Intestinal Bacterial Diversity Analysis Of Eight Marine Economic Animals

The intestinal bacterial diversity of Fenneropenaeus chinensis, Litopenaeus vannamei, Trachypenaeus curvirostris and Oratosquilla oratoria in Crustacea, and five fishes Sciaenops ocellatus, Hexagrammos otakii, Lateolabrax japonicus, Kareius bicoloratus and Scophthatmus maximus in Actinopterygii were analysed; phylogenetic trees were constructed based on the 16S rDNA gene of bacteria. The aim of this research is to analyze the intestinal bacterial diversity of these shrimps and fishes based on 16S rDNA gene, providing basic understanding of intestinal bacterial composition structure. The main work consists of the following three parts:1. Culture-independent method was used, and 16S rDNA was amplified via PCR from total genomic DNA extracted from intestinal microorganisms of F. chinensis, L. vannamei, T. curvirostris and O. oratoria, with clone libraries constructed later. Positive clones were digested by restriction endonuclease BsuRâ… and Hin6â… respectively, and then representative clones were sequenced. DNA sequence analysis and BLAST analysis indicated that more than 70% of the intestinal bacteria of F. chinensis, L. vannamei and T. curvirostris belonged to the Gammaproteobacteria, with Vibrio possessed the highest proportion. The gut of Oratosquilla oratoria had very low bacterial diversity, and most of the bacteria were uncultured ones. The intestinal bacteria of L. vannamei had the highest bacterial diversity, including Vibrio, Aeromonas, Photobacterium, Pseudoalteromonas, etc.2. Molecular method (RFLP analysis) was used to analyze the intestinal bacterial diversity of H. otakii, K. bicoloratus, S. ocellatus and L. japonicus. DNA sequence analysis indicated that more than 70% of the intestinal bacteria of these fishes belonged to the Gammaproteobacteria, and the statistical analysis revealed that the L. japonicus gut bacteria community had the highest diversity. The main bacterial composition of S. ocellatus was Vibrio, Photobacterium and Spiroplasma; the main bacterial composition of L. japonicus was Vibrio and Photobacterium; the main bacterial composition of H. otakii was Vibrio and Aliivibrio; the main bacterial composition of K. bicoloratus was Aeromonas and Mycoplasma.3. Molecular method was used to analyze the intestinal bacterial diversity of Scophthatmus maximus. 136 positive clones were selected randomly from the constructed 16S rDNA library to be digested by restriction-endonuclease (BsuRâ… ,Hin6â… å’ŒRsaâ… ), and 46 RFLP Patterns were obtained, with 36 of them had only one clone, indicating that the intestinal bacterial of Scophthatmus maximus had relatively higher diversity. The total intestinal microorganism genomic DNA of Scophthatmus maximus was isolated with high quality using in situ degradation, which was used for metagenomic sequcing.