50株麻风菌基因分型的初步研究

Preliminary study on the genotyping of Mycobacterium leprae on 50 isolates from China

目的了解中国麻风菌基因分型图谱,以选择合适麻风病流行病学研究的基因位点。方法从云南省文山州的37例、其他省市的13例麻风患者的皮损活检中,提取麻风菌DNA。以麻风菌基因组内的可变数目的串联重复序列(VNTR)为基因分型的基础,对11个位点进行聚合酶链反应,确定其重复序列的拷贝数。结果中国麻风菌株在GGT5、12-5、21-3、23-3位点的同一性高达 100％;12-5位点仅表现为3个拷贝;AC8、18-8、27-5、rpoT的同一性分别达97％、94％、97％、85％; GTA9、6-7、AC9位点有非常明显的多样性,但GTA9的多样性与其他国家不同。结论中国麻风菌株多数位点的基因型与印度、菲律宾的菌株一致,提示中国麻风菌株与印度和菲律宾的菌株相近; 12-5位点可作为区别中国与周边国家麻风菌株基因分型的标记物;VNTR基因分型方法可应用于麻风病传播链和分布的流行病学研究。

Objective To understand the genotypic mapping of Mycobacterium leprae identified in China and to compare with those from other countries to select suitable alleles for epidemiological investigation in the transmission chain of leprosy. Methods Various number of tandem repeat（VNTR） in genomie DNA of Mycobacterlum leprae was used in the present genotyping study. 33 skin biopsies from Wenshan prefecture, Yunnan province and 17 from other parts of China were studied. DNA extracted from skin biopsies of leprosy patients was subjected to PCR followed by agarose gel analysis and DNA sequencing to determine the number of repeats. Results Loci GGT-5,12-5,21-3 and 23-3 were as highly homogenous as 100% ; The homogeneity of locl AC-8, 18-8, 27-5 and rpoT were 97% , 94%, 97% and 85% respectively. Loci GTA-9,AC-9 and 6-7 showed significant allelic diversity in isolates and the diversity of GTA-9 in Mycobacterium leprae isolated from China was also different from those identified other countries. We had subjected locl GTA 9 and the ten loci to phylogenetic tree analysis respectively. Conclusion The present study revealed that the genotype of Mycobacterium leprae identified from China was close to the strains from the Philippines and India although a few loci were somehow differentiate. Locus 12-5 manifested as only 3 copies in China whereas 4-5 copies predominating in other countries. 12 5 locus might serve as a useful marker to diffrentiate Chinese strains from those in other countries. However, further study on the diversity of GTA-9 was needed in China. The molecular typing of Mycobacterium leprae from different geographic areas might be useful in studying the transmission of leprosy.