Combining polymerase chain reaction restriction enzyme analysis with phenotypic characters for mycobacteria identification in Taiwan.

SETTING

Many hospitals use the fully-automated BACTEC 960 Mycobacteria Growth Indicator Tube (MGIT) system and acid-fast staining to detect acid-fast bacilli (AFB) in clinical specimens; however, labour-intensive biochemical methods are used for further mycobacterial species identification.

OBJECTIVE

To develop a user-friendly algorithm for mycobacterial species identification from AFB smear-positive BACTEC tubes.

DESIGN

AFB smear-positive BACTEC tubes were collected and mycobacteria were isolated and identified by biochemical methods. The tubes were subgrouped by rpoB duplex polymerase chain reaction restriction enzyme analysis (rpoB DPRA). The results were combined with key phenotypic characters of mycobacteria isolated from the tubes to develop a species identification algorithm with 16S rDNA sequencing of the isolate being used as the gold standard method.

RESULTS

By rpoB DPRA, 441 AFB smear-positive BACTEC tubes were correctly subgrouped into 100 tubes containing Mycobacterium tuberculosis complex, 335 tubes containing non-tuberculous mycobacteria and six tubes containing both. A species identification algorithm was developed by combining the rpoB DPRA results of the tubes with growth rate, photoreactivity and two biochemical results of mycobacteria recovered from the tubes.

CONCLUSION

This user-friendly algorithm can be used for mycobacterial species identification from AFB smear-positive BACTEC tubes.