DISCRIMINATION BETWEEN DOMOIC-ACID-PRODUCING AND NONTOXIC FORMS OF THE DIATOM PSEUDONITZSCHIA-PUNGENS USING IMMUNOFLUORESCENCE

Separate polyclonal antibodies were developed against cell surface ant igens of the 2 forms of the pennate diatom Pseudonitzschia (previously known as Nitzschia) pungens, i.e. forma multiseries (the domoic-acid- producing form) and forma pungens (the nontoxic form). Positive antige nic reactions were visualized with epifluorescence microscopy, using a fluorescein isothiocyanate (FITC) indirect immunofluorescence assay. The assay successfully distinguished 31 clones of f. multiseries from the 17 clones of f. pungens tested, with no cross reactions of the ant isera between the 2 forms. The antisera were active against P. pungens cells isolated from Prince Edward Island and Nova Scotia, Canada, and Massachusetts, Rhode Island, Texas and Washington, USA. Of the 27 oth er clones tested from the genera Pseudonitzschia and Nitzschia, 2 othe r domoic-acid-producing species (P. australis and P. pseudodelicatissi ma) and 2 nontoxic species (P. subcurvata and P. fraudulenta) showed a slight positive response to the antisera, as did the domoic-acid-prod ucing pennate diatom Amphora coffaeiformis. These reactions are not gr eat enough to cause concern about misidentification, but pose question s about phylogenic relationships. Other representatives from the class Bacillariophyceae and from 8 other major classes of phytoplankton did not cross react with the antisera. Excellent labelling was obtained w ith live cells and those frozen at -60-degrees-C, or preserved in 2% g lutaraldehyde-paraformaldehyde, 2% borate-buffered formalin or 2% para formaldehyde. Immunofluorescence shows great promise as a technique to distinguish between the 2 forms of P. pungens for research and monito ring purposes.