The identification of Cryptosporidium species and Cryptosporidium parvum directly from whole faeces by analysis of a multiplex PCR of the 18S rRNA gene and by PCR/RFLP of the Cryptosporidium outer wall protein (COWP) gene

Abstract A multiplex polymerase chain reaction (PCR) procedure to amplify 18S rRNA gene fragments has been developed. Amplified DNA fragments of the expected size were obtained which were specific for Cryptosporidium parvum and Cryptosporidium wrairi (422 bp), Cryptosporidium baileyi (1106 bp) and Cryptosporidium muris (1346 bp). Cryptosporidium parvum and C. wrairi can be distinguished using a PCR/restriction fragment length polymorphism (RFLP) analysis of the Cryptosporidium outer wall protein (COWP) gene, and these two techniques were applied to DNA extracted from whole faeces using a simple and rapid procedure. Cryptosporidium parvum DNA was detected in the faeces of 72 humans and 24 calves where cryptosporidial oocysts were demonstrated using conventional light microscopy. The specific DNA fragments were not amplified using extracts of material containing other lower eukaryotic parasites.