Bidirectional redox cycling of phenazine-1-carboxylic acid by Citrobacter portucalensis MBL drives increased nitrate reduction

Abstract

Phenazines are secreted metabolites that microbes use in diverse ways, from quorum sensing to antimicrobial warfare to energy conservation. Phenazines are able to contribute to these activities due to their redox activity. The physiological consequences of cellular phenazine reduction have been extensively studied, but the counterpart phenazine oxidation has been largely overlooked. Phenazine-1-carboxylic acid (PCA) is common in the environment and readily reduced by its producers. Here, we describe its anaerobic oxidation by Citrobacter portucalensis strain MBL, which was isolated from topsoil in Falmouth, MA, and which does not produce phenazines itself. This activity depends on the availability of a suitable terminal electron acceptor, specifically nitrate or fumarate. When C. portucalensis MBL is provided reduced PCA and either nitrate or fumarate, it continuously oxidizes the PCA. We compared this terminal electron acceptor-dependent PCA-oxidizing activity of C. portucalensis MBL to that of several other γ-proteobacteria with varying capacities to respire nitrate and/or fumarate. We found that PCA oxidation by these strains in a fumarate-or nitrate-dependent manner is decoupled from growth and correlated with their possession of the fumarate or periplasmic nitrate reductases, respectively. We infer that bacterial PCA oxidation is widespread and genetically determined. Notably, reduced PCA enhances the rate of nitrate reduction to nitrite by C. portucalensis MBL beyond the stoichiometric prediction, which we attribute to C. portucalensis MBL’s ability to also reduce oxidized PCA, thereby catalyzing a complete PCA redox cycle. This bidirectionality highlights the versatility of PCA as a biological redox agent.