Utility of high-performance liquid chromatography analysis of mycolic acids and partial 16S rRNA gene sequencing for routine identification of Mycobacterium spp. in a national reference laboratory

Abstract 

High-performance liquid chromatography analysis of mycolic acids and partial gene sequencing for the first 500-bp 5′ end of the 16S rRNA gene were used singularly and in combination to evaluate the final identification of species. Examination of 200 cultures revealed 100 strains of slowly growing mycobacteria (SGM), 91 strains of rapidly growing mycobacteria (RGM), and 9 strains of other genera. SGM were discriminated in complexes with both methods for 56 strains, composed primarily of the Mycobacterium spp.: Mycobacterium avium, Mycobacterium terrae, and Mycobacterium simiae–Mycobacterium lentiflavum. For RGM, 73 strains were associated with complexes designated as Mycobacterium abscessus–Mycobacterium chelonae, Mycobacterium fortuitum–Mycobacterium peregrinum, and Mycobacterium mucogenicum–Mycobacterium phocaicum. Consistent identification of all the isolates differentiated to single species within the Mycobacterium genus was not possible with either test method. Sequencing results often distinguished complexes containing fewer species, and combining the results from each method increased the confidence of identifying the correct species.

Keywords: 16S rRNA gene sequencing, High-performance liquid chromatography, Mycolic acids