PT - JOURNAL ARTICLE AU - Rafael Rivera-Lugo AU - David Deng AU - Andrea Anaya-Sanchez AU - Sara Tejedor-Sanz AU - Valeria M Reyes Ruiz AU - Hans B Smith AU - Denis V Titov AU - John-Demian Sauer AU - Eric P Skaar AU - Caroline M Ajo-Franklin AU - Daniel A Portnoy AU - Samuel H Light TI - <em>Listeria monocytogenes</em> requires cellular respiration for NAD<sup>+</sup> regeneration and pathogenesis AID - 10.1101/2021.11.25.470024 DP - 2021 Jan 01 TA - bioRxiv PG - 2021.11.25.470024 4099 - http://biorxiv.org/content/early/2021/11/25/2021.11.25.470024.short 4100 - http://biorxiv.org/content/early/2021/11/25/2021.11.25.470024.full AB - Cellular respiration is essential for multiple bacterial pathogens and a validated antibiotic target. In addition to driving oxidative phosphorylation, bacterial respiration has a variety of ancillary functions that obscure its contribution to pathogenesis. We find here that the intracellular pathogen Listeria monocytogenes encodes two respiratory pathways which are partially functionally redundant and indispensable for pathogenesis. Loss of respiration decreased NAD+ regeneration, but this could be specifically reversed by heterologous expression of a water-forming NADH oxidase (NOX). NOX expression fully rescued intracellular growth defects and increased L. monocytogenes loads &gt;1,000-fold in a mouse infection model. Consistent with NAD+ regeneration maintaining L. monocytogenes viability and enabling immune evasion, a respiration-deficient strain exhibited elevated bacteriolysis within the host cytosol and NOX rescued this phenotype. These studies show that NAD+ regeneration, rather than oxidative phosphorylation, represents the primary role of L. monocytogenes respiration and highlight the nuanced relationship between bacterial metabolism, physiology, and pathogenesis.Competing Interest StatementDenis Titov is a co-inventor on a filed patent describing the use of NOX.