RAPD分析在Hp菌株鉴定中的应用  

Application of PCR based RAPD in differential diagnosis of strains of H.pylori

作　　者：王蔚虹[1] 阎赞华[1] 胡伏莲[1] 

机构地区：[1]北京医科大学第一医院消化内科

出　　处：《中华微生物学和免疫学杂志》1998年第6期434-436,共3页Chinese Journal of Microbiology and Immunology

基　　金：教委博士点专项基金

摘　　要：目的建立一种快速准确地鉴定不同幽门螺杆菌（Ｈｐ）菌株的ＲＡＰＤ分析方法，并评价它在区分Ｈｐ复发和再感染中的应用价值。方法用苯酚－氯仿方法提取ＨｐＤＮＡ，以一个１０ｎｔ的寡核苷酸引物，建立ＨｐＰＣＲ扩增体系，所得ＰＣＲ产物以２％琼脂糖凝胶电泳分析（ＲＡＰＤ分析）。同一菌株验证方法的重复性，对４例十二指肠溃疡伴Ｈｐ感染者三联治疗前后及根除后一年内再现的Ｈｐ菌株进行ＲＡＰＤ分析，鉴别复发和再感染。结果（１）同一Ｈｐ菌株于不同时间进行的２次ＲＡＰＤ分析，所得的ＰＣＲ产物完全一致，说明该方法具有良好的可重复性。（２）５０株不同的Ｈｐ菌株各产生不同的ＲＡＰＤ图谱，可按此图谱的不同将它们区分开来。（３）对４例十二指肠溃疡伴Ｈｐ感染治疗前及治疗后１年再现的菌株的ＲＡＰＤ分析，发现３例患者的菌株具有相同的ＲＡＰＤ图谱，证实为原菌株的复发，而１例患者的菌株则产生不同的ＲＡＰＤ图谱，推测为新菌株的再感染，４例患者随访１年均为十二指肠溃疡复发。结论ＲＡＰＤ分析只需单一的随机引物，方法简单、快速、经济、重复性好，可用于区分不同来源的Ｈｐ菌株。Objective To establish a rapid and sensitive polymerase chain reaction (PCR)based random amplified polymorphisc DNA (RAPD) fingerprinting method to distinguish different H.pylori isolates and evaluate its use in distinguishing recrudecence from reinfection of H.pylori after eradication. Methods H.pylori DNA was extracted and used for PCR based RAPD fingerprinting with a 10 nucleotide arbirary primer. The reproducibility and sensitivity of RAPD patterns was tested in 50 different clinical strains.Strains which isolated from 4 duodenal ulcer patients with H.pylori infection before treated with triple therapy and one year after eradication were tested with this method to confirm recrudecence or reinfection. Results RAPD fingerprinting yielded 10 to 20 main bands on the agarose gels.The profiles of the PCR products from the same isolates at two different times were indistinguishable. Fifty different clinical strains yielded different RAPD profile. By comparing 4 pairs of isolates before and after treatment,we confirmed that 3 patients with same RAPD profile belonged to recrudecence, and l patient with different RAPD profiles belonged to reinfection. Conclusions PCR based RAPD fingerprinting which only needs a single primer is a rapid, simple and reproducible method in distinguishing different H.pylori strains, especially in distinguishing recrudecence from reinfection of H.pylori .

关 键 词：幽门螺杆菌 多态现象 DNA 

分 类 号：R372[医药卫生—病原生物学] R378.99[医药卫生—基础医学]