采用Illumina MiSeq测序技术分析断奶幼兔盲肠微生物群落的多样性

Caecal Microbiota Diversity Analysis of Weaning Rabbits Using Illumina Miseq Sequencing Technology

本试验旨在采用Illumina MiSeq测序技术分析断奶幼兔盲肠微生物群落的多样性。选取30窝同日出生、胎次相同和平均窝重相近的健康新生仔兔,于14日龄开始补饲,断奶当日(30日龄)从30窝仔兔中选取48只体重相近的健康幼兔作为试验兔,饲喂不添加抗生素的基础饲粮。分别于断奶当日及断奶后1、2和3周随机选取6只健康幼兔屠宰,无菌采集盲肠内容物,从中选取3个样本提取微生物基因组总DNA,采用Illumina MiSeq测序技术检测其中的微生物群落多样性。结果显示:1)健康幼兔盲肠微生物群落α多样性指数指标(ACE值、Chao1值、Shannnon值)随日龄的增长有升高的趋势(P<0.10)。2)不同日龄健康幼兔盲肠微生物各菌门的相对丰度差异不显著(P>0.05),且均主要由厚壁菌门、拟杆菌门、变形菌门和疣微菌门组成;在科水平上,厚壁菌门主要由瘤胃球菌科和毛螺旋菌科组成,拟杆菌门主要由拟杆菌科和紫单胞菌科组成;在属水平上,拟杆菌属、Barnesiella、Akkermansia、普雷沃氏菌属和4种未知菌属为优势菌属;拟杆菌属和Parabacteroides在断奶当天的相对丰度较高,且与其他日龄组差异显著(P<0.05)。3)健康幼兔盲肠中均存在乳杆菌属,且相对丰度在各日龄组间差异不显著(P>0.05)。由此得出,断奶时健康幼兔盲肠微生物区系已基本建立,相对丰度最大的优势菌属为拟杆菌属,但断奶后随着日龄的增长,微生物组成趋于复杂。

The current study was conducted to evaluate the caecal microbiota diversity of weaning young rabbits by using Illumina Miseq sequencing technology.Thirty litters of newborn healthy rabbits with the same birthday and parity and similar average body weight were used,and forty-eight healthy rabbits with similar body weight were selected on the day of weaning（30days of age）.No antibiotics were used in the basal diet and all rabbits were fed restrictedly from 14 days of age to the end of the trial.Six healthy young rabbits were randomly selected and slaughtered on the day of weaning and 1,2and 3weeks after weaning,respectively.Three samples of caecal content at each sampling time were collected for extracting the microbial genomic total DNA.Microbiota diversity of caecal contents was evaluated using Illumina Miseq sequencing technology.The results showed as follows：1）the alpha diversity indexes（ACE value,Chao1 value and Shannnon value）of caecal microbiota of healthy young rabbits were tended to rise with age increasing（P〈0.10）.2）The caecal microbiota of healthy young rabbits at different age was mainly consisted of Firmicutes,Bacteroidetes,Proteobacteria and Verrucomicrobia at the phylum level,and their relative abundances were not significantly affected by age（P〉0.05）.Ruminococcaceae and Lachnospiraceae were the dominant families in Fimicutes,and Bacteroidetes was predominantly composed by Bacteroidaceae and Porphyromonadaceae.Bacteroides,Barnesiella,Akkermansia,Prevotella and four unclassified genus were the common dominant genera at the genera level.The relative abundances of Bacteroides and Parabacteroides on the day of weaning were higher which significantly higher than those of other ages（P〈0.05）.3）Lactobacillus existed in the caecum of healthy young rabbits,and its relative abundance was not significantly changed by age（P〉0.05）.Base on the results,it is concluded that the caecal microflora has been basically established and the relative abundance of Bacteroides is the highest at weaning,but microbial components tend to be complex after weaning with age increasing.