Isolation and Screening of Hydrogen-Oxidizing Bacteria from Mangrove Sediments for Efficient Single-Cell Protein Production Using CO2

The escalating global demand for large-scale, cost-effective, and sustainable high-quality protein has positioned single-cell protein (SCP) production from one-carbon (C1) gases as a highly promising solution. In this study, eight chemolithoautotrophic hydrogen-oxidizing bacteria (HOB) were isolated from mangrove sediments. Based on the 16S rRNA gene sequence analysis, they belonged to genera <i>Sulfurimonas</i>, <i>Sulfurovum</i>, <i>Thiomicrolovo</i>, and <i>Marinobacterium.</i> Among these, <i>Thiomicrolovo</i> sp. ZZH C-3 was identified as the most promising candidate for SCP production based on the highest biomass and protein content, and was selected for further characterization. Strain ZZH C-3 is a Gram-negative, short rod-shaped bacterium with multiple flagella. It can grow chemolithoautotrophically by using molecular hydrogen as an energy source and molecular oxygen as an electron acceptor. Genomic analysis further confirmed that ZZH C-3 harbors a complete reverse tricarboxylic acid (rTCA) cycle gene set for carbon fixation, and diverse hydrogenases (Group I, II, IV) for hydrogen oxidation. Subsequently, its cultivation conditions and medium composition for SCP production were systematically optimized using single-factor experiments and response surface methodology (RSM). Results showed that the optimal growth conditions were 28 °C, pH 7.0, and with 1 g/L (NH₄)₂SO₄ as the nitrogen source, 5-10% oxygen concentration, 9.70 mg/L FeSO₄·7H₂O, 0.17 g/L CaCl₂·2H₂O, and 1.90 mg/L MnSO₄·H₂O. Under the optimized conditions, strain ZZH C-3 achieved a maximum specific growth rate of 0.46 h^-1. After 28 h of cultivation, the optical density at 600 nm (OD₆₀₀) reached 0.94, corresponding to a biomass concentration of 0.60 g/L, and the protein content ranked at 73.56%. The biomass yield on hydrogen (Y_H2) was approximately 3.01 g/g H₂, with an average H₂-to-CO₂ consumption molar ratio of about 3.78. Compared to the model HOB <i>Cupriavidus necator</i>, strain ZZH C-3 exhibited a lower H₂/CO₂ consumption ratio, superior substrate conversion efficiency, and high protein content. Overall, this study not only validated the potential of mangrove HOB for SCP production but also offers new insights for future metabolic engineering strategies designed to enhance CO₂-to-biomass conversion efficiency.