用PCR-RFLP技术检测皮肤感染性肉芽肿组织中的分支杆菌

Detection and identification of Mycobacteria with polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP)from patients with Mycobacterial skin infections

目的 :探讨用聚酶链反应 -限制性片段长度多态性分析 (PCR -RFLP)方法快速检测皮肤感染性肉芽肿组织中的分支杆菌。方法 :皮肤感染性肉芽肿患者的组织标本 5份 ,提取组织DNA进行巢式PCR扩增 ,对PCR产物分别用HhaⅠ、MboⅠ、BstUⅠ 3种限制性内切酶进行酶切 ,聚丙烯酰胺凝胶电泳 ,进行RFLP分析 ,并与培养结果进行比较。结果 :从 2份临床诊断为游泳池肉芽肿的组织标本中检测出海分支杆菌 ,从 1份诊断为皮肤感染性肉芽肿的标本中检测出结核分支杆菌 ,此 3例结果与培养鉴定结果完全一致。另两份标本分别检测出结核分支杆菌和偶发分支杆菌。结论 :用PCR -RFLP方法可以快速鉴定皮肤感染性肉芽肿中的分支杆菌 ,可协助临床进行疾病的早期诊断和及时治疗。

Objective: To establish a rapid approach to the detection and identification of Mycobacteria from lesions of patients with suspected Mycobacterial infections. Methods: Specimens were obtained from five patients suspected to have Mycobacterial infections. DNA extracted from clinical samples was amplified by nested PCR. The PCR products were digested with Hha Ⅰ, MboⅠ, and BstUⅠ restriction enzymes and applied to PAGE. The species of Mycobacteria were determined by restriction fragment length polymorphism (RFLP) analysis. Identification of Mycobacteria culture was also performed in 3 patients. Results: M. marinum was found in two patients diagnosed as swimming pool granuloma. M. tuberculosis was found in one patient diagnosed as infectious skin granuloma. All these 3 Mycobacteria were confirmed by Mycobacteria culture. A strain of M. tuberculosis and a strain of M. fortuitum were detected in remain two patients. Conclusion: The results above indicate that PCR-RFLP analysis is rapid and reliable in detection and identification of different Mycobacteria species from skin tissues. Application of this method will be helpful for early diagnosis and treatment of Mycobacteria skin infections.