Hydrogen metabolic patterns driven by Clostridium-Streptococcus community shifts in a continuous stirred tank reactor.

The hydrogen (H2) production efficiency in dark fermentation systems is strongly dependent on the occurrence of metabolic pathways derived from the selection of microbial species that either consume molecular H2 or outcompete hydrogenogenic bacteria for the organic substrate. In this study, the effect of organic loading rate (OLR) on the H2 production performance, the metabolic pathways, and the microbial community composition in a continuous system was evaluated. Two bacterial genera, Clostridium and Streptococcus, were dominant in the microbial community depending on the OLR applied. At low OLR (14.7-44.1 gLactose/L-d), Clostridium sp. was dominant and directed the system towards the acetate-butyrate fermentation pathway, with a maximum H2 yield of 2.14 molH2/molHexose obtained at 29.4 gLactose/L-d. Under such conditions, the volumetric hydrogen production rate (VHPR) was between 3.2 and 11.6 LH2/L-d. In contrast, relatively high OLR (58.8 and 88.2 gLactose/L-d) favored the dominance of Streptococcus sp. as co-dominant microorganism leading to lactate production. Under these conditions, the formate production was also stimulated serving as a strategy to dispose the surplus of reduced molecules (e.g., NADH2+), which theoretically consumed up to 5.72 LH2/L-d. In such scenario, the VHPR was enhanced (13.7-14.5 LH2/L-d) but the H2 yield dropped to a minimum of 0.74 molH2/molHexose at OLR = 58.8 gLactose/L-d. Overall, this research brings clear evidence of the intrinsic occurrence of metabolic pathways detrimental for biohydrogen production, i.e., lactic acid fermentation and formate production, suggesting the use of low OLR as a strategy to control them.