Novel Syntrophic Isovalerate-Degrading Bacteria and Their Energetic Cooperation with Methanogens in Methanogenic Chemostats

Abstract

Isovalerate is an important intermediate in anaerobic degradation of proteins/amino acids. Little is known about how this compound is degraded due to challenges in cultivation and characterization of isovalerate-degrading bacteria, which are thought to symbiotically depend on methanogenic archaea. In this study, we successfully enriched novel syntrophic isovalerate degraders (uncultivated Clostridiales and Syntrophaceae members) through operation of mesophilic and thermophilic isovalerate-fed anaerobic reactors. Metagenomics- and metatranscriptomics-based metabolic reconstruction of novel putative syntrophic isovalerate metabolizers uncovered the catabolic pathway and byproducts (i.e., acetate, H2, and formate) of isovalerate degradation, mechanisms for electron transduction from isovalerate degradation to H2 and formate generation (via electron transfer flavoprotein; ETF), and biosynthetic metabolism. The identified organisms tended to prefer formate-based interspecies electron transfer with methanogenic partners. The byproduct acetate was further converted to CH4 and CO2 by either Methanothrix (mesophilic) and Methanosarcina (thermophilic), which employed different approaches for acetate degradation. This study presents insights into novel mesophilic and thermophilic isovalerate degraders and their interactions with methanogens.

Chasing the metabolism of novel syntrophic acetate-oxidizing bacteria in thermophilic methanogenic chemostats

“…Moreover, Methanosarcina higher expressed the genes involved in CO2-reducing pathway in ATL compared to PTL and VTL (Tables S3 and S4, [28,30]). These results implied that Methanosarcina played a multi-trophic functional role in thermophilic chemostats.…”

“…The completeness was calculated based on the number of expected marker genes present in MAGs, while the contamination based on the number of expected marker genes present in multiple copies. Genes were annotated using Prokka [81] and manual curation was performed as described previously [30]. Phylogenomic trees were built with PhyloPhlAn v0.99 ("-u" option) [82], and the tree was edited using iTOL [83].…”

“…One effective cultivation-independent approach to studying the physiology and in situ metabolism of uncultured organisms is the combination of metagenomics and metatranscriptomics [17,30,33,34]. In this study, we employed such approach to recover genomes (metagenome-assembled genomes, MAGs) and gene expression profiles of novel potential acetate degraders from fatty-acid-fed thermophilic anaerobic chemostats to investigate their catabolic pathways, energy conservation, and metabolic interactions with their methanogens partner.…”

“…Previous survey on syntrophic acetate metabolizers suggested that H2 has been regarded as electron carrier from acetate oxidizers to methanogens [4,14,27]. Several recent studies also suggested that formate transfer also play a role in propionate [28,29] and isovalerate [30] syntrophic degradation. Direct interspecies electron transfer (DIET) activity has been suggested in enrichment communities degrading propionate and butyrate [31,32].…”

Energy Availability Determines Strategy of Microbial Amino Acid Synthesis in Volatile Fatty Acid–Fed Anaerobic Methanogenic Chemostats

“…Compared with other communities, the available energy is relatively limited in the AMC system and thus plays fundamental roles in driving the evolution of the community members involved ( Lyu et al, 2018 ), as well as being associated with the metabolic interactions among these members ( Jackson and McInerney, 2002 ; Nobu et al, 2020 ). In our previous works, we enriched four AMCs in thermophilic methanogenic chemostats supplemented with acetate, propionate, butyrate, or isovalerate as sole carbon source (accordingly, the four chemostats were simplified as ATL, PTL, BTL, and VTL thereafter) ( Zheng et al, 2019 ; Chen et al, 2020a , b ). Because AA is one type of important public functions that have been reported in many microbial communities [e.g., in ocean ( Barbara and Mitchell, 2003 ) and in human gut ( Perna et al, 2019 ), as well as other AMC systems ( Embree et al, 2015 ; Hubalek et al, 2017 )], we analyzed the abilities of AA synthesis of syntrophic acetate-oxidizing bacteria (SAOBs) involved in our enriched AMCs and found that AA synthesis ability was deficient in most of the SAOBs in our AMCs ( Zeng et al, 2021 ).…”