Isolation and characterization of Enterococcus faecalis isolate VT-H1: A highly efficient hydrogen-producing bacterium from palm oil mill effluent (POME)

Facultative hydrogen-producing bacteria hold immense potential for fermentative hydrogen and valuable metabolite production. In this study, a mesophilic hydrogen-producing bacterium was isolated from palm oil mill effluent (POME) and identified as Enterococcus faecalis isolate VT-H1 using hybrid sequencing technologies. Our investigation encompassed the growth of the isolate and its capacity for hydrogen and volatile fatty acid (VFA) production. The results revealed that optimal growth occurred when glucose was utilized at 37 °C and initial pH of 7.0, resulting in a final OD₆₀₀ of 2.58. The isolate demonstrated exceptional hydrogen production capabilities, achieving a maximum cumulative hydrogen yield of 2.22 mol-H₂/mol-glucose at 30 °C and pH 7.0, outperforming previously reported E. faecalis strains by 5.84-fold. Kinetic analysis revealed favorable parameters for hydrogen production at 30 °C, with maximum hydrogen production (H_m = 2.63 mol-H₂/mol-glucose), maximum hydrogen production rate (R_m = 0.03 mol-H₂/mol-glucose·h), and lag-phase time (LPT = 18.34 h). Additionally, E. faecalis isolate VT-H1 exhibited significant butyric acid production (63.0 mM) alongside acetic acid (7.0 mM). Genome analysis unveiled the presence of key enzymes associated with hydrogen production, including formate hydrogen lyase (FHL) complex with formate dehydrogenase subunit alpha (FDH-α), bidirectional [NiFe] hydrogenase, NADH-dependent oxidoreductase subunit E, and NADH-quinone oxidoreductase subunit F. Moreover, genes associated with butyric acid production were also identified. These findings highlight the potential of E. faecalis isolate VT-H1 as a promising candidate for sustainable hydrogen and VFA production. Its unique characteristics, distinct genetic profile, and exceptional hydrogen and VFA production capabilities bring exciting opportunities for future biotechnological applications and further research to optimize and scale-up its potential for industrial use.