Inhibition of Virulence Factor Expression and Phage-Induced Lysis by Nitric Oxide in Escherichia coli O157:H7

Abstract

Our research uses computational and experimental methodologies in systems biology to address limitations of traditional antimicrobials. Traditional antimicrobials can cause phage-induced bacterial lysis, virulence factor expression, and extracellular protein release in Escherichia coli O157:H7 and are thus contraindicated for treatment. Previous research has demonstrated that nitric oxide (NO•), an effector of microbial cytotoxicity by macrophages, inhibits Stx2 expression in E. coli O157:H7 (SHIMIZU et al. 2012; VAREILLE et al. 2007). Because of this, we hypothesized that NO• may be able to reduce virulence factor expression and phage-induced lysis. We constructed fluorescent reporter plasmids of the promoter activities of three representative virulence factors – stx1, stx2, and katP, as well as sulA as a positive control. DNA damaging agents normally induce these genes concurrently with lysis. Except for KatP, we found that treatment with a NO• donor quenched DNA-damage induced expression of these genes in a global effect, and further investigation using KCN, a respiration inhibitor and chloramphenicol, a translation inhibitor, indicated that NO• is interfering with translation to cause this inhibitory effect. Furthermore, we also observed that NO• inhibits phage-induced lysis and extracellular protein release upon co-treatment with mitomycin C, ofloxacin, and hydrogen peroxide (H2O2). Additionally, we developed a preliminary NO• kinetic model in E. coli O157:H7 from a previous model in E. coli K-12 (ROBINSON and BRYNILDSEN 2013), which identified iron-sulfur cluster [Fe-S] damage as a potential mechanism for translation inhibition. This indicates that increasing NO• concentration in bacterial cells is a viable strategy for future research addressing the clinical problem of antimicrobial induced virulence factor expression and phage-induced lysis in E. coli O157:H7.