鱼腥草素钠对肺炎克雷伯菌耐药性和毒力的降低作用及其机制  

Reduction of drug resistance and virulence of Klebsiella pneumoniae by sodium houttuyfonate and its mechanisms

作　　者：赵赟安 赵一铮 沈琪 陈娇丽 赵芯米 武蓉珍 赵志钢 黄建胜 ZHAO Yun-an;ZHAO Yi-zheng;SHEN Qi;CHEN Jiao-li;ZHAO Xin-mi;WU Rong-zhen;ZHAO Zhi-gang;HUANG Jian-sheng(The Fifth Affiliated Hospital of Wenzhou Medical University,Lishui,Zhejiang 323000,China)

机构地区：[1]温州医科大学附属第五医院检验科,浙江丽水323000 [2]温州医科大学附属第五医院药学部,浙江丽水323000 [3]温州医科大学附属第五医院中医科,浙江丽水323000

出　　处：《中华医院感染学杂志》2024年第19期2887-2893,共7页Chinese Journal of Nosocomiology

基　　金：浙江省医药卫生科技计划基金资助项目(2023KY1368);浙江省基础公益研究计划省自然科学基金项目(LHDMY24H190001);丽水市重点研发计划基金资助项目(2020ZDYF11)。

摘　　要：目的研究鱼腥草素钠(SH)对肺炎克雷伯菌耐药性和毒力的影响。方法收集2021-2022年温州医科大学附属第五医院临床分离的肺炎克雷伯菌。采用聚合酶链式反应(PCR)检测碳青霉烯类耐药性肺炎克雷伯菌(CRKP)的耐药基因、多位点序列分型(MLST)明确ST型、微量肉汤稀释法分析菌株对亚胺培南(IPM)的最低抑菌浓度(MIC_(IPM)),综合耐药基因、ST分型和耐药表型结果筛选不同类型CRKP;PCR检测黏液表型调控基因(rpmA、rpmA2)、气杆菌素合成基因(iucA)、沙门菌素合成基因(iroB)和荚膜分型基因(wzi),筛选不同荚膜分型的高毒力肺炎克雷伯菌(hvKP);K-B纸片法联合微量肉汤稀释法分析SH作用前后CRKP对IPM的耐药性变化。血清杀菌实验和离心实验分析SH作用前后hvKP毒力及黏液产量变化。逆转录定量PCR(qRT-PCR)分析碳青霉烯耐药基因(bla_(KPC)、bla_(NDM))和毒力基因(galF)的mRNA表达变化。结果10株CRKP作为耐药性实验菌株,具有高中低三种耐药表型,9种ST型,携带有blaKPC-2,blaNDM-1,blaNDM-5不同耐药基因;7株不同荚膜分型hvKP联合标准hvKP(NUTH-K2044)作为毒力实验菌株;微量肉汤稀释法结果显示,SH作用后CRKP菌株的MICIPM降低至原来的1/4~1/2,K-B纸片法显示抑菌圈明显扩大;血清杀菌实验结果表明SH作用后hvKP在血清中的存活率降低(P<0.05),离心实验发现SH能抑制hvKP菌株黏液的产生;qRT-PCR定量结果证明SH能抑制bla_(KPC)、bla_(NDM)、galF基因mRNA的表达(P<0.05)。结论SH通过抑制耐药基因、毒力基因表达和黏液的合成,进而降低肺炎克雷伯菌的耐药性和毒力。OBJECTIVE To investigate the effects of Sodium houttuyfonate(SH)on drug resistance and virulence of Klebsiella pneumoniae.METHODS Klebsiella pneumoniae clinically isolated from the Fifth Affiliated Hospital of Wenzhou Medical University during 2021 to 2022 were collected.The drug resistance genes of carbapenem resistant Klebsiella pneumoniae(CRKP)was detected by PCR,the ST type was identified by multilocus sequence typing(MLST),the minimum inhibitory concentration(MIC)of the strain to imipenem(IPM)was analyzed by micro-broth dilution method,and different types of CRKP were screened by combining the results of drug resistance genes,ST typing and drug resistance phenotypes.PCR was used to detect mucus phenotype regulatory genes(rpmA and rpm A2),bacteriocin synthesis gene(iucA),Salmonellin synthesis genes(iroB),and capsule typing gene(wzi),and the highly virulent Klebsiella pneumoniae(hvKP)with different capsule typing were screened.K-B disk method combined with micro broth dilution method was used to analyze the changes in resistance of CRKP to IPM before and after SH treatment.The changes in the virulence and mucus production of hvKP before and after the action of SH were analyzed by serum sterilization experiments and centrifugation experiments.Reverse transcription quantitative PCR(qRT-PCR)was performed to analyze the changes in mRNA expression of carbapenem resistance genes(bla kPc,bla NDM)and virulence gene(galF).RESULTS Ten strains of CRKP were used as drug-resistant experimental strains,with high,medium and low three drug resistance phenotypes,nine ST type,carrying bla kc-2,bla NDM-1,bla NDM-5 different drug resistance genes.Seven strains with different capsule subtypes of hvKP combination with standard hvKP(NUTH-K2044)were used as virulence test strains.The results of the micro broth dilution method showed that after SH treatment,MIC of CRKP strain on IPM decreased to 1/4-1/2 of the original level,and the K-B disk method showed a significant expansion of the inhibitory zone.The results of serum bactericidal

关 键 词：鱼腥草素钠 肺炎克雷伯菌 耐药性 毒力 

分 类 号：R378[医药卫生—病原生物学]