Sulfate-reducing Bacterium Desulfovibrio Vulgaris Research Articles

Antibiofilm assay for antimicrobial peptides combating the sulfate-reducing bacteria Desulfovibrio vulgaris.

In medical, environmental, and industrial processes, the accumulation of bacteria in biofilms can disrupt many processes. Antimicrobial peptides (AMPs) are receiving increasing attention in the development of new substances to avoid or reduce biofilm formation. There is a lack of parallel testing of the effect against biofilms in this area, as well as in the testing of other antibiofilm agents. In this paper, a high-throughput screening was developed for the analysis of the antibiofilm activity of AMPs, differentiated into inhibition and removal of a biofilm. The sulfate-reducing bacterium Desulfovibrio vulgaris was used as a model organism.D. vulgaris represents an undesirable bacterium, which is considered one of the major triggers of microbiologically influenced corrosion. The application of a 96-well plate and steel rivets as a growth surface realizes real-life conditions and at the same time establishes a flexible, simple, fast, and cost-effective assay. All peptides tested in this study demonstrated antibiofilm activity, although these peptides should be individually selected depending on the addressed aim. For biofilm inhibition, the peptide DASamP1 is the most suitable, with a sustained effect for up to 21 days. The preferred peptides for biofilm removal are S6L3-33, in regard to bacteria reduction, and Bactenecin, regarding total biomass reduction.

Microbiologically influenced corrosion of CoCrFeMnNi high entropy alloy by sulfate-reducing bacterium Desulfovibrio vulgaris

Sulfate-reducing bacterium Desulfovibrio vulgaris promoted the microbiologically influenced corrosion (MIC) of CoCrFeMnNi high entropy alloy (HEA) and led to pitting corrosion. D. vulgaris caused the formation of metal sulfides and thinning of passive film. The corrosion resistance decreased, and pit depths increased when the carbon source reduced to 40%. The relative expression of hydrogenase genes hydA and hydB in the biofilms under 40% carbon source significantly increased, indicating that the indirect electron transfer mediated by 2H+/H2 might play an important role in promoting the HEA MIC by D. vulgaris under starvation.

Large-scale genetic characterization of the model sulfate-reducing bacterium, Desulfovibrio vulgaris Hildenborough.

Sulfate-reducing bacteria (SRB) are obligate anaerobes that can couple their growth to the reduction of sulfate. Despite the importance of SRB to global nutrient cycles and their damage to the petroleum industry, our molecular understanding of their physiology remains limited. To systematically provide new insights into SRB biology, we generated a randomly barcoded transposon mutant library in the model SRB Desulfovibrio vulgaris Hildenborough (DvH) and used this genome-wide resource to assay the importance of its genes under a range of metabolic and stress conditions. In addition to defining the essential gene set of DvH, we identified a conditional phenotype for 1,137 non-essential genes. Through examination of these conditional phenotypes, we were able to make a number of novel insights into our molecular understanding of DvH, including how this bacterium synthesizes vitamins. For example, we identified DVU0867 as an atypical L-aspartate decarboxylase required for the synthesis of pantothenic acid, provided the first experimental evidence that biotin synthesis in DvH occurs via a specialized acyl carrier protein and without methyl esters, and demonstrated that the uncharacterized dehydrogenase DVU0826:DVU0827 is necessary for the synthesis of pyridoxal phosphate. In addition, we used the mutant fitness data to identify genes involved in the assimilation of diverse nitrogen sources and gained insights into the mechanism of inhibition of chlorate and molybdate. Our large-scale fitness dataset and RB-TnSeq mutant library are community-wide resources that can be used to generate further testable hypotheses into the gene functions of this environmentally and industrially important group of bacteria.

Enhanced pitting corrosion resistance of CoCrFeMnNi high entropy alloy in the presence of Desulfovibrio vulgaris via nitrogen doping

• Desulfovibrio vulgaris accelerates pitting corrosion of HEA without nitrogen. • Nitrogen doping significantly inhibits pitting corrosion induced by D. vulgaris . • Nitrogen strengthens passive film by increasing corrosion resistant species. • Nitrogen doped HEAs exhibits less defects in passive films. Corrosion-resistant high nitrogen high entropy alloys (HEAs) were manufactured by pressurized metallurgy. This work revealed the inhibitory effect of nitrogen on pitting corrosion of HEAs caused by sulfate reducing bacterium Desulfovibrio vulgaris . Results indicated that HEA-0N was susceptible to pitting corrosion and sulfidation under attack of D. vulgaris , whereas the addition of nitrogen significantly decreased the pitting sensitivity. Pitting potential of HEA-0.52N and HEA-1.23N increased by 133% and 171%, respectively compared to HEA-0N in the presence of SRB. X-ray photoelectron spectroscopy results unveiled that nitrogen enriched in passive film and strengthened it by increasing fraction of Cr 2 O 3 and Fe 3+ ox . Surface of the nitrogen-alloyed HEAs exhibited less defective passive films as revealed by Mott-Schottky results. Nitrogen doping provides a novel insight into the design of microbial corrosion resistant HEA.

How a Formate Dehydrogenase Responds to Oxygen: Unexpected O2 Insensitivity of an Enzyme Harboring Tungstopterin, Selenocysteine, and [4Fe–4S] Clusters

The reversible two-electron interconversion of formate and CO2 is catalyzed by both nonmetallo- and metallo-formate dehydrogenases (FDHs). The latter group comprises molybdenum- or tungsten-containing enzymes with the metal coordinated by two equivalents of a pyranopterin cofactor, a cysteinyl or selenocysteinyl (Sec) ligand supplied by the polypeptide, and a catalytically essential terminal sulfido ligand. In addition, these biocatalysts incorporate one or more [4Fe–4S] clusters for facilitating long-distance electron transfer. However, an interesting dichotomy arises when attempting to understand how the metallo-FDHs react with O2. Whereas existing scholarship portrays these enzymes as being unable to perform in air due to extreme O2 lability of their metal centers, studies dating as far back as the 1930s emphasize that some of these systems exhibit formate oxidase (FOX) activity, coupling formate oxidation to O2 reduction. Therefore, to reconcile these conflicting views, we explored context-dependent functional linkages between metallo-FDHs and their cognate electron acceptors within the same organism vis-à-vis catalysis under atmospheric O2. Here, we report the discovery and characterization of an O2-insensitive FDH2 from the sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough (DvH) that ligates tungsten, Sec, and four [4Fe–4S] clusters. By advancing a robust expression platform for its recombinant production, we eliminate both the requirement of nitrate or azide during purification and reductive activation with thiols and/or formate prior to catalysis. Because the distinctive spectral signatures of formate-reduced DvH-FDH2 remain invariant under anaerobic and aerobic conditions, we benchmarked the enzyme activity in air, identifying CO2 as the catalytic product. Full reaction progress curve analysis discloses a high catalytic efficiency when probed with a high-potential artificial electron acceptor. Furthermore, we show that DvH-FDH2 enables near-stoichiometric hydrogen peroxide production without superoxide release to achieve O2 insensitivity. Notably, simultaneous electron transfer to cytochrome c and O2 reveals that metal-based electron bifurcation is operational in this system. Taken together, our work proves the co-occurrence of redox bifurcated FDH and FOX activities within a metalloenzyme scaffold. These findings set the stage for uncovering previously unknown O2-insensitive flavin-based electron bifurcation mechanisms, as well as for developing authentic formate/air biofuel cells, engineering O2-stable FDHs and biohybrid metallocatalysts, and discerning formate bioenergetics of gut microbiota.

Influence of Sulfate Reduction on Arsenic Migration and Transformation in Groundwater Environment

The sulfate-reducing bacteria-mediated reduction process is considered to be an important mechanism affecting arsenic migration and transformation in anaerobic environments. To investigate the effect of sulfate-reducing bacteria in a high-arsenic aquifer on arsenic migration and transformation, the typical sulfate-reducing bacteria Desulfovibrio vulgaris was selected for micro-cosmic experiments to simulate a groundwater environment with or without sulfate amendment. The reduction of Fe(III) and As(V) by Desulfovibrio vulgaris was identified, and Fe(III) and As(V) were reduced in both sulfate-free and sulfate-containing systems. However, the addition of 1 mM sulfate significantly enhanced Fe(III) and As(V) reduction. Compared with no sulfate addition, 1 mM sulfate increased the reduction rates of Fe(III) and As(V) by 111.9% and 402.2%, respectively. The sulfate process mediated by Desulfovibrio vulgaris also remarkably promoted arsenic release in sediments. These results indicated that sulfate concentration should be considered when sulfate reduction is used as a remediation method for arsenic pollution in groundwater.

Deletion Mutants, Archived Transposon Library, and Tagged Protein Constructs of the Model Sulfate-Reducing Bacterium Desulfovibrio vulgaris Hildenborough.

The dissimilatory sulfate-reducing deltaproteobacterium Desulfovibrio vulgaris Hildenborough (ATCC 29579) was chosen by the research collaboration ENIGMA to explore tools and protocols for bringing this anaerobe to model status. Here, we describe a collection of genetic constructs generated by ENIGMA that are available to the research community.

Sulfate reducing bacterium Desulfovibrio vulgaris caused severe microbiologically influenced corrosion of zinc and galvanized steel

The microbiologically influenced corrosion (MIC) of zinc and galvanized steel caused by a sulfate reducing bacterium (SRB) was investigated. After 7 days of incubation of Desulfovibrio vulgaris in 125 mL anaerobic vials (100 mL culture medium) at 37 °C, the sessile cell coverage on the galvanized steel was slightly higher than that on pure zinc: (1.9 ± 0.2) × 109 cells/cm2 vs. (9.0 ± 1.8) × 108 cells/cm2. The weight losses for galvanized steel and pure zinc were 31.5 ± 2.5 mg/cm2 and 35.4 ± 4.5 mg/cm2, respectively, which were 101 higher than that for carbon steel. The corrosion current densities of galvanized and pure zinc were 25.5 μA/cm2 and 100 μA/cm2, respectively after the 7-day incubation, confirming that galvanized steel was less prone to SRB MIC despite having a slightly higher sessile cell count. In both cases, the corrosion product was mainly ZnS. Three MIC mechanisms were possible for the severe corrosion against the two metals. Extracellular electron transfer MIC (EET-MIC) was thermodynamically favorable for zinc. Furthermore, in the presence of Zn coupons, H2 evolution in the headspace was 5.5 times higher than without Zn coupons, which suggested that proton attack and/or H2S attack also occurred in the corrosion process.

Effects of ferrous ion concentration on microbiologically influenced corrosion of carbon steel by sulfate reducing bacterium Desulfovibrio vulgaris

Ferrous ion (Fe2+) in a sulfate reducing bacteria (SRB) culture medium is known to enhance microbiologically influenced corrosion (MIC) of carbon steel, but the underlining mechanism is controversial. This work showed that it was due to better sessile cell growth that was likely attributed to Fe2+ detoxification of H2S. Two hundred ppm (w/w) initial Fe2+ in the ATCC 1249 medium achieved a 4.7 times higher Desulfovibrio vulgaris sessile cell count and 5.0 times higher C1018 carbon steel weight loss, compared to 20 ppm. Linear polarization resistance, electrochemical impedance spectroscopy and potentiodynamic polarization measurements corroborated weight loss and pitting data trends.

Migration and Transformation of Adsorbed Arsenic Mediated by Sulfate Reducing Bacteria

In the natural environment, arsenic (As) is mainly adsorbed on iron oxide minerals. The release of adsorbed arsenic from iron oxide minerals to the water is the main source of arsenic pollution. Microbes play a crucial role for this process. The purpose of this study was to investigate the effect of the sulfate-reducing bacteria Desulfovibrio vulgaris DP4 on the transformation and mobilization of As. The experimental results show that the released As concentration of the two systems is 0 μmol·L-1 at 0 h. Compared with the control, DP4 promotes the desorption of As(Ⅴ) before the 84 h incubation process. The released As concentration reaches the maximum value of 12.6 μmol·L-1 at 13 h, accounting for~79% of the initial total adsorbed As (16 μmol·L-1). The maximum released As concentration is~8.4 times higher than that of the control (1.5 μmol·L-1). After 84 hours, the concentration of the released As in the DP4 system is lower than the abiotic control, which suggests that the released As is readsorbed on the solid surface. During the incubation process, the As mobility is significantly correlated with Eh. The XRD results show that the crystallinity of the solid samples in the DP4 system decreases by~50%. In general, a lower crystallinity of the adsorbent indicates a higher adsorption capacity. This may be one important reason for the As readsorption after 84 h. In addition, the SEM shows that goethite is agglomerated by DP4 and the EDS results indicate that goethite is partially transformed to an Fe-S mineral. Based on XANES, arsenic-sulfur minerals were not detected in the solid phase, which further confirms the SEM-EDS results, that is, that Fe-S minerals formed in the solid phase, rather than As2S3 (AsS). The released As was readsorbed on the secondary iron mineral, resulting in a lower dissolved As concentration in the DP4 system than in the abiotic control. Furthermore, 19% As(Ⅲ) was detected in the solid phase while dissolved As(Ⅲ) was not determined during the incubation process. The results suggest that sulfate-reducing bacteria may directly reduce adsorbed As(Ⅴ) to As(Ⅲ).

Filling gaps in bacterial amino acid biosynthesis pathways with high-throughput genetics.

For many bacteria with sequenced genomes, we do not understand how they synthesize some amino acids. This makes it challenging to reconstruct their metabolism, and has led to speculation that bacteria might be cross-feeding amino acids. We studied heterotrophic bacteria from 10 different genera that grow without added amino acids even though an automated tool predicts that the bacteria have gaps in their amino acid synthesis pathways. Across these bacteria, there were 11 gaps in their amino acid biosynthesis pathways that we could not fill using current knowledge. Using genome-wide mutant fitness data, we identified novel enzymes that fill 9 of the 11 gaps and hence explain the biosynthesis of methionine, threonine, serine, or histidine by bacteria from six genera. We also found that the sulfate-reducing bacterium Desulfovibrio vulgaris synthesizes homocysteine (which is a precursor to methionine) by using DUF39, NIL/ferredoxin, and COG2122 proteins, and that homoserine is not an intermediate in this pathway. Our results suggest that most free-living bacteria can likely make all 20 amino acids and illustrate how high-throughput genetics can uncover previously-unknown amino acid biosynthesis genes.

Dispersal and inhibitory roles of mannose, 2-deoxy-d-glucose and N-acetylgalactosaminidase on the biofilm of Desulfovibrio vulgaris.

Biofilms of sulfate-reducing bacteria (SRB) are often the major cause of microbiologically influenced corrosion. The representative SRB Desulfovibrio vulgaris has previously been shown to have a biofilm that consists primarily of protein. In this study, by utilizing lectin staining, we identified that the biofilm of D. vulgaris also consists of the matrix components mannose, fucose and N-acetylgalactosamine (GalNAc), with mannose predominating. Based on these results, we found that the addition of mannose and the nonmetabolizable mannose analog 2-deoxy-d-glucose inhibits the biofilm formation of D. vulgaris as well as that of D. desulfuricans; both compounds also dispersed the SRB biofilms. In addition, the enzyme N-acetylgalactosaminidase, which degrades GalNAc, was effective in dispersing D. vulgaris biofilms. Therefore, by determining composition of the SRB biofilm, effective biofilm control methods may be devised.

Regulation, sensory domains and roles of two Desulfovibrio desulfuricans ATCC27774 Crp family transcription factors, HcpR1 and HcpR2, in response to nitrosative stress.

In silico analyses identified a Crp/Fnr family transcription factor (HcpR) in sulfate-reducing bacteria that controls expression of the hcp gene, which encodes the hybrid cluster protein and contributes to nitrosative stress responses. There is only one hcpR gene in the model sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough, but two copies in Desulfovibrio desulfuricans 27774, which can use nitrate as an alternative electron acceptor to sulfate. Structures of the D. desulfuricans hcpR1, hcpR2 and hcp operons are reported. We present evidence that hcp expression is regulated by HcpR2, not by HcpR1, and that these two regulators differ in both their DNA-binding site specificity and their sensory domains. HcpR1 is predicted to be a b-type cytochrome. HcpR1 binds upstream of the hcpR1 operon and its synthesis is regulated coordinately with hcp in response to NO. In contrast, hcpR2 expression was not induced by nitrate, nitrite or NO. HcpR2 is an iron-sulfur protein that reacts with NO and O2 . We propose that HcpR1 and HcpR2 use different sensory mechanisms to regulate subsets of genes required for defense against NO-induced nitrosative stress, and that diversification of signal perception and DNA recognition by these two proteins is a product of D. desulfuricans adaptation to its particular environmental niche.

Desulfovibrio vulgaris Growth Coupled to Formate-Driven H2 Production.

Formate is recognized as a superior substrate for biological H2 production by several bacteria. However, the growth of a single organism coupled to this energetic pathway has not been shown in mesophilic conditions. In the present study, a bioreactor with gas sparging was used, where we observed for the first time that H2 production from formate can be coupled with growth of the model sulfate-reducing bacterium Desulfovibrio vulgaris in the absence of sulfate or a syntrophic partner. In these conditions, D. vulgaris had a maximum growth rate of 0.078 h(-1) and a doubling time of 9 h, and the ΔG of the reaction ranged between -21 and -18 kJ mol(-1). This is the first report of a single mesophilic organism that can grow while catalyzing the oxidation of formate to H2 and bicarbonate. Furthermore, high volumetric and specific H2 production rates (125 mL L(-1) h(-1) and 2500 mL gdcw(-1) h(-1)) were achieved in a new bioreactor designed and optimized for H2 production. This high H2 production demonstrates that the nonconventional H2-producing organism D. vulgaris is a good biocatalyst for converting formate to H2.

Method for Fast Quantification of Pitting Using 3D Surface Parameters Generated with Infinite Focus Microscope

Empirical correlations that enable fast estimation of maximum and average pit depths and pitted area percentage from the standard 3D surface parameters obtainable with infinite focus microscopy were established in this study. Carbon steel coupons pitted by the sulfate-reducing bacterium Desulfovibrio vulgaris were used as the specimens. The maximum and average pit depths and the pitted area percentage were first determined from laborious measurements of individually manually identified pits on the specimens and then used as the basis for developing the correlations. A four-phase diagram, with average pit depth as the y-axis and pitted area percentage as the x-axis, was proposed as an effective visual tool to categorize the varying pitting and risk patterns. The study results afford more effective, quantitative characterization of pitting corrosion.

Extracellular Electron Transfer Is a Bottleneck in the Microbiologically Influenced Corrosion of C1018 Carbon Steel by the Biofilm of Sulfate-Reducing Bacterium Desulfovibrio vulgaris.

Carbon steels are widely used in the oil and gas industry from downhole tubing to transport trunk lines. Microbes form biofilms, some of which cause the so-called microbiologically influenced corrosion (MIC) of carbon steels. MIC by sulfate reducing bacteria (SRB) is often a leading cause in MIC failures. Electrogenic SRB sessile cells harvest extracellular electrons from elemental iron oxidation for energy production in their metabolism. A previous study suggested that electron mediators riboflavin and flavin adenine dinucleotide (FAD) both accelerated the MIC of 304 stainless steel by the Desulfovibrio vulgaris biofilm that is a corrosive SRB biofilm. Compared with stainless steels, carbon steels are usually far more prone to SRB attacks because SRB biofilms form much denser biofilms on carbon steel surfaces with a sessile cell density that is two orders of magnitude higher. In this work, C1018 carbon steel coupons were used in tests of MIC by D. vulgaris with and without an electron mediator. Experimental weight loss and pit depth data conclusively confirmed that both riboflavin and FAD were able to accelerate D. vulgaris attack against the carbon steel considerably. It has important implications in MIC failure analysis and MIC mitigation in the oil and gas industry.

Growth of the obligate anaerobe Desulfovibrio vulgaris Hildenborough under continuous low oxygen concentration sparging: impact of the membrane-bound oxygen reductases.

Although obligate anaerobe, the sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough (DvH) exhibits high aerotolerance that involves several enzymatic systems, including two membrane-bound oxygen reductases, a bd-quinol oxidase and a cc(b/o)o3 cytochrome oxidase. Effect of constant low oxygen concentration on growth and morphology of the wild-type, single (Δbd, Δcox) and double deletion (Δcoxbd) mutant strains of the genes encoding these oxygen reductases was studied. When both wild-type and deletion mutant strains were cultured in lactate/sulfate medium under constant 0.02% O2 sparging, they were able to grow but the final biomasses and the growth yield were lower than that obtained under anaerobic conditions. At the end of the growth, lactate was not completely consumed and when conditions were then switched to anaerobic, growth resumed. Time-lapse microscopy revealed that a large majority of the cells were then able to divide (over 97%) but the time to recover a complete division event was longer for single deletion mutant Δbd than for the three other strains. Determination of the molar growth yields on lactate suggested that a part of the energy gained from lactate oxidation was derived toward cells protection/repairing against oxidative conditions rather than biosynthesis, and that this part was higher in the single deletion mutant Δbd and, to a lesser extent, Δcox strains. Our data show that when DvH encounters oxidative conditions, it is able to stop growing and to rapidly resume growing when conditions are switched to anaerobic, suggesting that it enters active dormancy sate under oxidative conditions. We propose that the pyruvate-ferredoxin oxidoreductase (PFOR) plays a central role in this phenomenon by reversibly switching from an oxidative-sensitive fully active state to an oxidative-insensitive inactive state. The oxygen reductases, and especially the bd-quinol oxidase, would have a crucial function by maintaining reducing conditions that permit PFOR to stay in its active state.

Integrated Analysis of Transcriptomic and Proteomic Datasets Reveals Information on Protein Expressivity and Factors Affecting Translational Efficiency.

Integrated analysis of large-scale transcriptomic and proteomic data can provide important insights into the metabolic mechanisms underlying complex biological systems. In this chapter, we present methods to address two aspects of issues related to integrated transcriptomic and proteomic analysis. First, due to the fact that proteomic datasets are often incomplete, and integrated analysis of partial proteomic data may introduce significant bias. To address these issues, we describe a zero-inflated Poisson (ZIP)-based model to uncover the complicated relationships between protein abundances and mRNA expression levels, and then apply them to predict protein abundance for the proteins not experimentally detected. The ZIP model takes into consideration the undetected proteins by assuming that there is a probability mass at zero representing expressed proteins that were undetected owing to technical limitations. The model validity is demonstrated using biological information of operons, regulons, and pathways. Second, weak correlation between transcriptomic and proteomic datasets is often due to biological factors affecting translational processes. To quantify the effects of these factors, we describe a multiple regression-based statistical framework to quantitatively examine the effects of various translational efficiency-related sequence features on mRNA-protein correlation. Using the datasets from sulfate-reducing bacteria Desulfovibrio vulgaris, the analysis shows that translation-related sequence features can contribute up to 15.2-26.2% of the total variation of the correlation between transcriptomic and proteomic datasets, and also reveals the relative importance of various features in translation process.

Biofilm growth mode promotes maximum carrying capacity and community stability during product inhibition syntrophy.

Sulfate-reducing bacteria (SRB) can interact syntrophically with other community members in the absence of sulfate, and interactions with hydrogen-consuming methanogens are beneficial when these archaea consume potentially inhibitory H2 produced by the SRB. A dual continuous culture approach was used to characterize population structure within a syntrophic biofilm formed by the SRB Desulfovibrio vulgaris Hildenborough and the methanogenic archaeum Methanococcus maripaludis. Under the tested conditions, monocultures of D. vulgaris formed thin, stable biofilms, but monoculture M. maripaludis did not. Microscopy of intact syntrophic biofilm confirmed that D. vulgaris formed a scaffold for the biofilm, while intermediate and steady-state images revealed that M. maripaludis joined the biofilm later, likely in response to H2 produced by the SRB. Close interactions in structured biofilm allowed efficient transfer of H2 to M. maripaludis, and H2 was only detected in cocultures with a mutant SRB that was deficient in biofilm formation (ΔpilA). M. maripaludis produced more carbohydrate (uronic acid, hexose, and pentose) as a monoculture compared to total coculture biofilm, and this suggested an altered carbon flux during syntrophy. The syntrophic biofilm was structured into ridges (∼300 × 50 μm) and models predicted lactate limitation at ∼50 μm biofilm depth. The biofilm had structure that likely facilitated mass transfer of H2 and lactate, yet maximized biomass with a more even population composition (number of each organism) when compared to the bulk-phase community. Total biomass protein was equivalent in lactate-limited and lactate-excess conditions when a biofilm was present, but in the absence of biofilm, total biomass protein was significantly reduced. The results suggest that multispecies biofilms create an environment conducive to resource sharing, resulting in increased biomass retention, or carrying capacity, for cooperative populations.

CO2 exposure at pressure impacts metabolism and stress responses in the model sulfate-reducing bacterium Desulfovibrio vulgaris strain Hildenborough.

Geologic carbon dioxide (CO2) sequestration drives physical and geochemical changes in deep subsurface environments that impact indigenous microbial activities. The combined effects of pressurized CO2 on a model sulfate-reducing microorganism, Desulfovibrio vulgaris, have been assessed using a suite of genomic and kinetic measurements. Novel high-pressure NMR time-series measurements using 13C-lactate were used to track D. vulgaris metabolism. We identified cessation of respiration at CO2 pressures of 10 bar, 25 bar, 50 bar, and 80 bar. Concurrent experiments using N2 as the pressurizing phase had no negative effect on microbial respiration, as inferred from reduction of sulfate to sulfide. Complementary pressurized batch incubations and fluorescence microscopy measurements supported NMR observations, and indicated that non-respiring cells were mostly viable at 50 bar CO2 for at least 4 h, and at 80 bar CO2 for 2 h. The fraction of dead cells increased rapidly after 4 h at 80 bar CO2. Transcriptomic (RNA-Seq) measurements on mRNA transcripts from CO2-incubated biomass indicated that cells up-regulated the production of certain amino acids (leucine, isoleucine) following CO2 exposure at elevated pressures, likely as part of a general stress response. Evidence for other poorly understood stress responses were also identified within RNA-Seq data, suggesting that while pressurized CO2 severely limits the growth and respiration of D. vulgaris cells, biomass retains intact cell membranes at pressures up to 80 bar CO2. Together, these data show that geologic sequestration of CO2 may have significant impacts on rates of sulfate reduction in many deep subsurface environments where this metabolism is a key respiratory process.