Metatranscriptomic and thermodynamic insights into medium-chain fatty acid production using an anaerobic microbiome

Abstract

Importance 46Mixed communities of microbes play important roles in health, the environment, 47 agriculture, and biotechnology. While tapping the combined activity of organisms within 48 microbiomes may have advantages over the use of pure cultures for biomanufacturing, 49 harnessing the metabolism of these mixed cultures remains a major challenge. Here, we 50 predict metabolic functions of bacteria in a microbiome that produces medium-chain fatty 51 acids from a renewable feedstock. Our approach and findings can be used to engineer and 52 control microbiomes for improved biomanufacturing; to build synthetic mixtures of 53 microbes that produce valuable chemicals from renewable resources; and to better 54 understand microbial communities that contribute to health, agriculture, and the 55 environment. 56 microorganisms (1). In addition, MCFA producing bioreactors contain diverse communities 80 with members affiliated with the Firmicutes, particularly Clostridia and Lactobacilli, having 81 high abundance (4, 5, 11). However, the specific roles of the abundant microbiome 82 members are not well understood. 83Here, we utilize a combination of metagenomic, metatranscriptomic , pathway 84 modeling, and thermodynamic analyses to reconstruct the combined metabolic activity of a 85 microbiome converting lignocellulosic conversion residues to C6 and C8. In total, 37 high 86 quality draft metagenome-assembled genomes (MAGs) were assembled, and the gene 87 expression patterns of those representing the ten most abundant community members 88 during steady-state reactor operation were analyzed. Our results identify several MAGs in 89 the community that expressed genes predicted to be involved in complex carbohydrate 90 degradation and in the subsequent fermentation of degradation products to lactate and 91 acetate. Genes encoding enzymes capable of producing C6 and C8 from these fermentation 92 products were expressed by two abundant MAGs affiliated with the class Clostridia. Based 93 on a thermodynamic analysis of the proposed MCFA-producing pathways, we predict that 94 individual Clostridial MAGs use different substrates for MCFA production (lactate, versus a 95 combination of xylose, H2, and acetate). We also describe how new insights into MCFA 96 production could improve the biomanufacturing productivity of microbiomes. 97 Results 98 Microbiome characterization 99We previously described the establishment of a microbiome that produces MCFA in 100 a bioreactor that is continuously fed with the residues from lignocellulosic ethanol 101 production (4). The reactor feed contained high amounts of xylose, carbohydrate 102 6 oligomers, and uncharacterized organic matter (Fig. 1). Following an initial acclimation 103 period (Day 0 -Day 48), the quantity of produced organic acids stabilized (Fig. 1). To gain 104 insight into the microbial activities that were associated with this MCFA-producing 105 microbiome, samples were collected for metagenomic analysis at five different times (Days 106 12, 48, 84, 96, and 120), and RNA was pre...

Analysis of Human Gut Microbiome: Taxonomy and Metabolic Functions in Thai Adults

“…These observed results were consistent with the metabolic traits of butyrate-producing bacteria for butyrate production, which are normally identified in gut microbiome in healthy adults [32]. Moreover, we also found that Roseburia exhibited a high number of genes involved in xylulose-5-phosphate/fructose-6-phosphate phosphoketolase (Xfp, ECs: 4.1.2.9 and 4.1.2.22), which is the key catalytic enzyme in the hemicellulosic complex oligosaccharides fermentation pathway [34]. This finding is consistent with the most recent genome-scale metabolic reconstruction of the carbohydrate degradation and utilization pathways of Roseburia, which demonstrated the ability of Roseburia species to ferment pentoses (e.g., xylose and arabinose) and oligosaccharides (e.g., xylooligosaccharides, arabinoxylans and arabinogalactans) as carbon sources for growth through Xfp [35].…”

Temperature modulates stress response in mainstream anammox reactors

“…Relative transcript abundances as a proxy for expression levels were calculated from the total filtered mRNA reads from 56 RNA samples, which were individually mapped against the recovered MAGs, normalized for respective genome length and sequencing depth and expressed as transcripts per million (TPM) (Supplementary Table 2 ) 19 , 43 . The two most abundant anammox MAGs (~18% relative abundance) together were significantly ( p < 0.05, Student’s t -test) responsible for the highest transcriptional activity during the 20 °C and 14 °C regimes, respectively (Fig.…”

Product Inhibition and pH Affect Stoichiometry and Kinetics of Chain Elongating Microbial Communities in Sequencing Batch Bioreactors

“…Beside substrate uptake kinetics, SBRs can be used for the fundamental understanding and mathematical description of the effect of product toxicity and changing ethanol-to-acetate ratios on chain elongating organisms in dynamic environments. Omics techniques in combination with metabolic modeling could be used to identify labor division in microbial communities fed with a complex substrate and gain a deeper understanding of the effect of environmental conditions on the metabolic flux though microbial communities ( Scarborough et al, 2018 ). Lastly, integrated reactor setups can be exploited for testing the industrial potential of different cultivation strategies.…”