Activity and Diversity of Sulfate-Reducing Bacteria in a Petroleum Hydrocarbon-Contaminated Aquifer

Microbial sulfate reduction is an important metabolic activity in petroleum hydrocarbon (PHC)-contaminated aquifers. We quantified carbon source-enhanced microbial SO₄²⁻ reduction in a PHC-contaminated aquifer by using single-well push-pull tests and related the consumption of sulfate and added carbon sources to the presence of certain genera of sulfate-reducing bacteria (SRB). We also used molecular methods to assess suspended SRB diversity. In four consecutive tests, we injected anoxic test solutions (1,000 liters) containing bromide as a conservative tracer, sulfate, and either propionate, butyrate, lactate, or acetate as reactants into an existing monitoring well. After an initial incubation period, 1,000 liters of test solution-groundwater mixture was extracted from the same well. Average total test duration was 71 h. We measured concentrations of bromide, sulfate, and carbon sources in native groundwater as well as in injection and extraction phase samples and characterized the SRB population by using fluorescence in situ hybridization (FISH) and denaturing gradient gel electrophoresis (DGGE). Enhanced sulfate reduction concomitant with carbon source degradation was observed in all tests. Computed first-order rate coefficients ranged from 0.19 to 0.32 day⁻¹ for sulfate reduction and from 0.13 to 0.60 day⁻¹ for carbon source degradation. Sulfur isotope fractionation in unconsumed sulfate indicated that sulfate reduction was microbially mediated. Enhancement of sulfate reduction due to carbon source additions in all tests and variability of rate coefficients suggested the presence of specific SRB genera and a high diversity of SRB. We confirmed this by using FISH and DGGE. A large fraction of suspended bacteria hybridized with SRB-targeting probes SRB385 plus SRB385-Db (11 to 24% of total cells). FISH results showed that the activity of these bacteria was enhanced by addition of sulfate and carbon sources during push-pull tests. However, DGGE profiles indicated that the bacterial community structure of the dominant species did not change during the tests. Thus, the combination of push-pull tests with molecular methods provided valuable insights into microbial processes, activities, and diversity in the sulfate-reducing zone of a PHC-contaminated aquifer.     

Structural and Functional Dynamics of Sulfate-Reducing Populations in Bacterial Biofilms

We describe the combined application of microsensors and molecular techniques to investigate the development of sulfate reduction and of sulfate-reducing bacterial populations in an aerobic bacterial biofilm. Microsensor measurements for oxygen showed that anaerobic zones developed in the biofilm within 1 week and that oxygen was depleted in the top 200 to 400 μm during all stages of biofilm development. Sulfate reduction was first detected after 6 weeks of growth, although favorable conditions for growth of sulfate-reducing bacteria (SRB) were present from the first week. In situ hybridization with a 16S rRNA probe for SRB revealed that sulfate reducers were present in high numbers (approximately 10⁸ SRB/ml) in all stages of development, both in the oxic and anoxic zones of the biofilm. Denaturing gradient gel electrophoresis (DGGE) showed that the genetic diversity of the microbial community increased during the development of the biofilm. Hybridization analysis of the DGGE profiles with taxon-specific oligonucleotide probes showed that Desulfobulbus and Desulfovibrio were the main sulfate-reducing bacteria in all biofilm samples as well as in the bulk activated sludge. However, different Desulfobulbus and Desulfovibrio species were found in the 6th and 8th weeks of incubation, respectively, coinciding with the development of sulfate reduction. Our data indicate that not all SRB detected by molecular analysis were sulfidogenically active in the biofilm.     

Sulfate Reduction at a Lignite Seam: Microbial Abundance and Activity

In a combined isotope geochemical and microbiological investigation, a setting of multiple aquifers was characterized. Biologically mediated redox processes were observed in the aquifers situated in marine sands of Tertiary age and overlying Quaternary gravel deposits. Intercalated lignite seams define the aquitards, which separate the aquifers. Bacterial oxidation of organic matter is evident from dissolved inorganic carbon characterized by average carbon isotope values between ?18.4 per thousand and ?15.7 per thousand (PDB). Strongly positive sulfur isotope values of up to +50 per thousand (CTD) for residual sulfate indicate sulfate reduction under closed system conditions with respect to sulfate availability. Both, hydrochemical and isotope data are thus consistent with the recent activity of sulfate-reducing bacteria (SRB). Microbiological investigations revealed the presence of an anaerobic food chain in the aquifers. Most-probable-number (MPN) determinations for SRB and fermenting microorganisms reached highest values at the interface between aquifer and lignite seam (1.5 x 103 cells/g sediment dry mass). Five strains of SRB were isolated from highest MPN dilutions. Spore-forming bacteria appeared to dominate the SRB population. Sulfate reduction rates were determined by the 35S-radiotracer method. A detailed assessment indicates an increase in the reduction rate in proximity to the lignite seam, with a maximum turnover of 8.4 mM sulfate/a, suggesting that lignite-drived compounds represent the substrate for sulfate reduction.     

Effect of nitrite and nitrate on in situ sulfide production in an activated sludge immobilized agar gel film as determined by use of microelectrodes

Microelectrode, fluorescence in situ hybridization (FISH), and denaturing gradient gel electrophoresis (DGGE) analyses were used to investigate the effect of nitrite and nitrate on in situ sulfide production in an activated sludge immobilized agar gel film. Microelectrode measurements of O(2), H(2)S, NO(3)(-), NO(2)(-), and pH revealed that the addition of NO(2)(-) and NO(3)(-) forced sulfate reduction zones deeper in the agar gel and significantly reduced the in situ sulfide production levels. The sulfate reduction zone was consequently separated from O(2) and NO(2)(-) or NO(3)(-) respiration zones with increasing the concentrations of NO(2)(-) and NO(3)(-). These NO(2)(-) and NO(3)(-) treatments had only a transient effect on sulfide production. The in situ sulfide production quickly recovered to the previous levels when NO(2)(-) and NO(3)(-) were removed. The PCR-DGGE and FISH analyses revealed that 2-day-continuous addition of 500 microM NO(3)(-) did not change the metabolically active sulfate-reducing bacterial (SRB) community. On the basis of these data, it could be concluded that the addition of NO(2)(-) and NO(3)(-) did not kill SRB, but induced the interspecies competition for common carbon source (i.e., acetate) between nitrate-reducing heterotrophic bacteria and SRB and enhanced the oxidation of the produced sulfide, which were main possible causes of the suppression of in situ sulfide production in the agar gel.     

Use of a three-stage continuous culture system to study the effect of mucin on dissimilatory sulfate reduction and methanogenesis by mixed populations of human gut bacteria

A mixed culture of human fecal bacteria was grown for 120 days in a three-stage continuous culture system. To reproduce some of the nutritional and pH characteristics of the large gut, each vessel had a different operating volume (0.3, 0.5, and 0.8 liter) and pH (6.0, 6.5, and 7.0). A mixture of polysaccharides and proteins was used as carbon and nitrogen sources. Measurements of H2, CH4, S2-, sulfate reduction rates, sulfate-reducing bacteria (SRB), and volatile fatty acids were made throughout the experiment. After 48 days of running, porcine gastric mucin (5.8 g/day) was independently fed to vessel 1 of the multichamber system. The mucin was extensively degraded as evidenced by the stimulation of volatile fatty acid production. In the absence of mucin, sulfate-reducing activity was comparatively insignificant and methanogenesis was the major route for the disposal of electrons. The reverse occurred upon the addition of mucin; sulfate reduction predominated and methanogenesis was completely inhibited. This was attributed to release of sulfate from the mucin which enabled SRB to outcompete methanogenic bacteria for H2. SRB stimulated by mucin were acetate-utilizing Desulfobacter spp., lactate- and H2-utilizing Desulfovibrio spp., and propionate-utilizing Desulfobulbus spp. When the mucin pump was switched off, the multichamber system reverted to a state close to its original equilibrium. These data provide further evidence that sulfated polysaccharides such as mucin may be a source of sulfate for SRB in the human large gut.     

Use of a three-stage continuous culture system to study the effect of mucin on dissimilatory sulfate reduction and methanogenesis by mixed populations of human gut bacteria.

A mixed culture of human fecal bacteria was grown for 120 days in a three-stage continuous culture system. To reproduce some of the nutritional and pH characteristics of the large gut, each vessel had a different operating volume (0.3, 0.5, and 0.8 liter) and pH (6.0, 6.5, and 7.0). A mixture of polysaccharides and proteins was used as carbon and nitrogen sources. Measurements of H2, CH4, S2-, sulfate reduction rates, sulfate-reducing bacteria (SRB), and volatile fatty acids were made throughout the experiment. After 48 days of running, porcine gastric mucin (5.8 g/day) was independently fed to vessel 1 of the multichamber system. The mucin was extensively degraded as evidenced by the stimulation of volatile fatty acid production. In the absence of mucin, sulfate-reducing activity was comparatively insignificant and methanogenesis was the major route for the disposal of electrons. The reverse occurred upon the addition of mucin; sulfate reduction predominated and methanogenesis was completely inhibited. This was attributed to release of sulfate from the mucin which enabled SRB to outcompete methanogenic bacteria for H2. SRB stimulated by mucin were acetate-utilizing Desulfobacter spp., lactate- and H2-utilizing Desulfovibrio spp., and propionate-utilizing Desulfobulbus spp. When the mucin pump was switched off, the multichamber system reverted to a state close to its original equilibrium. These data provide further evidence that sulfated polysaccharides such as mucin may be a source of sulfate for SRB in the human large gut.     

Nested PCR-denaturing gradient gel electrophoresis approach to determine the diversity of sulfate-reducing bacteria in complex microbial communities

Here, we describe a three-step nested-PCR-denaturing gradient gel electrophoresis (DGGE) strategy to detect sulfate-reducing bacteria (SRB) in complex microbial communities from industrial bioreactors. In the first step, the nearly complete 16S rRNA gene was amplified using bacterial primers. Subsequently, this product was used as a template in a second PCR with group-specific SRB primers. A third round of amplification was conducted to obtain fragments suitable for DGGE. The largest number of bands was observed in DGGE patterns of products obtained with primers specific for the Desulfovibrio-Desulfomicrobium group, indicating a large diversity of these SRBs. In addition, members of other phylogenetic SRB groups, i.e., Desulfotomaculum, Desulfobulbus, and Desulfococcus-Desulfonema-Desulfosarcina, were detected. Bands corresponding to Desulfobacterium and Desulfobacter were not detected in the bioreactor samples. Comparative sequence analysis of excised DGGE bands revealed the identity of the community members. The developed three-step PCR-DGGE strategy is a welcome tool for studying the diversity of sulfate-reducing bacteria.     

Molecular- and cultivation-based analyses of microbial communities in oil field water and in microcosms amended with nitrate to control H2S production

Nitrate injection into oil fields is an alternative to biocide addition for controlling sulfide production (‘souring’) caused by sulfate-reducing bacteria (SRB). This study examined the suitability of several cultivation-dependent and cultivation-independent methods to assess potential microbial activities (sulfidogenesis and nitrate reduction) and the impact of nitrate amendment on oil field microbiota. Microcosms containing produced waters from two Western Canadian oil fields exhibited sulfidogenesis that was inhibited by nitrate amendment. Most probable number (MPN) and fluorescent in situ hybridization (FISH) analyses of uncultivated produced waters showed low cell numbers (≤10³ MPN/ml) dominated by SRB (>95% relative abundance). MPN analysis also detected nitrate-reducing sulfide-oxidizing bacteria (NRSOB) and heterotrophic nitrate-reducing bacteria (HNRB) at numbers too low to be detected by FISH or denaturing gradient gel electrophoresis (DGGE). In microcosms containing produced water fortified with sulfate, near-stoichiometric concentrations of sulfide were produced. FISH analyses of the microcosms after 55 days of incubation revealed that Gammaproteobacteria increased from undetectable levels to 5–20% abundance, resulting in a decreased proportion of Deltaproteobacteria (50–60% abundance). DGGE analysis confirmed the presence of Delta- and Gammaproteobacteria and also detected Bacteroidetes. When sulfate-fortified produced waters were amended with nitrate, sulfidogenesis was inhibited and Deltaproteobacteria decreased to levels undetectable by FISH, with a concomitant increase in Gammaproteobacteria from below detection to 50–60% abundance. DGGE analysis of these microcosms yielded sequences of Gamma- and Epsilonproteobacteria related to presumptive HNRB and NRSOB (Halomonas, Marinobacterium, Marinobacter, Pseudomonas and Arcobacter), thus supporting chemical data indicating that nitrate-reducing bacteria out-compete SRB when nitrate is added.     

水稻土中硫酸盐还原微生物研究进展

硫是水稻必需的营养元素之一.硫酸盐还原是硫元素生物地球化学循环中的关键步骤,在稻田土壤表层和水稻根际都十分活跃.介导硫酸盐还原过程的硫酸盐还原菌(sulfate- reducing bacteria, SRB)是稻田土壤中重要的功能菌群.它们不仅是硫元素生物地球化学循环的重要参与者,也是土壤中有机污染物降解的主要力量之一,发挥着重要的生态和环境功能.综述了稻田土壤中微生物参与的硫酸盐还原过程、SRB的生物多样性以及目前研究稻田土壤SRB主要采用的分子生态学方法,如末端限制性片段长度多样性(T-RFLP)、变性梯度凝胶电泳(DGGE)、实时荧光定量PCR(real-time PCR)、荧光原位杂交(FISH),并对水稻土壤中SRB的分子生态学研究方向进行了展望.     

A Study of the Relative Dominance of Selected Anaerobic Sulfate-Reducing Bacteria in a Continuous Bioreactor by Fluorescence <Emphasis Type="Italic">in Situ</Emphasis> Hybridization

The diversity and the community structure of sulfate-reducing bacteria (SRB) in an anaerobic continuous bioreactor used for treatment of a sulfate-containing wastewater were investigated by fluorescence in situ hybridization. Hybridization to the 16S rRNA probe EUB338 for the domain Bacteria was performed, followed by a nonsense probe NON338 as a control for nonspecific staining. Sulfate-reducing consortia were identified by using five nominally genus-specific probes (SRB129 for Desulfobacter, SRB221 for Desulfobacterium, SRB228 for Desulfotomaculum, SRB660 for Desulfobulbus, and SRB657 for Desulfonema) and four group-specific probes (SRB385 as a general SRB probe, SRB687 for Desulfovibrioaceae, SRB814 for Desulfococcus group, and SRB804 for Desulfobacteriaceae). The total prokaryotic population was determined by 4′,6-diamidino-2-phenylindole staining. Hybridization analysis using these 16S rRNA-targeted oligonucleotide probes showed that, of those microbial groupings investigated, Desulfonema, Desulfobulbus, spp., and Desulfobacteriaceae group were the main sulfate-reducing bacteria in the bioreactor when operated at steady state at 35°C, pH 7.8, and a 2.5-day residence time with feed stream containing 2.5 kg m⁻³ sulfate as terminal electron acceptor and 2.3 kg m⁻³ acetate as carbon source and electron donor.     

Analyses of spatial distributions of sulfate-reducing bacteria and their activity in aerobic wastewater biofilms.

The vertical distribution of sulfate-reducing bacteria (SRB) in aerobic wastewater biofilms grown on rotating disk reactors was investigated by fluorescent in situ hybridization (FISH) with 16S rRNA-targeted oligonucleotide probes. To correlate the vertical distribution of SRB populations with their activity, the microprofiles of O(2), H(2)S, NO(2)(-), NO(3)(-), NH(4)(+), and pH were measured with microelectrodes. In addition, a cross-evaluation of the FISH and microelectrode analyses was performed by comparing them with culture-based approaches and biogeochemical measurements. In situ hybridization revealed that a relatively high abundance of the probe SRB385-stained cells (approximately 10(9) to 10(10) cells per cm(3) of biofilm) were evenly distributed throughout the biofilm, even in the oxic surface. The probe SRB660-stained Desulfobulbus spp. were found to be numerically important members of SRB populations (approximately 10(8) to 10(9) cells per cm(3)). The result of microelectrode measurements showed that a high sulfate-reducing activity was found in a narrow anaerobic zone located about 150 to 300 microm below the biofilm surface and above which an intensive sulfide oxidation zone was found. The biogeochemical measurements showed that elemental sulfur (S(0)) was an important intermediate of the sulfide reoxidation in such thin wastewater biofilms (approximately 1,500 microm), which accounted for about 75% of the total S pool in the biofilm. The contribution of an internal Fe-sulfur cycle to the overall sulfur cycle in aerobic wastewater biofilms was insignificant (less than 1%) due to the relatively high sulfate reduction rate.     

Activity of sulfate-reducing bacteria in human periodontal pocket

Samples of subgingival dental tissues were examined for the presence of sulfate-reducing bacteria (SRB). Using enrichment cultures, SRBs were detected in 9 of 17 individuals. A pure culture of SRB was obtained from one sample collected from a patient with type IV periodontal disease. The characterization of this isolate showed that it belongs to the genus Desulfovibrio. The isolate used pyruvate, lactate, glucose, fructose, and ethanol as the sole source of carbon. However, the isolate was unable to use acetate and methanol as a carbon source, indicating it as an incomplete oxidizer unable to carry out the terminal oxidation of substrates. Apart from using sulfate as electron acceptor, the isolate also used thiosulfate and nitrate as an electron acceptor. It has the ability to use a variety of nitrogen sources, including ammonium chloride, nitrate, and glutamate. The optimum growth temperature of the isolate was 37 degrees C and the optimum pH for growth was 6.8. The SRB isolate contained the electron carrier desulfoviridin. The numbers of SRB in the mouth are assumed to be limited by sulfate. Potential sources of sulfate in the subgingival area include free sulfate in pocket fluid and glycosaminoglycans and sulfur-containing amino acids from periodontal tissues.     

Analysis of diversity and activity of sulfate-reducing bacterial communities in sulfidogenic bioreactors using 16S rRNA and dsrB genes as molecular markers

Here we describe the diversity and activity of sulfate-reducing bacteria (SRB) in sulfidogenic bioreactors by using the simultaneous analysis of PCR products obtained from DNA and RNA of the 16S rRNA and dissimilatory sulfite reductase (dsrAB) genes. We subsequently analyzed the amplified gene fragments by using denaturing gradient gel electrophoresis (DGGE). We observed fewer bands in the RNA-based DGGE profiles than in the DNA-based profiles, indicating marked differences in the populations present and in those that were metabolically active at the time of sampling. Comparative sequence analyses of the bands obtained from rRNA and dsrB DGGE profiles were congruent, revealing the same SRB populations. Bioreactors that received either ethanol or isopropanol as an energy source showed the presence of SRB affiliated with Desulfobulbus rhabdoformis and/or Desulfovibrio sulfodismutans, as well as SRB related to the acetate-oxidizing Desulfobacca acetoxidans. The reactor that received wastewater containing a diverse mixture of organic compounds showed the presence of nutritionally versatile SRB affiliated with Desulfosarcina variabilis and another acetate-oxidizing SRB, affiliated with Desulfoarculus baarsii. In addition to DGGE analysis, we performed whole-cell hybridization with fluorescently labeled oligonucleotide probes to estimate the relative abundances of the dominant sulfate-reducing bacterial populations. Desulfobacca acetoxidans-like populations were most dominant (50 to 60%) relative to the total SRB communities, followed by Desulfovibrio-like populations (30 to 40%), and Desulfobulbus-like populations (15 to 20%). This study is the first to identify metabolically active SRB in sulfidogenic bioreactors by using the functional gene dsrAB as a molecular marker. The same approach can also be used to infer the ecological role of coexisting SRB in other habitats.     

Distribution, diversity, and activity of sulfate-reducing bacteria in the water column in Gek-Gel Lake, Azerbaijan

The distribution and activity of sulfate-reducing bacteria (SRB) in the water column of the alpine meromictic Gek-Gel lake were studied. Apart from traditional microbiological methods based on cultivation and on measuring the process rates with radioactive labels, in situ fluorescent hybridization (FISH) was used, which enables identification and quantification without cultivating organisms. The peak rate of sulfate reduction, 0.486 microg S/(l day), was found in the chemocline at 33 m. The peak SRB number of 2.5 x 106 cells/ml, as determined by the end-point dilutions method on selective media, was found at the same depth. The phylogenetic position of the SRB, as determined by FISH, revealed the predominance of the Desulfovibrio spp., Desulfobulbus spp., and Desulfoarculus spp./Desulfomonile spp. groups. The numbers of spore-forming Desulfotomaculum spp. increased with depth. The low measured rates of sulfate reduction accompanied with high SRB numbers and the predominance of the groups capable of reducing a wide range of substrates permit us to propose utilization of electron acceptors other than sulfate as the main activity of the SRB in the water column.     

Nested PCR-Denaturing Gradient Gel Electrophoresis Approach To Determine the Diversity of Sulfate-Reducing Bacteria in Complex Microbial Communities

Here, we describe a three-step nested-PCR-denaturing gradient gel electrophoresis (DGGE) strategy to detect sulfate-reducing bacteria (SRB) in complex microbial communities from industrial bioreactors. In the first step, the nearly complete 16S rRNA gene was amplified using bacterial primers. Subsequently, this product was used as a template in a second PCR with group-specific SRB primers. A third round of amplification was conducted to obtain fragments suitable for DGGE. The largest number of bands was observed in DGGE patterns of products obtained with primers specific for the Desulfovibrio-Desulfomicrobium group, indicating a large diversity of these SRBs. In addition, members of other phylogenetic SRB groups, i.e., Desulfotomaculum, Desulfobulbus, and Desulfococcus-Desulfonema-Desulfosarcina, were detected. Bands corresponding to Desulfobacterium and Desulfobacter were not detected in the bioreactor samples. Comparative sequence analysis of excised DGGE bands revealed the identity of the community members. The developed three-step PCR-DGGE strategy is a welcome tool for studying the diversity of sulfate-reducing bacteria.     

Successional development of sulfate-reducing bacterial populations and their activities in an activated sludge immobilized agar gel film

A combination of fluorescence in situ hybridization (FISH), microprofiles, and denaturing gradient gel electrophoresis (DGGE) analysis of PCR-amplified 16S rDNA fragments followed by hybridization analysis with specific probes was applied to investigate successional development of sulfate-reducing bacteria (SRB) community structure and in situ sulfide production activity within an activated sludge immobilized agar gel film. In this model biofilm system, since biases arising from biofilm heterogeneity can be ignored, the population dynamics of SRB in the agar gel is directly related to physiological capability and in situ activity of SRB. Microelectrode measurements showed that an anoxic zone was already developed at the beginning (0 day), a first sulfide production of 0.054 mumol H2S m(-2) x s(-1) was detected during the first week, and the rate increased gradually to 0.221 mumol H2S m(-2) x s(-1) in the fifth week. The most active sulfide production zone moved upward to the chemocline and intensified with time to form a narrow zone with high volumetric sulfide production rates. This result coincided with the shift of the spatial distributions of SRB populations determined by FISH. In situ hybridization with probe SRB385 for mainly general SRB of the delta Proteobacteria plus some gram-positive bacteria and probe 660 for Desulfobulbus indicated that the most abundant populations of SRB were primarily restricted to near the oxic/anoxic interface (chemocline). A close observation of the development of the vertical distributions of SRB populations revealed that the cell numbers of Desulfobulbus tripled (from 0.5 x 10(8) to 1.5 x 10(8) cells cm(-3)) near the oxic/anoxic interface. Similar growth (from 1.0 x10(8) to 4.5 x 10(8) cells cm(-3)) of Desulfovibrio-like SRB that hybridized with probe SRB385 was observed. PCR-DGGE followed by hybridization analysis revealed that one Desulfobulbus strain was detected from the beginning, and another strain appeared after 1 week, coinciding with the first detected sulfide production. In addition, three strains hybridizing with probe 687 (possibly Desulfovibrio) were also dominant SRB in the agar gel.     

Microbial community analysis of rectal methanogens and sulfate reducing bacteria in two non-human primate species

Background  Methanogenesis by methanogenic Archaea and sulfate reduction by sulfate reducing bacteria (SRB) are the major hydrogenotrophic pathways in the human colon. Methanogenic status of mammals is suggested to be under evolutionary rather than dietary control. However, information is lacking regarding the dynamics of hydrogenotrophic microbial communities among different primate species.
Methods  Rectal swabs were collected from 10 sooty mangabeys ( Cercocebus atys ) and 10 baboons ( Papio hamadryas ). The diversity and abundance of methanogens and SRB were examined using PCR-denaturing gradient gel electrophoresis (DGGE) and real-time quantitative PCR (qPCR).
Results  The DGGE results revealed that intestinal Archaea and SRB communities differ between mangabeys and baboons. Phylogenetic analyses of Archaea DGGE bands revealed two distinct clusters with one representing a putative novel order of methanogenic Archaea. The qPCR detected a similar abundance of methanogens and SRB.
Conclusions  Intestinal Archaea and SRB coexist in these primates, and the community patterns are host species-specific.     

Spatial variability of sulfate reduction in a shallow aquifer

The distribution and metabolic activity of sulfate-reducing bacteria (SRB) in a shallow, suboxic aquifer were studied. A radioimaging technique was used to visualize and quantify the activity of sulfate reducers in sediments at a centimetre-level scale. The distribution of SRB metabolic activity was heterogeneous with areas showing little activity far outnumbering areas with high activity. Variation in sulfate-reducing activity was not statistically correlated with variation in depth, bacterial numbers, or the following sediment properties: sediment type (sand, peat or silt), grain size, permeability and hydraulic conductivity. Sulfate-reducing bacteria activity did vary significantly with sediment porosity (multivariate analysis, r = 0.48). We hypothesized that the small pore sizes associated with sediments with low porosity restricted the ability of SRB to grow to high numbers as well as their access to nutrients. To further explore the relationship between pore size and microbial metabolic activity, columns with varying pore diameters were constructed. Sulfate-reducing bacteria in the columns with the smallest pore diameters had the lowest rates of metabolism and SRB metabolic rates increased as the pore diameter increased. For the aquifer studied, sediment porosities and pore sizes were the main factor controlling SRB activity.     

Comparing lactate and glycerol as a single-electron donor for sulfate reduction in fluidized bed reactors

Among the greatest challenges to the full implementation of biological sulfate reduction are the cost and availability of the electron source. With the development of the biofuel industry, new organic substrates have become available. Therefore, this work sought to compare the performance of a sulfidogenic process utilizing either lactate or glycerol as the substrate for sulfate-reducing bacteria (SRB) growth. Although sulfate reduction is energetically more favorable with lactate, glycerol is a less expensive alternative because excess production is forecasted with the worldwide development of the biodiesel industry. Continuous experiments were performed in a fluidized bed (FB) reactor containing activated carbon as a carrier for a mixed bacterial population composed of sulfate-reducing and fermentative bacteria. During the lactate-fed phases, incomplete oxidation of lactate to acetate by SRB was the dominant metabolic pathway resulting in as much as 90 % sulfate reduction and high acetate concentrations (2.7 g L^?1). Conversely, in the glycerol-fed phases, glycerol degradation resulted from syntrophic cooperation between sulfate-reducing and fermentative bacteria that produce butyrate along with acetate (1.0 g L^?1) as oxidation products. To our knowledge, this is the first report of butyrate formation during sulfate reduction in a glycerol-fed continuous-flow reactor. Sulfate concentrations were reduced by about 90 % (from 2,000 to 100–300 mg L^?1) when glycerol was being fed to the reactor. Since the FB reactor was able to stand a change from lactate to glycerol, this reactor is recommended as the preferred option should glycerol be selected as a cost-effective alternative to lactate for continuous sulfate reduction.     

Diversity and vertical distribution of cultured and uncultured Deltaproteobacteria in an intertidal mud flat of the Wadden Sea

The diversity and distribution of Deltaproteobacteria in an intertidal mud flat of the German Wadden Sea was characterized by molecular biological techniques and cultivation. A 16S rRNA gene library generated with general primers (303 clones) suggested that sulfate-reducing bacteria (SRB) related to Desulfobulbaceae and Desulfosarcina were abundant. Fluorescence in situ hybridization (FISH) with probes targeting these groups was used to characterize their vertical distribution. The combination of FISH with catalysed reporter deposition (CARD-FISH) significantly enhanced the detection of selected subgroups of Deltaproteobacteria, particularly in deeper sediment layers. Up to 11% of all cells were assigned to SRB. Organisms related to Desulfosarcina and Desulfobulbaceae were the dominant SRB in the surface sediments. Two abundant subpopulations of Desulfosarcina-related bacteria were identified by FISH. The SRB community differed between the sampling site and a sandy intertidal flat chosen as a reference. Enrichments and MPN cultures inoculated with surface sediment were monitored by FISH. Nine strains of Deltaproteobacteria were isolated. Four strains were related to Desulfobulbaceae, such as Desulfobacterium catecholicum and Desulfocapsa spp. A subgroup including clone sequences and strains related to D. catecholicum could be detected in situ by a specific FISH probe. The first physiological experiments suggested specific functional roles for the isolates. Two strains utilized environmentally relevant compounds in coastal areas such as catechol and nitrate. One strain related to Desulfocapsa spp. disproportionated thiosulfate and might thus contribute to the sulfur isotope fractionation at the study site. A Fe(III)-reducing strain was obtained that affiliated with the Pelobacter-Desulphuromonas group. This group accounted for up to 6% of total cell numbers and even exceeded SRB numbers in upper sediment layers. These bacteria might substantially contribute to carbon mineralization via dissimilatory reduction of, e.g. Fe(III).