Key gene drives ethambutol resistance in Mycobacterium avium

Infections with the Mycobacterium avium-intracellulare complex (MAC) are the most common nontuberculous mycobacterial (NTM) illnesses, and ethambutol (EMB) is one of the main drugs used to treat them. Yet clinicians and researchers have had limited clarity about how often M. avium is resistant to EMB and what genetic changes cause that resistance. Ying Zhang and colleagues set out to fill that gap by testing clinical M. avium strains and then using laboratory methods to force and examine EMB resistance. They measured the minimum inhibitory concentration (MIC) of EMB on 40 M. avium clinical strains and found nearly all were intermediate or resistant. From a single EMB-susceptible isolate called strain 245 (MIC = 2 μg/mL), they generated multiple EMB-resistant mutants in the lab to study which genetic changes appeared when resistance developed. The team then sequenced those mutants to look for common mutations that could explain the loss of EMB susceptibility. The work was designed to identify direct genetic causes of resistance, a step toward better diagnostics and treatment decisions.

To probe mechanisms of resistance, the researchers first determined MIC values across the panel of clinical strains, finding 97.5% (39/40) were intermediate (MIC = 4 μg/mL) or resistant (MIC ≥ 8 μg/mL) to ethambutol (EMB). They chose the single susceptible clinical isolate, strain 245 (MIC = 2 μg/mL), to isolate EMB-resistant mutants and obtained 121 resistant mutants. Those mutants were analyzed by whole-genome sequencing or Sanger sequencing to identify genetic changes associated with resistance. Integrated analysis showed that 94.21% (114/121) of the mutants carried mutations in the ubiA gene. ubiA encodes DPPR (decaprenylphosphoryl-β-D-5-phosphoribose) synthase, an enzyme involved in cell wall biosynthesis that has been linked to high-level EMB resistance in M. tuberculosis. To test causality, the team performed complementation with the wild-type ubiA gene: this restored EMB susceptibility in EMB-resistant mutants and in a clinical strain 322 (EMB MIC = 64 μg/mL) carrying a ubiA mutation. Complementation reduced MICs from 32 μg/mL to 4 μg/mL and from 64 μg/mL to 8 μg/mL, respectively.

The study identifies ubiA mutation as the major mechanism of ethambutol resistance in Mycobacterium avium. That finding has two immediate implications. First, it explains why so many clinical M. avium isolates showed intermediate or high MICs to EMB in the authors’ testing: changes in a single, specific gene — ubiA — can account for most resistance that developed in the lab and appears in at least some clinical strains. Second, because the resistance is driven mainly by mutations in a defined gene, it becomes feasible to design molecular tests that look specifically for ubiA changes to provide rapid detection of EMB resistance. Faster, gene-based diagnostics could inform clinicians sooner about whether EMB is likely to work for a given patient and help avoid ineffective therapy. The study therefore points toward practical diagnostic tools and offers a clearer genetic target for surveillance of EMB resistance in this organism.