凡纳滨对虾和罗氏沼虾肠道微生物及抗生素抗性基因多样性分析

Diversity of gut microbiota and antibiotic resistance genes in Litopenaeus vannamei and Macrobrachium rosenbergii

探究凡纳滨对虾和罗氏沼虾肠道微生物及抗生素抗性基因(antibiotic resistance genes, ARGs)种类的差异。通过高通量测序和变性梯度凝胶电泳(denaturing gradient gel electrophoresis, DGGE)技术分析2种虾肠道微生物群落结构差异和微生物多样性,并运用PCR方法检测了2种虾肠道细菌常见38种ARGs的携带情况。结果显示,获得凡纳滨对虾和罗氏沼虾肠道细菌有效序列分别为42 795和40 713条,物种注释单元(operational taxonomic unit, OTU)数目分别为124和82,分类地位明确的细菌种类分别隶属5个门、17个属和5个门、16个属。凡纳滨对虾肠道细菌的优势类群为变形菌门,所占比例为75.45%,优势菌属为副球菌属(25.83%)和不动杆菌属(25.24%);罗氏沼虾肠道细菌的优势类群是厚壁菌门(49.74%),优势菌属为乳球菌属(49.01%)和弧菌属(29.98%)。凡纳滨对虾肠道细菌(2.19)Shannon指数高于罗氏沼虾肠道细菌(1.78),表明前者肠道细菌多样性大于后者。DGGE图谱的分析结果与高通量测序一致,2种虾肠道细菌种类差异很大。PCR结果显示,凡纳滨对虾肠道细菌携带15种ARGs,罗氏沼虾肠道细菌携带14种ARGs。本实验表明凡纳滨对虾肠道细菌的群落种类多样性、OTU丰富度、物种总数和ARGs种类均高于罗氏沼虾肠道细菌,为后续肠道微生物资源的挖掘提供了理论依据。

To compare the differences in gut microbiota and antibiotic resistance genes (ARGs) between Litopenaeus vannamei and Macrobrachium rosenbergii, microbiota community structure and microbiota diversity of two shrimps were analyzed by high-throughput sequencing and denaturing gradient gel electrophoresis (DGGE), and 38 kinds of ARGs in the gut bacteria of two shrimps were detected. The effective bacterial sequences of L. vanna- mei and M. rosenbergii were 42 795 and 40 713. The results showed that the numbers of operational taxonomic units (OTUs) were 124 and 82, and the bacterial types with clear classification status were 5 phyla, 17 genera and 5 phyla, 16 genera. The dominant group in the gut bacteria of L. vannamei is Proteobacteria, which accounts for 75.45%. The dominant genera are Paracoccus (25.83%) and Acinetobacter (25.24%). The dominant group in gut bacteria of M. rosenbergii is Firmicutes (49.74%), and the dominant genera are Lactococcus (49.01%) and Vibrio (29.98%). The Shannon index of the gut bacteria (2.19) of L. vannamei was higher than that of M. rosenbergii (1.78), indicating that the gut bacteria diversity of the former was greater than that of the latter. The analysis results of DGGE fingerprint were consistent with those of high-throughput sequencing, and the bacterial diversity of the two shrimps was significantly different. The PCR results showed that the gut bacteria of L. vannamei carried 15 ARGs and gut bacteria of M. rosenbergii carried 14 ARGs. Our study revealed that the community diversity, OTU richness, total number of species and ARGs in the gut bacteria of L. vannamei were higher than those of M. rosenbergii, which provided a theoretical basis for the subsequent excavation of gut microbial resources.