Journal: FEMS yeast research
We aim in this study to provide levels of susceptibility of 162 bloodstream isolates of non-Candida albicans and non-C. tropicalis species from a sentinel program conducted in 11 hospitals in Brazil. Additionally, we compared the broth microdilution (BMD) method of the European Committee of Susceptibility Testing (EUCAST) with Clinical Laboratory Standards Institute (CLSI) BMD method for fluconazole, itraconazole, voriconazole, and amphotericin B. The study included 103 C. parapsilosis, 38 C. glabrata, 8 C. orthopsilosis, and 7 C. krusei isolates, and single isolates of Pichia anomala, C. famata, C. lusitaniae, C. kefyr, C. guilliermondii, and C. metapsilosis. Of note, we observed cross-resistance between fluconazole and voriconazole for two isolates being one C. parapsilosis and one C. glabrata. Good essential agreement (EA) was observed between the EUCAST and the CLSI results for C. parapsilosis and for fluconazole, itraconazole, voriconazole, and amphotericin B, respectively: 98%, 99%, 98%, and 97%. Otherwise, for C. glabrata, the EA for fluconazole was 84.2% and for voriconazole 89.4%. Because data from Brazil are scarce, our results contribute to the consolidation of the database of candidemia agents and monitoring of trends in the profile of drug resistance.
The origin of modern fruits brought to microbial communities an abundant source of rich food based on simple sugars. Yeasts, especially Saccharomyces cerevisiae, usually become the predominant group in these niches. One of the most prominent and unique features and likely a winning trait of these yeasts is their ability to rapidly convert sugars to ethanol at both anaerobic and aerobic conditions. Why, when and how did yeast remodel their carbon metabolism to be able to accumulate ethanol under aerobic conditions and at the expense of decreasing biomass production? We hereby review the recent data on the carbon metabolism in Saccharomycetaceae species, and attempt to reconstruct the ancient environment, which could promote the evolution of alcoholic fermentation. We speculate that the first step towards the so-called alcoholic fermentation lifestyle was the exploration of anaerobic niches resulting in an increased metabolic capacity to degrade sugar to ethanol. The strengthened glycolytic flow had in parallel a beneficial effect on the microbial competition outcome, and later evolved as a “new” tool promoting the yeast competition ability under aerobic conditions. The basic aerobic alcoholic fermentation ability was subsequently “upgraded” in several lineages by evolving additional regulatory steps, like glucose repression in the S. cerevisiae clade, to achieve a more precise metabolic control. This article is protected by copyright. All rights reserved.
ABC-transporters with broad substrate specificity are responsible for pathogenic yeast resistance to antifungal compounds. Here we asked whether highly hydrophobic chemicals with delocalized positive charge can be used to overcome the resistance. Such molecules efficiently penetrate the plasma membrane and accumulate inside the cells. We reasoned that these properties can convert an active efflux of the compounds into a futile cycle thus interfering with the extrusion of the antibiotics. To test this we studied the effects of several alkylated rhodamines on the drug resistance of yeastSaccharomyces cerevisiae We found that octylrhodamine synergetically increases toxicity of Pd5p substrate - clotrimazole, while the others were less effective. Next, we compared the contributions of three major pleiotropic ABC-transporters (Pdr5p, Yor1p, Snq2p) on the accumulation of the alkylated rhodamines. While all of the tested compounds were extruded by Pdr5p, Yor1p and Snq2p showed narrower substrate specificity. Interestingly, among the tested alkylated rhodamines, inactivation of Pdr5p had a strongest effect on the accumulation of octylrhodamine inside the cells, which is consistent with the fact that clotrimazole is a substrate of Pdr5p. As alkylated rhodamines were shown to be non-toxic on mice, our study makes them potential components of pharmacological antifungal compositions.
Humans have acted as vectors for species and expanded their ranges since at least the dawn of agriculture. While relatively well characterized for macrofauna and macroflora, the extent and dynamics of human-aided microbial dispersal is poorly described. We studied the role which humans have played in manipulating the distribution of Saccharomyces cerevisiae, one of the world’s most important microbes, using whole genome sequencing. We include 52 strains representative of the diversity in New Zealand to the global set of genomes for this species. Phylogenomic approaches show an exclusively European origin of the New Zealand population, with a minimum of ten founder events mostly taking place over the last 1,000 years. Our results show that humans have expanded the range of S. cerevisiae and transported it to New Zealand where it was not previously present, where it has now become established in vineyards, but radiation to native forests appears limited.
Simultaneous fermentation of glucose and xylose can contribute to improved productivity and robustness of yeast-based processes for bioethanol production from lignocellulosic hydrolysates. This study explores a novel laboratory evolution strategy for identifying mutations that contribute to simultaneous utilization of these sugars in batch cultures of Saccharomyces cerevisiae. To force simultaneous utilization of xylose and glucose, the genes encoding glucose-6-phosphate isomerase (PGI1) and ribulose-5-phosphate epimerase (RPE1) were deleted in a xylose-isomerase-based xylose-fermenting strain with a modified oxidative pentose-phosphate pathway. Laboratory evolution of this strain in serial batch cultures on glucose-xylose mixtures yielded mutants that rapidly co-consumed the two sugars. Whole-genome sequencing of evolved strains identified mutations in HKX2, RSP5 and GAL83, whose introduction into a non-evolved xylose-fermenting S. cerevisiae strain improved co-consumption of xylose and glucose under aerobic and anaerobic conditions. Combined deletion of HXK2 and introduction of a GAL83G673T allele yielded a strain with a 2.5-fold higher xylose and glucose co-consumption ratio than its xylose-fermenting parental strain. These two modifications decreased the time required for full sugar conversion in anaerobic bioreactor batch cultures, grown on 20 g L-1 glucose and 10 g L-1 xylose, by over 24 h. This study demonstrates that laboratory evolution and genome resequencing of microbial strains engineered for forced co-consumption is a powerful approach for studying and improving simultaneous conversion of mixed substrates.
The brewing industry is experiencing a period of change and experimentation largely driven by customer demand for product diversity. This has coincided with a greater appreciation of the role of yeast in determining the character of beer and the widespread availability of powerful tools for yeast research. Genome analysis in particular has helped clarify the processes leading to domestication of brewing yeast and has identified domestication signatures that may be exploited for further yeast development. The functional properties of non-conventional yeast (both Saccharomyces and non-Saccharomyces) are being assessed with a view to creating beers with new flavours as well as producing flavoursome non-alcoholic beers. The discovery of the psychrotolerant S. eubayanus has stimulated research on de novo S. cerevisiae x S. eubayanus hybrids for low-temperature lager brewing and has led to renewed interest in the functional importance of hybrid organisms and the mechanisms that determine hybrid genome function and stability. The greater diversity of yeast that can be applied in brewing, along with an improved understanding of yeasts' evolutionary history and biology, is expected have a significant and direct impact on the brewing industry, with potential for improved brewing efficiency, product diversity and, above all, customer satisfaction.
Triacylglycerol (TAG) and glycogen are the two major metabolites for carbon storage in most eukaryotic organisms. We investigated the glycogen metabolism of the oleaginous Yarrowia lipolytica and found that this yeast accumulates up to 16% glycogen in its biomass. Assuming that elimination of glycogen synthesis would result in an improvement of lipid accumulation, we characterized and deleted the single gene coding for glycogen synthase, YlGSY1. The mutant was grown under lipogenic conditions with glucose and glycerol as substrates and we obtained up to 60% improvement in TAG accumulation compared to the wild-type strain. Additionally, YlGSY1 was deleted in a background that was already engineered for high lipid accumulation. In this obese background, TAG accumulation was also further increased. The highest lipid content of 52% was found after 3 days of cultivation in nitrogen-limited glycerol medium. Furthermore, we constructed mutants of Y. lipolytica and Saccharomyces cerevisiae that are deleted for both glycogen and TAG synthesis, demonstrating that the ability to store carbon is not essential. Overall, this work showed that glycogen synthesis is a competing pathway for TAG accumulation in oleaginous yeasts and that deletion of the glycogen synthase has beneficial effects on neutral lipid storage.
The architecture and regulation of Saccharomyces cerevisiae metabolic network are among the best studied owing to its widespread use in both basic research and industry. Yet, several recent studies have revealed notable limitations in explaining genotype-metabolic phenotype relations in this yeast, especially when concerning multiple genetic/environmental perturbations. Apparently unexpected genotype-phenotype relations may originate in the evolutionarily shaped cellular operating principles being hidden in common laboratory conditions. Predecessors of laboratory S. cerevisiae strains, the wild and the domesticated yeasts, have been evolutionarily shaped by highly variable environments, very distinct from laboratory conditions, and most interestingly by social life within microbial communities. Here we present a brief review of the genotypic and phenotypic peculiarities of S. cerevisiae in the context of its social lifestyle beyond laboratory environments. Accounting for this ecological context and the origin of the laboratory strains in experimental design and data analysis would be essential in improving the understanding of genotype-environment-phenotype relationships.
The yeast Saccharomyces cerevisiae is considered an important host for consolidated bioprocessing and the production of high titres of recombinant cellulases are required for efficient hydrolysis of lignocellulosic substrates to fermentable sugars. Since recombinant protein secretion profiles vary highly among different strain backgrounds, careful selection of robust strains with optimal secretion profiles are of crucial importance. Here, we construct and screen sets of haploid derivatives, isolated from natural strain isolates YI13, FINI and YI59, for improved general cellulase secretion. This report details a novel approach that combines secretion profiles of strains and phenotypic responses to stresses known to influence the secretion pathway for the development of a phenotypic screen to isolate strains with improved secretory capacities. A clear distinction was observed between the YI13 haploid derivatives and industrial and laboratory counterparts, Ethanol Red and S288c respectively. By using sub-lethal concentrations of the secretion stressor tunicamycin and cell wall stressor Congo Red, YI13 haploid derivative strains demonstrated tolerance profiles related to their heterologous secretion profiles. Our results demonstrate that a new screening technique combined with targeted mating approach can provide a pool of novel strains capable of high cellulase secretion.
Transcription of protein coding genes is a highly regulated process. In eukaryotes, it involves cross talk between the RNA polymerase II enzyme and different proteins of the transcriptional machinery. Twelve different subunits, Rpb1 to Rpb12, constitute RNA polymerase II. The sequence of the Rpb7 subunit is highly conserved across organisms. However, our knowledge and understanding of the role of Rpb7 in Schizosaccharomyces pombe is still limited. Therefore, in the present study we have studied the transcriptional response of Schizosaccharomyces pombe cells expressing reduced levels of rpb7+. Our global transcriptional analysis revealed that expression of genes belonging to different DNA repair pathways was down-regulated by reduced rpb7+ expression. It was observed that survival of S. pombe cells expressing low rpb7+ levels was compromised under genotoxic stress conditions. Rpb7 also exhibited genetic interaction with genes of various DNA repair pathways. Furthermore, the growth sensitivity of S. pombe cells with low rpb7+ levels under DNA damaging conditions was completely rescued by human Rpb7, indicating a functional conservation between these proteins. In summary, results from our whole-genome level gene expression analysis, as well as phenotypic and genetic experiments suggest a role for Rpb7 in DNA damage response in S. pombe.