Molecular Mechanisms of Intrinsic Streptomycin Resistance in Mycobacterium abscessus [PublishAheadOfPrint]

Streptomycin, the first drug used for the treatment of tuberculosis, shows limited activity against the highly resistant pathogen Mycobacterium abscessus. We recently identified two aminoglycoside-acetylating genes [aac(2') and eis2] which however do not affect susceptibility to streptomycin. This suggests the existence of a discrete mechanism of streptomycin resistance. M. abscessus BlastP analysis identified MAB_2385 as a close homologue of the 3' '-O-phosphotransferase [APH(3' ')] from the opportunistic pathogen Mycobacterium fortuitum, as a putative streptomycin resistance determinant. Heterologous expression of MAB_2385 in M. smegmatis increased the streptomycin minimal inhibitory concentration while the gene deletion mutant M. abscessus MAB_2385 showed increased streptomycin susceptibility. The minimal inhibitory concentration of other aminoglycosides was not altered in M. abscessus MAB_2385. This concludes that MAB_2385 is a specific and prime innate streptomycin resistance determinant in M. abscessus. We further explored the feasibility to apply an rpsL-based streptomycin counter selection to generate gene deletion mutants in M. abscessus. Spontaneous streptomycin resistant mutants of M. abscessus MAB_2385 were selected and we demonstrated that the wild-type rpsL is dominant over the mutated rpsL^K43R in merodiploid strains. In a proof of concept study we exploited this phenotype for construction of a targeted deletion mutant thereby establishing an rpsL-based counter selection method in M. abscessus.

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