产ESBLs和AmpC肺炎克雷伯菌医院感染分子流行病学研究

A molecular epidemiology study on nosocomial infection caused by ESBLs and AmpC-producing klebsiella pneumoniae

目的:研究调查临床分离的120株非重复肺炎克雷伯菌中产超广谱β-内酰胺酶(Extended spectrumβ-lactamases,ESBLs)和头孢菌素酶(Ampler class Cβ-lactamase,AmpC)菌株的发生率,以及ESBLs和AmpC的表型及基因型。方法:采用美国国家临床实验室标准委员会规定的ESBLs表型,筛选和确证ESBLs的发生率。以分析性等电聚焦、接合实验确定ESBLs和AmpC菌株中酶的表型。以质粒为模板,用β-内酰胺酶通用引物、blaTEM、blaSHV及blaAmpC作PCR扩增,并以染色体为模板,用blaAmpC为引物作PCR扩增以确定酶的基因型。结果:120株非重复的肺炎克雷伯菌中,产β-内酰胺酶的有93株(77.5%),其中15株产ESBLs(12.5%),9株产AmpC(7.5%)。医院感染ESBLs阳性率(25.5%)明显高于院外感染株(3.7%),所有产AmpC菌株均来自院内感染。所有菌株均对亚胺培南敏感。AmpC阳性菌株对β-内酰胺抗生素的耐药率高于AmpC阴性菌株。产ESBLs或AmpC的18株菌中有9株的编码基因可通过接合实验而转移。结论:肺炎克雷伯菌大多为产β-内酰胺酶菌株,其中产ESBLs和AmpC酶的频率较高,并常导致院内感染;其耐药质粒可通过接合转移给敏感菌,导致耐药性传播。

Objective：To study the prevalence, phenotypes and genotypes of ESBLs and AmpC-producing 120 nonrepetitive clinical isolates of K.pneumoniae. Methods： The occurrence of phenotype producing ESBLs was screened and determined according to the National Committee for Clinical Laboratory Standards （NCCLS）. The phenotypes of β -lactamase were studied by isolectric focusing and conjugation methods. The plasmid and genomic DNA extracted from the bacteria were used to determine genotype of the β-lactamases by PCR with bla, blaTEM, blaSHV and blaAmpC-specific primers. Results：The study showed that among 120 isolates, the β -lactamaseproducing strains, ESBLs-producing and AmpC-producing strains were 93 （77.5%）, 15 （ 12.5% ） and 9 （7.5%） isolates, respectively. The ESBLs-produeing strains were much more frequently found in nosoeomial isolates （25.5%） than that in community （3.7%）. And all AmpC-produeing strains were found in nosocomial isolates. None of them was resistant to imipenam. Furthermore, AmpC-producing strains were found to be more resistant to antibiotics than AmpC negative strains. The encoding genes could be transferred by conjugation and were found in 9 of 18 resistant strains producing both ESBLs and AmpC .Conclusion：A high incidence of ESBLs and AmpC -producing strains existed in K.pneumoniae. These β-lactamase producing strains had been the main pathogens of nosocomial infection. Otherwise, that the gene encoding ESBLs and AmpC could be transferred to susceptible strains by conjugation leading to the resistance spreading abroad.