Comparative Genomics Analysis Of Gardnerella Vaginalis Epidemic Strains In Shandong Province

Gardnerella vaginalis is a facultative anaerobes of the Bifidobacterium family.Gardnerella vaginalis cells are gram-variable but have Gram-positive cell wall.Gardnerella vaginalis is the main pathogen causing Bacterial vaginalis(BV)and is also associated with infertility,preterm birth,vertebral osteomyelitis,retinitis,and acute coxitis,which may increase the risk of sexually transmitted diseases,including HIV.At present,the pathogenesis of BV is not fully understood.Gardnerella vaginalis forms a biofilm on the vaginal epithelial cells and cannot be cleared by the human immune system or antibiotics,which is the main reason for BV recurrence and difficulty in curing.But Gardnerella vaginalis does not always cause BV.There is a high degree of genetic variability in Gardnerella vaginalis.The large differences in the symptoms of different patients with BV make people speculate that it may be caused by the genetic differences between different strains of Gardnerella vaginalis.Although there is no new association between Gardnerella vaginalis and virulence,there is genomic evidence of a higher prevalence of some genotypes in BV cases compared to clinically excluded BV women.Therefore,studying the genetic differences between different epidemic strains of Gardnerella vaginalis through genomic sequencing and comparative genomic analysis is of great significance for improving the diagnosis and treatment of BV.In this study,Gardnerella vaginalis strains from Shandong Province were isolated from clinical patient samples and 20 strains of Gardnerella vaginalis were obtained.Chaperone sequences can be used in phylogenetic studies and are widely used in the evolution of prokaryotes and eukaryotes.Based on the gene sequence of cpn60(encoding chaperone 60),a phylogenetic tree was constructed.With reference to the published Gardnerella vaginalis,the 20 isolated strains of Gardnerella vaginalis were divided into four subgroups of A,B,C,and D.1 strain belongs to subgroup A,6 strains belong to subgroup B,and 13 strains belong to subgroup C.Vaginolysin is a specific factor for Gardnerella vaginalis and is also a major virulence factor.The phylogenetic tree based on vly(encoding vaginolysin)sequence is divided into four subgroups of 1,2,3 and 4.Ten strains of Gardnerella vaginalis strains were screened for genome sequencing,and their biochemical reactions were tested.Sialidase activity experiments showed that the presence or absence of sialidase activity was not significantly related to the genotype.Drug resistance experiments have found that Gardnerella vaginalis has developed resistance to tetracyclines,nimidazoles,macrolides,and aminoglycoside antibiotics.The measurement of biofilm formation ability showed that the biofilm formation ability of different epidemic strains was quite different.Among them,cpn60 subgroup C was found to have a general medium biofilm formation ability.In this study,10 strains of Gardnerella vaginalis were subjected to genome sequencing and comparative genomics analysis.The genome size of 10 strains of Gardnerella vaginalis was between 1.54-1.75Mb,and the GC content of the genome was about 41%-43%.No plasmid was found,and 4 strains among them predicted 1-2 gene islands.The predicted resistance genes in Gardnerella vaginalis are the macrolide erythromycin resistance gene ermX,the lincosamide antibiotic resistance gene lsaC and the tetracycline resistance genes tetL and tetM,additionally found proteins encoding bleomycin and lanthionin antibiotics.The common virulence factors predicted in Gardnerella vaginalis are mainly related to adhesion,secretion system and iron absorption.However,the virulence factors that JNFY3,JNFY15 and JNFY17 lack are mostly related to bio film formation,which is consistent with their inability to form biofilms.Among 10 strains of Gardnerella vaginalis epidemic strains,1-2 secondary metabolite gene clusters were foun.Except for JNFY3,all predicted T3PKS gene clusters(type ? polyketid synthase),which may play an antibacterial and immune suppression role.This study also compared the differences in pathogenic genes between the epidemic strains in Shandong Province and the American model strain ATCC 14019.The epidemic strains in Shandong Province contain pathogenic factors that ATCC 14019 does not have.They include the Abi protein that may promote competitive exclusion and the TadE protein related to fimbria adhesion.ATCC 14019 encodes a methicillin resistance protein,but no genes encoding such proteins have been found in 10 strains of epidemic strains in Shandong province.Through the genomic sequencing and comparative genomics analysis of the 10 endemic strains of Gardnerella vaginalis obtained from clinical isolation and screening,as well as the comparison with the American model strain ATCC 14019,we have a basic understanding of the genomic characteristics,evolution and pathogenic genes of Gardnerella vaginalis.This has laid a theoretical foundation for our research on the pathogenic mechanism of Gardnerella vaginalis and also provided a basis for the accurate diagnosis and treatment of BV.