Wang Ya-Juan, Xiong Li-Kuan, Feng Ping
Center of Infection Disease, West China Hospital, Sichuan University, Chengdu 610041, China.
Sichuan Da Xue Xue Bao Yi Xue Ban. 2009 Nov;40(6):1130-4.
To evaluate the PCR-reverse line dot hybridization assay (RLB) based on 16S-23S rDNA internal transcribed spacer (ITS) sequence for detection and identification of mycobacterium species.
This study was performed in five different centers simultaneously, and the performance of PCR-RLB was estimated and verified by detection of 60 reference strains belonging to 50 mycobacterium species, 10 nonmycobacterial speices and 383 clinical isolates identified by immunochromatographic assay (ICA) combined sequencing and gas chromatography (GC).
The genus-specific probe hybridizied with the amplification of PCR of all mycobacteria and species-specific probes hybridized only with corresponding species, however, neither of them hybridized with the amplification of PCR of nonmycobacterial speices. Compared with ICA combined sequencing and GC, accuracy of PCR-RLB was nearly 100%, and PCR-RLB identified 380 of 383 in species-level successfully, including 11 mixed clinical isolates which could not be identified with other assays correctly.
The mycobacterial PCR- RLB based on 16S-23S rDNA ITS is rapid, convenient, sensitive and specific, and practical, and it is promising to be applied in clinical diagnosis.
评估基于16S - 23S rDNA内转录间隔区（ITS）序列的聚合酶链反应-反向线点杂交分析（RLB）用于分枝杆菌菌种检测和鉴定的效果。
本研究在五个不同中心同时进行，通过检测属于50种分枝杆菌菌种、10种非分枝杆菌菌种的60株参考菌株以及通过免疫层析分析（ICA）联合测序和气相色谱（GC）鉴定的383株临床分离株来评估和验证PCR - RLB的性能。
属特异性探针与所有分枝杆菌的PCR扩增产物杂交，种特异性探针仅与相应菌种杂交，然而，它们均不与非分枝杆菌菌种的PCR扩增产物杂交。与ICA联合测序和GC相比，PCR - RLB的准确率接近100%，且PCR - RLB成功在种水平鉴定了383株中的380株，包括11株其他检测方法无法正确鉴定的混合临床分离株。
基于16S - 23S rDNA ITS的分枝杆菌PCR - RLB快速、便捷、灵敏、特异且实用，有望应用于临床诊断。
