DNA转化和共培养研究淋球菌对大观霉素的耐药机制

Studying the mechanism of spectinomycin resistance in Neisseria gonorrhoeae using DNA transformation and co-culture

目的通过DNA转化和共培养研究淋球菌(NG)对大观霉素的耐药机制。方法将3株大观霉素耐药环丙沙星敏感的供体NG[大观霉素最小抑菌浓度(MIC)512 mg/L]与大观霉素敏感环丙沙星耐药的受体NG共培养,并将耐药菌株的DNA转化至大观霉素敏感的受体菌中。分别将供体菌、受体菌及共培养和DNA转化得到的NG的16sRNA基因扩增并测序,分析可能导致大观霉素耐药的基因突变。结果共培养后和DNA转化得到耐大观霉素的NG株(大观霉素MIC 512 mg/L),PCR扩增和DNA测序分析发现所有大观霉素耐药的NG均发现16sRNA基因胞嘧啶(C)1192胸腺嘧啶(T)突变,大观霉素敏感的受体菌未发现突变。结论共培养和DNA转化可以用于NG耐药机制研究,NG16SrRNA基因C1192T突变导致大观霉素对NG耐药。

Objective To study the mechanism of spectinomycin resistance in Neisseria gonorrhoeae by virtue of DNA transformation and co-culture.Methods 3 Neisseria gonorrhoeae stains resistant to spectinomycin（MIC=512 mg/L） and sensitive to ciprofloxacin as the donors were co-cultured with the strain as the receptor sensitive to spectinomycin and resistant to ciprofloxacin.DNA from 3 strains resistant to spectinomycin was transformed into the recipient strain sensitive to spectinomycin respectively.16SrRNA gene from all strains was amplified by polymerase chain reaction and was sequenced.Results New strains were acquired resistant to spectinomycin（MIC=512 mg/L） after co-culture and DNA transformation.PCR amplication and DNA sequencing showed that 16SrRNA gene C（cytosine）1192T（thymine） mutation occurred in all strains resistant to spectinomycin including donor strains and newly acquired strains.Mutation gene was transformed into recipient strain successfully.Conclusion DNA transformation and co-culture may be used to study resistant mechanism in Neisseria gonorrhoeae.16SrRNA gene C1192T mutation can lead to resistance to spectinomycin in Neisseria gonorrhoeae.