荚膜型和非荚膜型流感嗜血杆菌双重PCR检测方法的建立和临床应用  

Establishment and clinical application of double polymerase chain reaction for capsulate and non-capsulate Haemophilus influenzae

作　　者：赵瑛 王荣山 朱文珠 金洪星 金秀萍 范兴丽[4] 

机构地区：[1]绍兴市柯桥区中医医院检验科,浙江绍兴312030 [2]义乌市妇幼保健医院,浙江义乌322000 [3]绍兴市柯桥区中医医院儿科,浙江绍兴312030 [4]杭州医学院基础医学部,浙江杭州310053

出　　处：《中国卫生检验杂志》2017年第21期3102-3105,共4页Chinese Journal of Health Laboratory Technology

基　　金：浙江省科技计划项目(2015C33265);浙江省教育厅一般科研项目(Y201738453)

摘　　要：目的建立基于流感嗜血杆菌(Haemophilus influenzae,Hi)外膜蛋白P6编码基因omp6和荚膜编码基因bex A的双重PCR检测方法,并确定其检测限、灵敏度和特异度。方法根据Gen Bank公布流感嗜血杆菌的omp6和bex A基因核苷酸序列设计引物,用Hi标准菌株确定其检测限,双重PCR检测289例鼻窦炎患者鼻咽部分泌物(NPS)确定方法的灵敏度与特异度。结果 289例NPS培养阳性率为50.5%,14株为荚膜型;双重PCR检测omp6的阳性率为54.7%,bex A阳性16株。双重PCR与细菌培养及血清凝集试验符合率分别为95.8%和99.3%。检测时模板的下限为3.8 pg/μl。双重PCR检测omp6和bex A基因的灵敏度均为100.0%,特异度分别为91.6%和99.3%。结论建立的双重PCR是一种快速、特异和敏感的Hi感染的检测和分型方法,可代替细菌培养和血清凝集试验用于临床标本诊断。Objective To establish a double polymerase chain reaction（ PCR） method based on the outer membrane omp6 gene and capsule gene bex A of H. influenzae,and to master its detection limit,sensitivity and specificity. Methods According to the omp6 and bex A gene sequence of H. influenzae published by Gen Bank to design primers,and H. influenzae standard strains was used to verify the detection limit,and double PCR was used to detect NPS in 289 specimens for further evaluating the sensitivity and specificity. Results In 289 NPS specimens,the positive rate of H. influenzae were 50. 5% based on bacteria culture,including 14 capsulate H. influenzae; positive rate of H. influenzae was 54. 7% by double PCR,including 16 capsulate H. influenza. The coincident rates for double-PCR and the slide agglutination test was 95. 8% and 99. 3%,respectively. The lowest limit of detection was 3. 8 pg/μl. The sensitivity in the detection of omp6-PCR and bex A-PCR was both 100. 0%,and the specificity was 91. 6% and 99. 3%,respectively. Conclusion The established double PCR is rapid,specific,sensitive,and can replace the bacteria culture and serum agglutination test to diagnose clinical specimens.

关 键 词：流感嗜血杆菌 鼻窦炎 鼻咽部分泌物 双重聚合酶链式反应 细菌培养 

分 类 号：R378[医药卫生—病原生物学]