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Introduction: Desulfovibrio vulgaris Hildenborough is a gram-negative anaerobic bacterium belonging to the sulfate-reducing bacteria that exhibits highly versatile metabolism. By switching from one energy mode to another depending on nutrients availability in the environments„ it plays a central role in shaping ecosystems. Despite intensive efforts to study D. vulgaris energy metabolism at the genomic, biochemical and ecological level, bioenergetics in this microorganism remain far from being fully understood. Alternatively, metabolic modeling is a powerful tool to understand bioenergetics. However, all the current models for D. vulgaris appeared to be not easily adaptable to various environmental conditions. Methods: To lift off these limitations, here we constructed a novel transparent and robust metabolic model to explain D. vulgaris bioenergetics by combining whole-cell proteomic analysis with modeling approaches (Flux Balance Analysis). Results: The iDvu71 model showed over 0.95 correlation with experimental data. Further simulations allowed a detailed description of D. vulgaris metabolism in various conditions of growth. Altogether, the simulations run in this study highlighted the sulfate-to-lactate consumption ratio as a pivotal factor in D. vulgaris energy metabolism. Discussion: In particular, the impact on the hydrogen/formate balance and biomass synthesis is discussed. Overall, this study provides a novel insight into D. vulgaris metabolic flexibility. [ABSTRACT FROM AUTHOR]

Several species of microalgae can convert light energy into molecular hydrogen (H2) by employing enzymes of early phylogenetic origin, [FeFe]-hydrogenases, coupled to the photosynthetic electron transport chain. Bacterial [FeFe]-hydrogenases consist of a conserved domain that harbors the active site cofactor, the H-domain, and an additional domain that binds electron-conducting FeS clusters, the F-domain. In contrast, most algal hydrogenases characterized so far have a structurally reduced, so-termed M1-type architecture, which consists only of the H-domain that interacts directly with photosynthetic ferredoxin PetF as an electron donor. To date, only a few algal species are known to contain bacterial-type [FeFe]-hydrogenases, and no M1-type enzymes have been identified in these species. Here, we show that the chlorophycean alga Uronema belkae possesses both bacterial-type and algal-type [FeFe]-hydrogenases. Both hydrogenase genes are transcribed, and the cells produce H2 under hypoxic conditions. The biochemical analyses show that the two enzymes show features typical for each of the two [FeFe]-hydrogenase types. Most notable in the physiological context is that the bacterial-type hydrogenase does not interact with PetF proteins, suggesting that the two enzymes are integrated differently into the alga's metabolism. [ABSTRACT FROM AUTHOR]

IntroductionDesulfovibrio vulgaris Hildenborough is a gram-negative anaerobic bacterium belonging to the sulfate-reducing bacteria that exhibits highly versatile metabolism. By switching from one energy mode to another depending on nutrients availability in the environments„ it plays a central role in shaping ecosystems. Despite intensive efforts to study D. vulgaris energy metabolism at the genomic, biochemical and ecological level, bioenergetics in this microorganism remain far from being fully understood. Alternatively, metabolic modeling is a powerful tool to understand bioenergetics. However, all the current models for D. vulgaris appeared to be not easily adaptable to various environmental conditions.MethodsTo lift off these limitations, here we constructed a novel transparent and robust metabolic model to explain D. vulgaris bioenergetics by combining whole-cell proteomic analysis with modeling approaches (Flux Balance Analysis).ResultsThe iDvu71 model showed over 0.95 correlation with experimental data. Further simulations allowed a detailed description of D. vulgaris metabolism in various conditions of growth. Altogether, the simulations run in this study highlighted the sulfate-to-lactate consumption ratio as a pivotal factor in D. vulgaris energy metabolism.DiscussionIn particular, the impact on the hydrogen/formate balance and biomass synthesis is discussed. Overall, this study provides a novel insight into D. vulgaris metabolic flexibility.

Several species of microalgae can convert light energy into molecular hydrogen (H2) by employing enzymes of early phylogenetic origin, [FeFe]-hydrogenases, coupled to the photosynthetic electron transport chain. Bacterial [FeFe]-hydrogenases consist of a conserved domain that harbors the active site cofactor, the H-domain, and an additional domain that binds electron-conducting FeS clusters, the F-domain. In contrast, most algal hydrogenases characterized so far have a structurally reduced, so-termed M1-type architecture, which consists only of the H-domain that interacts directly with photosynthetic ferredoxin PetF as an electron donor. To date, only a few algal species are known to contain bacterial-type [FeFe]-hydrogenases, and no M1-type enzymes have been identified in these species. Here, we show that the chlorophycean alga Uronema belkae possesses both bacterial-type and algal-type [FeFe]-hydrogenases. Both hydrogenase genes are transcribed, and the cells produce H2 under hypoxic conditions. The biochemical analyses show that the two enzymes show features typical for each of the two [FeFe]-hydrogenase types. Most notable in the physiological context is that the bacterial-type hydrogenase does not interact with PetF proteins, suggesting that the two enzymes are integrated differently into the alga’s metabolism.

Aerobic hydrogen-oxidizing bacteria (HOB) is a group of active, abundant, and diverse microorganisms with representatives in nearly all phyla, distributed in soil exposed to atmospheric or elevated hydrogen. In this review, first, we discuss the fundamental physiology, isolation, and identification techniques for HOB. On this basis, the hydrogenase genetic organization and metabolic strategy of Cupriavidus necator, Mycobacterium smegmatis as representative HOB are summarized. Availability of hydrogen, oxygen, nutrients, and environmental variables such as temperature and moisture are key ecological factors influencing H2 oxidation activity, hydrogenase groups, and microbial composition. Finally, we systematically illustrate the ecological roles of HOB and the interactions between HOB and other soil microbiota, particularly rhizobia in agricultural soils and Cyanobacteria in deserts. Intensive studies should focus on the cell functioning regulated by hydrogenases and on the full play of ecological roles by competitive HOB consortiums in soil niches, which is expected to facilitate the biogeochemical process of carbon dioxide fixation and energy utilization. [ABSTRACT FROM AUTHOR]

Previous studies have indicated that hydrogen-rich water (HW) treatment can delay fruit ripening and senescence. However, little is known about the HW-delaying pulp breakdown. In this study, eight physiological characteristics revealed that HW treatment delayed both pericarp browning and pulp breakdown of litchi fruit. To gain a comprehensive understanding of the changes in litchi pulp, a combination of multiple metabolomics and gene expression analyses was conducted, assessing 67 primary metabolites, 103 volatiles, 31 amino acids, and 13 crucial metabolite-related genes. Results showed that HW treatment promoted starch degradation, decelerated cell wall degradation and glycolysis, and maintained the flavor and quality of litchi fruit. Furthermore, HW treatment stimulated the production of volatile alcohols, aldehydes, ketones, olefins, and amino acids, which might play a vital role in HW-delaying pulp breakdown. This study sheds light on the mechanism by which HW delayed pulp breakdown by investigating small molecule metabolites and metabolic pathways., 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 © 2024 Elsevier Ltd. All rights reserved.)

Molecular hydrogen is beneficial for fruits quality improvement. However, the mechanism involved, especially cellular metabolic responses, has not been well established. Here, the integrated widely targeted metabolomics analysis (UPLC-MS/MS) and biochemical evidence revealed that hydrogen-based irrigation could orchestrate, either directly or indirectly, an array of physiological responses in blueberry (Vaccinium spp.) during harvesting stage, especially for the delayed senescence in harvested stage (4 °C for 12 d). The hubs to these changes are wide-ranging metabolic reprogramming and antioxidant machinery. A total of 1208 distinct annotated metabolites were identified, and the characterization of differential accumulated metabolites (DAMs) revealed that the reprogramming, particularly, involves phenolic acids and flavonoids accumulation. These changes were positively matched with the transcriptional profiles of representative genes for their synthesis during the growth stage. Together, our findings open a new window for development of hydrogen-based agriculture that increases the shelf-life of fruits in a smart and sustainable manner., 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. Yan Zeng, Xu Cheng, and Pathier Didier are employees and hold ownership interests (including patents) in Air Liquide (China) R&D Co., Ltd., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Antibiotic fermentation residue, which is generated from the microbial antibiotic production process, has been a troublesome waste faced by the pharmaceutical industry. Dark fermentation is a potential technology to treat antibiotic fermentation residue in terms of renewable H 2 generation and waste management. However, the inherent antibiotic in antibiotic fermentation residue may inhibit its dark fermentation performance, and current understanding on this topic is limited. This investigation examined the impact of the inherent antibiotic on the dark H 2 fermentation of Cephalosporin C (CEPC) fermentation residue, and explored the mechanisms from the perspectives of bacterial communities and functional genes. It was found that CEP-C in the antibiotic fermentation residue significantly inhibited the H 2 production, with the H 2 yield decreasing from 17.2 mL/g-VS added to 12.5 and 9.6 mL/g-VS added at CEP-C concentrations of 100 and 200 mg/L, respectively. CEP-C also prolonged the H 2 -producing lag period. Microbiological analysis indicated that CEP-C remarkably decreased the abundances of high-yielding H 2 -producing bacteria, as well as downregulated the genes involved in hydrogen generation from the"pyruvate pathway" and"NADH pathway", essentially leading to the decline of H 2 productivity. The present work gains insights into how cephalosporin antibiotics influence the dark H 2 fermentation, and provide guidance for mitigating the inhibitory effects., 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 © 2024 Elsevier B.V. All rights reserved.)

Hydrogenedentota , a globally distributed bacterial phylum-level lineage, is poorly understood. Here, we established a comprehensive genomic catalog of Hydrogenedentota , including a total of seven clades (or families) with 179 genomes, and explored the metabolic potential and evolutionary history of these organisms. We show that a single genome, especially those belonging to Clade 6, often encodes multiple hydrogenases with genomes in Clade 2, which rarely encode hydrogenases being the exception. Notably, most members of Hydrogenedentota contain a group A3 [FeFe]-hydrogenase (BfuABC) with a non-canonical electron bifurcation mechanism, in addition to substrate-level phosphorylation and electron transport-linked phosphorylation pathways, in energy conservation. Furthermore, we show that BfuABC from Hydrogenedentota fall into five sub-types. Phylogenetic analysis reveals five independent routes for the evolution of BfuABC homologs in Hydrogenedentota . We speculate that the five sub-types of BfuABC might be acquired from Bacillota (synonym Firmicutes ) through separate horizontal gene transfer events. These data shed light on the diversity and evolution of bifurcating [FeFe]-hydrogenases and provide insight into the strategy of Hydrogenedentota to adapt to survival in various habitats., Importance: The phylum Hydrogenedentota is widely distributed in various environments. However, their physiology, ecology, and evolutionary history remain unknown, primarily due to the limited availability of the genomes and the lack of cultured representatives of the phylum. Our results have increased the knowledge of the genetic and metabolic diversity of these organisms and shed light on their diverse energy conservation strategies, especially those involving electron bifurcation with a non-canonical mechanism, which are likely responsible for their wide distribution. Besides, the organization and phylogenetic relationships of gene clusters coding for BfuABC in Hydrogenedentota provide valuable clues to the evolutionary history of group A3 electron bifurcating [FeFe]-hydrogenases., Competing Interests: The authors declare no conflict of interest.

Enhancing the availability of representative isolates from hydrothermal vents (HTVs) is imperative for comprehending the microbial processes that propel the vent ecosystem. In recent years, Campylobacteria have emerged as the predominant and ubiquitous taxon across both shallow and deep-sea vent systems. Nevertheless, only a few isolates have been cultured, primarily originating from deep-sea HTVs. Presently, no cultivable isolates of Campylobacteria are accessible in shallow water vent systems (<200 m), which exhibit markedly distinct environmental conditions from their deep-sea counterparts. In this study, we enriched a novel isolate (genus Sulfurospirillum , Campylobacteria) from shallow-water HTVs of Kueishan Island. Genomic and physiological analysis revealed that this novel Campylobacteria species grows on a variety of substrate and carbon/energy sources. The pan-genome and phenotypic comparisons with 12 previously isolated Sulfurospirillum species from different environments supported the identification of functional features in Sulfurospirillum genomes crucial for adaptation to vent environments, such as sulfur oxidation, carbon fixation, biofilm formation, and benzoate/toluene degradation, as well as diverse genes related with signal transportation. To conclude, the metabolic characteristics of this novel Campylobacteria augment our understanding of Campylobacteria spanning from deep-sea to shallow-water vent systems.IMPORTANCECampylobacteria emerge as the dominant and ubiquitous taxa within vent systems, playing important roles in the vent ecosystems. However, isolated representatives of Campylobacteria have been mainly from the deep-sea hydrothermal fields, leaving a significant knowledge gap regarding the functions, activities, and adaptation strategies of the vent microorganisms in shallow-water hydrothermal vents (HTVs). This study bridges this gap by providing insights into the phenomics and genomic diversity of genus Sulfurospirillum (order Campylobacterales, class Campylobacteria) based on data derived from a novel isolate obtained from shallow-water HTVs. Our mesophilic isolate of Sulfurospirillum not only augments the genus diversity of Campylobacteria pure cultures derived from vent systems but also serves as the inaugural reference isolate for Campylobacteria in shallow-water environments., Competing Interests: The authors declare no conflict of interest.

Oxidative stress in adipose tissue may alter the secretion pattern of adipocytokines and potentially promote atherosclerosis. However, the therapeutic role of hydrogen in adipose tissue under oxidative stress remains unclear. In this study, subcutaneous adipose tissue (SCAT) was collected from the mid-thoracic wounds of 12 patients who underwent open-heart surgery with a mid-thoracic incision. The adipose tissue was then immersed in a culture medium dissolved with hydrogen, which was generated using a hydrogen-generating device. The weight of the adipose tissue was measured before and after hydrogenation, and the tissue was immunostained for nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), and superoxide dismutase (SOD), which are markers of oxidative stress. The immunostaining results showed that HO-1 and Nrf2 expression levels were significantly decreased in the hydrogenated group, whereas SOD expression levels increased, but did not attain statistical significance. Image analysis of adipose tissue revealed that a reduction in adipocyte size. Furthermore, hydrogenated adipose tissue showed a trend toward increased gene expression levels of adiponectin and decreased gene expression levels of chemerin, an adipocytokine involved in adipogenesis. These results demonstrated the therapeutic potential of hydrogen gas for oxidative stress in adipose tissue and for reducing adipocyte size., (© 2024. The Author(s).)

Biofuel can be generated by different organisms using various substrates. The green alga Chlorococcum humicola OQ934050 exhibited the capability to photosynthesize carbonate carbon, maybe via the activity of carbonic anhydrase enzymes. The optimum treatment is C:N ratio of 1:1 (0.2 mmoles sodium carbonate and 0.2 mmoles sodium nitrate) as it induced the highest dry mass (more than 0.5 mg.mL -1 ). At this combination, biomass were about 0.2 mg/mL -1 carbohydrates, 0.085 mg/mL -1 proteins, and 0.16 mg/mL -1 oil of this dry weight. The C/N ratios of 1:1 or 10:1 induced up to 30% of the Chlorococcum humicola dry mass as oils. Growth and dry matter content were hindered at 50:1 C/N and oil content was reduced as a result. The fatty acid profile was strongly altered by the applied C.N ratios. The defatted leftovers of the grown alga, after oil extraction, were fermented by a newly isolated heterotrophic bacterium, identified as Bacillus coagulans OQ053202, to evolve hydrogen content as gas. The highest cumulative hydrogen production and reducing sugar (70 ml H 2 /g biomass and 0.128 mg/ml; respectively) were found at the C/N ratio of 10:1 with the highest hydrogen evolution efficiency (HEE) of 22.8 ml H 2 / mg reducing sugar. The optimum treatment applied to the Chlorococcum humicola is C:N ratio of 1:1 for the highest dry mass, up to 30% dry mass as oils. Some fatty acids were induced while others disappeared, depending on the C/N ratios. The highest cumulative hydrogen production and reducing sugar were found at the C/N ratio of 10:1., (© 2024. The Author(s).)

The use of enzymes to generate hydrogen, instead of using rare metal catalysts, is an exciting area of study in modern biochemistry and biotechnology, as well as biocatalysis driven by sustainable hydrogen. Thus far, the oxygen sensitivity of the fastest hydrogen-producing/exploiting enzymes, [FeFe]hydrogenases, has hindered their practical application, thereby restricting innovations mainly to their [NiFe]-based, albeit slower, counterparts. Recent exploration of the biodiversity of clostridial hydrogen-producing enzymes has yielded the isolation of representatives from a relatively understudied group. These enzymes possess an inherent defense mechanism against oxygen-induced damage. This discovery unveils fresh opportunities for applications such as electrode interfacing, biofuel cells, immobilization, and entrapment for enhanced stability in practical uses. Furthermore, it suggests potential combinations with cascade reactions for CO 2 conversion or cofactor regeneration, like NADPH, facilitating product separation in biotechnological processes. This work provides an overview of this new class of biocatalysts, incorporating unpublished protein engineering strategies to further investigate the dynamic mechanism of oxygen protection and to address crucial details remaining elusive such as still unidentified switching hot-spots and their effects. Variants with improved k cat as well as chimeric versions with promising features to attain gain-of-function variants and applications in various biotechnological processes are also presented.

Background: Clean energy hydrogen (H 2 ) produced from abundant lignocellulose is an alternative to fossil energy. As an essential influencing factor, there is a lack of comparison between constant temperatures (35, 55 and 65 °C) and gradient heating temperature (35 to 65 °C) on the H 2 production regulation potential from lignocellulose-rich straw via high-solid anaerobic digestion (HS-AD). More importantly, the microbial mechanism of temperature regulating H 2 accumulation needs to be investigated., Results: Constant 65 °C led to the lowest lignin residue (1.93%) and the maximum release of cellulose and hemicellulose, and the highest H 2 production (26.01 mL/g VS). H 2 production at 35 and 55 °C was only 14.56 and 24.13 mL/g VS, respectively. In order to further explore the potential of ultra-high temperature (65 °C), HS-AD was performed by gradient heating conditions (35 to 65 °C). However, compared to constant 65 °C, gradient heating conditions led to higher lignin residue (2.49%) and lower H 2 production (13.53 mL/g VS) than gradient heating conditions (47.98%). In addition, metagenomic analysis showed the cellulose/hemicellulose hydrolyzing bacteria and genes (mainly Thermoclostridium, and xynA, xynB, abfA, bglB and xynD), H 2- producing bacteria and related genes (mainly Thermoclostridium, and nifD, nifH and nifK), and microbial movement and metabolic functions were enriched at 65 °C. However, the enrichment of two-component systems under gradient heating conditions resulted in a lack of highly-enriched ultra-high-temperature cellulose/hemicellulose hydrolyzing genera and related genes but rather enriched H 2 consumption genera and genes (mainly Acetivibrio, and hyaB and hyaA) resulting in a weaker H 2 production., Conclusions: The lignin degradation process does not directly determine H 2 accumulation, which was actually regulated by bacteria/genes contributing to H 2 production/consumption. In addition, it is temperature that enhances the hydrolysis process of lignin rather than lignin-degrading enzymes, bacteria and genes by promoting microbial material transfer and metabolism. In terms of temperature, one of the key parameters of HS-AD for H 2 production, we developed an important regulatory strategy, enriched the theoretical basis of temperature regulation for H 2 production to further expanded the research horizon in this field. Video Abstract., (© 2024. The Author(s).)

The hydrogen molecule can effectively regulate plant growth and development, improving plant resistance to abiotic stresses. However, studies regarding the optimal concentration of hydrogen and the associated mechanisms of action in organisms are lacking. This study showed that the maximum germination rate of radish seeds decreased from 90 % to 50 % under the stress of cadmium ions (Cd 2+ ), and hydrogen nanobubble (NB) water significantly alleviated the stress effect of Cd 2+ on radish seed germination. A hydrogen concentration of 0.8 ppm had the best effect, reducing Cd 2+ accumulation in radish seeds by 63.23 % and increasing the maximum germination rate from 50 % to 65 %. At concentrations exceeding 1.2 ppm, the beneficial effect of hydrogen was weakened or even reversed. Consequently, we integrated the concept of the oxidative window into a REDOX balance model and demonstrated that an appropriate hydrogen concentration can effectively maintain the REDOX state within organisms. Transcriptome sequencing analysis revealed that hydrogen NB water modulated Cd 2+ absorption and accumulation in seeds by regulating cell wall components, alleviating oxidative stress through oxidoreductase activity, and enhancing nutrient synthesis and metabolism. This collectively alleviated the inhibitory effect of Cd 2+ on seed germination. This study is helpful for further understanding the effect of hydrogen concentration on the REDOX balance of seed germination, providing a theoretical basis for selecting hydrogen concentration to improve its effectiveness in agricultural fields., Competing Interests: Declaration of Competing Interest TThe 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 © 2024 Elsevier B.V. All rights reserved.)

Although the past decade has witnessed a rapid development of oxidoreductase-mimicking nanozymes, the mimicry of cofactors that play key roles in mediating electron and proton transfer remains limited. This study explores how surface Au-H species conjugated to Au nanoparticles (NPs) that imitate formate dehydrogenase (FDH) can serve as cofactors, analogous to NADH in natural enzymes, offering diverse possibilities for FDH-mimicking Au nanozymes to mimic various enzymes. Once O 2 is present, Au-H species assist Au NPs to complete the on-demand H 2 O 2 generation for cascade reactions. Alternatively, when oxidizing organic molecules are introduced as substrates, Au-H species confer nitro reductase- and aldehyde reductase-like activities on Au NPs under anaerobic conditions. Furthermore, similar to the dehydrogenase-NADH complex, Au NPs possessing Au-H species are gifted with esterase-like activity for ester hydrolysis. By revealing that Au-H species are prosthetic groups for FDH-mimicking Au nanozymes, this work may inspire explorations into future self-generated cofactor mimics for nanozymes, thereby circumventing the need for exogenous cofactors.

Microbial electrolysis cell (MEC) is gaining importance not only for effectively treating wastewater but also for producing hydrogen. The up-flow microbial electrolysis cell (UPMEC) is an innovative approach to enhance the efficiency, and substrate degradation. In this study, a baffled UPMEC with an anode divided into three regions by inserting the baffle (sieve) plates at varying distances from the cathode was designed. The effect of process parameters, such as flow rate (10, 15, and 20 mL/min), electrode area (50, 100, and 150 cm 2 ), and catholyte buffer concentration (50, 100, and 150 mM) were investigated using distillery wastewater as substrate. The experimental results showed a maximum of 0.6837 ± 0.02 mmol/L biohydrogen at 150 mM buffer, with 49 ± 1.0% COD reduction using an electrode of area 150 cm 2 . The maximum current density was 1335.94 mA/m 2 for the flow rate of 15 mL/min and surface area of 150 cm 2 . The results showed that at optimized flow rate and buffer concentration, maximum hydrogen production and effective treatment of wastewater were achieved in the baffled UPMEC. PRACTITIONER POINTS: Biohydrogen production from distillery wastewater was investigated in a baffled UPMEC. Flowrate, concentration and electrode areas significantly influenced the hydrogen production. Maximum hydrogen (0.6837±0.02mmol/L.day) production and COD reduction (49±1.0%) was achieved at 15 mL/min. Highest CHR of 95.37±1.9 % and OHR of 4.6±0.09 % was observed at 150 mM buffer concentration., (© 2024 Water Environment Federation.)

Biomethanation represents a promising approach for biomethane production, with biofilm-based processes like trickle bed reactors (TBRs) being among the most efficient solutions. However, maintaining stable performance can be challenging, and both pure and mixed culture approaches have been applied to address this. In this study, inocula enriched with hydrogenotrophic methanogens were introduced to to TBRs as bioaugmentation strategy to assess their impacts on the process performance and microbial community dynamics. Metagenomic analysis revealed a metagenome-assembled genome belonging to the hydrogenotrophic genus Methanobacterium, which became dominant during enrichment and successfully colonized the TBR biofilm after bioaugmentation. The TBRs achieved a biogas production with > 96 % methane. The bioaugmented reactor consumed additional H 2. This may be due to microbial species utilizing CO 2 and H 2 via various CO 2 reduction pathways. Overall, implementing bioaugmentation in TBRs showed potential for establishing targeted species, although challenges remain in managing H 2 consumption and optimizing microbial interactions., 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 © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The global climate change mainly caused by fossil fuels combustion promotes that zero-carbon hydrogen production through eco-friendly methods has attracted attention in recent years. This investigation explored the biohydrogen production by co-fermentation of corn straw (CS) and excess sludge (ES), as well as comprehensively analyzed the internal mechanism. The results showed that the optimal ratio of CS to ES was 9:1 (TS) with the biohydrogen yield of 101.8 mL/g VS, which was higher than that from the mono-fermentation of CS by 1.0-fold. The pattern of volatile fatty acids (VFAs) indicated that the acetate was the most preponderant by-product in all fermentation systems during the biohydrogen production process, and its yield was improved by adding appropriate dosage of ES. In addition, the content of soluble COD (SCOD) was reduced as increasing ES, while concentration of NH 4 + -N showed an opposite tendency. Microbial community analysis revealed that the microbial composition in different samples showed a significant divergence. Trichococcus was the most dominant bacterial genus in the optimal ratio of 9:1 (CS/ES) fermentation system and its abundance was as high as 41.8%. The functional genes prediction found that the dominant metabolic genes and hydrogen-producing related genes had not been significantly increased in co-fermentation system (CS/ES = 9:1) compared to that in the mono-fermentation of CS, implying that enhancement of biohydrogen production by adding ES mainly relied on balancing nutrients and adjusting microbial community in this study. Further redundancy analysis (RDA) confirmed that biohydrogen yield was closely correlated with the enrichment of Trichococcus., 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 © 2024 Elsevier Inc. All rights reserved.)

To evaluate the nutritional modes of orchids associated with 'rhizoctonia' fungi, analyses of hydrogen (δ 2 H), carbon (δ 13 C), and nitrogen (δ 15 N) stable isotope ratios are usually adopted. However, previous studies have not fully accounted for exchangeable hydrogens, which could affect these evaluations. Here, we performed standard δ 13 C, δ 15 N, and δ 2 H analyses on bulk samples. Additionally, we conducted δ 2 H analysis on α-cellulose and cellulose nitrate samples to investigate whether the heterogeneity of exchangeable hydrogens among plant species influences the assessment of nutritional modes. The δ 2 H of orchids were consistently higher than those of surrounding autotrophic plants, irrespective of the three pretreatments. Although the rhizoctonia-associated orchid exhibited lower δ 13 C, its δ 2 H was higher than those of the autotrophs. Notably, among all response variables, δ 15 N and δ 2 H exhibited high abilities for discriminating the nutritional modes of rhizoctonia-associated orchids. These results indicate that a time-efficient bulk sample analysis is an effective method for evaluating plant nutritional modes, as the heterogeneity of exchangeable hydrogens does not significantly impact the estimation. Using δ 15 N and δ 2 H benefits the assessment of partial mycoheterotrophy among rhizoctonia-associated orchids., (© 2024 The Author(s). New Phytologist © 2024 New Phytologist Foundation.)

Despite being classified as microaerophilic microorganisms, most Campylobacter species can grow anaerobically, using formate or molecular hydrogen (H 2 ) as electron donors, and various nitrogenous and sulfurous compounds as electron acceptors. Herein, we showed that both l-asparagine (l-Asn) and l-aspartic acid (l-Asp) bolster H 2 -driven anaerobic growth in several Campylobacter species, whereas the d-enantiomer form of both asparagine (d-Asn) and aspartic acid (d-Asp) only increased anaerobic growth in Campylobacter concisus strain 13826 and Campylobacter ureolyticus strain NCTC10941. A gene annotated as racD encoding for a putative d/l-Asp racemase was identified in the genome of both strains. Disruption of racD in Cc13826 resulted in the inability of the mutant strain to use either d-enantiomer during anaerobic growth. Hence, our results suggest that the racD gene is required for campylobacters to use either d-Asp or d-Asn. The use of d-Asp by various human opportunistic bacterial pathogens, including C. concisus, C. ureolyticus, and also possibly select strains of Campylobacter gracilis, Campylobacter rectus and Campylobacter showae, is significant, because d-Asp is an important signal molecule for both human nervous and neuroendocrine systems. To our knowledge, this is the first report of pathogens scavenging a d-amino acid essential for human health., Competing Interests: Declaration of competing interest The authors have no conflicts of interest., (Copyright © 2024 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.)

Hydrogen-ethanol co-production can significantly improve the energy conversion efficiency of corn stalk (CS). In this study, with CS as the raw material, the co-production characteristics of one-step and two-step photo-fermentation hydrogen production (PFHP) and ethanol production were investigated. In addition, the gas and liquid characteristics of the experiment were analyzed. The kinetics of hydrogen-ethanol co-production was calculated, and the economics of hydrogen and ethanol were analyzed. Results of the experiments indicated that the two-step hydrogen-ethanol co-production had the best hydrogen production performance when the concentration of CS was 25 g/L. The total hydrogen production was 350.08 mL, and the hydrogen yield was 70.02 mL/g, which was 2.45 times higher than that of the one-step method. The efficiency of hydrogen-ethanol co-production was 17.79 %, which was 2.76 times more efficient than hydrogen compared to fermentation with hydrogen. The result provides technical reference for the high-quality utilization of CS., 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 © 2024 Elsevier Ltd. All rights reserved.)

Furfural-tolerant and hydrogen-producing microbial consortia were enriched from soil, with hydrogen production of 259.84 mL/g-xylose under 1 g/L furfural stress. The consortia could degrade 2.5 g/L furfural within 24 h in the xylose system, more efficient than in the sugar-free system. Despite degradation of furfural to furfuryl alcohol, the release of reactive oxygen species and lactate dehydrogenase was also detected, suggesting that furfuryl alcohol is also a potential inhibitor of hydrogen production. The butyrate/acetate ratio was observed to decrease with increasing furfural concentration, leading to decreased hydrogen production. Furthermore, microbial community analysis suggested that dominated Clostridium butyricum was responsible for furfural degradation, while Clostridium beijerinckii reduction led to hydrogen production decrease. Overall, the enriched consortia in this study could efficiently degrade furfural and produce hydrogen, providing new insights into hydrogen-producing microbial consortia with furfural tolerance., 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 © 2024 Elsevier Ltd. All rights reserved.)

ABSTRACT P-bromophenol (4-BP) is a toxic halogenated phenolic organic compound. The conventional treatment processes for 4-BP elimination are costly and inefficient, with complete mineralization remaining a challenge for water treatment. To overcome these limitations, we investigated the treatment of 4-BP in a membrane biofilm reactor (MBfR) using hydrogen as an electron donor. The pathway of 4-BP degradation within the H 2 -MBfR was investigated through long-term operational experiments by considering the effect of nitrate and 4-BP concentrations, hydrogen partial pressure, static experiments, and microbial community diversity, which was studied using 16S rRNA. The results showed that H 2 -MBfR could quickly remove approximately 100% of 4-BP (up to 20 mg/L), with minimal intermediate product accumulation and 10 mg/L of nitrate continuously reduced. The microbial community structure showed that the presence of H 2 created an anaerobic environment, and Thauera was the dominant functional genus involved in the degradation of 4-BP. The genes encoding related enzymes were further enhanced. This study provides an economically viable and environmentally friendly bioremediation technique for water bodies that contain 4-BP and nitrates.

Lactic acid has been applied as a precursor for hydrogen (H 2 ) production from substrates rich in lactic acid bacteria (LAB), focusing on microbial interactions between producing and consuming LAB tested with model substrates. Therefore, this study evaluated the effect of single and combined lactic acid-consuming bacteria on mesophilic H 2 production in batch tests from lactic acid from fermented food waste (FW). Megasphaera elsdenii, Clostridium beijerinckii, and Clostridium butyricum were inoculated at different ratios (v/v). Additionally, thermal pretreated sludge (TPS) was added to the strain mixtures. The highest production was obtained with M. elsdenii, C. beijerinckii, and C. butyricum (17:66:17 ratio), obtaining 1629.0 mL/L reactor . The optimal mixture (68:32:0 of M. elsdenii and C. beijerinckii) enriched with TPS reached 1739.3 ± 98.6 mL H 2 /L reactor , consuming 98 % of lactic acid added. M. elsdenii and Clostridium strains enhance H 2 production from lactic acid as they persist in a microbial community initially dominated by LAB., 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 © 2024. Published by Elsevier Ltd.)

In order to further understand the effect of product inhibition on the metabolism of hydrogen production bacteria, and to seek an effective way to increase the hydrogen yield in fermentation, a simplified metabolic model of Ethanoligenens harbinense B49 was constructed to analyse the metabolic flux under acetate and ethanol inhibition separately and to analyse the flux changes of the nodes. Based on the changes in metabolic flux distribution, Glucose 6-phosphate (G6P), Pyruvate (PYR), and Acetyl-CoA (AcCoA) were identified as key nodes of hydrogen production in the metabolic network. Robustness analysis showed that G6P was flexible, while AcCoA and PYR were weakly rigid, indicating that acetate flux could be increased by adding inhibitors or using genetic manipulation. Furthermore, releasing inhibition of acetate could effectively increase hydrogen production. These findings suggested that the addition of acetate separation in ethanol-type fermentation process is expected to improve hydrogen production, which might be a promising way to full-scale produce biohydrogen in industrial applications. Further, for the first time, we report the effect of product inhibition on key nodes in the E. harbinense B49 hydrogen production metabolism network.

During in situ biomethanation, microbial communities can convert complex Organic Matter (OM) and H 2 into CH 4 . OM biodegradation was compared between Anaerobic Digestion (AD) and in situ biomethanation, in semi-continuous processes, using two inocula from the digester (D) and the post-digester (PoD) of an AD plant. The impact of H 2 on OM degradation was assessed using a fractionation method. Operational parameters included 20 days of hydraulic retention time and 1.5 g VS .L -1 .d -1 of organic loading rate. During in situ biomethanation, 485 NmL of H 2 were injected for each feeding (3 times a week). Maximum organic COD removal was 0.6 gCOD in AD control and at least 1.6 gCOD for in situ biomethanation. Therefore, COD removal was 2.5 times higher with H 2 injections. These results bring out the potential of H 2 injections during AD, not only for CO 2 consumption but also for better OM degradation., 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 © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

The conversion of carbon dioxide (CO 2 ) to methane (CH 4 ) is a strategy for sequestering CO 2 . Zero-valent iron (ZVI) has been proposed as an alternative electron donor for the CO 2 reduction to CH 4 . In this study, the effects of ZVI concentrations on the abiotic production of H 2 (without the action of microorganisms) in the first part and on the biological conversion of CO 2 to CH 4 using ZVI as a direct electron donor in the second part were examined. In the abiotic H 2 production, the increase in the ZVI concentration from 16 to 32, 64, and 96 g/L was found to have positive effects on both the amounts of H 2 generated and the rates of H 2 production because the extent of ZVI oxidation positively correlates with increasing surface area. Nevertheless, the increase in ZVI concentration from 96 to 224 g/L did not benefit the H 2 production because the ZVI dissolution was suppressed by the increasing aqueous pH above 10. In the bioconversion of CO 2 to CH 4 using ZVI as an electron donor, the main methanogenesis pathway occurred via hydrogenotrophic methanogenesis at pH 8.7-9.5 driven by the genus Methanobacterium of the class Methanobacteria. At ZVI concentrations of 64 g/L and above, the production of volatile fatty acid (VFA) became clear. Acetate was the main VFA, indicating the induction of homoacetogenesis at ZVI concentrations of 64 g/L and above. In addition, the presence of propionate as the second major VFA suggests the production of propionate from CO 2 and acetate under conditions with high H 2 partial pressure. The results indicated that the pathway for ZVI/CO 2 conversion to CH 4 was competitive between hydrogenotrophic methanogenesis and homoacetogenesis., 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 © 2024. Published by Elsevier Inc.)

Hydrogen gas (H 2 ), a gaseous signaling molecule, is involved in plant growth and development. This review collates emerging evidence to show that H 2 regulates the postharvest senescence of horticultural products through critical biochemical processes, including the improvement of antioxidant systems, the activation of cell wall metabolism, the promotion of energy metabolism, the inhibition of ethylene biosynthesis and the regulation of bacterial communities. Additionally, the interactions between H 2 and other signaling molecules are also discussed. This paper presents the current status of H 2 research in terms of its biological effects and safety in postharvest products by combining the research results on the molecular mechanisms of biological effects and H 2 signaling. The action mechanism of H 2 for postharvest preservation is also proposed, and it reflects the complexity and diversity of the pathways involved. Furthermore, a growing body of evidence has found a large number of downstream pathways or targets for the medical effects of H 2 . Therefore, the scientific and practical aspects of H 2 biology are proposed for the postharvest preservation of horticultural products., 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 © 2024 Elsevier Ltd. All rights reserved.)

Carbon capture and utilization (CCU) covers an array of technologies for valorizing carbon dioxide (CO 2 ). To date, most mature CCU technology conducted with capture agents operates against the CO 2 gradient to desorb CO 2 from capture agents, exhibiting high energy penalties and thermal degradation due to the requirement for thermal swings. This Perspective presents a concept of Bio-Integrated Carbon Capture and Utilization (BICCU), which utilizes methanogens for integrated release and conversion of CO 2 captured with capture agents. BICCU hereby substitutes the energy-intensive desorption with microbial conversion of captured CO 2 by the methanogenic CO 2 -reduction pathway, utilizing green hydrogen to generate non-fossil methane., (© 2024. The Author(s).)

Resource partitioning is central to the incredible productivity of microbial communities, including gigatons in annual methane emissions through syntrophic interactions. Previous work revealed how a sulfate reducer (Desulfovibrio vulgaris, Dv) and a methanogen (Methanococcus maripaludis, Mm) underwent evolutionary diversification in a planktonic context, improving stability, cooperativity, and productivity within 300-1000 generations. Here, we show that mutations in just 15 Dv and 7 Mm genes within a minimal assemblage of this evolved community gave rise to co-existing ecotypes that were spatially enriched within a few days of culturing in a fluidized bed reactor. The spatially segregated communities partitioned resources in the simulated subsurface environment, with greater lactate utilization by attached Dv but partial utilization of resulting H 2 by low affinity hydrogenases of Mm in the same phase. The unutilized H 2 was scavenged by high affinity hydrogenases of planktonic Mm, producing copious amounts of methane. Our findings show how a few mutations can drive resource partitioning amongst niche-differentiated ecotypes, whose interplay synergistically improves productivity of the entire mutualistic community., (© 2024. The Author(s).)

Various environmental factors, including H 2 availability, metabolic tradeoffs, optimal growth temperature, stochasticity, and hydrology, were examined to determine if they affect microbial competition between three autotrophic thermophiles. The thiosulfate reducer Desulfurobacterium thermolithotrophum ( T opt 72°C) was grown in mono- and coculture separately with the methanogens Methanocaldococcus jannaschii ( T opt 82°C) at 72°C and Methanothermococcus thermolithotrophicu s ( T opt 65°C) at 65°C at high and low H 2 concentrations. Both methanogens showed a metabolic tradeoff shifting from high growth rate-low cell yield at high H 2 concentrations to low growth rate-high cell yield at low H 2 concentrations and when grown in coculture with the thiosulfate reducer. In 1:1 initial ratios, D. thermolithotrophum outcompeted both methanogens at high and low H 2 , no H 2 S was detected on low H 2 , and it grew with only CO 2 as the electron acceptor indicating a similar metabolic tradeoff with low H 2 . When the initial methanogen-to-thiosulfate reducer ratio varied from 1:1 to 10 4 :1 with high H 2 , D. thermolithotrophum always outcompeted M. jannaschii at 72°C. However, M. thermolithotrophicus outcompeted D. thermolithotrophum at 65°C when the ratio was 10 3 :1. A reactive transport model that mixed pure hydrothermal fluid with cold seawater showed that hyperthermophilic methanogens dominated in systems where the residence time of the mixed fluid above 72°C was sufficiently high. With shorter residence times, thermophilic thiosulfate reducers dominated. If residence times increased with decreasing fluid temperature along the flow path, then thermophilic methanogens could dominate. Thermophilic methanogen dominance spread to previously thiosulfate-reducer-dominated conditions if the initial ratio of thermophilic methanogen-to-thiosulfate reducer increased., Importance: The deep subsurface is the largest reservoir of microbial biomass on Earth and serves as an analog for life on the early Earth and extraterrestrial environments. Methanogenesis and sulfur reduction are among the more common chemolithoautotrophic metabolisms found in hot anoxic hydrothermal vent environments. Competition between H2-oxidizing sulfur reducers and methanogens is primarily driven by the thermodynamic favorability of redox reactions with the former outcompeting methanogens. This study demonstrated that competition between the hydrothermal vent chemolithoautotrophs Methanocaldococcus jannaschii , Methanothermococcus thermolithotrophicus , and Desulfurobacterium thermolithotrophum is also influenced by other overlapping factors such as staggered optimal growth temperatures, stochasticity, and hydrology. By modeling all aspects of microbial competition coupled with field data, a better understanding is gained on how methanogens can outcompete thiosulfate reducers in hot anoxic environments and how the deep subsurface contributes to biogeochemical cycling., Competing Interests: The authors declare no conflict of interest.

ConspectusLife is an exergonic chemical reaction. The same was true when the very first cells emerged at life's origin. In order to live, all cells need a source of carbon, energy, and electrons to drive their overall reaction network (metabolism). In most cells, these are separate pathways. There is only one biochemical pathway that serves all three needs simultaneously: the acetyl-CoA pathway of CO 2 fixation. In the acetyl-CoA pathway, electrons from H 2 reduce CO 2 to pyruvate for carbon supply, while methane or acetate synthesis are coupled to energy conservation as ATP. This simplicity and thermodynamic favorability prompted Georg Fuchs and Erhard Stupperich to propose in 1985 that the acetyl-CoA pathway might mark the origin of metabolism, at the same time that Steve Ragsdale and Harland Wood were uncovering catalytic roles for Fe, Co, and Ni in the enzymes of the pathway. Subsequent work has provided strong support for those proposals.In the presence of Fe, Co, and Ni in their native metallic state as catalysts, aqueous H 2 and CO 2 react specifically to formate, acetate, methane, and pyruvate overnight at 100 °C. These metals (and their alloys) thus replace the function of over 120 enzymes required for the conversion of H 2 and CO 2 to pyruvate via the pathway and its cofactors, an unprecedented set of findings in the study of biochemical evolution. The reactions require alkaline conditions, which promote hydrogen oxidation by proton removal and are naturally generated in serpentinizing (H 2 -producing) hydrothermal vents. Serpentinizing hydrothermal vents furthermore produce natural deposits of native Fe, Co, Ni, and their alloys. These are precisely the metals that reduce CO 2 with H 2 in the laboratory; they are also the metals found at the active sites of enzymes in the acetyl-CoA pathway. Iron, cobalt and nickel are relicts of the environments in which metabolism arose, environments that still harbor ancient methane- and acetate-producing autotrophs today. This convergence indicates bedrock-level antiquity for the acetyl-CoA pathway. In acetogens and methanogens growing on H 2 as reductant, the acetyl-CoA pathway requires flavin-based electron bifurcation as a source of reduced ferredoxin (a 4Fe4S cluster-containing protein) in order to function. Recent findings show that H 2 can reduce the 4Fe4S clusters of ferredoxin in the presence of native iron, uncovering an evolutionary precursor of flavin-based electron bifurcation and suggesting an origin of FeS-dependent electron transfer in proteins. Traditionally discussed as catalysts in early evolution, the most common function of FeS clusters in metabolism is one-electron transfer, also in radical SAM enzymes, a large and ancient enzyme family. The cofactors and active sites in enzymes of the acetyl-CoA pathway uncover chemical antiquity in metabolism involving metals, methyl groups, methyl transfer reactions, cobamides, pterins, GTP, S -adenosylmethionine, radical SAM enzymes, and carbon-metal bonds. The reaction sequence from H 2 and CO 2 to pyruvate on naturally deposited native metals is maximally simple. It requires neither nitrogen, sulfur, phosphorus, RNA, ion gradients, nor light. Solid-state metal catalysts tether the origin of metabolism to a H 2 -producing, serpentinizing hydrothermal vent.

Background: Carbon monoxide (CO), hypothetically linked to prebiotic biosynthesis and possibly the origin of the life, emerges as a substantive growth substrate for numerous microorganisms. In anoxic environments, the coupling of CO oxidation with hydrogen (H 2 ) production is an essential source of electrons, which can subsequently be utilized by hydrogenotrophic bacteria (e.g., organohalide-respring bacteria). While Dehalococcoides strains assume pivotal roles in the natural turnover of halogenated organics and the bioremediation of chlorinated ethenes, relying on external H 2 as their electron donor and acetate as their carbon source, the synergistic dynamics within the anaerobic microbiome have received comparatively less scrutiny. This study delves into the intriguing prospect of CO serving as both the exclusive carbon source and electron donor, thereby supporting the reductive dechlorination of trichloroethene (TCE)., Results: The metabolic pathway involved anaerobic CO oxidation, specifically the Wood-Ljungdahl pathway, which produced H 2 and acetate as primary metabolic products. In an intricate microbial interplay, these H 2 and acetate were subsequently utilized by Dehalococcoides, facilitating the dechlorination of TCE. Notably, Acetobacterium emerged as one of the pivotal collaborators for Dehalococcoides, furnishing not only a crucial carbon source essential for its growth and proliferation but also providing a defense against CO inhibition., Conclusions: This research expands our understanding of CO's versatility as a microbial energy and carbon source and unveils the intricate syntrophic dynamics underlying reductive dechlorination., (© 2024. The Author(s).)

The fusion of hydrogenases and photosynthetic reaction centers (RCs) has proven to be a promising strategy for the production of sustainable biofuels. Type I (iron-sulfur-containing) RCs, acting as photosensitizers, are capable of promoting electrons to a redox state that can be exploited by hydrogenases for the reduction of protons to dihydrogen (H 2 ). While both [FeFe] and [NiFe] hydrogenases have been used successfully, they tend to be limited due to either O 2 sensitivity, binding specificity, or H 2 production rates. In this study, we fuse a peripheral (stromal) subunit of Photosystem I (PS I), PsaE, to an O 2 -tolerant [FeFe] hydrogenase from Clostridium beijerinckii using a flexible [GGS] 4 linker group ( Cb HydA1-PsaE). We demonstrate that the Cb HydA1 chimera can be synthetically activated in vitro to show bidirectional activity and that it can be quantitatively bound to a PS I variant lacking the PsaE subunit. When illuminated in an anaerobic environment, the nanoconstruct generates H 2 at a rate of 84.9 ± 3.1 µmol H 2 mg chl -1 h -1 . Further, when prepared and illuminated in the presence of O 2 , the nanoconstruct retains the ability to generate H 2 , though at a diminished rate of 2.2 ± 0.5 µmol H 2 mg chl -1 h -1 . This demonstrates not only that PsaE is a promising scaffold for PS I-based nanoconstructs, but the use of an O 2 -tolerant [FeFe] hydrogenase opens the possibility for an in vivo H 2 generating system that can function in the presence of O 2 ., Competing Interests: Competing interests statement:The authors declare no competing interest.

Thermophilic acetogenic bacteria have attracted attention as promising candidates for biotechnological applications such as syngas fermentation, microbial electrosynthesis, and methanol conversion. Here, we aimed to isolate and characterize novel thermophilic acetogens from diverse environments. Enrichment of heterotrophic and autotrophic acetogens was monitored by 16S rRNA gene-based bacterial community analysis. Seven novel Moorella strains were isolated and characterized by genomic and physiological analyses. Two Moorella humiferrea isolates showed considerable differences during autotrophic growth. The M. humiferrea LNE isolate (DSM 117358) fermented carbon monoxide (CO) to acetate, while the M. humiferrea OCP isolate (DSM 117359) transformed CO to hydrogen and carbon dioxide (H2 + CO2), employing the water-gas shift reaction. Another carboxydotrophic hydrogenogenic Moorella strain was isolated from the covering soil of an active charcoal burning pile and proposed as the type strain (ACPsT) of the novel species Moorella carbonis (DSM 116161T and CCOS 2103T). The remaining four novel strains were affiliated with Moorella thermoacetica and showed, together with the type strain DSM 2955T, the production of small amounts of ethanol from H2 + CO2 in addition to acetate. The physiological analyses of the novel Moorella strains revealed isolate-specific differences that considerably increase the knowledge base on thermophilic acetogens for future applications., (© The Author(s) 2024. Published by Oxford University Press on behalf of FEMS.)

We examined the microbial populations present in fecal samples of macropods capable of utilizing a mixture of hydrogen and carbon dioxide (70:30) percent. The feces samples were cultured under anaerobic conditions, and production of methane or acetic acids characteristic for methanogenesis and homoacetogenesis was measured. While the feces of adult macropods mainly produced methane from the substrate, the sample from a 2-month-old juvenile kangaroo only produced acetic acid and no methane. The stable highly enriched culture of the joey kangaroo was sequenced to examine the V3 and V4 regions of the 16S rRNA gene. The results showed that over 70% of gene copies belonged to the Clostridia class, with Paraclostridium and Blautia as the most predominant genera. The culture further showed the presence of Actinomyces spp., a genus which has only been identified in the GI tract of macropods in a few studies, and where none, to our knowledge, have been classified as homoacetogenic. The joey kangaroo mixed culture showed a doubling time of 3.54 h and a specific growth rate of 0.199/h, faster than what has been observed for homoacetogenic bacteria in general., Importance: Enteric methane emissions from cattle are a significant contributor to greenhouse gas emissions worldwide. Methane emissions not only contribute to climate change but also represent a loss of energy from the animal's diet. However, methanogens play an important role as hydrogen sink to rumen systems; without it, the performance of hydrolytic organisms diminishes. Therefore, effective strategies of methanogen inhibition would be enhanced in conjunction with the addition of alternative hydrogen sinks to the rumen. The significance of our research is to identify homoacetogens present in the GI tract of kangaroos and to present their performance in vitro , demonstrating their capability to serve as alternatives to rumen methanogens., Competing Interests: The authors declare no conflict of interest.

Molecular catalysts based on abundant elements that function in neutral water represent an essential component of sustainable hydrogen production. Artificial hydrogenases based on protein-inorganic hybrids have emerged as an intriguing class of catalysts for this purpose. We have prepared a novel artificial hydrogenase based on cobaloxime bound to a de novo three alpha-helical protein, α 3 C, via a pyridyl-based unnatural amino acid. The functionalized de novo protein was characterised by UV-visible, CD, and EPR spectroscopy, as well as MALDI spectrometry, which confirmed the presence and ligation of cobaloxime to the protein. The new de novo enzyme produced hydrogen under electrochemical, photochemical and reductive chemical conditions in neutral water solution. A change in hydrogen evolution capability of the de novo enzyme compared with native cobaloxime was observed, with turnover numbers around 80% of that of cobaloxime, and hydrogen evolution rates of 40% of that of cobaloxime. We discuss these findings in the context of existing literature, how our study contributes important information about the functionality of cobaloximes as hydrogen evolving catalysts in protein environments, and the feasibility of using de novo proteins for development into artificial metalloenzymes. Small de novo proteins as enzyme scaffolds have the potential to function as upscalable bioinspired catalysts thanks to their efficient atom economy, and the findings presented here show that these types of novel enzymes are a possible product.

Pulsed light illumination stands out as a noteworthy technique for photosynthetic H 2 production, playing a crucial role in eliminating O 2 and activating hydrogenase enzymes. However, further improvements are essential to make H 2 photoproduction suitable for future commercial applications. In our study, we observed a distinct enhancement in pulsed light-induced H 2 photoproduction in the unicellular green alga Chlamydomonas reinhardtii when treated with the optimal concentration of the mild O 2 scavenger Na 2 SO 3 . This improvement was a result of reduced O 2 content, increased hydrogenase enzyme activity, and suppressed H 2 -uptake activity. Furthermore, our findings indicate that exposing Na 2 SO 3 -treated C. reinhardtii to optimal light waveform continues to significantly boost pulsed light-induced H 2 photoproduction, attributed to the alleviation of impaired photosystem II activity. Altogether, the combined application of optimal sulfite concentration and light waveform effectively enhances pulsed light-induced photosynthetic H 2 production in the green alga C. reinhardtii., 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 © 2024 Elsevier B.V. All rights reserved.)

All aerobic life on Earth depends on oxygenic photosynthesis, occurring in both prokaryotic and eukaryotic organisms. This process can be divided into light reactions and carbon fixation. This special issue is a result of the International Conference on "Photosynthesis and Hydrogen Energy Research for Sustainability 2023," held in honor of Robert Blankenship, Győző Garab, Michael Grätzel, Norman Hüner, and Gunnar Öquist. After extensive discussions on various aspects of photosynthesis and hydrogen energy, eight high-quality papers were selected. These papers cover studies on abiotic stress, an overview of photosynthesis, thylakoid membrane lipid organization, energy transfer, and the genomics of both prokaryotic and eukaryotic photosynthesis, as well as biohydrogen production from cyanobacteria. The authors used new methods and techniques, likely bringing fresh ideas for improving biomass and crop yield., (© 2024. The Author(s), under exclusive licence to Springer Nature B.V.)

The impact of hydrogen (H 2 ) producing magnesium (Mg) incorporation into minced beef meat (MBM) on the quality and safety of the product was investigated. The H 2 -producing Mg (H2-P-Mg)-incorporated MBMs were vacuumed (VP) and stored at 4 °C for 12 days. Other MBMs were vacuumed and gassed with H 2 or N 2 . At the end of storage, the lowest browning index values were for H 2 and H2-P-Mg samples. H2- PMg and VP methods generally decreased the counts of mesophilic and psychrotrophic bacteria and yeast molds and restricted the formation of thiobarbituric acid reactive substances and biogenic amines. Heat mapping, PCA, and multivariate analysis methods confirmed chemical analysis results. The volatile compounds were at their highest levels in the control samples at the end of storage, followed by H 2 , N 2 , H2-P-Mg, and VP samples. Using the H2-P-Mg method in MBM preparation could protect the quality characteristics and safety of the product during cold storage., 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 © 2024 Elsevier Ltd. All rights reserved.)

Ciliate protozoa are an integral part of the rumen microbial community involved in a variety of metabolic processes. These processes are thought to be in part the outcome of interactions with their associated prokaryotic community. For example, methane production is enhanced through interspecies hydrogen transfer between protozoa and archaea. We hypothesize that ciliate protozoa are host to a stable prokaryotic community dictated by specific functions they carry. Here, we modify the microbial community by varying the forage-to-concentrate ratios and show that, despite major changes in the prokaryotic community, several taxa remain stably associated with ciliate protozoa. By quantifying genes belonging to various known reduction pathways in the rumen, we find that the bacterial community associated with protozoa is enriched in genes belonging to hydrogen utilization pathways and that these genes correspond to the same taxonomic affiliations seen enriched in protozoa. Our results show that ciliate protozoa in the rumen may serve as a hub for various hydrogenotrophic functions and a better understanding of the processes driven by different protozoa may unveil the potential role of ciliates in shaping rumen metabolism., (© 2024 The Author(s). Environmental Microbiology Reports published by John Wiley & Sons Ltd.)

Methane is the second most abundant climate-active gas, and understanding its sources and sinks is an important endeavour in microbiology, biogeochemistry, and climate sciences 1,2 . For decades, it was thought that methanogenesis, the ability to conserve energy coupled to methane production, was taxonomically restricted to a metabolically specialized group of archaea, the Euryarchaeota 1 . The discovery of marker genes for anaerobic alkane cycling in metagenome-assembled genomes obtained from diverse habitats has led to the hypothesis that archaeal lineages outside the Euryarchaeota are also involved in methanogenesis 3-6 . Here we cultured Candidatus Methanosuratincola verstraetei strain LCB70, a member of the archaeal class Methanomethylicia (formerly Verstraetearchaeota) within the phylum Thermoproteota, from a terrestrial hot spring. Growth experiments combined with activity assays, stable isotope tracing, and genomic and transcriptomic analyses demonstrated that this thermophilic archaeon grows by means of methyl-reducing hydrogenotrophic methanogenesis. Cryo-electron tomography revealed that Ca. M. verstraetei are coccoid cells with archaella and chemoreceptor arrays, and that they can form intercellular bridges connecting two to three cells with continuous cytoplasm and S-layer. The wide environmental distribution of Ca. M. verstraetei suggests that they might play important and hitherto overlooked roles in carbon cycling within diverse anoxic habitats., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Methanogenesis mediated by archaea is the main source of methane, a strong greenhouse gas, and thus is critical for understanding Earth's climate dynamics. Recently, genes encoding diverse methanogenesis pathways have been discovered in metagenome-assembled genomes affiliated with several archaeal phyla 1-7 . However, all experimental studies on methanogens are at present restricted to cultured representatives of the Euryarchaeota. Here we show methanogenic growth by a member of the lineage Korarchaeia within the phylum Thermoproteota (TACK superphylum) 5-7 . Following enrichment cultivation of 'Candidatus Methanodesulfokora washburnenis' strain LCB3, we used measurements of metabolic activity and isotope tracer conversion to demonstrate methanol reduction to methane using hydrogen as an electron donor. Analysis of the archaeon's circular genome and transcriptome revealed unique modifications in the energy conservation pathways linked to methanogenesis, including enzyme complexes involved in hydrogen and sulfur metabolism. The cultivation and characterization of this new group of archaea is critical for a deeper evaluation of the diversity, physiology and biochemistry of methanogens., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Rhodospirillum rubrum is a photosynthetic purple non-sulphur bacterium with great potential to be used for complex waste valorisation in biotechnological applications due to its metabolic versatility. This study investigates the production of hydrogen (H 2 ) and polyhydroxyalkanoates (PHA) by R. rubrum from syngas under photoheterotrophic conditions. An adaptive laboratory evolution strategy (ALE) has been carried out to improve the yield of the process. After 200 generations, two evolved strains were selected that showed reduced lag phase and enhanced poly-3-hydroxybutyrate (PHB) and H 2 synthesis compared to the parental strain. Genomic analysis of the photo-adapted (PA) variants showed four genes with single point mutations, including the photosynthesis gene expression regulator PpsR. The proteome of the variants suggested that the adapted variants overproduced H 2 due to a more efficient CO oxidation through the CO-dehydrogenase enzyme complex and confirmed that energy acquisition was enhanced through overexpression of the photosynthetic system and metal cofactors essential for pigment biosynthesis., 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 © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Methanogenic archaea are main contributors to methane emissions, and have a crucial role in carbon cycling and global warming. Until recently, methanogens were confined to Euryarchaeota, but metagenomic studies revealed the presence of genes encoding the methyl coenzyme M reductase complex in other archaeal clades 1-4 , thereby opening up the premise that methanogenesis is taxonomically more widespread. Nevertheless, laboratory cultivation of these non-euryarchaeal methanogens was lacking to corroborate their potential methanogenic ability and physiology. Here we report the isolation of a thermophilic archaeon LWZ-6 from an oil field. This archaeon belongs to the class Methanosuratincolia (originally affiliated with 'Candidatus Verstraetearchaeota') in the phylum Thermoproteota. Methanosuratincola petrocarbonis LWZ-6 is a strict hydrogen-dependent methylotrophic methanogen. Although previous metagenomic studies speculated on the fermentative potential of Methanosuratincolia members, strain LWZ-6 does not ferment sugars, peptides or amino acids. Its energy metabolism is linked only to methanogenesis, with methanol and monomethylamine as electron acceptors and hydrogen as an electron donor. Comparative (meta)genome analysis confirmed that hydrogen-dependent methylotrophic methanogenesis is a widespread trait among Methanosuratincolia. Our findings confirm that the diversity of methanogens expands beyond the classical Euryarchaeota and imply the importance of hydrogen-dependent methylotrophic methanogenesis in global methane emissions and carbon cycle., (© 2024. The Author(s), under exclusive licence to Springer Nature Limited.)

Cupriavidus necator is a facultative chemolithoautotrophic bacterium able to convert carbon dioxide into poly-3-hydroxybutyrate. This is highly promising as the conversion process allows the production of sustainable and biodegradable plastics. Poly-3-hydroxybutyrate accumulation is known to be induced by nutrient starvation, but information regarding the optimal stress conditions controlling the process is still heterogeneous and fragmentary. This study presents a comprehensive comparison of the effects of nutrient stress conditions, namely nitrogen, hydrogen, phosphorus, oxygen, and magnesium deprivation, on poly-3-hydroxybutyrate accumulation in C. necator DSM545. Nitrogen starvation exhibited the highest poly-3-hydroxybutyrate accumulation, achieving 54% of total cell dry weight after four days of nutrient stress, and a carbon conversion efficiency of 85%. The gas consumption patterns indicated flexible physiological mechanisms underlying polymer accumulation and depolymerization. These findings provide insights into strategies for efficient carbon conversion into bioplastics, and highlight the key role of C. necator for future industrial-scale applications., 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 © 2024 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Biological H 2 -assisted biogas upgrading has gained significant attention as an environmentally friendly substitute to common physico-chemical upgrading techniques, but is largely limited by the low solubility of H 2 . This study evaluated the design of a ceramic membrane contactor module for H 2 injection. H 2 dissolution was maintained at high efficiency by controlling gas supply and sludge recirculation rate, achieving a biogas quality of average 98.8% CH 4 during the stable operation phase with a 108% increase in the CH 4 production rate. This also outperforms conventional H 2 injection using diffuser sparging which could only achieve a biogas quality of 84% CH 4 content. Microbial community analysis found high Methanobacterium spp. abundance within the archaea at 95.2% at the end of the operation, allowing the dominance of the hydrogenotrophic methanogenesis pathway for high upgrading efficiencies. The system is a high-performance external membrane connector module coupled to common anaerobic digestion systems for biogas upgrading., 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 © 2024 Elsevier Ltd. All rights reserved.)

Achieving bacterial killing and osteogenic formation on an implant surface rarely occurs. In this study, a novel surface design-a palladium hydride (PdH x ) film that enables these two distinct features to coexist is introduced. The PdH x lattice captures protons in the extracellular microenvironment of bacteria, disrupting their normal metabolic activities, such as ATP synthesis, nutrient co-transport, and oxidative stress. This disruption leads to significant bacterial death, as evidenced by RNA sequence analysis. Additionally, the unique enzymatic activity and hydrogen-loading properties of PdH x activate the human antioxidant system, resulting in the rapid clearance of reactive oxygen species. This process reshapes the osteogenic immune microenvironment, promoting accelerated osteogenesis. These findings reveal that the downregulation of the NOD-like receptor signaling pathway is critical for activating immune cells toward M2 phenotype polarization. This novel surface design provides new strategies for modifying implant coatings to simultaneously prevent bacterial infection, reduce inflammation, and enhance tissue regeneration, making it a noteworthy contribution to the field of advanced materials., (© 2024 The Author(s). Advanced Materials published by Wiley‐VCH GmbH.)