血流感染高致病性肺炎克雷伯菌毒力因子及分子分型特点分析
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摘要
目的研究血流感染高致病性肺炎克雷伯菌的毒力基因和基因分型特点。方法采用PCR方法检测临床分离株血流感染高致病性肺炎克雷伯菌的高毒力基因,并进行荚膜血清型和ST分型;采用微量肉汤稀释法通过BD Phoenix全自动细菌鉴定/药敏系统对分离菌株进行药敏试验;应用加克拉维酸复合药(头孢他啶/克拉维酸或头孢噻肟/克拉维酸)与单药(头孢噻肟或头孢他啶)的药敏纸片组合进行肺炎克雷伯菌产ESBLs的表型确证试验。结果 115株血流感染肺炎克雷伯菌临床分离株分为高毒力肺炎克雷伯菌(Hvkp,占43.48%,50/115)和非Hvkp(non-HvKP,占56.52%,65/115)。与non-HvKP相比,HvKP更容易表现为高黏液性表型(44.00%VS 6.15%,P<0.01),且普遍携带rmpA(98.00%VS 4.61%,P<0.01)、rmpA2(50.00%VS 3.08%,P<0.01)、wcag(24.00%VS 2.15%,P<0.05)毒力基因。HvKP常为K1、K2荚膜血清型,non-HvKP常为K-nontypable血清型。本组血流感染肺炎克雷伯菌主要流行ST型别为ST23、ST65、ST37,其中Hvkp流行型为ST23、ST65和ST86。Hvkp对常用抗生素的敏感性好于non-HvKP,其中发现产ESBLs耐药hvKP菌株4株。结论血液感染高毒力肺炎克雷伯菌易表现为高黏液性表型且普遍携带rmpA毒力基因,因此应注意检测肺炎克雷伯菌黏液性状、荚膜血清型、毒力基因以及ST分型,以利于hvkp感染的识别。
Objectives To study the virulence genes and genotyping characteristics of hypermucoviscous Klebsiella pneumoniae isolated from patients with a bloodstream infection. Methods The virulence genes,capsular serotypes,and sequence types(STs)of K.pneumoniae strains were detected with PCR.A drug susceptibility test was performed with the BD Phoenix automated bacterial identification/susceptibility system using the broth dilution method.A drug sensitivity test was performed with a drug combination including clavulanic acid(ceftazidime + clavulanic acid or cefotaxime +clavulanic acid)and a single drug(cefotaxime or ceftazidime),and a confirmation test was performed to determine the phenotypes of ESBL-producing strains of K.pneumoniae. Results One hundred and fifteen strains of K.pneumoniae causing a bloodstream infection were divided into hypervirulent(Hv)K.pneumoniae(43.48%,50/115)and non-Hv K.pneumoniae(56.52%,65/115).Compared to non-Hv K.pneumoniae,Hv K.pneumoniae was more likely to exhibit a highly mucinous phenotype(44.00% vs 6.15%,P<0.01)and carry the virulence factors rmpA(98.00% vs.4.61%,P<0.01),rmpA2(50.00% vs.3.08 %,P<0.01),and wcag(24.00% vs.2.15%,P<0.05).Hv K.pneumoniaeusually expresses the K1 and K2 capsular serotypes while non-Hv K.pneumoniae expresses serotypes that are non-Ktypeable.In this study,the main STs of K.pneumoniae were ST23,ST65,and ST37,and the prevalent STs of Hv K.pneumoniae were ST23,ST65,and ST86.The sensitivity of Hv K.pneumoniae to antibiotics was significantly better than that of non-Hv K.pneumoniae,but ESBL-resistant Hv K.pneumoniae strains were found. Conclusion Laboratories should pay attention to determination of mucus characteristics and capsular serotypes,virulence genes,and STs of K.pneumoniae to assist clinicians in identifying an infection with Hv K.pneumoniae.
Objectives To study the virulence genes and genotyping characteristics of hypermucoviscous Klebsiella pneumoniae isolated from patients with a bloodstream infection. Methods The virulence genes,capsular serotypes,and sequence types(STs)of K.pneumoniae strains were detected with PCR.A drug susceptibility test was performed with the BD Phoenix automated bacterial identification/susceptibility system using the broth dilution method.A drug sensitivity test was performed with a drug combination including clavulanic acid(ceftazidime + clavulanic acid or cefotaxime +clavulanic acid)and a single drug(cefotaxime or ceftazidime),and a confirmation test was performed to determine the phenotypes of ESBL-producing strains of K.pneumoniae. Results One hundred and fifteen strains of K.pneumoniae causing a bloodstream infection were divided into hypervirulent(Hv)K.pneumoniae(43.48%,50/115)and non-Hv K.pneumoniae(56.52%,65/115).Compared to non-Hv K.pneumoniae,Hv K.pneumoniae was more likely to exhibit a highly mucinous phenotype(44.00% vs 6.15%,P<0.01)and carry the virulence factors rmpA(98.00% vs.4.61%,P<0.01),rmpA2(50.00% vs.3.08 %,P<0.01),and wcag(24.00% vs.2.15%,P<0.05).Hv K.pneumoniaeusually expresses the K1 and K2 capsular serotypes while non-Hv K.pneumoniae expresses serotypes that are non-Ktypeable.In this study,the main STs of K.pneumoniae were ST23,ST65,and ST37,and the prevalent STs of Hv K.pneumoniae were ST23,ST65,and ST86.The sensitivity of Hv K.pneumoniae to antibiotics was significantly better than that of non-Hv K.pneumoniae,but ESBL-resistant Hv K.pneumoniae strains were found. Conclusion Laboratories should pay attention to determination of mucus characteristics and capsular serotypes,virulence genes,and STs of K.pneumoniae to assist clinicians in identifying an infection with Hv K.pneumoniae.
引文
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[3]Shon AS,Russo TA.Hypervirulent Klebsiella pneumoniae:the next superbug?[J].Future Microbiol,2012,7(6):669.
[4]Diancourt L,Passet V,Verhoef J,et al.Multilocus sequence typing of Klebsiella pneumoniae nosocomial isolates[J].J Clin Microbiol,2005(43):4178-82.
[5]Candan ED,Aksoz N.Klebsiella pneumoniae:characteristics of carbapenem resistance and virulence factors[J].Acta Biochim Pol,2015(62):867.
[6]Li W,Sun GZ,Yu YH,et al.Increasing occurrence of antimicrobial-resistant hypervirulent(hypermucoviscous)Klebsiella pneumoniae isolates in China[J].Clin Infect Dis,2014(58):225-32.
[7]Yu WL,Lee MF,Chen CC,et al.Impacts of hypervirulence determinants on clinical features and outcomes of bacteremia caused by extended-spectrumβ-lactamase-producing Klebsiella pneumoniae[J].Microb Drug Resist,2017,23(3):376-83.
[8]Russo TA,Olson R,Macdonald U,et al.Aerobactin mediates virulence and accounts for increased siderophore production under iron-limiting conditions by hypervirulent(hypermucoviscous)Klebsiella pneumoniae[J].Infecti Immun,2014,82(6):2356.
[9]Russo TA,Shon AS,Beanan JM,et al.Hypervirulent K.Pneumoniae secretes more and more active iron-acquisition molecules than"Classical"K.Pneumoniae thereby enhancing its virulence[J].Plos One,2011,6(10):e26734.
[10]Enani MA,El-Khizzi NA.Community acquired Klebsiella pneumoniae,K1serotype.Invasive liver abscess with bacteremia and endophthalmitis[J].Saudi Med J,2012,33(7):782-6.
[11]Lin WH,Kao CY,Yang DC,et al.Clinical and microbiological characteristics of Klebsiella pneumoniae from community-acquired recurrent urinary tract infections[J].Eur J Clin Microbiol Infect Dis,2014,33(9):1533-9.
[12]Jung SW,Chae HJ,Park YJ,et al.Microbiological and clinical characteristics of bacteraemia caused by the hypermucoviscosity phenotype of Klebsiella pneumoniae in Korea[J].Epidemiol Infect,2013,141(2):334-40.
[13]Lin JC,Chang FY.The Distant metastasis of pyogenic liver abscess caused by klebsiella pneumoniae serotype K2and the underlying disease of diabetes mellitus should be carefully interpreted[J].Clin Infect Dis,2007,45(11):1531.
[14]Wu JH,Wu AM,Tsai CG,et al.Contribution of fucose-containing capsules in Klebsiella pneumoniae to bacterial virulence in mice[J].Exp Biol Med,2008(233):64-70.
[15]和晋渝.肺炎克雷伯菌的血清分型及毒力基因分布的研究[D].重庆:重庆医科大学,2012.
[16]Shon AS,Bajwa RP,Russo TA.Hypervirulent(hypermucoviscous)Klebsiella pneumoniae:A new and dangerous breed[J].Virulence,2013,4(2):107-18.
[17]Qu TT,Zhou JC,Jiang Y,et al.Clinical and microbiological characteristics of Klebsiella pneumoniae liver abscess in East China[J].BMC Infect Dis,2015(15):161.
[18]张改,黄德海,靳静,等.肺炎克雷伯菌噬菌体LH-02的生物学特性及基因组初步研究[J].中国病原生物学杂志,2016,11(1):9-12.
[19]Wang Q,Li B,Tsang AK,et al.Genotypic analysis of Klebsiella pneumoniae isolates in a Beijing hospital reveals high genetic diversity and clonal population structure of drug-resistant isolates[J].PLoS One,2013(8):e57091.