Killing of Gram-negative and Gram-positive bacteria by a bifunctional cell wall-targeting T6SS effector

Significance Previous studies have indicated that Gram-positive bacteria are not affected by type VI secretion serum (T6SS) intoxication. However, here we show that Acinetobacter baumannii employs its T6SS to kill different Gram-positive bacteria. Furthermore, we determined that killing was dependent on Tse4, a bifunctional effector possessing lytic transglycosylase and endopeptidase activities. Tse4 represents a broad family of modularly organized T6SS peptidoglycan-degrading effectors. In addition, we show that secretion of D-lysine by A. baumannii results in a pH increase, which greatly enhances Tse4 activity. These results expand the range of T6SS-mediated interbacterial interactions that may shape the composition of bacterial communities in the context of the human microbiota and polymicrobial infections. The type VI secretion system (T6SS) is a powerful tool deployed by Gram-negative bacteria to antagonize neighboring organisms. Here, we report that Acinetobacter baumannii ATCC 17978 (Ab17978) secretes D-lysine (D-Lys), increasing the extracellular pH and enhancing the peptidoglycanase activity of the T6SS effector Tse4. This synergistic effect of D-Lys on Tse4 activity enables Ab17978 to outcompete Gram-negative bacterial competitors, demonstrating that bacteria can modify their microenvironment to increase their fitness during bacterial warfare. Remarkably, this lethal combination also results in T6SS-mediated killing of Gram-positive bacteria. Further characterization revealed that Tse4 is a bifunctional enzyme consisting of both lytic transglycosylase and endopeptidase activities, thus representing a family of modularly organized T6SS peptidoglycan-degrading effectors with an unprecedented impact in antagonistic bacterial interactions.