某院分离的大肠埃希菌和肺炎克雷伯菌ESBLs基因检测及分型

Detection and genotyping of extendedspectrum βlactamases in Escherichia coli and Klebsiella pneumoniae isolated from a hospital

摘要:

目的了解某院分离的大肠埃希菌和肺炎克雷伯菌产生的超广谱β内酰胺酶（ESBLs）基因型。方法采用表型确证试验测定并收集该院产ESBLs大肠埃希菌（40株）和肺炎克雷伯菌（20株），提取质粒DNA。采用特异性引物扩增TEM、SHV和 CTXM系列基因，测序后进行序列分析。结果60株表型确证试验阳性的大肠埃希菌和肺炎克雷伯菌，聚合酶链反应（PCR）扩增均阳性，包括TEM、SHV、CTXM3组和CTXM9组4种基因型。大肠埃希菌中上述4种基因型阳性率分别为37.50%（15株）、2.50%（1株）、62.50%（25株）、50.00%（20株）；肺炎克雷伯菌上述4种基因型阳性率分别为40.00%（8株）、90.00%（18株）、65.00%（13株）、40.00%（8株）。100.00%的大肠埃希菌和80.00%的肺炎克雷伯菌产生blaCTXM，12.50%(5/40)的大肠埃希菌和25.00%(5/20)的肺炎克雷伯菌携带2种CTXM酶基因。23例TEM基因皆为TEM1型；19例SHV型基因包括SHV1型6株、SHV11型6株、SHV12型5株及SHV25型2株，仅SHV12为ESBLs基因，且均来源于肺炎克雷伯菌；66例CTXM型基因，其中CTXM14在大肠埃希菌和肺炎克雷伯菌的检出率分别为45.00%和35.00%，CTXM55检出率均为35.00%，CTXM15检出率分别为20.00%和15.00%，检出少量CTXM3、CTXM65、CTXM101及CTXM123基因型。结论该院分离的大肠埃希菌和肺炎克雷伯菌ESBLs基因型以CTXM 为主，其次为SHV12。CTXM基因型中以CTXM14最为常见，CTXM101及CTXM123型ESBLs为山东省首次检出。

Abstract:

ObjectiveTo investigate genotypes of extendedspectrum βlactamases（ESBLs） produced by Escherichia coli （E. coli） and Klebsiella pneumoniae (K. pneumoniae) isolated from a hospital.MethodsClinical isolates of ESBLsproducing E. coli (n=40) and K. pneumoniae (n=20) were collected according to ESBLs confirmatory test. Plasmid DNA was extracted by alkaline lysis method. PCR analysis was used to amplify TEM, SHV, and CTXM genes, sequence analysis was performed to identify βlactamase genotype.ResultsSixty confirmatory test positive E.coli and K.pneumoniae were positive for PCR amplification, including TEM,SHV, CTXM3 and CTXM9. The positive rate of above four genotypes among E.coli was 37.50%（15 isolates）,2.50%（1 isolate）,62.50%（25 isolates） and 50.00%（20 isolates）respectively; the positive rate of above four genotypes among K.pneumoniae was 40.00%（8 isolates）,90.00%（18 isolates）,65.00%（13 isolates） and 40.00%（8 isolates）respectively. 100.00% of E.coli and 80.00% of K.pneumoniae expressed blaCTXM, 12.50% (5/40) of E.coli and 25.00%(5/20)of K.pneumoniae carried two types of CTXM gene. Twentythree isolates with TEM gene were TEM1;nineteen isolates with SHV gene included 6 isolates of genotype SHV1, 6 of SHV11, 5 of SHV12 and 2 of SHV25，only SHV12 was ESBLs gene,all these types were from K.pneumoniae; 66 isolates were with CTXM gene, the detection rate of CTXM14 among E.coli and K. pneumoniae was 45.00% and 35.00% respectively, CTXM55 were both 35.00%，CTXM15 was 20.00% and 15.00% respectively，CTXM3, CTXM65, CTXM101 and CTXM123 genotypes accounted for fewer percentage of CTXM genotype.ConclusionThe majority of ESBL gene in E. coli and K.pneumoniae in this hospital is CTXM, the next is SHV12. The most prevalent ESBL genotype is CTXM14. CTXM101 and CTXM123 are initially detected in Shandong province.