应用RFLP和DGGE技术分析工厂化养殖凡纳滨对虾肠道微生物群落特征

Analysis of intestinal microecology of Litopenaeus vannamei in industrial aquaculture by RFLP and DGGE techniques

中文关键词: 凡纳滨对虾  肠道  微生物  工厂化养殖  PCR-RFLP  PCR-DGGE

英文关键词: Litopenaeus vannamei  Intestinal  Microbial  Industrial aquaculture  PCR-RFLP  PCR-DGGE

中文摘要:

      本研究应用PCR-RFLP(Polymerase Chain Reaction-Restriction Fragment Length Polymorphism)和PCR-DGGE(Polymerase Chain Reaction-Denaturing Gradient Gel Electrophoresis)技术，对工厂化养殖的凡纳滨对虾肠道菌群进行多样性分析。RFLP结果显示，8月样品中不同的克隆子为5个，其中以不可培养细菌（Unculturable bacteria）为主要优势菌，其次为鲁杰氏菌属菌种Ruegeria spp.和Rhodobacterales spp.。依据微生物多样性的覆盖率分析结果表明，所构建16S rDNA克隆文库的覆盖率为97.5%。10月样品中克隆子为8个，其中以不可培养细菌、芽孢杆菌属Bacillus spp.和弧菌属Vibrio spp.为主要优势菌属，其次为Photobacterium spp.和Neptunomonas spp.；所构建16S rDNA克隆文库的覆盖率为90.8%。应用DGGE分析8月和10月对虾肠道样品，菌群以不可培养细菌和弧菌属为主，其次为Streptomyces spp.、Ruegeria spp.、Enterococcus spp.和Photobacterium damselae。

英文摘要:

      The composition of bacterial community in the intestine of white shrimp Litopenaeus vannamei under industrial culture conditions was investigated by 16S rDNA-based molecular methods of denaturing gradient gel electrophoresis (DGGE) analysis and restriction fragment length polymorphism (RFLP) analysis. Sequencing results of cloned 16S rDNA amplicons revealed that unculturable bacteria were the dominant species, Ruegeria spp., and Rhodobacterales spp. were subdominant species in samples collected in August 2011. The coverage rate by the 16S rDNA clone library was 97.5%. Eight different RFLP patterns of the clones were obtained by restriction RFLP analysis from samples collected in October 2011. Sequencing results of cloned 16S rDNA amplicons revealed that unculturable bacteria, Bacillus spp. and Vibrio spp. were the dominant species; Photobacterium spp. and Neptunomonas spp. were subdominant species. The coverage rate by the 16S rDNA clone library was 90.8%. DGGE analysis was also carried out for the samples. Sequencing results of cloned 16S rDNA amplicons revealed that unculturable bacteria and Vibrio spp. were the dominant species. Comprehensive analysis of PCR-RFLP and PCR-DGGE experimental results would be helpful to understand the microbial components in the intestine of the white shrimp, and to comprehensively and objectively reveal the structure and diversity of white shrimp microbial community.