Studies on Bd0934 and Bd3507, Two Secreted Nucleases from , Reveal Sequential Release of Nucleases during the Predatory Cycle.

is an obligate predatory bacterium that invades and kills a broad range of Gram-negative prey cells, including human pathogens. Its potential therapeutic application has been the subject of increased research interest in recent years. However, an improved understanding of the fundamental molecular aspects of the predatory life cycle is crucial for developing this bacterium as a "living antibiotic." During intracellular growth, secretes an arsenal of hydrolases, which digest the content of the host cell to provide growth nutrients for the predator, e.g., prey DNA is completely degraded by the nucleases. Here, we have, on a genetic and molecular level, characterized two secreted DNases from , Bd0934 and Bd3507, and determined the temporal expression profile of other putative secreted nucleases. We conclude that Bd0934 and Bd3507 are likely a part of the predatosome but are not essential for the predation, host-independent growth, prey biofilm degradation, and self-biofilm formation. The detailed temporal expression analysis of genes encoding secreted nucleases revealed that these enzymes are produced in a sequential orchestrated manner. This work contributes to our understanding of the sequential breakdown of the prey nucleic acid by the nucleases secreted during the predatory life cycle of Antibiotic resistance is a major global concern with few available new means to combat it. From a therapeutic perspective, predatory bacteria constitute an interesting tool. They not only eliminate the pathogen but also reduce the overall pool of antibiotic resistance genes through secretion of nucleases and complete degradation of exogenous DNA. Molecular knowledge of how these secreted DNases act will give us further insight into how antibiotic resistance, and the spread thereof, can be limited through the action of predatory bacteria.