The present invention relates to a method for detecting and identifying mycobacterial species which comprises steps of amplifying 342 bp of rpoB gene fragments from clinically isolated mycobacterial using mycobacterial rpoB-specific PCR primers; sequencing 306 bp regions of the amplified 342 bp of rpoB gene fragments except the primer regions; and, inferring a phylogenetic tree with reference species. In accordance with the present invention, it was found that rpoB sequences from 44 mycobacterial species provide a basis for systematic phylogenetic relationship which can be used to identify clinically isolated mycobacteria that are pathogenic or potentially so. Accordingly, the amplification of rpoB DNA followed by automated sequencing and the analysis of phylogenetic relationships with the reference species can be used efficiently to detect and identify clinical isolates of mycobacteria which have not been identified by the conventional methods. In particular, this approach is useful for slowly growing, fastidious or uncultivable mycobacteria. Furthermore, in the case of M. tuberculosis, rifampin susceptibility can be simultaneously determined. Thus, the PCR- mediated comparative sequence analysis of rpoB DNA of the invention can be regarded as a reliable and rapid method for the diagnosis of mycobacterial infection.
展开▼
机译:本发明涉及一种检测和鉴定分枝杆菌种类的方法,该方法包括使用分枝杆菌rpoB特异性PCR引物从临床分离的分枝杆菌中扩增342bp的rpoB基因片段的步骤。除引物区外,对扩增的342 bp rpoB基因片段中的306 bp区域进行测序;并用参考物种推断出系统发育树。根据本发明,发现来自44种分枝杆菌属的rpoB序列提供了系统的系统发生关系的基础,其可用于鉴定致病的或潜在地如此的临床分离的分枝杆菌。因此,rpoB DNA的扩增,随后的自动测序以及与参考物种的系统发育关系分析可以有效地用于检测和鉴定尚未通过常规方法鉴定的分枝杆菌的临床分离株。特别是,这种方法对于缓慢生长,挑剔或无法培养的分枝杆菌很有用。此外,在 M的情况下。结核病,利福平敏感性可以同时测定。因此,本发明的rpoB DNA的PCR介导的比较序列分析可以被认为是诊断分枝杆菌感染的可靠且快速的方法。
展开▼