Iron Triggers λSo Prophage Induction and Release of Extracellular DNA in Shewanella oneidensis MR-1 Biofilms

Abstract

bProphages are ubiquitous elements within bacterial chromosomes and affect host physiology and ecology in multiple ways. We have previously demonstrated that phage-induced lysis is required for extracellular DNA (eDNA) release and normal biofilm formation in Shewanella oneidensis MR-1. Here, we investigated the regulatory mechanisms of prophage So spatiotemporal induction in biofilms. To this end, we used a functional fluorescence fusion to monitor So activation in various mutant backgrounds and in response to different physiological conditions. So induction occurred mainly in a subpopulation of filamentous cells in a strictly RecA-dependent manner, implicating oxidative stress-induced DNA damage as the major trigger. Accordingly, mutants affected in the oxidative stress response (⌬oxyR) or iron homeostasis (⌬fur) displayed drastically increased levels of phage induction and abnormal biofilm formation, while planktonic cells were not or only marginally affected. To further investigate the role of oxidative stress, we performed a mutant screen and identified two independent amino acid substitutions in OxyR (T104N and L197P) that suppress induction of So by hydrogen peroxide (H 2 O 2 ). However, So induction was not suppressed in biofilms formed by both mutants, suggesting a minor role of intracellular H 2 O 2 in this process. In contrast, addition of iron to biofilms strongly enhanced So induction and eDNA release, while both processes were significantly suppressed at low iron levels, strongly indicating that iron is the limiting factor. We conclude that uptake of iron during biofilm formation triggers So-mediated lysis of a subpopulation of cells, likely by an increase in iron-mediated DNA damage sensed by RecA.

Isolation and Characterization of a Shewanella Phage–Host System from the Gut of the Tunicate, Ciona intestinalis

“…This novel strain of Shewanella fidelis (3313) was sequenced, and its inducible prophages and a strain-specific lytic phage (SFCi1, which was isolated separately from seawater) were characterized. Previously, it has been shown that spontaneous prophage induction can augment biofilms in some strains of bacteria [28,29,30,31]; we demonstrate here that infection of S. fidelis 3313 by lytic phage SFCi1 also enhances biofilm formation in vitro in a similar DNA-dependent manner. …”

“…Activated prophages are known to influence biofilms by facilitating the transient liberation of extracellular DNA, which can then become a component of the biofilm matrix [21,31]. Biofilms can shelter microbial inhabitants from both physical and mechanical stressors, and consist mostly of microbially-derived polysaccharides, nucleic acids and lipids.…”

Defining the binding determinants of Shewanella oneidensis OxyR: Implications for the link between the contracted OxyR regulon and adaptation

“…In SoOxyR, the counterparts of these two cysteines are Cys-203 and Cys-212 respectively; mutation of either residue to serine locks SoOxyR into the reduced form (19,24). In addition, SoOxyR L197P mutant is found to be locked into the oxidized state (25) (18,19). Because the promoter is slightly leaky (26,27) Fig.…”

Impact of Spontaneous Prophage Induction on the Fitness of Bacterial Populations and Host-Microbe Interactions

“…In S. oneidensis, three prophages, MuSo1, MuSo2, and LambdaSo, jointly affect lysis and eDNA release, and consequently, a mutant devoid of all three phages is severely impaired in all stages of biofilm formation (70). Recent studies on this species have demonstrated that environmental iron levels are an important factor involved in inducing expression of LambdaSo in a RecA-dependent fashion during biofilm formation (71). Another highly interesting example for hostphage interactions is the lysogenic infection of P. aeruginosa PA14 with bacteriophage DMS3.…”