Heller, DanielleAmaya, IsabelMohamed, AleemAli, IlzatMavrodi, DmitriDeighan, PadraigSivanathan, Viknesh2022-07-142022-07-142022-06-21Danielle Heller, Isabel Amaya, Aleem Mohamed, Ilzat Ali, Dmitri Mavrodi, Padraig Deighan, Viknesh Sivanathan, Systematic overexpression of genes encoded by mycobacteriophage Waterfoul reveals novel inhibitors of mycobacterial growth, G3 Genes|Genomes|Genetics, 2022;, jkac140, https://doi.org/10.1093/g3journal/jkac140https://doi.org/10.1093/g3journal/jkac140http://hdl.handle.net/11603/25151Bacteriophages represent an enormous reservoir of novel genes, many of which are unrelated to existing entries in public databases and cannot be assigned a predicted function. Characterization of these genes can provide important insights into the intricacies of phage–host interactions and may offer new strategies to manipulate bacterial growth and behavior. Overexpression is a useful tool in the study of gene-mediated effects, and we describe here the construction of a plasmid-based overexpression library of a complete set of genes for Waterfoul, a mycobacteriophage closely related to those infecting clinically important strains of Mycobacterium tuberculosis and/or Mycobacterium abscessus. The arrayed Waterfoul gene library was systematically screened in a plate-based cytotoxicity assay, identifying a diverse set of 32 Waterfoul gene products capable of inhibiting the growth of the host Mycobacterium smegmatis and providing a first look at the frequency and distribution of cytotoxic products encoded within a single mycobacteriophage genome. Several of these Waterfoul gene products were observed to confer potent anti-mycobacterial effects, making them interesting candidates for follow-up mechanistic studies.8 pagesen-USThis item is likely protected under Title 17 of the U.S. Copyright Law. Unless on a Creative Commons license, for uses protected by Copyright Law, contact the copyright holder or the author.Attribution 4.0 International (CC BY 4.0)https://creativecommons.org/licenses/by/4.0/Text