Development of a multiplex PCR for the detection of Haplosporidium nelsoni, Haplosporidium costale and Perkinsus marinus in the eastern oyster (Crassostrea virginica, Gmelin, 1971).

Haplosporidium costale (SSO), Haplosporidium nelsoni (MSX) and Perkinsus marinus (Dermo) have caused oyster mortality on the North American east coast since the 1950s. Currently, the monitoring of oyster populations for these pathogens depends on histopathology for H. nelsoni, H. costale and the Ray/Mackin assay for P. marinus. In this study we describe the development and optimization of a multiplex polymerase chain reaction (MPCR) for the detection of H. nelsoni, H. costale and P. marinus. In addition, we determine its specificity and sensitivity. The MPCR clearly detects and differentiates the protozoan pathogens. There was no cross-reactivity between these species. The MPCR was able to detect DNA from H. nelsoni as low as 10 fg and P. marinus and H. costale as low as 1 pg. The MPCR allows for the detection of H. nelsoni, H. costale and P. marinus in a single PCR reaction. Copyright 2001 Academic Press.