2010-2011年复发和治疗抵抗麻风患者麻风杆菌耐药基因特征监测

Detection of drug-resistance associated mutations in patients with recurring or treatment.resistant leprosy during 2010-2011

目的探讨2010-2011年度复发和治疗抵抗麻风患者皮损中麻风杆菌对氨苯砜、利福平和氧氟沙星的耐药基因突变情况。方法收集2010-2011年由各省麻风防治机构诊断并上报的治疗抵抗及复发麻风患者的基本资料及皮损组织标本，提取患者皮损标本中的麻风杆菌DNA，对靶序列folP1、rpoB、gyrA3个片段经PCR扩增后测序比对，寻找突变位点。结果24例麻风患者中，11例为治疗抵抗患者，13例为复发患者。21例患者扩增的3个片段均为阳性，3例患者皮损内菌folP1片段有点突变，均为53：acc（苏氨酸Thr）→gcc（丙氨酸Ala），其中2例为治疗抵抗患者，1例为复发患者；1例复发患者皮损内菌rpoB片段有错义突变，为441：gat（天冬氨酸Asp）→aac（天冬酰胺Asn）；未发现有gyrA片段的错义突变。结论复发和治疗抵抗的麻风患者皮损中存在与氨苯砜和利福平耐药相关的folp1、rpoB片段突变菌，未发现与氧氟沙星耐药相关的gyrA片段突变菌。

Objective To detect gene mutations associated with dapsone-, rifampicin- and ofloxacin- resistance in lesions of patients with recurring or treatment-resistant leprosy collected from 2010 to 2011. Methods Clinical data and lesional specimens were collected during 2010-2011 from patients with recurring or treatment-resistant leprosy who were diagnosed and reported by provincial centers for leprosy control. Mycobacterium leprae DNA was extracted from the specimens and subjected to PCR for the amplification of folP1, rpoB and gyrA genes. The PCR products were directly sequenced and BLAST program was used to compare the sequence of isolated strains with the reference sequence in GenBank. Results Twenty-four patients were enrolled in this study, including 13 with recurring leprosy and 11 with treatment-resistant leprosy. Twenty-one patients showed positive PCR results in all the three regions. Of these PCR-positive specimens, 3 from 1 patient with recurring and 2 patients with resistant leprosy harbored a point mutation, aec （threonine）→ gcc （alanine）, at codon 53 in the floP1 gene, 1 from a patient with recurring leprosy harbored a missense mutation, gat （aspartic acid） → aac （asparagine）, at eodon 441 in the rpoB gene. Conclusions Mutations are detected in the folP1 and rpoB genes, which are associated with the resistance to dapsone and rifampicin respectively, but not in the ofloxaein resistance-associated gyrA gene, in Mycobacterium leprae isolates from patients with recurring or treatment-resistant leprosy.