Establishment and metabolic analysis of a model microbial community for understanding trophic and electron accepting interactions of subsurface anaerobic environments

<p>Abstract</p> <p>Background</p> <p>Communities of microorganisms control the rates of key biogeochemical cycles, and are important for biotechnology, bioremediation, and industrial microbiological processes. For this reason, we constructed a model microbial community comprised of three species dependent on trophic interactions. The three species microbial community was comprised of <it>Clostridium cellulolyticum</it>, <it>Desulfovibrio vulgaris </it>Hildenborough, and <it>Geobacter sulfurreducens </it>and was grown under continuous culture conditions. Cellobiose served as the carbon and energy source for <it>C. cellulolyticum</it>, whereas <it>D. vulgaris </it>and <it>G. sulfurreducens </it>derived carbon and energy from the metabolic products of cellobiose fermentation and were provided with sulfate and fumarate respectively as electron acceptors.</p> <p>Results</p> <p>qPCR monitoring of the culture revealed <it>C. cellulolyticum </it>to be dominant as expected and confirmed the presence of <it>D. vulgaris </it>and <it>G. sulfurreducens</it>. Proposed metabolic modeling of carbon and electron flow of the three-species community indicated that the growth of <it>C. cellulolyticum </it>and <it>D. vulgaris </it>were electron donor limited whereas <it>G. sulfurreducens </it>was electron acceptor limited.</p> <p>Conclusions</p> <p>The results demonstrate that <it>C. cellulolyticum</it>, <it>D. vulgaris</it>, and <it>G. sulfurreducens </it>can be grown in coculture in a continuous culture system in which <it>D. vulgaris </it>and <it>G. sulfurreducens </it>are dependent upon the metabolic byproducts of <it>C. cellulolyticum </it>for nutrients. This represents a step towards developing a tractable model ecosystem comprised of members representing the functional groups of a trophic network.</p

Impact of Terrestrial Input on Deep-Sea Benthic Archaeal Community Structure in South China Sea Sediments

© Copyright © 2020 Lai, Hedlund, Xie, Liu, Phelps, Zhang and Wang. Archaea are widespread in marine sediments and play important roles in the cycling of sedimentary organic carbon. However, factors controlling the distribution of archaea in marine sediments are not well understood. Here we investigated benthic archaeal communities over glacial-interglacial cycles in the northern South China Sea and evaluated their responses to sediment organic matter sources and inter-species interactions. Archaea in sediments deposited during the interglacial period Marine Isotope Stage (MIS) 1 (Holocene) were significantly different from those in sediments deposited in MIS 2 and MIS 3 of the Last Glacial Period when terrestrial input to the South China Sea was enhanced based on analysis of the long-chain n-alkane C31. The absolute archaeal 16S rRNA gene abundance in subsurface sediments was highest in MIS 2, coincident with high sedimentation rates and high concentrations of total organic carbon. Soil Crenarchaeotic Group (SCG; Nitrososphaerales) species, the most abundant ammonia-oxidizing archaea in soils, increased dramatically during MIS 2, likely reflecting transport of terrestrial archaea during glacial periods with high sedimentation rates. Co-occurrence network analyses indicated significant association of SCG archaea with benthic deep-sea microbes such as Bathyarchaeota and Thermoprofundales in MIS 2 and MIS 3, suggesting potential interactions among these archaeal groups. Meanwhile, Thermoprofundales abundance was positively correlated with total organic carbon (TOC), along with n-alkane C31 and sedimentation rate, indicating that Thermoprofundales may be particularly important in processing of organic carbon in deep-sea sediments. Collectively, these results demonstrate that the composition of heterotrophic benthic archaea in the South China Sea may be influenced by terrestrial organic input in tune with glacial-interglacial cycles, suggesting a plausible link between global climate change and microbial population dynamics in deep-sea marine sediments

Microbial Community Dynamics of Lactate Enriched Hanford Groundwaters

The Department of Energy site at Hanford, WA, has been historically impacted by U and Cr from the nuclear weapons industry. In an attempt to stimulate microbial remediation of these metals, in-situ lactate enrichment experiments are ongoing. In order to bridge the gap from the laboratory to the field, we inoculated triplicate anaerobic, continuous-flow glass reactors with groundwater collected from well Hanford 100-H in order to obtain a stable, enriched community while selecting for metal-reducing bacteria. Each reactor was fed from a single carboy containing defined media with 30 mM lactate at a rate of 0.223 ml/min under continuous nitrogen flow at 9 ml/min. Cell counts, organic acids, gDNA (for qPCR and pyrosequencing) and gases were sampled during the experiment. Cell counts remained low (less than 1x107 cells/ml) during the first two weeks of the experiment, but by day 20, had reached a density greater than 1x108 cells/ml. Metabolite analysis showed a decrease in the lactate concentrations over time. Pyruvate concentrations ranged from 20-40 uM the first week of the experiment then was undetectable after day 10. Likewise, formate appeared in the reactors during the first week with concentrations of 1.48-1.65 mM at day 7 then the concentrations decreased to 0.69-0.95 on day 10 and were undetectable on day 15. Acetate was present in low amounts on day 3 (0.15-0.33 mM) and steadily increased to 3.35-5.22 mM over time. Similarly, carbon dioxide was present in low concentrations early on and increased to 0.28-0.35 mM as the experiment progressed. We also were able to detect low amounts of methane (10-20 uM) during the first week of the experiment, but by day 10 the methane was undetectable. From these results and pyrosequencing analysis, we conclude that a shift in the microbial community dynamics occurred over time to eventually form a stable and enriched microbial community. Comprehensive investigations such as these allow for the examination of not only which nutrient source will accelerate site remediation, but also provide insight to evaluate remediation strategies through which enriched community members are important for bioremediation

Hexavalent Chromium Reduction under Fermentative Conditions with Lactate Stimulated Native Microbial Communities

This work conducted by ENIGMA- Ecosystems and Networks Integrated with Genes and Molecular Assemblies (http://enigma.lbl.gov), a Scientific Focus Area Program at Lawrence Berkeley National Laboratory. The submitted manuscript has been authored by a contractor of the U.S. Government under contract DE-AC05-00OR22725. Accordingly, the U.S. Government retains a nonexclusive, royalty-free license to publish or reproduce the published form of this contribution, or allow others to do so, for U.S. Government purposes.Conceived and designed the experiments: ACS JJM MP TJP SDB AVP DAE. Performed the experiments: ACS JJM ZKY. Analyzed the data: ACS JJM TY JDVN JZ DAE. Contributed reagents/materials/analysis tools: DAE TCH APA. Wrote the paper: ACS JJM DAE.Microbial reduction of toxic hexavalent chromium (Cr(VI)) in-situ is a plausible bioremediation strategy in electron-acceptor limited environments. However, higher Cr(VI) may impose stress on syntrophic communities and impact community structure and function. The study objectives were to understand the impacts of Cr(VI) concentrations on community structure and on the Cr(VI)-reduction potential of groundwater communities at Hanford, WA. Steady state continuous flow bioreactors were used to grow native communities enriched with lactate (30 mM) and continuously amended with Cr(VI) at 0.0 (No-Cr), 0.1 (Low-Cr) and 3.0 (High-Cr) mg/L. Microbial growth, metabolites, Cr(VI), 16S rRNA gene sequences and GeoChip based functional gene composition were monitored for 15 weeks. Temporal trends and differences in growth, metabolite profiles, and community composition were observed, largely between Low-Cr and High-Cr bioreactors. In both High-Cr and Low-Cr bioreactors, Cr(VI) levels were below detection from week 1 until week 15. With lactate enrichment, native bacterial diversity substantially decreased as Pelosinus spp., and Sporotalea spp., became the dominant groups, but did not significantly differ between Cr concentrations. The Archaea diversity also substantially decreased after lactate enrichment from Methanosaeta (35%), Methanosarcina (17%) and others, to mostly Methanosarcina spp. (95%). Methane production was lower in High-Cr reactors suggesting some inhibition of methanogens. Several key functional genes were distinct in Low-Cr bioreactors compared to High-Cr. Among the Cr resistant microbes, Burkholderia vietnamiensis, Comamonas testosterone and Ralstonia pickettii proliferated in Cr amended bioreactors. In-situ fermentative conditions facilitated Cr(VI) reduction, and as a result 3.0 mg/L Cr(VI) did not impact the overall bacterial community structure.Yeshttp://www.plosone.org/static/editorial#pee

SCFA lead lab technical assistance at Lawrence Berkeley National Laboratory: Baseline review of three groundwater plumes

During the closeout session, members of the technical assistance team conveyed to the site how impressed they were at the thoroughness of the site's investigation and attempts at remediation. Team members were uniformly pleased at the skilled detection work to identify sources, make quick remediation decisions, and change course when a strategy did not work well. The technical assistance team also noted that, to their knowledge, this is the only DOE site at which a world-class scientist has had primary responsibility for the environmental restoration activities. This has undoubtedly contributed to the successes observed and DOE should take careful note. The following overall recommendations were agreed upon: (1) The site has done a phenomenal job of characterization and identifying and removing source terms. (2) Technologies selected to date are appropriate and high impact, e.g. collection trenches are an effective remedial strategy for this complicated geology. The site should continue using technology that is adapted to the site's unique geology, such as the collection trenches. (3) The site should develop a better way to determine the basis of cleanup for all sites. (4) The sentinel well system should be evaluated and modified, if needed, to assure that the sentinel wells provide coverage to the current site boundary. Potential modifications could include installation, abandonment or relocation of wells based on the large amount of data collected since the original sentinel well system was designed. (5) Modeling to assist in remedial design and communication should continue. (6) The site should develop a plan to ensure institutional memory. (7) The most likely possibility for improving closure to 2006 is by removing the residual source of the Old Town plume and establishing the efficacy of remediation for the 51/64 plume

Low-substrate regulated microaerophilic behavior as a stress response of aquatic and soil bacteria

Low-substrate regulated microaerophilic behavior (LSRMB) was observed in 10–54% of the bacteria isolated from several fresh-water lakes or ponds, subsurface soils, activated sludge, and Antarctic dry valley soils. Five Pseudomonas and two Bacillus type species showed LSRMB. A subsurface Pseudomonas jessenii strain was used as a model to show the metabolic interaction between substrate and oxygen concentrations, cell band movement, and the appearance of unique stress lipids and proteins. When the oxygen in the P. jessenii culture medium was increased from 11% to 100% saturation under atmospheric condition, the concentration of 17:0 cyclopropane fatty acid, a stress indicator, increased five-fold, and four unique proteins were also detected. This stress response occurred only in low-substrate media. It is our hypothesis that LSRMB is a common but under-appreciated trait of many aquatic and soil bacteria

Insights into the structure of mixed CO2/CH4 in gas hydrates

The exchange of carbon dioxide for methane in natural gas hydrates is an attractive approach to harvesting CH4 for energy production while simultaneously sequestering CO2. In addition to the energy and environmental implications, the solid solution of clathrate hydrate (CH4)1–x(CO2)x·5.75H2O provides a model system to study how the distinct bonding and shapes of CH4 and CO2 influence the structure and properties of the compound. High-resolution neutron diffraction was used to examine mixed CO2/CH4 gas hydrates. CO2-rich hydrates had smaller lattice parameters, which were attributed to the higher affinity of the CO2 molecule interacting with H2O molecules that form the surrounding cages, and resulted in a reduction in the unit-cell volume. Experimental nuclear scattering densities illustrate how the cage occupants and energy landscape change with composition. These results provide important insights on the impact and mechanisms for the structure of mixed CH4/CO2 gas hydrate

Isolation and Characterization of Metal-Reducing Thermoanaerobacter Strains from Deep Subsurface Environments of the Piceance Basin, Colorado

Five bacterial strains were isolated from anaerobic enrichment cultures that had originated from inoculations with samples collected from the deep subsurface environments of the millions-of-years-old, geologically and hydrologically isolated Piceance Basin in Colorado. Small-subunit rRNA gene-based analyses indicated that all of these bacteria were closely related to Thermoanaerobacter ethanolicus, with similarities of 99.4 to 99.5%. Three isolates (X513, X514, and X561) from the five bacterial strains were used to examine physiological characteristics. These thermophilic bacteria were able to use acetate, glucose, hydrogen, lactate, pyruvate, succinate, and xylose as electron donors while reducing Fe(III), cobalt(III), chromium(VI), manganese(IV), and uranium(VI) at 60°C. One of the isolates (X514) was also able to utilize hydrogen as an electron donor for Fe(III) reduction. These bacteria exhibited diverse mineral precipitation capabilities, including the formation of magnetite (Fe(3)O(4)), siderite (FeCO(3)), rhodochrosite (MnCO(3)), and uraninite (UO(2)). The gas composition of the incubation headspace and the ionic composition of the incubation medium exerted profound influences on the types of minerals formed. The susceptibility of the thermophilic Fe(III)-reducing cultures to metabolic inhibitors specific for ferric reductase, hydrogenase, and electron transport indicated that iron reduction by these bacteria is an enzymatic process

Temperature-Dependent Oxygen and Carbon Isotopes Fractionations of Biogenic Siderite

Isotopic compositions of biogenic iron minerals may be used to infer environmental conditions under which bacterial iron reduction occurs. The major goal of this study is to examine temperature-dependent isotope fractionations associated with biogenic siderite (FeCO3). Experiments were performed by using both mesophilic (\u3c35°C) and thermophilic (\u3e45°C) iron-reducing bacteria. In addition, control experiments were performed to examine fractionations under nonbiologic conditions. Temperature-dependent oxygen isotope fractionation occurred between biogenic siderite and water from which the mineral was precipitated. Samples in thermophilic cultures (45–75°C) gave the best linear correlation, which can be described as 103 lnαsid-wt = 2.56 × 106 T−2 (K) + 1.69. This empirical equation agrees with that derived from inorganically precipitated siderite by Carothers et al. (1988) and may be used to approximate equilibrium fractionation. Carbon isotope fractionation between biogenic siderite and CO2, based on limited data, also varied with temperature and was consistent with the inorganically precipitated siderite of Carothers et al. (1988). These results indicate that temperature is a controlling factor for isotopic variations in biogenic minerals examined in this study. The temperature-dependent fractionations under laboratory conditions, however, could be complicated by other factors including incubation time and concentration of bicarbonate. Early precipitated siderite at 120-mM initial bicarbonate tended to be enriched in 18O. Siderite formed at \u3c30 mM of bicarbonate tended to be depleted in 18O. Other\u3evariables, such as isotopic compositions of water, types of bacterial species, or bacterial growth rates, had little effect on the fractionation. In addition, siderite formed in abiotic controls had similar oxygen isotopic compositions as those of biogenic siderite at the same temperature, suggesting that microbial fractionations cannot be distinguished from abiotic fractionations under conditions examined here