Molecular basis of intrinsic macrolide resistance in the Mycobacterium tuberculosis complex

Abstract

The intrinsic resistance of the Mycobacterium tuberculosis complex (MTC) to most antibiotics, including macrolides, is generally attributed to the low permeability of the mycobacterial cell wall. However, nontuberculous mycobacteria (NTM) are much more sensitive to macrolides than members of the MTC. A search for macrolide resistance determinants within the genome of M. tuberculosis revealed the presence of a sequence encoding a putative rRNA methyltransferase. The deduced protein is similar to Erm methyltransferases, which confer macrolide-lincosamide-streptogramin (MLS) resistance by methylation of 23S rRNA, and was named ErmMT. The corresponding gene, ermMT (erm37), is present in all members of the MTC but is absent in NTM species. Part of ermMT is deleted in some vaccine strains of Mycobacterium bovis BCG, such as the Pasteur strain, which lack the RD2 region. The Pasteur strain was susceptible to MLS antibiotics, whereas MTC species harboring the RD2 region were resistant to them. The expression of ermMT in the macrolide-sensitive Mycobacterium smegmatis and BCG Pasteur conferred MLS resistance. The resistance patterns and ribosomal affinity for erythromycin of Mycobacterium host strains expressing ermMT, srmA (monomethyltransferase from Streptomyces ambofaciens), and ermE (dimethyltransferase from Saccharopolyspora erythraea) were compared, and the ones conferred by ErmMT were similar to those conferred by SrmA, corresponding to the MLS type I phenotype. These results suggest that ermMT plays a major role in the intrinsic macrolide resistance of members of the MTC and could be the first example of a gene conferring resistance by target modification in mycobacteria.

FIG. 1.

Comparison of the amino acid sequence of ErmMT (Erm37 subgroup, accession no. CAA98396) with those of ErmCD (ErmX subgroup) from Corynebacterium diphtheriae (accession no. AAA98484), SrmA (ErmO subgroup) from Streptomyces ambofaciens (accession no. CAA11706), ErmE (ErmE subgroup) from Saccharopolyspora erythraea (accession no. CAB60001), ErmC′ (ErmC subgroup) from Bacillus subtilis (accession no. AAA98136), and ErmAM (ErmB subgroup) from Streptococcus pneumoniae (accession no. S12727). The structures of ErmC′ and ErmAM are known (Protein Data Bank accession no. 2ERC and 1YUB). The alignment was made using the MultAlign program (12). Conserved regions are shaded (grey, moderate conservation; black, identical residues). The positions and approximate extents of the eight conserved motifs found in other DNA and RNA methyltransferases are indicated by arrows. Motif VII is weakly conserved in the Erm family.

FIG. 2.

Schematic representation of the ermMT regions of M. tuberculosis, M. bovis, and M. bovis BCG Moreau compared to that of M. bovis BCG Pasteur (RD2 deletion).

FIG. 3.

Binding of erythromycin to ribosomes extracted from M. smegmatis strains harboring the empty vector (pMIP12, ▪), pOMV16 (ermMT, ○), pOMV20 (ermE, ▴), and pOMV30 (srmA, •).