[Composition and changes of intestinal flora in septic mouse model].

OBJECTIVE

To investigate the dynamic changes of intestinal flora in septic model mice.

METHODS

Forty-two male SPF C57BL/6 mice were selected, the sepsis model was reproduced by cecal ligation and puncture (CLP), and the experimental mice were divided into CLP 6-12 hours group (n = 9) and 1, 2, 3 days group (all n = 10) and Sham group (n = 3) according to the time points after modeling. Intestinal flora 16S rRNA sequencing was carried out within feces from the colonic lumen of mice, and the effective sequences were clustered to obtain an operational classification unit (OTU) for statistical analysis of biological information, including Alpha diversity analysis, species composition analysis, principal coordinate analysis (PCoA analysis) and species difference analysis (LEfSe analysis), the dynamic changes of intestinal flora after CLP modeling were analyzed.

RESULTS

Compared with Sham group, with the prolongation of modeling time, the Alpha diversity of intestinal flora decreased, showing a decrease in community richness index (3 days after CLP: Chao index 1 was 367.9±162.6 vs. 508.3±105.9, Ace index was 372.5±151.9 vs. 498.8±104.2), Shannon index decreased in community diversity index (3 days after CLP: 2.57±1.06 vs. 4.30±0.57, Simpson index increased (3 days after CLP: 0.26±0.19 vs. 0.04±0.03), suggesting that with the progress of CLP modeling time, the richness and diversity of intestinal flora decreased. According to species composition analysis, in OTU level, OTU 633 accounted for the highest proportion in CLP 1 day group (24.79%), OTU 1016 was dominant in CLP 2 days and 3 days groups, and the highest proportion was 61.75% in CLP 3 days group; in genus level, the abundance of norank_f_Muribaculaceae showed a slight increase and then a significant decrease with time, the abundance of Escherichia-Shigella increased significantly in CLP 2 days and 3 days groups, the abundance of Lactobacillus increased first and then decreased with time, the abundance of Bacteroides showed a trend of gradual increase with time. PCoA analysis suggested that CLP 6-12 hours group had a higher structural similarity with Sham group. The flora structure changed gradually with the time after modeling, and the change was significant after 3 days of CLP. LEfSe analysis indicated that the main components that caused the differences among the groups were g_norank_f_Muribaculaceae, g_Prevotellaceae_UCG-001, s_uncultured_Bacteroidales_bacterium_g_norank_f_Muribaculaceae, g_Parabacteroides, Escherichia-Shigella and OTU 1016. The differences in abundance of Escherichia-Shigella and OTU 016 among the five groups ranked first in the genus level and OTU level respectively; the abundance of Escherichia-Shigella gradually increased from 0.01% (0%, 0.02%) in 6-12 hours after CLP to 44.79% (3.71%, 53.75%) in 3 days with time, and the abundance of OTU 1016 increased from 0.01% (0%, 0.02%) in 6-12 hours after CLP to 44.69% (3.66%, 53.64%) in 3 days with time.

CONCLUSIONS

Intestinal dysbiosis occurred in all CLP model groups, the diversity of the flora gradually decreased with the progress of modeling time, Escherichia-Shigella gradually became the dominant bacteria.