New Insights on the Ecology of Free-living, Heterotrophic Nanoflagellates Basedon the Use of Molecular Biological Approaches

摘要

Nanoplanktonic protists comprise a diverse assemblage of flagellate species.Current methods for identifying small protists do not readily permit identification and enumeration of nanoplanktonic flagellates in cultures or field samples. The aim of this study was to apply molecular biological techniques for identification and enumeration of nanoflagellate taxa in water samples. Restriction fragment length polymorphism (RFLP) analysis of small subunit rDNA differentiated cultures of heterotrophic nanoflagellates according to established taxonomic classification at the generic and species level. In addition, this method permitted discrimination of polymorphic forms of Paraphysomonas species. A fluorescent in situ hybridization method that uses rRNA-targeted oligonucleotide probes for counting protists in cultures and environmental water samples was developed. This method gave estimates of nanoplankton abundances that might be more representative of natural abundances than estimates obtained by fluorochrome stains. Abundances of the cosmopolitan heterotrophic flagellate, paraphysomonas imperforata, enumerated using species-specific probes, were found to be extremely low in coastal environments. Water temperature, the absolute abundance of P. imperforata, and nanoplankton grazers appeared to play a role in determining P. imperforata dominance. Results from this study also suggest that enrichment cultivation can select for nanoflagellates that may not be representative of abundant oceanic species.