Use of molecular probes to assess geographic distribution of Pfiesteria species.

We have developed multiple polymerase chain reaction (PCR)-based methods for the detection of Pfiesteria sp. in cultures and environmental samples. More than 2,100 water and sediment samples from estuarine sites of the U.S. Atlantic and gulf coasts were assayed for the presence of Pfiesteria piscicida Steidinger & Burkholder and Pfiesteria shumwayae Glasgow & Burkholder by PCR probing of extracted DNA. Positive results were found in about 3% of samples derived from routine monitoring of coastal waters and about 8% of sediments. The geographic range of both species was the same, ranging from New York to Texas. Pfiesteria spp. are likely common and generally benign inhabitants of coastal areas, but their presence maintains a potential for fish and human health problems.

Heteroduplex mobility assay-guided sequence discovery: elucidation of the small subunit (18S) rDNA sequences of Pfiesteria piscicida and related dinoflagellates from complex algal culture and environmental sample DNA pools.

The newly described heterotrophic estuarine dinoflagellate Pfiesteria piscicida has been linked with fish kills in field and laboratory settings, and with a novel clinical syndrome of impaired cognition and memory disturbance among humans after presumptive toxin exposure. As a result, there is a pressing need to better characterize the organism and these associations. Advances in Pfiesteria research have been hampered, however, by the absence of genomic sequence data. We employed a sequencing strategy directed by heteroduplex mobility assay to detect Pfiesteria piscicida 18S rDNA "signature" sequences in complex pools of DNA and used those data as the basis for determination of the complete P. piscicida 18S rDNA sequence. Specific PCR assays for P. piscicida and other estuarine heterotrophic dinoflagellates were developed, permitting their detection in algal cultures and in estuarine water samples collected during fish kill and fish lesion events. These tools should enhance efforts to characterize these organisms and their ecological relationships. Heteroduplex mobility assay-directed sequence discovery is broadly applicable, and may be adapted for the detection of genomic sequence data of other novel or nonculturable organisms in complex assemblages.