Differential protein expression during growth on model and commercial mixtures of naphthenic acids in Pseudomonas fluorescens Pf‐5

Naphthenic acids (NAs) are carboxylic acids with the formula (CnH2n+ZO2) and are among the most toxic, persistent constituents of oil sands process-affected waters (OSPW), produced during oil sands extraction. Currently, the proteins and mechanisms involved in NA biodegradation are unknown. Using LC-MS/MS shotgun proteomics, we identified proteins overexpressed during the growth of Pseudomonas fluorescens Pf-5 on a model NA (4′-n-butylphenyl)-4-butanoic acid (n-BPBA) and commercial NA mixture (Acros). By day 11, >95% of n-BPBA was degraded. With Acros, a 17% reduction in intensity occurred with 10–18 carbon compounds of the Z family −2 to −14 (major NA species in this mixture). A total of 554 proteins (n-BPBA) and 631 proteins (Acros) were overexpressed during growth on NAs, including several transporters (e.g., ABC transporters), suggesting a cellular protective response from NA toxicity. Several proteins associated with fatty acid, lipid, and amino acid metabolism were also overexpressed, including acyl-CoA dehydrogenase and acyl-CoA thioesterase II, which catalyze part of the fatty acid beta-oxidation pathway. Indeed, multiple enzymes involved in the fatty acid oxidation pathway were upregulated. Given the presumed structural similarity between alkyl-carboxylic acid side chains and fatty acids, we postulate that P. fluorescens Pf-5 was using existing fatty acid catabolic pathways (among others) during NA degradation

Metagenomic Analyses Reveal That Energy Transfer Gene Abundances Can Predict the Syntrophic Potential of Environmental Microbial Communities

Hydrocarbon compounds can be biodegraded by anaerobic microorganisms to form methane through an energetically interdependent metabolic process known as syntrophy. The microorganisms that perform this process as well as the energy transfer mechanisms involved are difficult to study and thus are still poorly understood, especially on an environmental scale. Here, metagenomic data was analyzed for specific clusters of orthologous groups (COGs) related to key energy transfer genes thus far identified in syntrophic bacteria, and principal component analysis was used in order to determine whether potentially syntrophic environments could be distinguished using these syntroph related COGs as opposed to universally present COGs. We found that COGs related to hydrogenase and formate dehydrogenase genes were able to distinguish known syntrophic consortia and environments with the potential for syntrophy from non-syntrophic environments, indicating that these COGs could be used as a tool to identify syntrophic hydrocarbon biodegrading environments using metagenomic data

In situ detection of an aerobic alkane metabolites in subsurface environments

Article deposited according to the Frontiers publishing policy for [Frontiers in Microbiological Chemistry]: http://www.frontiersin.org/about/faq, [June 10, 2013].YesFunding provided by the Open Access Authors Fund

Assessing Microbial Corrosion Risk on Offshore Crude Oil Production Topsides under Conditions of Nitrate and Nitrite Treatment for Souring

Oilfield souring is a detrimental effect caused by sulfate-reducing microorganisms that reduce sulfate to sulfide during their respiration process. Nitrate or nitrite can be used to mitigate souring, but may also impart a corrosion risk. Produced fluids sampled from the topside infrastructure of two floating, production, storage, and offloading (FPSO) vessels (Platform A and Platform B) were assessed for microbial corrosion under nitrate and nitrite breakthrough conditions using microcosm tests incubated at 54 °C. Microbial community compositions on each individual FPSO were similar, while those between the two FPSO vessels differed. Platform B microbial communities responded as expected to nitrate breakthrough conditions, where nitrate-reducing activity was enhanced and sulfate reduction was inhibited. In contrast, nitrate treatments of Platform A microbial communities were not as effective in preventing sulfide production. Nitrite breakthrough conditions had the strongest sulfate reduction inhibition in samples from both platforms, but exhibited the highest pitting density. Live experimental replicates with no nitrate or nitrite additive yielded the highest general corrosion rates in the study (up to 0.48 mm/year), while nitrate- or nitrite-treated fluids revealed general corrosion rates that are considered low or moderate (<0.12 mm/year). Overall, the results of this study provide a description of nitrogen- and sulfur-based microbial activities under thermophilic conditions, and their risk for MIC that can occur along fluid processing lines on FPSO topsides that process fluids during offshore oil production operations

<title>Weapons team engagement trainer: a transfer of high-tech military training technology to the law enforcement community</title>

Six years ago at SPIE, a team of government researchers and engineers unveiled a new, military, Weapons Team Engagement Trainer (WTET).\u27 At that time, potential applications of this prototype military training device to civilian law enforcement training were realized. Subsequent action was taken under the Federal Technology Transfer Act of 1986,2 enabling the transfer of WTET to the private sector, through a cooperative agreement between: The Office of Naval Research (ONR), NAWCTSD, and the commercial weapons training organization Firearms Training Systems, Inc. (FATS). Planning also began for release of a commercial WTET system. The government research and development facility and the National Institute of Justice (NIJ) formed a cooperative agreement to make the prototype system available to military, federal, and local law enforcement agencies for use in Orlando, Florida-until a commercial version could become available. This cooperative effort has provided evidence of the effectiveness and realism of WTET with law enforcement personnel. This paper offers, a technical description of the improvements made to WTET, a brief explanation of the commercialization process, a suminaiy of the evaluations conducted to date, and insight into how that information has been used in the development of the commercial version

Combined Use of Diagnostic Fumarate Addition Metabolites and Genes Provides Evidence for Anaerobic Hydrocarbon Biodegradation in Contaminated Groundwater

The widespread use of hydrocarbon-based fuels has led to the contamination of many natural environments due to accidental spills or leaks. While anaerobic microorganisms indigenous to many fuel-contaminated groundwater sites can play a role in site remediation (e.g., monitored natural attenuation, MNA) via hydrocarbon biodegradation, multiple lines of evidence in support of such bioremediation are required. In this study, we investigated two fuel-contaminated groundwater sites for their potential to be managed by MNA. Microbial community composition, biogeochemical indicators, fumarate addition metabolites, and genes diagnostic of both alkane and alkyl-monoaromatic hydrocarbon activation were assessed. Fumarate addition metabolites and catabolic genes were detected for both classes of hydrocarbon biodegradation at both sites, providing strong evidence for in situ anaerobic hydrocarbon biodegradation. However, relevant metabolites and genes did not consistently co-occur within all groundwater samples. Using newly designed mixtures of quantitative polymerase chain reaction (qPCR) primers to target diverse assA and bssA genes, we measured assA gene abundances ranging from 105&ndash;108 copies/L, and bssA gene abundances ranging from 105&ndash;1010 copies/L at the sites. Overall, this study demonstrates the value of investigating fuel-contaminated sites using both metabolites and genes diagnostic of anaerobic hydrocarbon biodegradation for different classes of hydrocarbons to help assess field sites for management by MNA

Time Course-Dependent Methanogenic Crude Oil Biodegradation: Dynamics of Fumarate Addition Metabolites, Biodegradative Genes, and Microbial Community Composition

Biodegradation of crude oil in subsurface petroleum reservoirs has adversely impacted most of the world's oil, converting this resource to heavier forms that are of lower quality and more challenging to recover. Oil degradation in deep reservoir environments has been attributed to methanogenesis over geological time, yet our understanding of the processes and organisms mediating oil transformation in the absence of electron acceptors remains incomplete. Here, we sought to identify hydrocarbon activation mechanisms and reservoir-associated microorganisms that may have helped shape the formation of biodegraded oil by incubating oilfield produced water in the presence of light (°API = 32) or heavy crude oil (°API = 16). Over the course of 17 months, we conducted routine analytical (GC, GC-MS) and molecular (PCR/qPCR of assA and bssA genes, 16S rRNA gene sequencing) surveys to assess microbial community composition and activity changes over time. Over the incubation period, we detected the formation of transient hydrocarbon metabolites indicative of alkane and alkylbenzene addition to fumarate, corresponding with increases in methane production and fumarate addition gene abundance. Chemical and gene-based evidence of hydrocarbon biodegradation under methanogenic conditions was supported by the enrichment of hydrocarbon fermenters known to catalyze fumarate addition reactions (e.g., Desulfotomaculum, Smithella), along with syntrophic bacteria (Syntrophus), methanogenic archaea, and several candidate phyla (e.g., “Atribacteria”, “Cloacimonetes”). Our results reveal that fumarate addition is a possible mechanism for catalyzing the methanogenic biodegradation of susceptible saturates and aromatic hydrocarbons in crude oil, and we propose the roles of community members and candidate phyla in our cultures that may be involved in hydrocarbon transformation to methane in crude oil systems