天津地区肺炎患儿肺炎支原体耐药检测及分析

Molecular Detection and Analysis of Resistance in Mycoplasma Pneumoniae in Children Hospitalized with Pneumonia in Tianjin

【摘要】目的了解天津地区住院肺炎患儿肺炎支原体（MP）耐大环内酯类抗菌药物情况及主要的耐药机制。方法收集天津市儿童医院2010年10月一2011年12月确诊为肺炎患儿的肺泡灌洗液标本237份，提取DNA，采用聚合酶链反应（PCR）对其进行肺炎支原体病原检测，从检测阳性标本中按每月随机抽取3—5份共50份进行巢式PCR反应，电泳，在紫外灯下切胶纯化回收产物并测序，测序结果与NCBI已登录的MP标准株M12923SrRNA基因作对比，观察其有无耐药基因位点的突变。结果237份标本中MP阳性率为54．43％（129／237）；50份测序结果显示，其中4份的测序结果与NCBI已登录的MP标准株M12923SrRNA基因序列一致，其余46份均出现2063位A—G突变，未见其他位点的突变，耐药率为92％（46／50）。结论天津地区MP耐大环内酯类药物现状严重，其主要的耐药机制为23srRNAV区产生A2063G点突变。

Objective To study the characterization and molecular mechanisms of macrolide-resistant mycoplasma pneumoniae （MP） in children hospitalized with pneumonia in Tianjin. Methods A total 237 of alveolar lavage fluid speci- mens were collected from children with pneumonia hospitalized in Tianjin children＇s hospital from October 2010 to Decem- ber 2011. The DNA of them was extracted. The genomic fragments of the strains were amplified by using polymerase chain re- action （PCR） method with specific primers. The positive strains were selected randomly to be amplified with nest PCR meth- od. The amplifiable production was electrophoresised and purified and the 23 S rRNA gene sequences were sent to sequenc- ing. The results of sequencing were compared with the sequence of MP M129 to detect the mutations. Results Among 237 specimens, 129 samples were MP positive after detecting by PCR with the primers. The positive rate was 54.43% （129/237）. Of the 50 strains that were selected to sequence, 4 strains and the standard strains were completely same. The other 46 strains showed A-to-G transition at position 2063. There was no other site of mutation. The percentage of macrolide-resistant was 92% （46/50）. Conclusion The macrolide-resistance in mycoplasma pneumoniae is serious in Tianjin. The major molecular mechanism is A-to-G transition at position 2063 of the 23 S rRNA domain V.