Estimation Of Giant Panda's Intestinal Flora Structure In Intestine Based On ERIC-PCR And 16s RDNA-RFLP Technologies

The subadult panda's intestinal tract harbors a complex microbial ecosystem which plays a key role in nutrition and health. Although this macrobiotic has been studied in fairly detail by culture techniques and plate count in subadult panda's feces suggest that mass of the observable bacteria cannot be cultivated. In order to find OTU the diversity of subadult panda's intestinal flora, investigate the structural variation and changes, and find OTU whether the structure of intestinal flora community in subadult panda was related to the seasonal change. In this study, we analyzed the characteristics of subadult panda's intestinal flora by ERIC-PCR based fingerprint and 16S rDNA-RFLP. The contents are summarized as follows:1 To investigate the similarity and stability of subadult giant pandas'intestinal flora in four seasons. Total DNA of microbial community was extracted from three subadult giant pandas'feces samples and then analyzed by ERIC-PCR. Major bands of the ERIC-PCR fingerprint were purified, cloned, sequenced and alignment. Results show that intestinal flora was very similar between different subadult panda individuals in same season, but microbial community of individuals in different seasons tended to be low similarity. The season might affect the diversity of subadult giant panda's intestinal flora. Escherichia coli and Citrobacter koseri were the predominant bacteria in intestinal flora of subadult panda. Conclusion:The preliminary results suggest that the diversity of the subadult giant panda's intestinal flora is related with the season in a certain extent.2. The diversity of subadult panda's intestinal flora was analyzed by 16S rDNA cloning library. The amplified 16S rDNA sequences were analyzed by RFLP (Restriction Fragment Length Polymorphism). PCR-RFLP analysis was carried OTU with restriction enzyme Hinfl and Mspl, and 29 OTUs (Operational Taxonomic Units) were obtained from these samples. The results of diversity Shannon-Wiener index, Coverage C, and species richness calculated indicated that bacteria diversity in subadult panda's intestinal were very rich. Sequence analysis of 7 superiority OTUs showed diverse phyla of bacteria in the 16S rDNA cloning library, which consisted of Escherichia coli, Uncultured bacterium, Shigella flexneri and Citrobacter freundii. The dominant group of the cloning library was Escherichia coli (47.5% of the total clones). More than 13.75% of the total clones showed high similarity to uncultured bacteria. The results suggest that diversity of subadult panda's intestinal flora is abundant and exist a large amount of unknown bacteria.3. The results obtained by the two methods were basically accordance, and both of them had advantages and disadvantages. This article indicated that combining 16S rDNA cloning sequences and the analysis of ERIC-PCR fingerprinting profiles were timely, rapid and accurate molecular test facility of diversification in Giant Panda's intestinal flora.