Disproportionate Carbon Dioxide Efflux in Bacterial Metabolic Pathways for Different Organic Substrates Leads to Variable Contribution to Carbon-Use Efficiency.

Microbial organic matter turnover is an important contributor to the terrestrial carbon dioxide (CO2) budget. Partitioning of organic carbons into biomass relative to CO2 efflux, termed carbon-use efficiency (CUE), is widely used to characterize organic carbon cycling by soil microorganisms. Recent studies challenge proposals of CUE dependence on the oxidation state of the substrate carbon and implicate instead metabolic strategies. Still unknown are the metabolic mechanisms underlying variability in CUE. We performed a multiomics investigation of these mechanisms in Pseudomonas putida, a versatile soil bacterium of the Gammaproteobacteria, processing a mixture of plant matter derivatives. Our 13C-metabolomics data captured substrate carbons into different metabolic pathways: cellulose-derived sugar carbons in glycolytic and pentose-phosphate pathways; lignin-related aromatic carbons in the tricarboxylic acid cycle. Subsequent 13C-metabolic flux analysis revealed a 3-fold lower investment of sugar carbons in CO2 efflux compared to aromatic carbons, in agreement with reported substrate-dependent CUE. Proteomics analysis revealed enzyme-level regulation only for substrate uptake and initial catabolism, which dictated downstream fluxes through CO2-producing versus biomass-synthesizing reactions. Metabolic partitioning as shown here explained the substrate-dependent CUE calculated from reported metabolic flux analyses of other bacteria, further supporting a metabolism-guided perspective for predicting the microbial conversion of accessible organic matter to CO2 efflux.

Energy flux couples sulfur isotope fractionation to proteomic and metabolite profiles in Desulfovibrio vulgaris.

Microbial sulfate reduction is central to the global carbon cycle and the redox evolution of Earth's surface. Tracking the activity of sulfate reducing microorganisms over space and time relies on a nuanced understanding of stable sulfur isotope fractionation in the context of the biochemical machinery of the metabolism. Here, we link the magnitude of stable sulfur isotopic fractionation to proteomic and metabolite profiles under different cellular energetic regimes. When energy availability is limited, cell-specific sulfate respiration rates and net sulfur isotope fractionation inversely covary. Beyond net S isotope fractionation values, we also quantified shifts in protein expression, abundances and isotopic composition of intracellular S metabolites, and lipid structures and lipid/water H isotope fractionation values. These coupled approaches reveal which protein abundances shift directly as a function of energy flux, those that vary minimally, and those that may vary independent of energy flux and likely do not contribute to shifts in S-isotope fractionation. By coupling the bulk S-isotope observations with quantitative proteomics, we provide novel constraints for metabolic isotope models. Together, these results lay the foundation for more predictive metabolic fractionation models, alongside interpretations of environmental sulfur and sulfate reducer lipid-H isotope data.

Membrane lipid and expression responses of Saccharolobus islandicus REY15A to acid and cold stress.

Archaea adjust the number of cyclopentane rings in their glycerol dibiphytanyl glycerol tetraether (GDGT) membrane lipids as a homeostatic response to environmental stressors such as temperature, pH, and energy availability shifts. However, archaeal expression patterns that correspond with changes in GDGT composition are less understood. Here we characterize the acid and cold stress responses of the thermoacidophilic crenarchaeon Saccharolobus islandicus REY15A using growth rates, core GDGT lipid profiles, transcriptomics and proteomics. We show that both stressors result in impaired growth, lower average GDGT cyclization, and differences in gene and protein expression. Transcription data revealed differential expression of the GDGT ring synthase grsB in response to both acid stress and cold stress. Although the GDGT ring synthase encoded by grsB forms highly cyclized GDGTs with ≥5 ring moieties, S. islandicus grsB upregulation under acidic pH conditions did not correspond with increased abundances of highly cyclized GDGTs. Our observations highlight the inability to predict GDGT changes from transcription data alone. Broader analysis of transcriptomic data revealed that S. islandicus differentially expresses many of the same transcripts in response to both acid and cold stress. These included upregulation of several biosynthetic pathways and downregulation of oxidative phosphorylation and motility. Transcript responses specific to either of the two stressors tested here included upregulation of genes related to proton pumping and molecular turnover in acid stress conditions and upregulation of transposases in cold stress conditions. Overall, our study provides a comprehensive understanding of the GDGT modifications and differential expression characteristic of the acid stress and cold stress responses in S. islandicus.

Seasonal shifts in community composition and proteome expression in a sulphur-cycling cyanobacterial mat.

Seasonal changes in light and physicochemical conditions have strong impacts on cyanobacteria, but how they affect community structure, metabolism, and biogeochemistry of cyanobacterial mats remains unclear. Light may be particularly influential for cyanobacterial mats exposed to sulphide by altering the balance of oxygenic photosynthesis and sulphide-driven anoxygenic photosynthesis. We studied temporal shifts in irradiance, water chemistry, and community structure and function of microbial mats in the Middle Island Sinkhole (MIS), where anoxic and sulphate-rich groundwater provides habitat for cyanobacteria that conduct both oxygenic and anoxygenic photosynthesis. Seasonal changes in light and groundwater chemistry were accompanied by shifts in bacterial community composition, with a succession of dominant cyanobacteria from Phormidium to Planktothrix, and an increase in diatoms, sulphur-oxidizing bacteria, and sulphate-reducing bacteria from summer to autumn. Differential abundance of cyanobacterial light-harvesting proteins likely reflects a physiological response of cyanobacteria to light level. Beggiatoa sulphur oxidation proteins were more abundant in autumn. Correlated abundances of taxa through time suggest interactions between sulphur oxidizers and sulphate reducers, sulphate reducers and heterotrophs, and cyanobacteria and heterotrophs. These results support the conclusion that seasonal change, including light availability, has a strong influence on community composition and biogeochemical cycling of sulphur and O2 in cyanobacterial mats.

Metaproteomics reveals functional partitioning and vegetational variation among permafrost-affected Arctic soil bacterial communities.

Microbial activity in Arctic soils controls the cycling of significant stores of organic carbon and nutrients. We studied in situ processes in Alaskan soils using original metaproteomic methods in order to relate important heterotrophic functions to microbial taxa and to understand the microbial response to Arctic greening. Major bacterial groups show strong metabolic specialization in organic topsoils. α-/ß-/γ-Proteobacteria specialized in the acquisition of small, soluble compounds, whereas Acidobacteria, Actinobacteria, and other detritosphere groups specialized in the degradation of plant-derived polymers. α-/ß-/γ-Proteobacteria dominated the expression of transporters for common root exudates and limiting nitrogenous compounds, supporting an ecological model of dependence upon plants for carbon and competition with plants for nitrogen. Detritosphere groups specialized in distinct substrates, with Acidobacteria producing the most enzymes for hemicellulose depolymerization. Acidobacteria was the most active group across the three plant ecotypes sampled-the largely nonvascular, lower biomass intertussock and the largely vascular, higher biomass tussock and shrub. Functional partitioning among bacterial groups was stable between plant ecotypes, but certain functions associated with α-/ß-/γ-Proteobacteria were more strongly expressed in higher biomass ecotypes. We show that refined metaproteomic approaches can elucidate soil microbial ecology as well as biogeochemical trajectories of major carbon stocks. IMPORTANCE The Arctic is warming twice as fast as the rest of the planet, and Arctic soils currently store twice as much carbon as the entire atmosphere-two facts that make understanding how Arctic soil microbial communities are responding to climate change particularly urgent. Greening of vegetation cover across the Arctic landscape is one of the most prominent climate-driven shifts in Arctic terrestrial ecology, with potentially profound effects on biogeochemical cycling by the soil microbiome. Here we use metaproteomics to document microbial metabolic functions that drive soil carbon and nutrient cycling processes in an Arctic tundra landscape. We identify functional roles among bacterial taxonomic groups that are largely stable across vegetation types, with certain functions strongly expressed by rhizosphere groups reflecting a community metabolic response to greening.

Chitin utilization by marine picocyanobacteria and the evolution of a planktonic lifestyle.

Marine picocyanobacteria Prochlorococcus and Synechococcus, the most abundant photosynthetic cells in the oceans, are generally thought to have a primarily single-celled and free-living lifestyle. However, while studying the ability of picocyanobacteria to supplement photosynthetic carbon fixation with the use of exogenous organic carbon, we found the widespread occurrence of genes for breaking down chitin, an abundant source of organic carbon that exists primarily as particles. We show that cells that encode a chitin degradation pathway display chitin degradation activity, attach to chitin particles, and show enhanced growth under low light conditions when exposed to chitosan, a partially deacetylated soluble form of chitin. Marine chitin is largely derived from arthropods, which underwent major diversifications 520 to 535 Mya, close to when marine picocyanobacteria are inferred to have appeared in the ocean. Phylogenetic analyses confirm that the chitin utilization trait was acquired at the root of marine picocyanobacteria. Together this leads us to postulate that attachment to chitin particles allowed benthic cyanobacteria to emulate their mat-based lifestyle in the water column, initiating their expansion into the open ocean, seeding the rise of modern marine ecosystems. Subsequently, transitioning to a constitutive planktonic life without chitin associations led to cellular and genomic streamlining along a major early branch within Prochlorococcus. Our work highlights how the emergence of associations between organisms from different trophic levels, and their coevolution, creates opportunities for colonizing new environments. In this view, the rise of ecological complexity and the expansion of the biosphere are deeply intertwined processes.

Isotopic Signatures of Carbon Transfer in a Proterozoic Analogue Microbial Mat.

Modern microbial mats are potential analogues for Proterozoic ecosystems, yet only a few studies have characterized mats under low-oxygen conditions that are relevant to Proterozoic environments. Here, we use protein-stable isotope fingerprinting (P-SIF) to determine the protein carbon isotope (δ13C) values of autotrophic, heterotrophic, and mixotrophic organisms in a benthic microbial mat from the low-oxygen Middle Island Sinkhole, Lake Huron, USA (MIS). We also measure the δ13C values of the sugar moieties of exopolysaccharides (EPS) within the mat to explore the relationships between cyanobacterial exudates and heterotrophic anabolic carbon uptake. Our results show that Cyanobacteria (autotrophs) are 13C-depleted, relative to sulfate-reducing bacteria (heterotrophs), and 13C-enriched, relative to sulfur oxidizing bacteria (autotrophs or mixotrophs). We also find that the pentose moieties of EPS are systematically enriched in 13C, relative to the hexose moieties of EPS. We hypothesize that these isotopic patterns reflect cyanobacterial metabolic pathways, particularly phosphoketolase, that are relatively more active in low-oxygen mat environments, rather than oxygenated mat environments. This results in isotopically more heterogeneous C sources in low-oxygen mats. While this might partially explain the isotopic variability observed in Proterozoic mat facies, further work is necessary to systematically characterize the isotopic fractionations that are associated with the synthesis of cyanobacterial exudates. IMPORTANCE The δ13C compositions of heterotrophic microorganisms are dictated by the δ13C compositions of their organic carbon sources. In both modern and ancient photosynthetic microbial mats, photosynthetic exudates are the most likely source of organic carbon for heterotrophs. We measured the δ13C values of autotrophic, heterotrophic, and mixotrophic bacteria as well as the δ13C value of the most abundant photosynthetic exudate (exopolysaccharide) in a modern analogue for a Proterozoic environment. Given these data, future studies will be better equipped to estimate the most likely carbon source for heterotrophs in both modern environments as well as in Proterozoic environments preserved in the rock record.

Complex regulation in a Comamonas platform for diverse aromatic carbon metabolism.

Critical to a sustainable energy future are microbial platforms that can process aromatic carbons from the largely untapped reservoir of lignin and plastic feedstocks. Comamonas species present promising bacterial candidates for such platforms because they can use a range of natural and xenobiotic aromatic compounds and often possess innate genetic constraints that avoid competition with sugars. However, the metabolic reactions of these species are underexplored, and the regulatory mechanisms are unknown. Here we identify multilevel regulation in the conversion of lignin-related natural aromatic compounds, 4-hydroxybenzoate and vanillate, and the plastics-related xenobiotic aromatic compound, terephthalate, in Comamonas testosteroni KF-1. Transcription-level regulation controls initial catabolism and cleavage, but metabolite-level thermodynamic regulation governs fluxes in central carbon metabolism. Quantitative 13C mapping of tricarboxylic acid cycle and cataplerotic reactions elucidates key carbon routing not evident from enzyme abundance changes. This scheme of transcriptional activation coupled with metabolic fine-tuning challenges outcome predictions during metabolic manipulations.

Tracking nitrogen allocation to proteome biosynthesis in a marine microbial community.

Microbial growth in many environments is limited by nitrogen availability, yet there is limited understanding of how complex communities compete for and allocate this resource. Here we develop a broadly applicable approach to track biosynthetic incorporation of 15N-labelled nitrogen substrates into microbial community proteomes, enabling quantification of protein turnover and N allocation to specific cellular functions in individual taxa. Application to oligotrophic ocean surface water identifies taxa-specific substrate preferences and a distinct subset of protein functions undergoing active biosynthesis. The cyanobacterium Prochlorococcus is the most effective competitor for acquisition of ammonium and urea and shifts its proteomic allocation of N over the day/night cycle. Our approach reveals that infrastructure and protein-turnover functions comprise substantial biosynthetic demand for N in Prochlorococcus and a range of other microbial taxa. The direct interrogation of the proteomic underpinnings of N limitation with 15N-tracking proteomics illuminates how nutrient stress differentially influences metabolism in co-existing microbes.

DsrC is involved in fermentative growth and interacts directly with the FlxABCD-HdrABC complex in Desulfovibrio vulgaris Hildenborough.

DsrC is a key protein in dissimilatory sulfur metabolism, where it works as co-substrate of the dissimilatory sulfite reductase DsrAB. DsrC has two conserved cysteines in a C-terminal arm that are converted to a trisulfide upon reduction of sulfite. In sulfate-reducing bacteria, DsrC is essential and previous works suggested additional functions beyond sulfite reduction. Here, we studied whether DsrC also plays a role during fermentative growth of Desulfovibrio vulgaris Hildenborough, by studying two strains where the functionality of DsrC is impaired by a lower level of expression (IPFG07) and additionally by the absence of one conserved Cys (IPFG09). Growth studies coupled with metabolite and proteomic analyses reveal that fermentation leads to lower levels of DsrC, but impairment of its function results in reduced growth by fermentation and a shift towards more fermentative metabolism during sulfate respiration. In both respiratory and fermentative conditions, there is increased abundance of the FlxABCD-HdrABC complex and Adh alcohol dehydrogenase in IPFG09 versus the wild type, which is reflected in higher production of ethanol. Pull-down experiments confirmed a direct interaction between DsrC and the FlxABCD-HdrABC complex, through the HdrB subunit. Dissimilatory sulfur metabolism, where sulfur compounds are used for energy generation, is a key process in the ecology of anoxic environments, and is more widespread among bacteria than previously believed. Two central proteins for this type of metabolism are DsrAB dissimilatory sulfite reductase and its co-substrate DsrC. Using physiological, proteomic and biochemical studies of Desulfovibrio vulgaris Hildenborough and mutants affected in DsrC functionality, we show that DsrC is also relevant for fermentative growth of this model organism and that it interacts directly with the soluble FlxABCD-HdrABC complex that links the NAD(H) pool with dissimilatory sulfite reduction.

Proteome Expression and Survival Strategies of a Proteorhodopsin-Containing Vibrio Strain under Carbon and Nitrogen Limitation.

Photoheterotrophy is a widespread mode of microbial metabolism, notably in the oligotrophic surface ocean, where microbes experience chronic nutrient limitation. One especially widespread form of photoheterotrophy is based on proteorhodopsin (PR), which uses light to generate proton motive force that can drive ATP synthesis, flagellar movement, or nutrient uptake. To clarify the physiological benefits conferred by PR under nutrient stress conditions, we quantified protein-level gene expression of Vibrio campbellii CAIM 519 under both carbon and nitrogen limitation and under both light and dark conditions. Using a novel membrane proteomics strategy, we determined that PR expression is higher under C limitation than N limitation but is not light regulated. Despite expression of PR photosystems, V. campbellii does not exhibit any growth or survival advantages in the light and only a few proteins show significant expression differences between light and dark conditions. While protein-level proteorhodopsin expression in V. campbellii is clearly responsive to nutrient limitation, photoheterotrophy does not appear to play a central role in the survival physiology of this organism under these nutrient stress conditions. C limitation and N limitation, however, result in very different survival responses: under N-limited conditions, viability declines, cultivability is lost rapidly, central carbon flux through the Entner-Doudoroff pathway is increased, and ammonium is assimilated via the GS-GOGAT pathway. In contrast, C limitation drives cell dwarfing with maintenance of viability, as well as utilization of the glyoxylate shunt, glutamate dehydrogenase and anaplerotic C fixation, and a stringent response mediated by the Pho regulon. IMPORTANCE Understanding the nutrient stress responses of proteorhodopsin-bearing microbes like Vibrio campbellii yields insights into microbial contributions to nutrient cycling, lifestyles of emerging pathogens in aquatic environments, and protein-level adaptations implemented during times of nutrient limitation. In addition to its broad taxonomic and geographic prevalence, the physiological role of PR is diverse, so we developed a novel proteomics strategy to quantify its expression at the protein level. We found that proteorhodopsin expression levels in this wild-type photoheterotroph under these experimental conditions, while higher under C than under N limitation, do not afford measurable light-driven growth or survival advantages. Additionally, this work links differential protein expression patterns between C- and N-limited cultures to divergent stationary-phase survival phenotypes.

Analogous Metabolic Decoupling in Pseudomonas putida and Comamonas testosteroni Implies Energetic Bypass to Facilitate Gluconeogenic Growth.

Gluconeogenic carbon metabolism is not well understood, especially within the context of flux partitioning between energy generation and biomass production, despite the importance of gluconeogenic carbon substrates in natural and engineered carbon processing. Here, using multiple omics approaches, we elucidate the metabolic mechanisms that facilitate gluconeogenic fast-growth phenotypes in Pseudomonas putida and Comamonas testosteroni, two Proteobacteria species with distinct metabolic networks. In contrast to the genetic constraint of C. testosteroni, which lacks the enzymes required for both sugar uptake and a complete oxidative pentose phosphate (PP) pathway, sugar metabolism in P. putida is known to generate surplus NADPH by relying on the oxidative PP pathway within its characteristic cyclic connection between the Entner-Doudoroff (ED) and Embden-Meyerhoff-Parnas (EMP) pathways. Remarkably, similar to the genome-based metabolic decoupling in C. testosteroni, our 13C-fluxomics reveals an inactive oxidative PP pathway and disconnected EMP and ED pathways in P. putida during gluconeogenic feeding, thus requiring transhydrogenase reactions to supply NADPH for anabolism in both species by leveraging the high tricarboxylic acid cycle flux during gluconeogenic growth. Furthermore, metabolomics and proteomics analyses of both species during gluconeogenic feeding, relative to glycolytic feeding, demonstrate a 5-fold depletion in phosphorylated metabolites and the absence of or up to a 17-fold decrease in proteins of the PP and ED pathways. Such metabolic remodeling, which is reportedly lacking in Escherichia coli exhibiting a gluconeogenic slow-growth phenotype, may serve to minimize futile carbon cycling while favoring the gluconeogenic metabolic regime in relevant proteobacterial species. IMPORTANCE Glycolytic metabolism of sugars is extensively studied in the Proteobacteria, but gluconeogenic carbon sources (e.g., organic acids, amino acids, aromatics) that feed into the tricarboxylic acid (TCA) cycle are widely reported to produce a fast-growth phenotype, particularly in species with biotechnological relevance. Much remains unknown about the importance of glycolysis-associated pathways in the metabolism of gluconeogenic carbon substrates. Here, we demonstrate that two distinct proteobacterial species, through genetic constraints or metabolic regulation at specific metabolic nodes, bypass the oxidative PP pathway during gluconeogenic growth and avoid unnecessary carbon fluxes by depleting protein investment into connected glycolysis pathways. Both species can leverage instead the high TCA cycle flux during gluconeogenic feeding to meet NADPH demand. Importantly, lack of a complete oxidative pentose phosphate pathway is a widespread metabolic trait in Proteobacteria with a gluconeogenic carbon preference, thus highlighting the important relevance of our findings toward elucidating the metabolic architecture in these bacteria.

Carbon substrate re-orders relative growth of a bacterium using Mo-, V-, or Fe-nitrogenase for nitrogen fixation.

Biological nitrogen fixation is catalyzed by the molybdenum (Mo), vanadium (V) and iron (Fe)-only nitrogenase metalloenzymes. Studies with purified enzymes have found that the 'alternative' V- and Fe-nitrogenases generally reduce N2 more slowly and produce more byproduct H2 than the Mo-nitrogenase, leading to an assumption that their usage results in slower growth. Here we show that, in the metabolically versatile photoheterotroph Rhodopseudomonas palustris, the type of carbon substrate influences the relative rates of diazotrophic growth based on different nitrogenase isoforms. The V-nitrogenase supports growth as fast as the Mo-nitrogenase on acetate but not on the more oxidized substrate succinate. Our data suggest that this is due to insufficient electron flux to the V-nitrogenase isoform on succinate compared with acetate. Despite slightly faster growth based on the V-nitrogenase on acetate, the wild-type strain uses exclusively the Mo-nitrogenase on both carbon substrates. Notably, the differences in H2 :N2 stoichiometry by alternative nitrogenases (~1.5 for V-nitrogenase, ~4-7 for Fe-nitrogenase) and Mo-nitrogenase (~1) measured here are lower than prior in vitro estimates. These results indicate that the metabolic costs of V-based nitrogen fixation could be less significant for growth than previously assumed, helping explain why alternative nitrogenase genes persist in diverse diazotroph lineages and are broadly distributed in the environment.

Metabolic and biogeochemical consequences of viral infection in aquatic ecosystems.

Ecosystems are controlled by 'bottom-up' (resources) and 'top-down' (predation) forces. Viral infection is now recognized as a ubiquitous top-down control of microbial growth across ecosystems but, at the same time, cell death by viral predation influences, and is influenced by, resource availability. In this Review, we discuss recent advances in understanding the biogeochemical impact of viruses, focusing on how metabolic reprogramming of host cells during lytic viral infection alters the flow of energy and nutrients in aquatic ecosystems. Our synthesis revealed several emerging themes. First, viral infection transforms host metabolism, in part through virus-encoded metabolic genes; the functions performed by these genes appear to alleviate energetic and biosynthetic bottlenecks to viral production. Second, viral infection depends on the physiological state of the host cell and on environmental conditions, which are challenging to replicate in the laboratory. Last, metabolic reprogramming of infected cells and viral lysis alter nutrient cycling and carbon export in the oceans, although the net impacts remain uncertain. This Review highlights the need for understanding viral infection dynamics in realistic physiological and environmental contexts to better predict their biogeochemical consequences.

Nitrogen sourcing during viral infection of marine cyanobacteria.

The building blocks of a virus derived from de novo biosynthesis during infection and/or catabolism of preexisting host cell biomass, and the relative contribution of these 2 sources has important consequences for understanding viral biogeochemistry. We determined the uptake of extracellular nitrogen (N) and its biosynthetic incorporation into both virus and host proteins using an isotope-labeling proteomics approach in a model marine cyanobacterium Synechococcus WH8102 infected by a lytic cyanophage S-SM1. By supplying dissolved N as 15N postinfection, we found that proteins in progeny phage particles were composed of up to 41% extracellularly derived N, while proteins of the infected host cell showed almost no isotope incorporation, demonstrating that de novo amino acid synthesis continues during infection and contributes specifically and substantially to phage replication. The source of N for phage protein synthesis shifted over the course of infection from mostly host derived in the early stages to more medium derived later on. We show that the photosystem II reaction center proteins D1 and D2, which are auxiliary metabolic genes (AMGs) in the S-SM1 genome, are made de novo during infection in an apparently light-dependent manner. We also identified a small set of host proteins that continue to be produced during infection; the majority are homologs of AMGs in S-SM1 or other viruses, suggesting selective continuation of host protein production during infection. The continued acquisition of nutrients by the infected cell and their utilization for phage replication are significant for both evolution and biogeochemical impact of viruses.

Proteomic and Isotopic Response of Desulfovibrio vulgaris to DsrC Perturbation.

Dissimilatory sulfate reduction is a microbial energy metabolism that can produce sulfur isotopic fractionations over a large range in magnitude. Calibrating sulfur isotopic fractionation in laboratory experiments allows for better interpretations of sulfur isotopes in modern sediments and ancient sedimentary rocks. The proteins involved in sulfate reduction are expressed in response to environmental conditions, and are collectively responsible for the net isotopic fractionation between sulfate and sulfide. We examined the role of DsrC, a key component of the sulfate reduction pathway, by comparing wildtype Desulfovibrio vulgaris DSM 644T to strain IPFG07, a mutant deficient in DsrC production. Both strains were cultivated in parallel chemostat reactors at identical turnover times and cell specific sulfate reduction rates. Under these conditions, sulfur isotopic fractionations between sulfate and sulfide of 17.3 ± 0.5‰ or 12.6 ± 0.5‰ were recorded for the wildtype or mutant, respectively. The enzymatic machinery that produced these different fractionations was revealed by quantitative proteomics. Results are consistent with a cellular-level response that throttled the supply of electrons and sulfur supply through the sulfate reduction pathway more in the mutant relative to the wildtype, independent of rate. We conclude that the smaller fractionation observed in the mutant strain is a consequence of sulfate reduction that proceeded at a rate that consumed a greater proportion of the strains overall capacity for sulfate reduction. These observations have consequences for models of sulfate reducer metabolism and how it yields different isotopic fractionations, notably, the role of DsrC in central energy metabolism.

Closely related viruses of the marine picoeukaryotic alga Ostreococcus lucimarinus exhibit different ecological strategies.

In marine ecosystems, viruses are major disrupters of the direct flow of carbon and nutrients to higher trophic levels. Although the genetic diversity of several eukaryotic phytoplankton virus groups has been characterized, their infection dynamics are less understood, such that the physiological and ecological implications of their diversity remain unclear. We compared genomes and infection phenotypes of the two most closely related cultured phycodnaviruses infecting the widespread picoprasinophyte Ostreococcus lucimarinus under standard- (1.3 divisions per day) and limited-light (0.41 divisions per day) nutrient replete conditions. OlV7 infection caused early arrest of the host cell cycle, coinciding with a significantly higher proportion of infected cells than OlV1-amended treatments, regardless of host growth rate. OlV7 treatments showed a near-50-fold increase of progeny virions at the higher host growth rate, contrasting with OlV1's 16-fold increase. However, production of OlV7 virions was more sensitive than OlV1 production to reduced host growth rate, suggesting fitness trade-offs between infection efficiency and resilience to host physiology. Moreover, although organic matter released from OlV1- and OlV7-infected hosts had broadly similar chemical composition, some distinct molecular signatures were observed. Collectively, these results suggest that current views on viral relatedness through marker and core gene analyses underplay operational divergence and consequences for host ecology.