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Sporomusa ovata is a Gram-negative acetogen of the Sporomusaceae family with a unique physiology. This anerobic bacterium is a core microbial catalyst for advanced CO 2 -based biotechnologies including gas fermentation, microbial electrosynthesis, and hybrid photosystem. Until now, no genetic tools exist for S. ovata , which is a critical obstacle to its optimization as an autotrophic chassis and the acquisition of knowledge about its metabolic capacities. Here, we developed an electroporation protocol for S. ovata . With this procedure, it became possible to introduce replicative plasmids such as pJIR751 and its derivatives into the acetogen. This system was then employed to demonstrate the feasibility of heterologous expression by introducing a functional β-glucuronidase enzyme under the promoters of different strengths in S. ovata . Next, a recombinant S. ovata strain producing the non-native product acetone both from an organic carbon substrate and from CO 2 was constructed. Finally, a replicative plasmid capable of integrating itself on the chromosome of the acetogen was developed as a tool for genome editing, and gene deletion was demonstrated. These results indicate that S. ovata can be engineered and provides a first-generation genetic toolbox for the optimization of this biotechnological workhorse.IMPORTANCE S. ovata harbors unique features that make it outperform most microbes for autotrophic biotechnologies such as a capacity to acquire electrons from different solid donors, a low H 2 threshold, and efficient energy conservation mechanisms. The development of the first-generation genetic instruments described in this study is a key step toward understanding the molecular mechanisms involved in these outstanding metabolic and physiological characteristics. In addition, these tools enable the construction of recombinant S. ovata strains that can synthesize a wider range of products in an efficient manner., Competing Interests: The authors declare no conflict of interest.

We have previously demonstrated abnormal gut microbial composition in castration-resistant prostate cancer (CRPC) patients, here we revealed the mechanism of gut microbiota-derived short-chain fatty acids (SCFAs) as a mediator linking CRPC microbiota dysbiosis and prostate cancer (PCa) progression. By using transgenic TRAMP mouse model, PCa patient samples, in vitro PCa cell transwell and macrophage recruitment assays, we examined the effects of CRPC fecal microbiota transplantation (FMT) and SCFAs on PCa progression. Our results showed that FMT with CRPC patients' fecal suspension increased SCFAs-producing gut microbiotas such as Ruminococcus, Alistipes, Phascolarctobaterium in TRAMP mice, and correspondingly raised their gut SCFAs (acetate and butyrate) levels. CRPC FMT or SCFAs supplementation significantly accelerated mice's PCa progression. In vitro, SCFAs enhanced PCa cells migration and invasion by inducing TLR3-triggered autophagy that further activated NF-κB and MAPK signalings. Meanwhile, autophagy of PCa cells released higher level of chemokine CCL20 that could reprogramme the tumor microenvironment by recruiting more macrophage infiltration and simultaneously polarizing them into M2 type, which in turn further strengthened PCa cells invasiveness. Finally in a cohort of 362 PCa patients, we demonstrated that CCL20 expression in prostate tissue was positively correlated with Gleason grade, pre-operative PSA, neural/seminal vesical invasion, and was negatively correlated with post-operative biochemical recurrence-free survival. Collectively, CRPC gut microbiota-derived SCFAs promoted PCa progression via inducing cancer cell autophagy and M2 macrophage polarization. CCL20 could become a biomarker for prediction of prognosis in PCa patients. Intervention of SCFAs-producing microbiotas may be a useful strategy in manipulation of CRPC., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Inc.)

The involvement of host immunity in the gut microbiota-mediated colonization resistance to Clostridioides difficile infection (CDI) is incompletely understood. Here, we show that interleukin (IL)-22, induced by colonization of the gut microbiota, is crucial for the prevention of CDI in human microbiota-associated (HMA) mice. IL-22 signaling in HMA mice regulated host glycosylation, which enabled the growth of succinate-consuming bacteria Phascolarctobacterium spp. within the gut microbiome. Phascolarctobacterium reduced the availability of luminal succinate, a crucial metabolite for the growth of C. difficile, and therefore prevented the growth of C. difficile. IL-22-mediated host N-glycosylation is likely impaired in patients with ulcerative colitis (UC) and renders UC-HMA mice more susceptible to CDI. Transplantation of healthy human-derived microbiota or Phascolarctobacterium reduced luminal succinate levels and restored colonization resistance in UC-HMA mice. IL-22-mediated host glycosylation thus fosters the growth of commensal bacteria that compete with C. difficile for the nutritional niche.

Background Pediatric hypertension is recognized as an emerging global health concern. Although new guidelines are developed for facilitating clinical management, the reasons for the prevalence of hypertension in children remain unknown. Genetics and environmental factors do not fully account for the growing incidence of pediatric hypertension. Because stable bacterial flora in early life are linked with health outcomes later in life, we hypothesized that reshaping of gut microbiota in early life affects blood pressure (BP) of pediatric subjects. Methods and Results To test this hypothesis, we administered amoxicillin, the most commonly prescribed pediatric antibiotic, to alter gut microbiota of young, genetically hypertensive rats (study 1) and dams during gestation and lactation (study 2) and recorded their BP. Reshaping of microbiota with reductions in Firmicutes/Bacteriodetes ratio were observed. Amoxicillin treated rats had lower BP compared with untreated rats. In young rats treated with amoxicillin, the lowering effect on BP persisted even after antibiotics were discontinued. Similarly, offspring from dams treated with amoxicillin showed lower systolic BP compared with control rats. Remarkably, in all cases, a decrease in BP was associated with lowering of Veillonellaceae, which are succinate-producing bacteria. Elevated plasma succinate is reported in hypertension. Accordingly, serum succinate was measured and found lower in animals treated with amoxicillin. Conclusions Our results demonstrate a direct correlation between succinate-producing gut microbiota and early development of hypertension and indicate that reshaping gut microbiota, especially by depleting succinate-producing microbiota early in life, may have long-term benefits for hypertension-prone individuals.

Microbial electrosynthesis is a bioprocess where microbes reduce CO 2 into multicarbon chemicals with electrons derived from the cathode of a bioelectrochemical reactor. Developing a highly productive microbial electrosynthesis reactor requires excellent electrical connection between the electrochemical setup, the cathode, and the microbes. Copper is a highly conductive cathode material widely employed in electrochemical apparatuses. However, the antimicrobial properties of copper limit its usage for bioelectrochemistry. Here, biocompatible reduced graphene oxide coated on copper foam is synthesized as a cathode material for the microbial electrosynthesis of acetate from CO 2 . Dense and electroactive Sporomusa ovata biofilms form on the surface of reduced graphene oxide-coated copper foam electrodes while only scattered and damaged cells cover uncoated copper electrodes. Besides the formation of metabolically-active biofilms, acetate production rate from CO 2 is 21.3 and 43.5-fold higher with this novel composite cathode compared with an uncoated copper foam cathode and a reversed cathode made of reduced graphene oxide foam coated with copper, respectively. The results demonstrate that reduced graphene oxide can be employed as a biocompatible and conductive buffer between microbes and bactericidal electrode materials with excellent electrochemical property to enable highly performant microbial electrosynthesis., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Maternal dietary supplementation during gestation and lactation improves the health of piglets. The purpose of this study was to determine the effect of different levels of methionine (Met) supplementation in the sows' diet during late gestation and lactation on piglets. Thirty sows were randomly divided into three groups and fed the following diets from day 90 of gestation to day 21 of lactation: (a) control group (a basal diet containing 0.36% Met), (b) 0.48% Met group (a basal diet with additional 0.12% Met), and (c) 0.60% Met group (a basal diet with additional 0.24% Met). On day 21 after farrowing, piglets of average body weight (n = 10 per group) were selected for sample collection. The results showed that the 0.48% Met and 0.60% Met diets significantly lowered the malondialdehyde content in the piglets' serum (P < 0.05). In addition, the glutathione peroxidase content was significantly increased in the 0.48% Met group (P < 0.05) and the total glutathione content was significantly reduced in the 0.60% Met group (P < 0.05) compared to the control group. Furthermore, Met supplementation of the sows' diet was associated with alterations in 37 plasma metabolites in the piglets. In the piglets' intestinal microbiota, the relative abundances of Phascolarctobacterium and Bacteroidetes in the 0.48% Met group were higher than those in the other two groups (P < 0.05). Our results suggest that a diet including 0.48% Met during late gestation and lactation can maintain the health of piglets by increasing the antioxidant capacity and changing the intestinal microbiota composition, but a higher level of Met supplementation may increase the potential risk to piglets.

Adaptive laboratory evolution (ALE) is an approach enabling the development of novel characteristics in microbial strains via the application of a constant selection pressure. This method is also an efficient tool to acquire insights on molecular mechanisms responsible for specific phenotypes. ALE experiments have mainly been conducted with heterotrophic microbes to study, for instance, cell metabolism with different multicarbon substrates, tolerance to solvents, pH variation, and high temperature. Here, we describe employing an ALE method to generate Sporomusa ovata strains growing faster autotrophically and reducing CO 2 into acetate more efficiently. Strains developed via this ALE method were also used to gain knowledge on the autotrophic metabolism of S. ovata as well as other acetogenic bacteria.

The objective of this study was to determine whether a combination of crude glycerin (CG) and soyabean oil (SO) could be used to partially replace maize in the diet of Nellore steers while maintaining optimum feed utilisation. Eight castrated Nellore steers fitted with ruminal and duodenal cannulas were used in a double 4×4 Latin square design balanced for residual effects, in a factorial arrangement (A×B), when factor A corresponded to the provision of SO, and factor B to the provision of CG. Steers feed SO and CG showed similar DM intake, DM, organic matter and neutral-detergent fibre digestibility to that of steers fed diets without oil and without glycerine (P>0·05). Both diets with CG additions reduced the acetate:propionate ratio and increased the proportion of iso-butyrate, butyrate, iso-valerate and valerate (P<0·05). Steers fed diets containing SO had less total N excretion (P<0·001) and showed greater retained N expressed as % N intake (P=0·022). SO and CG diet generated a greater ruminal abundance of Prevotella, Succinivibrio, Ruminococcus, Syntrophococcus and Succiniclasticum. Archaea abundance (P=0·002) and total ciliate protozoa were less in steers fed diets containing SO (P=0·011). CG associated with lipids could be an energy source, which is a useful strategy for the partial replacement of maize in cattle diets, could result in reduced total N excretion and ruminal methanogens without affecting intake and digestibility.

Acetogenic bacteria (i.e., acetogens) produce acetate from CO 2 during anaerobic chemoautotrophic growth. Because acetogens fix CO 2 with high energy efficiency, they have been investigated as biocatalysts of CO 2 conversion into valuable chemicals. Recent studies revealed that some acetogens are capable of extracellular electron transfer (EET), which enables electron exchange between microbial cells and extracellular solid materials. Thus, acetogens are promising candidates as biocatalysts in recently developed bioelectrochemical technologies, including microbial electrosynthesis (MES), in which useful chemicals are biologically produced from CO 2 using electricity as the energy source. In microbial photoelectrosynthesis, a variant of MES technology, the conversion of CO 2 into organic compounds is achieved using light as the sole energy source without an external power supply. In this mini-review, we introduce the general features of bioproduction and EET of acetogens and describe recent progress and future prospects of MES technologies based on the EET capability of acetogens.

The aim of this study is to obtain a systems-level understanding of the interactions between Dehalococcoides and corrinoid-supplying microorganisms by analyzing community structures and functional compositions, activities, and dynamics in trichloroethene (TCE)-dechlorinating enrichments. Metagenomes and metatranscriptomes of the dechlorinating enrichments with and without exogenous cobalamin were compared. Seven putative draft genomes were binned from the metagenomes. At an early stage (2 days), more transcripts of genes in the Veillonellaceae bin-genome were detected in the metatranscriptome of the enrichment without exogenous cobalamin than in the one with the addition of cobalamin. Among these genes, sporulation-related genes exhibited the highest differential expression when cobalamin was not added, suggesting a possible release route of corrinoids from corrinoid producers. Other differentially expressed genes include those involved in energy conservation and nutrient transport (including cobalt transport). The most highly expressed corrinoid de novo biosynthesis pathway was also assigned to the Veillonellaceae bin-genome. Targeted quantitative PCR (qPCR) analyses confirmed higher transcript abundances of those corrinoid biosynthesis genes in the enrichment without exogenous cobalamin than in the enrichment with cobalamin. Furthermore, the corrinoid salvaging and modification pathway of Dehalococcoides was upregulated in response to the cobalamin stress. This study provides important insights into the microbial interactions and roles played by members of dechlorinating communities under cobalamin-limited conditions. IMPORTANCE The key chloroethene-dechlorinating bacterium Dehalococcoides mccartyi is a cobalamin auxotroph, thus acquiring corrinoids from other community members. Therefore, it is important to investigate the microbe-microbe interactions between Dehalococcoides and the corrinoid-providing microorganisms in a community. This study provides systems-level information, i.e., taxonomic and functional compositions and dynamics of the supportive microorganisms in dechlorinating communities under different cobalamin conditions. The findings shed light on the important roles of Veillonellaceae species in the communities compared to other coexisting community members in producing and providing corrinoids for Dehalococcoides species under cobalamin-limited conditions., (Copyright © 2017 American Society for Microbiology.)

The Sporomusa genus comprises anaerobic spore-forming acetogenic bacteria that stain Gram-negative. Sporomusa species typically grow with one-carbon substrates and N-methylated compounds. In the degradation of these compounds methyltransferases are involved. In addition, Sporomusa species can grow autotrophically with H2 and CO2 , and use a variety of sugars for acetogenic growth. Here we describe a genome analysis of Sporomusa strain An4 and a proteome analysis of cells grown under five different conditions. Comparison of the genomes of Sporomusa strain An4 and Sporomusa ovata strain H1 indicated that An4 is a S. ovata strain. Proteome analysis showed a high abundance of several methyltransferases, predominantly trimethylamine methyltransferases, during growth with betaine, whereas trimethylamine is one of the main end-products of betaine degradation. In methanol degradation methyltransferases are also involved. In methanol-utilizing methanogens, two methyltransferases catalyse methanol conversion, methyltransferase 1 composed of subunits MtaB and MtaC and methyltransferase 2, also called MtaA. The two methyltransferase 1 subunits MtaB and MtaC were highly abundant when strain An4 was grown with methanol. However, instead of MtaA a methyltetrahydrofolate methyltransferase was synthesized. We propose a novel methanol degradation pathway in Sporomusa strain An4 that uses a methyltetrahydrofolate methyltransferase instead of MtaA., (© 2015 Society for Applied Microbiology and John Wiley & Sons Ltd.)

A co-culture system comprising an acetogenic bacterium, Sporomusa ovata DSMZ2662, and a denitrifying bacterium, Pseudomonas stutzeri JCM20778, enabled denitrification using H2 as the sole external electron donor and CO2 as the sole external carbon source. Acetate produced by S. ovata supported the heterotrophic denitrification of P. stutzeri. A nitrogen balance study showed the reduction of nitrate to nitrogen gas without the accumulation of nitrite and nitrous oxide in the co-culture system. S. ovata did not show nitrate reduction to ammonium in the co-culture system. Significant proportions of the consumed H2 were utilized for denitrification: 79.9 ± 4.6% in the co-culture system containing solid-phase humin and 62.9±11.1% in the humin-free co-culture system. The higher utilization efficiency of hydrogen in the humin-containing system was attributed to the higher denitrification activity of P. stutzeri under the acetate deficient conditions. The nitrogen removal rate of the humin-containing co-culture system reached 0.19 kg NO3(-)-N·m(-3)·d(-1). Stable denitrification activity for 61 days of successive sub-culturing suggested the robustness of this co-culture system. This study provides a novel strategy for the in situ enhancement of microbial denitrification., (Copyright © 2016 The Society for Biotechnology, Japan. Published by Elsevier B.V. All rights reserved.)

To provide new insight into the dark fermentation process, a multi-lateral study was performed to study the microbiology of 20 different lab-scale bioreactors operated in four different countries (Brazil, Chile, Mexico, and Uruguay). Samples (29) were collected from bioreactors with different configurations, operation conditions, and performances. The microbial communities were analyzed using 16S rRNA genes 454 pyrosequencing. The results showed notably uneven communities with a high predominance of a particular genus. The phylum Firmicutes predominated in most of the samples, but the phyla Thermotogae or Proteobacteria dominated in a few samples. Genera from three physiological groups were detected: high-yield hydrogen producers (Clostridium, Kosmotoga, Enterobacter), fermenters with low-hydrogen yield (mostly from Veillonelaceae), and competitors (Lactobacillus). Inocula, reactor configurations, and substrates influence the microbial communities. This is the first joint effort that evaluates hydrogen-producing reactors and operational conditions from different countries and contributes to understand the dark fermentation process.

Cobalamin and other corrinoids are essential cofactors for many organisms. The majority of microbes with corrinoid-dependent enzymes do not produce corrinoids de novo, and instead must acquire corrinoids produced by other organisms in their environment. However, the profile of corrinoids produced in corrinoid-dependent microbial communities, as well as the exchange and modification of corrinoids among community members have not been well studied. In this study, we applied a newly developed liquid chromatography tandem mass spectrometry-based corrinoid detection method to examine relationships among corrinoids, their lower ligand bases and specific microbial groups in microbial communities containing Dehalococcoides mccartyi that has an obligate requirement for benzimidazole-containing corrinoids for trichloroethene respiration. We found that p-cresolylcobamide ([p-Cre]Cba) and cobalamin were the most abundant corrinoids in the communities. It suggests that members of the family Veillonellaceae are associated with the production of [p-Cre]Cba. The decrease of supernatant-associated [p-Cre]Cba and the increase of biomass-associated cobalamin were correlated with the growth of D. mccartyi by dechlorination. This supports the hypothesis that D. mccartyi is capable of fulfilling its corrinoid requirements in a community through corrinoid remodelling, in this case, by importing extracellular [p-Cre]Cba and 5,6-dimethylbenzimidazole (DMB) (the lower ligand of cobalamin), to produce cobalamin as a cofactor for dechlorination. This study also highlights the role of DMB, the lower ligand produced in all of the studied communities, in corrinoid remodelling. These findings provide novel insights on roles played by different phylogenetic groups in corrinoid production and corrinoid exchange within microbial communities. This study may also have implications for optimizing chlorinated solvent bioremediation., (© 2014 Society for Applied Microbiology and John Wiley & Sons Ltd.)

Microbial electrosynthesis (MES) systems with mixed cultures often generate a variety of gaseous and soluble chemicals. Methane is the primary end product in mixed-culture MES because it is the thermodynamically most favorable reduction product of CO2. Here, we show that the peptaibol alamethicin selectively suppressed the growth of methanogens in mixed-culture MES systems, resulting in a shift of the solution and cathode communities to an acetate-producing system dominated by Sporomusa, a known acetogenic genus in MES systems. Archaea in the methane-producing control were dominated by Methanobrevibacter species, but no Archaea were detected in the alamethicin-treated reactors. No methane was detected in the mixed-culture reactors treated with alamethicin over 10 cycles (∼ 3 days each). Instead, acetate was produced at an average rate of 115 nmol ml(-1) day(-1), similar to the rate reported previously for pure cultures of Sporomusa ovata on biocathodes. Mixed-culture control reactors without alamethicin generated methane at nearly 100% coulombic recovery, and no acetate was detected. These results show that alamethicin is effective for the suppression of methanogen growth in MES systems and that its use enables the production of industrially relevant organic compounds by the inhibition of methanogenesis., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Numerous bacteria assemble proteinaceous microcompartments to isolate certain biochemical reactions within the cytoplasm. The assembly, structure, contents, and functions of these microcompartments are active areas of research. Here we show that the Gram-negative sporulating bacterium Acetonema longum synthesizes propanediol utilization (PDU) microcompartments when starved or grown on 1,2-propanediol (1,2-PD) or rhamnose. Electron cryotomography of intact cells revealed that PDU microcompartments are highly irregular in shape and size, similar to purified PDU microcompartments from Salmonella enterica serovar Typhimurium LT2 that were imaged previously. Homology searches identified a 20-gene operon in A. longum that contains most of the structural, enzymatic, and regulatory genes thought to be involved in PDU microcompartment assembly and function. Transcriptional data on PduU and PduC, which are major structural and enzymatic proteins, respectively, as well as imaging, indicate that PDU microcompartment synthesis is induced within 24 h of growth on 1,2-PD and after 48 h of growth on rhamnose.

Corrinoids are essential cofactors of reductive dehalogenases in Dehalococcoides mccartyi, an important bacterium in bioremediation, yet sequenced D. mccartyi strains do not possess the complete pathway for de novo corrinoid biosynthesis. Pelosinus sp. and Desulfovibrio sp. have been detected in dechlorinating communities enriched from contaminated groundwater without exogenous cobalamin corrinoid. To investigate the corrinoid-related interactions among key members of these communities, we constructed consortia by growing D. mccartyi strain 195 (Dhc195) in cobalamin-free, trichloroethene (TCE)- and lactate-amended medium in cocultures with Desulfovibrio vulgaris Hildenborough (DvH) or Pelosinus fermentans R7 (PfR7) and with both in tricultures. Only the triculture exhibited sustainable dechlorination and cell growth when a physiological level of 5,6-dimethylbenzimidazole (DMB), the lower ligand of cobalamin, was provided. In the triculture, DvH provided hydrogen while PfR7 provided corrinoids to Dhc195, and the initiation of dechlorination and Dhc195 cell growth was highly dependent on the growth of PfR7. Corrinoid analysis indicated that Dhc195 imported and remodeled the phenolic corrinoids produced by PfR7 into cobalamin in the presence of DMB. Transcriptomic analyses of Dhc195 showed the induction of the CbiZ-dependent corrinoid-remodeling pathway and BtuFCD corrinoid ABC transporter genes during corrinoid salvaging and remodeling. In contrast, another operon annotated to encode a putative iron/cobalamin ABC transporter (DET1174-DET1176) was induced when cobalamin was exogenously provided. Interestingly, a global upregulation of phage-related genes was observed when PfR7 was present. These findings provide insights into both the gene regulation of corrinoid salvaging and remodeling in Dhc195 when it is grown without exogenous cobalamin and microbe-to-microbe interactions in dechlorinating microbial communities.

Propionic acid is presently mainly produced by chemical synthesis. For many applications, especially in feed and food industries, a fermentative production of propionic acid from cheap and renewable resources is of large interest. In this work, we investigated the use of a co-culture to convert household flour to propionic acid. Batch and fed-batch fermentations of hydrolyzed flour and a process of simultaneous saccharification and fermentation were examined and compared. Fed-batch culture with substrate limitation was found to be the most efficient process, reaching a propionic acid concentration of 30 g/L and a productivity of 0.33 g/L*h. This is the highest productivity so far achieved with free cells on media containing flour hydrolysate or glucose as carbon source. Batch culture and culture with controlled saccharification and fermentation delivered significantly lower propionic acid production (17-20 g/L) due to inhibition by the intermediate product lactate. It is concluded that co-culture fermentation of flour hydrolysate can be considered as an appealing bioprocess for the production of propionic acid.

Phenolyl cobamides are unique members of a class of cobalt-containing cofactors that includes vitamin B12 (cobalamin). Cobamide cofactors facilitate diverse reactions in prokaryotes and eukaryotes. Phenolyl cobamides are structurally and chemically distinct from the more commonly used benzimidazolyl cobamides such as cobalamin, as the lower axial ligand is a phenolic group rather than a benzimidazole. The functional significance of this difference is not well understood. Here we show that in the bacterium Sporomusa ovata, the only organism known to synthesize phenolyl cobamides, several cobamide-dependent acetogenic metabolisms have a requirement or preference for phenolyl cobamides. The addition of benzimidazoles to S. ovata cultures results in a decrease in growth rate when grown on methanol, 3,4-dimethoxybenzoate, H2 plus CO2, or betaine. Suppression of native p-cresolyl cobamide synthesis and production of benzimidazolyl cobamides occur upon the addition of benzimidazoles, indicating that benzimidazolyl cobamides are not functionally equivalent to the phenolyl cobamide cofactors produced by S. ovata. We further show that S. ovata is capable of incorporating other phenolic compounds into cobamides that function in methanol metabolism. These results demonstrate that S. ovata can incorporate a wide range of compounds as cobamide lower ligands, despite its preference for phenolyl cobamides in the metabolism of certain energy substrates. To our knowledge, S. ovata is unique among cobamide-dependent organisms in its preferential utilization of phenolyl cobamides.

Microbial electrosynthesis, the conversion of carbon dioxide to organic molecules using electricity, has recently been demonstrated for acetogenic microorganisms, such as Sporomusa ovata. The energy for reduction of carbon dioxide originates from the hydrolysis of water on the anode, requiring a sufficiently low potential. Here we evaluate the use of sulfide as an electron source for microbial electrosynthesis. Abiotically oxidation of sulfide on the anode yields two electrons. The oxidation product, elemental sulfur, can be further oxidized to sulfate by Desulfobulbus propionicus, generating six additional electrons in the process. The eight electrons generated from the combined abiotic and biotic steps were used to reduce carbon dioxide to acetate on a graphite cathode by Sporomusa ovata at a rate of 24.8 mmol/day · m(2). Using a strain of Desulfuromonas as biocatalyst on the anode resulted in an acetate production rate of 49.9 mmol/day · m(2), with a Coulombic efficiency of over 90%. These results demonstrate that sulfide can serve effectively as an alternative electron donor for microbial electrosynthesis.

Pelosinus spp. are fermentative firmicutes that were recently reported to be prominent members of microbial communities at contaminated subsurface sites in multiple locations. Here we report metabolic characteristics and their putative genetic basis in Pelosinus sp. strain HCF1, an isolate that predominated anaerobic, Cr(VI)-reducing columns constructed with aquifer sediment. Strain HCF1 ferments lactate to propionate and acetate (the methylmalonyl-coenzyme A [CoA] pathway was identified in the genome), and its genome encodes two [NiFe]- and four [FeFe]-hydrogenases for H(2) cycling. The reduction of Cr(VI) and Fe(III) may be catalyzed by a flavoprotein with 42 to 51% sequence identity to both ChrR and FerB. This bacterium has unexpected capabilities and gene content associated with reduction of nitrogen oxides, including dissimilatory reduction of nitrate to ammonium (two copies of NrfH and NrfA were identified along with NarGHI) and a nitric oxide reductase (NorCB). In this strain, either H(2) or lactate can act as a sole electron donor for nitrate, Cr(VI), and Fe(III) reduction. Transcriptional studies demonstrated differential expression of hydrogenases and nitrate and nitrite reductases. Overall, the unexpected metabolic capabilities and gene content reported here broaden our perspective on what biogeochemical and ecological roles this species might play as a prominent member of microbial communities in subsurface environments.

Pelosinus fermentans JBW45 is an anaerobic, lactate-fermenting bacterium isolated from Cr(VI)-contaminated groundwater at the Hanford Nuclear Reservation 100-H site (Washington) that was collected after stimulation with a polylactate compound. The genome sequence of this organism will provide insight into the metabolic potential of a predominant population during stimulation for metal-reducing conditions.

Pelosinus fermentans 16S rRNA gene sequences have been reported from diverse geographical sites since the recent isolation of the type strain. We present the genome sequence of the P. fermentans type strain R7 (DSM 17108) and genome sequences for two new strains with different abilities to reduce iron, chromate, and uranium.

Two anaerobic bacterial strains, designated SHI-1(T) and SHI-2, were isolated from chlorinated solvent-contaminated groundwater. They were found to be identical in phenotypic properties and shared high (98.5-99.8 %) pairwise 16S rRNA gene sequence similarity. Multiple 16S rRNA genes were found to be present in the isolates as well as Pelosinus fermentans DSM 17108(T) and Sporotalea propionica DSM 13327(T). Strains SHI-1(T) and SHI-2 could be differentiated from their closest phylogenetic relatives, P. fermentans DSM 17108(T) and S. propionica DSM 13327(T), on the basis of their phenotypic and phylogenetic properties. The isolates were Gram-negative, spore-forming, motile rods with peritrichous flagella. Growth occurred at 10-42 °C and pH 5.5-8.5. Fermentative growth was observed on Casamino acids, fructose, fumarate, glucose, glycerol, pyruvate and yeast extract. The major organic acids produced from glucose and glycerol fermentation were propionate and acetate. The major organic acids produced from fermentation of fumarate were propionate, acetate and succinate. The major cellular fatty acids were summed feature 4 (consisting of C(15:1)ω8c and/or C(15:2)), summed feature 8 (consisting of C(17:1)ω8c and/or C(17:2)) and C(14:0) dimethyl aldehyde. The polar lipids comprised aminophospholipids, including phosphatidylethanolamine and phosphatidylserine, and an unknown phospholipid. The genomic DNA G+C content was 39.2 mol%. We propose that strains SHI-1(T) and SHI-2 are assigned to a novel species of the genus Pelosinus, with the name Pelosinus defluvii sp. nov. (type strain SHI-1(T) = NRRL Y-59407(T) = LMG 25549(T)). The description of the genus Pelosinus is emended. We also propose the transfer of S. propionica to the genus Pelosinus as Pelosinus propionicus comb. nov. (type strain TmPN3(T) = DSM 13327(T) = ATCC BAA-626(T)), on the basis of phylogenetic, chemotaxonomic and phenotypic properties.

Isolation, cultivation, and characterization of the intestinal microorganisms are important for understanding the comprehensive physiology of the human gastrointestinal (GI) tract microbiota. Here, we isolated two novel bacterial strains, YIT 12067(T) and YIT 12068, from the feces of healthy human adults. Phylogenetic analysis indicated that they belonged to the same species and were most closely related to Phascolarctobacterium faecium ACM 3679(T), with 91.4% to 91.5% 16S rRNA gene sequence similarities, respectively. Substrate availability tests revealed that the isolates used only succinate; they did not ferment any other short-chain fatty acids or carbohydrates tested. When these strains were cocultured with the xylan-utilizing and succinate-producing bacterium Paraprevotella xylaniphila YIT 11841(T), in medium supplemented with xylan but not succinate, their cell numbers became 2 to 3 orders of magnitude higher than those of the monoculture; succinate became undetectable, and propionate was formed. Database analysis revealed that over 200 uncultured bacterial clones from the feces of humans and other mammals showed high sequence identity (>98.7%) to YIT 12067(T). Real-time PCR analysis also revealed that YIT 12067(T)-like bacteria were present in 21% of human fecal samples, at an average level of 3.34 × 10(8) cells/g feces. These results indicate that YIT 12067(T)-like bacteria are distributed broadly in the GI tract as subdominant members that may adapt to the intestinal environment by specializing to utilize the succinate generated by other bacterial species. The phylogenetic and physiological properties of YIT 12067(T) and YIT 12068 suggest that these strains represent a novel species, which we have designated Phascolarctobacterium succinatutens sp. nov.

Two hallmarks of the Firmicute phylum, which includes the Bacilli and Clostridia classes, are their ability to form endospores and their "Gram-positive" single-membraned, thick-cell-wall envelope structure. Acetonema longum is part of a lesser-known family (the Veillonellaceae) of Clostridia that form endospores but that are surprisingly "Gram negative," possessing both an inner and outer membrane and a thin cell wall. Here, we present macromolecular resolution, 3D electron cryotomographic images of vegetative, sporulating, and germinating A. longum cells showing that during the sporulation process, the inner membrane of the mother cell is inverted and transformed to become the outer membrane of the germinating cell. Peptidoglycan persists throughout, leading to a revised, "continuous" model of its role in the process. Coupled with genomic analyses, these results point to sporulation as a mechanism by which the bacterial outer membrane may have arisen and A. longum as a potential "missing link" between single- and double-membraned bacteria., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Microbial electrosynthesis, a process in which microorganisms use electrons derived from electrodes to reduce carbon dioxide to multicarbon, extracellular organic compounds, is a potential strategy for capturing electrical energy in carbon-carbon bonds of readily stored and easily distributed products, such as transportation fuels. To date, only one organism, the acetogen Sporomusa ovata, has been shown to be capable of electrosynthesis. The purpose of this study was to determine if a wider range of microorganisms is capable of this process. Several other acetogenic bacteria, including two other Sporomusa species, Clostridium ljungdahlii, Clostridium aceticum, and Moorella thermoacetica, consumed current with the production of organic acids. In general acetate was the primary product, but 2-oxobutyrate and formate also were formed, with 2-oxobutyrate being the predominant identified product of electrosynthesis by C. aceticum. S. sphaeroides, C. ljungdahlii, and M. thermoacetica had high (>80%) efficiencies of electrons consumed and recovered in identified products. The acetogen Acetobacterium woodii was unable to consume current. These results expand the known range of microorganisms capable of electrosynthesis, providing multiple options for the further optimization of this process.

The possibility of providing the acetogenic microorganism Sporomusa ovata with electrons delivered directly to the cells with a graphite electrode for the reduction of carbon dioxide to organic compounds was investigated. Biofilms of S. ovata growing on graphite cathode surfaces consumed electrons with the reduction of carbon dioxide to acetate and small amounts of 2-oxobutyrate. Electrons appearing in these products accounted for over 85% of the electrons consumed. These results demonstrate that microbial production of multicarbon organic compounds from carbon dioxide and water with electricity as the energy source is feasible.

A strictly anaerobic, mesophilic and aminolytic strain (WN036(T)) was isolated from a methanogenic reactor treating waste from cattle farms. Cells were Gram-positive cocci, often occurred in pairs and were non-motile. Although spore formation was not confirmed by microscopic observation of cells, the strain produced thermotolerant cells. The optimum temperature for growth was 35-37 degrees C and the optimum pH was 6.7. Oxidase, catalase and nitrate-reducing activities were negative. The novel strain did not ferment carbohydrates and grew in PY medium without additional substrates. The strain utilized l-glutamate, l-glutamine, l-histidine and l-arginine as growth substrates. Major fermentation products were acetate and butyrate with a small amount of propionate. The genomic DNA G+C content was 32.5 mol%. The major cellular fatty acids were C(17 : 1)omega8, C(18 : 1)omega7 DMA and C(16 : 0). The diagnostic diamino acid of the cell-wall peptidoglycan was lysine. Glutamic acid, glycine, alanine and aspartic acid were also detected in the cell-wall peptidoglycan. On the basis of 16S rRNA gene sequences, the most closely related species to strain WN036(T) were Peptoniphilus asaccharolyticus ATCC 14965(T) (89.8 %) and Peptoniphilus indolicus ATCC 29427(T) (89.6 %). Based on the differences in the phenotypic and phylogenetic characteristics of strain WN036(T) compared with those of closely related species, a novel genus and species, Anaerosphaera aminiphila gen. nov., sp. nov., is proposed. The type strain is WN036(T) (=JCM 15094(T)=DSM 21120(T)).

A strictly anaerobic, sluggishly motile, spore-forming, thermophilic bacterium, designated strain AeG(T), was isolated from microbial mats colonizing a runoff channel formed by free-flowing thermal waters of a bore well (New Lorne Bore; registered number 17263) in the Great Artesian Basin, Australia. Cells of strain AeG(T) were curved rods (2.0-10.0x0.8-1.0 mum) and stained Gram-negative. The strain grew optimally in tryptone-yeast extract-citrate medium at 55 degrees C (range for growth between 45 and 60 degrees C) and pH 7.0 (range for growth between pH 6.5 and 8.0). Citrate and malate, but no other organic acids, carbohydrates or amino acids could be used in the presence of up to 0.1 % yeast extract. Although yeast extract and/or tryptone were required for growth on citrate, they did not support growth as sole carbon sources. Strain AeG(T) reduced thiosulfate and sulfite in the presence of 0.2 % yeast extract, but not Fe(III), Mn(IV), sulfate, elemental sulfur, nitrate or nitrite. Growth was inhibited by chloramphenicol, streptomycin, tetracycline, penicillin and ampicillin and in the presence of NaCl concentrations >1 %. The DNA G+C content was 55.4+/-1.0 mol% as determined by the thermal denaturation method. 16S rRNA gene sequence analysis indicated that strain AeG(T) was a member of the family Veillonellaceae, class 'Clostridia', phylum 'Firmicutes' and was most closely related to members of the genus Propionispora (mean 16S rRNA gene sequence similarity value to type strains was 90.8 %). Based on these results, strain AeG(T) is considered to represent a novel species in a new genus, for which the name Sporolituus thermophilus gen. nov., sp. nov. is proposed. The type strain of the type species is AeG(T) (=JCM 15556(T)=KCTC 5668(T)).

A novel antibiotic naphthalecin was purified and isolated from the cells of an anaerobic bacterium isolated from a soil sample. This antibiotic contained a naphthalene moiety, so named as naphthalecin, and showed antibacterial activity against gram positive species. The producing strain, an obligate anaerobe, was identified as a new species of the genus Sporotalea. Identification of the bacterium, cultivation, purification, structure determination, and antibacterial activity are shown.

Three strains of anaerobic, non-spore-forming, Gram-negative coccobacilli (YIT 11816T, YIT 11817 and YIT 11818) were isolated from human faeces. On the basis of 16S rRNA gene sequence similarity, these strains were shown to belong to the family Alcaligenaceae and to be related to the type strain of Sutterella stercoricanis (94.9 %) and to Sutterella wadsworthensis WAL 7877 (94.3 %); the similarity to strains of any other species with a validly published name within the family Alcaligenaceae was less than 92 %. Biochemical data supported the affiliation of these strains to the genus Sutterella. These strains therefore represent a novel species, for which the name Sutterella parvirubra sp. nov. is proposed; the type strain is YIT 11816T (=DSM 19354T =JCM 14724T). The cells of another isolate, strain YIT 11815T, were non-spore-forming, Gram-negative, very large rods, 1x5-200 microm in size, with or without a central, subterminal or terminal swelling of 2-4 microm diameter when grown in a broth medium supplemented with glucose. Based on comparative 16S rRNA gene sequencing, this bacterium is a member of the family Acidaminococcaceae, and most closely related to Megamonas hypermegale (95.3 % similarity to the type strain). Interestingly, the 16S rRNA gene sequence of strain YIT 11815T showed 99 % similarity to sequences of uncultured colonic bacteria. A 16S rRNA gene sequence divergence value of >3 % from known cultured species suggested that isolate YIT 11815T represents a novel species, for which the name Megamonas funiformis sp. nov. is proposed; the type strain is YIT 11815T (=DSM 19343T =JCM 14723T).

An unusual propionigenic bacterium was isolated from the intestinal tract of the soil-feeding termite Thoracotermes macrothorax. Strain TmPN3 is a motile, long rod that stains gram-positive, but reacts gram-negative in the KOH test. It forms terminal endospores and ferments lactate, glucose, lactose, fructose, and pyruvate to propionate and acetate via the methyl-malonyl-CoA pathway. Propionate and acetate are formed at a ratio of 2:1, typical of most propionigenic bacteria. Under a H(2)/CO(2) atmosphere, the fermentation product pattern of glucose, fructose, and pyruvate shifts towards propionate formation at the expense of acetate. Cell suspensions reduce oxygen with lactate, glucose, glycerol, or hydrogen as electron donor. In the presence of oxygen, the product pattern of lactate fermentation shifts from propionate to acetate production. 16S rRNA gene sequence analysis showed that strain TmPN3 is a firmicute that clusters among the Acidaminococcaceae, a subgroup of the Clostridiales comprising obligately anaerobic, often endospore-forming bacteria that possess an outer membrane. Based on phenotypic differences and less than 92% sequence similarity to the 16S rRNA gene sequence of its closest relative, the termite hindgut isolate Acetonema longum, strain TmPN3(T) is proposed as the type species of a new genus, Sporotalea propionica gen. nov. sp. nov. (DSM 13327(T), ATCC BAA-626(T)).

Acetogens were enumerated from root homogenates of the black needlerush Juncus roemerianus obtained from a nearly pristine salt marsh. An isolated colony, ST1, yielded acetogenic activity and was initially thought to be a pure culture; however, ST1 was subsequently found to be composed of an aerotolerant fermentative anaerobe (RC) and an acetogen (RS(T)) ((T) indicates type strain). The two spore-forming mesophiles were separated by selective cultivation under conditions favouring the growth of either RC or RS(T). The 16S rRNA gene sequence of RC was 99 % similar to that of Clostridium intestinale, indicating that RC was a new isolate of this clostridial species. The rRNA gene sequence most similar to that of RS(T) was only 96 % similar to that of RS(T) and was from a species of the acetogenic genus Sporomusa, indicating that RS(T) was a new sporomusal species; the name Sporomusa rhizae sp. nov. is proposed. RC grew at the expense of saccharides. H(2)-forming butyrate fermentation was the primary catabolism utilized by RC under anoxic conditions, while homolactate fermentation was the primary catabolism under oxic conditions. RC consumed O(2) and tolerated 20 % O(2) in the headspace of shaken broth cultures. In contrast, RS(T) was acetogenic, utilized H(2), lactate and formate, did not utilize saccharides, and could not tolerate high concentrations of O(2). RS(T) grew by trophic interaction with RC on saccharides via the uptake of H(2), and, to a lesser extent, lactate and formate produced by RC. Co-cultures of the two organisms yielded high amounts of acetate. These results indicate that (i) previously uncharacterized species of Sporomusa are associated with Juncus roots and (ii) trophic links to O(2)-consuming aerotolerant anaerobes might contribute to the in situ activities and survival strategies of acetogens in salt marsh rhizospheres, a habitat subject to gradients of plant-derived O(2).

Histamine is an inflammatory agent that contributes to bovine laminitis. Cattle fed silage-containing rations often have large populations of Allisonella histaminiformans, but this obligate histidine-decarboxylating bacterium could not be isolated from cattle fed timothy hay. The growth of A. histaminiformans was stimulated by yeast extract, protein hydrolysates, and water-soluble extracts of alfalfa or corn silage. Extracts of alfalfa were more potent than corn silage. Because growth and histamine production were not stimulated by Casamino Acids or a mixture of purified amino acids, it appeared that A. histaminiformans requires peptides. The idea that A. histaminiformans requires peptides is consistent with the observation that alfalfa silages often have a large amount of peptide nitrogen.

A Gram-negative, spore-forming anaerobe, KS(T), was isolated from an enrichment culture that was set up for anaerobic degradation of the aliphatic polyester poly(propylene adipate). The strain had the cellular organization of Sporomusa, vibrio-shaped cells and terminal round spores, and fermented sugars and sugar alcohols to propionic and acetic acid. Based on the morphological and physiological features as well as on a 16S rRNA gene similarity of 98 %, it was grouped with Propionispora vibrioides. A relatively low DNA-DNA hybridization value with the type strain of this species (47 %), and differences in substrate utilization and spore morphology, suggested that the strain should be classified in a separate species, Propionispora hippei sp. nov., with KS(T) as the type strain (=DSM 15287(T)=ATCC BAA-665(T)).

Previously undescribed, homoacetogenic bacteria were isolated from gut homogenates of the soil-feeding termite Thoracotermes macrothorax. The isolates were slightly curved, banana-shaped rods (0.6-0.7x1.3-7.0 micro m) and were motile by one or more lateral flagella. In older cultures, cells formed club-like sporangia that developed into terminal, heat-resistant endospores. Cells stained Gram-positive but were Gram-negative in the KOH test. The isolates were mesophilic and grew homoacetogenically on H(2)/CO(2) and L-lactate. Strain TmAO3(T), which was characterized further, also grew homoacetogenically on pyruvate, citrate, L-alanine, D-mannitol, ethanol, formate and methanol. Succinate was decarboxylated to propionate; fumarate, L-malate and oxaloacetate were fermented to propionate and acetate. Hexoses were not used as substrates. Resting cells had a large capacity for hydrogen-dependent oxygen reduction [826 nmol min(-1) (mg protein)(-1)], which enabled them to initiate growth in non-reduced basal medium that originally contained up to 1.5 kPa oxygen in the headspace, although growth commenced only after the medium had been rendered anoxic. Redox difference spectra of cell extracts indicated the presence of membrane-bound b-type cytochrome(s). Comparative 16S rRNA gene sequence analysis revealed that strain TmAO3(T) belongs to a subgroup of the phylum of Gram-positive bacteria that is characterized by low DNA G+C content and a Gram-negative cell wall. It is related most closely to representatives of the genus SPOROMUSA: Based on morphological and physiological properties and on 16S rRNA gene sequence similarity of 94-97 % to other Sporomusa species, the isolates are assigned to Sporomusa aerivorans sp. nov. (type strain, TmAO3(T)=DSM 13326(T)=ATCC BAA-625(T)).

The objectives of this study were to examine the effects of growth substrate and extracellular pH on phosphoenolpyruvate-dependent glucose phosphorylation as well as to examine how maltose is phosphorylated by the ruminal bacterium Megasphaera elsdenii B159. Phosphoenolpyruvate-dependent glucose phosphorylation by toluene-treated cells was constitutive, and glucose phosphorylation was reduced by 69% at pH 5.0. When toluene-treated cells were incubated in histidine buffer, little maltose phosphorylation occurred in the absence of inorganic phosphate. However, the addition of increasing concentrations of either potassium or sodium phosphate increased maltose phosphorylation. Maximal phosphorylation activity was observed at between 25 and 50 mM of either inorganic phosphate source. Compared with the control incubations, maltose phosphorylation was increased over threefold with 25 mM of either potassium or sodium phosphate. Phosphoglucomutase activity was detected in cell extracts of M. elsdenii B159, and this enzyme had a K(m) of 3.2 mM for glucose-1-P and a V(max) of 1836 nmol of NADP(+) reduced/mg of protein per min. Maltose was also hydrolyzed by an inducible maltase (K(m), 1.19 mM). To our knowledge, this is the first report of a maltose phosphorylase and a maltase in M. elsdenii.

The effect of Levucell SC, a strain of Saccharomyces cerevisiae marked as a feed additive for ruminants, was investigated in vitro on lactate metabolism by the ruminal bacteria Streptococcus bovis and Megasphaera elsdenii. The coculture between 10(7) live cells x mL(-1) of SC and a Streptococcus bovis strain in the presence of glucose reduced lactate production by the bacterial strain. Live yeast cells were able to compete with Streptococcus bovis for glucose utilization in strictly anaerobic conditions, so less glucose was available for the bacterium. SC also stimulated L-lactate utilization by a strain of M. elsdenii. The effect depended on the concentration of yeast cells added. Bacterial growth and fermentation end-product concentrations were also increased in the presence of SC. Some amino acids and vitamins, but not dicarboxylic acids, stimulated the bacterial specific activity of L-lactate uptake. SC was able to provide amino acids to M. elsdenii. In a coculture of Streptococcus bovis and M. elsdenii on glucose, the reduction of lactate concentration was improved by SC, the same trend being observed when maltose or soluble starch were used as carbon and energy source. These results indicate that SC can be a very useful tool to reduce lactate accumulation in vitro during fermentation of soluble sugars.

Mixed ruminal bacteria convert trans-aconitate to tricarballylate, a tricarboxylic acid which chelates blood divalent cations and decreases their availability (J. B. Russell and P. J. Van Soest, Appl. Environ. Microbiol. 47:155-159, 1984). Decreases in blood magnesium in turn cause a potentially fatal disease known as grass tetany. trans-Aconitate was stoichiometrically reduced to tricarballylate by Selenomonas ruminantium, a common ruminal bacterium in grass-fed ruminants (J. B. Russell, Appl. Environ. Microbiol. 49:120-126, 1985). When mixed ruminal bacteria were enriched with trans-aconitate, a trans-aconitate-oxidizing bacterium was also isolated (G. M. Cook, F. A. Rainey, G. Chen, E. Stackebrandt, and J. B. Russell, Int. J. Syst. Bacteriol. 44:576-578, 1994). The trans-aconitate-oxidizing bacterium was identified as Acidaminococcus fermentans, and it converted trans-aconitate to acetate, a nontoxic end product of ruminal fermentation. When S. ruminantium and A. fermentans were cocultured with trans-aconitate and glucose, tricarballylate never accumulated and all the trans-aconitate was converted to acetate. Continuous-culture studies (dilution rate, 0.1 h-1) likewise indicated that A. fermentans could outcompete S. ruminantium for trans-aconitate. When mixed ruminal bacteria were incubated in vitro with 10 mM trans-aconitate for 24 h, 45% of the trans-aconitate was converted to tricarballylate. Tricarballylate production decreased 50% if even small amounts of A. fermentans were added to the incubation mixes (0.01 mg of protein per mg of mixed bacterial protein). When A. fermentans (2 g of bacterial protein) was added directly to the rumen, the subsequent conversion of trans-aconitate to tricarballylate decreased 50%, but this effect did not persist for more than 18 h.(ABSTRACT TRUNCATED AT 250 WORDS)

Physiological concentrations [< 11 mM] of formate do not violate the metabolism of S. bovis and M. elsdenii. A significant inhibition is caused by concentrations of 22 and 44 mM. In this case the process of ammonia formation in S. bovis is inhibited more pronouncedly. Peculiar effects of formate (11 mM) on LDH, FDH, aconitase, isocitrate dehydrogenase, fumarase, L-MDH and malic-enzyme have been stated. The changes show that it enhances assimilation of sugars fermented to lactate in S. bovis, in contrast to M. elsdenii, where it activates the utilization of lactic acid. During the log-phase S. bovis utilized only 11.5% of [14C] H2O2, whereas M. elsdenii uses 33.4% of it. The major amount of the label is transferred from intracellular inclusions to nucleic acids (in S. bovis--74.7%, in M. elsdenii--87%) and then incorporated into low molecular substances (23.5 and 11.9%, respectively), the rest being incorporated into proteins and lipids.

Production of the amino acid 2-aminobutyrate was studied in four strains of Megasphaera elsdenii grown on a lactate-based growth medium containing Bacto-casamino acids and yeast extract. Supplementation with threonine increased the production of 2-aminobutyrate in three of the four strains, but no substantial increase in production was noted with serine, methionine, or aspartate, all of which are potential sources for the precursor of 2-aminobutyrate, 2-oxobutyrate. L-Cycloserine, an inhibitor of alanine transaminases, decreased both alanine and 2-aminobutyrate production, suggesting that 2-aminobutyrate synthesis may share the same metabolic pathway as alanine synthesis or that 2-oxobutyrate can act as a substrate for alanine transaminases. Decreases in the production of 2-aminobutyrate were associated with a reduction in the catabolism of branched-chain amino acids in two of the four strains.

The objective of this study was to determine the effects of an Aspergillus oryzae fermentation extract (Amaferm) as well as other factors on lactate utilization by the ruminal bacterium Megasphaera elsdenii B159. Addition of Amaferm or a filter-sterilized Amaferm filtrate stimulated L-lactate uptake by both M. elsdenii and the ruminal selenomonad strain H18. Growth of M. elsdenii in medium that contained DL-lactate (2 g/L), Trypticase, and yeast extract was only slightly stimulated by the addition of 5% (vol/vol) Amaferm filtrate after 24 h. However, growth of M. elsdenii in a similar medium lacking Trypticase and yeast extract was increased over twofold by the addition of either 2 or 5% (vol/vol) Amaferm filtrate. These results suggest that Amaferm provides growth factors (i.e., amino acids, B vitamins) to support growth of M. elsdenii on lactate. There was no inhibition of L-lactate uptake when lactate-grown cells of M. elsdenii were incubated with excess (10 mM) glucose, sucrose, or maltose. In addition, when cells were grown on glucose, sucrose, or maltose rather than lactate there was little difference in L-lactate uptake, suggesting that L-lactate transport in M. elsdenii is not subject to catabolite repression by these soluble sugars. Both K+ and Na+ had little effect on L-lactate uptake. Uptake was unaffected at extracellular pH values between 6.0 and 8.0, whereas pH values of 5.0 and 4.0 increased uptake. In addition, L-lactate uptake was inhibited between 34 and 61% by protonophores. These results suggest that protons may be involved in the uptake of L-lactate by M. elsdenii B159.

(R)-2-Hydroxyglutaryl-CoA dehydratase (HGDA/B) from Acidaminococcus fermentans requires an activator protein for activity. This activator (HGDC) has not yet been purified from its natural source due to its low concentration combined with an extreme sensitivity towards oxygen. Gene expression in Escherichia coli identified an open reading frame (780 bp) as the gene encoding HGDC. Dehydratase activity was stimulated at least tenfold by cell-free extracts of E. coli cells transformed with a plasmid carrying hgdC. On the chromosome the hgdC gene is located just before hgdA and hgdB.

A rumen strain of Megasphaera elsdenii was grown on glucose and lactate in monensin-free and monensin-supplemented medium (10 mg/L). Monensin had no effect on growth rate, growth yields, metabolic pattern and composition of cells. Growth yields of dry matter and protein were higher in cultures supplied with glucose than in cultures supplied with lactate. The bacterium compensated the lower gain of energy from fermentation of lactate by rapid utilization of this substrate. Cells grown on glucose contained more saccharide and less protein than lactate-grown cells.