Despite the clear ecological significance of the microbiomes inhabiting groundwater and connected ecosystems, our current understanding of their habitats, functionality, and the ecological processes controlling their assembly have been limited. In this study, an efficient pipeline combining geochemistry, high-throughput FluidigmTM functional gene amplification and sequencing was developed to analyze the suspended and attached microbial communities inhabiting five groundwater monitoring wells in the Illinois Basin, USA. The dominant taxa in the suspended and the attached microbial communities exhibited significantly different spatial and temporal changes in both alpha- and beta-diversity. Further analyses of representative functional genes affiliated with N2 fixation (nifH), methane oxidation (pmoA), and sulfate reduction (dsrB, and aprA), suggested functional redundancy within the shallow aquifer microbiomes. While more diversified functional gene taxa were observed for the suspended microbial communities than the attached ones except for pmoA, different levels of changes over time and space were observed between these functional genes. Notably, deterministic and stochastic ecological processes shaped the assembly of microbial communities and functional gene reservoirs differently. While homogenous selection was the prevailing process controlling assembly of microbial communities, the neutral processes (e.g., dispersal limitation, drift and others) were more important for the functional genes. The results suggest complex and changing shallow aquifer microbiomes, whose functionality and assembly vary even between the spatially proximate habitats and fractions. This research underscored the importance to include all the interface components for a more holistic understanding of the biogeochemical processes in aquifer ecosystems, which is also instructive for practical applications.

Regulatory uncertainty is a component of carbon storage projects that can significantly affect project resources and timelines. The Illinois Basin – Decatur Project (IBDP) is a one-million tonne, deep-saline CO2 storage project led by the Midwest Geologic Sequestration Consortium (MGSC) that was initiated before the United States carbon storage regulatory framework was fully developed. To address uncertainty in the evolving regulatory environment, a comprehensive, risk-based, monitoring strategy was used that was expected to be over and above the anticipated new regulatory requirements. Ten years after its initiation, the IBDP has successfully completed the pre-injection and injection phases and is currently in the post-injection site care and monitoring phase. The IBDP experienced two lengthy permitting processes that proved to be the rate-limiting factor for the project, increased the length of time before the project could begin injection, and required additional resources to conduct the project. Future projects are likely to continue to have significant amounts of regulatory uncertainty as their respective project designs and monitoring programs will need to be evaluated by regulators on a case-by-case basis. Operators and regulators alike can benefit from frequent proactive communications during the permitting process in order to significantly reduce the length of the permitting process, maintain the critical safeguards of the Class VI rules, and ultimately meet timely goals for reductions to atmospheric CO2 emissions.

This study aimed to identify phenotypic and genotypic resistance profiles of bacteria isolated from soil and manure samples obtained from a farm where tetracycline and tylosin were used extensively. Samples were collected from the manure and soil, before and after manure application. All of the 151 bacteria were identified based on BLAST results. Minimum inhibitory concentrations were determined by agar dilution and E-test methods. Class-specific primers were used to amplify 6 erythromycin and 7 tetracycline resistance (Tc-r) genes for 46 tetracycline-and 18 erythromycin-resistant isolates. Tcr gene sequence identity between different species of isolates was identified to investigate the dissemination of resistance genes. All of the 151 bacteria belonged to 4 different phyla: Proteobacteria, Bacteroidetes, Actinobacteria, and Firmicutes. The majority of the tetracycline-resistant isolates were present in Pseudomonadaceae. Twenty of the isolates, representing 14 different genera, were positive for Tcr genes. Only 2 isolates possessed the erm(Q) gene. This study probably is the first to describe the presence of tetracycline genes in members of the genera Simplicispira and Agrococcus. Results of the study suggest that the diversity of Tcr gene-carrying bacteria are increasing. Furthermore, tetracycline-resistant bacteria could be detected 3 months after the manure application.

Recently, brine samples from the Cambrian-age Mount Simon Formation (the deepest, most inaccessible sedimentary rock formation of the Illinois Basin) and the overlying Ironton-Galesville Formation were collected as part of a major research effort evaluating the feasibility of sequestration of carbon dioxide in deep geologic formations. Halide and halide/cation ratios (especially Cl/Br and Na/Br ratios) from groundwater samples collected during this investigation suggest that the brines of the Cambrian-age strata formed by the evaporation of seawater well beyond the point of halite precipitation. The Cl/Br and Na/Br ratios, the presence of Mississippi-Valley-Type (MVT) ore mineralization in close proximity to the Illinois Basin, and the tectonic history of the region and the Illinois Basin suggest that components of ore-forming brines and perhaps crystalline basement brine are likely still present within the Mount Simon Formation. Halide and cation/halide ratio plots show that these brines have mixed with and have been diluted by subaerially evaporated seawater, seawater and dilute groundwater. Movement of brines out of the Mount Simon Formation and/or exchange with brines of other formations is constrained by the overlying, siltstone-and shale-rich Eau Claire Formation, a low-permeability layer.The most plausible interpretation of the halide and halide/cation ratio data is that the brines of the Cambrian-age strata were introduced to the Illinois Basin from outside of the basin, perhaps when the Illinois Basin was connected to the Arkoma (Oklahoma and Arkansas) and Black Warrior Basins (Alabama and Mississippi) via the Reelfoot Rift during Cambrian and early Ordovician time. In addition, the presence of some percentage of high NaCl, low Cl/Br brines from the crystalline basement is suggested given the geochemical relationships of the halide and cation/halide ratios and the tectonic history of the Illinois Basin. Finally, halide and cation/halide ratios determined by this investigation, and regional geochemical evidence and hydrogeologic modeling (by others) suggest that the brines of these strata probably were affected by regional hydrothermal activity during Permian time that was responsible for the MVT ore deposits of the Midwestern U. S. Thus, the brines of the deepest strata of the Illinois Basin constitute a different, more complex type of fluid than those found elsewhere in the basin. Halide and halide-cation ratios suggest that these deep brines are dominated by residual evaporitic brine (possibly originating as ore-forming brines) with dilution by seawater and dilute groundwater. Other components may include subaerially evaporated seawater and crystalline basement brines. Published by Elsevier Ltd.

The Midwest Geological Sequestration Consortium is conducting the Illinois Basin - Decatur Project (TBDP), a large-scale carbon capture and storage (CCS) project in Decatur, Illinois, USA. An extensive Monitoring, Verification, and Accounting (MVA) program has been implemented, and a deep monitoring well, VW#1, was drilled to a depth of 2,201 in (7,264 ft). The well was perforated in eleven zones ranging in depth from 1,499 to 2,153 m (4917 to 7061 if). Fluid samples were collected during three sampling events using two different sampling methods. This paper describes the well construction, well completion, sample collection procedures, and selected preinjection sampling results from VW#1. Prior to sampling, particular care was taken to remove non-native fluids from each of the sampling zones. Density, specific conductance, bromide concentrations, and bromide-chloride ratios have been used as indicators of sample representativeness.Fluid compositional data and formation pressure data have been collected from VW#1. These high-quality, field-based data are essential to establish hydrochemical conditions in and above the storage reservoir prior to CO2 injection, indicate the response of the reservoir to injection, provide detailed geochemical model inputs, and provide insights for on-going work at the Illinois State Geological Survey about the origin of Illinois Basin brines. Primary brine constituents are chloride, sodium, and calcium with average total dissolved solids concentrations of 190,000 mg/L in the Mt. Simon Sandstone and 65,600 mg/L in Ironton-Galesville Formation (above the injection reservoir). (C) 2013 The Authors. Published by Elsevier Ltd. Open access under CC BY-NC-ND license.

A low-diversity microbial community, dominated by the γ-proteobacterium Halomonas sulfidaeris, was detected in samples of warm saline formation porewater collected from the Cambrian Mt. Simon Sandstone in the Illinois Basin of the North American Midcontinent (1.8 km/5872 ft burial depth, 50°C, pH 8, 181 bars pressure). These highly porous and permeable quartz arenite sandstones are directly analogous to reservoirs around the world targeted for large-scale hydrocarbon extraction, as well as subsurface gas and carbon storage. A new downhole low-contamination subsurface sampling probe was used to collect in situ formation water samples for microbial environmental metagenomic analyses. Multiple lines of evidence suggest that this H. sulfidaeris-dominated subsurface microbial community is indigenous and not derived from drilling mud microbial contamination. Data to support this includes V1-V3 pyrosequencing of formation water and drilling mud, as well as comparison with previously published microbial analyses of drilling muds in other sites. Metabolic pathway reconstruction, constrained by the geology, geochemistry and present-day environmental conditions of the Mt. Simon Sandstone, implies that H. sulfidaeris-dominated subsurface microbial community may utilize iron and nitrogen metabolisms and extensively recycle indigenous nutrients and substrates. The presence of aromatic compound metabolic pathways suggests this microbial community can readily adapt to and survive subsurface hydrocarbon migration.

The use of carbon dioxide (CO2) for enhanced oil recovery (EOR) is being tested for oil fields in the Illinois Basin, USA. While this technology has shown promise for improving oil production, it has raised some issues about the safety of CO2 injection and storage. The Midwest Geological Sequestration Consortium (MGSC) organized a Monitoring, Verification, and Accounting (MVA) team to develop and deploy monitoring programs at three EOR sites in Illinois, Indiana, and Kentucky, USA. MVA goals include establishing baseline conditions to evaluate potential impacts from CO2 injection, demonstrating that project activities are protective of human health and the environment, and providing an accurate accounting of stored CO2. This paper focuses on the use of MVA techniques in monitoring a small CO2 leak from a supply line at an EOR facility under real-world conditions.The ability of shallow monitoring techniques to detect and quantify a CO2 leak under real-world conditions has been largely unproven. In July of 2009, a leak in the pipe supplying pressurized CO2 to an injection well was observed at an MGSC EOR site located in west-central Kentucky. Carbon dioxide was escaping from the supply pipe located approximately 1 m underground. The leak was discovered visually by site personnel and injection was halted immediately. At its largest extent, the hole created by the leak was approximately 1.9 m long by 1.7 m wide and 0.7 m deep in the land surface. This circumstance provided an excellent opportunity to evaluate the performance of several monitoring techniques including soil CO2 flux measurements, portable infrared gas analysis, thermal infrared imagery, and aerial hyperspectral imagery.Valuable experience was gained during this effort. Lessons learned included determining 1) hyperspectral imagery was not effective in detecting this relatively small, short-term CO2 leak, 2) even though injection was halted, the leak remained dynamic and presented a safety risk concern during monitoring activities and, 3) the atmospheric and soil monitoring techniques used were relatively cost-effective, easily and rapidly deployable, and required minimal manpower to set up and maintain for short-term assessments. However, characterization of CO2 distribution near the land surface resulting from a dynamic leak with widely variable concentrations and fluxes was challenging. (C) 2011 Published by Elsevier Ltd.

The Midwest Geological Sequestration Consortium is conducting a large-scale carbon capture and storage (CCS) project in Decatur, Illinois, USA to demonstrate the ability of a deep saline formation to store one million tonnes of carbon dioxide (CO2) from an ethanol facility. Beginning in early 2011, CO2 will be injected at a rate of 1,000 tonnes/day for three years into the Mount Simon Sandstone at a depth of approximately 2,100 meters.An extensive Monitoring, Verification, and Accounting (MVA) program has been undertaken for the Illinois Basin Decatur Project (IBDP) and is focused on the 0.65 km(2) project site. Goals include establishing baseline conditions to evaluate potential impacts from CO2 injection, demonstrating that project activities are protective of human health and the environment, and providing an accurate accounting of stored CO2. MVA efforts are being conducted pre-, during, and post-CO2 injection. Soil and net CO2 flux monitoring has been conducted for more than one year to characterize near-surface CO2 conditions. More than 2,200 soil CO2 flux measurements have been manually collected from a network of 118 soil rings since June 2009. Three ring types have been evaluated to determine which type may be the most effective in detecting potential CO2 leakage. Bare soil, shallow-depth rings were driven 8 cm into the ground and were prepared to minimize surface vegetation in and near the rings. Bare soil, deep-depth rings were prepared similarly, but were driven 46 cm. Natural-vegetation, shallow-depth rings were driven 8 cm and are most representative of typical vegetation conditions. Bare-soil, shallow-depth rings had the smallest observed mean flux (1.78 mu mol m(-2)s(-1)) versus natural-vegetation, shallow-depth rings (3.38 mu mol m(-2)s(-1)). Current data suggest bare ring types would be more sensitive to small CO2 leak signatures than natural ring types because of higher signal to noise ratios.An eddy covariance (EC) system has been in use since June 2009. Baseline data from EC monitoring is being used to characterize pre-injection conditions, and may then be used to detect changes in net exchange CO2 fluxes (F-c) that could be the result of CO2 leakage into the near-surface environment during or following injection. When injection at IBDP begins, soil and net CO2 monitoring efforts will have established a baseline of near-surface conditions that will be important to help demonstrate the effectiveness of storage activities. (C) 2011 Published by Elsevier Ltd.

The development of the Illinois Basin-Decatur USA test site for a 1 million tonne injection of CO2 into the Mount Simon Sandstone saline reservoir beginning in 2011 has been a multiphase process requiring a wide array of personnel and resources that began in 2003. The process of regional characterization took two years as part of a Phase I effort focused on the entire Illinois Basin, located in Illinois, Indiana, and Kentucky, USA. Seeking the cooperation of an industrial source of CO2 and site selection within the Basin took place during Phase II while most of the concurrent research emphasis was on a set of small-scale tests of Enhanced Oil Recovery (EOR) and CO2 injection into a coal seam. Phase III began the commitment to the 1 million-tonne test site development through the collaboration of the Archer Daniels Midland Company (ADM) who is providing a site, the CO2, and developing a compression facility, of Schlumberger Carbon Services who is providing expertise for operations, drilling, geophysics, risk assessment, and reservoir modelling, and of the Illinois State Geological Survey (ISGS) whose geologists and engineers lead the Midwest Geological Sequestration Consortium (MGSC). Communications and outreach has been a collaborative effort of ADM, ISGS and Schlumberger Carbon Services. The Consortium is one of the seven Regional Carbon Sequestration Partnerships, a carbon sequestration research program supported by the National Energy Technology Laboratory of the U.S. Department of Energy. (C) 2011 Published by Elsevier Ltd.

RNA methylase genes are common antibiotic resistance determinants for multiple drugs of the macrolide, lincosamide, and streptogramin B (MLSB) families. We used molecular methods to investigate the diversity, distribution, and abundance of MLSB methylases in waste lagoons and groundwater wells at two swine farms with a history of tylosin (a macrolide antibiotic structurally related to erythromycin) and tetracycline usage. Phylogenetic analysis guided primer design for quantification of MLSB resistance genes found in tylosin-producing Streptomyces (tlr(B), tlr(D)) and commensal/pathogenic bacteria (erm(A), erm(B), erm(C), erm(F), erm(G), erm(Q)). The near absence of tlr genes at these sites suggested a lack of native antibiotic-producing organisms. The gene combination erm(ABCF) was found in all lagoon samples analyzed. These four genes were also detected with high frequency in wells previously found to be contaminated by lagoon leakage. A weak correlation was found between the distribution of erm genes and previously reported patterns of tetracycline resistance determinants, suggesting that dissemination of these genes into the environment is not necessarily linked. Considerations of gene origins in history (i.e., phylogeny) and gene distributions in the landscape provide a useful "molecular ecology" framework for studying environmental spread of antibiotic resistance.

Public perception of carbon capture and sequestration (CCS) projects represents a potential barrier to commercialization. Outreach to stakeholders at the local, regional, and national level is needed to create familiarity with and potential acceptance of CCS projects. This paper highlights the Midwest Geological Sequestration Consortium (MGSC) multi-level outreach approach which interacts with multiple stakeholders. The MGSC approach focuses on external and internal communication. External communication has resulted in building regional public understanding of CCS. Internal communication, through a project Risk Assessment process, has resulted in enhanced team communication and preparation of team members for outreach roles.

For the Illinois Basin - Decatur Project, we evaluated risks to project values that include safety, research, and public acceptance as well as successfully storing CO2. Starting with a FEP list, experts in six working groups identified risks and characterized the Likelihood and Severity of negative impact. Evaluations were made both by group consensus and by later independent scoring.Results are being used to rationalize and shape risk- reduction measures, especially those involving well engineering and subsurface characterization. Risk evaluation influences the plans for monitoring and external communications, and will inform the construction and quantitative attribution of flow simulations and system models. (C) 2009 Elsevier Ltd. All rights reserved.

Antibiotics are used in animal livestock production for therapeutic treatment of disease and at subtherapeutic levels for growth promotion and improvement of feed efficiency. It is estimated that approximately 75% of antibiotics are not absorbed by animals and are excreted in waste. Antibiotic resistance selection occurs among gastrointestinal bacteria, which are also excreted in manure and stored in waste holding systems. Land application of animal waste is a common disposal method used in the United States and is a means for environmental entry of both antibiotics and genetic resistance determinants. Concerns for bacterial resistance gene selection and dissemination of resistance genes have prompted interest about the concentrations and biological activity of drug residues and break-down metabolites, and their fate and transport. Fecal bacteria can survive for weeks to months in the environment, depending on species and temperature, however, genetic elements can persist regardless of cell viability. Phylogenetic analyses indicate antibiotic resistance genes have evolved, although some genes have been maintained in bacteria before the modern antibiotic era. Quantitative measurements of drug residues and levels of resistance genes are needed, in addition to understanding the environmental mechanisms of genetic selection, gene acquisition, and the spatiotemporal dynamics of these resistance genes and their bacterial hosts. This review article discusses an accumulation of findings that address aspects of the fate, transport, and persistence of antibiotics and antibiotic resistance genes in natural environments, with emphasis on mechanisms pertaining to soil environments following land application of animal waste effluent.