多药耐药铜绿假单胞菌β-内酰胺酶基因与转座子、整合子遗传标记研究

β-Lactamase and Genetic Markers of Transposon and Integron in Multidrug Resistant Pseudomonas aeruginosa

目的研究铜绿假单胞菌连续分离株的β-内酰胺酶基因、孔膜蛋白oprD2基因及整合子和转座子介导的各种耐药基因的分布状况。方法纸片扩散法测定铜绿假单胞菌对17种抗菌药物的敏感性,采用PCR方法检测20株铜绿假单胞菌β-内酰胺酶编码基因、孔膜蛋白oprD2基因、Tn21/Tn501型转座子编码基因merA、Ⅰ类整合子编码基因qacEΔ1-sul1;采用PCR直接全自动荧光法进行阳性基因测序。结果20株铜绿假单胞菌检出TEM、OXA-2群、OXA-10群、CARB 4种β-内酰胺酶基因,未检出质粒型AmpC酶和金属β-内酰胺酶基因;膜孔蛋白编码基因oprD2均为缺失型,Tn21/Tn501型转座子遗传标记MerA阳性7株(35.0%),Ⅰ类整合子遗传标记qacEΔ1-sul1阳性10株(50.0%);2号株作OXA-2群阳性基因测序,与OXA-21型接近,但仍存在2个氨基酸序列差别,可确认为新亚型。结论多药耐药铜绿假单胞菌β-内酰胺酶类抗菌药物的耐药主要与TEM、OXA、CARB型耐药基因有关,整合子与转座子参与了铜绿假单胞菌的耐药和多药耐药,并发现1种铜绿假单胞菌OXA基因新亚型。

OBJECTIVE To explore the distribution of β-lactamase and drug resistant gene mediated by transposon and integron in continuous isolates of Pseudornonas aeruginosa. METHODS The antibiotics susceptibility was tested by K-B method; the β-lactamase and the outer membrane protein gene oprD2 were detected by polymerase chain reaction（PCR） ; the genotype of merA encoding Tn21/Tn501 type trandsposon and qacE△1-sul1 encoding Ⅰ type integron was detected by PCR and the positive genes were amplified and sequenced by PCR fluorescence spectrophotometry. RESULTS The TEM type, OXA-2, OXA-10 and CARB of β-lactamase genes had been detected in 20 strains of P. aeruginosa, but plasmid-mediated ampC enzyme and metallo-β-lactamase had not been detected, the gene oprD2 encoding porins was not detected. The gene merA was detected in 7 strains（35.0%）, and the qacE△-sul1 had been detected in 10 strains（50.0%）. The gene OXA-2 in the isolate No. 2 was sequenced, and translated into amino acid, and the translated amino acid sequence was compared with GenBank, the results indicated that amino acid sequence of OXA in the isolate No. 2 was simlar to that in GenBank, exsited 2 different amino acid sequences, so was confirmed as a new subtype. CONCLUSIONS Resistance to β-lactamase compounds in P. aeruginosa of our hospital is related to TEM, OXA and CARB genes, and integron and transposon contribute to the drug resistance and multidrug resistance in P. aeruginosa.