Bioenergetic State of Escherichia coli Controls Aminoglycoside Susceptibility.

Aminoglycosides (AG) have been used against Gram-negative bacteria for decades. Yet, how bacterial metabolism and environmental conditions modify AG toxicity is poorly understood. Here, we show that the level of AG susceptibility varies depending on the nature of the respiratory chain that Escherichia coli uses for growth, i.e., oxygen, nitrate, or fumarate. We show that all components of the fumarate respiratory chain, namely, hydrogenases 2 and 3, the formate hydrogenlyase complex, menaquinone, and fumarate reductase are required for AG-mediated killing under fumarate respiratory conditions. In addition, we show that the AAA+ ATPase RavA and its Von Wildebrand domain-containing partner, ViaA, are essential for AG to act under fumarate respiratory conditions. This effect was true for all AG that were tested but not for antibiotics from other classes. In addition, we show that the sensitizing effect of RavA-ViaA is due to increased gentamicin uptake in a proton motive force-dependent manner. Interestingly, the sensitizing effect of RavA-ViaA was prominent in poor energy conservation conditions, i.e., with fumarate, but dispensable under high energy conservation conditions, i.e., in the presence of nitrate or oxygen. We propose that RavA-ViaA can facilitate uptake of AG across the membrane in low-energy cellular states. Antibiotic resistance is a major public health, social, and economic problem. Aminoglycosides (AG) are known to be highly effective against Gram-negative bacteria, but their use is limited to life-threatening infections because of their nephrotoxicity and ototoxicity at therapeutic dose. Elucidation of AG-sensitization mechanisms in bacteria would allow reduced effective doses of AG. Here, we have identified the molecular components involved in anaerobic fumarate respiration that are required for AG to kill. In addition to oxidoreductases and menaquinone, this includes new molecular players, RavA, an AAA+ ATPase, and ViaA, its partner that has the VWA motif. Remarkably, the influence of RavA-ViaA on AG susceptibility varies according to the type of bioenergetic metabolism used by E. coli. This is a significant advance because anaerobiosis is well known to reduce the antibacterial activity of AG. This study highlights the critical importance of the relationship between culture conditions, metabolism, and antibiotic susceptibility.