Researches On The Gut Microbiota Structure Of Diet-induced Obesity Rats

In recent decades, the population with obesity has increased in an astonishing speed. Obesity and the related disorders, including type II diabetes, hypertension, cardiovascular pathology and nonalcoholic fatty liver disease, hazard enormously the health and the life quality of the humanity. Although there are a lot of researches which demonstrate the relationship between obesity and gene mutation, since the genotype of humanity would not change significantly in a relatively short time, many researchers have diverted their attention to the environmental factors. As the second genome of human, intestinal flora has commensal relationship of mutually benefit with the host. It has been broadly recognized that there exists relations between obesity and microbial constitution with their impact on metabolic and physiological tune in the host. Therefore, exploration of the relationship between microflora and obesity has become the focus of the academia in the world.In this research, we divide a batch of SD rats randomly into two groups: the first group, diet-induced obesity (DIO) group, was fed with high-fat-diet (HFD), while the second group, control group, was fed with normal chow diet. We sampled their feces every week from the start point to the seventh week. Some methods of molecular microbial ecology, ERIC-PCR, community hybridization, quantitative PCR and denaturing gradient gel electrophoresis (DGGE) etc., combined with multivariate statistical analysis (PCA, PLS) were applied to analyze the microbiota structure of the samples from both time points. We attempt to demonstrate the change and difference of intestinal microflora between DIO group and control group and discover a critical factor and molecular marker which can differentiate the two groups.First, we research on the intestinal microbiota structure of diet-induced obesity rats and the specific molecular markers. ERIC-PCR combined with community hybridization were used to analyze the difference of fecal microbiota structure between the two groups, while multivariate statistical analysis was used to identify the most important bands causing the difference, which were sequenced afterwards and aligned to find the species with the closest relationship. Finally, we devised a pair of specific primers according to the sequence to validate the results. As a result, we find that there exists significant difference between DIO and control group and a molecular marker which existed only in DIO group was identified, which can be used as a marker of the gut microbiota structure of DIO rats.Second, we conducted researches on intestinal Bifidobacterium group of diet-induced obesity rats by group specific PCR-DGGE. We use Bifidobacterium group specific PCR-DGGE to demonstrate the difference of Bifidobacterium species between the two groups and real-time quantitative PCR to clarify the difference in quantity. The critical bands were also sequenced to identify its species. As a result, we find that Bifidobacterium species between the two groups is significantly different and a critical band has a similarity of 100% with Bifidobacterium minimum, which is probably relative to obesity.Through the researches above, we demonstrate the difference of intestinal microbiota structure between DIO and normal rats and found a specific gene fragment of DIO rats. Then, we analyze the structural difference of Bifidobacterium group between DIO and normal rats and identified a species of Bifidobacterium which is specifically found in DIO group. These results can help us further understand the relationship between obesity and intestinal microbiota structure of the host.