The Homologous Gene of Chromosomal Virulence D (chvD) Presents High Resolution as a Novel Biomarker in Mycobacterium Species Identification

Abstract

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.