Elevated energy costs of biomass production in mitochondrial respiration-deficient Saccharomyces cerevisia.

Microbial growth requires energy for maintaining the existing cells and producing components for the new ones. Microbes therefore invest a considerable amount of their resources into proteins needed for energy harvesting. Growth in different environments is associated with different energy demands for growth of yeast Saccharomyces cerevisiae, although the cross-condition differences remain poorly characterized. Furthermore, a direct comparison of the energy costs for the biosynthesis of the new biomass across conditions is not feasible experimentally; computational models, on the contrary, allow comparing the optimal metabolic strategies and quantify the respective costs of energy and nutrients. Thus in this study, we used a resource allocation model of S. cerevisiae to compare the optimal metabolic strategies between different conditions. We found that S. cerevisiae with respiratory-impaired mitochondria required additional energetic investments for growth, while growth on amino acid-rich media was not affected. Amino acid supplementation in anaerobic conditions also was predicted to rescue the growth reduction in mitochondrial respiratory shuttle-deficient mutants of S. cerevisiae. Collectively, these results point to elevated costs of resolving the redox imbalance caused by de novo biosynthesis of amino acids in mitochondria. To sum up, our study provides an example of how resource allocation modeling can be used to address and suggest explanations to open questions in microbial physiology.

Assessment of quorum sensing effects of tyrosol on fermentative performance by chief ethnic fermentative yeasts from northeast India.

AIM: Tyrosol, a quorum sensing molecule in yeasts, was reported to reduce lag phase and induces hyphae formation during cell proliferation. However, evidence of any enhancing effect of tyrosol in cellular proliferation within fermentative environment is unclear. In this investigation, selected yeast cells were assessed for their ability to synthesize tyrosol followed by examining the role of the molecule during fermentation. METHODS AND RESULTS: Tyrosols were characterized in four fermentative yeasts viz., Saccharomyces cerevisiae, Wickerhamomyces anomalus, Candida glabrata and Candida tropicalis isolated from traditional fermentative cakes of northeast India. All the isolates synthesized tyrosol while C. tropicalis exhibited filamentous growth in response to tyrosols retrieved from other isolates. Purified tyrosols showed protective behaviour in C. tropicalis and S. cerevisiae under ethanol mediated oxidative stress. During fermentation, tyrosol significantly enhanced growth of W. anomalus in starch medium while C. tropicalis exhibited growth enhancement in starch and glucose sources. The chief fermentative yeast S. cerevisiae showed notable enhancement in fermentative capacity in starch medium under the influence of tyrosol con-commitment of ethanol production. CONCLUSION: The study concludes that tyrosol exerts unusual effect in cellular growth and fermentative ability of both Saccharomyces and non-Saccharomyces yeasts. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report of expression of tyrosol by non-conventional yeasts, where the molecule was found to exert enhancing effect during fermentation, thereby augmenting the process of metabolite production during traditional fermentation.

Characterization of Saccharomyces uvarum (Beijerinck, 1898) and related hybrids: assessment of molecular markers that predict the parent and hybrid genomes and a proposal to name yeast hybrids.

The use of the nuclear DNA reassociation technique has led taxonomists to consider Saccharomyces uvarum a synonym of S. bayanus. The latter, however, is not a species but a hybrid harbouring S. eubayanus (Seu) and S. uvarum (Su) subgenomes with a minor DNA contribution from S. cerevisiae (Sc). To recognize genetically pure lines of S. uvarum and putative interspecies hybrids among so-called S. bayanus strains present in public culture collections, we propose the use of four markers that were defined from the S. bayanus CBS 380T composite genome, namely SeuNTS2 (rDNA), ScMAL31, MTY1 and SuMEL1. Saccharomyces carlsbergensis CBS 1513 was found to be similar to S. bayanus except that it carries the SeuMEL1 allele. Different marker combinations revealed that among 33 strains examined only a few were similar to CBS 380T, but many pure S. uvarum lines and putative Su/Seu-related hybrids occurred. Our results demonstrated that these hybrids were erroneously considered authentic S. bayanus and therefore the varietal state 'Saccharomyces bayanus var. uvarum comb. nov. Naumov' is not valid. Our markers constitute a tool to get insights into the genomic makeup of Saccharomyces interspecies hybrids. We also make a proposal to name those hybrids that may also be applicable to other fungal hybrids.

Survey of cryptic unstable transcripts in yeast.

BACKGROUND: Cryptic unstable transcripts (CUTs) are a largely unexplored class of nuclear exosome degraded, non-coding RNAs in budding yeast. It is highly debated whether CUT transcription has a functional role in the cell or whether CUTs represent noise in the yeast transcriptome. We sought to ascertain the extent of conserved CUT expression across a variety of Saccharomyces yeast strains to further understand and characterize the nature of CUT expression. RESULTS: We sequenced the WT and rrp6Δ transcriptomes of three S.cerevisiae strains: S288c, Σ1278b, JAY291 and the S.paradoxus strain N17 and utilized a hidden Markov model to annotate CUTs in these four strains. Utilizing a four-way genomic alignment we identified a large population of CUTs with conserved syntenic expression across all four strains. By identifying configurations of gene-CUT pairs, where CUT expression originates from the gene 5' or 3' nucleosome free region, we observed distinct gene expression trends specific to these configurations which were most prevalent in the presence of conserved CUT expression. Divergent pairs correlate with higher expression of genes, and convergent pairs correlate with reduced gene expression. CONCLUSIONS: Our RNA-seq based method has greatly expanded upon previous CUT annotations in S.cerevisiae underscoring the extensive and pervasive nature of unstable transcription. Furthermore we provide the first assessment of conserved CUT expression in yeast and globally demonstrate possible modes of CUT-based regulation of gene expression.

Saccharomyces boulardii and bismuth subsalicylate as low-cost interventions to reduce the duration and severity of cholera.

We conducted a randomised single-blinded clinical trial of 100 cholera patients in Port-au-Prince, Haiti to determine if the probiotic Saccharomyces cerevisiae var. boulardii and the anti-diarrhoeal drug bismuth subsalicylate (BS) were able to reduce the duration and severity of cholera. Subjects received either: S. boulardii 250 mg, S. boulardii 250 mg capsule plus BS 524 mg tablet, BS 524 mg, or two placebo capsules every 6 hours alongside standard treatment for cholera. The length of hospitalisation plus the number and volume of emesis, stool and urine were recorded every 6 hours until the study subject was discharged (n = 83), left against medical advice (n = 11), or requested removal from the study (n = 6). There were no reported deaths or adverse study-related events. There were no statistically significant differences between the study arms and the outcomes of interest.

Comparative analysis of DNA word abundances in four yeast genomes using a novel statistical background model.

Previous studies have shown that the identification and analysis of both abundant and rare k-mers or "DNA words of length k" in genomic sequences using suitable statistical background models can reveal biologically significant sequence elements. Other studies have investigated the uni/multimodal distribution of k-mer abundances or "k-mer spectra" in different DNA sequences. However, the existing background models are affected to varying extents by compositional bias. Moreover, the distribution of k-mer abundances in the context of related genomes has not been studied previously. Here, we present a novel statistical background model for calculating k-mer enrichment in DNA sequences based on the average of the frequencies of the two (k-1) mers for each k-mer. Comparison of our null model with the commonly used ones, including Markov models of different orders and the single mismatch model, shows that our method is more robust to compositional AT-rich bias and detects many additional, repeat-poor over-abundant k-mers that are biologically meaningful. Analysis of overrepresented genomic k-mers (4≤k≤16) from four yeast species using this model showed that the fraction of overrepresented DNA words falls linearly as k increases; however, a significant number of overabundant k-mers exists at higher values of k. Finally, comparative analysis of k-mer abundance scores across four yeast species revealed a mixture of unimodal and multimodal spectra for the various genomic sub-regions analyzed.

Biodiversity of indigenous Saccharomyces populations from old wineries of south-eastern Sicily (Italy): preservation and economic potential.

In recent years, the preservation of biodiversity has become an important issue. Despite much public discussion, however, current practices in the food industry seldom take account of its potential economic importance: on the contrary, the introduction of industrialized agriculture practices over large areas has often resulted in a dramatic reduction in biodiversity.In this paper, we report on the remarkable degree of biodiversity in the wine yeast populations naturally present in a small area of Sicily (Italy) where traditional (non-industrial) winery practices are still in place. Out of more than 900 Saccharomyces yeast isolates recovered from late spontaneous fermentations, we detected at least 209 strains. Most interestingly, when evaluated at the fermentation and technological level, a number of isolates were found to be superior to industrial yeast strains. Out of a selected group, isolates from two strains were used for experimental fermentations in a winery environment and the quality of the wines produced was assessed at the technological, quality and sensory levels. Given that the characteristics of the wines produced were found to be industrially appealing, the study demonstrated the economic potential of preserving the patrimony of Sicilian yeast biodiversity and highlighted the importance of maintaining traditional wine making practices.

Safety assessment studies of probiotic Saccharomyces boulardii strain Unique 28 in Sprague-Dawley rats.

Strains of Saccharomyces boulardii, a probiotic yeast, have been found to be effective in the treatment of diarrhoea, inflammatory bowel disease, irritable bowel syndrome and other conditions. In the present study, Unique 28, a strain of S. boulardii isolated and characterised in our laboratory, was evaluated for its safety assessment. Acute and subacute toxicity tests were performed in rats. The dose of Unique 28 (5×10(9) cfu/g) fed orally was, up to 6,500 mg per kg of b.w. (body weight) for acute toxicity and up to 1,300 mg per kg of b.w. for sub-acute toxicity studies. This dose was well tolerated and there was no morbidity or any kind of toxic clinical symptoms displayed either in male or female rats. Moreover, the results of sub-acute toxicity studies using Unique 28 administered for 14 weeks indicated that there were no clear unwanted treatment related effects. Overall results of this toxicology assessment indicate that Unique 28 is safe for human consumption.

Unifying vertical and nonvertical evolution: a stochastic ARG-based framework.

Evolutionary biologists have introduced numerous statistical approaches to explore nonvertical evolution, such as horizontal gene transfer, recombination, and genomic reassortment, through collections of Markov-dependent gene trees. These tree collections allow for inference of nonvertical evolution, but only indirectly, making findings difficult to interpret and models difficult to generalize. An alternative approach to explore nonvertical evolution relies on phylogenetic networks. These networks provide a framework to model nonvertical evolution but leave unanswered questions such as the statistical significance of specific nonvertical events. In this paper, we begin to correct the shortcomings of both approaches by introducing the "stochastic model for reassortment and transfer events" (SMARTIE) drawing upon ancestral recombination graphs (ARGs). ARGs are directed graphs that allow for formal probabilistic inference on vertical speciation events and nonvertical evolutionary events. We apply SMARTIE to phylogenetic data. Because of this, we can typically infer a single most probable ARG, avoiding coarse population dynamic summary statistics. In addition, a focus on phylogenetic data suggests novel probability distributions on ARGs. To make inference with our model, we develop a reversible jump Markov chain Monte Carlo sampler to approximate the posterior distribution of SMARTIE. Using the BEAST phylogenetic software as a foundation, the sampler employs a parallel computing approach that allows for inference on large-scale data sets. To demonstrate SMARTIE, we explore 2 separate phylogenetic applications, one involving pathogenic Leptospirochete and the other Saccharomyces.

An economic comparison of different fermentation configurations to convert corn stover to ethanol using Z. mobilis and Saccharomyces.

Numerous routes are being explored to lower the cost of cellulosic ethanol production and enable large-scale production. One critical area is the development of robust cofermentative organisms to convert the multiple, mixed sugars found in biomass feedstocks to ethanol at high yields and titers without the need for processing to remove inhibitors. Until such microorganisms are commercialized, the challenge is to design processes that exploit the current microorganisms' strengths. This study explored various process configurations tailored to take advantage of the specific capabilities of three microorganisms, Z. mobilis 8b, S. cerevisiae, and S. pastorianus. A technoeconomic study, based on bench-scale experimental data generated by integrated process testing, was completed to understand the resulting costs of the different process configurations. The configurations included whole slurry fermentation with a coculture, and separate cellulose simultaneous saccharification and fermentation (SSF) and xylose fermentations with none, some or all of the water to the SSF replaced with the fermented liquor from the xylose fermentation. The difference between the highest and lowest ethanol cost for the different experimental process configurations studied was $0.27 per gallon ethanol. Separate fermentation of solid and liquor streams with recycle of fermented liquor to dilute the solids gave the lowest ethanol cost, primarily because this option achieved the highest concentrations of ethanol after fermentation. Further studies, using methods similar to ones employed here, can help understand and improve the performance and hence the economics of integrated processes involving enzymes and fermentative microorganisms.

Alignment uncertainty and genomic analysis.

The statistical methods applied to the analysis of genomic data do not account for uncertainty in the sequence alignment. Indeed, the alignment is treated as an observation, and all of the subsequent inferences depend on the alignment being correct. This may not have been too problematic for many phylogenetic studies, in which the gene is carefully chosen for, among other things, ease of alignment. However, in a comparative genomics study, the same statistical methods are applied repeatedly on thousands of genes, many of which will be difficult to align. Using genomic data from seven yeast species, we show that uncertainty in the alignment can lead to several problems, including different alignment methods resulting in different conclusions.

Role of ASCA and the NOD2/CARD15 mutation Gly908Arg in predicting increased surgical costs in Crohn's disease patients: a project of the European Collaborative Study Group on Inflammatory Bowel Disease.

BACKGROUND: NOD2/CARD15, the first identified susceptibility gene in Crohn's disease (CD), is associated with ileal stenosis and increased frequency of surgery. Anti-Saccharomyces cerevisiae antibody (ASCA), a serological marker for CD, is associated with ileal location and a high likelihood for surgery. We hypothesized that the presence of ASCA and NOD2/CARD15 mutations could predict increased health care cost in CD. METHODS: CD patients in a prospectively designed community-based multinational European and Israeli cohort (n = 228) followed for mean 8.3 (SD 2.6) years had blood drawn for measurement of ASCA (IgG, IgA), Arg702Trp, Gly908Arg, and Leu1007fsinsC. Days spent in the hospital and the costs of medical and surgical hospitalizations and medications were calculated. RESULTS: The median duration of surgical hospitalizations was longer in Gly908Arg-positive than -negative patients, 3.5 and 1.5 days/patient-year (P < 0.01), and in ASCA-positive than -negative patients, 1.1 and 0 days/patient-year (P < 0.001). Median surgical hospitalization cost was 1,580 euro/patient-year in Gly908Arg-positive versus 0 euro/patient-year in -negative patients (P < 0.01), and 663 euro/patient-year in ASCA-positive versus 0 euro/patient-year in -negative patients (P < 0.001). Differences in cost of medications between groups were not significant. The effect of Gly908Arg was expressed in countries with higher Gly908Arg carriage rates. ASCA raised surgical costs independently of the age at diagnosis of disease. Arg702Trp and Leu1007fsinsC did not affect the cost of health care. CONCLUSIONS: Since CD patients positive for Gly908Arg and ASCA demonstrated higher health care costs, it is possible that measurement of Gly908Arg and ASCA at disease diagnosis can forecast the expensive CD patients.

Bayesian estimation of concordance among gene trees.

Multigene sequence data have great potential for elucidating important and interesting evolutionary processes, but statistical methods for extracting information from such data remain limited. Although various biological processes may cause different genes to have different genealogical histories (and hence different tree topologies), we also may expect that the number of distinct topologies among a set of genes is relatively small compared with the number of possible topologies. Therefore evidence about the tree topology for one gene should influence our inferences of the tree topology on a different gene, but to what extent? In this paper, we present a new approach for modeling and estimating concordance among a set of gene trees given aligned molecular sequence data. Our approach introduces a one-parameter probability distribution to describe the prior distribution of concordance among gene trees. We describe a novel 2-stage Markov chain Monte Carlo (MCMC) method that first obtains independent Bayesian posterior probability distributions for individual genes using standard methods. These posterior distributions are then used as input for a second MCMC procedure that estimates a posterior distribution of gene-to-tree maps (GTMs). The posterior distribution of GTMs can then be summarized to provide revised posterior probability distributions for each gene (taking account of concordance) and to allow estimation of the proportion of the sampled genes for which any given clade is true (the sample-wide concordance factor). Further, under the assumption that the sampled genes are drawn randomly from a genome of known size, we show how one can obtain an estimate, with credibility intervals, on the proportion of the entire genome for which a clade is true (the genome-wide concordance factor). We demonstrate the method on a set of 106 genes from 8 yeast species.

Scanning sequences after Gibbs sampling to find multiple occurrences of functional elements.

BACKGROUND: Many DNA regulatory elements occur as multiple instances within a target promoter. Gibbs sampling programs for finding DNA regulatory elements de novo can be prohibitively slow in locating all instances of such an element in a sequence set. RESULTS: We describe an improvement to the A-GLAM computer program, which predicts regulatory elements within DNA sequences with Gibbs sampling. The improvement adds an optional "scanning step" after Gibbs sampling. Gibbs sampling produces a position specific scoring matrix (PSSM). The new scanning step resembles an iterative PSI-BLAST search based on the PSSM. First, it assigns an "individual score" to each subsequence of appropriate length within the input sequences using the initial PSSM. Second, it computes an E-value from each individual score, to assess the agreement between the corresponding subsequence and the PSSM. Third, it permits subsequences with E-values falling below a threshold to contribute to the underlying PSSM, which is then updated using the Bayesian calculus. A-GLAM iterates its scanning step to convergence, at which point no new subsequences contribute to the PSSM. After convergence, A-GLAM reports predicted regulatory elements within each sequence in order of increasing E-values, so users have a statistical evaluation of the predicted elements in a convenient presentation. Thus, although the Gibbs sampling step in A-GLAM finds at most one regulatory element per input sequence, the scanning step can now rapidly locate further instances of the element in each sequence. CONCLUSION: Datasets from experiments determining the binding sites of transcription factors were used to evaluate the improvement to A-GLAM. Typically, the datasets included several sequences containing multiple instances of a regulatory motif. The improvements to A-GLAM permitted it to predict the multiple instances.

Yeast (different sources and levels) as protein source in diets of reared piglets: effects on protein digestibility and N-metabolism.

The aim of this study was to examine the feeding value of different yeasts as a substitute for soya bean meal, the main protein source in diets of weaned piglets. Tested two yeasts were already available on the market, Saccharomyces cerevisiae and Kluyveromyces lactis (beer and milk yeast), which replaced 40% of the soya bean meal in the diets. Furthermore, a yeast (Kluyveromyces fragilis) grown on whey, a side-product of cheese production, was used in increasing concentrations in the diets, so that increasing amounts of the soya bean meal (20%, 40% and 60%) could be replaced. As proved in these experiments, a replacement of 60% of the soya protein with whey yeast protein had positive effects on the performances (daily weight gain) and on the N-metabolism and did not have negative effects on the health or the faeces consistency. The whey yeast stands out because of its high protein quality (N-digestibility and N-retention). Furthermore, the replacement of soya bean meal with highly digestible yeasts is welcomed under the aspect of animal health, because of the reduction of anti-nutritive soya components (stachyose, glycinin) in diets of weaned piglets. The controlled production conditions of the yeasts result in a high feed safety; in addition, the yeast as an end-of-pipe-product is a resource conserving and valuable feed. A main stimulus for the use of yeasts, however, in a food production controlled by economic standpoints, is their price and the costs of other competing feeds.

[Technology for the whole utilization of brewer's yeast in food industry]. / Tecnología para la utilización integral de la levadura de cerveza en la industria alimenticia.

A flexible scheme for the fractionation of brewer's yeast was developed. The procedure allows the production of different products such as: dry yeast flakes, dry yeast pills, yeast-extract based table sauce, yeast protein concentrates and soy-like sauce. The investment required for the processing of one ton per day is below 2 million dollars with an overall profitability higher than 53%. Investment is recovered in 0.75 years. The production of food ingredients from yeast upgrades its biomass about 25 fold. Present procedure is compared with other biomass fractionation processes taking into account the utilization of all technological streams where the process becomes environmentally friendly since effluent production significantly lower than similar technologies.

Overexpression of multisubunit replication factors in yeast.

Facile genetic and biochemical manipulation coupled with rapid cell growth and low cost of growth media has established the yeast Saccharomyces cerevisiae as a versatile workhorse. This article describes the use of yeast expression systems for the overproduction of complex multipolypeptide replication factors. The regulated overexpression of these factors in yeast provides for a readily accessible and inexpensive source of these factors in large quantities. The methodology is illustrated with the five-subunit replication factor C. Whole-cell extracts are prepared by blending yeast cells with glass beads or frozen yeast with dry ice. Procedures are described that maximize the yield of these factors while minimizing proteolytic degradation.

A microbiology investigation on probiotic pharmaceutical products used for human health.

Many and different probiotic pharmaceutical products are presently commercialised in the world. On this regard, a microbiological investigation was carried out to screen the microorganisms incorporated into these products, commonly used for human health. After determination of the cell number and viability of bacteria, several experiments were performed in vitro in order to characterise the microorganisms and to evaluate their probiotic value. Among all the strains identified, best results were obtained with Lactobacillus rhamnosus, Enterococcus faecium and Saccharomyces cerevisiae as far as regards growth rates, pH and bile salts tolerance. Moreover, the identification profiles of microorganisms showed a better reliability for the products containing a single species whereas the ones composed of different strains were usually not satisfactory. In some cases, the presence of Lactobacillus and Saccharomyces species was in disagreement with the claimed composition of the product and some species of lactobacilli, bifidobacteria and streptococci were found not viable. In defined mixed cultures experiments, the antagonism of Lactobacillus acidophilus and Enterococcus faecium versus Yersinia enterocolitica was demonstrated and explained as acid and/or antimicrobials production.

Atypical regions in large genomic DNA sequences.

Large genomic DNA sequences contain regions with distinctive patterns of sequence organization. We describe a method using logarithms of probabilities based on seventh-order Markov chains to rapidly identify genomic sequences that do not resemble models of genome organization built from compilations of octanucleotide usage. Data bases have been constructed from Escherichia coli and Saccharomyces cerevisiae DNA sequences of > 1000 nt and human sequences of > 10,000 nt. Atypical genes and clusters of genes have been located in bacteriophage, yeast, and primate DNA sequences. We consider criteria for statistical significance of the results, offer possible explanations for the observed variation in genome organization, and give additional applications of these methods in DNA sequence analysis.

Evaluation of two commercialized systems for the rapid identification of medically important yeasts.

A total of 77 recent clinical isolates of Candida albicans and other medically important yeasts were identified by two different commercial tests, Rapidec albicans (API-bioMérieux) and Fongiscreen 4H (Sanofi Diagnostics Pasteur), and conventional mycological methods. The strains were from 13 different species of yeasts and consisted of strains of 36 C. albicans, three of Candida famata, nine of Candida (Torulopsis) glabrata, five of Candida guilliermondii, two of Candida kefyr, three of Candida krusei, one of Candida lusitaniae, four of Cryptococcus neoformans, five of Candida parapsilosis, six of Candida tropicalis, one of Candida viswanathii, one of Rhodotorula rubra and one of Saccharomyces cerevisiae. According to the reactivity profiles of the isolates, identification was always correct with Fongiscreen 4H and was correct in 97.3% of the strains with Rapidec albicans. The latter test did not identify two C. albicans isolates that were correctly identified by Fongiscreen 4H. Both methods (97.3% correlation) were very useful for identification of C. albicans achieving the aim of their manufacturers. Additionally, Fongiscreen 4H was very useful for the identification of three other species of yeasts: C. glabrata, C. tropicalis and Cr. neoformans. The results of our study indicate that the accuracy of Rapidec albicans and Fongiscreen 4H is similar to that of the conventional methods used in this study for the identification of C. albicans. The same is true of Fongiscreen 4H in the identification of C. glabrata, C. tropicalis and Cr. neoformans. Both tests could be rapid and easy-to-perform tools in the clinical microbiology laboratory, but differences in cost must be taken into account.