Novel approach in developing Saccharomyces cerevisiae hybrid bioethanol producers by mating of natural isolates having desirable traits

Bioethanol production from lignocellulosic hydrolysates requires a producer strain that tolerates both the presence of growth and fermentation inhibitors and high ethanol concentrations. Therefore, we constructed heterozygous intraspecies hybrid diploids of Saccharomyces cerevisiae by crossing two natural S. cerevisiae isolates, YIIc17_E5 and UWOPS87-2421, a good ethanol producer found in wine and a strain from the flower of the cactus Opuntia megacantha resistant to inhibitors found in lignocellulosic hydrolysates, respectively. Hybrids grew faster than parental strains in the absence and in the presence of acetic and levulinic acids and 2-furaldehyde, inhibitors frequently found in lignocellulosic hydrolysates, and the overexpression of YAP1 gene increased their survival. Furthermore, although originating from the same parental strains, hybrids displayed different fermentative potential in a CO2 production test, suggesting genetic variability that could be used for further selection of desirable traits. Therefore, our results suggest that the construction of intraspecies hybrids coupled with the use of genetic engineering techniques is a promising approach for improvement or development of new biotechnologically relevant strains of S. cerevisiae. Moreover, it was found that the success of gene targeting (gene targeting fidelity) in natural S. cerevisiae isolates (YIIc17_E5α and UWOPS87-2421α) was strikingly lower than in laboratory strains and the most frequent off-targeting event was targeted chromosome duplication.

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Metabolic Engineering of Wine Strains of Saccharomyces cerevisiae

Genes ◽

10.3390/genes11090964 ◽

2020 ◽

Vol 11 (9) ◽

pp. 964

Author(s):

Mikhail A. Eldarov ◽

Andrey V. Mardanov

Keyword(s):

Saccharomyces Cerevisiae ◽

Metabolic Engineering ◽

Wine Yeast ◽

Strain Engineering ◽

Starter Cultures ◽

Detailed Knowledge ◽

Wine Quality ◽

Wine Strain ◽

Natural Isolates ◽

Genomic Editing

Modern industrial winemaking is based on the use of starter cultures of specialized wine strains of Saccharomyces cerevisiae yeast. Commercial wine strains have a number of advantages over natural isolates, and it is their use that guarantees the stability and reproducibility of industrial winemaking technologies. For the highly competitive wine market with new demands for improved wine quality, it has become increasingly critical to develop new wine strains and winemaking technologies. Novel opportunities for precise wine strain engineering based on detailed knowledge of the molecular nature of a particular trait or phenotype have recently emerged due to the rapid progress in genomic and “postgenomic” studies with wine yeast strains. The review summarizes the current achievements of the metabolic engineering of wine yeast, the results of recent studies and the prospects for the application of genomic editing technologies for improving wine S. cerevisiae strains.

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Ploidy-Regulated Variation in Biofilm-Related Phenotypes in Natural Isolates of Saccharomyces cerevisiae

G3 Genes|Genome|Genetics ◽

10.1534/g3.114.013250 ◽

2014 ◽

Vol 4 (9) ◽

pp. 1773-1786 ◽

Cited By ~ 23

Author(s):

E. A. Hope ◽

M. J. Dunham

Keyword(s):

Saccharomyces Cerevisiae ◽

Natural Isolates

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Population Analysis and Evolution of Saccharomyces cerevisiae Mitogenomes

Microorganisms ◽

10.3390/microorganisms8071001 ◽

2020 ◽

Vol 8 (7) ◽

pp. 1001

Author(s):

Daniel Vieira ◽

Soraia Esteves ◽

Carolina Santiago ◽

Eduardo Conde-Sousa ◽

Ticiana Fernandes ◽

...

Keyword(s):

Saccharomyces Cerevisiae ◽

Yeast Species ◽

Phenotypic Diversity ◽

Population Analysis ◽

Nuclear Genome ◽

Reference Sequence ◽

Evolutionary Analysis ◽

Novel Approach ◽

History Of ◽

First Time

The study of mitogenomes allows the unraveling of some paths of yeast evolution that are often not exposed when analyzing the nuclear genome. Although both nuclear and mitochondrial genomes are known to determine phenotypic diversity and fitness, no concordance has yet established between the two, mainly regarding strains’ technological uses and/or geographical distribution. In the current work, we proposed a new method to align and analyze yeast mitogenomes, overcoming current difficulties that make it impossible to obtain comparable mitogenomes for a large number of isolates. To this end, 12,016 mitogenomes were considered, and we developed a novel approach consisting of the design of a reference sequence intended to be comparable between all mitogenomes. Subsequently, the population structure of 6646 Saccharomyces cerevisiae mitogenomes was assessed. Results revealed the existence of particular clusters associated with the technological use of the strains, in particular regarding clinical isolates, laboratory strains, and yeasts used for wine-associated activities. As far as we know, this is the first time that a positive concordance between nuclear and mitogenomes has been reported for S. cerevisiae, in terms of strains’ technological applications. The results obtained highlighted the importance of including the mtDNA genome in evolutionary analysis, in order to clarify the origin and history of yeast species.

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Enforced Mutualism Leads to Improved Cooperative Behavior between Saccharomyces cerevisiae and Lactobacillus plantarum

Microorganisms ◽

10.3390/microorganisms8081109 ◽

2020 ◽

Vol 8 (8) ◽

pp. 1109

Author(s):

S. Christine du Toit ◽

Debra Rossouw ◽

Maret du Toit ◽

Florian F. Bauer

Keyword(s):

Saccharomyces Cerevisiae ◽

Amino Acids ◽

Lactobacillus Plantarum ◽

Species Interactions ◽

Cooperative Behavior ◽

Synthetic Medium ◽

Grape Juice ◽

Growth Patterns ◽

Microbial Interactions ◽

Novel Approach

Saccharomyces cerevisiae and Lactobacillus plantarum are responsible for alcoholic and malolactic fermentation, respectively. Successful completion of both fermentations is essential for many styles of wine, and an understanding of how these species interact with each other, as well as the development of compatible pairings of these species, will help to manage the process. However, targeted improvements of species interactions are difficult to perform, in part because of the chemical and biological complexity of natural grape juice. Synthetic ecological systems reduce this complexity and can overcome these difficulties. In such synthetic systems, mutualistic growth of different species can be enforced through the reciprocal exchange of essential nutrients. Here, we implemented a novel approach to evolve mutualistic traits by establishing a co-dependent relationship between S. cerevisiae BY4742Δthi4 and Lb. plantarum IWBT B038 by omitting different combinations of amino acids from a chemically defined synthetic medium simulating standard grape juice. After optimization, the two species were able to support the growth of each other when grown in the absence of appropriate combinations of amino acids. In these obligatory mutualistic conditions, BY4742Δthi4 and IWBT B038 were co-evolved for approximately 100 generations. The selected evolved isolates showed improved mutualistic growth and the growth patterns under non-selective conditions indicate the emergence of mutually beneficial adaptations independent of the synthetic selection pressure. The combined use of synthetic ecology and co-evolution is a promising strategy to better understand and biotechnologically improve microbial interactions.

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A novel approach for the improvement of ethanol fermentation by Saccharomyces cerevisiae

Canadian Journal of Microbiology ◽

10.1139/w10-032 ◽

2010 ◽

Vol 56 (6) ◽

pp. 495-500 ◽

Cited By ~ 5

Author(s):

Lihua Hou ◽

Xiaohong Cao ◽

Chunling Wang

Keyword(s):

Saccharomyces Cerevisiae ◽

Ethanol Production ◽

Genome Engineering ◽

Ethanol Conversion ◽

Mitotic Chromosomes ◽

Gene Encoding ◽

Very High Gravity ◽

Novel Approach ◽

Very High Gravity Fermentation ◽

Tetraploid Strain

Fermentation properties under the control of multiple genes are difficult to alter with traditional methods in Saccharomyces cerevisiae . Here, a novel genome engineering approach is developed to improve ethanol production in very high gravity fermentation with 300 g/L glucose as the carbon source. This strategy involved constructing aneuploid strains on the base of tetraploid cells. The tetraploid strain was constructed by using the plasmid YCplac33-GHK, which harbored the HO gene encoding the site-specific Ho endonucleases. The aneuploid strain, WT4-M, was selected and screened after the tetraploid cells were treated with methyl benzimidazole-2-yl-carbamate to induce loss of mitotic chromosomes. It was found that aneuploid strain WT4-M not only exhibited an increase in ethanol production and osmotic and thermal tolerance, but also an improvement in the sugar–ethanol conversion rate. Notably, WT4-M provided up to 9.8% improvement in ethanol production compared with the control strain. The results demonstrated that the strategy of aneuploidy was valuable for creating yeast strains with better fermentation characteristics.

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Microbial Synergy via an Ethanol-Triggered Pathway

Molecular and Cellular Biology ◽

10.1128/mcb.24.9.3874-3884.2004 ◽

2004 ◽

Vol 24 (9) ◽

pp. 3874-3884 ◽

Cited By ~ 81

Author(s):

Michael G. Smith ◽

Shelley G. Des Etages ◽

Michael Snyder

Keyword(s):

Saccharomyces Cerevisiae ◽

Caenorhabditis Elegans ◽

Dimethyl Sulfoxide ◽

Cell Physiology ◽

Low Doses ◽

Microbial Interaction ◽

Signaling Molecule ◽

Bacterial Physiology ◽

Diffusible Factor ◽

Natural Isolates

ABSTRACT We have discovered a microbial interaction between yeast, bacteria, and nematodes. Upon coculturing, Saccharomyces cerevisiae stimulated the growth of several species of Acinetobacter, including, A. baumannii, A. haemolyticus, A. johnsonii, and A. radioresistens, as well as several natural isolates of Acinetobacter. This enhanced growth was due to a diffusible factor that was shown to be ethanol by chemical assays and evaluation of strains lacking ADH1, ADH3, and ADH5, as all three genes are involved in ethanol production by yeast. This effect is specific to ethanol: methanol, butanol, and dimethyl sulfoxide were unable to stimulate growth to any appreciable level. Low doses of ethanol not only stimulated growth to a higher cell density but also served as a signaling molecule: in the presence of ethanol, Acinetobacter species were able to withstand the toxic effects of salt, indicating that ethanol alters cell physiology. Furthermore, ethanol-fed A. baumannii displayed increased pathogenicity when confronted with a predator, Caenorhabditis elegans. Our results are consistent with the concept that ethanol can serve as a signaling molecule which can affect bacterial physiology and survival.

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