The Efficient Technology Of Detecting Plankton Diversity And Its Application In The Study Of Brown Tide In Bohai

Eukaryotic plankton plays an important role in the water ecosystems of materialcycles, energy flow and information transfer, so study its structure will contribute tothe development of water environment. Molecular biology has fast and accuratecharacteristics compared to traditional microscopic analysis and it provides a newmethod for studying plankton diversity. Conventional molecular biology methodsinclude clone library, DGGE, RFLP and so on. But complex experimental procedures,high costs and low coverage become its drawback. Next-generation sequencing iswidely used in the area of genomics and transcriptome because of its efficiency,accuracy and high throughput.In this manuscript, we invented a strategy for the amplification and paired-endsequencing of V9regions from environmental eukaryotic18S rDNA gene ampliconlibraries. Then we recommended an approach for estimating plankton richness byclustering the reads into operational taxonomic units and uncovering taxonomicinformation. This invention had created a new method to explore plankton diversitybecause of its simple, fast, cost-effective, high throughput and sensitivitycharacteristics.As important sea farms, seasides of Shanhaiguan, Funing, Xinkaikou, Letingstrongly improve the local sea farm industry. However, the eutrophication trulyincrease the frequency of red tides, especially the brown tide had discovered since2009with brown tide algae diameter around2μm and maximum cell density around109cell/L. Due to the fact that this kind of tide could tremendously prevent foodintake of bay scallop (Argopecten irradians), economic losses happened during thattime. The morphology character of brown tide algae is obscure, using optics methodto observe is hard and the bias is usually high. In the mean time, traditional molecular method has some drawbacks such as time consuming, higher cost and narrowcoverage. In this study, nanoplankton samples (Shanhaiguan, Funing, Xinkaikou andLeting) were collected during the brown tide in2011and amplicon libraries of V9hypervariable region of18S ribosomal DNA were built and sequenced by Hiseq2000.Based on the analysis, we obtained235-255phytoplankton species and35-97zooplankton,nanoplankton diversity varies between different stations. The diversityindex analysis showed that seaside area of Leting had the highest diversity andspecies evenness. Xinkaikou had the highest species richness whereas Shanhaiguanhad the lowest species evenness, richness and diversity. The result strongly revealedthe high sensitivity and widely coverage of this method. Then we found that higherphytoplankton species were dinophyceae, chlorophyta, bacillariophyta and higherzooplankton species were ciliophora, metazoa, cercozoa, euglenozoa. The planktoncommunity transformed maybe due to the change of physical-chemical elements andbiological factors during brown tide.In the mean time, we did analysis of this brown tide algae, results also showedthat Aureococcus anophagefferens and Minutocellus polymorphus had relativelyhigher density in all four stations. The two species had different distribution indifferent stations, in seaside area of Shanhaiguan, Aureococcus anophagefferens hadhigher density, next to is XinkaiKou, third is Leting, Funing had the lowest density. Incontrast, in terms of Minutocellus polymorphus, the density from highest to lowest isas follows: Funing, Leting, Shanhai Guan, Xinkai Kou. According to the aboveanalysis, A. anophagefferens and M. polymorphus are likely to co-occur during thebrown tide in the Bohai Sea. This study will contribute to a better understanding ofthe mechanism behind the brown tide break.