Application of combined morphological-molecular approaches to the identification of planktonic protists from environmental samples

The value of molecular databases for unicellular eukaryotic identification and phylogenetic reconstruction is predicated on the availability of sequences and accuracy of taxonomic identifications that accompany those sequences. Biased representation of sequences is due in part to the differing ability to isolate and culture various groups of protists. Techniques that allow for parallel single-cell morphological and molecular identifications have been reported for a few groups of unicellular protists. We have sought to explore how those techniques can be adapted to work across a greater phylogenetic diversity of taxa. Twelve morphologically diverse and abundant members of limnetic microplankton, including ciliates, dinoflagellates, cryptophytes, stramenopiles, and synurophytes, were targeted for analysis. These cells were captured directly from environmental samples, identified, and prepared for sequence analyses using variations of single-cell extraction techniques depending on their size, mobility, and the absence or presence of the cell wall. The application of these techniques yielded a strong congruence between the morphological and molecular identifications of the targeted taxa. Challenges to the single-cell approach in some groups are discussed. The general ability to obtain DNA sequences and morphological descriptions from individual cells should open new avenues to studying either rare or difficult to culture taxa, even directly at the point of collection (e.g. remote locations or shipboard).