The first step in the infection of fungal pathogens in humans is the adhesion of the pathogen to host tissue cells or abiotic surfaces such as catheters and implants. One of the main players involved in this are the expressed cell wall adhesins. Here, we review the Flo adhesin family and their involvement in the adhesion of these yeasts during human infections. Firstly, we redefined the Flo adhesin family based on the domain architectures that are present in the Flo adhesins and their functions, and set up a new classification of Flo adhesins. Next, the structure, function, and adhesion mechanisms of the Flo adhesins whose structure has been solved are discussed in detail. Finally, we identified from Pfam database datamining yeasts that could express Flo adhesins and are encountered in human infections and their adhesin architectures. These yeasts are discussed in relation to their adhesion characteristics and involvement in infections.
Flocculation or cell aggregation is a well-appreciated characteristic of industrial brewer’s strains, since it allows removal of the cells from the beer in a cost-efficient and environmentally-friendly manner. However, many industrial strains are non-flocculent and genetic interference to increase the flocculation characteristics are not appreciated by the consumers. We applied adaptive laboratory evolution (ALE) to three non-flocculent, industrial Saccharomyces cerevisiae brewer’s strains using small continuous bioreactors (ministats) to obtain an aggregative phenotype, i.e., the “snowflake” phenotype. These aggregates could increase yeast sedimentation considerably. We evaluated the performance of these evolved strains and their produced flavor during lab scale beer fermentations. The small aggregates did not result in a premature sedimentation during the fermentation and did not result in major flavor changes of the produced beer. These results show that ALE could be used to increase the sedimentation behavior of non-flocculent brewer’s strains.
Lager beer fermentations rely on specific polyploid hybrids between Saccharomyces cerevisiae and Saccharomyces eubayanus falling into the two groups of S. carlsbergensis/Saaz-type and S. pastorianus/Frohberg-type. These strains provide a terroir to lager beer as they have long traditional associations and local selection histories with specific breweries. Lager yeasts share, based on their common origin, several phenotypes. One of them is low transformability, hampering the gene function analyses required for proof-of-concept strain improvements. PCR-based gene targeting is a standard tool for manipulating S. cerevisiae and other ascomycetes. However, low transformability paired with the low efficiency of homologous recombination practically disable targeted gene function analyses in lager yeast strains. For genetic manipulations in lager yeasts, we employed a yeast transformation protocol based on lithium-acetate/PEG incubation combined with electroporation. We first introduced freely replicating CEN/ARS plasmids carrying ScRAD51 driven by a strong heterologous promoter into lager yeast. RAD51 overexpression in the Weihenstephan 34/70 lager yeast was necessary and sufficient in our hands for gene targeting using short-flanking homology regions of 50 bp added to a selection marker by PCR. We successfully targeted two independent loci, ScADE2/YOR128C and ScHSP104/YLL026W, and confirmed correct integration by diagnostic PCR. With these modifications, genetic alterations of lager yeasts can be achieved efficiently and the RAD51-containing episomal plasmid can be removed after successful strain construction.
Non-Saccharomyces species have been recognized for their beneficial contribution to fermented food and beverages based on their volatile compound formation and their ability to ferment glucose into ethanol. At the end of fermentation brewer’s yeast flocculate which provides an easy means of separation of yeasts from green beer. Flocculation in Saccharomyces cerevisiae requires a set of flocculation genes. These FLO-genes, FLO1, FLO5, FLO9, FLO10, and FLO11, are located at telomeres and transcription of these adhesins is regulated by Flo8 and Mss11. Here, we show that Saccharomycopsis fermentans, an ascomycete yeast distantly related to S. cerevisiae, possesses a very large FLO/ALS-like Sequence (FAS) family encompassing 34 genes. Fas proteins are variable in size and divergent in sequence and show similarity to the Flo1/5/9 family. Fas proteins show the general build with a signal peptide, an N-terminal carbohydrate binding PA14 domain, a central region differing by the number of repeats and a C-terminus with a consensus sequence for GPI-anchor attachment. Like FLO genes in S. cerevisiae, FAS genes are mostly telomeric with several paralogs at each telomere. We term such genes that share evolutionary conserved telomere localization “telologs” and provide several other examples. Adhesin expression in S. cerevisiae and filamentation in Candida albicans is regulated by Flo8 and Mss11. In Saccharomycopsis we identified only a single protein with similarity to Flo8 based on sequence similarity and the presence of a LisH domain.
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