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At least 667 records · Page 37

Use of immunomagnetic separation for the detection of Desulfovibrio vulgaris from environmental samples

Immunomagnetic separation (IMS) has proved highly efficient for recovering microorganisms from heterogeneous samples. Current investigation targeted the separation of viable cells of the sulfate-reducing bacterium, Desulfovibrio vulgaris. Streptavidin-coupled paramagnetic beads and biotin labeled antibodies raised against surface antigens of this microorganism were used to capture D. vulgaris cells in both bioreactor grown laboratory samples and from extremely low-biomass environmental soil and subsurface drilling samples. Initial studies on detection, recovery efficiency and viability for IMS were performed with laboratory grown D. vulgaris cells using various cell densities. Efficiency of cell isolation and recovery (i.e., release of the microbial cells from the beads following separation) was followed by microscopic imaging and acridine orange direct counts (AODC). Excellent recovery efficiency encouraged the use of IMS to capture Desulfovibrio spp. cells from low-biomass environmental samples. The environmental samples were obtained from a radionuclide-contaminated site in Germany and the chromium (VI)-contaminated Hanford site, an ongoing bioremediation project of the U.S. Department of Energy. Field deployable IMS technology may greatly facilitate environmental sampling and bioremediation process monitoring and enable transcriptomics and proteomics/metabolomics-based studies directly on cells collected from the field.

09 BIOMASS FUELS↗

Iron acquisition and mineral transformation by cyanobacteria living in extreme environments

Iron is an essential micronutrient for most living organisms, including cyanobacteria. These microorganisms have been found in Earth's driest polar and non-polar deserts, including the Atacama Desert, Chile. Iron-containing minerals were identified in colonized rock substrates from the Atacama Desert, however, the interactions between microorganisms and iron minerals remain unclear. In the current study, we determined that colonized gypsum rocks collected from the Atacama Desert contained both magnetite and hematite phases. A cyanobacteria isolate was cultured on substrates consisting of gypsum with embedded magnetite nanoparticles. Transmission electron microscopy imaging revealed a significant reduction in the size of magnetite nanoparticles due to their dissolution, which occurred around the microbial biofilms. Concurrently, hematite was detected, likely from the oxidation of the magnetite nanoparticles. Higher cell counts and production of siderophores were observed in cultures with magnetite nanoparticles suggesting that cyanobacteria were actively acquiring iron from the magnetite nanoparticles. Magnetite dissolution and iron acquisition by the cyanobacteria was further confirmed using large bulk magnetite crystals, uncovering a survival strategy of cyanobacteria in these extreme environments.

59 BASIC BIOLOGICAL SCIENCES↗

Oxygen demand for the stabilization of the organic fraction of municipal solid waste in passively aerated bioreactors

Highlights: • The use of an passively aerated reactor enables effective stabilization of OFMSW. • Convective air flow does not inhibit the aerobic stabilization of waste. • The use of an passively aerated reactor reduces the heat loss due to convection. • The volume of supplied air exceeds 1.7–2.88 times the microorganisms demand. - Abstract: Conventional aerobic waste treatment technologies require the use of aeration devices that actively transport air through the stabilized waste mass, which greatly increases operating costs. In addition, improperly operated active aeration systems, may have the adverse effect of cooling the stabilized biomass. Because active aeration can be a limiting factor for the stabilization process, passive aeration can be equally effective and less expensive. Unfortunately, there are few reports documenting the use of passive aeration systems in municipal waste stabilization. There have been doubts raised as to whether a passive aeration system provides enough oxygen to the organic matter mineralization processes. In this paper, the effectiveness of aeration during aerobic stabilization of four different organic fractions of municipal waste in a reactor with an integrated passive ventilation system and leachate recirculation was analyzed. For the study, four fractions separated by a rotary screen were chosen. Despite the high temperatures in the reactor, the air flow rate was below 0.016 m{sup 3}/h. Using Darcy’s equation, theoretical values of the air flow rate were estimated, depending on the intensity of microbial metabolism and the amount of oxygen required for the oxidation of organic compounds. Calculations showed that the volume of supplied air exceeded the microorganisms demand for oxidation and endogenous activity by 1.7–2.88-fold.

54 ENVIRONMENTAL SCIENCES↗

Phase Preference by Active, Acetate-Utilizing Bacteria at the Rifle, CO Integrated Field Research Challenge Site

Previous experiments at the Rifle, Colorado Integrated Field Research Challenge (IFRC) site demonstrated that field-scale addition of acetate to groundwater reduced the ambient soluble uranium concentration. In this report, sediment samples collected before and after acetate field addition were used to assess the active microbes via {sup 13}C acetate stable isotope probing on 3 phases [coarse sand, fines (8-approximately 150 {micro}m), groundwater (0.2-8 {micro}m)] over a 24-day time frame. TRFLP results generally indicated a stronger signal in {sup 13}C-DNA in the 'fines' fraction compared to the sand and groundwater. Before the field-scale acetate addition, a Geobacter-like group primarily synthesized {sup 13}C-DNA in the groundwater phase, an alpha Proteobacterium primarily grew on the fines/sands, and an Acinetobacter sp. and Decholoromonas-like OTU utilized much of the {sup 13}C acetate in both groundwater and particle-associated phases. At the termination of the field-scale acetate addition, the Geobacter-like species was active on the solid phases rather than the groundwater, while the other bacterial groups had very reduced newly synthesized DNA signal. These findings will help to delineate the acetate utilization patterns of bacteria in the field and can lead to improved methods for stimulating distinct microbial populations in situ.

15 GEOTHERMAL ENERGY↗

Using complex resistivity imaging to infer biogeochemical processes associated with bioremediation of a uranium-contaminated aquifer

Experiments at the Department of Energy’s Rifle Integrated Field Research Challenge (IFRC) site near Rifle, Colorado (USA) have demonstrated the ability to remove uranium from groundwater by stimulating the growth and activity of Geobacter species through acetate amendment. Prolonging the activity of these strains in order to optimize uranium bioremediation has prompted the development of minimally-invasive and spatially-extensive monitoring methods diagnostic of their in situ activity and the end products of their metabolism. Here we demonstrate the use of complex resistivity imaging for monitoring biogeochemical changes accompanying stimulation of indigenous aquifer microorganisms during and after a prolonged period (100+ days) of acetate injection. A thorough raw-data statistical analysis of discrepancies between normal and reciprocal measurements and incorporation of a new power-law phase-error model in the inversion were used to significantly improve the quality of the resistivity phase images over those obtained during previous monitoring experiments at the Rifle IRFC site. The imaging results reveal spatiotemporal changes in the phase response of aquifer sediments, which correlate with increases in Fe(II) and precipitation of metal sulfides (e.g., FeS) following the iterative stimulation of iron and sulfate reducing microorganism. Only modest changes in resistivity magnitude were observed over the monitoring period. The largest phase anomalies (>40 mrad) were observed hundreds of days after halting acetate injection, in conjunction with accumulation of Fe(II) in the presence of residual FeS minerals, reflecting preservation of geochemically reduced conditions in the aquifer – a prerequisite for ensuring the long-term stability of immobilized, redox-sensitive contaminants, such as uranium.

54 ENVIRONMENTAL SCIENCES↗

Biodegradation test of SPS-LS blends as polymer electrolyte membrane fuel cells

Sulfonated polystyrene (SPS) can be applied as a proton exchange membrane fuel cell due to its fairly good chemical stability. In order to be applied as polymer electrolyte membrane fuel cells (PEMFCs), membrane polymer should have a good ionic conductivity, high proton conductivity, and high mechanical strength. Lignosulfonate (LS) is a complex biopolymer which has crosslinks and sulfonate groups. SPS-LS blends with addition of SiO{sub 2} are used to increase the proton conductivity and to improve the mechanical properties and thermal stability. However, the biodegradation test of SPS-LS blends is required to determine whether the application of these membranes to be applied as an environmentally friendly membrane. In this study, had been done the synthesis of SPS, biodegradability test of SPS-LS blends with variations of LS and SiO{sub 2} compositions. The biodegradation test was carried out in solid medium of Luria Bertani (LB) with an activated sludge used as a source of microorganism at incubation temperature of 37°C. Based on the results obtained indicated that SPS-LS-SiO{sub 2} blends are more decomposed by microorganism than SPS-LS blends. This result is supported by analysis of weight reduction percentage, functional groups with Fourier Transform Infrared (FTIR) Spectroscopy, and morphological surface with Scanning Electron Microscopy (SEM)

36 MATERIALS SCIENCE↗

Viability of Cladosporium herbarum spores under 157 nm laser and vacuum ultraviolet irradiation, low temperature (10 K) and vacuum

Ultraviolet photons can damage microorganisms, which rarely survive prolonged irradiation. In addition to the need for intact DNA, cell viability is directly linked to the functionality of the cell wall and membrane. In this work, Cladosporium herbarum spore monolayers exhibit high viability (7%) when exposed to 157 nm laser irradiation (412 kJm⁻²) or vacuum-ultraviolet irradiation (110–180 nm) under standard pressure and temperature in a nitrogen atmosphere. Spore viability can be determined by atomic-force microscopy, nano-indentation, mass, μ-Raman and attenuated reflectance Fourier-transform far-infrared spectroscopies and DNA electrophoresis. Vacuum ultraviolet photons cause molecular damage to the cell wall, but radiation resistance in spores arises from the activation of a photon-triggered signaling reaction, expressed via the exudation of intracellular substances, which, in combination with the low penetration depth of vacuum-ultraviolet photons, shields DNA from radiation. Resistance to phototoxicity under standard conditions was assessed, as was resistance to additional environmental stresses, including exposure in a vacuum, under different rates of change of pressure during pumping time and low (10 K) temperatures. Vacuum conditions were far more destructive to spores than vacuum-ultraviolet irradiation, and UV-B photons were two orders of magnitude more damaging than vacuum-ultraviolet photons. The viability of irradiated spores was also enhanced at 10 K. This work, in addition to contributing to the photonic control of the viability of microorganisms exposed under extreme conditions, including decontamination of biological warfare agents, outlines the basis for identifying bio-signaling in vivo using physical methodologies.

36 MATERIALS SCIENCE↗

Kinetic study of treatment of wastewater contains food preservative agent by anaerobic baffled reactor : An overview

The characteristic of wastewater of food industries with preservative substances is high content of organic substances, degradable and high total suspended solid. High organic content in this waste forced the treatment is biologically and pointed out to anaerobic treatment. Anaerobic showed the better performance of degradation than aerobic for high content organic and also for toxic materials. During that day the treatment of food wastewater is aerobically which is high consume of energy required and high volume of sludge produced. The advantage of anaerobic is save high energy, less product of sludge, less requirement of nutrients of microorganism and high efficiency reduction of organic load. The high efficiency of reduction will reduce the load of further treatment, so that, the threshold limit based on the regulation would be easy to achieve. Research of treatment of wastewater of food industries would be utilized by both big scale industries and small industries using addition of preservative substances. The type reactor of anaerobic process is anaerobic baffled reactor that will give better contact between wastewater and microorganism in the sludge. The variables conducted in this research are the baffled configuration, sludge height, preservative agent contents, hydralic retention time and influence of micro nutrients. The respons of this research are the COD effluent, remaining preservative agent, pH, formation of volatile fatty acid and total suspended solid. The result of this research is kinetic model of the anaerobic baffled reactor, reaction kinetic of preservative agent degradation and technology of treatment wastewater contains preservative agent. The benefit of this research is to solve the treatment of wastewater of food industries with preservative substance in order to achieve wastewater limit regulation and also to prevent the environmental deterioration.

37 INORGANIC, ORGANIC, PHYSICAL, AND ANALYTICAL CH↗

Mutant alcohol dehydrogenase leads to improved ethanol tolerance in Clostridium thermocellum

Clostridium thermocellum is a thermophilic, obligately anaerobic, Gram-positive bacterium that is a candidate microorganism for converting cellulosic biomass into ethanol through consolidated bioprocessing. Ethanol intolerance is an important metric in terms of process economics, and tolerance has often been described as a complex and likely multigenic trait for which complex gene interactions come into play. Here, we resequence the genome of an ethanol-tolerant mutant, show that the tolerant phenotype is primarily due to a mutated bifunctional acetaldehyde-CoA/alcohol dehydrogenase gene (adhE), hypothesize based on structural analysis that cofactor specificity may be affected, and confirm this hypothesis using enzyme assays. Biochemical assays confirm a complete loss of NADH-dependent activity with concomitant acquisition of NADPH-dependent activity, which likely affects electron flow in the mutant. The simplicity of the genetic basis for the ethanol-tolerant phenotype observed here informs rational engineering of mutant microbial strains for cellulosic ethanol production.

Brown, Steven D.↗

Microbially induced precipitation of silica by anaerobic methane-oxidizing consortia and implications for microbial fossil preservation

Authigenic carbonate minerals can preserve biosignatures of microbial anaerobic oxidation of methane (AOM) in the rock record. It is not currently known whether the microorganisms that mediate sulfate-coupled AOM—often occurring as multicelled consortia of anaerobic methanotrophic archaea (ANME) and sulfate-reducing bacteria (SRB)—are preserved as microfossils. Electron microscopy of ANME-SRB consortia in methane seep sediments has shown that these microorganisms can be associated with silicate minerals such as clays [Chen et al., Sci. Rep. 4, 1–9 (2014)], but the biogenicity of these phases, their geochemical composition, and their potential preservation in the rock record is poorly constrained. Long-term laboratory AOM enrichment cultures in sediment-free artificial seawater [Yu et al., Appl. Environ. Microbiol. 88, e02109-21 (2022)] resulted in precipitation of amorphous silicate particles (~200 nm) within clusters of exopolymer-rich AOM consortia from media undersaturated with respect to silica, suggestive of a microbially mediated process. The use of techniques like correlative fluorescence in situ hybridization (FISH), scanning electron microscopy with energy dispersive X-ray spectroscopy (SEM-EDS), and nanoscale secondary ion mass spectrometry (nanoSIMS) on AOM consortia from methane seep authigenic carbonates and sediments further revealed that they are enveloped in a silica-rich phase similar to the mineral phase on ANME-SRB consortia in enrichment cultures. Like in cyanobacteria [Moore et al., Geology 48, 862–866 (2020)], the Si-rich phases on ANME-SRB consortia identified here may enhance their preservation as microfossils. Finally, the morphology of these silica-rich precipitates, consistent with amorphous-type clay-like spheroids formed within organic assemblages, provides an additional mineralogical signature that may assist in the search for structural remnants of microbial consortia in rocks which formed in methane-rich environments from Earth and other planetary bodies.

54 ENVIRONMENTAL SCIENCES↗

Clear as mud redefined: Tunable transparent mineral scaffolds for visualizing microbial processes below ground

Microbes inhabiting complex porous microenvironments in sediments and aquifers catalyze reactions that are critical to global biogeochemical cycles and ecosystem health. However, the opacity and complexity of porous sediment and rock matrices have considerably hindered the study of microbial processes occurring within these habitats. Here, we generated microbially compatible, optically transparent mineral scaffolds to visualize and investigate microbial colonization and activities occurring in these environments, in laboratory settings and in situ. Using inexpensive synthetic cryolite mineral, we produced optically transparent scaffolds mimicking the complex 3D structure of sediments and rocks by adapting a suspension-based, freeze-casting technique commonly used in materials science. Fine-tuning of parameters, such as freezing rate and choice of solvent, provided full control of pore size and architecture. The combined effects of scaffold porosity and structure on the movement of microbe-sized particles, tested using velocity tracking of fluorescent beads, showed diverse yet reproducible behaviors. The scaffolds we produced are compatible with epifluorescence microscopy, allowing the fluorescence-based identification of colonizing microbes by DNA-based staining and fluorescence in situ hybridization (FISH) to depths of 100 µm. Additionally, Raman spectroscopy analysis indicates minimal background signal in regions used for measuring deuterium and 13 C enrichment in microorganisms, highlighting the potential to directly couple D 2 O or 13 C stable isotope probing and Raman-FISH for quantifying microbial activity at the single-cell level. To demonstrate the relevance of cryolite scaffolds for environmental field studies, we visualized their colonization by diverse microorganisms within rhizosphere sediments of a coastal seagrass plant using epifluorescence microscopy. The tool presented here enables highly resolved, spatially explicit, and multimodal investigations into the distribution, activities, and interactions of underground microbes typically obscured within opaque geological materials until now.

36 MATERIALS SCIENCE↗

Microbial Lithotrophic Oxidation of Structural Fe(II) in Biotite

Microorganisms are known to participate in the weathering of primary phyllosilicate minerals through production of organic ligands and acids, and through uptake of products of weathering. Here we show that a lithotrophic Fe(II)-oxidizing, nitrate-reducing enrichment culture (Straub, 6 1996) can grow via oxidation of structural Fe(II) in biotite, a Fe(II)-rich trioctahedral mica found in granitic rocks. Oxidation of silt/clay sized biotite particles was detected by a decrease in extractable Fe(II) content and simultaneous nitrate reduction. Moessbauer spectroscopy confirmed structural Fe(II) oxidation. Approximately 107 cells were produced per {micro}mol Fe(II) oxidized, in agreement with previous estimates of the growth yield of lithoautotrophic circumneutral-pH Fe(II)-oxidizing bacteria. Microbial oxidation of structural Fe(II) resulted in biotite alterations similar to those found in nature, including decrease in unit cell b-dimension toward dioctahedral levels and iron and potassium release. The demonstration of microbial oxidation of structural Fe(II) suggests that microorganisms may be directly responsible for the initial step in the weathering of biotite in granitic aquifers and the plant rhizosphere.

54 ENVIRONMENTAL SCIENCES↗

Genomic encyclopedia of sugar utilization pathways in the Shewanella genus

To address a practically and fundamentally important challenge of reconstruction of carbohydrate utilization machinery in any microorganism directly from its genomic sequence, we have established a subsystems-based comparative approach and applied it to 19 genomes from the Shewanella genus. The key stages of our approach include: (i) a homology-based identification of gene candidates using a genomic compilation of ~500 known components of sugar catabolic pathways; (ii) functional assignment of orthologs and prediction of alternative genes and pathway variants based on genomic (operons, regulons) and functional (subsystems, pathways) context analysis; (iii) validation of bioinformatic predictions by a combination of biochemical, genetic and physiological experiments. The obtained genomic encyclopedia of sugar utilization includes ~170 protein families (mostly metabolic enzymes, transporters and transcriptional regulators) spanning 17 distinct pathways with a mosaic distribution across Shewanella species providing insights into their ecophysiology and adaptive evolution. The reconstructed catabolic pathways are significantly enriched by nonorthologous gene replacements and alternative biochemical routes. Phenotypic assays revealed a remarkable consistency between predicted and observed phenotype, an ability to utilize an individual sugar as a sole source of carbon and energy, over the entire matrix of tested strains and sugars. In addition to improving our knowledge of genomics, functional organization and evolution of the sugar catabolome, this study confirmed the efficiency of the established approach, which is scalable and applicable to other groups of microorganisms.

59 BASIC BIOLOGICAL SCIENCES↗

Clostridium thermocellum ATCC27405 transcriptomic, metabolomic and proteomic profiles after ethanol stress

Clostridium thermocellum is a candidate consolidated bioprocessing biocatalyst, which is a microorganism that expresses enzymes for both cellulose hydrolysis and its fermentation to produce fuels such as lignocellulosic ethanol. However, C. thermocellum is relatively sensitive to ethanol compared to ethanologenic microorganisms such as yeast and Zymomonas mobilis that are used in industrial fermentations but do not possess native enzymes for industrial cellulose hydrolysis. In this study, C. thermocellum was grown to mid-exponential phase and then treated with ethanol to a final concentration of 3.9 g/L to investigate its physiological and regulatory responses to ethanol stress. Samples were taken pre-shock and 2, 12, 30, 60, 120, and 240 min post-shock, and from untreated control fermentations for systems biology analyses. Cell growth was arrested by ethanol supplementation with intracellular accumulation of carbon sources such as cellobiose, and sugar phosphates, including fructose-6-phosphate and glucose-6-phosphate. The largest response of C. thermocellum to ethanol shock treatment was in genes and proteins related to nitrogen uptake and metabolism, which is likely important for redirecting the cells physiology to overcome inhibition and allow growth to resume. This study suggests possible avenues for metabolic engineering and provides comprehensive, integrated systems biology datasets that will be useful for future metabolic modeling and strain development endeavors.

59 BASIC BIOLOGICAL SCIENCES↗

Bacterial and Archaea Community Present in the Pine Barrens Forest of Long Island, NY: Unusually High Percentage of Ammonia Oxidizing Bacteria

Of the few preserved areas in the northeast of United States, the soil in the Pine Barrens Forests presents a harsh environment for the microorganisms to grow and survive. In the current study we report the use of clustering methods to scientifically select the sampling locations that would represent the entire forest and also report the microbial diversity present in various horizons of the soil. Sixty six sampling locations were selected across the forest and soils were collected from three horizons (sampling depths). The three horizons were 0-10 cm (Horizon O); 11-25 cm (Horizon A) and 26-40 cm (Horizon B). Based on the total microbial substrate utilization pattern and K-means clustering analysis, the soil in the Pine Barrens Forest can be classified into four distinct clusters at each of the three horizons. One soil sample from each of the four clusters were selected and archaeal and bacterial populations within the soil studied using pyrosequencing method. The results show the microbial communities present in each of these clusters are different. Within the microbial communities present, microorganisms involved in nitrogen cycle occupy a major fraction of microbial community in the soil. High level of diversity was observed for nitrogen fixing bacteria. In contrast, Nitrosovibrio and Nitrosocaldus spp are the single bacterial and archaeal population respectively carrying out ammonia oxidation in the soil.

54 ENVIRONMENTAL SCIENCES↗

Non-contiguous finished genome sequence of Bacteroides coprosuis type strain (PC139 T )

Bacteroides coprosuis Whitehead et al. 2005 belongs to the genus Bacteroides, which is a member of the family Bacteroidaceae. Members of the genus Bacteroides in general are known as beneficial protectors of animal guts against pathogenic microorganisms, and as contributors to the degradation of complex molecules such as polysaccharides. B. coprosuis itself was isolated from a manure storage pit of a swine facility, but has not yet been found in an animal host. The species is of interest solely because of its isolated phylogenetic location. The genome of B. coprosuis is already the 5(th) sequenced type strain genome from the genus Bacteroides. The 2,991,798 bp long genome with its 2,461 protein-coding and 78 RNA genes and is a part of the Genomic Encyclopedia of Bacteria and Archaea project.

59 BASIC BIOLOGICAL SCIENCES↗