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Target properties of CO2 capture adsorbents that would ensure economic viability of bioenergy with carbon capture and storage (BECCS) are defined. The key role of sorbent lifetime in the process cost is demonstrated, and an optimal heat of adsorption for BECCS is postulated through a balance of adsorbent-adsorbate affinity and regeneration energy demand. Using an exponential decay model of sorbent capacity increases the process cost and results in an optimum sorbent lifetime. To ensure a levelized cost of carbon below $100/tonne-CO2, adsorbents should be designed to have working capacities above 0.75 mol/kg, lifetimes over 2 years, heats of adsorption of approximately -40 kJ/mol, and exponential degradation decay constants below 5 × 10-6 cycle-1 (equivalent to a half-life of 1.3 years). Our model predicts a BECCS process cost of $65/t-CO2 can be achieved with a degradation-resistant adsorbent, $40/kg sorbent cost, 2.0 mol/kg working capacity, -40 kJ/mol heat of adsorption, and at least a 2 year lifetime.

Recent advances in the utilization of hydrogen as an alternative fuel source to conventional fossil fuels have led to a search for a renewable process of producing hydrogen. Most hydrogen today is produced from hydrocarbons in a process that also releases high levels of carbon dioxide and carbon monoxide, two established greenhouse gasses; because of this harmful means of production, research has been directed toward using anaerobic digestion to produce useful levels of hydrogen gas. Anaerobic systems have been shown to produce a biogas that is easily used in producing energy, but certain processes can be performed to further enhance the concentrations of hydrogen. These processes include the inhibition of microorganisms that lower hydrogen concentrations and the constant removal of hydrogen to promote hydrogen-producing bacteria. Experimental designs and large-scale applications have shown this process to be environmentally viable with limited, but promising, economic potential. With a constant increase for the need of hydrogen gas, the sustainable production of hydrogen is becoming more important. This review explores some of the recent research on this topic and explores the processes behind using anaerobic digestion for hydrogen production.

PURPOSE OF REVIEW: Glycopeptide antibiotics such as vancomycin are frequently utilized to treat resistant Gram-positive infections such as methicillin-resistant Staphylococcus aureus. The current literature on glycopeptide and lipoglycopeptide structure, hypersensitivity and potential cross-reactivity was reviewed, highlighting implications for safe prescribing. RECENT FINDINGS: Structurally similar, glycopeptides could theoretically cross-react. Immediate reactions to vancomycin include non-IgE-mediated reactions (e.g. red man syndrome) and IgE-mediated hypersensitivity (e.g. anaphylaxis), sharing clinical features. Vancomycin can activate mast cells via MAS-related G-protein-coupled receptor X2, an IgE-independent receptor implicated in non-IgE reactions. In-vivo and in-vitro testing for suspected IgE-mediated reactions to glycopeptides remain ill-defined. Vancomycin is increasingly recognized to cause severe cutaneous adverse reactions (SCAR), with drug reaction with eosinophilia and systemic symptoms (DRESS) predominantly reported. Vancomycin DRESS has been associated with HLA-A32:-01, with a number needed to prevent of 1 in 74. Data demonstrating cross-reactivity amongst glycopeptides and lipoglycopeptides is limited to case reports/series. SUMMARY: Further studies and in-vivo/in-vitro diagnostics are required for better differentiation between IgE and non-IgE glycopeptide reactions. Despite its association with vanomycin DRESS, utility of pharmacogenomic screening for HLA-A32: 01 is ill-defined. Although HLA-A32:01 has been associated with vancomycin DRESS, its utility for pharmacogenomic screening is ill defined. Further clinical and immunological cross-reactivity data for glycopeptide/lipoglycopeptide antibiotics is required.

Nicotinamide riboside (NR) is a naturally occurring form of vitamin B3 shown to preferentially elevate the nicotinamide adenine dinucleotide (NAD+) metabolome compared to other vitamin B3 forms (nicotinic acid and nicotinamide). Although daily requirements of vitamin B3 are typically met through the diet, recent studies have shown that additional supplementation with NR may be an effective method to counter the age-related decline in NAD+ levels as NR bypasses the rate-limiting step in NAD+ biosynthesis. Furthermore, pharmaceutical applications of NR for age-related disorders have been proposed. In this study, the safety of a high-purity, nature-identical, synthetic NR (NR-E), manufactured under the guidelines of good manufacturing practices for dietary supplements (21 CFR 111) as well as for drugs (21 CFR 210), was investigated in a 90-day oral toxicity study in Sprague Dawley rats at 300, 500, and 1,200 mg/kg/d. There were no mortality or clinical observations attributable to the test substance at any dose. A small but statistically significant decrease in body weight was observed at day 92 in the 1,200 mg/kg/d NR-treated male rats only. In contrast to a previously published safety assessment using a different synthetic NR (NIAGEN), whose no-observed-adverse-effect-level (NOAEL) was reported to be 300 mg/kg/d, there were no adverse changes in clinical pathology parameters and no notable macroscopic or microscopic findings or treatment-related effects at similar doses. In the current study, the NOAEL for systemic toxicity of NR-E in Sprague-Dawley rats was conservatively determined to be 500 mg/kg/d for males (solely based on body weight) and 1,200 mg/kg/d for females.

Certain transcription factors are proposed to form functional interactions with RNA to facilitate proper regulation of gene expression. Sox2, a transcription factor critical for maintenance of pluripotency and neurogenesis, has been found associated with several lncRNAs, although it is unknown whether these interactions are direct or via other proteins. Here we demonstrate that human Sox2 interacts directly with one of these lncRNAs with high affinity through its HMG DNA-binding domain in vitro. These interactions are primarily with double-stranded RNA in a non-sequence specific fashion, mediated by a similar but not identical interaction surface. We further determined that Sox2 directly binds RNA in mouse embryonic stem cells by UV-cross-linked immunoprecipitation of Sox2 and more than a thousand Sox2-RNA interactions in vivo were identified using fRIP-seq. Together, these data reveal that Sox2 employs a high-affinity/low-specificity paradigm for RNA binding in vitro and in vivo., Some transcription factors have been proposed to functionally interact with RNA to facilitate proper regulation of gene expression. Here the authors demonstrate that human Sox2 interact directly and with high affinity to RNAs through its HMG DNA-binding domain.

The multi-heme c-type cytochrome OmcS is one of the central components used for extracellular electron transport in the Geobacter sulfurreducens strain DL-1, but its role in other microbes, including other strains of G. sulfurreducens, is currently a matter of debate. Therefore, we investigated the function of OmcS in the G. sulfurreducens strain KN400, which is even more effective in extracellular electron transfer than the DL-1 strain. We found that deleting omcS from strain KN400 did not negatively impact the rate of Fe(III) oxide reduction and that the cells expressed conductive filaments. Replacing the wild-type pilin gene with the aro-5 pilin gene eliminated the OmcS-deficient strain’s ability to transport electrons to insoluble electron acceptors and diminished filament conductivity. These results are consistent with the concept that electrically conductive pili are the primary conduit for long-range electron transfer in G. sulfurreducens and closely related species. These findings, coupled with the lack of OmcS homologs in other microbes capable of extracellular electron transfer, suggest that OmcS is not a common critical component for extracellular electron transfer. IMPORTANCE OmcS has been widely studied and noted to be one of the key components for extracellular electron exchange by the Geobacter sulfurreducens strain DL-1. However, the true importance of OmcS warrants further investigation because it is well known that few bacteria, even within the Geobacteraceae family, contain OmcS homologs, and many bacteria that are capable of extracellular electron transfer lack an abundance of any type of outer surface c-type cytochrome. In addition, there is debate about the importance of OmcS filaments in the mechanism of extracellular electron transport to insoluble electron acceptors by G. sulfurreducens. It has been suggested that filaments comprised of OmcS rather than e-pili are the predominant conductive filaments expressed by G. sulfurreducens. However, the results presented here, along with multiple other sources of evidence, indicate that OmcS filaments cannot be the primary, conductive, protein nanowires expressed by G. sulfurreducens.

Microbial extracellular electron transfer plays an important role in diverse biogeochemical cycles, metal corrosion, bioelectrochemical technologies, and anaerobic digestion. Evaluation of electron uptake from pure Fe(0) and stainless steel indicated that, in contrast to previous speculation in the literature, Desulfovibrio ferrophilus and Desulfopila corrodens are not able to directly extract electrons from solid-phase electron-donating surfaces. D. ferrophilus grew with Fe(III) as the electron acceptor, but Dp. corrodens did not. D. ferrophilus reduced Fe(III) oxide occluded within porous alginate beads, suggesting that it released a soluble electron shuttle to promote Fe(III) oxide reduction. Conductive atomic force microscopy revealed that the D. ferrophilus pili are electrically conductive and the expression of a gene encoding an aromatics-rich putative pilin was upregulated during growth on Fe(III) oxide. The expression of genes for multi-heme c-type cytochromes was not upregulated during growth with Fe(III) as the electron acceptor, and genes for a porin-cytochrome conduit across the outer membrane were not apparent in the genome. The results suggest that D. ferrophilus has adopted a novel combination of strategies to enable extracellular electron transport, which may be of biogeochemical and technological significance.

Direct interspecies electron transfer (DIET) between bacteria and methanogenic archaea appears to be an important syntrophy in both natural and engineered methanogenic environments. However, the electrical connections on the outer surface of methanogens and the subsequent processing of electrons for carbon dioxide reduction to methane are poorly understood. Here, we report that the genetically tractable methanogen Methanosarcina acetivorans can grow via DIET in coculture with Geobacter metallireducens serving as the electron-donating partner. Comparison of gene expression patterns in M. acetivorans grown in coculture versus pure-culture growth on acetate revealed that transcripts for the outer-surface multiheme c-type cytochrome MmcA were higher during DIET-based growth. Deletion of mmcA inhibited DIET. The high aromatic amino acid content of M. acetivorans archaellins suggests that they might assemble into electrically conductive archaella. A mutant that could not express archaella was deficient in DIET. However, this mutant grew in DIET-based coculture as well as the archaellum-expressing parental strain in the presence of granular activated carbon, which was previously shown to serve as a substitute for electrically conductive pili as a conduit for long-range interspecies electron transfer in other DIET-based cocultures. Transcriptomic data suggesting that the membrane-bound Rnf, Fpo, and HdrED complexes also play a role in DIET were incorporated into a charge-balanced model illustrating how electrons entering the cell through MmcA can yield energy to support growth from carbon dioxide reduction. The results are the first genetics-based functional demonstration of likely outer-surface electrical contacts for DIET in a methanogen. IMPORTANCE The conversion of organic matter to methane plays an important role in the global carbon cycle and is an effective strategy for converting wastes to a useful biofuel. The reduction of carbon dioxide to methane accounts for approximately a third of the methane produced in anaerobic soils and sediments as well as waste digesters. Potential electron donors for carbon dioxide reduction are H2 or electrons derived from direct interspecies electron transfer (DIET) between bacteria and methanogens. Elucidating the relative importance of these electron donors has been difficult due to a lack of information on the electrical connections on the outer surfaces of methanogens and how they process the electrons received from DIET. Transcriptomic patterns and gene deletion phenotypes reported here provide insight into how a group of Methanosarcina organisms that play an important role in methane production in soils and sediments participate in DIET.

Tissue culture approaches are widely used in crop plants for the purposes of micropropagation, regeneration of plants through organogenesis, obtaining pathogen-free plantlets from meristem culture, and developing genetically modified plants. In this research, we evaluated variables that can influence the success of shoot growth and plantlet production in tissue cultures of drug-typeCannabis sativaL. (marijuana). Various sterilization methods were tested to ensure shoot development from nodal explants by limiting the frequency of contaminating endophytes, which otherwise caused the death of explants. Seven commercially grown tetrahydrocannabinol (THC)-containing cannabis genotypes (strains) showed significant differences in response to shoot growth from meristems and nodal explants on Murashige and Skoog (MS) medium containing thidiazuron (1 μM) and naphthaleneacetic acid (0.5 μM) plus 1% activated charcoal. The effect of Driver and Kuniyuki Walnut (DKW) or MS basal salts in media on shoot length and leaf numbers from nodal explants was compared and showed genotype dependency with regard to the growth response. To obtain rooted plantlets, shoots from meristems and nodal explants of genotype Moby Dick were evaluated for rooting, following the addition of sodium metasilicate, silver nitrate, indole-3-butyric acid (IBA), kinetin, or 2,4-D. Sodium metasilicate improved the visual appearance of the foliage and improved the rate of rooting. Silver nitrate also promoted rooting. Following acclimatization, plantlet survival in hydroponic culture, peat plugs, and rockwool substrate was 57, 76, and 83%, respectively. The development of plantlets from meristems is described for the first time inC. sativaand has potential for obtaining pathogen-free plants. The callogenesis response of leaf explants of 11 genotypes on MS medium without activated charcoal was 35% to 100%, depending on the genotype; organogenesis was not observed. The success in recovery of plantlets from meristems and nodal explants is influenced by cannabis genotype, degree of endophytic contamination of the explants, and frequency of rooting. The procedures described here have potential applications for research and commercial utility to obtain plantlets in stage 1 tissue cultures ofC. sativa.

Syntrophic interspecies electron exchange is essential for the stable functioning of diverse anaerobic microbial communities. Hydrogen/formate interspecies electron transfer (HFIT), in which H2 and/or formate function as diffusible electron carriers, has been considered to be the primary mechanism for electron transfer because most common syntrophs were thought to lack biochemical components, such as electrically conductive pili (e-pili), necessary for direct interspecies electron transfer (DIET). Here we report that Syntrophus aciditrophicus, one of the most intensively studied microbial models for HFIT, produces e-pili and can grow via DIET. Heterologous expression of the putative S. aciditrophicus type IV pilin gene in Geobacter sulfurreducens yielded conductive pili of the same diameter (4 nm) and conductance of the native S. aciditrophicus pili and enabled long-range electron transport in G. sulfurreducens. S. aciditrophicus lacked abundant c-type cytochromes often associated with DIET. Pilin genes likely to yield e-pili were found in other genera of hydrogen/formate-producing syntrophs. The finding that DIET is a likely option for diverse syntrophs that are abundant in many anaerobic environments necessitates a reexamination of the paradigm that HFIT is the predominant mechanism for syntrophic electron exchange within anaerobic microbial communities of biogeochemical and practical significance.Access opt

Chemical activation experiments and computational methods have been used to study the unimolecular reactions of C2H5CH2Br and C2D5CHFBr with 90 and 93 kcal mol–1 of vibrational energy, respectively...

By mimicking iron(Fe)-based phosphorus (P) immobilization in natural environments, an Fe-retrofitted UCT-MBR involving in situ vivianite crystallization for removing and recovering P from sewage was developed, and its performance was examined in this work. We show that dosing of ferrihydrite, once biological P uptake reached its limit, enabled effective ongoing P removal; whereas conventional conditions in the anaerobic chamber of the University of Cape Town (UCT) system (i.e., a sludge retention time of hours and a completely mixed sludge phase) was insufficient for a satisfactory Fe(III) bioreduction, with the overaccumulation of Fe(III) as fine particles finally resulting in severe membrane fouling and collapse in P removal. The enhancement of reductive conditions in the anaerobic chamber by lowering agitation and adding biocarriers to favor Fe(III) reduction was found to be effective in enabling ongoing P removal and recovery. The average level of effluent P was as low as 0.18 mg/L for a period of 258 d under this condition. Using chemical and spectroscopic methods, the P product was identified as primarily vivianite: Fe

Disinfection byproducts are formed during most drinking water treatment and presently number >800, some of which are implicated in human health outcomes including bladder cancer and infertility, with unknown mechanisms of action. In particular, it is not yet understood whether these compounds can disrupt the estrogen-signaling pathway through binding to the human estrogen receptor (ER). In the present study, 21 disinfection byproducts, selected for their predicted involvement in endocrine-related diseases and their structural diversity, were individually evaluated for their binding affinity to the human ER and in silico, and then a subset of these chemicals was studied in binary mixtures with the known weak estrogen, 4-n-nonylphenol. Individually, 9 of the 21 disinfection byproducts were able to weakly bind to the ER, with affinities ranging from log median inhibitory concentration values of -3.83 to -2.19 M. In binary mixtures, the chemicals followed concentration addition, with their weak binding affinities having little contribution to the overall mixture affinity. These results demonstrate the variety of small-molecule disinfection byproduct structures that are capable of binding to the ER, and that their weak binding can still be of importance when overall human exposure to mixtures of disinfection byproducts in disinfected drinking water is considered. Environ Toxicol Chem 2019;9999:1-9. © 2019 SETAC.

The gas-phase unimolecular reactions of C2D5CHFCl molecules with 94 kcal mol-1 of vibrational energy have been studied by the chemical-activation experimental technique and by electronic-structure computations. Products from the reaction of C2D5CHFCl molecules, formed by the recombination of C2D5 and CHFCl radicals in a room temperature bath gas, were measured by gas chromatography-mass spectrometry. The 2,1-DCl (81%) and 1,1-HCl (17%) elimination reactions are the principal processes, but 2,1-DF and 1,1-HF elimination reactions also are observed. Comparison of experimental rate constants to calculated statistical rate constants provides threshold energies. The potential surfaces associated with C2D5(F)C: + HCl and C2D5(Cl)C: + HF reactions are of special interest because hydrogen-bonded adducts with HCl and HF with dissociation energies of 6.4 and 9.3 kcal mol-1, respectively, are predicted by calculations. The relationship between the geometries and threshold energies of transition states for 1,1-HCl elimination and carbene:HCl adducts is complex, and previous studies of related molecules, such as CD3CHFCl, CD2ClCHFCl, C2D5CHCl2, and halogenated methanes are included in the computational analysis. Extensive calculations for CH3CHFCl as a model for 1,1-HCl reactions illustrate properties of the exit-channel potential energy surface. Since the 1,1-HCl transition state is submerged relative to dissociation of the adduct, inner and outer transition states should be considered for analysis of rate constants describing 1,1-HCl elimination and addition reactions of carbenes to HCl.

It is known that the physiology of Methanosarcina species can differ significantly, but the ecological impact of these differences is unclear. We recovered two strains of Methanosarcina from two different ecosystems with a similar enrichment and isolation method. Both strains had the same ability to metabolize organic substrates and participate in direct interspecies electron transfer but also had major physiological differences. Strain DH-1, which was isolated from an anaerobic digester, used H2 as an electron donor. Genome analysis indicated that it lacks an Rnf complex and conserves energy from acetate metabolism via intracellular H2 cycling. In contrast, strain DH-2, a subsurface isolate, lacks hydrogenases required for H2 uptake and cycling and has an Rnf complex for energy conservation when growing on acetate. Further analysis of the genomes of previously described isolates, as well as phylogenetic and metagenomic data on uncultured Methanosarcina in anaerobic digesters and diverse soils and sediments, revealed a physiological dichotomy that corresponded with environment of origin. The physiology of type I Methanosarcina revolves around H2 production and consumption. In contrast, type II Methanosarcina species eschew H2 and have genes for an Rnf complex and the multiheme, membrane-bound c-type cytochrome MmcA, shown to be essential for extracellular electron transfer. The distribution of Methanosarcina species in diverse environments suggests that the type I H2-based physiology is well suited for high-energy environments, like anaerobic digesters, whereas type II Rnf/cytochrome-based physiology is an adaptation to the slower, steady-state carbon and electron fluxes common in organic-poor anaerobic soils and sediments. IMPORTANCE Biogenic methane is a significant greenhouse gas, and the conversion of organic wastes to methane is an important bioenergy process. Methanosarcina species play an important role in methane production in many methanogenic soils and sediments as well as anaerobic waste digesters. The studies reported here emphasize that the genus Methanosarcina is composed of two physiologically distinct groups. This is important to recognize when interpreting the role of Methanosarcina in methanogenic environments, especially regarding H2 metabolism. Furthermore, the finding that type I Methanosarcina species predominate in environments with high rates of carbon and electron flux and that type II Methanosarcina species predominate in lower-energy environments suggests that evaluating the relative abundance of type I and type II Methanosarcina may provide further insights into rates of carbon and electron flux in methanogenic environments.

Heterotrophic nitrifying and aerobic denitrifying bacteria (HNADB) are important for partial nitrification treatment of high strength ammonium leachate. However, conditions for their optimal performance in mixed reactor systems have yet to be determined. In this study, optimal parameters were identified and included free ammonia (FA) concentrations below 40 mg/L, a dissolved oxygen concentration of 1.2 mg/L, a carbon to nitrogen ratio of 5 and a reflux ratio of 4. These conditions were applied to a continuous anoxic/oxic membrane moving biofilm reactor treating raw incineration leachate with high total ammonium nitrogen (TAN = 1400 mg/L). Ammonium conversion and nitrogen removal efficiencies of 99% and 86% were achieved. Autotrophic ammonia oxidizing bacteria were inhibited at FA concentrations above 25 mg/L. HNADB, particularly Paracoccus species, contributed to ammonium conversion at high FA (25–40 mg/L). These results show that leachate with high TAN and FA can be treated using parameters that support the growth of HNADB.

The “International Research Experience for Students (IRES)” at Doane University (DU) located in Crete, Nebraska, exposed undergraduate science, technology, engineering, and mathematics (STEM) students to international research at the Karlsruhe Institute of Technology (KIT) in Germany. The international collaboration team included three undergraduate researchers per year from DU, one faculty member and one postdoctoral fellow from DU, two faculty mentors at KIT, and several graduate, post-doctoral, and technical staff at KIT. Prior to departure to Germany, the students received extensive research training, as well as culture and language preparation from the mentors at DU. While in Germany, the students received an in-depth orientation to Karlsruhe, Germany, Europe, the research setting at KIT, and the international collaborators. The eight week summer projects over three years involved nanolithography, nano- to microsized array fabrication, organic synthesis using click chemistry, and surface modifications for sensing and other biomedical research applications. When the students returned from Germany, they continued to conduct research at DU and train other undergraduate students using the expertise acquired from KIT. The DU research students, including the IRES scholars, learned oral and written communication skills. They presented their KIT and DU research results at weekly seminars and at local and national meetings. An external assessment firm evaluated the program, the students, and mentors on a yearly basis before and after the summer research. This enabled all participants to continuously improve the learning objectives and the program execution including three program adjustments implemented in year 2 or 3. The survey data shows that the IRES program provided an enriching experience for the students in research and international culture and established a successful base of collaboration for mentors.

High concentrations of nitrous oxide were recovered from partial nitrification treated leachate in a microbial electrolysis cell (MEC) inoculated with a nosZ-deficient strain of Pseudomonas aeruginosa. N2O conversion efficiencies > 90% were achieved when a potential of 0.8 V was applied to the MEC. The ΔnosZ strain was enriched in the 0.8 V MEC, but Achromobacter dominated the non-current control. Nitric oxide reductase genes were highly expressed by ΔnosZ cells growing in the 0.8 V MEC, consistent with enhanced nitrous oxide production rates. Concentrations of phenazine derivatives and transcripts from phenazine biosynthesis genes were also high in the 0.8 V MEC. Phenazine derivatives are known to act as electron shuttles, enhance biofilm formation, and help ward off competitors, thereby increasing the survivability of the ΔnosZ strain in the MEC. These results show that applied current stabilized growth of the ΔnosZ strain in the reactor and allowed it to sustainably generate high concentrations of nitrous oxide.

Cholesterol seems to play a central role in the augmentation of saporin-based immunotoxin (IT) cytotoxicity by triterpenoid saponins. Endolysosomal escape has been proposed as one mechanism for the saponin-mediated enhancement of targeted toxins. We investigated the effects of lipid depletion followed by repletion on Saponinum album (SA)-induced endolysosomal escape of Alexa Fluor labelled saporin and the saporin-based immunotoxin OKT10-SAP, directed against CD38, in Daudi lymphoma cells. Lipid deprived cells showed reduced SA-induced endolysosomal escape at two concentrations of SA, as determined by a flow cytometric method. The repletion of membrane cholesterol by low density lipoprotein (LDL) restored SA-induced endolysosomal escape at a concentration of 5 &micro, g/mL SA but not at 1 &micro, g/mL SA. When LDL was used to restore the cholesterol levels in lipid deprived cells, the SA augmentation of OKT10-SAP cytotoxicity was partially restored at 1 &micro, g/mL SA and fully restored at 5 &micro, g/mL SA. These results suggest that different mechanisms of action might be involved for the two different concentrations of SA and that endosomal escape may not be the main mechanism for the augmentation of saporin IT cytotoxicity by SA at the sub-lytic concentration of 1 &micro, g/mL SA.

The synaptic vesicle protein 2A (SV2A) is an integral 12-transmembrane domain glycoprotein located in the membrane of synaptic vesicles and widely distributed throughout the brain. In 2004, SV2A was shown to be the molecular target of the antiepileptic drug levetiracetam. High-affinity SV2A ligands have been described by the pharmaceutical company UCB more recently, and this enabled the rapid development of SV2A positron emission tomography (PET) radioligands. Because synaptic vesicle proteins, such as SV2A, have previously been used as biomarkers of synaptic density, PET imaging of SV2A may have broad utility across neuropathological diseases. In this chapter we review the current status of SV2A PET imaging.

The formation of vivianite (Fe3(PO4)2·8H2O) in iron (Fe)-dosed wastewater treatment facilities has the potential to develop into an economically feasible method of phosphorus (P) recovery. In this work, a long-term steady FeIII-dosed University of Cape Town process-membrane bioreactor (UCT-MBR) system was investigated to evaluate the role of Fe transformations in immobilizing P via vivianite crystallization. The highest fraction of FeII, to total Fe (Fetot), was observed in the anaerobic chamber, revealing that a redox condition suitable for FeIII reduction was established by improving operational and configurational conditions. The supersaturation index for vivianite in the anaerobic chamber varied but averaged ∼4, which is within the metastable zone and appropriate for its crystallization. Vivianite accounted for over 50% of the Fetot in the anaerobic chamber, and its oxidation as it passed through the aerobic chambers was slow, even in the presence of high dissolved oxygen concentrations at circumneutral pH. This study has shown that the high stability and growth of vivianite crystals in oxygenated activated sludge can allow for the subsequent separation of vivianite as a P recovery product.

The multi-heme c-type cytochrome OmcS, is one of the central components for extracellular electron transport in Geobacter sulfurreducens strain DL-1, but its role in other microbes, including other strains of G. sulfurreducens is currently a matter of debate. Therefore, we investigated the function of OmcS in G. sulfurreducens strain KN400, which is even more effective in extracellular electron transfer than strain DL-1. We found that deleting omcS from strain KN400 did not negatively impact the rate of Fe(III) oxide reduction and did not affect the strain’s ability to accept electrons via direct interspecies electron transfer. The OmcS-deficient strain also continued to produce conductive filaments, consistent with the concept that electrically conductive pili are the primary conduit for long-range electron transfer in G. sulfurreducens and closely related species. These findings, coupled with the lack of OmcS homologs in most other microbes capable of extracellular electron transfer, suggest that OmcS is not a common critical component for extracellular electron transfer.

Geobacter anodireducens is unique in that it can generate high current densities in bioelectrochemical systems (BES) operating under high salt conditions. This ability is important for the development of BES treating high salt wastewater and microbial desalination cells. Therefore, the genome of G. anodireducens was characterized to identify proteins that might allow this strain to survive in high salt BES. Comparison to other Geobacter species revealed that 81 of its 87 c-type cytochromes had homologs in G. soli and G. sulfurreducens. Genes coding for many extracellular electron transfer proteins were also detected, including the outer membrane c-type cytochromes OmcS and OmcZ and the soluble c-type cytochrome PgcA. G. anodireducens also appears to have numerous membrane complexes involved in the translocation of protons and sodium ions and channels that provide protection against osmotic shock. In addition, it has more DNA repair genes than most Geobacter species, suggesting that it might be able to more rapidly repair DNA damage caused in high salt and low pH anode environments. Although this genomic analysis provides invaluable insight into mechanisms used by G. anodireducens to survive in high salt BES, genetic, transcriptomic, and proteomic studies will need to be done to validate their roles.

Biological treatment of high salinity organic wastewater is a significant challenge because many microorganisms involved in the anaerobic digestion process cannot survive high osmotic pressures. In order to alleviate some of the stresses associated with the treatment of high salinity wastewater, two lab-scale up-flow anaerobic sludge bed reactors with or without magnetite (100 g/L) were used to treat high salinity organic wastewater. This study showed that the bioreactor amended with magnetite had higher chemical oxygen demand removal efficiencies (90.2% ± 0.54% vs 73.1% ± 1.9%) and methane production rates (4082 ± 334 ml (standard temperature and atmospheric pressure, STP)/d vs 2640 ± 120 ml (STP)/d) than the non-amended control reactor. In addition, the consumption of volatile fatty acids (20.9 ± 3.4 mM vs 61.7 ± 2.0 mM) was accelerated. Microbial community analysis revealed that the addition of magnetite caused the enrichment of many bacterial genera known to form robust biofilms (i.e. Pseudomonas) that are also capable of extracellular electron transfer and methanogens from the genus Methanosarcina which have been shown to participate in direct interspecies electron transfer. These results show that magnetite addition could enhance the performance of anaerobic digesters treating high salinity wastewater.

Chaperone proteins are required to maintain the overall fold and function of proteins in the cell. As part of the Hsp70 family, Ssa1 acts to maintain cellular proteostasis through a variety of diverse pathways aimed to preserve the native conformation of target proteins, thereby preventing aggregation and future states of cellular toxicity. Studying the structural dynamics of Ssa1 in vitro is essential to determining their precise mechanisms and requires the development of purification methods that result in highly pure chaperones. Current methods of expressing and purifying Ssa1 utilize affinity tagged constructs expressed in Escherichia coli, however, expression in an exogenous source produces proteins that lack post-translational modifications leading to undesired structural and functional effects. Current protocols to purify Ssa1 from Saccharomyces cerevisiae require large amounts of starting material, multiple steps of chromatography, and result in low yield. Our objective was to establish a small-scale purification of Ssa1 expressed from its endogenous source, Saccharomyces cerevisiae, with significant yield and purity. We utilized a protein A affinity tag that was previously used to purify large protein complexes from yeast, combined with magnetic Dynabeads that are conjugated with rabbit immunoglobulin G (IgG). Our results show that we can produce native, highly pure, active Ssa1 via this one-step purification with minimal amounts of starting material, and this Ssa1-protein A fusion does not alter cellular phenotypes. This methodology is a significant improvement in Ssa1 purification and will facilitate future experiments that will elucidate the biochemical and biophysical properties of Hsp70 chaperones.

Anaerobically treated leachate from municipal solid waste incineration plants contains extremely high free ammonia (FA) concentrations, which can hinder short-cut nitrification. pH adjustments made...

The five unimolecular HX and DX (X = F, Cl) elimination pathways of CD2ClCHFCl* were examined using a chemical activation technique; the molecules were generated with 92 kcal mol–1 of vibrational energy in a room-temperature bath gas by a combination of CD2Cl and CHFCl radicals. The total unimolecular rate constant was 9.7 × 107 s–1, and branching fractions for each channel were 0.52 (2,1-DCl), 0.29 (1,1-HCl), 0.10 (2,1-DF), 0.07 (1,1-HF), and 0.02 (1,2-HCl). Comparison of the individual experimental rate constants to calculated statistical rate constants gave threshold energies for each process as 63, 72, 66, 73, and 70 kcal mol–1, listed in the same order as the branching fractions. The 1,1-HCl and 1,1-HF reactions gave carbenes, CD2Cl(F)C: and CD2Cl(Cl)C:, respectively, as products, which have hydrogen-bonded complexes with HCl or HF in the exit channel of the potential energy surface. These carbenes have energy in excess of the threshold energy for D atom migration to give CDCl═CDF and CDCl═CDCl, and ...

Anaerobic digestion of municipal solid waste (MSW) incineration leachate is limited by high organic loading rates, high concentrations of complex organic matter, and the presence of compounds like ammonia, heavy metals, and salts that can inhibit microbial activity. The purpose of this study was to investigate whether the addition of magnetite could help reactors treating MSW incineration leachate overcome these limitations and promote efficient conversion of complex organic matter to methane. These studies showed that magnetite amendments improved chemical oxygen demand (COD) removal efficiencies (78.8% vs 89.0%) and methane production rates (3.7 m3STP/(m3·d) vs 4.8 m3STP/(m3·d)) at an organic loading rate (OLR) of 18.2 kg COD/(m3·d). Significant differences in microbial community structures were also observed between non-amended and magnetite-amended reactors. The majority of sequences from the magnetite-amended reactor clustered with acetogenic and mixed acid fermentative bacteria. Sequences from bacteria capable of extracellular electron transfer and methanogens from the genera Methanosarcina and Methanosaeta were also more abundant in magnetite-amended reactors. It is possible that these species were participating in direct interspecies electron transfer (DIET) facilitated by magnetite amendments. In addition, ammonia nitrogen, calcium, and heavy metals were lower in effluent collected from magnetite-amended reactors than non-amended controls, suggesting that magnetite adsorbed these inhibitory compounds and created an environment more amenable to microbial growth. These results are significant and should be used to develop strategies designed to optimize bio-methanogenic treatment of complex waste with elevated contaminant concentrations.

Nitrous oxide (N2O) is formed during wastewater nitrogen removal processes. It is a strong greenhouse gas, however, if properly captured it can also be used as a renewable energy source. In this study, a nosZ-deficient strain of Pseudomonas aeruginosa was constructed. During growth under denitrifying conditions, the nosZ-deficient strain was more highly transcribing other genes from the denitrification pathway (narG, nirS, and norB) than the wild-type strain. This strain could also convert 85% of NO2−-N to N2O when it was grown with acetate compared to 2−-N concentrations (700 mg/L) was inoculated with this strain, the N2O conversion efficiencies were >73% and N2O comprised 73~81% of the biogas being generated. The energy yield from wastewater in bioaugmented reactors also reached levels as high as 1260 kJ/m3. These results are significant and show that bioaugmentation of reactors during denitrification treatment processes with nosZ-deficient strains of Pseudomonas or other core denitrifying bacteria might be an effective way to enhance N2O recovery.

Solid supported colorimetric sensing arrays have the advantage of portability and ease of use when deployed in the field, such as crime scenes, disaster zones, or in war zones, but many sensor arrays require complex fabrication methods. Here, we report a practical method for the fabrication of 4 × 4 colorimetric sensor arrays, which are printed on nylon membranes, using a commercially available inkjet printer. In order to test the efficacy of the printed arrays, they were exposed to 43 analytes at concentrations ranging from 0.001 to 3.0 M for a total of 559 samples of inorganic and organic acids or bases including hydrochloric, acetic, phthalic, malonic, picric, and trifluoroacetic acid, ammonium hydroxide, sodium hydroxide, lysine, and water as the control. Colorimetric data from the imaged arrays was analyzed with linear discriminant analysis and k-nearest neighbors to determine the analyte and concentration with ∼88-90% accuracy. Overall, the arrays have impressive analytical power to identify a variety of analytes at different concentrations while being simple to fabricate.

The recombination of CF3 and CHF2 radicals in a room-temperature bath gas was used to prepare vibrationally excited CF3CHF2* molecules with 101 kcal mol–1 of vibrational energy. The subsequent 1,2-H atom transfer and 1,1-HF and 1,2-HF elimination reactions were observed as a function of bath gas pressure by following the CHF3, CF3(F)C: and C2F4 product concentrations by gas chromatography using a mass spectrometer as the detector. The singlet CF3(F)C: concentration was measured by trapping the carbene with trans-2-butene. The experimental rate constants are 3.6 × 104, 4.7 × 104, and 1.1 × 104 s–1 for the 1,2-H atom transfer and 1,1-HF and 1,2-HF elimination reactions, respectively. These experimental rate constants were matched to statistical RRKM calculated rate constants to assign threshold energies (E0) of 88 ± 2, 88 ± 2, and 87 ± 2 kcal mol–1 to the three reactions. Pentafluoroethane is the only fluoroethane that has a competitive H atom transfer decomposition reaction, and it is the only example with...

Growth of bacterial and archaeal species capable of interspecies electron exchange was stimulated by addition of conductive materials (carbon cloth or granular activated carbon (GAC)) to anaerobic digesters treating dog food (a substitute for the dry-organic fraction of municipal solid waste (OFMSW)). Methane production (772-1428mmol vs

PurposeThe aim of this work was to develop a quantitative, flow cytometric method for tracking the endolysosomal escape of a fluorescently labelled saporin toxin.MethodsFlow cytometric measurements of fluorescent pulse width and height were used to track the endocytic uptake into Daudi cells of a fluorescently labelled saporin toxin and the saporin based immunotoxin, OKT10-SAP. Subsequently, measurement of changes in pulse width were used to investigate the effect of a triterpenoid saponin on the endolysosomal escape of internalised toxin into the cytosol. Live cell confocal microscopy was used to validate the flow cytometry data.ResultsIncreased endolysosomal escape of saporin and OKT10-SAP was observed by confocal microscopy in cells treated with saponin. Fluorescent pulse width measurements were also able to detect and quantify escape more sensitively than confocal microscopy. Saponin induced endolysosomal escape could be abrogated by treatment with chloroquine, an inhibitor of endolysosomal acidification. Chloroquine abrogation of escape was also mirrored by a concomitant abrogation of cytotoxicity.ConclusionsPoor endolysosomal escape is often a rate limiting step for the cytosolic delivery of protein toxins and other macromolecules. Pulse width analysis offers a simple method to semi-quantify the endolysosomal escape of this and similar molecules into the cytosol.

Anaerobic dynamic membrane bioreactors (AnDMBRs) can improve the efficiency of organic matter removal during wastewater treatment at a low cost. However, application of AnDMBRs for treatment of high-strength wastewater is usually unsuccessful. This study investigated whether use of conductive carbon cloth as the supporting material in an AnDMBR permits higher organic loading rates for treatment of brewery wastewater than non-conductive polyester cloth. The AnDMBR with carbon cloth operated stably with a COD removal efficiency of 98% even when high concentrations of influent COD (10,000 mg/L) were provided, while the polyester cloth reactor deteriorated when reactors were fed only 5000 mg/L influent COD. Microorganisms capable of direct interspecies electron transfer (DIET), including Geobacter and Methanothrix species, dominated the surface of the carbon cloth. These results demonstrate that carbon cloth provides an excellent supporting material for AnDMBRs by stimulating growth of microorganisms that can directly transport electrons to and from conductive materials.

In this study, nitrous oxide was recovered from a lab-scale moving-bed biofilm reactor (MBBR) treating partial nitrification-treated leachate supplemented with a nosZ-deficient strain of Pseudomonas aeruginosa. Batch culture tests with the nosZ-deficient strain determined that the threshold for free nitrous acid (FNA) inhibition was 0.016 mg/L and that FNA concentrations above this threshold severely inhibited denitrification and transcription of genes from the dissimilatory nitrate reduction pathway (narG, nirS, and norB). High nitrite removal and N2O conversion efficiencies (>95%) were achieved with long-term operation of this MBBR. N2O accounted for the majority of biogas (80%) produced when the MBBR was fed partial nitrification-treated leachate with high nitrite concentrations and the drainage ratio was adjusted to 30%. Bacterial community analysis revealed that the nosZ-deficient Pseudomonas strain remained metabolically active and was primarily responsible for denitrification processes in the reactor. This study presents a promising method for N2O recovery from incineration leachate.

In order to illustrate the impact that application of positive or negative potential to conductive materials can have on direct interspecies electron transfer (DIET) and reactor performance under high organic loading rates, three continuous laboratory-scale reactors with carbon-cloth electrodes poised at +0.7 V, −0.7 V (vs. Ag/AgCl) and no-potential were fed high concentrations of ethanol wastewater. While exoelectrogens and methanogens that are capable of DIET were significantly enriched in poised reactors, they performed worse than the non-current control. Volatile fatty acids (VFAs) accumulated more rapidly in the positively then negatively poised reactor, but neither could withstand high-loading rates. These results demonstrate that applying potential to conductive materials had a negative effect on anaerobic digestion under high-loading conditions.

The discovery of a methanogen that can conserve energy to support growth solely from the oxidation of organic carbon coupled to the reduction of an extracellular electron acceptor expands the possible environments in which methanogens might thrive. The potential importance of c-type cytochromes for extracellular electron transfer to syntrophic bacterial partners and/or Fe(III) minerals in some Archaea was previously proposed, but these studies with Methanosarcina acetivorans provide the first genetic evidence for cytochrome-based extracellular electron transfer in Archaea. The results suggest parallels with Gram-negative bacteria, such as Shewanella and Geobacter species, in which multiheme outer-surface c-type cytochromes are an essential component for electrical communication with the extracellular environment. M. acetivorans offers an unprecedented opportunity to study mechanisms for energy conservation from the anaerobic oxidation of one-carbon organic compounds coupled to extracellular electron transfer in Archaea with implications not only for methanogens but possibly also for Archaea that anaerobically oxidize methane., Extracellular electron exchange in Methanosarcina species and closely related Archaea plays an important role in the global carbon cycle and enhances the speed and stability of anaerobic digestion by facilitating efficient syntrophic interactions. Here, we grew Methanosarcina acetivorans with methanol provided as the electron donor and the humic analogue, anthraquione-2,6-disulfonate (AQDS), provided as the electron acceptor when methane production was inhibited with bromoethanesulfonate. AQDS was reduced with simultaneous methane production in the absence of bromoethanesulfonate. Transcriptomics revealed that expression of the gene for the transmembrane, multiheme, c-type cytochrome MmcA was higher in AQDS-respiring cells than in cells performing methylotrophic methanogenesis. A strain in which the gene for MmcA was deleted failed to grow via AQDS reduction but grew with the conversion of methanol or acetate to methane, suggesting that MmcA has a specialized role as a conduit for extracellular electron transfer. Enhanced expression of genes for methanol conversion to methyl-coenzyme M and the Rnf complex suggested that methanol is oxidized to carbon dioxide in AQDS-respiring cells through a pathway that is similar to methyl-coenzyme M oxidation in methanogenic cells. However, during AQDS respiration the Rnf complex and reduced methanophenazine probably transfer electrons to MmcA, which functions as the terminal reductase for AQDS reduction. Extracellular electron transfer may enable the survival of methanogens in dynamic environments in which oxidized humic substances and Fe(III) oxides are intermittently available. The availability of tools for genetic manipulation of M. acetivorans makes it an excellent model microbe for evaluating c-type cytochrome-dependent extracellular electron transfer in Archaea.

While studies have shown that anaerobic co-digestion of chicken manure (CM) and corn stover (CS) is an efficient method to treat these agricultural wastes, the microbial ecology of these systems and optimal parameters for the digestion process are yet to be determined. In this study, the effects of different initial substrate concentrations and CS : CM mixture ratios on co-digestion and microbial community structure were evaluated. Results demonstrated that both the highest cumulative methane yields and methane production rates were obtained from reactors with a CS : CM ratio of 1 : 1 during hemi-solid-state anaerobic digestion (HSS-AD). Cumulative methane yields and methane production rates were 24.8% and 42% lower in solid-state anaerobic digestion (SS-AD) reactors using the same CS : CM ratios. Analysis of microbial community structures revealed that cellulolytic bacteria and a diversity of syntrophic microorganisms capable of direct interspecies electron transfer (DIET) and hydrogen interspecies transfer (HIT) were enriched in the best-performing reactors. Methanosarcina species also dominated during HSS-AD, and their presence was positively correlated with methane production in the reactors.

Bioelectrochemical conversion of CO2 to CH4 is a promising way to increase the calorific value of biogas produced during anaerobic digestion. There are two groups of methanogens enriched in these systems, hydrogenotrophs and acetoclastic methanogens that can also directly accept electrons from an electrode or another microorganism. In this study, a microbial electrolysis cell (MEC) poised at −500 mV (vs. SHE) was operated for biogas upgrading. Methane content in the biogas increased from 71% to >90%, and 8.2% of the CO2 was converted to methane. Methanothrix, an acetoclastic methanogen that can participate in direct electron transfer (DET), and Azonexus, an acetate-oxidizing electrogen, were enriched on the cathode. Transcriptomics revealed that Methanothrix on the cathode were using the CO2 reduction pathway, while Methanothrix in the bulk sludge were using the acetate decarboxylation pathway for production of methane. These results show that stimulation of DET in MEC enhances biogas-upgrading processes.

The glucocorticoid receptor (GR) binds the noncoding RNA Gas5 via its DNA-binding domain (DBD) with functional implications in pro-apoptosis signaling. Here, we report a comprehensive in vitro binding study where we have determined that GR-DBD is a robust structure-specific RNA-binding domain. GR-DBD binds to a diverse range of RNA hairpin motifs, both synthetic and biologically derived, with apparent mid-nanomolar affinity while discriminating against uniform dsRNA. As opposed to dimeric recognition of dsDNA, GR-DBD binds to RNA as a monomer and confers high affinity primarily through electrostatic contacts. GR-DBD adopts a discrete RNA-bound state, as assessed by NMR, distinct from both free and DNA-bound. NMR and alanine mutagenesis suggest a heightened involvement of the C-terminal α-helix of the GR-DBD in RNA-binding. RNA competes for binding with dsDNA and occurs in a similar affinity range as dimer binding to the canonical DNA element. Given the prevalence of RNA hairpins within the transcriptome, our findings strongly suggest that many RNAs have potential to impact GR biology.

The anaerobic corrosion of iron structures is expensive to repair and can be a safety and environmental concern. It has been known for over 100 years that the presence of anaerobic respiratory microorganisms can accelerate iron corrosion. Multiple studies have suggested that there are sulfate reducers, methanogens, and acetogens that can directly accept electrons from Fe(0) to support sulfate or carbon dioxide reduction. However, all of the strains studied can also use H2 as an electron donor for growth, which is known to be abiotically produced from Fe(0). Furthermore, no proteins definitely shown to function as extracellular electrical contacts with Fe(0) were identified. The studies described here demonstrate that direct electron transfer from Fe(0) can support anaerobic respiration. They also map out a simple genetic approach to the study of iron corrosion mechanisms in other microorganisms. A better understanding of how microorganisms promote iron corrosion is expected to lead to the development of strategies that can help reduce adverse impacts from this process., The concept that anaerobic microorganisms can directly accept electrons from Fe(0) has been controversial because direct metal-microbe electron transfer has previously only been indirectly inferred. Fe(0) oxidation was studied with Geobacter sulfurreducens strain ACL, an autotrophic strain that was previously shown to grow with electrons derived from a graphite cathode as the sole electron donor. Strain ACL grew with Fe(0) as the sole electron donor and fumarate as the electron acceptor. However, it appeared that at least a portion of the electron transfer was via H2 produced nonenzymatically from the oxidation of Fe(0) to Fe(II). H2, which accumulated in abiotic controls, was consumed during the growth of strain ACL, the cells were predominately planktonic, and genes for the uptake hydrogenase were highly expressed. Strain ACLHF was constructed to prevent growth on H2 or formate by deleting the genes for the uptake of hydrogenase and formate dehydrogenases from strain ACL. Strain ACLHF also grew with Fe(0) as the sole electron donor, but H2 accumulated in the culture, and cells heavily colonized Fe(0) surfaces with no visible planktonic growth. Transcriptomics suggested that the outer surface c-type cytochromes OmcS and OmcZ were important during growth of strain ACLHF on Fe(0). Strain ACLHF did not grow on Fe(0) if the gene for either of these cytochromes was deleted. The specific attachment of strain ACLHF to Fe(0), coupled with requirements for known extracellular electrical contacts, suggest that direct metal-microbe electron transfer is the most likely option for Fe(0) serving as an electron donor.

Conservation of energy to support growth solely from extracellular electron transfer was demonstrated for the first time in a methanogen.Methanosarcina acetivoransgrew with methanol as the sole electron donor and the extracellular electron acceptor anthraquione-2,6-disulfonate (AQDS) as the sole electron acceptor when methane production was inhibited with bromoethanesulfonate. Transcriptomics revealed that transcripts for the gene for the transmembrane, multi-heme,c-type cytochrome MmcA were 4-fold higher in AQDS-respiring cells versus methanogenic cells. A strain in which the gene for MmcA was deleted failed to grow via AQDS reduction whereas strains in which other cytochrome genes were deleted grew as well as the wild-type strain. The MmcA-deficient strain grew with the conversion of methanol or acetate to methane, suggesting that MmcA has a specialized role as a conduit for extracellular electron transfer. Enhanced expression of genes for methanol conversion to methyl-coenzyme M and components of the Rnf complex suggested that methanol is oxidized to carbon dioxide in AQDS-respiring cells through a pathway that is similar to methyl-coenezyme M oxidation in methanogenic cells. However, during AQDS respiration the Rnf complex and reduced methanophenazine probably transfer electrons to MmcA, which functions as the terminal reductase for AQDS reduction. Extracellular electron transfer may enable survival of methanogens in dynamic environments in which oxidized humic substances and Fe(III) oxides are intermittently available. The availability of tools for genetic manipulation ofM. acetivoransmakes it an excellent model microbe for evaluatingc-type cytochrome-dependent extracellular electron transfer in Archaea.ImportanceExtracellular electron exchange inMethanosarcinaspecies and closely related Archaea plays an important role in the global carbon cycle and can enhance the speed and stability of anaerobic digestion, an important bioenergy strategy. The potential importance ofc-type cytochromes for extracellular electron transfer to syntrophic bacterial partners and/or Fe(III) minerals in some Archaea has been suspected for some time, but the studies withMethanosarcina acetivoransreported here provide the first genetic evidence supporting this hypothesis. The results suggest parallels with Gram-negative bacteria, such asShewanellaandGeobacterspecies, in which outer-surfacec-type cytochromes are an essential component for electrical communication with the extracellular environment.M. acetivoransoffers an unprecedented opportunity to study mechanisms for energy conservation from the anaerobic oxidation of one-carbon organic compounds coupled to extracellular electron transfer in Archaea with implications not only for methanogens, but possibly also for anaerobic methane oxidation.

The raw fresh leachate from municipal solid waste (MSW) incineration plants contains high concentrations of volatile fatty acids (VFAs), ammonia and metals, all compounds that severely limit anaerobic digestion treatment efficiencies. These inhibitory compounds make reactor systems unstable, causing reactor start-up periods to take more than 100 days, even when the leachate is diluted significantly. In this study, granular activated carbon (GAC) was incorporated into a bioreactor fed with raw incineration leachate. Addition of GAC allowed direct treatment of raw incineration leachate without any start-up acclimation period, while the non-amended control reactor soured immediately and collapsed within 17 days. When hydraulic retention time (HRT) of the GAC-amended reactor was stepwise decreased to increase organic loading rates (OLR) to 25.0 kgCOD/(m3·d), COD removal efficiencies remained stable at >90%. Metagenomic analysis of the GAC-amended reactor revealed that Geobacter and Methanosarcina, species known to participate in direct interspecies electron transfer (DIET), were more abundant in the GAC-amended reactor than the seed sludge. In addition, the abundance of genes coding for proteins thought to be involved in DIET such as electrically conductive pili and the outer membrane c-type cytochrome, OmcS, increased significantly, while genes involved in fermentation, and nitrate (narG) and sulfate (dsrA) reduction dropped significantly as the experiment progressed. These results are significant because this is the first detailed investigation into the metabolic capabilities of microbial communities involved in efficient treatment of raw incineration leachate within biomethanogenic reactors that did not require a long start-up period.

We have previously shown that antibody-dependent cellular cytotoxicity (ADCC) cooperates with immunotoxin (IT)-mediated killing of human leukaemia cells in an severe combined immunodeficient (SCID) mouse model of human T-cell acute lymphoblastic leukaemia (SCID-HSB-2 mice), but not in an equivalent non-obese diabetic (NOD)/SCID mouse model. In these earlier studies, we reasoned that diminished ADCC due to the functional deficit in natural killer (NK) cell activity in NOD/SCID mice resulted in a failure of effective perforin/granzyme-mediated cytotoxicity necessary for the delivery of the augmentative effect. Poly-inosinic-cytidylic acid [poly (I:C)] is a synthetic dsRNA toll-like receptor 3 (TLR3) agonist that possesses a number of biological properties that includes the in vivo activation of NK cells. We show here that intravenous (i.v.) injection of SCID mice with [poly (I:C)] results in characteristic time-related changes in serum interleukin 2 (IL-2), IL-12, and interferon &gamma, (INF&gamma, ) cytokine levels that are consistent with TLR3 driven activation of SCID mouse NK cells. Concomitantly, there are changes in the expression levels of CD2, CD16/32 (Fc&gamma, RII/RIII), CD161 (NK1.1), and F4/80 in the bulk splenocyte population. These observed changes correlate with an increase in the in vitro lytic capabilities of putative NK cells from within the splenocyte population of [poly (I:C)] treated SCID mice. We demonstrate that the in vivo activation of NK cells with [poly (I:C)] in SCID mice bearing disseminated human T-cell leukaemia xenografts resulted in a significant improvement in the therapeutic activity exerted by an intact murine monoclonal antibody against human CD7. This was also seen for a saporin-based immunotoxin constructed with the same intact antibody (HB2-SAPORIN), but not with an F(ab&rsquo, )2 derivative of the same antibody or of an IT constructed with the same F(ab&rsquo, )2 HB2 antibody derivative. This study further demonstrates the previously reported reinforcing role of ADCC for the therapeutic activity of IT in an SCID mouse model of human T-ALL and the potential to significantly boost this further with [poly (I:C)]. Our study provides the rationale to justify the exploration of the clinical utility of IT based therapeutics in combination with TLR3 agonists, such as [poly (I:C)], for the treatment of haematological, and possibly other, malignancies.

Biogas produced from anaerobic digestion usually contains 30%-50% CO2, much of which must be removed, before utilization. Bioelectrochemical biogas upgrading approaches show promise, however, they have not yet been optimized for practical applications. In this study, a bioelectrochemical system with low energy input (applied cathode potential of -0.5 V vs. standard hydrogen electrode, SHE) was used for in-situ biogas upgrading. High efficiency CO2 conversion (318.5 mol/d/m2) was achieved when the system was operated with an organic load of 1.7 kgCOD/(m3 d). Methane content in the upgraded biogas was 97.0% and CO2 concentrations stayed below 3%, which is comparable to biogas upgraded with more expensive and less sustainable physiochemical approaches. The high efficiency of this approach could likely be attributed to a significant enrichment of Methanothrix (92.7%) species on the cathode surface that were expressing genes involved in both acetogenic methanogenesis and direct electron transfer (DET). Electromethanogenesis by these organisms also increased proton consumption and created a higher pH that increased the solubility of CO2 in the bioreactor. In addition, CO2 removal from the biogas was likely further enhanced by an enrichment of Actinobacillus species known to be capable of CO2 fixation. Artificial neural network (ANN) models were also used to estimate CH4 production under different loading conditions. The ANN architecture with 10 neurons at hidden layers fit best with a mean square error of 6.06 × 10−3 and R2 of 0.99.

In this study, an Escherichia coli (E. coli) whole-cell biosensor for the specific detection of bioavailable arsenic was developed by placing a green fluorescent protein (GFP) reporter gene under the control of the ArsR1 (GSU2952) regulatory circuit from Geobacter sulfurreducens. E. coli cells only emitted green fluorescence in the presence of arsenite and were more sensitive to arsenite when they were grown in M9 supplemented medium compared to LB medium. Under optimal test conditions, the Geobacter arsR1 promoter had a detection limit of 0.01 μM arsenite and the GFP expression was linear within a range of 0.03–0.1 μM (2.25–7.5 μg/l). These values were well below World Health Organization’s drinking water quality standard, which is 10 μg/l. The feasibility of using this whole-cell biosensor to detect arsenic in water samples, such as arsenic polluted tap water and landfill leachate was verified. The biosensor was determined to be just as sensitive as atomic fluorescence spectrometry. This study examines the potential applications of biosensors constructed with Geobacter ArsR-Pars regulatory circuits and provides a rapid and cost-effective tool that can be used for arsenic detection in water samples.

Triterpenoid saponins augment the cytotoxicity of saporin based immunotoxins. It is postulated that this results from a saponin-mediated increase in the endolysosomal escape of the toxin to the cytosol, but this remains to be confirmed. To address this issue, we used a number of pharmacological inhibitors of endocytic processes as probes to investigate the role played by saponin in the endolysosomal escape of fluorescently labeled saporin and a saporin based immunotoxin targeted against CD38 on human lymphoma and leukemia cell lines. Endolysosomal escape of the toxin was measured by flow cytometric pulse shape analysis. These results were compared to the effects of the various inhibitors on the saponin-mediated augmentation of toxin and immunotoxin cytotoxicity. Inhibitors of clathrin-mediated endocytosis, micropinocytosis, and endosomal acidification abrogated the saponin-induced increase in the endolysosomal escape of the toxin into the cytosol, suggesting that these processes may be involved in the internalization of saponin to the same endolysosomal vesicle as the toxin. Alternatively, these processes may play a direct role in the mechanism by which saponin promotes toxin escape from the endolysosomal compartment to the cytosol. Correlation with the effects of these inhibitors on the augmentation of cytotoxicity provides additional evidence that endolysosomal escape is involved in driving augmentation.