Abstract
This study describes the dynamics and complexity of microbial communities producing hydrogen-rich fermentation gas from sugar-beet molasses in five packed-bed reactors (PBRs). The bioreactors constitute a part of a system producing hydrogen from the by-products of the sugar-beet industry that has been operating continuously in one of the Polish sugar factories. PBRs with different working volumes, packing materials, construction and inocula were tested. This study focused on analysis (based on 16S rRNA profiling and shotgun metagenomics sequencing) of the microbial communities selected in the PBRs under the conditions of high (>100 cm3/g COD of molasses) and low (<50 cm3/g COD of molasses) efficiencies of hydrogen production. The stability and efficiency of the hydrogen production are determined by the composition of dark fermentation microbial communities. The most striking difference between the tested samples is the ratio of hydrogen producers to lactic acid bacteria. The highest efficiency of hydrogen production (130–160 cm3/g COD of molasses) was achieved at the ratios of HPB to LAB ≈ 4:2.5 or 2.5:1 as determined by 16S rRNA sequencing or shotgun metagenomics sequencing, respectively. The most abundant Clostridium species were C. pasteurianum and C. tyrobutyricum. A multiple predominance of LAB over HPB (3:1–4:1) or clostridia over LAB (5:1–60:1) results in decreased hydrogen production. Inhibition of hydrogen production was illustrated by overproduction of short chain fatty acids and ethanol. Furthermore, concentration of ethanol might be a relevant marker or factor promoting a metabolic shift in the DF bioreactors processing carbohydrates from hydrogen-yielding toward lactic acid fermentation or solventogenic pathways. The novelty of this study is identifying a community balance between hydrogen producers and lactic acid bacteria for stable hydrogen producing systems. The balance stems from long-term selection of hydrogen-producing microbial community, operating conditions such as bioreactor construction, packing material, hydraulic retention time and substrate concentration. This finding is confirmed by additional analysis of the proportions between HPB and LAB in dark fermentation bioreactors from other studies. The results contribute to the advance of knowledge in the area of relationships and nutritional interactions especially the cross-feeding of lactate between bacteria in dark fermentation microbial communities.
Highlights
Hydrolysis and acidogenesis are the two initial steps of anaerobic digestion whereby complex organic matter is decomposed and processed mainly to short-chain fatty acids, alcohols, carbon dioxide and hydrogen, the identity and proportion of which depend on the type of fermentation
As it has been highlighted in a recent excellent review paper (Castelló et al, 2020), hydrogen-producing microbial communities show (i) a relatively low diversity in comparison to methanogenic communities and (ii) tendencies to change, which is reflected in the instability of hydrogen production
The differences observed in the composition of the microbial communities with unstable hydrogen production are not related to the emergence of new taxa of bacteria
Summary
Hydrolysis and acidogenesis are the two initial steps of anaerobic digestion whereby complex organic matter is decomposed and processed mainly to short-chain fatty acids, alcohols, carbon dioxide and hydrogen, the identity and proportion of which depend on the type of fermentation. Dark fermentation (DF) is a part of the acidogenic step of anaerobic digestion which involves conversion of an organic substrate to hydrogen, carbon dioxide and non-gaseous products including acetic and butyric acids. In the clostridial-type fermentation, pyruvate:ferredoxin oxidoreductase (PFOR) oxidizes pyruvate to acetyl-CoA in the presence of ferredoxin (Fd) that is simultaneously reduced. In the Enterobacter-type fermentation, hydrogen can be formed in the reactions of NADH oxidation by NFOR as described for the clostridial-type fermentation (Seppälä et al, 2011; Nasr et al, 2017; Maeda et al, 2018)
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