Characterization Of Oropharyngeal Bacterial Flora In Patients With Cirrhosis

Objective:To establish an applicable solution for DNA concentration for the study of oropharynx microbial community structure. To profile the oropharyngeal bacterial flora in patients with cirrhosis and Cirrhosis complicated by pneumonia.Methods:First, oropharyngeal swabs were collected from healthy controls, patients with cirrhosis and cirrhosis complicated with pneumonia. Then, DNA was extracted from the swabs and was enriched by the whole genome amplification (WGA) technology. Third, the DNA fingerprint was obtained by PCR-denaturing gradient gel electrophoresis (PCR-DGGE) with the V3variable region of16S rRNA gene general primers. The DGGE profiles were compared in diversity analysis and principal component analysis. At the same time, the bands were excised for sequencing, and were compared in gray and occurrence. Results:(1)After WGA enrichment combined with PCR-DGGE, we successfully won the oropharyngeal bacterial flora fingerprint.(2)The oropharynx advantage bacterium group of the Liver cirrhosis complicated with pneumonia group has the highest diversity, followed by liver cirrhosis group and healthy controls.Principal component analysis showed that the liver cirrhosis complicated with pneumonia group, cirrhosis group and healthy controls respectively gathered themselves together, distinguish each other well,people infected with the same pathogenic bacteria gather together.(3) Compared with healthy controls, cirrhosis group have a variation in seven bacterial flora. The level of Campylobacter32.4was reduce, the level of Haemophilus34.5, Streptococcus/Lactobacillus45.5, Selenomonas61.3and Olsenella90.9were increase, the presence of Bulleidia26.5and Streptococcus33.0was increase.Conclusion:These results indicated that the WGA method based on phi29V2DNA polymerase can be used to successfully enrich oropharyngeal mucosal microbiome DNA for DGGE profile analysis. Oropharyngeal bacterial flora of patients with cirrhosis was significantly differed from that of healthy.7bacteria were found to be involved in liver cirrhotic disease, were speculated to be predisposing factors for subsequent lung infection susceptibility. But, this still need to be verified with large samples.