John D. Coates

Organic compounds with a negative nominal oxidation state of carbon (NOSC) are thermodynamically recalcitrant in anaerobic ecosystems, but few studies have measured the influence of NOSC on carbon degradation rates, gaseous product yields, or microbiome composition. We amended anaerobic rice paddy sediment microcosms with water-soluble monomeric organic carbon compounds varying in NOSC. Consistent with thermodynamic and stoichiometric predictions, negative NOSC compounds are catabolized more slowly but produce more methane per mole of carbon. Negative NOSC microbiomes have higher alpha diversity, more syntrophs and methanogens, and fewer fermentative bacteria. Strikingly, fermentative bacterial taxa display genomically encoded NOSC catabolic preferences both in the lab and field. Negative NOSC-preferring fermenters have longer predicted doubling times, consistent with the thermodynamic recalcitrance of their preferred substrates. We propose that microbial NOSC catabolic preferences may reflect the thermodynamic niche of microorganisms and we anticipate that extending research on microbial catabolic preferences to a greater variety of organic carbon substrates and diverse microbiomes will improve our understanding of microbial carbon cycling and trait evolution.

Polyhydroxyalkanoates (PHAs) are widespread microbial storage polymers increasingly recognized for roles beyond carbon and energy storage, including redox homeostasis and stress physiology. While PHA accumulation is classically associated with stationary-phase metabolism under severe nutrient imbalance, comparatively little is known about growth-associated PHA synthesis in facultative anaerobes with unusual respiratory strategies. Here, we investigated polyhydroxybutyrate (PHB) metabolism in Azospira suillum PS, a genetically tractable perchlorate-reducing bacterium capable of both aerobic and anaerobic respiration. Using physiological growth experiments, targeted gene deletions, PHB extractions, and intracellular redox measurements, we examined PHB accumulation under varying respiratory and nutrient conditions. We demonstrate that PHB accumulates during exponential growth under moderately nitrogen-limited conditions, both aerobically and during perchlorate respiration, representing a rare example of growth-associated PHB synthesis under anaerobic conditions. Genomic analysis revealed four phaC homologs, one of which could not be deleted under the experimental conditions tested and co-localized with phaB and phaR . Redox profiling further revealed a strong positive correlation between PHB accumulation and intracellular NADPH/NADP ⁺ ratios. Together, these findings expand the physiological contexts in which PHB synthesis is known to occur and highlight perchlorate-respiring bacteria as underexplored model systems for studying growth-integrated carbon and redox storage strategies.

The anaerobic conversion of organic matter to methane and carbon dioxide typically relies on obligate syntrophic interactions between bacteria and methanogenic archaea, where interspecies electron transfer enables thermodynamically constrained reactions to proceed near equilibrium. Syntrophs often couple the oxidation of fermentation products such as fatty acids and alcohols to the reduction of protons to form hydrogen (H ₂ ). These reactions can only proceed if low H ₂ concentrations are maintained by H ₂ -consuming syntrophic partners. Here, we describe “lithosyntrophy,” a mode of syntrophic interaction in which electrons that drive hydrogenotrophic methanogenesis originate from an inorganic compound rather than from the canonical organic substrates. Candidatus Phosphitivorax anaerolimi Phox-21 oxidizes phosphite (HPO ^{₃ 2−} , oxidation state +3) to phosphate coupled to hydrogenogenesis in an obligate energetic dependency on a hydrogenotrophic methanogen, Methanoculleus sp. Physiology experiments, thermodynamic calculations, genomic annotation, and metaproteomics analysis collectively revealed a mechanism for syntrophic phosphite oxidation in Phox-21. In this pathway, electrons derived from phosphite drive H ₂ production via an electron-confurcating hydrogenase. Unlike previously characterized acetogenic phosphite oxidizers, Phox-21 is a mixotroph that assimilates acetate to form biomass. Lithosyntrophic phosphite oxidizers may play important roles both in transferring reducing equivalents as well as biologically available phosphorus to other members of their surrounding microbial communities, establishing a previously unrecognized metabolic and biogeochemical link between the phosphorus and carbon redox cycles in anoxic ecosystems.

The anaerobic conversion of organic matter to methane and carbon dioxide typically relies on obligate syntrophic interactions between bacteria and methanogenic archaea, where interspecies hydrogen (H ₂ ) transfer enables thermodynamically constrained reactions to proceed near equilibrium. Syntrophs couple the oxidation of fermentation products such as fatty acids and alcohols to the reduction of protons to form H ₂ . These reactions can only proceed if low H ₂ concentrations are maintained by H ₂ -consuming syntrophic partners. Here, we describe "lithosyntrophy," a novel mode of syntrophic interaction in which electrons that drive hydrogenotrophic methanogenesis originate from an inorganic compound rather than from the canonical organic substrates. Candidatus Phosphitivorax anaerolimi strain Phox-21 oxidizes phosphite (HPOH ₃ ^2- , oxidation state +3) to phosphate coupled to hydrogenogenesis in an obligate energetic dependency on a hydrogenotrophic methanogen, Methanoculleus sp. Physiology experiments, thermodynamic calculations, genomic annotation, and metaproteomics analysis collectively revealed a mechanism for syntrophic phosphite oxidation in Phox-21, which requires phosphite, acetate, and CO ₂ as co-substrates. In this pathway, electrons derived from phosphite drive H ₂ production via an electron-confurcating hydrogenase. Unlike previously characterized acetogenic phosphite oxidizers that grow without exogenous acetate, Phox-21 requires acetate to regenerate AMP, a cofactor required by the phosphite dehydrogenase, PtdF. Lithosyntrophic phosphite oxidizers may play important roles both in transferring reducing equivalents as well as biologically available phosphorus to other members of their surrounding microbial communities. We infer that lithosyntrophic DPO emerged before acetoclastic methanogenesis and was a major sink for acetate in the Archaean when phosphite was more abundant.

SAR11 bacteria (order Pelagibacterales ) are oligotrophs and often the most abundant bacterioplankton in aquatic environments. A subset of sequenced SAR11 genomes, predominantly in the brackish and freshwater SAR11 subclades, contain homologs of pha genes, which in other organisms confer the ability to store carbon and energy via polyhydroxyalkanoate (PHA) polymers. Here, we investigated the relevance of PHA production to SAR11 biology. Phylogenetics showed that Pha proteins occurred on a long branch and provided evidence for origin at the common ancestor of the brackish IIIa and freshwater LD12 subclades, followed by horizontal transfer within SAR11. Using the LD12 representative “ Candidatus Fonsibacter ubiquis” strain LSUCC0530, we found that a large minority of LSUCC0530 cells contained a single Nile red-staining granule, confirmed that the cells produced polyhydroxybutyrate (PHB), and estimated the total PHB content in the cells. We heterologously expressed the LSUCC0530 phaCAB locus in Escherichia coli , finding it to be functional and the likely origin of the PHB. We also determined that, irrespective of changes to carbon, nitrogen, and phosphorus concentrations, a similar fraction of LSUCC0530 cells generated PHB granules and expression of the phaCAB locus remained constant. We suggest that PHB synthesis in LSUCC0530 may be constitutively active due to the slow growth dynamics and minimal regulation that characterize SAR11 bacteria. This is the first characterization of polymer storage in SAR11, providing new insights into the likely fitness advantage for cells harboring this metabolism.

Microorganisms are essential drivers of earth’s geochemical cycles. However, the significance of elemental redox cycling mediated by microorganisms is often underestimated beyond the most well-studied nutrient cycles. Phosphite, (per)chlorate, and iodate are each considered esoteric substrates metabolized by microorganisms. However, recent investigations have indicated that these metabolisms are widespread and ubiquitous, affirming a need to continue studying the underlying microbiology to understand their biogeochemical effects and their interface with each other and our biosphere. This review focuses on combining canonical techniques of culturing microorganisms with modern omic approaches to further our understanding of obscure metabolic pathways and elucidate their importance in global biogeochemical cycles. Using these approaches, marker genes of interest have already been identified for phosphite, (per)chlorate, and iodate using traditional microbial physiology and genetics. Subsequently, their presence was queried to reveal the distribution of metabolic pathways in the environment using publicly available databases. In conjunction with each other, computational and experimental techniques provide a more comprehensive understanding of the location of these microorganisms, their underlying biochemistry and genetics, and how they tie into our planet’s geochemical cycles.

The genus Denitromonas is currently a non-validated taxon that has been identified in several recent publications as members of microbial communities arising from marine environments. Very little is known about the biology of Denitromonas spp., and no pure cultures are presently found in any culture collections. The current epitaph of Denitromonas was given to the organism under the assumption that all members of this genus are denitrifying bacteria. This study performs phenotypic and genomic analyses on three new Denitromonas spp. isolated from tidal mudflats in the San Francisco Bay. We demonstrate that Denitromonas spp. are indeed all facultative denitrifying bacteria that utilize a variety of carbon sources such as acetate, lactate, and succinate. In addition, individual strains also use the esoteric electron acceptors perchlorate, chlorate, and iodate. Both 16S and Rps/Rpl phylogenetic analyses place Denitromonas spp. as a deep branching clade in the family Zoogloeaceae , separate from either Thauera spp., Azoarcus spp., or Aromatoleum spp. Genome sequencing reveals a G + C content ranging from 63.72% to 66.54%, and genome sizes range between 4.39 and 5.18 Mb. Genes for salt tolerance and denitrification are distinguishing features that separate Denitromonas spp. from the closely related Azoarcus and Aromatoleum genera. IMPORTANCE: The genus Denitromonas is currently a non-validated taxon that has been identified in several recent publications as members of microbial communities arising from marine environments. Very little is known about the biology of Denitromonas spp., and no pure cultures are presently found in any culture collections. The current epitaph of Denitromonas was given to the organism under the assumption that all members of this genus are denitrifying bacteria. This study performs phenotypic and genomic analyses on three Denitromonas spp., Denitromonas iodatirespirans sp. nov.—a novel iodate-reducing bacterium—and two novel perchlorate-reducing bacteria, Denitromonas halophila and Denitromonas ohlonensis , isolated from San Francisco Bay intertidal mudflats.