Oxidative stress responses in biofilms

Abstract

Oxidizing agents are low-molecular-weight molecules that oxidize other substances by accepting electrons from them. They include reactive oxygen species (ROS), such as superoxide anions (O₂^-), hydrogen peroxide (H₂O₂), and hydroxyl radicals (HO^-), and reactive chlorine species (RCS) including sodium hypochlorite (NaOCl) and its active ingredient hypochlorous acid (HOCl), and chloramines. Bacteria encounter oxidizing agents in many different environments and from diverse sources. Among them, they can be produced endogenously by aerobic respiration or exogenously by the use of disinfectants and cleaning agents, as well as by the mammalian immune system. Furthermore, human activities like industrial effluent pollution, agricultural runoff, and environmental activities like volcanic eruptions and photosynthesis are also sources of oxidants. Despite their antimicrobial effects, bacteria have developed many mechanisms to resist the damage caused by these toxic molecules. Previous research has demonstrated that growing as a biofilm particularly enhances bacterial survival against oxidizing agents. This review aims to summarize the current knowledge on the resistance mechanisms employed by bacterial biofilms against ROS and RCS, focussing on the most important mechanisms, including the formation of biofilms in response to oxidative stressors, the biofilm matrix as a protective barrier, the importance of detoxifying enzymes, and increased protection within multi-species biofilm communities. Understanding the complexity of bacterial responses against oxidative stress will provide valuable insights for potential therapeutic interventions and biofilm control strategies in diverse bacterial species.

Keywords: Antimicrobial resistance; Biofilms; Hydrogen peroxide; Hypochlorous acid; Oxidative stress; Reactive chlorine species; Reactive oxygen species.

Fig. 1

Sources of oxidative stress in bacteria and biofilms. Bacteria encounter toxic reactive species such as reactive oxygen and chlorine species (ROS and RCS, respectively) from diverse sources. (a) These toxic molecules can be endogenously produced by aerobic respiration, in which molecular oxygen (O₂) acquires electrons and is converted into superoxide anions (O₂⁻), hydrogen peroxide (H₂O₂), and hydroxyl radicals (HO⁻). (b) ROS and RCS can also be produced by the mammalian innate immune system during phagocytosis. (c) Some antibiotics, such as quinolones and β-lactams, are also known to induce oxidative stress in bacterial cells through many different mechanisms. (d) Plants can also generate toxic oxygen species during photosynthesis or by their roots as, for example, a response to stress. (e) Human activities, including agriculture and the release of industrial effluent pollution, are also important sources of oxidants. Furthermore, disinfectants and cleaning agents such as bleach in wastewater treatment, as well as in industrial, domestic, and hospital settings, are also important sources of ROS and RCS. On a bigger scale, (f) UVA radiation and (g) volcanic gases can also generate toxic reactive species. Cl-, Br⁻, and SCN⁻: chloride, bromide, and thiocyanate anions, respectively; HOCl: hypochlorous acid; HOBr: hypobromous acid; HOSCN: hypothiocyanite; SOD: superoxide dismutase; MPO: myeloperoxidase. Created with BioRender.com.

Fig. 2

Stimulation of biofilm formation by oxidative stress. Oxidative stress induces cell adhesion by inducing changes in cell morphology by, for example, the development of rugose variants and an increase in cell hydrophobicity. Furthermore, these toxic species increase the production of the EPS matrix and its components as well as cyclic-di-GMP. These processes induce the initial step of biofilm development (step 1). Additionally, oxidants such as NaOCl disrupt the biofilm matrix, promoting the release of single cells or cell aggregates, which can then colonize other sites (step 3). Biofilm formation process described by the three-step inclusive model proposed by Sauer et al. [2]. Created with BioRender.com.

Fig. 3

Summary of the oxidative stress response mechanisms employed by biofilms against ROS and RCS.