Abstract:For rapid, accurate and high throughput detection of Haplosporidium nelsoni, pyrosequencing analysis coupled with PCR amplification of the target sequence was developed. H. nelsoni DNA sequence was obtained by the OIE reference PCR method. Pyrosequencing special primers were designed targeting the conserved region of the sequence. The DNA of Haplosporidium-positive oyster samples was chosen to amplify the target sequence using pyrosequencing primers, and the sequence was analyzed by PyroMarkTM ID System. BLAST online showed that the sequence was specific for H. nelsoni. Oyster samples were detected by both the PCR-pyrosequencing method and the OIE reference PCR method. The results showed that the PCR-pyrosequencing detection method could identify H. nelsoni and the result was consistent with the OIE reference PCR examination. The method meets the requirements of H.nelsoni quarantine and provides a new approach for the examination of other animal diseases.
