布鲁氏菌利福平耐药株rpoB基因突变特征分析

Mutation characteristics of rpoB gene in rifampicin-resistant Brucella strains

目的分析布鲁氏菌利福平耐药株rpoB基因的突变特征。方法选取分离自新疆维吾尔自治区的4株布鲁氏菌利福平耐药株(JSY-26、G-9、WSY-13、AW-3株)DNA,采用PCR扩增利福平rpoB基因,并对核苷酸序列进行测序。以布鲁氏菌利福平耐药标准株(RB51株)和利福平敏感株(ALT-8株)的rpoB基因序列为参照,应用Mega 7.0软件比对分析4株布鲁氏菌利福平耐药株rpoB基因利福平耐药决定区(rifampicin resistance determination region,RRDR)内外的碱基突变位点及类型。结果经序列比对分析,JSY-26和WSY-13株均在rpoB基因RRDR 1576 bp处发生单碱基点突变,碱基由鸟嘌呤(guanine,G)突变为腺嘌呤(adenine,A);G-9株在rpoB基因RRDR 1606 bp处发生单碱基点突变,碱基由胞嘧啶(cytosine,C)突变为A。AW-3株在rpoB基因RRDR外2536、2537、2626、2636、2654 bp处出现3种类型共5处突变,分别为插入突变3处[胸腺嘧啶(thymine,T)插入1次、C插入2次],缺失突变1处(C缺失),单碱基点突变1处(由G突变为C)。结论布鲁氏菌利福平耐药株rpoB基因RRDR突变以单碱基点突变为主,RRDR外则出现多位点插入和缺失突变。

Objective To analyze the mutation characteristics of rpoB gene in rifampicin-resistant Brucella strains.Methods DNA of 4 rifampicin-resistant Brucella strains(JSY-26,G-9,WSY-13 and AW-3)isolated from Xinjiang Uygur Autonomous Region was selected,rifampicin rpoB gene was amplified by PCR and its nucleotide sequence was sequenced.The rpoB gene sequences of rifampicin-resistant Brucella standard strain(RB51)and sensitive strain(ALT-8)were used as reference,the mutation sites and types of the rpoB gene inside and outside the rifampicin resistance determination region(RRDR)of the 4 rifampicin-resistant Brucella strains were analyzed by Mega 7.0 software.Results Through sequence alignment,both JSY-26 and WSY-13 strains underwent a single base point mutation at the RRDR 1576 bp of the rpoB gene,with the base changing from guanine(G)to adenine(A).The G-9 strain underwent a single base point mutation at the RRDR 1606 bp of the rpoB gene,with the base changing from cytosine(C)to A.The AW-3 strain showed 5 mutations of 3 types outside rpoB gene RRDR at 2536,2537,2626,2636 and 2654 bp,namely 3 insertion mutations[thymine(T)insertion once and C insertion twice],1 deletion mutation(C deletion),and 1 single base point mutation(from G to C mutation).Conclusion The RRDR mutations in the rpoB gene of the rifampicin-resistant Brucella strains are mainly characterized by single base point mutations,while multiple insertion and deletion mutations occur outside the RRDR.