1887

Abstract

Rapid identification of the two major species of associated with human infections, and , is important for investigating outbreaks of cryptosporidiosis. This study reports the development and validation of a real-time PCR TaqMan procedure for detection of species and identification of and in stool specimens. This procedure comprised a generic TaqMan assay targeting the 18S rRNA for sensitive detection of species, as well as two other TaqMan assays for identification of and . The generic species assay can be duplexed with the -specific assay. The generic species assay was able to detect ten species and did not cross-react with a panel of ten other protozoan parasites. The generic species assay could detect 1–10 oocysts in a 300 μl stool specimen, whilst each of the species-specific TaqMan assays had detection sensitivities that were approximately tenfold higher. The 18S rRNA assay was found to detect species in 49/55 DNA extracts from stool specimens containing either or . The TaqMan assay correctly identified in 24/31 validation panel specimens containing this species. The -specific assay correctly identified in 21/24 validation panel specimens containing this species. This real-time PCR procedure was used to detect and identify and in stool specimens from outbreak investigations in the USA and Botswana, resulting in identification of and/or in 66/67 stool specimens shown to be positive for these species using other techniques. From the outbreak specimens tested, the TaqMan procedure was found to have a specificity of 94 %. This TaqMan PCR procedure should be a valuable tool for the laboratory diagnosis of cryptosporidiosis caused by and during outbreak investigations.

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2008-09-01
2019-11-14
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