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Background  Mycobacteria can be quickly and simply identified by PCR restriction-enzyme analysis (PRA), but misidentification can occur because of similarities in band sizes that are critical for discriminating among species. Capillary electrophoresis can provide computer-aided band discrimination. The aim of this research was to develop an algorithm for identifying mycobacteria by combined  rpo B duplex PRA (DPRA) and  hsp65  PRA with capillary electrophoresis.    Results  Three hundred and seventy-six acid-fast bacillus smear-positive BACTEC cultures, including 200  Mycobacterium tuberculosis  complexes (MTC) and 176 non-tuberculous mycobacteria (NTM) were analyzed. With combined  hsp65  and  rpo B DPRA, the accuracy rate was 100% (200 isolates) for the MTC and 91.4% (161 isolates) for the NTM. Among the discordant results (8.6%) for the NTM, one isolate of  Mycobacterial species  and an isolate of  M. flavescens  were found as new sub-types in  hsp65  PRA.    Conclusions  This effective and novel identification algorithm using combined  rpo B DPRA and  hsp65  PRA with capillary electrophoresis can rapidly identify mycobacteria and find new sub-types in  hsp65  PRA. In addition, it is complementary to 16 S rDNA sequencing.

bmc microbiology

Combined rpo B duplex PCR and hsp65 PCR restriction fragment length polymorphism with capillary electrophoresis as an effective algorithm for identification of Mycobacterial species from clinical isolates