Jeon, B. S. | bioRxiv Science

bioRxiv2025

Steering the chain-elongating microbiome to specific medium-chain carboxylates with ethanol and lactate as co-electron donors: maximizing C8 or C6

Microbial chain elongation is a sustainable process to convert organic residues into valuable biochemicals via anaerobic fermentation. The operating conditions for the bioreactor and the ecological interactions among functional populations are crucial in controlling this process, but they have not been completely ascertained. Here, we unraveled two operating conditions (i.e., environmental factors): (1) the substrate ratio of ethanol and lactate as co-electron donors, and (2) the temperature, which affected both the product specificity (i.e., function) and microbial dynamics of chain-elongating microbiomes in a continuously fed bioreactor with product extraction. Specifically, we found that the increase in the substrate ratio of ethanol to lactate shifted the microbiomes toward n-caprylate (C8) production, while the slightly higher operating temperatures of 37{degrees}C or 42{degrees}C were advantageous to n-caproate (C6) production. We detected a core microbiome that was similar for all environmental conditions and the two bioreactors, consisting of populations from Sphaerochaeta spp., Caproiciproducens spp., and Oscillospiraceae. Besides the core microbiome, we observed positive correlations between Erysipelaclostridiaceae UCG-004, Bacteroides spp., Oscillospiraceae NK4A214, Rikenellaceae RC9, and Pseudoclavibacter spp. with n-caprylate production. Similar populations compared to the core microbiome were positively correlated with n-caproate production. We showed that we can steer microbiomes toward a high specificity of certain medium-chain carboxylates. Abstract Graphics O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=134 SRC="FIGDIR/small/637006v1_ufig1.gif" ALT="Figure 1"> View larger version (60K): org.highwire.dtl.DTLVardef@70557eorg.highwire.dtl.DTLVardef@fcee81org.highwire.dtl.DTLVardef@18e64feorg.highwire.dtl.DTLVardef@1888a8d_HPS_FORMAT_FIGEXP M_FIG C_FIG SYNOPSISGenerating medium-chain carboxylates is a promising open-culture biotechnology production platform for converting organic waste streams into biofuels and chemicals.

bioengineering↗

bioRxiv2024

The gases H2 and O2 in open-culture reactors influence the performance and microbiota of chain elongation into n-caproate and n-caprylate.

Medium-chain carboxylates are used in various industrial applications. These chemicals are typically extracted from palm oil, which is deemed not sustainable. Recent research has focused on microbial chain elongation using reactors to produce medium-chain carboxylates, such as n-caproate (C6) and n-caprylate (C8), from organic substrates such as wastes. Even though the production of n-caproate is relatively well-characterized, bacteria and metabolic pathways that are responsible for n-caprylate production are not. Here, three 5-L reactors with continuous membrane-based liquid-liquid extraction (i.e., pertraction) were fed ethanol and acetate and operated for an operating period of 234 days with different operating conditions. Metagenomic and metaproteomic analyses were employed. n-Caprylate production rates and reactor microbiomes differed between reactors even when operated similarly due to differences in H2 and O2 between the reactors. The complete reverse {beta}-oxidation pathway was present and expressed by several bacterial species in the Clostridia class. Several Oscillibacter spp., including Oscillibacter valericigenes, were positively correlated with n-caprylate production rates, while Clostridium kluyveri was positively correlated with n-caproate production. Pseudoclavibacter caeni, which is a strictly aerobic bacterium, was abundant across all the operating periods, regardless of n-caprylate production rates. This study provides insight into microbiota that are associated with n-caprylate production in open-culture reactors and provides ideas for further work. ImportanceMicrobial chain elongation pathways in open-culture biotechnology systems can be utilized to convert organic waste and industrial side streams into valuable industrial chemicals. Here, we investigated the microbiota and metabolic pathways that produce medium-chain carboxylates, including n-caproate (C6) and n-caprylate (C8), in reactors with in-line product extraction. Although the reactors in this study were operated similarly, different microbial communities dominated and were responsible for chain elongation. We found that different microbiota were responsible for n-caproate or n-caprylate production, and this can inform engineers on how to operate the systems better. We also observed which changes in operating conditions steered the production toward and away from n-caprylate, but more work is necessary to ascertain a mechanistic understanding that could be predictive. This study provides pertinent research questions for future work.

microbiology↗