海南发现4株产NDM-1多重耐药的肺炎克雷伯菌

目的分离和鉴定海南地区临床上发现的耐碳青霉烯类药物的肺炎克雷伯菌携带新德里金属β-内酰胺酶-1（NDM-1）基因的细菌。方法使用法国梅里埃API20E生化鉴定条和VITEK2全自动细菌鉴定仪及其配套的GNS试剂，鉴定出临床上耐亚胺培南和美洛培南的肺炎克雷伯菌；并经改良Hodge试验和亚胺培南，亚胺培南＋EDTA纸片法协同试验检测产金属酶类碳青霉烯酶，对目的菌株进行NDM-1耐药基因PCR特异性扩增，扩增后产物进行琼脂糖凝胶电泳和测序鉴定；同时将目的菌株进行质粒接合转移试验分析其耐药机制，并对接合前后的药敏结果进行比较分析。结果从13株耐亚胺培南或美洛培南的肺炎克雷伯菌中，经亚胺培南，亚胺培南＋EDTA纸片法协同试验检出4株阳性，确定为产金属酶类碳青霉烯酶细菌。该4株产金属酶的肺炎克雷伯菌的改良Hodge试验均为阴性，NDM-1基因PCR扩增产物经琼脂糖凝胶电泳和测序分析，确认此4株目标菌株均为携带NDM-1的耐药基因的耐药菌株；这4株接合子E．coli J53（NDM-1）对所有β-内酰胺类药物的MIC值较受体菌E．coli J53耐药性均有很大提高，经PCR扩增其接合子NDM-1基因片段分析证实4株接合子均接合成功。结论海南发现的4株耐碳青霉烯酶类的肺炎克雷伯菌为携带blaNDM-1基因，且其NDM-1耐药基因可通过质粒在不同菌株之间传递。

Isolation and identification of four multiresistant Klebsiella pneumonia strains producing NDM-1 in Hainan province of China.

Objective To isolate and identify reduced carbapenem susceptible Khbsiella pneumonia with NDM-1 in Hainan. Method The KlebsieUa pneumonia resistant to imiperem and meropenem were identified by using BioMerieux API 20E identification system and automatic VITEK-2 dentification system. The modified Hedge test and EDTA-disk synergy test were used to confirm the ability of hydrolyzing carbapenemase, and the NDM-1 gene was confirmed by PCR and DNA sequence analysis. After sequenced,the bacteria with NDM-1 gene were took to the plasmid DNA preparations and conjugation experiment were used to determine the mechanism of drug resistance. Results Among 13iImiperem or meropenem-resistant Khbsiella pneumonia,4 were positive in IP/IP＋EDTA-disk synergy test but negative in Modified Hedge Test. All of the 4 strains carrying NDM-1 gene could be probed by PCR, and the plasmids in the 4 strains were transferred into the drug resistance to β-lactams could be enhanced by NDM-1 transferred through E.Coli J53. Conclusion The susceptibility of four Klebsiella pneumonia strains with NDM-1 gene in Hainan to carbapenem were isolated and NDM-1 gene can be transferred through plasmids and all of 4 strains possesse a special plasmid which can be transferred into the E. coli J53 by Conjunction Trail.