RESUMO
OBJECTIVES: To genetically characterize an unusual genotype of Cryptosporidium from the stools of humans with diarrhoea and to identify risk factors in the affected patients. METHODS: DNA was extracted from human faeces where Cryptosporidium oocysts were detected by light microscopy. Cryptosporidial gene fragments from six different loci were analysed by PCR alone, PCR/RFLP and by DNA sequencing. Oocysts were characterized by light and immunofluorescence microscopy and epidemiological data was collected from the affected patients. RESULTS: Analysis of the Cryptosporidium oocyst wall protein (COWP) gene amplified from > 2000 human faecal samples identified 19 patients all of which produced an unusual RFLP profile. Subsequent DNA sequence analysis of this and an additional four genetic loci (including 18S rRNA sequences) confirmed these as a homogeneous group which was genetically distinct from Cryptosporidium parvum. The isolates were identified as Cryptosporidium meleagridis since the gene sequences were identical to those from this species recovered from birds. Conventional microscopy showed oocysts indistinguishable from C. parvum and reacted strongly with two different commercially available anti-oocyst monoclonal antibodies. None of the patients showed risk factors unusual for cryptosporidiosis; however, ten of the cases occurred during the summer/autumn, six had a history of foreign travel, four were co-infected with Giardia, two were HIV positive, and six were without identifiable immunocompromising factors. CONCLUSIONS: This study further confirms that C. meleagridis, in addition to C. parvum, is involved in human disease. The study also highlights the lack of basic information on the host range of this genus of parasites, the complexity of the transmission routes involved in human cryptosporidiosis, and the value of molecular techniques in identify hitherto unrecognised differences in Cryptosporidium from human faeces.
