Kamal Raj, Dayal R, Katoch V M, Katoch K
National JALMA Institute for Leprosy and Other Mycobacterial Diseases (ICMR), Tajganj, Agra, India.
Lepr Rev. 2006 Jun;77(2):141-6.
Nucleic acid sequences of Mycobacterium leprae were detected using gene probes hybridizing with targeting ribosomal RNA (16S rRNA), ribosomal DNA (16S rDNA) and gene amplification techniques (PCR) in skin lesion of paediatric leprosy patients and the effect of treatment on the by these methods. Eighty paediatric leprosy patients were included in the study. Most cases (79%) were between 9 and 16 years of age. Cases were divided into three groups according to treatment status, viz. untreated (30), undergoing treatment (30), and at the end of treatment (20). Clinical and slit smear examination for acid fast bacilli (AFB) was performed and nucleic acids were extracted and fractionated from skin biopsies. M. leprae specific 16S rRNA and 16S rDNA was detected by hybridization with gene probes whereas the 36 kDa gene sequence was detected by a gene amplification assay (PCR). The cases were classified as paucibacillary (PB) and multibacillary (MB) by the standard criteria of WHO (1988). Positivity of 16S rRNA in PB cases decreased from 60% in untreated to 10.5% after 4-8 months of treatment whereas for 16S r DNA, it decreased from 50% to 21%, for PCR from 70% to 36.8% for the same specimen, and all became negative at 1 year. Similar trends were seen in MB cases where positivity in smear positive untreated cases decreased from 100% to 56.2% with 16S rRNA and 42.8% with 16S rDNA and PCR, respectively, after 9-12 months of treatment and all became negative at 2 years, except one case which remained positive with PCR. Similar results were observed in smear negative MB cases, 100% positivity detected by 16S r RNA and PCR, 75% detected by 16S rDNA decreased to zero after 9-12 months of therapy. This study suggests the potential usefulness of gene probes targeting 16S rRNA and 16S rDNA and PCR as supportive molecular tools for diagnosis of smear negative evolving MB disease and also monitoring the response to treatment, these observations however, needs to be validated in prospective follow up studies.
利用与靶向核糖体RNA（16S rRNA）、核糖体DNA（16S rDNA）杂交的基因探针以及基因扩增技术（PCR），检测小儿麻风病患者皮肤病变中麻风分枝杆菌的核酸序列，并观察这些方法对治疗效果的影响。本研究纳入了80例小儿麻风病患者。大多数病例（79%）年龄在9至16岁之间。根据治疗状态将病例分为三组，即未治疗组（30例）、正在治疗组（30例）和治疗结束组（20例）。进行了临床检查和抗酸杆菌（AFB）涂片检查，并从皮肤活检组织中提取和分离核酸。通过与基因探针杂交检测麻风分枝杆菌特异性16S rRNA和16S rDNA，而通过基因扩增试验（PCR）检测36 kDa基因序列。根据世界卫生组织（1988年）的标准将病例分为少菌型（PB）和多菌型（MB）。PB病例中16S rRNA的阳性率从未治疗时的60%降至治疗4至8个月后的10.5%，而16S rDNA的阳性率从50%降至21%，同一标本的PCR阳性率从70%降至36.8%，且在1年时全部转阴。MB病例也观察到类似趋势，涂片阳性的未治疗病例在治疗9至12个月后，16S rRNA的阳性率从100%降至56.2%，16S rDNA和PCR的阳性率分别降至42.8%，除1例PCR仍为阳性外，所有病例在2年时均转阴。在涂片阴性的MB病例中也观察到类似结果，16S rRNA和PCR检测的阳性率为100%，16S rDNA检测的阳性率为75%，治疗9至12个月后降至零。本研究表明，靶向16S rRNA和16S rDNA的基因探针以及PCR作为支持性分子工具，对于诊断涂片阴性的进展性MB疾病以及监测治疗反应具有潜在用途，然而，这些观察结果需要在前瞻性随访研究中得到验证。
