Microbial transformations of organic carbon (OC) generate a large flux of CO2 into the atmosphere and influence the C balance of terrestrial and aquatic ecosystems. Yet, inherent heterogeneity in natural environments precludes direct quantification of multiple microbial C fluxes that underlie CO2 production. Here we used a continuous flow bioreactor coupled with a stable C isotope analyzer to determine the effects of temperature and C availability (cellobiose concentration) on C fluxes and 13C discrimination of a microbial population growing at steady-state in a homogeneous, well-mixed environment. We estimated C uptake affinity and C use efficiency (CUE) to characterize the physiological responses of microbes to changing environmental conditions. Temperature increased biomass-C specific respiration rate and C uptake affinity at lower C availability, but did not influence those parameters at higher C availability. CUE decreased non-linearly with increasing temperature. The non-linear, negative relationship between CUE and temperature was more pronounced under lower C availability than under relatively high C availability. We observed stable isotope fractionation between C substrate and microbial biomass C (7~12‰ depletion), and between microbial biomass and respired CO2 (4~10‰ depletion). Microbial discrimination against 13C-containing cellobiose during C uptake was influenced by temperature and C availability, while discrimination during respiration was only influenced by C availability. Shifts in C uptake affinity with temperature and C availability may have modified uptake-induced 13C fractionation. By stressing the importance of C availability on temperature responses of microbial C fluxes, C uptake affinity, CUE, and isotopic fractionation, this study contributes to a fundamental understanding of C flow through microbes. This will help guide parameterization of microbial responses to varying temperature and C availability within Earth-system models.

Iron (Fe) oxides can stabilize organic carbon (OC) through adsorption and co-precipitation, while microbial Fe reduction can disrupt Fe-bound OC (Fe-OC) and further increase OC mineralization. The net effects of OC preservation and mineralization mediated by Fe oxides are still unclear, especially for old carbon (formed from plant litters over millions of years) and crystalline Fe oxides. Accelerating the recovery of wetland carbon sinks is critical for mitigating climate change and achieving carbon neutrality. Quantifying the net effect of Fe-mediated OC mineralization and preservation is vital for understanding the role of crystalline Fe oxides in carbon cycling and promoting the recovery of soil carbon sinks. Here, we explored the OC balances mediated by hematite (Hem) and lignite addition (Lig) to freshwater wetland (FW, rich in C and Fe) and saline-alkaline wetland (SW, poor in C and Fe) soil slurries, incubated under anaerobic conditions. Results showed that Lig caused net OC accumulation (FW: 5.9 ± 3.6 mg g-1; SW: 8.3 ± 3.2 mg g-1), while Hem caused dramatic OC loss, particularly in the FW soils. Hem inhibited microbial Fe(III) reduction by decreasing the relative abundance of Fe respiration reducers, while substantially enhancing OC mineralization through the shift in the microbial community structure of FW soils. Lig resulted in carbon emission, but its contribution to preservation by the formation of Fe-OC was far higher than that which caused OC loss. We concluded that crystalline Fe oxide addition solely favored the increase of OC mineralization by adjusting the microbial community structure, while old carbon enriched with an aromatic and alkyl promoted Fe-OC formation and further increased OC persistence. Our findings could be employed for wetland restoration, particularly for the recovery of soil carbon sinks.

Using a pulse chase (13)CO2 plant labeling experiment we compared the flow of plant carbon into macromolecular fractions of rhizosphere soil microorganisms. Time dependent (13)C dilution patterns in microbial cellular fractions were used to calculate their turnover time. The turnover times of microbial biomolecules were found to vary: microbial RNA (19 h) and DNA (30 h) turned over fastest followed by chloroform fumigation extraction-derived soluble cell lysis products (14 days), while phospholipid fatty acids (PLFAs) had the slowest turnover (42 days). PLFA/NLFA (13)C analyses suggest that both mutualistic arbuscular mycorrhizal and saprophytic fungi are dominant in initial plant carbon uptake. In contrast, high initial (13)C enrichment in RNA hints at bacterial importance in initial C uptake due to the dominance of bacterial derived RNA in total extracts of soil RNA. To explain this discrepancy, we observed low renewal rate of bacterial lipids, which may therefore bias lipid fatty acid based interpretations of the role of bacteria in soil microbial food webs. Based on our findings, we question current assumptions regarding plant-microbe carbon flux and suggest that the rhizosphere bacterial contribution to plant assimilate uptake could be higher. This highlights the need for more detailed quantitative investigations with nucleic acid biomarkers to further validate these findings.

Oceanic crust is a massive potential habitat for microbial life on Earth, yet our understanding of this ecosystem is limited due to difficulty in access. In particular, measurements of rates of microbial activity are sparse. We used stable carbon isotope incubations of crustal samples, coupled with functional gene analyses, to examine the potential for carbon fixation on oceanic crust. Both seafloor-exposed and subseafloor basalts were recovered from different mid-ocean ridge and hot spot environments (i.e., the Juan de Fuca Ridge, the Mid-Atlantic Ridge, and the Loihi Seamount) and incubated with (13)C-labeled bicarbonate. Seafloor-exposed basalts revealed incorporation of (13)C-label into organic matter over time, though the degree of incorporation was heterogeneous. The incorporation of (13)C into biomass was inconclusive in subseafloor basalts. Translating these measurements into potential rates of carbon fixation indicated that 0.1-10 nmol C g(-1) rock d(-1) could be fixed by seafloor-exposed rocks. When scaled to the global production of oceanic crust, this suggests carbon fixation rates of 10(9)-10(12) g C year(-1), which matches earlier predictions based on thermodynamic calculations. Functional gene analyses indicate that the Calvin cycle is likely the dominant biochemical mechanism for carbon fixation in basalt-hosted biofilms, although the reductive acetyl-CoA pathway and reverse TCA cycle likely play some role in net carbon fixation. These results provide empirical evidence for autotrophy in oceanic crust, suggesting that basalt-hosted autotrophy could be a significant contributor of organic matter in this remote and vast environment.

Nitrogen based eutrophication of ecosystems is a global problem that gains momentum through a growing global population. The water quality of nitrate or ammonium contaminated rivers and streams cannot always be amended in centralized waste water treatment plants. Field denitrification plants were suggested as a solution for a decentralized reduction of nitrate to dinitrogen. Here, stable and cheap organic carbon sources serve as carbon and electron source for a microbial community. Still, our knowledge on the impact of these organic carbon sources on the development and diversity of these cultures is sparse. Moreover, the stability of these denitrification plants at different nitrate loading rates especially in the higher concentration regime were not tested so far. In this study, we compare the fate of carbon and nitrogen as well as the microbial community of wood pellet (WP) (pressed sawdust), wheat straw, and wood chips (WC) based laboratory denitrification reactors. Our study reveals that the diversity and composition of the community is strongly dependent on the carbon source. The diversity decreased in the order WC, wheat straw, and WPs. The three reactor types were characterized by different nitrate reduction kinetics and were affected differently by high nitrate loading rates. While the nitrate reduction kinetics were negatively influenced by higher nitrate doses in the wheat straw reactors, WPs as carbon source sustained the opposite trend and WC lead to an overall slower but concentration independent nitrate reduction rate. Counterintuitively, the concentration of soluble organic carbon was highest in the WP reactors but methane emission was not detectable. This is corroborated by the microbial diversity data in which methanogenic species were highly underrepresented compared to the other two reactor types. In contrary, the methane emissions in the wheat straw and WC reactors were comparable to each other.

The unique symbiosis between a vertebrate salamander, Ambystoma maculatum, and unicellular green alga, Oophila amblystomatis, involves multiple modes of interaction. These include an ectosymbiotic interaction where the alga colonizes the egg capsule, and an intracellular interaction where the alga enters tissues and cells of the salamander. One common interaction in mutualist photosymbioses is the transfer of photosynthate from the algal symbiont to the host animal. In the A. maculatum-O. amblystomatis interaction, there is conflicting evidence regarding whether the algae in the egg capsule transfer chemical energy captured during photosynthesis to the developing salamander embryo. In experiments where we took care to separate the carbon fixation contributions of the salamander embryo and algal symbionts, we show that inorganic carbon fixed by A. maculatum embryos reaches 2% of the inorganic carbon fixed by O. amblystomatis algae within an egg capsule after 2 h in the light. After 2 h in the dark, inorganic carbon fixed by A. maculatum embryos is 800% of the carbon fixed by O. amblystomatis algae within an egg capsule. Using photosynthesis inhibitors, we show that A. maculatum embryos and O. amblystomatis algae compete for available inorganic carbon within the egg capsule environment. Our results confirm earlier studies suggesting a role of heterotrophic carbon fixation during vertebrate embryonic development. Our results also show that the considerable capacity of developing A. maculatum embryos for inorganic carbon fixation precludes our ability to distinguish any minor role of photosynthetically transferred carbon from algal symbionts to host salamanders using bicarbonate introduced to the egg system as a marker.

The European Climate Law recently codified the goal for European climate neutrality by 2050, highlighting the need for sustainable farming practices within a robust and transparent carbon dioxide equivalent (CO2e) accounting system. In the present study, a series of equations were proposed for the estimation of CO2e emissions from corn silage fermentation. Systematic review of previous meta-analyses of corn silage fermentation identified the mean and standard deviation statistics for key model inputs of acetic acid, ethanol, lactic acid, ammonia, and volatile-corrected dry matter loss. Estimates of CO2e emissions were determined for a mock dataset comprising 1,000 iterations of randomly-generated values for each metric in accordance with mean and variance statistics of the source data. Estimates for CO2e emissions of corn silage based on meta-analysis review of laboratory experiments were 1.9 ± 5.6% (GWP20) and 0.2 ± 5.5% (GWP100) of silage dry matter. Furthermore, model results demonstrated a precedent for CO2 recycling by silage microorganisms, which was supported by genome annotation of strains belonging to common silage species. Linear model equations for GWP20 and GWP100 with inputs and outputs in mg kg-1 silage dry matter were developed, where inputs are acetic acid (A), ethanol (E), lactic acid (L), and volatile corrected dry matter loss (DV). Linear equations are (for GWP20; Eq. 11): GWP 20 = - 3626.1 - 0.04343 A + 0.8011 E - 0.03173 L + 1.46573 D V and for GWP100; Eq. 12: GWP 100 = - 8526.10 - 0.22403 A - 0.11963 E - 0.03173 L + 1.46573 D V . .

Legume crops in rice cultivation are typically rotated and incorporated into the soil as green manure to improve soil fertility. Biochar has recently been co-incorporated with green manure to simultaneously stimulate soil organic carbon (SOC) mineralization and increase carbon (C) sequestration. However, few studies examine the effects of the co-incorporation of biochar and green manure on C cycling and the underlying microbial mechanisms in paddy fields. In this study, the effects of the co-incorporation of green manure and biochar on C mineralization, dissolved organic carbon (DOC) characteristics, and microbial community structures were investigated. A pot study was conducted with three treatments: inorganic NPK (NPK), inorganic NPK + green manure (GM), and inorganic NPK + green manure + biochar (GMC). Organic amendments significantly increased cumulative C mineralization, with amounts in the order GMC (3,434 mg·kg-1) > GM (2,934 mg·kg-1) > NPK (2,592 mg·kg-1). Fertilizer treatments had similar effects on DOC concentrations, with amounts in the order GMC (279 mg·kg-1) > GM (255 mg·kg-1) > NPK (193 mg·kg-1). According to fluorescence spectra, the highest microbial humic acid-like fraction and biological index were also in GMC. Co-incorporation of green manure and biochar shifted the composition of bacterial and fungal communities but more importantly, increased fungal network complexity and decreased bacterial network complexity. The increase in fungal network complexity with the increase in DOC concentrations and microbially derived components was the dominant factor in promoting C mineralization. Overall, this study reveals the underlying biochemical mechanism, the interaction between DOC and fungal network of C cycling in paddy soil under the co-incorporation of green manure and biochar management, and provides fundamental knowledge for exploring effective approaches to improve soil fertility and health in the future.

Despite several lines of observational evidence, there is a lack of consensus on whether higher fungal:bacterial (F:B) ratios directly cause higher soil carbon (C) storage. We employed RNA sequencing, protein profiling and isotope tracer techniques to evaluate whether differing F:B ratios are associated with differences in C storage. A mesocosm (13)C labeled foliar litter decomposition experiment was performed in two soils that were similar in their physico-chemical properties but differed in microbial community structure, specifically their F:B ratio (determined by PLFA analyses, RNA sequencing and protein profiling; all three corroborating each other). Following litter addition, we observed a consistent increase in abundance of fungal phyla; and greater increases in the fungal dominated soil; implicating the role of fungi in litter decomposition. Litter derived (13)C in respired CO2 was consistently lower, and residual (13)C in bulk SOM was higher in high F:B soil demonstrating greater C storage potential in the F:B dominated soil. We conclude that in this soil system, the increased abundance of fungi in both soils and the altered C cycling patterns in the F:B dominated soils highlight the significant role of fungi in litter decomposition and indicate that F:B ratios are linked to higher C storage potential.

Anaerobic Ni-containing carbon-monoxide dehydrogenases (Ni-CODHs) catalyze the reversible conversion between carbon monoxide and carbon dioxide as multi-enzyme complexes responsible for carbon fixation and energy conservation in anaerobic microbes. However, few biochemically characterized model enzymes exist, with most Ni-CODHs remaining functionally unknown. Here, we performed phylogenetic and structure-based Ni-CODH classification using an expanded dataset comprised of 1942 non-redundant Ni-CODHs from 1375 Ni-CODH-encoding genomes across 36 phyla. Ni-CODHs were divided into seven clades, including a novel clade. Further classification into 24 structural groups based on sequence analysis combined with structural prediction revealed diverse structural motifs for metal cluster formation and catalysis, including novel structural motifs potentially capable of forming metal clusters or binding metal ions, indicating Ni-CODH diversity and plasticity. Phylogenetic analysis illustrated that the metal clusters responsible for intermolecular electron transfer were drastically altered during evolution. Additionally, we identified novel putative Ni-CODH-associated proteins from genomic contexts other than the Wood-Ljungdahl pathway and energy converting hydrogenase system proteins. Network analysis among the structural groups of Ni-CODHs, their associated proteins and taxonomies revealed previously unrecognized gene clusters for Ni-CODHs, including uncharacterized structural groups with putative metal transporters, oxidoreductases, or transcription factors. These results suggested diversification of Ni-CODH structures adapting to their associated proteins across microbial genomes.

An anaerobic thermophilic strain (strain PCO) was isolated from a syngas-converting enrichment culture. Syngas components cannot be used by strain PCO, but the new strain is very tolerant to carbon monoxide (pCO = 1.7 × 10(5) Pa, 100% CO). 16S rRNA gene analysis and DNA-DNA hybridization revealed that strain PCO is a strain of Thermoanaerobacter thermohydrosulfuricus. The physiology of strain PCO and other Thermoanaerobacter species was compared, focusing on their tolerance to carbon monoxide. T. thermohydrosulfuricus, T. brockii subsp. finnii, T. pseudethanolicus, and T. wiegelii were exposed to increased CO concentrations in the headspace, while growth, glucose consumption and product formation were monitored. Remarkably, glucose conversion rates by Thermoanaerobacter species were not affected by CO. All the tested strains fermented glucose to mainly lactate, ethanol, acetate, and hydrogen, but final product concentrations differed. In the presence of CO, ethanol production was generally less affected, but H2 production decreased with increasing CO partial pressure. This study highlights the CO resistance of Thermoanaerobacter species.

Organisms of the candidate phylum Saccharibacteria have frequently been detected as active members of hydrocarbon degrading communities, yet their actual role in hydrocarbon degradation remained unclear. Here, we analyzed three enrichment cultures of hydrocarbon-amended groundwater samples using genome-resolved metagenomics to unravel the metabolic potential of indigenous Saccharibacteria. Community profiling based on ribosomal proteins revealed high variation in the enrichment cultures suggesting little reproducibility although identical cultivation conditions were applied. Only 17.5 and 12.5% of the community members were shared between the three enrichment cultures based on ribosomal protein clustering and read mapping of reconstructed genomes, respectively. In one enrichment, two Saccharibacteria strains dominated the community with 16.6% in relative abundance and we were able to recover near-complete genomes for each of them. A detailed analysis of their limited metabolism revealed the capacity for peptide degradation, lactate fermentation from various hexoses, and suggests a scavenging lifestyle with external retrieval of molecular building blocks. In contrast to previous studies suggesting that Saccharibacteria are directly involved in hydrocarbon degradation, our analyses provide evidence that these organisms can be highly abundant scavengers acting rather as organic carbon sinks than hydrocarbon degraders in these communities.

Mass blooms of purple sulfur bacteria growing seasonally on green stranded macroalgae have a major impact on the microbial composition and functionality of intertidal mats. To explore the active anoxygenic phototrophic community in purple bacterial mats from the Roscoff Aber Bay (Brittany, France), we conducted a combined approach including molecular and high-resolution secondary ion mass spectrometry (NanoSIMS) analyses. To investigate the dynamics of carbon and nitrogen assimilation activities, NanoSIMS was coupled with a stable isotope probing (SIP) experiment and a compound specific isotope analysis (CSIA) of fatty acid methyl ester (FAME). Sediment samples were incubated with 13C- and/or 15N-labeled acetate, pyruvate, bicarbonate and ammonium. NanoSIMS analysis of 13C - and 15N -incubated samples showed elevated incorporations of 13C - and 15N in the light and of 13C -acetate in the dark into dense populations of spherical cells that unambiguously dominated the mats. These results confirmed CSIA data that ranked vaccenic acid, an unambiguous marker of purple sulfur bacteria, as the most strongly enriched in the light after 13C -acetate amendment and indicated that acetate uptake, the most active in the mat, was not light-dependent. Analysis of DNA- and cDNA-derived pufM gene sequences revealed that Thiohalocapsa-related clones dominated both libraries and were the most photosynthetically active members of the mat samples. This study provides novel insights into the contribution of purple sulfur bacteria to the carbon cycle during their seasonal developments at the sediment surface in the intertidal zone.

The metallic low and intermediate level radioactive waste generally consists of carbon steel and stainless steels. The corrosion rate of carbon steel in deep groundwater is typically low, unless the water is very acidic or microbial activity in the environment is high. Therefore, the assessment of microbially induced corrosion of carbon steel in deep bedrock environment has become important for evaluating the safety of disposal of radioactive waste. Here we studied the corrosion inducing ability of indigenous microbial community from a deep bedrock aquifer. Carbon steel coupons were exposed to anoxic groundwater from repository site 100 m depth (Olkiluoto, Finland) for periods of 3 and 8 months. The experiments were conducted at both in situ temperature and room temperature to investigate the response of microbial population to elevated temperature. Our results demonstrate that microorganisms from the deep bedrock aquifer benefit from carbon steel introduced to the nutrient poor anoxic deep groundwater environment. In the groundwater incubated with carbon steel the planktonic microbial community was more diverse and 100-fold more abundant compared to the environment without carbon steel. The betaproteobacteria were the most dominant bacterial class in all samples where carbon steel was present, whereas in groundwater incubated without carbon steel the microbial community had clearly less diversity. Microorganisms induced pitting corrosion and were found to cluster inside the corrosion pits. Temperature had an effect on the species composition of microbial community and also affected the corrosion deposits layer formed on the surface of carbon steel.

Carbon monoxide (CO), well known as a toxic gas, is increasingly recognized as a key metabolite and signaling molecule. Microbial utilization of CO is quite common, evidenced by the rapid escalation in description of new species of CO-utilizing bacteria and archaea. Carbon monoxide dehydrogenase (CODH), the protein complex that enables anaerobic CO-utilization, has been well-characterized from an increasing number of microorganisms, however the regulation of multiple CO-related gene clusters in single isolates remains unexplored. Many species are extraordinarily resistant to high CO concentrations, thriving under pure CO at more than one atmosphere. We hypothesized that, in strains that can grow exclusively on CO, both carbon acquisition via the CODH/acetyl CoA synthase complex and energy conservation via a CODH-linked hydrogenase must be differentially regulated in response to the availability of CO. The CO-sensing transcriptional activator, CooA is present in most CO-oxidizing bacteria. Here we present a genomic and phylogenetic survey of CODH operons and cooA genes found in CooA-containing bacteria. Two distinct groups of CooA homologs were found: one clade (CooA-1) is found in the majority of CooA-containing bacteria, whereas the other clade (CooA-2) is found only in genomes that encode multiple CODH clusters, suggesting that the CooA-2 might be important for cross-regulation of competing CODH operons. Recombinant CooA-1 and CooA-2 regulators from the prototypical CO-utilizing bacterium Carboxydothermus hydrogenoformans were purified, and promoter binding analyses revealed that CooA-1 specifically regulates the hydrogenase-linked CODH, whereas CooA-2 is able to regulate both the hydrogenase-linked CODH and the CODH/ACS operons. These studies point to the ability of dual CooA homologs to partition CO into divergent CO-utilizing pathways resulting in efficient consumption of a single limiting growth substrate available across a wide range of concentrations.

Marine diazotrophs are a diverse group with key roles in biogeochemical fluxes linked to primary productivity. The unicellular, diazotrophic cyanobacterium Cyanothece is widely found in coastal, subtropical oceans. We analyze the consequences of diazotrophy on growth efficiency, compared to NO3 --supported growth in Cyanothece, to understand how cells cope with N2-fixation when they also have to face carbon limitation, which may transiently affect populations in coastal environments or during blooms of phytoplankton communities. When grown in obligate diazotrophy, cells face the double burden of a more ATP-demanding N-acquisition mode and additional metabolic losses imposed by the transient storage of reducing potential as carbohydrate, compared to a hypothetical N2 assimilation directly driven by photosynthetic electron transport. Further, this energetic burden imposed by N2-fixation could not be alleviated, despite the high irradiance level within the cultures, because photosynthesis was limited by the availability of dissolved inorganic carbon (DIC), and possibly by a constrained capacity for carbon storage. DIC limitation exacerbates the costs on growth imposed by nitrogen fixation. Therefore, the competitive efficiency of diazotrophs could be hindered in areas with insufficient renewal of dissolved gases and/or with intense phytoplankton biomass that both decrease available light energy and draw the DIC level down.

Carbon monoxide (CO) is commonly known as a toxic gas, yet both cultivation studies and emerging genome sequences of bacteria and archaea establish that CO is a widely utilized microbial growth substrate. In this study, we determined the prevalence of anaerobic carbon monoxide dehydrogenases ([Ni,Fe]-CODHs) in currently available genomic sequence databases. Currently, 185 out of 2887, or 6% of sequenced bacterial and archaeal genomes possess at least one gene encoding [Ni,Fe]-CODH, the key enzyme for anaerobic CO utilization. Many genomes encode multiple copies of [Ni,Fe]-CODH genes whose functions and regulation are correlated with their associated gene clusters. The phylogenetic analysis of this extended protein family revealed six distinct clades; many clades consisted of [Ni,Fe]-CODHs that were encoded by microbes from disparate phylogenetic lineages, based on 16S rRNA sequences, and widely ranging physiology. To more clearly define if the branching patterns observed in the [Ni,Fe]-CODH trees are due to functional conservation vs. evolutionary lineage, the genomic context of the [Ni,Fe]-CODH gene clusters was examined, and superimposed on the phylogenetic trees. On the whole, there was a correlation between genomic contexts and the tree topology, but several functionally similar [Ni,Fe]-CODHs were found in different clades. In addition, some distantly related organisms have similar [Ni,Fe]-CODH genes. Thermosinus carboxydivorans was used to observe horizontal gene transfer (HGT) of [Ni,Fe]-CODH gene clusters by applying Kullback-Leibler divergence analysis methods. Divergent tetranucleotide frequency and codon usage showed that the gene cluster of T. carboxydivorans that encodes a [Ni,Fe]-CODH and an energy-converting hydrogenase is dissimilar to its whole genome but is similar to the genome of the phylogenetically distant Firmicute, Carboxydothermus hydrogenoformans. These results imply that T carboxydivorans acquired this gene cluster via HGT from a relative of C. hydrogenoformans.

Carbon monoxide (CO)-metabolizing Clostridium spp. were enriched from the biomass of a butanol-producing reactor. After six successive biomass transfers, ethanol production reached as much as 11.8 g/L with minor accumulation of acetic acid, under intermittent gas feeding conditions and over a wide pH range of 6.45-4.95. The molar ratio of ethanol to acetic acid exceeded 1.7 after the lag phase of 11 days and reached its highest value of 8.6 during the fermentation process after 25 days. Although butanol production was not significantly enhanced in the enrichment, the biomass was able to convert exogenous butyric acid (3.2 g/L) into butanol with nearly 100% conversion efficiency using CO as reducing power. This suggested that inhibition of butanol production from CO was caused by the lack of natural butyric acid production, expectedly induced by unsuitable pH values due to initial acidification resulting from the acetic acid production. The enriched Clostridium population also converted glucose to formic, acetic, propionic, and butyric acids in batch tests with daily pH adjustment to pH 6.0. The Clostridium genus was enriched with its relative abundance significantly increasing from 7% in the inoculum to 94% after five successive enrichment steps. Unidentified Clostridium species showed a very high relative abundance, reaching 73% of the Clostridium genus in the enriched sludge (6th transfer).

There is widespread interest in how geochemistry affects the genomic makeup of microbial communities, but the possible impacts of oxidation-reduction (redox) conditions on the chemical composition of biomacromolecules remain largely unexplored. Here we document systematic changes in the carbon oxidation state, a metric derived from the chemical formulas of biomacromolecular sequences, using published metagenomic and metatranscriptomic datasets from 18 studies representing different marine and terrestrial environments. We find that the carbon oxidation states of DNA, as well as proteins inferred from coding sequences, follow geochemical redox gradients associated with mixing and cooling of hot spring fluids in Yellowstone National Park (USA) and submarine hydrothermal fluids. Thermodynamic calculations provide independent predictions for the environmental shaping of the gene and protein composition of microbial communities in these systems. On the other hand, the carbon oxidation state of DNA is negatively correlated with oxygen concentration in marine oxygen minimum zones. In this case, a thermodynamic model is not viable, but the low carbon oxidation state of DNA near the ocean surface reflects a low GC content, which can be attributed to genome reduction in organisms adapted to low-nutrient conditions. We also present evidence for a depth-dependent increase of oxidation state at the species level, which might be associated with alteration of DNA through horizontal gene transfer and/or selective degradation of relatively reduced (AT-rich) extracellular DNA by heterotrophic bacteria. Sediments exhibit even more complex behavior, where carbon oxidation state minimizes near the sulfate-methane transition zone and rises again at depth; markedly higher oxidation states are also associated with older freshwater-dominated sediments in the Baltic Sea that are enriched in iron oxides and have low organic carbon. This geobiochemical study of carbon oxidation state reveals a new aspect of environmental information in metagenomic sequences, and provides a reference frame for future studies that may use ancient DNA sequences as a paleoredox indicator.

Fructooligosaccharides (FOSs) metabolism in Lactobacillus plantarum is controlled by two gene clusters, and the global regulator catabolite control protein A (CcpA) may be involved in the regulation. To understand the mechanism, this study focused on the regulation relationships of CcpA toward target genes and the binding effects on the catabolite responsive element (cre). First, reverse transcription-PCR analysis of the transcriptional organization of the FOS-related gene clusters showed that they were organized in three independent polycistronic units. Diauxic growth, hierarchical utilization of carbohydrates and repression of FOS-related genes were observed in cultures containing FOS and glucose, suggesting carbon catabolite repression (CCR) control in FOS utilization. Knockout of ccpA gene eliminated these phenomena, indicating the principal role of this gene in CCR of FOS metabolism. Furthermore, six potential cre sites for CcpA binding were predicted in the regions of putative promoters of the two clusters. Direct binding was confirmed by electrophoretic mobility shift assays in vitro and chromatin immunoprecipitation in vivo. The results of the above studies suggest that CcpA is a vital regulator of FOS metabolism in L. plantarum and that CcpA-dependent CCR regulates FOS metabolism through the direct binding of CcpA toward the cre sites in the promoter regions of FOS-related clusters.