Ecological Interactions Between Metals and Microbes

Analyses of chromium resistant microbes. Culturable xylene-degrading and chromate-resistant microbes were obtained from chronically cocontaminated soil using a microcosm enrichment technique, and shown to correlate to dominant soil populations using culture independent techniques. The soil microbial community proved able to mount a respiratory response to addition of xylene in the presence of chromate. The majority of isolates belonged to the ubiquitous but poorly studied high %G+C Gram positive genus Arthrobacter, and exhibited considerable genotypic and phenotypic variability. Phenotypic assays uncovered a wide variation in the levels of chromate resistance, even between very closely related strains. Primers designed against conserved motifs in the known chrA chromate efflux gene failed to detect similar sequences among the chromate resistant Arthrobacter isolates obtained through enrichment

Ecological Interactions Between Metals and Microbes That Impact Bioremediation

Previous work showed the correlation between bacterial biomass, population structure and the amount of lead, chromium and aromatic compounds present along a 21.6 m transect in which the concentrations of both heavy metals (Pb and Cr) and aromatic compounds varied 2-3 orders of magnitude. This work suggested that (a) biomass level was better correlated to the level of biodegradable organic C than the level of heavy metals, (b) microbial community composition differed between highly contaminated soils and uncontaminated ones, and (c) substantial microbial activity was found even in the highly contaminated soils. One confounding factor in these analyses was that the contaminated soils contained Pb, Cr, and aromatic hydrocarbons. Therefore, it was difficult to determine which factors were most important in the shifts of microbial community composition. Therefore, experiments were conducted in microcosms in which individual factors could be systematically varied. In this case, soils were used from the Seymour, IN site which had low levels of contamination, and the microbial community had little chance to adapt to heavy metals or aromatic compounds

Ecological Interactions Between Metals and Microbes That Impact Bioremediation

Bacterial Community Diversity at a Mixed Waste Contaminated Site The correlation between bacterial population structure and lead, chromium and organic compounds present along a 21.6 m transect was examined. There was a gradient of heavy metal (Cr and Pb) and petroleum hydrocarbon contamination in these soils. A 16S rDNA analysis method and fatty acid methyl esters derived from phospholipids (PLFA) analysis were used to compare microbial communities. Soil microbial DNA was extracted and community fingerprint patterns for each sample location were produced by DGGE separation of the V3 region of the 16S rRNA genes amplified by PCR. Visual analysis of DGGE patterns indicated that sample locations with high concentrations of total toluene (12,000 mg kg-1), xylenes (8,000 mg kg-1), methylene chloride (10,000 mg kg-1), lead (17,000 mg kg-1) and chromium (3,200 mg kg-1) have a different community composition from the community with lower metals (200 mg kg-1) and organics (1200 mg kg-1) content. Microbial biomass, indicated by total phospholipid-P, was greatest in soils with highest organic contamination

Ecological Interactions Between Metals and Microbes That Impact Bioremediation

Samples have been obtained from (a) soil highly contaminated with Cr (tannery site) and (b) soils contaminated with petroleum, Cr, and Pb (Seymour, IN). Microcosm experiments with the tannery site soil indicated that microbial biomass (assayed as phospholipid-phosphate) and activity (assayed as carbon dioxide evolution) were primarily determined by organic carbon availability, but not total Cr concentration. The toxicity of metals to the indigenous microbial populations of the Seymour soils was determined by measuring microbial activity (incorporation of tritiated leucine into protein) of cells extracted from soil particles in solutions of increasing metal concentration. Although total Cr concentration varied 100-fold in these soils, the inhibition constant for Cr toxicity varied < 3-fold. Of additional interest in one soil was the dose-response function; the response suggests the soil contains a complex mixture of microbes with different Cr resistance levels. Cr and Pb resistant bacteria have been isolated from these soil samples. In Arthrobacter sp. Cr15, Cr resistance was spontaneously lost at a frequency of ca. 0.5% after growth for 20 generations in non-selective medium. The wild-type contained a 60 kb plasmid. In two Cr sensitive strains, restriction fragment analysis has shown that 15 kb of the plasmid have been lost. Matings between the wild type and cured strains result in transfer of the Cr resistance phenotype at a frequency of 1%

High-level chromate resistance in Arthrobacter sp. strain FB24 requires previously uncharacterized accessory genes

<p>Abstract</p> <p>Background</p> <p>The genome of <it>Arthrobacter </it>sp. strain FB24 contains a chromate resistance determinant (CRD), consisting of a cluster of 8 genes located on a 10.6 kb fragment of a 96 kb plasmid. The CRD includes <it>chrA</it>, which encodes a putative chromate efflux protein, and three genes with amino acid similarities to the amino and carboxy termini of ChrB, a putative regulatory protein. There are also three novel genes that have not been previously associated with chromate resistance in other bacteria; they encode an oxidoreductase (most similar to malate:quinone oxidoreductase), a functionally unknown protein with a WD40 repeat domain and a lipoprotein. To delineate the contribution of the CRD genes to the FB24 chromate [Cr(VI)] response, we evaluated the growth of mutant strains bearing regions of the CRD and transcript expression levels in response to Cr(VI) challenge.</p> <p>Results</p> <p>A chromate-sensitive mutant (strain D11) was generated by curing FB24 of its 96-kb plasmid. Elemental analysis indicated that chromate-exposed cells of strain D11 accumulated three times more chromium than strain FB24. Introduction of the CRD into strain D11 conferred chromate resistance comparable to wild-type levels, whereas deletion of specific regions of the CRD led to decreased resistance. Using real-time reverse transcriptase PCR, we show that expression of each gene within the CRD is specifically induced in response to chromate but not by lead, hydrogen peroxide or arsenate. Higher levels of <it>chrA </it>expression were achieved when the <it>chrB </it>orthologs and the WD40 repeat domain genes were present, suggesting their possible regulatory roles.</p> <p>Conclusion</p> <p>Our findings indicate that chromate resistance in <it>Arthrobacter </it>sp. strain FB24 is due to chromate efflux through the ChrA transport protein. More importantly, new genes have been identified as having significant roles in chromate resistance. Collectively, the functional predictions of these additional genes suggest the involvement of a signal transduction system in the regulation of chromate efflux and warrants further study.</p

Pan-viral serology implicates enteroviruses in acute flaccid myelitis.

Since 2012, the United States of America has experienced a biennial spike in pediatric acute flaccid myelitis (AFM)1-6. Epidemiologic evidence suggests non-polio enteroviruses (EVs) are a potential etiology, yet EV RNA is rarely detected in cerebrospinal fluid (CSF)2. CSF from children with AFM (n = 42) and other pediatric neurologic disease controls (n = 58) were investigated for intrathecal antiviral antibodies, using a phage display library expressing 481,966 overlapping peptides derived from all known vertebrate and arboviruses (VirScan). Metagenomic next-generation sequencing (mNGS) of AFM CSF RNA (n = 20 cases) was also performed, both unbiased sequencing and with targeted enrichment for EVs. Using VirScan, the viral family significantly enriched by the CSF of AFM cases relative to controls was Picornaviridae, with the most enriched Picornaviridae peptides belonging to the genus Enterovirus (n = 29/42 cases versus 4/58 controls). EV VP1 ELISA confirmed this finding (n = 22/26 cases versus 7/50 controls). mNGS did not detect additional EV RNA. Despite rare detection of EV RNA, pan-viral serology frequently identified high levels of CSF EV-specific antibodies in AFM compared with controls, providing further evidence for a causal role of non-polio EVs in AFM

Multi-Scale Mass Transfer Processes Controlling Natural Attenuation and Engineered Remediation: An IFRC Focused on Hanford?s 300 Area Uranium Plume January 2010 to January 2011

The Integrated Field Research Challenge (IFRC) at the Hanford Site 300 Area uranium (U) plume addresses multi-scale mass transfer processes in a complex subsurface biogeochemical setting where groundwater and riverwater interact. A series of forefront science questions on reactive mass transfer motivates research. These questions relate to the effect of spatial heterogeneities; the importance of scale; coupled interactions between biogeochemical, hydrologic, and mass transfer processes; and measurements and approaches needed to characterize and model a mass-transfer dominated biogeochemical system. The project was initiated in February 2007, with CY 2007, CY 2008, CY 2009, and CY 2010 progress summarized in preceding reports. A project peer review was held in March 2010, and the IFRC project acted upon all suggestions and recommendations made in consequence by reviewers and SBR/DOE. These responses have included the development of 'Modeling' and 'Well-Field Mitigation' plans that are now posted on the Hanford IFRC web-site, and modifications to the IFRC well-field completed in CY 2011. The site has 35 instrumented wells, and an extensive monitoring system. It includes a deep borehole for microbiologic and biogeochemical research that sampled the entire thickness of the unconfined 300 A aquifer. Significant, impactful progress has been made in CY 2011 including: (i) well modifications to eliminate well-bore flows, (ii) hydrologic testing of the modified well-field and upper aquifer, (iii) geophysical monitoring of winter precipitation infiltration through the U-contaminated vadose zone and spring river water intrusion to the IFRC, (iv) injection experimentation to probe the lower vadose zone and to evaluate the transport behavior of high U concentrations, (v) extended passive monitoring during the period of water table rise and fall, and (vi) collaborative down-hole experimentation with the PNNL SFA on the biogeochemistry of the 300 A Hanford-Ringold contact and the underlying redox transition zone. The modified well-field has functioned superbly without any evidence for well-bore flows. Beyond these experimental efforts, our site-wide reactive transport models (PFLOTRAN and eSTOMP) have been updated to include site geostatistical models of both hydrologic properties and adsorbed U distribution; and new hydrologic characterization measurements of the upper aquifer. These increasingly robust models are being used to simulate past and recent U desorption-adsorption experiments performed under different hydrologic conditions, and heuristic modeling to understand the complex functioning of the smear zone. We continued efforts to assimilate geophysical logging and 3D ERT characterization data into our site wide geophysical model, with significant and positive progress in 2011 that will enable publication in 2012. Our increasingly comprehensive field experimental results and robust reactive transport simulators, along with the field and laboratory characterization, are leading to a new conceptual model of U(VI) flow and transport in the IFRC footprint and the 300 Area in general, and insights on the microbiological community and associated biogeochemical processes influencing N, S, C, Mn, and Fe. Collectively these findings and higher scale models are providing a unique and unparalleled system-scale understanding of the biogeochemical function of the groundwater-river interaction zone

Genome-Scale Modeling of Light-Driven Reductant Partitioning and Carbon Fluxes in Diazotrophic Unicellular Cyanobacterium Cyanothece sp. ATCC 51142

Genome-scale metabolic models have proven useful for answering fundamental questions about metabolic capabilities of a variety of microorganisms, as well as informing their metabolic engineering. However, only a few models are available for oxygenic photosynthetic microorganisms, particularly in cyanobacteria in which photosynthetic and respiratory electron transport chains (ETC) share components. We addressed the complexity of cyanobacterial ETC by developing a genome-scale model for the diazotrophic cyanobacterium, Cyanothece sp. ATCC 51142. The resulting metabolic reconstruction, iCce806, consists of 806 genes associated with 667 metabolic reactions and includes a detailed representation of the ETC and a biomass equation based on experimental measurements. Both computational and experimental approaches were used to investigate light-driven metabolism in Cyanothece sp. ATCC 51142, with a particular focus on reductant production and partitioning within the ETC. The simulation results suggest that growth and metabolic flux distributions are substantially impacted by the relative amounts of light going into the individual photosystems. When growth is limited by the flux through photosystem I, terminal respiratory oxidases are predicted to be an important mechanism for removing excess reductant. Similarly, under photosystem II flux limitation, excess electron carriers must be removed via cyclic electron transport. Furthermore, in silico calculations were in good quantitative agreement with the measured growth rates whereas predictions of reaction usage were qualitatively consistent with protein and mRNA expression data, which we used to further improve the resolution of intracellular flux values

MicroRNA Dysregulation in the Spinal Cord following Traumatic Injury

Spinal cord injury (SCI) triggers a multitude of pathophysiological events that are tightly regulated by the expression levels of specific genes. Recent studies suggest that changes in gene expression following neural injury can result from the dysregulation of microRNAs, short non-coding RNA molecules that repress the translation of target mRNA. To understand the mechanisms underlying gene alterations following SCI, we analyzed the microRNA expression patterns at different time points following rat spinal cord injury