牙鲆(Paralichthysolivaceus)仔稚幼鱼肠道菌群结构比较分析

Composition of Intestinal Bacterial Community of Japanese Flounder(Paralichthys olivaceus) During Early Life Stages

采用MiSeq 16S rRNA高通量测序技术和生物信息学分析方法,构建了牙鲆(Paralichthys olivaceus)工厂化人工育苗模式下仔稚幼鱼阶段6个不同发育时期18个样品的16S rRNA基因测序文库,共获得7462个OTU(Operational Taxonomic Unit),分类为42个菌门972个菌属。对肠道菌群的形成过程及结构多样性变化分析显示,牙鲆初孵仔鱼的菌群组成多样性丰富,体内的优势菌为变形菌门(Proteobacteria)、厚壁菌门(Firmicutes)和拟杆菌门(Bacteroidetes);在9日龄和21日龄摄食轮虫(Rotifer)和卤虫(Artemia sp.)幼体样品中,肠道的优势菌群结构较单一,变形菌门成为此时期肠道的优势菌群;45日龄摄食配合饲料后,肠道中变形菌门的相对丰度显著降低,厚壁菌门和拟杆菌门的相对丰度明显增大,成为肠道菌群的优势菌群。在属水平的菌群结构中发现,牙鲆仔稚幼鱼肠道优势菌群的种类和数量都发生了较大变化,在9日龄和21日龄时期肠道中弧菌属(Vibrio)相对丰度最高,到45日龄后相对丰度锐减到最低水平;拟杆菌属(Bacteroides)和普氏菌属(Prevotella)在80日龄后达到较高水平,成为肠道优势菌属;厚壁菌门的8个菌属在80–115日龄时期均发展成为优势菌属,定植于牙鲆的肠道。本研究揭示了工厂化人工育苗模式下牙鲆仔稚幼鱼肠道菌群结构及演替规律。

A healthy microbial community in digestive tract is critically important during early life stages of fish. To examine the microbial diversity in the gastrointestinal tract of Japanese flounder （Paralichthys olivaceus）, we surveyed the intestinal bacteria of P. olivaceus during the larvae and juvenile stages. In this study, Illumina MiSeq of 16S rRNA and biological information analysis method were used to explore the intestinal bacterial community composition in indoor tank-culture system. Sampling was carried out throughout the larvae and juvenile stages for six time points. The obtained 7462 operational taxonomic units （OTUs） were classified into 42 bacterial species and 972 genera. The results indicated that larval fish （1 day-post-hatch; dph） displayed a distinct and diversified gut microflora, with Proteobacteria, Bacteroidete and Firmicutes dominating the community structure. During the age of 9 and 21 dph, Proteobacteria population increased significantly and remained to be the main species. By comparison, Firmicutes had fast turnover rate and increased sharply at 45 dph after feeding formulated food. The change of intestinal dominant microflora genus was also significant. The relative abundance of Vibrio was the highest during the period of live feed （9 and 21 dph sampling）, and was the lowest level after feeding formulated feed （115 dph）. By contrast, the Bacteroides and Prevotella became the predominant intestinal bacteria at 80 dph, whereas Firmicutes became a core microbiota at 80 dph. Blautia and other related genus colonized in the intestinal tract of P. olivaceus and became the dominant microbiota between 80 and 115 dph. The results revealed the change and establishment of microbiota during the transition stages from larvae to juvenile in response to live and formulated feed. Our results provided a database for analyzing the role of the intestinal micro ecological system of P. olivaceus at larvae and juvenile stages and have important implication for larval production ofP. olivaceus.