Study On The Correlation Between Vaginal Microbiome Diversity And High Risk HPV Infection And Cervical Disease

Part I: Correlation of between vaginal microbiome diversity and high-risk HPV infectionObjective: To investigate the correlation between vaginal microbiome diversity and cervical high-risk HPV(HR-HPV)infection.Methods: Select healthy female at the physical examination center of Renji Hospital of Shanghai Jiaotong University School of Medicine or community health centre.Based on the status of cervical HR-HPV infection,divide them into normal group and infection group,and collect vaginal secretions,then use bacteria 16 S rRNA v3-v4 region gene to amplification with PCR and second-generation high-through sequencing to analysis vaginal microorganisms,using bioinformatics analysis methods to study the vaginal microbiota with ? diversity,? diversity,and use LEfSe and LDA discriminant analysis to detect biomarkers species for each group.Results: 68 samples in the normal group and 75 in the infected group have been analyzed,lactobacilli were the predominant species in both groups.Compared with the normal group,the number of lactobacilli in the infected group decreased but there was no significant difference(p <0.05),Bacillus,Shigella,Prevotella,Giantococcus,Cilia and other species increased in infected groups,and the differences were statistically significant;compared with the normal group,the species richness indecies such as Sobs index,ACE index were increased,the species diversity index Shannon index increased,Simpson index decreased in the infected group,the difference was statistically significant;Based on the weighted-unifrac distance algorithm to calculate PCoA,we can see the species distribution is more dispersed in the infected group compared to normal group;Using LEfSe and LDA discriminant analysis to detect the characteristic species in each group,Bacillus,Chi Herobacterium and marine bacilli can be used as 4biomarker species of the infected group,and Lactobacillus,Gardnerella Vaginal were the normal group biomarkers.Conclusion: 1 Lactobacillus was the predominant specie in both high-risk HPV infection group and normal control group.Compared to normal group,Lactobacillus was reduced in infected group and Bacillus,Prevotella,Megasococcus increased.2 Species diversity indices and richness indices increased in infected group,and the difference was statistically significant.3 Bacillus,Shigella,and ocean bacilli can be regarded as biomarkers species in the infected group.Part II: Study on the vaginal microbiota between HPV16 / 18 infection group and other high-risk HPV infection groupObjective: Analyze composition characters of the vaginal microbiota of HPV16 / 18 infection group and other high-risk HPV infection group,and explore the causes of the difference in their pathogenicity in terms of vaginal microbiota.Methods: 84 adult women who were examined at the medical center of Renji Hospital affiliated to School of Medicine,Shanghai Jiaotong University or from the community healthy center were enrolled in this study.According to the results of their cervical HPV infection,they were divided into HPV16 / 18 infection group and other high-risk HPV infection group,Vaginal secretions were collected and their vaginal microbiota composition was analyzed,bacterial 16 S rRNA v3-v4 region gene were amplificated in PCR and next-generation sequencing technology were used to define the compositional characteristics of vaginalmicroecology between the two groupsResults: 35 samples were enrolled in HPV16 / 18 infection group and 49 samples in other high-risk HPV infection group.Both groups had the dominant bacterium of Lactobacillus,and the strains of Prevotella,Bacillus,Streptococcus,and other Gram-negative bacteria were detected in both groups,the composition of the flora were no significant difference between the two groups,diversity index(Chao index,Ace index)and richness index(Shannon index,Simpson index)were similar in the two groups,and there were no difference in Statistical significance between the two groups.UnrecordedMethylococci bacteria can be regarded as biomarkers for HPV16 / 18 infection by LEfSe and LDA analysis.conclusion: 1 In both HPV16 / 18 infection group and other high-risk HPV infection group,lacticebllus are dominant in the vaginal microbiota,and both groups detected Prevotella,Bacillus,Streptococcus,Macrobacterium and other anaerobic bacteria and Gram-negative bacteria.2 There was no significant difference in species composition,species diversity indecies and richness indecies between HPV16 / 18 and other high-risk HPV infection groups,the conclusion may needs to be further verificated in larger sample size3 Methylococcus aureus Unrecorded bacteria can be used as a marker of HPV16 / 18 infection.Part III: Correlation between vaginal microbiota and different stages of cervical diseaseObjective: To analyze the vaginal microbiota distribution and community structure of women with cervical lesions at different stages,to studythe dynamic changes during the development of cervical lesions and to explore the relationship between the dynamic changes of vaginal flora and the progression of cervical lesions.Methods: The adult female vaginal secretion samples from the cervical clinics or physical examination centers of Renji Hospital Affiliated to Shanghai Jiao Tong University were collected for cervical HPV test,cervical TCT test and vaginal biopsy when necessary.The samples of vaginal secretions were collected and sequenced.According to the above tests results,68 cases of Normal group,51 cases of LSIL group,23 cases of HSIL group,10 cases of cervical cancer group were enrolled,vaginal secretions samples were extracted total DNA,bacterial 16 SrRNA V3-V4 region used for PCR and next generation sequencing technology Illumina Miseq platform were used to detect the deep sequences of microbiota,bioinformatics methods were used to study the vaginal microbiological composition of each sample,species diversity index(Chao index,Ace index)and richness index(Shannon index,Simpson index)were used to anlysis the species diversity and richness of each group,The statistical differences of species composition of each group was detected,and the biomarker species of each group was analyzed by LEfSe and LDA methods.Result:Lactobacillus was the predominant species in each group,but with the progress of cervical disease,the proportion of lactobacilli gradually decreased from 40.7% in the normal group to 18.1% in the cancer group,while the proportion of Bacillus and other anaerobic bacteria gradually increased.With the progress of cervical disease,vaginal microbiota diversity indices and richness indices significantly increased,but in LSIL and HSIL group,the difference between the two groups was not statistically significant.There was no significant species in the LSIL group by LEfSe and LDA analysis,butPrevotella and Megasphaera were biomarkers of the HSIL group,while in the cancer group,Bacillus ?Sneathia?Fusobacterium?Acidovorax?Porphyromonas and Oceanobacillus were regarded as biomarker speciesConclusion: 1 vaginal microbiota was the dominant in Lactobacillus,with a relatively low diversty co exist with other microorganisms,and lactobacillus iners was the main lactbacillus in this region.2 With the progress of cervical disease,vaginal microbiota showed a decline in the proportion of lactbacillus,anaerobic bacteroa and gram-negative bacteria increased,and microbial diversity and richness showed an upward trend.3 Prevotella,Megasphaera were HSIL group biomarker species,whereas in the cancer group,Bacillus? Sneathia? Fusobacterium? Acidovorax? Porphyromonas ?Oceanobacillus were biomarker species...