Objective: To evaluate the resolution of chromosomal virulence D (chvD ) as a novel marker for mycobacterial species identification.
Methods: A segment of chvD (652 bp) was amplified by PCR from 63 mycobacterial reference strains, 163 nontuberculous mycobacterial clinical isolates, and 16 M. tuberculosis complex (MTBC) clinical isolates. A phylogenetic tree based on the reference strains was constructed by the neighbor-joining and IQ-tree methods. Comparative sequence analysis of the homologous chvD gene efficiently differentiated the species within the genus Mycobacterium . Slowly growing Mycobacterium (SGM) and rapidly growing Mycobacterium (RGM) were separated in the phylogenetic tree based on the chvD gene.
Results: The sequence discrepancies were obvious between M. kansasii and M. gastri , M. chelonae and M. abscessus , and M. avium and M. intracellulare , none of which could be achieved by 16S ribosomal RNA (rRNA) homologous gene alignment. Furthermore, chvD manifested larger intraspecies diversity among members of M. intracellulare subspecies. A total of 174 of the 179 (97.21%) clinical isolates, consisting of 12 mycobacterial species, were identified correctly by chvD blast. Four M. abscessus subsp. abscessus were identified as M. abscessus subsp. bolletii by chvD . MTBC isolates were indistinguishable, because they showed 99.84%– 100% homology.
Conclusion: Homologous chvD is a promising gene marker for identifying mycobacterial species, and could be used for highly accurate species identification among mycobacteria.
Keywords: species identification, Mycobacterium , chromosomal virulence D, phylogenetic tree