Comparison of advanced methodologies for diatom identification within dynamic coastal communities

Diatom community composition has a critical influence on global ocean health and ecological processes. Developing accurate and efficient methods for diatom identification under dynamic environmental conditions is essential to understanding the implications of diatom community changes. Two developing methods for identifying and enumerating phytoplankton, cell imaging and molecular sequencing, are experiencing rapid advancements. This study aims to compare diatom taxonomic composition results within natural assemblages derived from rapidly advancing methods, FlowCam imaging and metabarcoding of the V4 region of the 18S rRNA gene, with traditional light microscopy cell counting techniques. All three methods were implemented in tandem to analyze changes in dynamic diatom assemblages within simulated upwelling experiments conducted in the California upwelling zone. The results of this study indicate that, summed across all samples, DNA sequencing detected four times as many genera as morphology-based methods, thus supporting previous findings that DNA sequencing is the most powerful method for analyzing species richness. Results indicate that all three methods returned comparable relative abundance for the most abundant genera. However, the three methods did not return comparable absolute abundance, primarily due to barriers in deriving quantities in equal units. Overall, this study indicates that at the semi-quantitative level of relative abundance measurements, FlowCam imaging and metabarcoding of the V4 region of the 18S rRNA gene yield comparable results with light microscopy but at the qualitative and quantitative levels, enumeration metrics diverge, and thus method selection and cross-method comparison should be performed with caution.