snakes (eight species) were Cryptosporidium-seroconverted; however, the differences in Ab titer among species were not significant. The distribution of absorbance values did not conform to a normal distribution (P < 0.05); mean Ab titer of nine snakes with cryptosporidiosis was significantly higher than mean Ab titer of the remaining ten snakes (P < 0.01). The monoclonal Ab test showed that the stools of 17 (89%) of 19 seropositive snakes contained Cryptosporidium oocysts; nine of those 19 (47%) stools were positive by acid-fast-stained (AFS) direct wet smear (DWS). A significant positive relationship between Cryptosporidium-seroconversion and snake age was observed (P < 0.03); the mean age of seropositive snakes was significantly higher than the mean age of seronegative animals (P < 0.01). The proposed assay is fast, reproducible, could be used for retrospective and prospective seroepizootiological studies, and assist ophidian centers in establishing Cryptosporidium-free collections. After standardization, the assay can be used for large-scale screening of captive snakes.
authors.affiliation: Department of Molecular Microbiology and Immunology, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, MD 21205, USA. tgraczyk@phnet.sph.jhu.edu
authors.forename: T K
authors.initials: TK
authors.lastname: Graczyk
authors.forename: M R
authors.initials: MR
authors.lastname: Cranfield
mesh_terms: D000375:Aging; D000818:Animals; D000913:Antibodies, Protozoan / Q000097:blood*; D003457:Cryptosporidiosis / Q000175:diagnosis / Q000276:immunology / Q000662:veterinary*; D003458:Cryptosporidium / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D007136:Immunoglobulins; D011817:Rabbits; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D012911:Snakes / Q000276:immunology*; D013045:Species Specificity
chemical_list: D000913:Antibodies, Protozoan; D007136:Immunoglobulins