Complex Responses to Hydrogen Peroxide and Hypochlorous Acid by the Probiotic Bacterium Lactobacillus reuteri

Abstract

Inflammatory diseases of the gut are associated with increased intestinal oxygen concentrations and high levels of inflammatory oxidants, including hydrogen peroxide (H2O2) and hypochlorous acid (HOCl), which are antimicrobial compounds produced by the innate immune system. This contributes to dysbiotic changes in the gut microbiome, including increased populations of proinflammatory enterobacteria (Escherichia coli and related species) and decreased levels of health-associated anaerobic Firmicutes and Bacteroidetes. The pathways for H2O2 and HOCl resistance in E. coli have been well studied, but little is known about how commensal and probiotic bacteria respond to inflammatory oxidants. In this work, we have characterized the transcriptomic response of the anti-inflammatory, gut-colonizing Gram-positive probiotic Lactobacillus reuteri to both H2O2 and HOCl. L. reuteri mounts distinct but overlapping responses to each of these stressors, and both gene expression and survival were strongly affected by the presence or absence of oxygen. Oxidative stress response in L. reuteri required several factors not found in enterobacteria, including the small heat shock protein Lo18, polyphosphate kinase 2, and RsiR, an L. reuteri-specific regulator of anti-inflammatory mechanisms. IMPORTANCE Reactive oxidants, including hydrogen peroxide and hypochlorous acid, are antimicrobial compounds produced by the immune system during inflammation. Little is known, however, about how many important types of bacteria present in the human microbiome respond to these oxidants, especially commensal and other health-associated species. We have now mapped the stress response to both H2O2 and HOCl in the intestinal lactic acid bacterium Lactobacillus reuteri.

Environment-Specific Probiotic Supernatants Modify the Metabolic Activity and Survival of Streptococcus mutans in vitro

“…It has also been shown that L. reuteri is capable of producing H 2 O 2 (Kang et al, 2011;Basu et al, 2019), and in our experiments, L. reuteri produced H 2 O 2 in each MM variation. H 2 O 2 is mainly produced by Pox (as described) and NADH oxidase (Nox) (Hertzberger et al, 2014), while it is unclear which enzymatic pathway was relevant in our setting.…”

The Cu(II) Reductase RclA Protects Escherichia coli against the Combination of Hypochlorous Acid and Intracellular Copper

“…1 ; see Data Set S1 in the supplemental material), including members of the Bacteroidetes , Clostridiaceae , and Lactobacillaceae , where their physiological roles are unknown. Expression of the rclA homolog of the probiotic Lactobacillus reuteri is induced modestly by HOCl, but an L. reuteri rclA mutant is not sensitive to HOCl stress ( 36 ). It is unclear, however, whether this is because RclA has a different physiological function in L. reuteri (possibly related to Cu homeostasis) or because RclA requires either strong induction or the presence of RclB and RclC to protect against HOCl under laboratory growth conditions.…”

“…1 ) and is found almost exclusively in bacteria known to colonize epithelial surfaces (see Data Set S1 in the supplemental material), suggesting that it may play an important role in host-microbe interactions in many species. Bacteria encoding RclA homologs include Gram-negative species (e.g., Salmonella enterica ), Gram-positive species (e.g., Streptococcus sanguinis ), obligate anaerobes (e.g., Clostridium perfringens ), facultative anaerobes (e.g., Staphylococcus aureus ), pathogens (e.g., Enterococcus faecalis ), commensals (e.g., Bacteroides thetaiotaomicron ), and probiotics (e.g., Lactobacillus reuteri ) ( 36 ), suggesting that RclA’s function may be broadly conserved and not specific to a single niche or type of host-microbe interaction. RclR, RclB, and RclC are much less widely conserved and are found only in certain species of proteobacteria, primarily members of the Enterobacteriaceae ( Fig.…”

“…thetaiotaomicron), and probiotics (e.g., Lactobacillus reuteri) (36), suggesting that RclA's function may be broadly conserved and not specific to a single niche or type of host-microbe interaction. RclR, RclB, and RclC are much less widely conserved and are found only in certain species of proteobacteria, primarily members of the Enterobacteriaceae ( Fig.…”

The Cu(II) reductase RclA protectsEscherichia coliagainst the combination of hypochlorous acid and intracellular copper

“…Bacteroides thetaiotaomicron ), and probiotics ( e.g. Lactobacillus reuteri ) (36), suggesting that RclA’s function may be broadly conserved and not specific to a single niche or type of host-microbe interaction. RclR, RclB, and RclC are much less widely conserved, and are found only in certain species of proteobacteria, primarily members of the Enterobacteriaceae (Figure 1 and Supplemental Table 1).…”