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New rapid scheme for distinguishing subspecies of the Mycobacterium abscessus group and identification of Mycobacterium massiliense with inducible clarithromycin resistance

ABSTRACT: Mycobacterium abscessus [M. abscessus (sensu lato) or M. abscessus group] comprises three closely related taxa with taxonomic status under revision: M. abscessus sensu stricto, M. bolletii and M. massiliense. We describe here a simple, robust and cost effective PCR-based method for distinguishing among M. abscessus, M. massiliense and bolletii. Based on the M. abscessus ATCC 19977T genome, discriminatory regions were identified between M. abscessus and M. massiliense from array-based comparative genomic hybridization. A typing scheme using PCR primers designed for four of these locations was applied to 46 well-characterized clinical isolates comprising 29 M. abscessus, 15 M. massiliense and 2 M. bolletii previously identified by multi-target sequencing. Interestingly, 2 isolates unequivocally identified as M. massiliense were shown to have a full length erm(41) instead of the expected gene deletion and showed inducible clarithromycin resistance after 14 days. We propose using this PCR-based typing scheme combined with erm(41) PCR for a straightforward identification of M. abscessus, M. massiliense and M. bolletii and assessment of inducible clarithromycin resistance. This method can be easily implemented into a routine workflow providing subspecies level identification within 24 hours of isolation of M. abscessus group.

ORGANISM(S): Mycobacteroides abscessus Mycobacteroides abscessus subsp. bolletii

PROVIDER: GSE43629 | GEO | 2013/07/17

SECONDARY ACCESSION(S): PRJNA186855

REPOSITORIES: GEO

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Project description:Members of the Mycobacterium (M.) abscessus complex (MABC) are rapidly growing mycobacteria showing smooth and/or rough colony morphotype. While not as virulent as M. tuberculosis, they can cause soft tissue infection and fatal pulmonary disease, especially in patients with cystic fibrosis. Diagnosing MABC pulmonary disease is challenging since the isolation of M. abscessus from respiratory samples is in itself not diagnostic and the clinical features are often non-specific. Immunologic assays, which could aid in the understanding and diagnosis of the disease, are not available. In this study eight rough and six smooth colony morphotype isolates were collected from seven clinical MABC strains and the M. abscessus reference strain ATCC19977, as six strains showed both morphotypes simultaneously and two strains only showed a rough morphotype. Clinical isolates were submitted to whole genome sequencing. Quantitative proteomic analysis was performed on bacterial lysates and the culture supernatant of all 14 isolates. Supernatant proteins present in all isolates were compared in a BLAST search against other clinically significant mycobacterial species to determine species-specific proteins of MABC. In silico B- and T-cell epitope prediction was performed for species-specific proteins. All clinical strains were found to be M. abscessus ssp. abscessus. Six of seven rough colony clinical isolates contained genetic changes in the MAB_4099c gene, which is a likely genetic basis for the rough morphotype. Proteomic analysis detected 3 137 different proteins in total of which 79 proteins were found in the culture supernatants of all isolates. BLAST analyses of these 79 proteins identified 12 of those exclusively encoded by all members of MABC plus M. immunogenum. In silico prediction of epitopes predicted B- and T-cell epitopes in all these 12 species-specific proteins, rendering them promising candidates for future studies on immune pathogenesis and immune diagnostic tools for MABC disease.

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New rapid scheme for distinguishing subspecies of the Mycobacterium abscessus group and identification of Mycobacterium massiliense with inducible clarithromycin resistance

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