女性患者解脲脲原体抗四环素耐药基因回复突变的研究

Study on the back mutation of Ureaplasma urealyticum anti-tetracycline resistance gene in female patients

目的了解女性解脲脲原体的感染率,分析其中解脲脲原体对四环素耐药性的回复突变的情况及可能原因。方法对解脲脲原体临床株分离并进行药敏实验,统计感染率。使用试剂盒提取四环素敏感株的基因组DNA,并通过PCR和琼脂糖凝胶电泳检测tetM耐药基因片段,了解其中解脲脲原体临床株回复突变的现象;对携tetM耐药基因的四环素回复突变株tet(M)基因克隆并测序,使用Blast与Genbank标准序列(U08812.1)进行对比分析,探索回复突变的主要位点。结果解脲脲原体检出率为24.81%,其中对四环素的耐药率为9.23%,四环素敏感株中tetM耐药基因的携带率为42.37%(n=25),tetM基因全长及前导肽序列测序的三个样品与标准序列对比后,确认其在前导序列均存在不同程度的位点突变。结论解脲脲原体的四环素敏感株携带有tetM的耐药基因,该基因前导序列的突变可能是其四环素耐药回复突变的原因之一,探索耐药基因对临床抗生素使用具有一定指导意义。

Objective To understand the infection rate of Ureaplasma urealyticum in women,and analyze the reversion mutation of Ureaplasma urealyticum resistance to tetracycline and explore the possible mutational situations and reasons.Methods Ureaplasma urealyticum were isolated and the infection rate was calculated by susceptibility test.Genomic DNA of tetracycline sensitive strains was extracted by kits,and tetM drug-resistant gene fragments were detected by PCR and agarose gel electrophoresis to understand the recovery mutation of the clinical strain of Ureaplasma urealyticum.The whole gene of tetM was cloned and sequenced with tetM-resistant genes.Blast and Genbank standard sequences(U08812.1) was used for comparative analysis.Results The detection rate of Ureaplasma urealyticum in this experiment was 24.81%;among which,the drug resistance rate to tetracycline was 9.23%,and the carrying rate of tetM resistant gene in tetracycline sensitive strain was 42.37%(n=25).Total length and leader peptide sequence of three samples were sequenced.And standard contrast after confirm its leading sequences had different degrees of point mutations.Conclusions Tetracycline sensitive strains of Ureaplasma urealyticum in carried tetM resistance genes,and the mutation of the leading sequence of this gene might be one of the reasons for the reverse mutation of tetM resistance.The exploration of drug resistance genes has a certain guiding significance for the clinical use of antibiotics.