Epidemic Characteristics And Homology Analysis Of Clinical Carbapenem-Resistant Klebsiella Pneumoniae

Objective1.To understand the clinical characteristics and epidemic status of carbapenem-resistant Klebsiella pneumoniae in the Affiliated Hospital of Yangzhou Hospital.2.The phenotype of resistance genes and the resistance of the commonly used antibiotics carried by CRKP strain in the hospital were analyzed,and the transfer characteristics of drug-resistant genes were estimated by conjugation transfer experiment,so as to provide a theoretical basis for the prevention and treatment of drug-resistant bacteria.3.To explore the ST phenotype,capsular serotype,drug resistance gene and virulence gene carriage of CRKP in the hospital,so as to improve the understanding from the molecular level,and provide scientific basis and theoretical support for the prevention.Methods1.Sixty non-repetitive CRKP strains isolated from clinical samples from January2019 to June 2021 in the Affiliated Hospital of Yangzhou Hospital were collected.VITEK-2 compact automatic microbiological identification instrument was used for strain identification.The clinical data of the infected patients were consulted,and the sex,age,hospitalization department,specimen type,basic disease,clinical diagnosis,outcome,hospitalization time,invasive operation before specimen collection and antibiotic exposure were analyzed.2.Experimental strains were tested for susceptibility to clinically common antimicrobial agents by the broth microdilution method.3.Polymerase chain reaction was used to detect the phenotype of drug resistance genes carried by CRKP strains,and the plasmid conjugation transfer test of NDM and KPC resistance genes was carried out.4.illumina gene sequencing was used to analyze the ST phenotype,capsule serotype,drug resistance gene and virulence gene carriage of 18 NDM-1 and KPC-2carbapenemase strains.Results1.The main sources of the 60 CRKP strains were sputum and blood,accounting for 80.0%(48 / 60)and 10.0%(6 / 60),respectively,and the main source areas were intensive care medicine and respiratory medicine,accounting for 63.3%(38 / 60)and13.3%(8 / 60),respectively.Most of the infected patients were elderly men over 60 years old.2.Before the samples were collected,51 patients had a history of antibiotic exposure,mainly β-lactamase inhibitors,cephalosporins and carbapenems,relatively less quinolones,and the least aminoglycosides,accounting for 3.7%,45 patients(83.3%)underwent invasive operation.3.In vitro susceptibility results showed that all the experimental strains showed100% resistance to ciprofloxacin,meropenem,gentamicin and tigecycline,and more than 95% resistance to doxycycline,chloramphenicol and aztreonam,but they still had high sensitivity to polymyxin.4.PCR and plasmid conjugation transfer test results: 90.0%(54/60)strains carried KPC-2 gene,and 30.0%(18/60)carried NDM-1 gene.The plasmid conjugation transfer test results showed that the successful rate of plasmid transfer of CRKP strains carrying only KPC-2 type was low,but in 18 CRKP strains with coexisting KPC-2 and NDM-1 carbapenemase genes,16 were positive for the conjugation transfer experiments.5.illumina gene sequencing results: among 18 CRKP strains with KPC-2 and NDM-1 carbapenemase genes coexisting,MLST typing was identified as ST11,and the serum capsule type was KL64,and they all carried β-Lactams(bla CTX-M-65,bla KPC-2),tetracyclines [tet(A)],aminoglycosides(rmt B),sulfa(sul2),trimethoprim(dfr A14),fosfomycin(fos A)and quinolones(Qnr S1)resistance genes,only 1 strain was found to contain β-Lactams(bla ADC-25,bla OXA-23,bla OXA-66),aminoglycosides[aph(3 ’)-Ic,str B,str A],tetracyclines [tet(B)],macrolides [mph(E),msr(E)] drug resistance genes,the main virulence genes carried were yersiniabatin,aerobactin,rmp A,rmp A2.Conclusion1.The CRKP strains isolated in the hospital were mainly derived from ICU and respiratory department,the specimens are mainly sputum and blood,and the infected patients are mainly elderly men aged over 60 years old;antibiotic exposure and invasive treatment may be the main factors for the generation and spread of drug-resistant strains in the hospital.2.18 CRKP strains with coexisting KPC-2 and NDM-1 carbapenemase genes belonged to the ST11 phenotype,and the capsule serotype was KL64,suggesting that ST11-KL64 CRKP is the dominant strain in the hospital.3.The 18 strains of CRKP with coexisting KPC-2 and NDM-1 carbapenemase genes isolated in the hospital carried multiple virulence genes and drug resistance genes,and the combination of multiple virulence phenotypes and drug resistance genes leaded to the high pathogenicity of CR-hv KP.4.The CRKP strains isolated in this hospital had a high resistance rate to common antibiotics.However,the results of in vitro experiments showed that they were sensitive to polymyxin,which could be used as an altenative treatment option for ST11-KL64 CRKP infection.