广州地区革兰阴性杆菌CTX-M和OXA型广谱β-内酰胺酶基因分型研究

目的调查广州地区临床分离的革兰阴性杆菌中CTX-M型超广谱β-内酰胺酶(ESBLs)和OXA型广谱β-内酰胺酶(beta-lactamase,Bla)的主要基因型别及其流行情况.方法按照NCCLS 2001年标准筛选广州地区临床分离菌株的ESBLs表型;用聚合酶链反应(PCR)扩增法和DNA测序法进行ESBLs基因序列分析.结果PCR扩增结果显示,CTX-M1、CTX-M2、CTX-M9群和OXA的总阳性率在本地区临床分离的临床检测ESBLs阳性的革兰阴性杆菌中分别为9.96%、0、35.5%和1.6%,同时检出≥两种ESBLs基因的菌株数64株,阳性率为5.9%;序列分析进一步证实了CTX-M型ESBLs的具体型别,包括CTX-M-3、CTX-M-22、CTX-M-9、CTX-M-14、CTX-M-17、CTX-M-18、CTX-M-21、CTX-M-24、TOHO-2和TOHO-3,其比例分别为3.23%、3.7%、4.99%、3.32%、2.21%、3.69%、2.95%、4.43%、1.85%和1.48%;其中以CTX-M-9和CTX-M-24阳性率较高;在本地区只检出1种OXA型Bla-OXA-2/PSE-2(1.38%,15/1084);另外,还检出了8株无法具体归类的CTX-M-9型ESBLs,占0.74%;CTX-M型基因主要分布于大肠埃希菌和肺炎克雷伯菌中,分别占42.8%和36.3%,而OXA基因则主要分布于铜绿假单胞菌中,占80%.结论本地区CTX-M类ESBLs也较为常见,其中尤以CTX-M-9和CTX-M-24为主,暂无CTX-M2群ESBLs,可能存在1种或多种新的CTX-M型ESBLs.

Genotypes of CTX-M-like-extended-spectrum and OXA-like-spectrum Beta-lactamase in Clinical Isolates of Gram-negative Bacilli in Guangzhou

OBJECTIVE To investigate the main genotypes and the epidemic characteristics of CTX-M-like-extended-spectrum beta-lactamases(ESBLs) and OXA-like-spectrum beta-lactamases(Bla) of the common Gram-negative bacilli in Guangzhou. METHODS The ESBLs phenotype of 3500 clinical isolates was primarily screened and confirmed by the NCCLS phenotypic confirmatory test according to guidelines of NCCLS(2001).PCR was performed on ESBLs phenotype-positive clinical isolates using primer specific for CTX-M1,CTX-M2, CTX-M9 and OXA,respectively,and the PCR products were purified and sequenced by ABI prism 377 DNA sequencer. RESULTS The positive rates of PCR results for CTX-M1,CTX-M2,CTX-M9,OXA,and two or more of them in all the ESBLs-phenotype-positive clinical Gram-negative isolates were 9.96%(108/1084),0,35.5%(385/1085),1.6%(17/1085),and 5.9%(64/1084),respectively.The PCR products for CTX-M1 and CTX-M9 were further identified as CTX-M-3(3.23%),CTX-M-22(3.7%),CTX-M-9(4.99%),CTX-M-14(3.32%),CTX-M-17(2.21%),CTXM-18(3.69%),CTX-M-21(2.95%),CTX-M-24(4.43%),TOHO-2(1.85%),and TOHO-3(1.48%).The prevalence of CTX-M-9 and CTX-M-24 was the highest in Guangzhou.All PCR products for OXA were further identified as OXA-2/PSE-2 gene(1.38%,15/1084).Besides above 8 strains which producing an unclassified CTX-M-like gene were detected in the present study(0.74%,8/1084).Among them,42.8%(151/353) of CTX-M-producing strains were Escherichia coli,36.3%(128/353) of CTX-M-producing strains were Klebsiella pneumoniae,while 80%(12/15) of OXA-producing strains were Pseudomonas aeruginosa. CONCLUSIONS CTX-M-like ESBLs,especially CTX-M-9 and CTX-M-24,are the prevalent genotypes of ESBLs of clinical Gram-negative bacilli in Guangzhou.But the CTX-M2-like ESBLs could not be detected in our city.Finally,one or more kinds of novel CTX-M-like ESBLs might be existed in Guangzhou.