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Item Genomic Features and Pervasive Negative Selection in Rhodanobacter Strains Isolated from Nitrate and Heavy Metal Contaminated Aquifer(American Society for Microbiology, 2022-02) Peng, Mu; Wang, Dongyu; Lui, Lauren M.; Nielsen, Torben; Tian, Renmao; Kempher, Megan L.; Tao, Xuanyu; Pan, Chongle; Chakraborty, Romy; Deutschbauer, Adam M.; Thorgersen, Michael P.; Adams, Michael W. W.; Fields, Matthew W.; Hazen, Terry C.; Arkin, Adam P.; Zhou, Aifen; Zhou, JizhongDespite the dominance of Rhodanobacter species in the subsurface of the contaminated Oak Ridge Reservation (ORR) site, very little is known about the mechanisms underlying their adaptions to the various stressors present at ORR. Recently, multiple Rhodanobacter strains have been isolated from the ORR groundwater samples from several wells with varying geochemical properties.Item Characterization of subsurface media from locations up- and down-gradient of a uranium-contaminated aquifer(Elsevier BV, 2020-05) Moon, Ji-Won; Paradis, Charles J.; Joyner, Dominique C.; von Netzer, Frederick; Majumder, Erica L.; Dixon, Emma R.; Podar, Mircea; Ge, Xiaoxuan; Walian, Peter J.; Smith, Heidi J.; Wu, Xiaoqin; Zane, Grant M.; Walker, Kathleen F.; Thorgersen, Michael P.; Poole, Farris L. II; Lui, Lauren M.; Adams, Benjamin G.; De León, Kara B.; Brewer, Sheridan S.; Williams, Daniel E.; Lowe, Kenneth A.; Rodriguez, Miguel; Mehlhorn, Tonia L.; Pfiffner, Susan M.; Chakraborty, Romy; Arkin, Adam P.; Wall, Judy D.; Fields, Matthew W.; Adams, Michael W.W.; Stahl, David A.; Elias, Dwayne A.; Hazen, Terry C.The processing of sediment to accurately characterize the spatially-resolved depth profiles of geophysical and geochemical properties along with signatures of microbial density and activity remains a challenge especially in complex contaminated areas. This study processed cores from two sediment boreholes from background and contaminated core sediments and surrounding groundwater. Fresh core sediments were compared by depth to capture the changes in sediment structure, sediment minerals, biomass, and pore water geochemistry in terms of major and trace elements including pollutants, cations, anions, and organic acids. Soil porewater samples were matched to groundwater level, flow rate, and preferential flows and compared to homogenized groundwater-only samples from neighboring monitoring wells. Groundwater analysis of nearby wells only revealed high sulfate and nitrate concentrations while the same analysis using sediment pore water samples with depth was able to suggest areas high in sulfate-and nitrate-reducing bacteria based on their decreased concentration and production of reduced by-products that could not be seen in the groundwater samples. Positive correlations among porewater content, total organic carbon, trace metals and clay minerals revealed a more complicated relationship among contaminant, sediment texture, groundwater table, and biomass. The fluctuating capillary interface had high concentrations of Fe and Mn-oxides combined with trace elements including U, Th, Sr, Ba, Cu, and Co. This suggests the mobility of potentially hazardous elements, sediment structure, and biogeochemical factors are all linked together to impact microbial communities, emphasizing that solid interfaces play an important role in determining the abundance of bacteria in the sediments.Item Mechanism Across Scales: A Holistic Modeling Framework Integrating Laboratory and Field Studies for Microbial Ecology(Frontiers Media SA, 2021-03) Lui, Lauren M.; Majumder, Erica L.-W.; Smith, Heidi J.; Carlson, Hans K.; von Netzer, Frederick; Fields, Matthew W.; Stahl, David A.; Zhou, Jizhong; Hazen, Terry C.; Baliga, Nitin S.; Adams, Paul D.; Arkin, Adam P.Over the last century, leaps in technology for imaging, sampling, detection, high-throughput sequencing, and -omics analyses have revolutionized microbial ecology to enable rapid acquisition of extensive datasets for microbial communities across the ever-increasing temporal and spatial scales. The present challenge is capitalizing on our enhanced abilities of observation and integrating diverse data types from different scales, resolutions, and disciplines to reach a causal and mechanistic understanding of how microbial communities transform and respond to perturbations in the environment. This type of causal and mechanistic understanding will make predictions of microbial community behavior more robust and actionable in addressing microbially mediated global problems. To discern drivers of microbial community assembly and function, we recognize the need for a conceptual, quantitative framework that connects measurements of genomic potential, the environment, and ecological and physical forces to rates of microbial growth at specific locations. We describe the Framework for Integrated, Conceptual, and Systematic Microbial Ecology (FICSME), an experimental design framework for conducting process-focused microbial ecology studies that incorporates biological, chemical, and physical drivers of a microbial system into a conceptual model. Through iterative cycles that advance our understanding of the coupling across scales and processes, we can reliably predict how perturbations to microbial systems impact ecosystem-scale processes or vice versa. We describe an approach and potential applications for using the FICSME to elucidate the mechanisms of globally important ecological and physical processes, toward attaining the goal of predicting the structure and function of microbial communities in chemically complex natural environments.Item Impact of hydrologic boundaries on microbial planktonic and biofilm communities in shallow terrestrial subsurface environments(2018-09) Smith, Heidi J.; Zelaya, Anna J.; De León, Kara B.; Chakraborty, R.; Elias, Dwayne A.; Hazen, Terry C.; Arkin, Adam P.; Cunningham, Alfred B.; Fields, Matthew W.Subsurface environments contain a large proportion of planetary microbial biomass and harbor diverse communities responsible for mediating biogeochemical cycles important to groundwater used by human society for consumption, irrigation, agriculture and industry. Within the saturated zone, capillary fringe and vadose zones, microorganisms can reside in two distinct phases (planktonic or biofilm), and significant differences in community composition, structure and activity between free-living and attached communities are commonly accepted. However, largely due to sampling constraints and the challenges of working with solid substrata, the contribution of each phase to subsurface processes is largely unresolved. Here, we synthesize current information on the diversity and activity of shallow freshwater subsurface habitats, discuss the challenges associated with sampling planktonic and biofilm communities across spatial, temporal and geological gradients, and discuss how biofilms may be constrained within shallow terrestrial subsurface aquifers. We suggest that merging traditional activity measurements and sequencing/-omics technologies with hydrological parameters important to sediment biofilm assembly and stability will help delineate key system parameters. Ultimately, integration will enhance our understanding of shallow subsurface ecophysiology in terms of bulk-flow through porous media and distinguish the respective activities of sessile microbial communities from more transient planktonic communities to ecosystem service and maintenance.Item Microbial Functional Gene Diversity Predicts Groundwater Contamination and Ecosystem Functioning(2018-02) He, Zhili; Zhang, Ping; Wu, Linwei; Rocha, Andrea M.; Tu, Qichao; Shi, Zhou; Wu, Bo; Qin, Yujia; Wang, Jianjun; Yan, Qingyun; Curtis, Daniel; Ning, Daliang; Van Nostrand, Joy D.; Wu, Liyou; Yang, Yunfeng; Elias, Dwayne A.; Watson, David B.; Adams, Michael W. W.; Fields, Matthew W.; Alm, Eric J.; Hazen, Terry C.; Adams, Paul D.; Arkin, Adam P.; Zhou, JizhongContamination from anthropogenic activities has significantly impacted Earth\'s biosphere. However, knowledge about how environmental contamination affects the biodiversity of groundwater microbiomes and ecosystem functioning remains very limited. Here, we used a comprehensive functional gene array to analyze groundwater microbiomes from 69 wells at the Oak Ridge Field Research Center (Oak Ridge, TN), representing a wide pH range and uranium, nitrate, and other contaminants. We hypothesized that the functional diversity of groundwater microbiomes would decrease as environmental contamination (e.g., uranium or nitrate) increased or at low or high pH, while some specific populations capable of utilizing or resistant to those contaminants would increase, and thus, such key microbial functional genes and/or populations could be used to predict groundwater contamination and ecosystem functioning. Our results indicated that functional richness/diversity decreased as uranium (but not nitrate) increased in groundwater. In addition, about 5.9% of specific key functional populations targeted by a comprehensive functional gene array (GeoChip 5) increased significantly (P < 0.05) as uranium or nitrate increased, and their changes could be used to successfully predict uranium and nitrate contamination and ecosystem functioning. This study indicates great potential for using microbial functional genes to predict environmental contamination and ecosystem functioning.IMPORTANCE Disentangling the relationships between biodiversity and ecosystem functioning is an important but poorly understood topic in ecology. Predicting ecosystem functioning on the basis of biodiversity is even more difficult, particularly with microbial biomarkers. As an exploratory effort, this study used key microbial functional genes as biomarkers to provide predictive understanding of environmental contamination and ecosystem functioning. The results indicated that the overall functional gene richness/diversity decreased as uranium increased in groundwater, while specific key microbial guilds increased significantly as uranium or nitrate increased. These key microbial functional genes could be used to successfully predict environmental contamination and ecosystem functioning. This study represents a significant advance in using functional gene markers to predict the spatial distribution of environmental contaminants and ecosystem functioning toward predictive microbial ecology, which is an ultimate goal of microbial ecology.Item Unintended Laboratory-Driven Evolution Reveals Genetic Requirements for Biofilm Formation by Desulfovibrio vulgaris Hildenborough.(2017-10) De Leon, K. B.; Zane, Grant M.; Trotter, V. V.; Krantz, Gregory; Arkin, Adam P.; Butland, G. P.; Walian, P. J.; Fields, Matthew W.; Wall, Judy D.Biofilms of sulfate-reducing bacteria (SRB) are of particular interest as members of this group are culprits in corrosion of industrial metal and concrete pipelines as well as being key players in subsurface metal cycling. Yet the mechanism of biofilm formation by these bacteria has not been determined. Here we show that two supposedly identical wild-type cultures of the SRB Desulfovibrio vulgaris Hildenborough maintained in different laboratories have diverged in biofilm formation. From genome resequencing and subsequent mutant analyses, we discovered that a single nucleotide change within DVU1017, the ABC transporter of a type I secretion system (T1SS), was sufficient to eliminate biofilm formation in D. vulgaris Hildenborough. Two T1SS cargo proteins were identified as likely biofilm structural proteins, and the presence of at least one (with either being sufficient) was shown to be required for biofilm formation. Antibodies specific to these biofilm structural proteins confirmed that DVU1017, and thus the T1SS, is essential for localization of these adhesion proteins on the cell surface. We propose that DVU1017 is a member of the lapB category of microbial surface proteins because of its phenotypic similarity to the adhesin export system described for biofilm formation in the environmental pseudomonads. These findings have led to the identification of two functions required for biofilm formation in D. vulgaris Hildenborough and focus attention on the importance of monitoring laboratory-driven evolution, as phenotypes as fundamental as biofilm formation can be altered.Item Expression profiling of hypothetical genes in Desulfovibrio vulgaris leads to improved functional annotation(2009-03) Elias, Dwayne A.; Mukhopadhyay, A.; Joachimiak, M. P.; Drury, Elliott C.; Redding, Alyssa M.; Yen, Huei-Che B.; Fields, Matthew W.; Hazen, Terry C.; Arkin, Adam P.; Keasling, J. D.; Wall, Judy D.Hypothetical (HyP) and conserved HyP genes account for >30% of sequenced bacterial genomes. For the sulfate-reducing bacterium Desulfovibrio vulgaris Hildenborough, 347 of the 3634 genes were annotated as conserved HyP (9.5%) along with 887 HyP genes (24.4%). Given the large fraction of the genome, it is plausible that some of these genes serve critical cellular roles. The study goals were to determine which genes were expressed and provide a more functionally based annotation. To accomplish this, expression profiles of 1234 HyP and conserved genes were used from transcriptomic datasets of 11 environmental stresses, complemented with shotgun LC–MS/MS and AMT tag proteomic data. Genes were divided into putatively polycistronic operons and those predicted to be monocistronic, then classified by basal expression levels and grouped according to changes in expression for one or multiple stresses. One thousand two hundred and twelve of these genes were transcribed with 786 producing detectable proteins. There was no evidence for expression of 17 predicted genes. Except for the latter, monocistronic gene annotation was expanded using the above criteria along with matching Clusters of Orthologous Groups. Polycistronic genes were annotated in the same manner with inferences from their proximity to more confidently annotated genes. Two targeted deletion mutants were used as test cases to determine the relevance of the inferred functional annotations.Item Dynamic Succession of Groundwater Sulfate-Reducing Communities during Prolonged Reduction of Uranium in a Contaminated Aquifer(2017-04) Zhang, Ping; He, Zhili; Van Nostrand, Joy D.; Qin, Yujia; Deng, Ye; Wu, Liyou; Tu, Qichao; Wang, Jianjun; Schadt, Christopher W.; Fields, Matthew W.; Hazen, Terry C.; Arkin, Adam P.; Stahl, David A.; Zhou, JizhongTo further understand the diversity and dynamics of SRB in response to substrate amendment, we sequenced genes coding for the dissimilatory sulfite reductase (dsrA) in groundwater samples collected after an emulsified vegetable oil (EVO) amendment, which sustained U(VI)-reducing conditions for one year in a fast-flowing aquifer. EVO amendment significantly altered the composition of groundwater SRB communities. Sequences having no closely related-described species dominated (80%) the indigenous SRB communities in nonamended wells. After EVO amendment, Desulfococcus, Desulfobacterium, and Desulfovibrio, known for long-chain-fatty-acid, short-chain-fatty-acid and H2 oxidation and U(VI) reduction, became dominant accounting for 7 ± 2%, 21 ± 8%, and 55 ± 8% of the SRB communities, respectively. Succession of these SRB at different bioactivity stages based on redox substrates/products (acetate, SO4–2, U(VI), NO3–, Fe(II), and Mn(II)) was observed. Desulfovibrio and Desulfococcus dominated SRB communities at 4–31 days, whereas Desulfobacterium became dominant at 80–140 days. By the end of the experiment (day 269), the abundance of these SRB decreased but the overall diversity of groundwater SRB was still higher than non-EVO controls. Up to 62% of the SRB community changes could be explained by groundwater geochemical variables, including those redox substrates/products. A significant (P < 0.001) correlation was observed between groundwater U(VI) concentrations and Desulfovibrio abundance. Our results showed that the members of SRB and their dynamics were correlated significantly with slow EVO biodegradation, electron donor production and maintenance of U(VI)-reducing conditions in the aquifer.Item Global transcriptional, physiological, and metabolite analyses of the responses of Desulfovibrio vulgaris Hildenborough to salt adaptation(2009-12) He, Zhili; Zhou, Aifen; Baidoo, Edward E. K.; He, Q.; Joachimiak, M. P.; Benke, P.; Phan, R.; Mukhopadhyay, A.; Hemme, C. L.; Huang, K.; Alm, E. J.; Fields, Matthew W.; Wall, Judy D.; Stahl, David A.; Hazen, Terry C.; Keasling, J. D.; Arkin, Adam P.; Zhou, JizhongThe response of Desulfovibrio vulgaris Hildenborough to salt adaptation (long-term NaCl exposure) was examined by performing physiological, global transcriptional, and metabolite analyses. Salt adaptation was reflected by increased expression of genes involved in amino acid biosynthesis and transport, electron transfer, hydrogen oxidation, and general stress responses (e.g., heat shock proteins, phage shock proteins, and oxidative stress response proteins). The expression of genes involved in carbon metabolism, cell growth, and phage structures was decreased. Transcriptome profiles of D. vulgaris responses to salt adaptation were compared with transcriptome profiles of D. vulgaris responses to salt shock (short-term NaCl exposure). Metabolite assays showed that glutamate and alanine accumulated under salt adaptation conditions, suggesting that these amino acids may be used as osmoprotectants in D. vulgaris. Addition of amino acids (glutamate, alanine, and tryptophan) or yeast extract to the growth medium relieved salt-related growth inhibition. A conceptual model that links the observed results to currently available knowledge is proposed to increase our understanding of the mechanisms of D. vulgaris adaptation to elevated NaCl levels.Item Impact of elevated nitrate on sulfate-reducing bacteria: A comparative study of Desulfovibrio vulgaris(2010-05) He, Q.; He, Zhili; Joyner, D. C.; Joachimiak, M. P.; Price, M. N.; Yang, Zamin K.; Yen, Huei-Che B.; Hemme, C. L.; Chen, W.; Fields, Matthew W.; Stahl, David A.; Keasling, J. D.; Keller, M.; Arkin, Adam P.; Hazen, Terry C.; Wall, Judy D.; Zhou, JizhongSulfate-reducing bacteria have been extensively studied for their potential in heavy-metal bioremediation. However, the occurrence of elevated nitrate in contaminated environments has been shown to inhibit sulfate reduction activity. Although the inhibition has been suggested to result from the competition with nitrate-reducing bacteria, the possibility of direct inhibition of sulfate reducers by elevated nitrate needs to be explored. Using Desulfovibrio vulgaris as a model sulfate-reducing bacterium, functional genomics analysis reveals that osmotic stress contributed to growth inhibition by nitrate as shown by the upregulation of the glycine/betaine transporter genes and the relief of nitrate inhibition by osmoprotectants. The observation that significant growth inhibition was effected by 70mM NaNO3 but not by 70mM NaCl suggests the presence of inhibitory mechanisms in addition to osmotic stress. The differential expression of genes characteristic of nitrite stress responses, such as the hybrid cluster protein gene, under nitrate stress condition further indicates that nitrate stress response by D. vulgaris was linked to components of both osmotic and nitrite stress responses. The involvement of the oxidative stress response pathway, however, might be the result of a more general stress response. Given the low similarities between the response profiles to nitrate and other stresses, less-defined stress response pathways could also be important in nitrate stress, which might involve the shift in energy metabolism. The involvement of nitrite stress response upon exposure to nitrate may provide detoxification mechanisms for nitrite, which is inhibitory to sulfate-reducing bacteria, produced by microbial nitrate reduction as a metabolic intermediate and may enhance the survival of sulfate-reducing bacteria in environments with elevated nitrate level.