半巢式聚合酶链反应在浆膜腔细菌感染诊断中的初步应用  

The Clinical Application of the Semi-Nested Polymerase Chain Reaction in Diagnosis of Bacterial Infection in Serous Cavity

作　　者：周跃[1] 王建民[2] 沈轶瑶[1] 汤伟[1] 顾宇峰[1] 

机构地区：[1]南通大学附属医院感染性疾病科,南通226001 [2]南通大学医学院微生物学教研室,南通226001

出　　处：《热带医学杂志》2007年第11期1073-1075,共3页Journal of Tropical Medicine

摘　　要：目的观察革兰分型半巢式聚合酶链反应(PCR)在浆膜腔细菌感染诊断中的应用价值。方法对30例浆膜腔积液标本,以细菌16SrRNA基因为靶序列,采用一对通用引物和一条革兰阴性菌特异性引物,以半巢式PCR方法进行检测,并同时作细菌培养。结果以通用引物对30例标本作第1次PCR,11例扩增出371bp长度的DNA片段,阳性率为36.7%;对阳性PCR产物再以革兰阴性菌特异性引物作第2次PCR,9例扩增出353bp的DNA片段即为革兰阴性菌。对此30例标本作细菌培养,阳性率为16.7%,低于PCR扩增法。5例标本细菌分离结果与半巢式PCR革兰分型结果相同。结论革兰分型半巢式PCR方法能够灵敏地检测细菌并作出革兰阴性、阳性分型,对于浆膜腔感染的早期诊断具有较好的应用价值。Objective To explore the clinical importance of the semi-nested polymerase chain reaction （seminested PCR） for the detection of bacterium and identification of Gram type in diagnosis of bacterial infection in serous cavity. Methods The method of semi-nested PCR, using a pair of universal primers and a Gram negative type （G-） specific primer targeted at the 16S rRNA gene, was adopted to amplify the DNA of bacterium and to identify its Gram type. 30 clinical specimens obtained from patients＇ serous cavity were simultaneously examined by the semi-nested PCR method and bacterial culture method. Results At the first PCR, using the universal primers, 371 bp DNA fragments were amplified from 11 cases out of 30 samples, with a positive rate of 36.7%. At the second PCR, adopting the positive products of the first PCR as the templates, 353 bp DNA fragments were amplified from 9 cases and all were identified as G^- bacterial infection. The positive rate detected by bacterial culture method was 16.7% and was lower than that of the semi-nested PCR method. The Gram typing results of bacteria of 5 samples detected by the method of bacterial culture were the same as that detected by the method of the semi-nested PCR. Conclusion The method of the semi-nested PCR method is more sensitive than the bacterial culture method. This diagnosis method is rapid and can identify the Gram type of bacteria at the same time, so the method is very suitable for clinical diagnosis of bacterial infection of serous cavity.

关 键 词：细菌感染 半巢式聚合酶链反应 16SRRNA基因 细菌培养 浆膜腔 

分 类 号：Q555[生物学—生物化学]