Variable pH and subsequent change in pCO2 modulates the biofilm formation, synthesis of extracellular polymeric substances, and survivability of a marine bacterium Bacillus stercoris GST-03

Abstract

Biofilm-forming bacteria adhere to the substrates and engage in the nutrient cycling process. However, environmental conditions may interrupt the biofilm formation ability, which ultimately may affect various biogeochemical cycles. The present study reports the effect of varying pH and subsequent change in pCO₂ on the survivability, biofilm formation, and synthesis of extracellular polymeric substances (EPS) of a biofilm-forming marine bacterium Bacillus stercoris GST-03 isolated from the Bhitarkanika mangrove ecosystem, Odisha, India. Understanding the pH-dependent alteration in EPS constituents, and associated functional groups of a marine bacterium will provide better insight into the adaptability of the bacteria in future ocean acidification scenarios. The strain was found to tolerate and form biofilm up to pH 4, with the maximum biofilm formation at pH 6. EPS yield and the synthesis of the key components of the EPS, including carbohydrate, protein, and lipid, were found maximum at pH 6. Changes in biofilm formation patterns and various topological parameters at varying pH/pCO₂ conditions were observed. A cellular chaining pattern was observed at pH 4, and maximum biofilm formation was obtained at pH 6 with biomass of 5.28582 ± 0.5372 μm³/μm² and thickness of 9.982 ± 1.5288 μm. Structural characterization of EPS showed changes in various functional groups of constituent macromolecules with varying pH. The amorphous nature of the EPS and the changes in linkages and associated functional groups (-R₂CHOR, -CH₃, and -CH₂) with pH variation was confirmed. EPS showed a two-step degradation with a maximum weight loss of 59.147% and thermal stability up to 480 °C at pH 6. The present work efficiently demonstrates the role of EPS in providing structural and functional stability to the biofilm in varying pH conditions. The findings will provide a better understanding of the adaptability of marine bacteria in the future effect of ocean acidification.

Keywords: Biofilm; Extracellular polymeric substances; Marine bacteria; Ocean acidification; pH.