The Prevalence Of The Antibiotic Resistance Genes And Virulence Determinants Among Clinical Enterococci In Baotou,Inner Mongolia

Objective The aim of this study is to investigate the distribution and resistance of Enterococci isolated from clinical specimens in Baotou,Inner Mongolia.We also discussed the epidemiological characteristics of these isolates,which provides a reference for the standardized clinical use of antimicrobial drugs and prevents the emergence and spread of resistant Enterococci.Additionally,we further investigate the resistant mechanism of Enterococcus by detecting the distribution of resistance genes and virulence determinants.Method Screening for resistance to antimicrobial drugs among 117 enterococcal clinical isolates was performed by the agar-screening method.Out of the 117 enterococcal isolates,50 were selected for further detecting the distribution of resistance genes(aac(6’)-Ie-aph(2’’)-Ia、aph(2’’)-Ib、aph(2’’)-Ic、aph(2’’)-Id、aph(3’)-IIIa、ant(6’)-Ia、erm(B)、MazEF、vanA、vanB、tet(L))and virulence determinants(cylA、gelE、esp、 ace、agg、 efaA)using polymerase chain reaction(PCR)-based methods.Furthermore,a PCR method was used to detect the presence of drug resistant genes PBP5、gyrA and parC,then sequencing of the PCR products was performed.Results(1)Analysis of drug resistance: The clinical isolates of enterococci were resistant to 9 types of antibiotics used in clinical practice.We found that more than half of the strains were resistant to three or more antibiotics.Our results also indicated the resistance rate to be higher in E.faecium than E.faecalis.(2)Distribution of pathogenic gene: We detected cylA、gelE、esp、ace、agg genes in 18%,28%,16%,2% and 0% isolates tested.Our findings also found that 92.9% E.faecalis isolates harbored efaA gene.(3)Distribution of resistance gene: The prevalence of AME gene among the tested strains with high-level gentamicin resistance was as follows:aac(6’)-Ie-aph(2’’)-Ia(89.3%)、aph(2’’)-Ib(0)、aph(2’’)-Ic(7%),aph(2’’)-Id(10.7%)、aph(3’)-IIIa(23.3%).In addition,the prevalence of AME gene among the tested strains with high-level streptomycin resistance was as follows: aph(3’)-IIIa(53.3%)、ant(6’)-Ia(66.7%).Out of enterococcal isolates that were erythromycin resistance in our study,83.3% isolates were found to carry ermB gene.Moreover,73.91% and 4.27% isolates were found to carry tet(L)and MazEF genes among isolates with tetracycline resistance and vancomycin resistance,respectively.(4)Mutation analysis of resistance gene:we found that all E.faecium isolates withpenicillin resistance harbored PBP5 gene.We also found two novel amino acid mutations in the C-terminal region of PBP5,which are largely responsible for binding to penicillin.In addition,87.5% and 91.30% isolates with fluoroquinolone antibiotic resistance carried ParC and gyrA genes,which were further sequenced and compared with reference sequences obtained from GenBank.Conclusions(1)The clinical isolates of enterococci in Baotou is resistant to nine antibacterial agents used in clinical practice in different degrees.Some multiple drug-resistant strains resist even more drugs.Also,different antimicrobial resistances pattern was observed.(2)A significant correlation was found between resistance genes and virulence determinants with antibiotic resistance among the clinical isolates of enterococci.(3)Our results showed that important amino acid changes in PBP5,gyrA and parC located at the binding cavity are largely responsible for binding of ligands.