Molecular Analysis of the 18S rRNA Gene of Cryptosporidium serpentis in a Wild-Caught Corn Snake ( Elaphe guttata guttata ) and a Five-Species Restriction Fragment Length Polymorphism- Based Assay That Can Additionally Discern C. parvum from C. wrairi

Abstract

An adult wild-caught corn snake (Elaphe guttata guttata) was presented for humane euthanasia and necropsy because of severe cryptosporidiosis. The animal was lethargic and >5% dehydrated but in good flesh. Gastric lavage was performed prior to euthanasia. Histopathologic findings included gastric mucosal hypertrophy and a hemorrhagic erosive gastritis. Numerous 5- to 7-μm-diameter round extracellular organisms were associated with the mucosal hypertrophy. A PCR, acid-fast stains, Giemsa stains, and an enzyme immunoassay were all positive for Cryptosporidiumspp. PCR and restriction fragment length polymorphism (RFLP) analysis on gastric lavage and gastric mucosal specimens, and subsequent sequencing of the 18S rRNA gene, enabled a distinct molecular characterization of the infecting organism as Cryptosporidium serpentis. Until recently, studies on snakeCryptosporidium have relied on host specificity and gross and histopathologic observations to identify the infecting species. A multiple alignment of our sequence against recently published sequences of the 18S rRNA gene of C. serpentis (GenBank accession no.AF093499 , AF093500 , and AF093501 [L. Xiao et al., unpublished data, 1998]) revealed 100% homology with the C. serpentis(Snake) sequence (AF093499 ) previously described by Xiao et al. An RFLP method to differentiate the five presently sequenced strains of Cryptosporidium at this locus was developed. This assay, which uses SpeI andSspI, complements a previously reported assay by additionally distinguishing the bovine strain ofCryptosporidium from Cryptosporidium wrairi.

Detection ofCryptosporidiumspecies in feces or gastric contents from snakes and lizards as determined by polymerase chain reaction analysis and partial sequencing of the 18S ribosomal RNA gene

“…As reported earlier, the clinical signs reported in some Cryptosporidium-positive reptiles are consistent with chronic gastrointestinal disease of subtle clinical onset. 11,12,17 However, not all animals with gastrointestinal diseases were positive for cryptosporidia. In particular, swelling of the abdomen, gastrointestinal wall thickening, malodorous feces, and diarrhea can be caused by other microorganisms, such as flagellates or enteropathogenic bacteria.…”

“…8,10,16 Other methods used for the differentiation of certain Cryptosporidium species usually comprise important human pathogenic, or possibly zoonotic, pseudoparasitic species and genotypes, but not all reptilian pathogenic types. 11,13,21 Polymerase chain reaction (PCR) assays for the detection and subsequent sequencing of Cryptosporidium species have been described, usually based on a relatively long amplicon. 1,21 The primers of the present study have originally been desi gned for identification of cryptosporidian species from formalin-fixed, paraffin-embedded tissue samples.…”

“…8,10,16 Other methods used for the differentiation of certain Cryptosporidium species usually comprise important human pathogenic, or possibly zoonotic, pseudoparasitic species and genotypes, but not all reptilian pathogenic types. 11,13,21…”

Detection and Genotyping of Cryptosporidium Spp. in Biofilms and Cattle Feces From the Black River Watershed, Jamaica

“…Therefore, the high degree of sequence homology presented in the 18S rRNA enabled the use of primers that could amplify a wide range of Cryptosporidium spp. isolated from environment sources (Kimbell, Miller, Chavez, & Altman, 1999).…”

Cryptosporidium and Giardia as water contaminant pathogens in Hungary

“…C. serpentis infection in lizards is usually asymptomatic, whereas the infection in snakes frequently causes clinical disease (gastric hyperplasia, postprandial regurgitation and firm midbody swelling or chronic debilitating enteritis) and pathological changes (Brownstein et al 1977, Cranfield et al 1999, Kimbell et al 1999. No pathological changes were found in the intestine and cloacae of adult lizards infected by C. saurophilum, but weight loss, abdominal swelling, and mortality occurred in some colonies of juvenile geckos Infection occurs when cysts are ingested orally by a susceptible host.…”