Picophytoplankton identification by flow cytometry and high-throughput sequencing in a clean reservoir

Understanding picophytoplankton variations that play important roles in the material circulation and energy flow are critical to assessing overall status of waterbody, especially for clean reservoirs which remain a relatively stable community structure and high species diversity due to lower nitrogen and phosphorus nutrients. However, their response to key environmental factors and tightly acting microbial remains poorly understood. Traditional quantification methods are limited, such as chlorophyll-a, turbidity and microscope. There are still many defects with present molecular analysis. In this study, a flow cytometric analysis and high-throughput sequencing combination methodology was developed and tested on clean water from a reservoir, by a monthly dynamic for a vegetative period April-September in 2019 to improve the accuracy of dynamic monitoring for the picophytoplankton system. More species of Pico-Cyanobacteria and Pico-Eukaryotes were discovered. The increased percentage of pigment compounds from 8.2% to 76.3% proves the effective reduce of heterotrophic disturbing and enrichment of target populations. Picophytoplankton that was previously neglected due to their low relative abundance has once again entered the scope of our eyes. Phytoplankton were divided into three categories. The first one was the highly abundant and frequently present taxa, the second one was the low-abundance but highlytransient population, and the third one was the low abundance and stable group. Synechococcus, Emiliania, Tetraselmis and Thalassiosira were dominant picophytoplankton and displayed obvious temporal and spatial distribution characteristics. Pico-PE rich Cyanobacteria and Nano-Eukaryotes with high transience abnormally increased in summer. Temperature, ammonia-N, nitrate-N, turbidity and total nitrogen were most influencing factors, while some picophytoplankton with special physiological structure showed distinct competitive advantages in the microbial community. As for the off-flavor compounds, the concentration of 2-methylisoborneol and geosmin were high even 66.7% and 20.8% of the samples exceeded their olfactory threshold. Chrysochromuina, Planktothrix and Microcystis might be the potential producers.

Automated summary

This study developed a new methodology combining flow cytometric analysis and high-throughput sequencing for monitoring picophytoplankton in a clean reservoir, discovering more species of Pico-Cyanobacteria and Pico-Eukaryotes to improve monitoring accuracy.