Metabolic versatility of anaerobic sludge towards platform chemical production from waste glycerol.

Waste glycerol is produced in excess by several industries, such as during biodiesel production. In this work, the metabolic versatility of anaerobic sludge was explored towards waste glycerol valorization. By applying different environmental (methanogenic and sulfate-reducing) conditions, three distinct microbial cultures were obtained from the same inoculum (anaerobic granular sludge), with high microbial specialization, within three different phyla (Thermodesulfobacteriota, Euryarchaeota and Pseudomonadota). The cultures are capable of glycerol conversion through different pathways: (i) glycerol conversion to methane by a bacterium closely related to Solidesulfovibrio alcoholivorans (99.8% 16S rRNA gene identity), in syntrophic relationship with Methanofollis liminatans (98.8% identity), (ii) fermentation to propionate by Propionivibrio pelophilus strain asp66 (98.6% identity), with a propionate yield of 0.88 mmol mmol-1 (0.71 mg mg-1) and a propionate purity of 80-97% and (iii) acetate production coupled to sulfate reduction by Desulfolutivibrio sulfoxidireducens (98.3% identity). In conclusion, starting from the same inoculum, we could drive the metabolic and functional potential of the microbiota towards the formation of several valuable products that can be used in industrial applications or as energy carriers. KEY POINTS: Versatility of anaerobic cultures was explored for waste glycerol valorization Different environmental conditions lead to metabolic specialization Biocommodities such as propionate, acetate and methane were produced.

Alcohol dehydrogenase system acts as the sole pathway for methanol oxidation in Desulfofundulus kuznetsovii strain TPOSR.

Desulfofundulus kuznetsovii is a thermophilic, spore-forming sulphate-reducing bacterium in the family Peptococcaceae. In this study, we describe a newly isolated strain of D. kuznetsovii, strain TPOSR, and compare its metabolism to the type strain D. kuznetsovii 17T. Both strains grow on a large variety of alcohols, such as methanol, ethanol and propane-diols, coupled to the reduction of sulphate. Strain 17T metabolizes methanol via two routes, one involving a cobalt-dependent methyl transferase and the other using a cobalt-independent alcohol dehydrogenase. However, strain TPOSR, which shares 97% average nucleotide identity with D. kuznetsovii strain 17T, lacks several genes from the methyl transferase operon found in strain 17T. The gene encoding the catalytically active methyl transferase subunit B is missing, indicating that strain TPOSR utilizes the alcohol dehydrogenase pathway exclusively. Both strains grew with methanol during cobalt starvation, but growth was impaired. Strain 17T was more sensitive to cobalt deficiency, due to the repression of its methyl transferase system. Our findings shed light on the metabolic diversity of D. kuznetsovii and their metabolic differences of encoding one or two routes for the conversion of methanol.

Moorella caeni sp. nov., isolated from thermophilic anaerobic sludge from a methanol-fed reactor.

Strain AMPT has been previously suggested as a strain of the species Moorella thermoacetica Jiang et al. 2009 (based on the high 16S rRNA gene identity, 98.3 %). However, genome-based phylogenetic analysis of strain AMPT reveals that this bacterium is in fact a novel species of the genus Moorella. Genome relatedness indices between strain AMPT and Moorella thermoacetica DSM 521T were below the minimum threshold values required to consider them members of the same species (digital DNA-DNA hybridization, 52.2 % (<70%); average nucleotide identity, 93.2 % (<95%)). Based on phylogenetic and phenotypic results we recommend that strain AMPT (DSM 21394T=JCM 35360T) should be classified as representing new species, for which we propose the name Moorella caeni sp. nov.

Comparative genomics and proteomics of Eubacterium maltosivorans: functional identification of trimethylamine methyltransferases and bacterial microcompartments in a human intestinal bacterium with a versatile lifestyle.

Eubacterium maltosivorans YIT is a human intestinal isolate capable of acetogenic, propionogenic and butyrogenic growth. Its 4.3-Mb genome sequence contains coding sequences for 4227 proteins, including 41 different methyltransferases. Comparative proteomics of strain YIT showed the Wood-Ljungdahl pathway proteins to be actively produced during homoacetogenic growth on H2 and CO2 while butyrogenic growth on a mixture of lactate and acetate significantly upregulated the production of proteins encoded by the recently identified lctABCDEF cluster and accessory proteins. Growth on H2 and CO2 unexpectedly induced the production of two related trimethylamine methyltransferases. Moreover, a set of 16 different trimethylamine methyltransferases together with proteins for bacterial microcompartments were produced during growth and deamination of the quaternary amines, betaine, carnitine and choline. Growth of strain YIT on 1,2-propanediol generated propionate with propanol and induced the formation of bacterial microcompartments that were also prominently visible in betaine-grown cells. The present study demonstrates that E. maltosivorans is highly versatile in converting low-energy fermentation end-products in the human gut into butyrate and propionate whilst being capable of preventing the formation of the undesired trimethylamine by converting betaine and other quaternary amines in bacterial microcompartments into acetate and butyrate.

Principles, Advances, and Perspectives of Anaerobic Digestion of Lipids.

Several problems associated with the presence of lipids in wastewater treatment plants are usually overcome by removing them ahead of the biological treatment. However, because of their high energy content, waste lipids are interesting yet challenging pollutants in anaerobic wastewater treatment and codigestion processes. The maximal amount of waste lipids that can be sustainably accommodated, and effectively converted to methane in anaerobic reactors, is limited by several problems including adsorption, sludge flotation, washout, and inhibition. These difficulties can be circumvented by appropriate feeding, mixing, and solids separation strategies, provided by suitable reactor technology and operation. In recent years, membrane bioreactors and flotation-based bioreactors have been developed to treat lipid-rich wastewater. In parallel, the increasing knowledge on the diversity of complex microbial communities in anaerobic sludge, and on interspecies microbial interactions, contributed to extend the knowledge and to understand more precisely the limits and constraints influencing the anaerobic biodegradation of lipids in anaerobic reactors. This critical review discusses the most important principles underpinning the degradation process and recent key discoveries and outlines the current knowledge coupling fundamental and applied aspects. A critical assessment of knowledge gaps in the field is also presented by integrating sectorial perspectives of academic researchers and of prominent developers of anaerobic technology.

Anaerobic microbial methanol conversion in marine sediments.

Methanol is an ubiquitous compound that plays a role in microbial processes as a carbon and energy source, intermediate in metabolic processes or as end product in fermentation. In anoxic environments, methanol can act as the sole carbon and energy source for several guilds of microorganisms: sulfate-reducing microorganisms, nitrate-reducing microorganisms, acetogens and methanogens. In marine sediments, these guilds compete for methanol as their common substrate, employing different biochemical pathways. In this review, we will give an overview of current knowledge of the various ways in which methanol reaches marine sediments, the ecology of microorganisms capable of utilizing methanol and their metabolism. Furthermore, through a metagenomic analysis, we shed light on the unknown diversity of methanol utilizers in marine sediments which is yet to be explored.

Comparative proteomics of Geobacter sulfurreducens PCAT in response to acetate, formate and/or hydrogen as electron donor.

Geobacter sulfurreducens is a model bacterium to study the degradation of organic compounds coupled to the reduction of Fe(III). The response of G. sulfurreducens to the electron donors acetate, formate, hydrogen and a mixture of all three with Fe(III) citrate as electron acceptor was studied using comparative physiological and proteomic approaches. Variations in the supplied electron donors resulted in differential abundance of proteins involved in the citric acid cycle (CAC), gluconeogenesis, electron transport, and hydrogenases and formate dehydrogenase. Our results provided new insights into the electron donor metabolism of G. sulfurreducens. Remarkably, formate was the preferred electron donor compared to acetate, hydrogen, or acetate plus hydrogen. When hydrogen was the electron donor, formate was formed, which was associated with a high abundance of formate dehydrogenase. Notably, abundant proteins of two CO2 fixation pathways (acetyl-CoA pathway and the reversed oxidative CAC) corroborated chemolithoautotrophic growth of G. sulfurreducens with formate or hydrogen and CO2 , and provided novel insight into chemolithoautotrophic growth of G. sulfurreducens.

The bacterial sulfur cycle in expanding dysoxic and euxinic marine waters.

Dysoxic marine waters (DMW, < 1 µM oxygen) are currently expanding in volume in the oceans, which has biogeochemical, ecological and societal consequences on a global scale. In these environments, distinct bacteria drive an active sulfur cycle, which has only recently been recognized for open-ocean DMW. This review summarizes the current knowledge on these sulfur-cycling bacteria. Critical bottlenecks and questions for future research are specifically addressed. Sulfate-reducing bacteria (SRB) are core members of DMW. However, their roles are not entirely clear, and they remain largely uncultured. We found support for their remarkable diversity and taxonomic novelty by mining metagenome-assembled genomes from the Black Sea as model ecosystem. We highlight recent insights into the metabolism of key sulfur-oxidizing SUP05 and Sulfurimonas bacteria, and discuss the probable involvement of uncultivated SAR324 and BS-GSO2 bacteria in sulfur oxidation. Uncultivated Marinimicrobia bacteria with a presumed organoheterotrophic metabolism are abundant in DMW. Like SRB, they may use specific molybdoenzymes to conserve energy from the oxidation, reduction or disproportionation of sulfur cycle intermediates such as S0 and thiosulfate, produced from the oxidation of sulfide. We expect that tailored sampling methods and a renewed focus on cultivation will yield deeper insight into sulfur-cycling bacteria in DMW.

Propionate Production from Carbon Monoxide by Synthetic Cocultures of Acetobacterium wieringae and Propionigenic Bacteria.

Gas fermentation is a promising way to convert CO-rich gases to chemicals. We studied the use of synthetic cocultures composed of carboxydotrophic and propionigenic bacteria to convert CO to propionate. So far, isolated carboxydotrophs cannot directly ferment CO to propionate, and therefore, this cocultivation approach was investigated. Four distinct synthetic cocultures were constructed, consisting of Acetobacterium wieringae (DSM 1911T) and Pelobacter propionicus (DSM 2379T), Ac. wieringae (DSM 1911T) and Anaerotignum neopropionicum (DSM 3847T), Ac. wieringae strain JM and P. propionicus (DSM 2379T), and Ac. wieringae strain JM and An. neopropionicum (DSM 3847T). Propionate was produced by all the cocultures, with the highest titer (∼24 mM) being measured in the coculture composed of Ac. wieringae strain JM and An. neopropionicum, which also produced isovalerate (∼4 mM), butyrate (∼1 mM), and isobutyrate (0.3 mM). This coculture was further studied using proteogenomics. As expected, enzymes involved in the Wood-Ljungdahl pathway in Ac. wieringae strain JM, which are responsible for the conversion of CO to ethanol and acetate, were detected; the proteome of An. neopropionicum confirmed the conversion of ethanol to propionate via the acrylate pathway. In addition, proteins related to amino acid metabolism and stress response were highly abundant during cocultivation, which raises the hypothesis that amino acids are exchanged by the two microorganisms, accompanied by isovalerate and isobutyrate production. This highlights the importance of explicitly looking at fortuitous microbial interactions during cocultivation to fully understand coculture behavior. IMPORTANCE Syngas fermentation has great potential for the sustainable production of chemicals from wastes (via prior gasification) and flue gases containing CO/CO2. Research efforts need to be directed toward expanding the product portfolio of gas fermentation, which is currently limited to mainly acetate and ethanol. This study provides the basis for a microbial process to produce propionate from CO using synthetic cocultures composed of acetogenic and propionigenic bacteria and elucidates the metabolic pathways involved. Furthermore, based on proteomics results, we hypothesize that the two bacterial species engage in an interaction that results in amino acid exchange, which subsequently promotes isovalerate and isobutyrate production. These findings provide a new understanding of gas fermentation and a coculturing strategy for expanding the product spectrum of microbial conversion of CO/CO2.

A metabolic and physiological design study of Pseudomonas putida KT2440 capable of anaerobic respiration.

BACKGROUND: Pseudomonas putida KT2440 is a metabolically versatile, HV1-certified, genetically accessible, and thus interesting microbial chassis for biotechnological applications. However, its obligate aerobic nature hampers production of oxygen sensitive products and drives up costs in large scale fermentation. The inability to perform anaerobic fermentation has been attributed to insufficient ATP production and an inability to produce pyrimidines under these conditions. Addressing these bottlenecks enabled growth under micro-oxic conditions but does not lead to growth or survival under anoxic conditions. RESULTS: Here, a data-driven approach was used to develop a rational design for a P. putida KT2440 derivative strain capable of anaerobic respiration. To come to the design, data derived from a genome comparison of 1628 Pseudomonas strains was combined with genome-scale metabolic modelling simulations and a transcriptome dataset of 47 samples representing 14 environmental conditions from the facultative anaerobe Pseudomonas aeruginosa. CONCLUSIONS: The results indicate that the implementation of anaerobic respiration in P. putida KT2440 would require at least 49 additional genes of known function, at least 8 genes encoding proteins of unknown function, and 3 externally added vitamins.

Acetotrophic sulfate-reducing consortia develop active biofilms on zeolite and glass beads in batch cultures at initial pH 3.

Sulfate-reducing microbial communities remain a suitable option for the remediation of acid mine drainage using several types of carrier materials and appropriate reactor configurations. However, acetate prevails as a product derived from the incomplete oxidation of most organic substrates by sulfate reducers, limiting the efficiency of the whole process. An established sulfate-reducing consortium, able to degrade acetate at initial acidic pH (3.0), was used to develop biofilms over granular activated carbon (GAC), glass beads, and zeolite as carrier materials. In batch assays using glycerol, biofilms successfully formed on zeolite, glass beads, and GAC with sulfide production rates of 0.32, 0.26, and 0.14 mmol H2S/L·d, respectively, but only with glass beads and zeolite, acetate was degraded completely. The planktonic and biofilm communities were determined by the 16S rRNA gene analysis to evaluate the microbial selectivity of the carrier materials. In total, 46 OTUs (family level) composed the microbial communities. Ruminococcaceae and Clostridiaceae families were present in zeolite and glass beads, whereas Peptococcaceae was mostly enriched on zeolite and Desulfovibrionaceae on glass beads. The most abundant sulfate reducer in the biofilm of zeolite was Desulfotomaculum sp., while Desulfatirhabdium sp. abounded in the planktonic community. With glass beads, Desulfovibrio sp. dominated the biofilm and the planktonic communities. Our results indicate that both materials (glass beads and zeolite) selected different key sulfate-reducing microorganisms able to oxidize glycerol completely at initial acidic pH, which is relevant for a future application of the consortium in continuous bioreactors to treat acidic streams. KEY POINTS: • Complete consumption of glycerol and acetate at acidic pH by sulfate reduction. • Glass beads and zeolite are suitable materials to form sulfate-reducing biofilms. • Acetotrophic sulfate-reducing bacteria attached to zeolite preferably.

Genome-guided analysis allows the identification of novel physiological traits in Trichococcus species.

BACKGROUND: The genus Trichococcus currently contains nine species: T. flocculiformis, T. pasteurii, T. palustris, T. collinsii, T. patagoniensis, T. ilyis, T. paludicola, T. alkaliphilus, and T. shcherbakoviae. In general, Trichococcus species can degrade a wide range of carbohydrates. However, only T. pasteurii and a non-characterized strain of Trichococcus, strain ES5, have the capacity of converting glycerol to mainly 1,3-propanediol. Comparative genomic analysis of Trichococcus species provides the opportunity to further explore the physiological potential and uncover novel properties of this genus. RESULTS: In this study, a genotype-phenotype comparative analysis of Trichococcus strains was performed. The genome of Trichococcus strain ES5 was sequenced and included in the comparison with the other nine type strains. Genes encoding functions related to e.g. the utilization of different carbon sources (glycerol, arabinan and alginate), antibiotic resistance, tolerance to low temperature and osmoregulation could be identified in all the sequences analysed. T. pasteurii and Trichococcus strain ES5 contain a operon with genes encoding necessary enzymes for 1,3-PDO production from glycerol. All the analysed genomes comprise genes encoding for cold shock domains, but only five of the Trichococcus species can grow at 0 °C. Protein domains associated to osmoregulation mechanisms are encoded in the genomes of all Trichococcus species, except in T. palustris, which had a lower resistance to salinity than the other nine studied Trichococcus strains. CONCLUSIONS: Genome analysis and comparison of ten Trichococcus strains allowed the identification of physiological traits related to substrate utilization and environmental stress resistance (e.g. to cold and salinity). Some substrates were used by single species, e.g. alginate by T. collinsii and arabinan by T. alkaliphilus. Strain ES5 may represent a subspecies of Trichococcus flocculiformis and contrary to the type strain (DSM 2094T), is able to grow on glycerol with the production of 1,3-propanediol.

Multiple and flexible roles of facultative anaerobic bacteria in microaerophilic oleate degradation.

Anaerobic degradation of long-chain fatty acids (LCFA) involves syntrophic bacteria and methanogens, but facultative anaerobic bacteria (FAB) might have a relevant role as well. Here we investigated oleate degradation by a syntrophic synthetic co-culture of Syntrophomonas zehnderi (Sz) and Methanobacterium formicicum (Mf) and FAB (two oleate-degrading Pseudomonas spp. I1 + I2). Sz + Mf were first cultivated in a continuous bioreactor under strict anaerobic conditions. Thereafter, I1 + I2 were inoculated and microaerophilic conditions were provided. Methane and acetate were the main degradation products by Sz + Mf in anaerobiosis and by Sz + Mf + I1 + I2 in microaerophilic conditions. However, acetate production from oleate was higher in microaerophilic conditions (5% O2 ) with the four microorganisms together (0.41 ± 0.07 mmol day-1 ) than in anaerobiosis with Sz + Mf (0.23 ± 0.05 mmol day-1 ). Oleate degradation in batch assays was faster by Sz + Mf + I1 + I2 (under microaerophilic conditions) than by Sz + Mf alone (under strict anaerobic conditions). I1 + I2 were able to grow with oleate and with intermediates of oleate degradation (hydrogen, acetate and formate). This work highlights the importance of FAB, particularly Pseudomonas sp., in anaerobic reactors treating oleate-based wastewater, because they accelerate oleate conversion to methane, by protecting strict anaerobes from oxygen toxicity and also by acting as alternative hydrogen/formate and acetate scavengers for LCFA-degrading anaerobes.

Effect of nickel, cobalt, and iron on methanogenesis from methanol and cometabolic conversion of 1,2-dichloroethene by Methanosarcina barkeri.

Methanogens are responsible for the last step in anaerobic digestion (AD), in which methane (a biofuel) is produced. Some methanogens can cometabolize chlorinated pollutants, contributing for their removal during AD. Methanogenic cofactors involved in cometabolic reductive dechlorination, such as F430 and cobalamin, contain metal ions (nickel, cobalt, iron) in their structure. We hypothesized that the supplementation of trace metals could improve methane production and the cometabolic dechlorination of 1,2-dichloroethene (DCE) by pure cultures of Methanosarcina barkeri. Nickel, cobalt, and iron were added to cultures of M. barkeri growing on methanol and methanol plus DCE. Metal amendment improved DCE dechlorination to vinyl chloride (VC): assays with 20 µM of Fe3+ showed the highest final concentration of VC (5× higher than in controls without Fe3+ ), but also in assays with 5.5 µM of Co2+ and 5 µM of Ni2+ VC formation was improved (3.5-4× higher than in controls without the respective metals). Dosing of metals could be useful to improve anaerobic removal of chlorinated compounds, and more importantly decrease the detrimental effect of DCE on methane production in anaerobic digesters.

Insight into the sulfur metabolism of Desulfurella amilsii by differential proteomics.

Many questions regarding proteins involved in microbial sulfur metabolism remain unsolved. For sulfur respiration at low pH, the terminal electron acceptor is still unclear. Desulfurella amilsii is a sulfur-reducing bacterium that respires elemental sulfur (S0 ) or thiosulfate, and grows by S0 disproportionation. Due to its versatility, comparative studies on D. amilsii may shed light on microbial sulfur metabolism. Requirement of physical contact between cells and S0 was analyzed. Sulfide production decreased by around 50% when S0 was trapped in dialysis membranes, suggesting that contact between cells and S0 is beneficial, but not strictly needed. Proteome analysis was performed under the aforementioned conditions. A Mo-oxidoreductase suggested from genome analysis to act as sulfur reductase was not detected in any growth condition. Thiosulfate and sulfite reductases showed increased abundance in thiosulfate-reducing cultures, while rhodanese-like sulfurtransferases were highly abundant in all conditions. DsrE and DsrL were abundantly detected during thiosulfate reduction, suggesting a modified mechanism of sulfite reduction. Proteogenomics suggest a different disproportionation pathway from what has been reported. This work points to an important role of rhodaneses in sulfur processes and these proteins should be considered in searches for sulfur metabolism in broader fields like meta-omics.

Trichococcus shcherbakoviae sp. nov., isolated from a laboratory-scale anaerobic EGSB bioreactor operated at low temperature.

A new species of the genus Trichococcus, strain Art1T, was isolated from a psychrotolerant syntrophic propionate-oxidizing consortium, obtained before from a low-temperature EGSB reactor fed with a mixture of VFAs (acetate, propionate and butyrate). The 16S rRNA gene sequence of strain Art1T was highly similar to those of other Trichococcus species (99.7-99.9 %) but digital DNA-DNA hybridization values were lower than those recommended for the delineation of a novel species, indicating that strain Art1T is a novel species of the genus Trichococcus. Cells of strain Art1T are non-motile cocci with a diameter of 0.5-2.0 µm and were observed singularly, in pairs, short chains and irregular conglomerates. Cells of Art1T stained Gram-positive and produced extracellular polymeric substances . Growth was optimal at pH 6-7.5 and cells could grow in a temperature range of from -2 to 30 °C (optimum 25-30 °C). Strain Art1T can degrade several carbohydrates, and the main products from glucose fermentation are lactate, acetate, formate and ethanol. The genomic DNA G+C content of strain Art1T is 46.7 %. The major components of the cellular fatty acids are C16 : 1 ω9c, C16 : 0 and C18 : 1 ω9c. Based on genomic and physiological characteristics of strain Art1T, a new species of the genus Trichococcus, Trichococcusshcherbakoviae, is proposed. The type strain of Trichococcusshcherbakoviae is Art1T (=DSM 107162T = VKM B-3260T).

Novel haloalkaliphilic methanotrophic bacteria: An attempt for enhancing methane bio-refinery.

Methane bioconversion into products with a high market value, such as ectoine or hydroxyectoine, can be optimized via isolation of more efficient novel methanotrophic bacteria. The research here presented focused on the enrichment of methanotrophic consortia able to co-produce different ectoines during CH4 metabolism. Four different enrichments (Cow3, Slu3, Cow6 and Slu6) were carried out in basal media supplemented with 3 and 6% NaCl, and using methane as the sole carbon and energy source. The highest ectoine accumulation (∼20 mg ectoine g biomass-1) was recorded in the two consortia enriched at 6% NaCl (Cow6 and Slu6). Moreover, hydroxyectoine was detected for the first time using methane as a feedstock in Cow6 and Slu6 (∼5 mg g biomass-1). The majority of the haloalkaliphilic bacteria identified by 16S rRNA community profiling in both consortia have not been previously described as methanotrophs. From these enrichments, two novel strains (representing novel species) capable of using methane as the sole carbon and energy source were isolated: Alishewanella sp. strain RM1 and Halomonas sp. strain PGE1. Halomonas sp. strain PGE1 showed higher ectoine yields (70-92 mg ectoine g biomass-1) than those previously described for other methanotrophs under continuous cultivation mode (∼37-70 mg ectoine g biomass-1). The results here obtained highlight the potential of isolating novel methanotrophs in order to boost the competitiveness of industrial CH4-based ectoine production.

Comparative proteome analysis of propionate degradation by Syntrophobacter fumaroxidans in pure culture and in coculture with methanogens.

Syntrophobacter fumaroxidans is a sulfate-reducing bacterium able to grow on propionate axenically or in syntrophic interaction with methanogens or other sulfate-reducing bacteria. We performed a proteome analysis of S. fumaroxidans growing with propionate axenically with sulfate or fumarate, and in syntrophy with Methanospirillum hungatei, Methanobacterium formicicum or Desulfovibrio desulfuricans. Special attention was put on the role of hydrogen and formate in interspecies electron transfer (IET) and energy conservation. Formate dehydrogenase Fdh1 and hydrogenase Hox were the main confurcating enzymes used for energy conservation. In the periplasm, Fdh2 and hydrogenase Hyn play an important role in reverse electron transport associated with succinate oxidation. Periplasmic Fdh3 and Fdh5 were involved in IET. The sulfate reduction pathway was poorly regulated and many enzymes associated with sulfate reduction (Sat, HppA, AprAB, DsrAB and DsrC) were abundant even at conditions where sulfate was not present. Proteins similar to heterodisulfide reductases (Hdr) were abundant. Hdr/Flox was detected in all conditions while HdrABC/HdrL was exclusively detected when sulfate was available; these complexes most likely confurcate electrons. Our results suggest that S. fumaroxidans mainly used formate for electron release and that different confurcating mechanisms were used in its sulfidogenic metabolism.

Novel energy conservation strategies and behaviour of Pelotomaculum schinkii driving syntrophic propionate catabolism.

Under methanogenic conditions, short-chain fatty acids are common byproducts from degradation of organic compounds and conversion of these acids is an important component of the global carbon cycle. Due to the thermodynamic difficulty of propionate degradation, this process requires syntrophic interaction between a bacterium and partner methanogen; however, the metabolic strategies and behaviour involved are not fully understood. In this study, the first genome analysis of obligately syntrophic propionate degraders (Pelotomaculum schinkii HH and P. propionicicum MGP) and comparison with other syntrophic propionate degrader genomes elucidated novel components of energy metabolism behind Pelotomaculum propionate oxidation. Combined with transcriptomic examination of P. schinkii behaviour in co-culture with Methanospirillum hungatei, we found that formate may be the preferred electron carrier for P. schinkii syntrophy. Propionate-derived menaquinol may be primarily re-oxidized to formate, and energy was conserved during formate generation through newly proposed proton-pumping formate extrusion. P. schinkii did not overexpress conventional energy metabolism associated with a model syntrophic propionate degrader Syntrophobacter fumaroxidans MPOB (i.e., CoA transferase, Fix and Rnf). We also found that P. schinkii and the partner methanogen may also interact through flagellar contact and amino acid and fructose exchange. These findings provide new understanding of syntrophic energy acquisition and interactions.

Eubacterium maltosivorans sp. nov., a novel human intestinal acetogenic and butyrogenic bacterium with a versatile metabolism.

A novel anaerobic, non-spore-forming bacterium was isolated from a faecal sample of a healthy adult. The isolate, designated strain YIT, was cultured in a basal liquid medium under a gas phase of H2/CO2 supplemented with yeast extract (0.1 g l-1). Cells of strain YIT were short rods (0.4-0.7×2.0-2.5 µm), appearing singly or in pairs, and stained Gram-positive. Catalase activity and gelatin hydrolysis were positive while oxidase activity, indole formation, urease activity and aesculin hydrolysis were negative. Growth was observed within a temperature range of 20-45 °C (optimum, 35-37 °C), and a pH range of 5.0-8.0 (optimum pH 7.0-7.5). Doubling time was 2.3 h when grown with glucose at pH 7.2 and 37 °C. Besides acetogenic growth, the isolate was able to ferment a large range of monomeric sugars with acetate and butyrate as the main end products. Strain YIT did not show respiratory growth with sulfate, sulfite, thiosulfate or nitrate as electron acceptors. The major cellular fatty acids of the isolate were C16 : 0 and C18 : 0. The genomic DNA G+C content was 47.8 mol%. Strain YIT is affiliated to the genus Eubacterium, sharing highest levels of 16S rRNA gene similarity with Eubacterium limosum ATCC 8486T (97.3 %), Eubacterium callanderi DSM 3662T (97.5 %), Eubacterium aggregans DSM 12183T (94.4 %) and Eubacterium barkeri DSM 1223T (94.8 %). Considering its physiological and phylogenetic characteristics, strain YIT represents a novel species within the genus Eubacterium, for which the name Eubacterium maltosivorans sp. nov. is proposed. The type strain is YIT (=DSM 105863T=JCM 32297T).