Respiration-deficient Mutants Research Articles

Importance of Intracellular Energy Status on High-Hydrostatic-Pressure Inactivation of sake Yeast Saccharomyces cerevisiae.

The HHP inactivation behaviors of Niigata sake yeast Saccharomyces cerevisiae strain S9arg and its aerobic respiratory-deficient mutant strains were investigated after cultivating them in a YPD media containing 2% to 15% glucose, as well as in moromi mash, in a laboratory-scale sake brewing process. The piezotolerance of strain S9arg, shown after cultivation in a YPD medium containing 2% glucose, decreased to become piezosensitive with increasing glucose concentrations in YPD media. In contrast, the piezosensitivity of a mutant strain UV1, shown after cultivation in the YPD medium containing 2% glucose, decreased to become piezotolerant with increasing glucose concentrations in the YPD medium. The intracellular ATP concentrations were analyzed for an S. cerevisiae strain with intact aerobic respiratory ability, as well as for strain UV1. The higher concentration of ATP after cultivation suggested a higher energy status and may be closely related to higher piezotolerance for the yeast strains. The decreased piezotolerance of strain S9arg observed after a laboratory-scale sake brewing test may be due to a lower energy status resulting from a high glucose concentration in moromi mash during the early period of brewing, as well as a lower aeration efficiency during the brewing process, compared with cultivation in a YPD medium containing 2% glucose.

Severe ethanol stress induces the preferential synthesis of mitochondrial disaggregase Hsp78 and formation of DUMPs in Saccharomyces cerevisiae

Severe ethanol stress (>9% v/v) induces pronounced translation repression in yeast cells. However, some proteins, which are exceptionally synthesized even under translation repression, play important roles in ethanol tolerance. These proteins are expected to provide important clues for elucidating the survival strategies of yeast cells under severe ethanol stress. In this study, we identified Hsp78 as a protein effectively synthesized under severe ethanol stress. As Hsp78 is involved in mitochondrial protein quality control, we investigated the effect of severe ethanol stress on mitochondrial proteins and found that Ilv2, Kgd1, and Aco1 aggregated with Hsp78 under severe ethanol stress, forming mitochondrial deposition sites for denatured proteins, called DUMPs (Deposits of Unfolded Mitochondrial Proteins). Aggregation of mitochondrial proteins and formation of DUMPs were accelerated in hsp78∆ cells compared with those in wild-type cells. During the recovery process after ethanol removal, aggregated Ilv2 and DUMP levels rapidly decreased in wild-type cells but were maintained for a long time (>180 min) in hsp78Δ cells. Furthermore, the frequency of respiration-deficient mutants caused by severe ethanol stress was higher in hsp78∆ cells than in wild-type cells. These results indicate that severe ethanol stress damaged mitochondrial proteins and that Hsp78 was preferentially synthesized to cope with the damage, thereby suppressing the rapid increase in aggregated protein levels under stress and achieving proper clearance of aggregated proteins during the recovery process. This study provides novel insights into the adverse effects of ethanol on mitochondria and yeast response to severe ethanol stress.

Respiratory Deficient Mutants Induced in Baker's Yeast by the Action of Acridine Orange and some Plant Growth Hormones طفرات نقص التنفس المستحدثة فی خمیرة الخباز کنتیجة للمعاملة بالأکریدین وهرمونات النمو النباتیة

The present study aimed to investigate the mutagenic effect of acriflavine and some plant growth hormones on mitochondrial DNA of Saccharomyces cerevisiae. Two yeast isolates isolated from different sources were used in this work. The results of acriflavine treatments showed significant toxicity on the viability of yeast cells via a dose – response appeared. This means that the survival of cells was decreased with the increasing in acriflavine concentrations. This is a clear evidence for the toxic action of acriflavine on yeast cells. At the concentration of 1 ppm the parent cells of isolate – A retain the normal respiratory abilities, while the isolate – B was less affected. The high rate of petite mutants appeared when the cells were exposed to 4 ppm. Isolate – B was more sensitive to acriflavine than isolate – A regarding the decline in the viability of cells and respiratory deficient mutants induced. In addition, kinetin and naphthyl acetic acid showed a dose – response for the viability of yeast cells without respiratory deficient mutants induced. This indicated that both plant growth hormones enhances the division of yeast cells leading the cells to continue growing. In addition, plant growth hormones plays several vital roles in biochemical processes related to cell cycle of yeast cells. The signaling molecules regulated cell growth must received significant attention because it is alarm sounds for the potential of carcinogenicity.

Quantitative multi-omics analysis of the effects of mitochondrial dysfunction on lipid metabolism in Saccharomyces cerevisiae.

In this study, combined genome, transcriptome, and metabolome analysis was performed for eight Saccharomyces cerevisiae mitochondrial respiration-deficient mutants. Each mutant exhibited a unique nuclear genome mutation pattern; the nuclear genome mutations, and thus potentially affected genes and metabolic pathways, showed a co-occurrence frequency of ≤ 3 among the eight mutants. For example, only a lipid metabolism-related pathway was likely to be affected by the nuclear genome mutationsin one of the mutants. However, large deletions in the mitochondrial genome were the shared characteristic among the eight mutants. At the transcriptomic level, lipid metabolism was the most significantly enriched Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway for differentially expressed genes (DEGs) co-occurring in both ≥ 4 and ≥ 5 mutants. Any identified DEG enriched in lipid metabolism showed the same up-/down-regulated pattern among nearly all eight mutants. Further, 126 differentially expressed lipid species (DELS) were identified, which also showed the same up-/down-regulated pattern among nearly all investigated mutants. It was conservatively demonstrated that the similar change pattern of lipid metabolism in the entire investigated mutant population was attributed to mitochondrial dysfunction. The change spectrum of lipid species was presented, suggesting that the number and change degree of up-regulated lipid species were higher than those of down-regulated lipid species. Additionally, energy storage lipids increased in content and plasma-membrane phospholipid compositions varied in the relative proposition. The results for the genome, transcriptome, and lipidome were mutually validated, which provides quantitative data revealing the roles of mitochondria from a global cellular perspective.

Co-culture strategy for improved 2G bioethanol production using a mixture of sugarcane molasses and bagasse hydrolysate as substrate

Mixed sugar co-fermentation represents an interesting fermentation strategy to improve bioethanol production from low-cost raw materials, such as sugarcane bagasse. The aim of this study was to evaluate a mixture of sugars (hexoses and pentoses) from sugarcane bagasse hemicellulosic hydrolysate diluted with sugarcane molasses as substrate to be converted either in monoculture or co-culture fermentations. A co-culture fermentation technique using hexoses-fermenting yeasts (Saccharomyces cerevisiae: from Santa Adélia Mill, an industrial strain (SA) and a respiratory-deficient mutant strain (ScP)) together with xylose-fermenting yeasts (Scheffersomyces stipitis (SS) and Spathaspora passalidarum (SP)) was carried out to enhance bioethanol production. Co-culture fermentation in a medium with a high mixed sugar concentration (100 g.L−1) resulted in a complete sugar consumption, ethanol titer of 30.2 g.L−1 and increased ethanol productivity (Qp =4.44 g.L−1. h−1) using wild type strains (SP + SA). In contrast, the maximum ethanol titer (49.2 g.L−1) was reached during co-culture performed with S. stipitis and the respiratory-deficient mutant strain of S. cerevisiae, however ethanol productivity (Qp =0.60 g.L−1. h−1) decreased due to the poor fermentation performance of the mutant strain. This work suggests that improved efficiency and good scalability of the mixed sugar fermentation process makes an important basis for the economic viability of bioethanol production from lignocelluloses.

A genome-wide view of mutations in respiration-deficient mutants of Saccharomyces cerevisiae selected following carbon ion beam irradiation.

Mitochondrial dysfunction in Saccharomyces cerevisiae was selected as a marker of ion penetration following carbon ion beam (CIB) irradiation. Respiration-deficient mutants were screened. Following confirmation of negligible spontaneous mutation, eight genetically stable S. cerevisiae respiration-deficient mutant strains and a control strain were resequenced with ~ 200-fold read depth. Strategies were established to identify and validate the particular mutations induced by CIB irradiation. In the nuclear genome, CIB irradiation mainly caused base substitutions and some small (< 100bp) insertions/deletions (indels), which were widely distributed across the chromosomes. Although mitochondrial dysfunction was selected as a screening marker, variants in the nuclear genome were detected at a high frequency (10-7) relative to spontaneous mutations (10-9). The transition to transversion ratio for base substitutions was 0.746, which was less than that of spontaneous mutations. In the mitochondrial genome, there were very large deletions including substantial gene areas, resulting in extremely low read coverage. Meanwhile, every mutant possessed a distinctive mutation pattern, for both the nuclear and the mitochondrial genome. Nuclear genomes contained scanty mitochondrial respiration-related genes that were potentially affected by verified mutations, suggesting that variants in the mitochondrial genome may be the main drivers of respiratory deficiencies. Our study confirmed the previous finding that heavy ion beam (HIB) irradiation mainly induces substantial base substitutions and some small indels but also yielded some novel findings, in particular, novel structural variants in the mitochondrial genomes. These data will enhance the understanding of HIB-induced damage and mutations and aid in the HIB-based microbial mutation breeding.

Nickel Oxide Nanoparticles Trigger Caspase- and Mitochondria-Dependent Apoptosis in the Yeast Saccharomyces cerevisiae.

The expansion of the industrial use of nickel oxide (NiO) nanoparticles (NPs) raises concerns about their potential adverse effects. Our work aimed to investigate the mechanisms of toxicity induced by NiO NPs, using the yeast Saccharomyces cerevisiae as a cell model. Yeast cells exposed to NiO NPs exhibited typical hallmarks of regulated cell death (RCD) by apoptosis [loss of cell proliferation capacity (cell viability), exposure of phosphatidylserine at the outer cytoplasmic membrane leaflet, nuclear chromatin condensation, and DNA damage] in a process that required de novo protein synthesis. The execution of yeast cell death induced by NiO NPs is Yca1p metacaspase-dependent. NiO NPs also induced a decrease in the mitochondrial membrane potential and an increase in the frequency of respiratory-deficient mutants, which supports the involvement of mitochondria in the cell death process. Cells deficient in the apoptosis-inducing factor ( aif1Δ) displayed higher tolerance to NiO NPs, which reinforces the involvement of mitochondria in RCD by apoptosis. In summary, this study shows that NiO NPs induce caspase- and mitochondria-dependent apoptosis in yeast. Our results warn about the possible harmful effects associated with the use of NiO NPs.

Evaluation of fermentation potential of wild and UV-mutated yeasts screened from traditional murcha

Murcha (an amylolytic starter) from different parts of Eastern Nepal were screened for fermentative yeasts. The most potential one was UV-mutated (8W lamp at λ = 254 nm and an intensity of 44.21 Wm-2 for 5-50 s) to study the effect of mutation on growth and fermentation properties. Respiratory-deficient mutants (RDMs) that resulted from the mutation were identified by triphenyl tetrazolium chloride (TTC) overlay technique and replica-plated for further isolation. Cell growth, substrate utilization, and ethanol yield of the mutants were compared with normal cells by carrying out fermentation in high-test cane molasses broth of 30 °Bx. An exhaustive screening of the samples resulted in only two murcha viz., from Laxmimarga (LM) and Udayapur (UD), having the desirable fermentation properties. UV-mutation study of UD and LM yeasts (both identified as strains of Saccharomyces cerevisiae) showed 8-12% survival and ~ 22% RDMs yield of the survived cells. Out of the 8 randomly selected RDMs, only UDm4 (colony No. 4 from UD) showed fermentation properties worth further investigation. Comparison of UD, LM and UDm4 by fermenting molasses (high test) broth of 30°Bx showed the least growth of UDm4 but the highest alcohol yield (9% and 16% more compared to UD and LM, respectively). The present finding indicates that it is possible to improve fermentation properties of feral yeasts from murcha by relatively simple UV-mutation approach. Finding the right mutant (the selective screening part), however, may involve considerable time and effort.

Mitochondrial dysfunction reduces yeast replicative lifespan by elevating RAS-dependent ROS production by the ER-localized NADPH oxidase Yno1.

Mitochondrial dysfunction leads to the accumulation of reactive oxygen species (ROS) which is associated with cellular dysfunction, disease etiology, and senescence. Here, we used the eukaryotic model Saccharomyces cerevisiae, commonly studied for cellular aging, to demonstrate how defective mitochondrial function affects yeast replicative lifespan (RLS). We show that RLS of respiratory-deficient cells decreases significantly, indicating that the maintenance of RLS requires active respiration. The shortening of RLS due to mitochondrial dysfunction was not related to the accumulation of extrachromosomal ribosomal DNA circles, a well-known cause of aging in yeast. Instead, intracellular ROS and oxidatively damaged proteins increased in respiratory-deficient mutants. We show that, while the protein kinase A activity is not elevated, ROS generation in respiratory-deficient cells depends on RAS signaling pathway. The ER-localized NADPH oxidase Yno1 also played a role in producing ROS. Our data suggest that a severe defect in mitochondrial respiration accelerates cellular aging by disturbing protein homeostasis in yeast.

Nucleo-mitochondrial interaction of yeast in response to cadmium sulfide quantum dot exposure

Cell sensitivity to quantum dots (QDs) has been attributed to a cascade triggered by oxidative stress leading to apoptosis. The role and function of mitochondria in animal cells are well understood but little information is available on the complex genetic networks that regulate nucleo-mitochondrial interaction. The effect of CdS QD exposure in yeast Saccharomyces cerevisiae was assessed under conditions of limited lethality (<10%), using cell physiological and morphological endpoints. Whole-genomic array analysis and the screening of a deletion mutant library were also carried out. The results showed that QDs: increased the level of reactive oxygen species (ROS) and decreased the level of reduced vs oxidized glutathione (GSH/GSSG); reduced oxygen consumption and the abundance of respiratory cytochromes; disrupted mitochondrial membrane potentials and affected mitochondrial morphology. Exposure affected the capacity of cells to grow on galactose, which requires nucleo-mitochondrial involvement. However, QDs exposure did not materially induce respiratory deficient (RD) mutants but only RD phenocopies. All of these cellular changes were correlated with several key nuclear genes, including TOM5 and FKS1, involved in the maintenance of mitochondrial organization and function. The consequences of these cellular effects are discussed in terms of dysregulation of cell function in response to these “pathological mitochondria”.

The respiratory chain provides salt stress tolerance by maintaining a low NADH/NAD+ ratio in Zymomonas mobilis.

The respiratory chain of ethanol-producing Zymomonas mobilis shows an unusual physiological property in that it is not involved in energy conservation, even though this organism has a complete electron transport system. We reported previously that respiratory-deficient mutants (RDMs) of Z. mobilis exhibit higher growth rates and enhanced ethanol productivity under aerobic and high-temperature conditions. Here, we demonstrated that the salt tolerance of RDM strains was drastically decreased compared with the wild-type strain. We found that the NADH/NAD+ ratio was maintained at low levels in both the wild-type and the RDM strains under non-stress conditions. However, the ratio substantially increased in the RDM strains in response to salt stress. Complementation of the deficient respiratory-chain genes in the RDM strains resulted in a decrease in the NADH/NAD+ ratio and an increase in the growth rate. In contrast, expression of malate dehydrogenase, activity of which increases the supply of NADH, in the RDM strains led to an increased NADH/NAD+ ratio and resulted in poor growth. Taken together, these results suggest that the respiratory chain of Z. mobilis functions to maintain a low NADH/NAD+ ratio when the cells are exposed to environmental stresses, such as salinity.

Deletion of the MED13 and CDK8 subunits of the Mediator improves the phenotype of a long-lived respiratory deficient mutant of Podospora anserina

In Podospora anserina, the loss of function of the cytochrome segment of the mitochondrial respiratory chain is viable. This is due to the presence in this organism, as in most filamentous fungi, of an alternative respiratory oxidase (AOX) that provides a bypass to the cytochrome pathway. However mutants lacking a functional cytochrome pathway present multiple phenotypes including poorly colored thin mycelium and slow growth. In a large genetic screen based on the improvement of these phenotypes, we isolated a large number of independent suppressor mutations. Most of them led to the constitutive overexpression of the aox gene. In this study, we characterize a new suppressor mutation that does not affect the production of AOX. It is a loss-of-function mutation in the gene encoding the MED13 subunit of the kinase module of the Mediator complex. Inactivation of the cdk8 gene encoding another subunit of the same module also results in partial suppression of a cytochrome-deficient mutant. Analysis of strains lacking the MED13 or CDK8 subunits points to the importance of these subunits as regulators involved in diverse physiological processes such as growth, longevity and sexual development. Interestingly, transcriptional analyses indicate that in P. anserina, loss of the respiratory cytochrome pathway results in the up-regulation of glycolysis-related genes revealing a new type of retrograde regulation. The loss of MED13 augments the up-regulation of some of these genes.

Influence of Altered NADH Metabolic Pathway on the Respiratory-deficient Mutant of Rhizopus oryzae and its L-lactate Production.

Respiratory-deficient mutants of Rhizopus oryzae (R. oryzae) AS 3.3461 were acquired by ultraviolet (UV) irradiation to investigate changes in intracellular NADH metabolic pathway and its influence on the fermentation characteristics of the strain. Compared with R. oryzae AS 3.3461, the intracellular ATP level of the respiratory-deficient strain UV-1 decreased by 52.7 % and the glucose utilization rate rose by 8.9 %; When incubated for 36 h, the activities of phosphofructokinase (PFK), hexokinase (HK), and pyruvate kinase (PK) in the mutant rose by 74.2, 7.2, and 12.0 %, respectively; when incubated for 48 h, the intracellular NADH/NAD(+) ratio of the mutant rose by 14.6 %; when a mixed carbon source with a glucose/gluconic acid ratio of 1:1 was substituted to culture the mutant, the NADH/NAD(+) ratio decreased by 4.6 %; the ATP content dropped by 27.6 %; the lactate dehydrogenase (LDH) activity rose by 22.7 %; and the lactate yield rose by 11.6 %. These results indicated that changes to the NADH metabolic pathway under a low-energy charge level can effectively increase the glycolytic rate and further improve the yield of L-lactate of R. oryzae.

Isolation of Chlamydomonas reinhardtii mutants with altered mitochondrial respiration by chlorophyll fluorescence measurement

The unicellular green alga Chlamydomonas reinhardtii is a model organism for studying energetic metabolism. Most mitochondrial respiratory-deficient mutants characterized to date have been isolated on the basis of their reduced ability to grow in heterotrophic conditions. Mitochondrial deficiencies are usually partly compensated by adjustment of photosynthetic activity and more particularly by transition to state 2. In this work, we explored the opportunity to select mutants impaired in respiration and/or altered in dark metabolism by measuring maximum photosynthetic efficiency by chlorophyll fluorescence analyses (FV/FM). Out of about 2900 hygromycin-resistant insertional mutants generated from wild type or from a mutant strain deficient in state transitions (stt7 strain), 22 were found to grow slowly in heterotrophic conditions and 8 of them also showed a lower FV/FM value. Several disrupted coding sequences were identified, including genes coding for three different subunits of respiratory-chain complex I (NUO9, NUOA9, NUOP4) or for isocitrate lyase (ICL1). Overall, the comparison of respiratory mutants obtained in wild-type or stt7 genetic backgrounds indicated that the FV/FM value can be used to isolate mutants severely impaired in dark metabolism.

Bioethanol production from Miscanthus using thermotolerant Saccharomyces cerevisiae mbc 2 isolated from the respiration-deficient mutants

One major obstacle of simultaneous saccharification and fermentation (SSF) is that enzymatic hydrolysis in SSF is conducted at non-optimal temperature. To break through this bottleneck, thermotolerant Saccharomyces cerevisiae strains mbc 1-4 were screened by spot-assay from respiration-deficient mutants induced by ethidium bromide. When four strains placed in liquid medium, the strains exhibited significant increased growth and fermentability at 42 °C in comparison to the host. Consequently, S. cerevisiae mbc 2 was selected to be the most suitable for bioethanol production at 42 °C. Miscanthus was pretreated with 1.5 M NaOH at 150 °C for 30 min in 1 L-vessel. As a result, 42.6% of solids containing 77.9% cellulose were recovered. Next, SSF was conducted with pretreated Miscanthus containing 3% glucan to estimate the fermentability of S. cerevisiae mbc 2 at 42 °C. In results, Ethanol concentration and theoretical ethanol yield by S. cerevisiae mbc 2 in 48 h were 15.3 g/L and 90.1%, respectively, whereas those by the control were 8.3 g/L and 49.3%, respectively. In addition, SSF with increased substrate concentration upto 9% glucan resulted in 86.3% and 82.2% of theoretical ethanol yield, respectively. Thus, our approach for SSF with high solid loading using thermotolerant S. cerevisiae mbc 2 contributes to process and microbial strain development for bioethanol production.

Enhanced hexose fermentation by Saccharomyces cerevisiae through integration of stoichiometric modeling and genetic screening

In order to determine beneficial gene deletions for ethanol production by the yeast Saccharomyces cerevisiae, we performed an in silico gene deletion experiment based on a genome-scale metabolic model. Genes coding for two oxidative phosphorylation reactions (cytochrome c oxidase and ubiquinol cytochrome c reductase) were identified by the model-based simulation as potential deletion targets for enhancing ethanol production and maintaining acceptable overall growth rate in oxygen-limited conditions. Since the two target enzymes are composed of multiple subunits, we conducted a genetic screening study to evaluate the in silico results and compare the effect of deleting various portions of the respiratory enzyme complexes. Over two-thirds of the knockout mutants identified by the in silico study did exhibit experimental behavior in qualitative agreement with model predictions, but the exceptions illustrate the limitation of using a purely stoichiometric model-based approach. Furthermore, there was a substantial quantitative variation in phenotype among the various respiration-deficient mutants that were screened in this study, and three genes encoding respiratory enzyme subunits were identified as the best knockout targets for improving hexose fermentation in microaerobic conditions. Specifically, deletion of either COX9 or QCR9 resulted in higher ethanol production rates than the parental strain by 37% and 27%, respectively, with slight growth disadvantages. Also, deletion of QCR6 led to improved ethanol production rate by 24% with no growth disadvantage. The beneficial effects of these gene deletions were consistently demonstrated in different strain backgrounds and with four common hexoses. The combination of stoichiometric modeling and genetic screening using a systematic knockout collection was useful for narrowing a large set of gene targets and identifying targets of interest.

Mentha piperitaessential oil induces apoptosis in yeast associated with both cytosolic and mitochondrial ROS-mediated damage

Mentha piperita (MP), also known as peppermint, is an aromatic and medicinal plant widely used in the food industry, perfumery and cosmetic, pharmacy and traditional medicine. Its essential oil (EO) displays antimicrobial activity against a range of bacteria and fungi. In this study, we found that MP EO lethal cytotoxicity is associated with increased levels of intracellular reactive oxygen species, mitochondrial fragmentation and chromatin condensation, without loss of the plasma membrane integrity, indicative of an apoptotic process. Overexpression of cytosolic catalase and superoxide dismutases reverted the lethal effects of the EO and of its major component menthol. Conversely, deficiency in Sod1p (cytosolic copper-zinc-superoxide dismutase) greatly increased sensitivity to both agents, but deficiency in Sod2p (mitochondrial manganese superoxide dismutase) only induced sensitivity under respiratory growth conditions. Mentha piperita EO increased the frequency of respiratory deficient mutants indicative of damage to the mitochondrial genome, although increase in mitochondrial thiol oxidation does not seem to be involved in the EO toxicity.

Respiratory-deficient mutants of the unicellular green alga Chlamydomonas: A review

Genetic manipulation of the unicellular green alga Chlamydomonas reinhardtii is straightforward. Nuclear genes can be interrupted by insertional mutagenesis or targeted by RNA interference whereas random or site-directed mutagenesis allows the introduction of mutations in the mitochondrial genome. This, combined with a screen that easily allows discriminating respiratory-deficient mutants, makes Chlamydomonas a model system of choice to study mitochondria biology in photosynthetic organisms. Since the first description of Chlamydomonas respiratory-deficient mutants in 1977 by random mutagenesis, many other mutants affected in mitochondrial components have been characterized. These respiratory-deficient mutants increased our knowledge on function and assembly of the respiratory enzyme complexes. More recently some of these mutants allowed the study of mitochondrial gene expression processes poorly understood in Chlamydomonas.In this review, we update the data concerning the respiratory components with a special focus on the assembly factors identified on other organisms. In addition, we make an inventory of different mitochondrial respiratory mutants that are inactivated either on mitochondrial or nuclear genes.

Dynamic model‐based analysis of furfural and HMF detoxification by pure and mixed batch cultures of S. cerevisiae and S. stipitis

Inhibitory compounds that result from biomass hydrolysis are an obstacle to the efficient production of second-generation biofuels. Fermentative microorganisms can reduce compounds such as furfural and 5-hydroxymethyl furfural (HMF), but detoxification is accompanied by reduced growth rates and ethanol yields. In this study, we assess the effects of these furan aldehydes on pure and mixed yeast cultures consisting of a respiratory deficient mutant of Saccharomyces cerevisiae and wild-type Scheffersomyces stipitis using dynamic flux balance analysis. Uptake kinetics and stoichiometric equations for the intracellular reduction reactions associated with each inhibitor were added to genome-scale metabolic reconstructions of the two yeasts. Further modification of the S. cerevisiae metabolic network was necessary to satisfactorily predict the amount of acetate synthesized during HMF reduction. Inhibitory terms that captured the adverse effects of the furan aldehydes and their corresponding alcohols on cell growth and ethanol production were added to attain qualitative agreement with batch experiments conducted for model development and validation. When the two yeasts were co-cultured in the presence of the furan aldehydes, inoculums that reduced the synthesis of highly toxic acetate produced by S. cerevisiae yielded the highest ethanol productivities. The model described here can be used to generate optimal fermentation strategies for the simultaneous detoxification and fermentation of lignocellulosic hydrolysates by S. cerevisiae and/or S. stipitis.

Co-operative inhibitory effects of hydrogen peroxide and iodine against bacterial and yeast species.

BackgroundHydrogen peroxide and iodine are powerful antimicrobials widely used as antiseptics and disinfectants. Their antimicrobial properties are known to be enhanced by combining them with other compounds. We studied co-operative inhibitory activities (synergism, additive effects and modes of growth inhibition) of hydrogen peroxide and iodine used concurrently against 3 bacterial and 16 yeast species.ResultsSynergistic or additive inhibitory effects were shown for hydrogen peroxide and iodine mixtures against all 19 species used in the study. Both biocides were mostly cidal individually and in mixtures against Pseudomonas aeruginosa and Staphylococcus aureus. Both compounds manifested static inhibitory effects individually, but their mixtures were synergistically cidal for Saccharomyces cerevisiae and Escherihia coli. Cells of S. cerevisiae treated with hydrogen peroxide and iodine-hydrogen peroxide mixture produced increased numbers of respiratory deficient mutants indicating genotoxic effects.ConclusionIodine and hydrogen peroxide used concurrently interact synergistically or additively against a range of prokaryotic and eukaryotic microorganisms. The study provides an insight as to how these traditional antimicrobials could be used more effectively for disinfection and antisepsis. In addition, a simple approach is proposed for scoring genotoxicity of different biocides by using the budding yeast system.