Mycobacterium abscessus (Mab) is a rapidly growing NTM causing skin and soft tissue infections and pulmonary infections in patients with chronic lung damage. It stands apart as one of the most antibiotic resistant microbial species, making its infections incredibly difficult to treat. A combination of an oral macrolide and the aminoglycoside, amikacin, comprises the frontline treatment against Mab. Therapy is prolonged and low cure rates are deplorable. The poor efficacies of these antibiotics result from various mechanisms of intrinsic resistance induced in Mab upon exposure to drugs as well as the host environment. Recently we showed that MabHflX, a conserved ribosome-associated GTPase, is required for macrolide-lincosamide resistance (4). The absence of HflX in a DMs_hflX deletion strain results in an increased population of 70S ribosomes suggesting that HflX is involved in dissociation of ribosomes stalled in the presence of antibiotics. However, the detailed mechanism of HflX-mediated antibiotic resistance, as well as the mechanisms by which antibiotic-bound ribosomal subunits are recycled remain unknown. In another study we demonstrated a role for ARE-ABCF proteins in macrolide/lincosamide resistance.
In Aim 1 of this proposal we will use a combination of genetic, biochemical and structural approaches to determine the mechanisms of HflX and ABCF-mediated macrolide-lincosamide resistance. In another independent study, aminoglycoside resistance in zinc-starved M. smegmatis was found to originate from ribosome hibernation, which involves binding of mycobacterial protein Y (MPY). We hypothesize that a low-zinc host environment would similarly result in MPY-dependent ribosome hibernation during Mab infection, and confer resistance to aminoglycosides including amikacin.
In Aim 2 of this proposal, we will determine the role of MabMPY in aminoglycoside resistance of which could potentially explain the observed discord between in vitro efficiency and in vivo efficacy of the current treatments (Aim 2). The extreme innate antibiotic resistance of Mab presents a unique opportunity to study the convergence of multiple resistance mechanisms in this pathogen. An in-depth understanding of these various mechanisms is critical in the development of new therapeutic approaches towards treatment of Mab infections.
Macrolides and aminoglycosides comprise the frontline treatment against M. abscessus. However induction of resistance pathways either by exposure to the drugs or the host environment result in a poor treatment efficacy. In this proposal we will undertake an in- depth structure-function characterization of two novel resistance pathways involving the ribosome associated proteins- HflX and ABCFs. This work will also provide the first demonstration of changes in bacteria within the host environment that result in increased resistance to aminoglycosides.
