Study On Cow Endometritis Associated Microbiota And Its Interaction Mechanisms With The Host

Endometritis is a common obstetrical disease, which leads to huge economic losses in dairy industry. Reproductive tracts microecology plays an important role in the occurrence of cow endometritis, therefore, exploring its mechanism not only can lay theoretical basis for developing probiotics used for the prevention and treatment of cow endometritis, but also provide reference for clarifying pathogenesis of endometritis. In the present study, in order to clarify cow endometritis associated microbiota and its interaction mechanisms with the host, change of bacterial diversity in reproductive tracts of healthy cows and endometritis cows were investigated using PCR-DGGE and Real-time PCR, then, RNA-seq technique was used to analyse genomic expression profiling of cow endometrial cellsUterine samples were collected from healthy cows and endometritis cows on days 7, 14, 25, 36, and 46 postpartum and then PCR-DGGE was used to investigate their microbial community structure. The results showed that uterine bacterial diversity in healthy and endometritis groups followed the same succession rule: increasing firstly then decreasing. On day 7, Helcococcus ovis, Fusobacterium necrophorum and Parvimonas micra were only detected in endometritis group, while Ureaplasma diversum, Parasporobacterium paucivorans, Methanotorris igneus and Clostridium perfringens were detected in both groups. On day 14, Ureaplasma diversum and Helcococcus ovis were only detected in healthy group, while Porphyromonas levii, Bacteroides heparinolyticus, Methanotorris igneus and Escherichia coli were detected in both groups. On day 25, Porphyromonas levii, Methanotorris igneus and Pseudomonas mendocina were detected in both groups. On day 36, bacterial diversity in endometritis group was significantly higher than healthy group, and Porphyromonas levii, Micrococcus luteus, Escherichia fergusonii, Actinobacillus seminis, Methanotorris igneus and Fusobacterium necrophorum were only detected in endometritis group. On day 46, bacterial diversity in both groups decreased sinificantly and there was no significant difference between the two groups.Based on results of PCR-DGGE, number of six bacteria in uterus of healthy and endometritis cows postpartum was investigated using Real-time PCR. The results showed that number of the bacteria changed dynamically as different stage of postpartum and the number was almost the same in healthy and endometritis cows on day 46. Number of Ureaplasma diversum was the highest among the bacteria and deacreased as the time expanded, furthermore, the number in endometritis group was higher than healthy group. Number of Fusobacterium necrophorum in both groups increased firstly then decreased, which was higher in endometritis group than healthy group from day 7 to day 36. There was no significant difference on number of Methanotorris igneus and Pseudomonas mendocina between healthy and endometritis groups during different stages of postpartum. On day 25, number of Porphyromonas levii in endometritis group was significantly higher than healthy group. Number of Helcococcus ovis increased firstly then decreased, and it was higher in endometrtis group than healthy group on day 14 and 25. Combined with results of PCR-DGGE, Fusobacterium necrophorum and Porphyromonas levii may be the main pathogenic bacteria of cow endometritis.Results of RNA-seq showed that 108 genes were differentially expressed inlcuding 90 up-regulated genes and 18 down-regulated genes in cow endometrial cells of LPS treatment group compared to control group. Results of GO classification and enrichment of 108 different genes showed that, 24 GO terms were annotated to the biological process, 15 GO terms were annotated to the cell component, and 10 GO terms were annotated to the molecular function. 68 GO terms were enrichmented to the biological process, 4 GO terms were enrichmented to the cell component and 8 GO terms were enrichmented to the molecular function.The KEGG results revealed that 20 signaling pathway were more highly expressed in LPS treated groups including TNF signaling pathway, NOD-like receptor signaling pathway and NF-kappa B signaling pathway which are closely related to inflammation process. Then four genes were selected to verify the results of RNA-seq using Real-time PCR and the results showed that RNA-seq conducted in this study was accurate.In conclusion, the present study demonstrated change of bacterial diversity in uterus of healhty cows and endometritis cows and their difference using PCR-DGGE and Real-time PCR, Futhermore, genomic expression profiling of cow endometrial cells was analysized using RNA-seq technique. The results not only lay a foundation for calrify pathogenesis of endometritis, but also provided reference for developing probiotics to prevent and treat cow endometritis.