临床分离CRKP耐药和毒力基因检测及分子进化分析

 目的 了解临床耐碳青霉烯类肺炎克雷伯菌(CRKP)的耐药和毒力基因携带情况,为临床防治提供依据。方法 收集某院2020年3月—2021年3月临床标本分离的CRKP 36株,对菌株进行药敏鉴定,采用聚合酶链反应(PCR)扩增检测耐药基因、荚膜血清型基因、毒力基因,采用多位点序列分型(MLST)方法对菌株进行序列分型(ST分型),基于wzi测序结果进行血清型分型和分子进化分析。结果 36株CRKP均检出bla_KPC基因,未检出bla_IMP、bla_VIM、bla_OXA-48基因。黏液丝试验均为阴性,未检出K1、K2、K5、K20、K57五种常见荚膜血清型,36株CRKP rmpA2、wcaG、ybtS、aerobactin、iutA、iroN、ycf、mrkD、mrkA、silS、uge、Plvpk、fimH、wzi基因检出率为100%;TerW为86.11%;fimA、magA均为阴性。MLST结果分析显示,36株均为ST11型。仅32株成功测序wzi基因,wzi分型结果为K14.K64 96.88%(31/32),K24 3.12%(1/32)。基于测序结果构建分子进化树,结果显示32株菌中31株100%同源,1株与浙江菌株KP18069 99%同源。结论 该院CRKP对碳青霉烯类耐药的主要机制是携带bla_KPC基因,优势ST分型为ST11,wzi分型以K14.K64为主,且携带了大量的毒力基因。分子进化树提示存在同源性感染,怀疑有输入性传播,应及时采取防控措施,防止耐碳青霉烯类高毒力肺炎克雷伯菌医院传播。

Antimicrobial resistance,virulence genes and molecular evolution of clinically isolated CRKP

Objective To understand antimicrobial resistance and virulence gene carrying of clinically isolated carbapenem-resistant Klebsiella pneumoniae (CRKP), so as to provide basis for clinical prevention and treatment.Methods 36 strains of clinically isolated CRKP in a hospital from March 2020 to March 2021 were collected, antimicrobial susceptibility identification of strains were performed, antimicrobial resistance genes, capsular serotype genes and virulence genes were detected by polymerase chain reaction (PCR) amplification, sequence typing (ST) was performed by multilocus sequence typing, serotyping and molecular evolutionary tree analysis were performed based on wzi sequencing results.Results 36 CRKP strains were all detected bla_KPC gene, but bla_IMP, bla_VIM, and bla_OXA-48 genes were not found. String test was negative, four common capsule serotypes K1, K2, K5, K20, and K57 were not found, detection rates of rmpA2, wcaG, ybtS, aerobactin, iutA, iroN, ycf, mrkD, mrkA, silS, uge, Plvpk, fimH, and wzi genes of 36 CRKP strains were 100%; TerW was 86.11%; fimA and magA were both negative. MLST analysis showed that 36 strains were all ST11, only 32 strains were successfully sequenced wzi gene. wzi typing results were K14. K64 96.88% (31/32) and K24 3.12% (1/32). Molecular evolutionary tree analysis based on sequencing results showed that 31 of 32 strains were 100% homologous, and 1 strain was 99% homologous with Zhejiang strain KP18069.Conclusion The main mechanism of resistance of CRKP to carbapenems in this hospital is bla_KPC gene, the dominant ST is ST11, and wzi type is mainly K14.K64, which carries a large number of virulence genes. Molecular evolutionary tree indicates that there is homologous infection, and it is suspected that there is import transmission. Prevention and control measures should be taken in time to prevent the transmission of carbapenem-resistant hypervirulent Klebsiella pneumoniae (CR-hvKP) in the hospital.