Identification of 54 mycobacterial species by PCR-restriction fragment length polymorphism analysis of the hsp65 gene

A total of 121 reference and clinical strains of both slowly and rapidly gr owing mycobacteria belonging to 54 species were studied for restriction fra gment length polymorphism of a PCR-amplified 439-bp segment of the gene enc oding the 65-kDa heat shock protein. Restriction digests were separated by 10% polyacrylamide gel electrophoresis (PAGE). By including a size standard in each sample, the restriction fragment profile was calculated using a co mputer-aided comparison program. An algorithm describing these 54 species ( including 22 species not previously described) is proposed. We found that t his assay based on 10% PAGE provided a more precise estimate than that base d on agarose gel electrophoresis of the real size of restriction fragments as deduced from the sequence analysis and allowed identification of mycobac teria whose PCR-restriction fragment length polymorphism analysis patterns were unequivocally identified by fragments shorter than 60 bp.