Molecular Identification of Rare Clinical Mycobacteria by Application of ۱۶S-۲۳S Spacer Region Sequencing

Objective(s) In addition to several molecular methods and in particular ۱۶S rDNA analysis, the application of a more discriminatory genetic marker, i.e., ۱۶S-۲۳S internal transcribed spacer gene sequence has had a great impact on identification and classification of mycobacteria. In the current study we aimed to apply this sequencing power to conclusive identification of some Iranian clinical strains of mycobacteria. Materials and Methods The test strains consisted of nineteen mycobacterial isolates which were initially identified by the use of conventional phenotypic techniques and molecular methods and subjected to further definitive identification using the ۱۶S-۲۳S internal transcribed spacer gene sequencing. Results Out of ۱۹ studied strains, ۷ isolates were found to be rapidly growing and ۱۲ isolates as slowly growing mycobacteria. With the exception of one isolate, i.e., the isolate HNTM۸۷, which yielded a distinct ITS sequence incomparable with all previously identified mycobacteria, the remaining isolates produced the sequences similar to the established mycobacteria and were clearly identified and differentiated from closely related taxa. A phylogenetic tree based on maximum parsimony analysis of ۱۶S-۲۳S internal transcribed spacer gene sequences constructed showing the relatedness of Iranian clinical isolates with the closely related type species of mycobacteria. Conclusion This study showed that the ۱۶S-۲۳S internal transcribed spacer gene of the genus Mycobacterium exhibits a high variation which is of value for discriminating closely related taxa and could be used independently or in combination with ۱۶S rDNA sequencing to delineate the true identity of rare mycobacterial species.

Keywords:

Internal Transcribed Spacer , Mycobacterium , Sequence Analysis