Molecular Characterization and Identification serves to develop and employ molecular diagnostics including primers for polymerase chain reaction (PCR) and DNA probes, that target Pfiesteria species, related dinoflagellates, or Amphanomyces invadens.
DNA was isolated from clonal cultures isolated at VIMS and from other sources following identification by scanning electron microscopy (SEM). Three ribosomal RNA gene regions were amplified by PCR from these clonal-culture DNAs and sequenced to identify regions of sequence that were unique to each species or genus. The DNA sequences were also compared to determine the genetic relationships among the Pfiesteria-like dinoflagellates (PLOs).
Based on DNA-sequence differences we have developed sensitive and specific molecular diagnostics for Pfiesteria piscicida, Pfiesteria shumwayae, and other PLOs including "Lucy"-like and Cryptoperidiniopsis species and for A. invadans. These molecular tools were initially tested and optimized on DNA from clonal cultures and were shown to specifically amplify the targeted DNA and did not cross amplify the DNA of other species. In addition, the probes and primers have been show to specifically target these organisms in environmental samples and are currently being used in environmental monitoring programs.
Principal Investigator: Kimberly Reece
- Seon-Sook An
- Gail Scott
- Lidia Sandoval