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目的分析脓肿分支枝杆菌的embB基因,以探讨其耐乙胺丁醇的分子机制。方法用16S rRNA基因序列分析法鉴定5株脓肿分枝杆菌临床株,测定乙胺丁醇对临床株及标准株(ATCC 19977)的最低抑菌浓度(MIC)。PCR扩增embB基因的全序列,将所测序列进行生物信息学分析。结果乙胺丁醇对5株脓肿分支杆菌标准株和临床株的MIC均为128μg/mL,属高度耐药。从脓肿分支枝杆菌的标准株和临床株均扩增出约3 200 bp片段,与GenBank中脓肿分支枝杆菌标准株的embB基因大小一致。5株临床株与标准株比较,其核苷酸序列存在9个点突变,在突变位点所编码的氨基酸序列中,仅第18位、87位、770位密码子编码与标准株不同的氨基酸。6株脓肿分支枝杆菌的embB基因与对EMB敏感的结核分支杆菌标准株(H37RV)的相应基因序列比较,第303-305位密码子的核苷酸序列存在差异,但仅第303、304位核苷酸编码的氨基酸序列不同,第306位密码子的核苷酸序列无差异。结论脓肿分支杆菌对乙胺丁醇的耐药并非embB基因的突变所致,为embB基因天然存在结构的不同,属于天然耐药。结构的差异与第306位密码子无关,可能与第303、304密码子有关。
Objective To analyze the embB gene of Mycobacterium tuberculosis in abscess to explore the molecular mechanism of its resistance to ethambutol. Methods Five clinical strains of Mycobacterium abscess were identified by 16S rRNA gene sequence analysis. The minimum inhibitory concentration (MIC) of ethambutol to clinical strains and standard strains (ATCC 19977) was determined. The whole sequence of embB gene was amplified by PCR, and the bioinformatics analysis was carried out. Results Ethambutol had a MIC of 128 μg / mL against 5 strains of Mycobacterium abscess and clinical strains, which were highly resistant. The 3 200 bp fragment was amplified from both standard and clinical strains of Mycobacterium abscess, which was consistent with the size of the embB gene of Mycobacterium abscessus in GenBank. Five strains of clinical isolates had 9 point mutations in the nucleotide sequence compared with the standard strains. Only 18th, 87th and 770th codons in the amino acid sequence encoded by the mutation sites encoded amino acids different from the standard strains . The nucleotide sequence of codons 303-305 of 6 strains of Mycobacterium abscess were compared with that of EMB-sensitive standard strain of Mycobacterium tuberculosis (H37RV), but only 303,304 Nucleotide-encoded amino acid sequence is different, the 306th codon nucleotide sequence no difference. Conclusion The resistance to ethambutol in Mycobacterium abscess is not caused by the mutation of embB gene. Because of the different natural structure of embB gene, it belongs to natural resistance. The difference in structure is not related to the 306th codon, which may be related to the 303,304 codon.