An attempt to semi-quantify potentially toxic diatoms of the genus Pseudo-nitzschia in Tokyo Bay, Japan by using massively parallel sequencing technology

Currently, no tool is available that precisely identifies and quantifies all 48 species of Pseudo-nitzschia simultaneously. In the present study, we used massively parallel sequencing-based technology with genus -specific Pseudo-nitzschia primers to investigate the distribution and population dynamics of the genus in Tokyo Bay, Japan. Furthermore, we attempted to quantify the abundance of each species using both relative abundance data detected in the massively parallel sequencing-based survey, and cell count data, obtained by light microscopy. Several species of Pseudo-nitzschia were detected continuously from July 10 to September 9, 2016 in the sampling location. Cell densities ranged from 2-300 cells mL(-1) (sample no.=10). Fourteen operational taxonomic units (OTUs) attributable to Pseudonitzschia were detected, and 99.9% of sequences detected by massively parallel sequencing belonged to the genus. Each of the most abundant OTUs comprised a single species, and were identified, in decreasing order of abundance, as F. multistriata (Takano) Takano, P. pungens (Grunow ex Cleve) Hasle, P. fraudulenta (Cleve) Hasle, P. multiseries (Hasle) Hasle, and P. galaxiae Lundholm & Moestrup. Species identification was possible in 9 of the 14 OTUs detected, of which several were detected in Tokyo Bay for the first time. The cell abundance ranges of the five most abundant species were estimated as 1,976-246,395, 0.03-52,999, 5-42,820, 4-24,617, and 0.03-1,007 cells L-1, respectively. The detection limit was 0.03 cells L-1.