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目的了解重庆地区儿童结核病中结核分枝杆菌临床分离株的数目可变串联重复序列(variable numbertandem repeats,VNTR)的分布特征,寻找适合的VNTR位点组合。方法采用多位点串联重复序列(multiple locus VNTRanalysis,MLVA)分型方法,选择标化的24个VNTR位点,对101例结核分枝杆菌临床分离株DNA进行检测,结果采用BioNumerics 6.1数据库软件进行聚类分析和单位点Hunter-Gaston分辨率指数(Hunter-Gaston index,HGI)分析,并比较分析国际推荐的分组(12、15、24位点)的基因分型鉴定能力。结果 MLVA分析结果显示24个VNTR位点在不同菌株中存在明显的多态性,101例结核分枝杆菌临床分离株可分为1个基因群83种基因型,其中69种基因型只有1个菌株,占68.32%(69/101),另有32株临床分离株表现出14种基因型,占31.68%(32/101),成簇率为17.82%;24个VNTR位点的HGI具有较大差异(0.168～0.829),HGI能达到0.5以上的VNTR位点数有16个;24个VNTR位点进行不同的位点组合:12、15个和24个位点组合的HGI分别为0.995、0.996、0.996。结论 重庆地区儿童结核病中结核分枝杆菌存在基因多态性,国际推荐的15个VNTR位点组合的MLVA分型方法适用于其流行病学的研究。
Objective To understand the distribution of variable numbertandem repeats (VNTRs) in clinical isolates of tuberculosis in children with tuberculosis in Chongqing and to find a suitable combination of VNTR loci. Methods Multiple locus VNTRanalysis (MLVA) typing method was used to detect the DNA of 101 clinical isolates of Mycobacterium tuberculosis from 24 VNTR loci. The results were analyzed using BioNumerics 6.1 database software Cluster analysis and Hunter-Gaston index (HGI) analysis, and compared the genotyping ability of the internationally recommended group (12,15,24). Results The results of MLVA analysis showed that there were obvious polymorphisms in 24 VNTR loci in different strains. 101 clinical isolates of Mycobacterium tuberculosis were divided into 83 genotypes of 1 gene group, of which 69 genotypes had only one (68.32%, 69/101). Another 32 isolates showed 14 genotypes, accounting for 31.68% (32/101) with clustering rate of 17.82%. The HGI of 24 VNTR loci (0.168-0.829). There were 16 VNTR loci with HGI up to 0.5. 24 VNTR loci were grouped in different loci. The HGI of 12, 15 and 24 loci were 0.995 and 0.996, respectively , 0.996. Conclusion The genotypes of Mycobacterium tuberculosis in children with tuberculosis in Chongqing are polymorphic. The MLVA genotyping method of 15 internationally recommended combinations of VNTR loci is suitable for its epidemiological study.