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Energy is an important requirement for population growth, technological progress and urbanisation. Worldwide energy demand has been projected to increase 5-fold by 2100. Fulfilment of these energy requirements cannot be solely reliant on fossil fuels, such as oil, coal and natural gas, on account of their adverse environmental impacts and concomitant depletion of natural resources. As a result multiple approaches for generating alternative energy are being explored globally.In this review paper, focused on the viability of waste especially MSW being a source for bioenergy products such as methane gas, bio-enzyme, biofuel and bio-fertiliser production. This review also focuses on the environmental impacts of MSW, the effect of MSW pre-treatments and properties (physical and chemical) on bioenergy products production.

Miscanthus is a fast-growing perennial grass that attracts significant attention for its potential application as a feedstock for bioethanol production. This report explores the difference in the lignocellulosic composition of various Miscanthus cultivars, including Miscanthus × giganteus cultivated at the same location (mainly Lincoln, UK). It also assesses the sugar release profiles and mineral composition profiles of five Miscanthus cultivars harvested over a growing period from November 2018 to February 2019. The results showed that Miscanthus × giganteus contains approximately 45.5% cellulose, 29.2% hemicellulose and 23.8% lignin (dry weight, w/w). Other cultivars of Miscanthus also contain high quantities of carbohydrates (cellulose 41.1–46.0%, hemicellulose 24.3–32.6% and lignin 21.4–24.9%). Pre-treatment of Miscanthus using dilute acid followed by enzymatic hydrolysis released 63.7–80.2% of the theoretical glucose content. Fermentation of a hydrolysate of Miscanthus × giganteus using Saccharomyces cerevisiae NCYC2592 produced 13.58 ± 1.11 g/L of ethanol from 35.13 ± 0.46 g/L of glucose, corresponding to a yield of 0.148 g/g dry weight Miscanthus biomass. Scanning electron microscopy was used to study the morphology of raw and hydrolysed Miscanthus samples, which provided visual proof of Miscanthus lignocellulose degradation in these processes. The sugar release profile showed that a consequence of Miscanthus plant growth is an increase in difficulty in releasing monosaccharides from the biomass. The potassium, magnesium, sodium, sulphur and phosphorus contents in various Miscanthus cultivars were analysed. The results revealed that these elements were slowly lost from the plants during the latter part of the growing season, for a specific cultivar, until February 2019.

This review shows the potential of algae in combating both air and water pollution and the production of biofuel feedstock in a much cleaner manner. Various research articles published in leading journals have been studied thoroughly to give a critical mentioning of various methods of cultivation in different reactors. It provides a comprehensive narration about a) selection of suitable algal species b) selection of a suitable mode for growing c) selection of suitable reactor type and d) carbon bio-fixation. This review found that hybrid airlift photobioreactors are highly efficient because it exploits the benefits of two different types of reactors to overcome the drawbacks. It also provides salient features in the selection of suitable microalgal species. The mixotrophic culture mode is found to be best for the integrated approach. The models used to calculate carbon dioxide bio-fixation rate would be useful for engineers and researchers to quantify the carbon removal.

Inhibitor tolerance is one of the key challenges in lignocellulosic bioethanol production. The presence of inhibitors in a lignocellulosic hydrolysate induces stress in microorganisms and thus reduces bioethanol synthesis efficiency. In this study, 166 marine yeasts isolated from different marine environments were compared with 78 terrestrial yeasts for their tolerance to inhibitory compounds such as acetic acid, formic acid, furfural, vanillin and salt, which are commonly found in hydrolysates derived from lignocellulosic materials. Marine yeasts showed higher tolerance to all the inhibitors tested than terrestrial yeasts. The most tolerant marine yeast was Wickerhamomyces anomalus M15 which had IC50 values of 10.7% (w/w) and 83.9 mM for salt and acetic acid, respectively, while those for an industrial terrestrial yeast Saccharomyces cerevisiae NCYC2592 were 6.0% (w/w) and 75.9 mM, respectively.Statistical analysis revealed that marine yeast clustered separately from terrestrial yeast. In fermentation, using simulated wheat straw hydrolysates containing inhibitors and prepared using seawater, three selected marine yeasts Saccharomyces cerevisiae AZ65, W. anomalus M15, and Candidamembranifaciens M2 produced 23–24 g L1 bioethanol from 60 g L1 glucose, while terrestrial yeast Saccharomyces cerevisiae produced only 12.5 g L1 bioethanol. Marine yeasts were significantly more tolerant to the presence of inhibitory compounds and are phenotypically distinct from terrestrial yeasts.Significantly, a higher quantity of bioethanol was obtained in fermentation using selected marine yeasts with media containing inhibitory compounds. The study has highlighted the potential of finding microorganisms with industrially favourable characteristics in marine ecosystems and other adverse and challenging environments

This review introduces a new approach of completely marine based bioethanol production by analyzing and evaluating the recent trends in bioethanol fermentations using algae, marine microorganisms and the replacement of freshwater with seawater. Both macroalgae and microalgae have been successfully used for bioethanol production. Marine yeasts showed excellent tolerance to salt and inhibitors, and fit for seawater fermentation. The combination of marine biomass, marine microorganism and seawater has a potential for a greener bioethanol production.

Lignocellulosic feedstocks for biorefinery are likely to be seasonal and the supply of feedstock to cellulosic biorefineries remains a challenge. One way to overcome this is by utilizing a mixed feedstock which facilitates the maintenance of a year-round feedstock supply. This study investigated the impact of mixing of three industrially relevant cellulosic feedstocks—wheat straw, willow and Miscanthus using two major performance indicators—sugar yield and fermentation inhibitor production. A microwave hydrothermal pre-treatment regime of 200 °C for 5 min was applied to each feedstock individually and to 1:1 (w/w) mixes and the predicted sugar yield in the mixes was compared to the observed values. All the mixes resulted in improved sugar yields with willow + Miscanthus (15.4%, p = 0.015) and wheat + willow (13.6%, p = 0.010) showing a statistically significant improvement. Saccharification kinetics, inhibitor production, impact on yeast metabolic activity and growth were compared and no adverse impacts of mixing were observed. The use of mixed feedstocks in a hot water based commercial production of biofuels is unlikely to have any adverse effects on productivity and may indeed prove beneficial.

Worldwide significant amounts of food waste are generated daily causing serious environmental issues, occupying land and requiring expenditure of resources for its treatment. A smart method for handling this food waste problem is the development of novel processes targeting the conversion of this waste into value added products. Although valorization of food waste to biofuels, biochemicals and bio-polymers have been widely investigated, the utilization of food waste streams into biofertiliser has not been intensively reviewed. Conversion of food waste, especially agriculture residues into biofertiliser would reduce its environmental impact, improve nutrition levels of the soil, decrease requirements for synthetic chemical fertiliser and have a direct benefit on food production. This paper reviews recent progress in the field regarding the production of biofertiliser from food waste, using anaerobic digestion, aerobic composting, chemical hydrolysis, in situ degradation and direct burning methods. This review also highlights the latest field applications of biofertiliser derived from various food waste streams. It confirms that the technology for the conversion of food waste to biofertilisers is viable, but the production efficiency could be improved with better process control strategies, strict quality controls, development of a smart product distribution system and adoption of advanced technologies. Field tests have indicated that biofertilisers which are obtained in proper managed AD plants are safe and could partially replace the use of chemical fertilisers in field application.

Treatment of Municipal Solid Waste (MSW) becomes an increasing problem worldwide; however, MSW contains high lignocellulosic content that can be further processed for the generation of electricity, transportation fuel and other commercially important products. This study aims to explore the possibility of hydrolyzing MSW into a sugar solution, which could then be fermented into liquid biofuels, such as bioethanol. Two commercial cellulase enzymes, Cellic® CTec2, and a fresh prepared fungal filtrate were used to hydrolyse MSW. Results revealed that CTec2 displayed the best hydrolysis efficiency. Use of an alkali pre-treatment on MSW before enzymatic hydrolysis failed to show any improvement in terms of release of sugars, an assessment of the efficacy of addition of surfactants revealed that addition of tween 80 or polyethylglycol had a no significant effect on release of sugars. Results revealed that by increasing the solid liquid ratio the amount of glucose increased, Data generated by this study was analysed using design expert stat program.

Analysis of the chemical components of Municipal solid waste (MSW) indicated that the presence of high concentrations of toxic heavy metals. The aim of this study was to select the best leaching condition, in order to generate a compost product, which adheres to the environmental standards for safe use. To determine the role of organic acids produced in the bioleaching process, chemical leaching experiments were compared using inorganic (H2SO4) or organic acids (oxalic, and gluconic acids). These results revealed that addition of H2SO4 correlated with a lower retained yield when comparing with addition of the other acids. Bioleaching was then studied by culturing fungi on the MSW. Addition of Aspergillus niger correlated with an efficient removal of heavy metal elements. The effect of pH was also determined and assays at pH 5.5 using a clay medium (32.4%) or at pH 2.5 using a sorghum medium (28%) were characterised by a lower retained yield. In the above conditions, we looked at elemental ions which have been deemed beneficial to the environment (P and K) and those which are toxic (Cd, Cu,Cr, Ni, Pb and Zn), these elements were measured in the solid fractions.

A novel seawater-based pretreatment process was developed to improve the hydrolysis yield of brown (Laminaria digitata), green (Ulva linza) and red (Porphyra umbilicalis) macroalgae. Pre-treated with 5% sulphuric acid at 121 °C, 15 minutes, L. digitata, U. linza and P. umbilicalis liberated 64.63 ± 0.30%, 69.19 ± 0.11% and 63.03 ± 0.04% sugar in seawater compared with 52.82 ± 0.16%, 45.93 ± 0.37% and 48.60 ± 0.07% in reverse-osmosis water, respectively. Low hydrolysis yields (2.6–11.7%) were observed in alkali and hydrothermal pretreatment of macroalgae, although seawater led to relatively higher yields. SEM images of hydrolyzed macroalgae showed that reverse-osmosis water caused contortions in the remaining cell walls following acid and hydrothermal pre-treatments in the L. digitata and U. linza samples. Fed-batch fermentations using concentrated green seaweed hydrolysates and seawater with marine yeast Wickerhamomyces anomalus M15 produced 48.24 ± 0.01 g/L ethanol with an overall yield of 0.329 g/g available sugars. Overall, using seawater in hydrolysis of seaweed increased sugar hydrolysis yield and subsequent bioethanol production.

Industrial production of bioethanol from lignocellulosic materials (LCM&prime, s) is reliant on a microorganism being tolerant to the stresses inherent to fermentation. Previous work has highlighted the importance of a cytochrome oxidase chaperone gene (COX20) in improving yeast tolerance to acetic acid, a common inhibitory compound produced during pre-treatment of LCM&rsquo, s. The presence of acetic acid has been shown to induce oxidative stress and programmed cell death, so the role of COX20 in oxidative stress was determined. Analysis using flow cytometry revealed that COX20 expression was associated with reduced levels of reactive oxygen species (ROS) in hydrogen peroxide and metal-induced stress, and there was a reduction in apoptotic and necrotic cells when compared with a strain without COX20. Results on the functionality of COX20 have revealed that overexpression of COX20 induced respiratory growth in &Delta, imp1 and &Delta, cox18, two genes whose presence is essential for yeast respiratory growth. COX20 also has a role in protecting the yeast cell against programmed cell death.

Background Hydrolysates derived from lignocellulosic material contain a complex mix of inhibitory compounds dependant on the type of biomass and the pre-treatment process employed. These inhibitors prevent the subsequent fermentation of available sugars by yeast into ethanol. Results Inhibitory compounds normally work synergistically to reduce metabolic output, rates of budding and viability; however, it was observed in this study that the presence of weak acids actually improved tolerance to hydroxymethyl furfural (HMF) and furfural in Saccharomyces cerevisiae . The protective role of weak acids in HMF or furfural stressed cells was only apparent with relatively low concentrations of acetic acid (20 mM), however, there was an improvement in glucose utilisation and ethanol production when compared with HMF or furfural stressed cells. Focusing on HMF stressed cells quantitative trait loci (QTL) analysis identified a region on chromosome VI related to the enhanced tolerance to HMF in the presence of acetic acid. Two genes FET5 and HAC1 located in this region were up-regulated under the combined stress of acetic acid with HMF stress and null mutants exhibited a return to HMF sensitivity. Conclusions Presence of acetic acid helps yeast cells overcome HMF stress, QTL analysis identified two genes on a loci on chromosome VI, knocking out these genes returns the cell to HMF sensitivity.

Bioethanol generated from household, agricultural or forestry waste is currently being pursued as the second generation biofuel with wheat straw being highlighted as a promising lignocellulosic residue feedstock in the UK. Natural fermentative ecosystems such as agricultural manure and forest floors containing decomposing wood and leaf litter are potential sources of yeast strains with suitable phenotypic characteristics for improved production of bioethanol. Identifying yeast with innate fermentation capabilities could potentially contain a pool of genes that confer resistance to the chemical constituents of lignocellulosic biomass. Wild yeasts were analysed using a phenotypic microarray for utilization of xylose and tolerance to pre-treatment inhibitors. Assays identified yeast which had stress tolerant phenotypes and yeast with xylose utilization capabilities. Performance fermentations confirmed desirable phenotypes such as conversion of xylose into ethanol and inhibitor tolerance, making these yeasts of interest for further industrial exploration and exploitation.

Both neuronal acetylcholine and nonneuronal acetylcholine have been demonstrated to modulate inflammatory responses. Studies investigating the role of acetylcholine in the pathogenesis of bacterial infections have revealed contradictory findings with regard to disease outcome. At present, the role of acetylcholine in the pathogenesis of fungal infections is unknown. Therefore, the aim of this study was to determine whether acetylcholine plays a role in fungal biofilm formation and the pathogenesis of Candida albicans infection. The effect of acetylcholine on C. albicans biofilm formation and metabolism in vitro was assessed using a crystal violet assay and phenotypic microarray analysis. Its effect on the outcome of a C. albicans infection, fungal burden, and biofilm formation were investigated in vivo using a Galleria mellonella infection model. In addition, its effect on modulation of host immunity to C. albicans infection was also determined in vivo using hemocyte counts, cytospin analysis, larval histology, lysozyme assays, hemolytic assays, and real-time PCR. Acetylcholine was shown to have the ability to inhibit C. albicans biofilm formation in vitro and in vivo . In addition, acetylcholine protected G. mellonella larvae from C. albicans infection mortality. The in vivo protection occurred through acetylcholine enhancing the function of hemocytes while at the same time inhibiting C. albicans biofilm formation. Furthermore, acetylcholine also inhibited inflammation-induced damage to internal organs. This is the first demonstration of a role for acetylcholine in protection against fungal infections, in addition to being the first report that this molecule can inhibit C. albicans biofilm formation. Therefore, acetylcholine has the capacity to modulate complex host-fungal interactions and plays a role in dictating the pathogenesis of fungal infections.

Candida albicans metabolic activity in the presence and absence of acetylcholine was measured using phenotypic microarray analysis. Acetylcholine inhibited C. albicans biofilm formation by slowing metabolism independent of biofilm forming capabilities. Phenotypic microarray analysis can therefore be used for screening compound libraries for novel anti-fungal drugs and measuring antifungal resistance.

Biofuels are expected to play a role in replacing crude oil as a liquid transportation fuel, and research into butanol has highlighted the importance of this alcohol as a fuel. Butanol has a higher energy density than ethanol, butanol–gasoline blends do not separate in the presence of water, and butanol is miscible with gasoline (Szulczyk, Int J Energy Environ 1(1):2876–2895, 40). Saccharomyces cerevisiae has been used as a fermentative organism in the biofuel industry producing ethanol from glucose derived from starchy plant material; however, it typically cannot tolerate butanol concentrations greater than 2 % (Luong, Biotechnol Bioeng 29 (2):242–248, 27). 90 Saccharomyces spp. strains were screened for tolerance to 1-butanol via a phenotypic microarray assay and we observed significant variation in response with the most tolerant strains (S. cerevisiae DBVPG1788, S. cerevisiae DBVPG6044 and S. cerevisiae YPS128) exhibiting tolerance to 4 % 1-butanol compared with S. uvarum and S. castelli strains, which were sensitive to 3 % 1-butanol. Response to butanol was confirmed using traditional yeast methodologies such as growth; it was observed that fermentations in the presence of butanol, when using strains with a tolerant background, were significantly faster. Assessing for genetic rationale for tolerance, it was observed that 1-butanol-tolerant strains, when compared with 1-butanol-sensitive strains, had an up-regulation of RPN4, a transcription factor which regulates proteasome genes. Analysing for the importance of RPN4, we observed that a Δrpn4 strain displayed a reduced rate of fermentation in the presence of 1-butanol when compared with the BY4741 background strain. This data will aid the development of breeding programmes to produce better strains for future bio-butanol production.

Inhibitors released by the breakdown of plant cell walls prevent efficient conversion of sugar into ethanol. The aim of this study was to develop a fast and reliable inhibitor sensitivity assay for ethanologenic yeast strains. The assay comprised bespoke 96-well plates containing inhibitors in isolation or combination in a format that was compatible with the Phenotypic Microarray Omnilog reader (Biolog, hayward, CA, USA). A redox reporter within the assay permits analysis of inhibitor sensitivity in aerobic and/or anaerobic conditions. Results from the assay were verified using growth on spot plates and tolerance assays in which maintenance of viability was assessed. The assay allows for individual and synergistic effects of inhibitors to be determined. It was observed that the presence of both acetic and formic acid significantly inhibited the yeast strains assessed, although this impact could be partially mitigated by buffering to neutral pH. Scheffersomyces stipitis, Candida spp., and Pichia guilliermondii demonstrated increased sensitivity to short chain weak acids at concentrations typically present in lignocellulosic hydrolysates. S. cerevisiae exhibited robustness to short chain weak acids at these concentrations. However, S. stipitis, Candida spp., and P. guilliermondii displayed increased tolerance to HMF when compared to that observed for S. cerevisiae. The results demonstrate that the phenotypic microarray assay developed in the current study is a valuable tool that can be used to identify yeast strains with desirable resistance to inhibitory compounds found in lignocellulosic hydrolysates.

The potential of using sorghum milling waste for the development of a biorefining strategy for the production of bioethanol was investigated. Both red and white sorghum were processed using a traditional Nigerian wet-milling process to sorghum flour. The sorghum milling waste, sorghum bran, was hydrolysed using both enzymatic and dilute acid hydrolysis to produce a generable fermentation feedstock. The hydrolysates were subsequently investigated for fermentative biofuel production. Following a hydrolysis step, a medium containing ˜61 g/L glucose was obtained. Trace presence of inhibitors was detected in the hydrolysates and sufficient nitrogen content to support microorganism growth and bioethanol production. In test bioethanol production experiments using the sorghum milling waste derived hydrolysates only, 24.35 g/L bioethanol was produced by a yeast Kluyveromyces marxianus, equivalent to a yield of 0.15 g bioethanol per g of sorghum milling waste.

Yeast, like other eukaryotes, contains a complete mitochondrial thioredoxin system comprising a thioredoxin (Trx3) and a thioredoxin reductase (Trr2). Mitochondria are a main source of reactive oxygen species (ROS) in eukaryotic organisms, and this study investigates the role of Trx3 in regulating cell death during oxidative stress conditions.We have previously shown that the redox state of mitochondrial Trx3 is buffered by the glutathione redox couple such that oxidized mitochondrial Trx3 only accumulates in mutants simultaneously lacking Trr2 and a glutathione reductase (Glr1). We show here that the redox state of mitochondrial Trx3 is important for yeast growth and its oxidation in a glr1 trr2 mutant induces programmed cell death. Apoptosis is dependent on the Yca1 metacaspase, since loss of YCA1 abrogates cell death induced by oxidized Trx3. Our data also indicate a role for a mitochondrial 1-cysteine (Cys) peroxiredoxin (Prx1) in the oxidation of Trx3, since Trx3 does not become oxidized in glr1 trr2 mutants or in a wild-type strain exposed to hydrogen peroxide in the absence of PRX1.This study provides evidence that the redox state of a mitochondrial thioredoxin regulates yeast apoptosis in response to oxidative stress conditions. Moreover, the results identify a signaling pathway, where the thioredoxin system functions in both antioxidant defense and in controlling cell death.Mitochondrial Prx1 functions as a redox signaling molecule that oxidizes Trx3 and promotes apoptosis. This would mean that under conditions where Prx1 cannot detoxify mitochondrial ROS, it induces cell death to remove the affected cells.

The nature of and factors effecting sodium interactions with psyllium were investigated in vitro. In a batch extraction system, psyllium mucilage gel retained at least 50% of sodium across a range of concentrations (5–300 mg sodium per g psyllium) and pH (2–10) environments. FTIR and Na NMR analyses of psyllium gels indicated that binding was complex with non-specific multi-site interactions. The potential use of psyllium husk as a binding agent for the reduction of bioavailable sodium was therefore evaluated. The binding of sodium at physiologically relevant conditions (pH 1.2 (stomach) and 6.8 (intestine)) was studied in a gastrointestinal tract (GIT) pH simulated model. Results show consistently high sodium retention (∼50%) across the GIT model and less than 20% loss of bound sodium under the simulated intestinal pH conditions after repeated washings.

Yeasts that are present in marine environments have evolved to survive hostile environments that are characterized by high exogenous salt content, high concentrations of inhibitory compounds, and low soluble carbon and nitrogen levels. Therefore, yeasts isolated from marine environments could have interesting characteristics for industrial applications. However, the application of marine yeast in research or industry is currently very limited owing to the lack of a suitable isolation method. Current methods for isolation suffer from fungal interference and/or low number of yeast isolates. In this paper, an efficient and nonlaborious isolation method has been developed and successfully isolated large numbers of yeasts without bacterial or fungal growth. The new method includes a three-cycle enrichment step followed by an isolation step and a confirmation step. Using this method, 116 marine yeast strains were isolated from 14 marine samples collected in the UK, Egypt, and the USA. These strains were further evaluated for the utilization of fermentable sugars (glucose, xylose, mannitol, and galactose) using a phenotypic microarray assay. Seventeen strains with higher sugar utilization capacity than the reference terrestrial yeast Saccharomyces cerevisiae NCYC 2592 were selected for identification by sequencing of the ITS and D1/D2 domains. These strains belonged to six species: S. cerevisiae, Candida tropicalis, Candida viswanathii, Wickerhamomyces anomalus, Candida glabrata, and Pichia kudriavzevii. The ability of these strains for improved sugar utilization using seawater-based media was confirmed and, therefore, they could potentially be utilized in fermentations using marine biomass in seawater media, particularly for the production of bioethanol and other biochemical products.

This paper explores the possibility of using an industrially processed municipal solid waste (MSW) for cellulase enzyme production via solid state fermentation (SSF) by Trichoderma reesei and Aspergillus niger. Both fungi grew well on the MSW substrate and production of cellulase enzymes was optimized for temperature, moisture content, inoculation and period of incubation. The effect of additional minerals, and alternative carbon and nitrogen sources were also examined. Following optimization a cellulase activity of 26.10 ± 3.09 FPU/g could be produced using T. reesei at 30°C with a moisture content of 60% with an inoculums of 0.5 million spores/g and incubation for 168 hours. Addition of extra nitrogen and/or carbon did not improve cellulase accumulation. Acid or alkali pretreatment of MSW led to reduced cellulase production. Crude enzymes produced from MSW by T. reesei were evaluated for their ability to release glucose from MSW. A cellulose hydrolysis yield of 24.7% was achieved, which was close to that obtained using a commercial enzyme. Results demonstrated that MSW can be used as an inexpensive lignocellulosic material for the production of cellulase enzymes.

Cellular mechanisms that maintain redox homeostasis are crucial, providing buffering against oxidative stress. Glutathione, the most abundant low molecular weight thiol, is considered the major cellular redox buffer in most cells. To better understand how cells maintain glutathione redox homeostasis, cells of Saccharomyces cerevisiae were treated with extracellular oxidized glutathione (GSSG), and the effect on intracellular reduced glutathione (GSH) and GSSG were monitored over time. Intriguingly cells lacking GLR1 encoding the GSSG reductase in S. cerevisiae accumulated increased levels of GSH via a mechanism independent of the GSH biosynthetic pathway. Furthermore, residual NADPH-dependent GSSG reductase activity was found in lysate derived from glr1 cell. The cytosolic thioredoxin-thioredoxin reductase system and not the glutaredoxins (Grx1p, Grx2p, Grx6p, and Grx7p) contributes to the reduction of GSSG. Overexpression of the thioredoxins TRX1 or TRX2 in glr1 cells reduced GSSG accumulation, increased GSH levels, and reduced cellular glutathione E(h)'. Conversely, deletion of TRX1 or TRX2 in the glr1 strain led to increased accumulation of GSSG, reduced GSH levels, and increased cellular E(h)'. Furthermore, it was found that purified thioredoxins can reduce GSSG to GSH in the presence of thioredoxin reductase and NADPH in a reconstituted in vitro system. Collectively, these data indicate that the thioredoxin-thioredoxin reductase system can function as an alternative system to reduce GSSG in S. cerevisiae in vivo.

Several studies have implicated the NF-kappaB inhibitor-like protein 1 (NFkBIL1) gene located in the class III region of the major histocompatibility complex (MHC) as a possible susceptibility locus for rheumatoid arthritis (RA). Based on limited homology, it has been suggested to be a member of the inhibitor of NF-kappaB (IkappaB) family of proteins, but a role in mRNA processing has also been proposed. We have investigated the expression of NFkBIL1 in RA synovial tissue and characterized its function. Real-time PCR showed the two NFkBIL1 mRNA splice variants are expressed in a tissue-specific manner. Dual immunofluorescent staining of human RA synovium with polyclonal anti-NFkBIL1 antibodies and anti-CD68, anti-CD3 or anti-factor VIII showed that NFkBIL1 was expressed in the rheumatoid synovial lining and sub-lining layers and co-localized in CD68+ and CD3+, but not Factor VIII+ cells. Confocal microscopy of cultured synovial fibroblasts revealed expression in speckled nuclear and homogenous cytoplasmic distributions, suggesting shuttling between the cytoplasmic and nuclear compartments. Functional tests showed that NFkBIL1 isoforms were incapable of associating with NF-kappaB and did not inhibit it, thus disproving the hypothesis that NFkBIL1 functions as an IkappaB. Affinity purification of endogenous NFkBIL1 proteins and co-immunoprecipitation experiments showed that NFkBIL1 can associate with mRNA and with three protein partners, identified by mass spectrometry as leukophysin, translation elongation factor 1 alpha and CTP synthase I. These data support a potential role for NFkBL1 in the pathogenesis of RA and indicates that it may be involved in mRNA processing or the regulation of translation.

Highlights • Storage stability of unwashed potato crisps fried in sunflower oil’s was compared. • Oxidation products were determined, with and without flushing with nitrogen gas. • Crisps fried in SO were the least stable and those using HOSO the most stable. • Presence of nitrogen gas slowed down the oxidation rate after frying., Unwashed, sliced, batch-fried potato crisps have a unique texture and are growing in popularity in the UK/EU premium snack food market. In this study, the storage stability of unwashed sliced (high surface starch) potatoes (crisps) fried in regular sunflower oil (SO) or in high oleic sunflower oil (HOSO) was compared over accelerated shelf life testing (45 °C, 6 weeks); with and without nitrogen gas flushing. Primary oxidation products (lipid hydroperoxides) were measured with a ferrous oxidation-xylenol orange (FOX) assay and volatile secondary oxidation products (hexanal) were quantified by using solid phase micro-extraction gas chromatography mass spectrometry (HS-SPME-GC/MS). Results revealed that crisps fried in SO were the least stable. Flushing the stored crisps with nitrogen gas proved to be effective in slowing down the oxidation rate after frying with sunflower oil, significantly stabilizing the crisps. However, crisps fried in HOSO were the most stable, with the lowest rate of development of oxidation markers, and this has previously not been shown for crisps with a high free starch content.

Aims Climate change is exerting an increasingly profound effect on grape composition, microbiology, chemistry and the sensory aspects of wine. Identification of autochthonous yeasts tolerant to stress could help to alleviate this effect. Methods and Results Tolerance to osmotic pressure, ethanol and pH of 94 Saccharomyces cerevisiae strains and 29 strains non-Saccharomyces from the warm climate region DO ‘Vinos de Madrid’ (Spain) using phenotypic microarray and their fermentative behaviour were studied. The screening highlighted 12 strains of S. cerevisiae isolated from organic cellars with improved tolerance to osmotic stress, high ethanol concentrations and suitable fermentative properties. Screening of non-Saccharomyces spp. such as Lanchacea thermotolerans, Torulaspora delbrueckii, Schizosaccharomyces pombe and Mestchnikowia pulcherrima also highlighted tolerance to these stress conditions. Conclusions This study confirmed the adaptation of native strains to the climatic conditions in each area of production and correlated these adaptations with good fermentation properties. Screening has revealed that identifying yeast strains adapted to fermentation stresses is an important approach for making quality wines in very warm areas. Significance and Impact of the Study The results have special relevance because it is a pioneering study that has approached the problem of climate change for wines from a microbiological aspect and has analysed the situation in situ in wineries from a warm climate zone. Resistant strains were found with good biological properties; studying these strains for their stress response mechanisms during fermentation will be of interest to the wine making industry.

Phage display techniques using random peptide interactions have supported the role of mammalian glutathione transferase (GST) as part of a signalling pathway for both oxidative stress and an apoptosis pathway. Little is known about the interaction of nonmammalian GST with other proteins. GSTs have been implicated in the development of chronic nematode infections by neutralising cytotoxic products arising from host immune initiated reactive oxygen species (ROS) assault. In this study we attached one of the key GSTs expressed in the model nematode Caenorhabditis elegans to an affinity support matrix and directly identified major interacting proteins by two-dimensional electrophoresis and peptide mass fingerprinting before and following oxidative stress. Nematode GST does not appear to be a stand-alone enzyme and interacts with many types of proteins in both normal and ROS stress conditions. Pull-down proteomic presents a flexible, label free, rapid and economical assay without specialised ligand fishing equipment to identify protein binding partners.

Phenotype microarray (PM) technology provides an insight into the metabolic profiling of microbial cells within 96-well plate system. The PM assay allows for cells to be assessed for utilisation of nutrients or sensitivity to toxic compounds. The assay utilises a redox sensitive tetrazolium dye which becomes irreversibly reduced upon detection of cellular metabolic output, detection is synchronous with a colour change from colourless to purple. Output from PM technology can be measured visually or quantified by reader the absorbance in each well. PM technology has highlighted differences in growth requirements, nutrient utilisation, sensitivity to toxins, and genetic diversity in bacteria, fungi and mammalian cells.

Background Bioethanol fermentations follow traditional beverage fermentations where the yeast is exposed to adverse conditions such as oxidative stress. Lignocellulosic bioethanol fermentations involve the conversion of pentose and hexose sugars into ethanol. Environmental stress conditions such as osmotic stress and ethanol stress may affect the fermentation performance; however, oxidative stress as a consequence of metabolic output can also occur. However, the effect of oxidative stress on yeast with pentose utilising capabilities has yet to be investigated. Results Assaying for the effect of hydrogen peroxide-induced oxidative stress on Candida, Pichia and Scheffersomyces spp. has demonstrated that these yeast tolerate hydrogen peroxide-induced oxidative stress in a manner consistent with that demonstrated by Saccharomyces cerevisiae. Pichia guillermondii appears to be more tolerant to hydrogen peroxide-induced oxidative stress when compared to Candida shehatae, Candida succiphila or Scheffersomyces stipitis. Conclusions Sensitivity to hydrogen peroxide-induced oxidative stress increased in the presence of minimal media; however, addition of amino acids and nucleobases was observed to increase tolerance. In particular adenine increased tolerance and methionine reduced tolerance to hydrogen peroxide-induced oxidative stress.

Background Protein-SH groups are amongst the most easily oxidized residues in proteins, but irreversible oxidation can be prevented by protein glutathionylation, in which protein-SH groups form mixed disulphides with glutathione. Glutaredoxins and thioredoxins are key oxidoreductases which have been implicated in regulating glutathionylation/deglutathionylation in diverse organisms. Glutaredoxins have been proposed to be the predominant deglutathionylase enzymes in many plant and mammalian species, whereas, thioredoxins have generally been thought to be relatively inefficient in deglutathionylation. Results We show here that the levels of glutathionylated proteins in yeast are regulated in parallel with the growth cycle, and are maximal during stationary phase growth. This increase in glutathionylation is not a response to increased reactive oxygen species generated from the shift to respiratory metabolism, but appears to be a general response to starvation conditions. Our data indicate that glutathionylation levels are constitutively high in all growth phases in thioredoxin mutants and are unaffected in glutaredoxin mutants. We have confirmed that thioredoxins, but not glutaredoxins, catalyse deglutathionylation of model glutathionylated substrates using purified thioredoxin and glutaredoxin proteins. Furthermore, we show that the deglutathionylase activity of thioredoxins is required to reduce the high levels of glutathionylation in stationary phase cells, which occurs as cells exit stationary phase and resume vegetative growth. Conclusions There is increasing evidence that the thioredoxin and glutathione redox systems have overlapping functions and these present data indicate that the thioredoxin system plays a key role in regulating the modification of proteins by the glutathione system.

The soluble global proteome of adult nematode Heligmosomoides polygyrus (H. p.) bakeri, a hookworm laboratory model was compared for the first time in the intestines of a slow-responder mouse host strain (C57/BL10) that is known to support a primary parasite infection for many months, and rapid-responder mouse host (SWR) that is known to eliminate the nematode infection by week 6 postinfection. At week 4 postinfection, major adult nematode proteins selectively produced following establishment of infection in C57/BL10 hosts include several globin forms, calreticulin and a phosphatidylethanolamine-binding protein. The increased synthesis of forms of myosin, actin and troponin in the nematode living in the rapid-responder SWR host may relate to the attempted reorganisation or repair of the cytoskeleton and/or muscle layer in the host immune initiated, increased mucus production and smooth muscle activity within intestinal environment. Initial evidence suggests weakly antigenic forms of globins dominant in the cytosol of H. p. bakeri adults in the intestinal environment compared to their low production in a related free-living nematode. The demonstration of considerable plasticity within a parasitic nematode proteome provides a molecular basis for the previously observed phenotypic plasticity within different host environments. Proteome plasticity has relevance to the efficiency of future vaccine and drug therapy, and the continued failure of defined antigen vaccines in mammalian populations.

Background The use of a microwave synthesis reactor has allowed kinetic data for the hydrothermal reactions of straw biomass to be established from short times, avoiding corrections required for slow heating in conventional reactors, or two-step heating. Access to realistic kinetic data is important for predictions of optimal reaction conditions for the pretreatment of biomass for bioethanol processes, which is required to minimise production of inhibitory compounds and to maximise sugar and ethanol yields. Results The gravimetric loss through solubilisation of straw provided a global measure of the extent of hydrothermal deconstruction. The kinetic profiles of furan and lignin-derived inhibitors were determined in the hydrothermal hydrolysates by UV analysis, with concentrations of formic and acetic acid determined by HPLC. Kinetic analyses were either carried out by direct fitting to simple first order equations or by numerical integration of sequential reactions. Conclusions A classical Arrhenius activation energy of 148 kJmol−1 has been determined for primary solubilisation, which is higher than the activation energy associated with historical measures of reaction severity. The gravimetric loss is primarily due to depolymerisation of the hemicellulose component of straw, but a minor proportion of lignin is solubilised at the same rate and hence may be associated with the more hydrophilic lignin-hemicellulose interface. Acetic acid is liberated primarily from hydrolysis of pendant acetate groups on hemicellulose, although this occurs at a rate that is too slow to provide catalytic enhancement to the primary solubilisation reactions. However, the increase in protons may enhance secondary reactions leading to the production of furans and formic acid. The work has suggested that formic acid may be formed under these hydrothermal conditions via direct reaction of sugar end groups rather than furan breakdown. However, furan degradation is found to be significant, which may limit ultimate quantities generated in hydrolysate liquors.

Saccharomyces cerevisiae is the micro-organism of choice for the conversion of fermentable sugars during beverage or bioethanol fermentations. These fermentations are characterised by high osmotic stress on a yeast cell, with selected brewing fermentations beginning at 20–25% fermentable sugars and bioethanol fermentations at 13% fermentable sugars. RCK2 encodes for a MAPKAP (MAPK-activated protein kinase) enzyme and was identified on a locus by QTL analysis in yeast cells under osmotic stress, RCK2 expression was placed under a tetracycline regulatable vector and rescued glucose, sorbitol or glycerol induced osmotic stress in an rck2 null strain. A strain overexpressing RCK2 had significantly faster fermentation rates when compared with the empty vector control strain. Presence of RCK2 increased rates of glucose utilisation (~40 g glucose in first 8 h) during a 15% glucose fermentation and concurrent production of ethanol when compared with empty vector controls. Tolerance to osmotic stress using the tetracycline regulatable vectors could be turned off with the addition of tetracycline returning a rck2 null strain back to osmotic sensitivity.

BACKGROUND: During industrial fermentation of lignocellulose residues to produce bioethanol, microorganisms are exposed to a number of factors that influence productivity. These include inhibitory compounds produced by the pre-treatment processes required to release constituent carbohydrates from biomass feed-stocks and during fermentation, exposure of the organisms to stressful conditions. In addition, for lignocellulosic bioethanol production, conversion of both pentose and hexose sugars is a pre-requisite for fermentative organisms for efficient and complete conversion. All these factors are important to maximise industrial efficiency, productivity and profit margins in order to make second-generation bioethanol an economically viable alternative to fossil fuels for future transport needs. RESULTS: The aim of the current study was to assess Saccharomyces yeasts for their capacity to tolerate osmotic, temperature and ethanol stresses and inhibitors that might typically be released during steam explosion of wheat straw. Phenotypic microarray analysis was used to measure tolerance as a function of growth and metabolic activity. Saccharomyces strains analysed in this study displayed natural variation to each stress condition common in bioethanol fermentations. In addition, many strains displayed tolerance to more than one stress, such as inhibitor tolerance combined with fermentation stresses. CONCLUSIONS: Our results suggest that this study could identify a potential candidate strain or strains for efficient second generation bioethanol production. Knowledge of the Saccharomyces spp. strains grown in these conditions will aid the development of breeding programmes in order to generate more efficient strains for industrial fermentations.