HYPHAEdelity: a quantitative image analysis tool for assessing peripheral whole colony filamentation.

The yeast Saccharomyces cerevisiae, also known as brewer's yeast, can undergo a reversible stress-responsive transition from individual ellipsoidal cells to chains of elongated cells in response to nitrogen- or carbon starvation. Whole colony morphology is frequently used to evaluate phenotypic switching response; however, quantifying two-dimensional top-down images requires each pixel to be characterized as belonging to the colony or background. While feasible for a small number of colonies, this labor-intensive assessment process is impracticable for larger datasets. The software tool HYPHAEdelity has been developed to semi-automate the assessment of two-dimensional whole colony images and quantify the magnitude of peripheral whole colony yeast filamentation using image analysis tools intrinsic to the OpenCV Python library. The software application functions by determining the total area of filamentous growth, referred to as the f-measure, by subtracting the area of the inner colony boundary from the outer-boundary area associated with hyphal projections. The HYPHAEdelity application was validated against automated and manually pixel-counted two-dimensional top-down images of S. cerevisiae colonies exhibiting varying degrees of filamentation. HYPHAEdelity's f-measure results were comparable to areas determined through a manual pixel enumeration method and found to be more accurate than other whole colony filamentation software solutions.

Disparity in pseudohyphal morphogenic switching response to the quorum sensing molecule 2-phenylethanol in commercial brewing strains of Saccharomyces cerevisiae.

Saccharomyces cerevisiae can undergo filamentous growth in response to specific environmental stressors, particularly nitrogen-limitation, whereby cells undergo pseudohyphal differentiation, a process where cells transition from a singular ellipsoidal appearance to multicellular filamentous chains from the incomplete scission of the mother-daughter cells. Previously, it was demonstrated that filamentous growth in S. cerevisiae is co-regulated by multiple signaling networks, including the glucose-sensing RAS/cAMP-PKA and SNF pathways, the nutrient-sensing TOR pathway, the filamentous growth MAPK pathway, and the Rim101 pathway, and can be induced by quorum-sensing aromatic alcohols, such as 2-phenylethanol. However, the prevalent research on the yeast-pseudohyphal transition and its induction by aromatic alcohols in S. cerevisiae has been primarily limited to the strain Σ1278b. Due to the prospective influence of quorum sensing on commercial fermentation, the native variation of yeast-to-filamentous phenotypic transition and its induction by 2-phenylethanol in commercial brewing strains was investigated. Image analysis software was exploited to enumerate the magnitude of whole colony filamentation in 16 commercial strains cultured on nitrogen-limiting SLAD medium; some supplemented with exogenous 2-phenylethanol. The results demonstrate that phenotypic switching is a generalized, highly varied response occurring only in select brewing strains. Nevertheless, strains exhibiting switching behavior altered their filamentation response to exogenous concentrations of 2-phenylethanol.

Influence of cell surface characteristics on adhesion of Saccharomyces cerevisiae to the biomaterial hydroxylapatite.

The influence of the physicochemical properties of biomaterials on microbial cell adhesion is well known, with the extent of adhesion depending on hydrophobicity, surface charge, specific functional groups and acid-base properties. Regarding yeasts, the effect of cell surfaces is often overlooked, despite the fact that generalisations may not be made between closely related strains. The current investigation compared adhesion of three industrially relevant strains of Saccharomyces cerevisiae (M-type, NCYC 1681 and ALY, strains used in production of Scotch whisky, ale and lager, respectively) to the biomaterial hydroxylapatite (HAP). Adhesion of the whisky yeast was greatest, followed by the ale strain, while adhesion of the lager strain was approximately 10-times less. According to microbial adhesion to solvents (MATS) analysis, the ale strain was hydrophobic while the whisky and lager strains were moderately hydrophilic. This contrasted with analyses of water contact angles where all strains were characterised as hydrophilic. All yeast strains were electron donating, with low electron accepting potential, as indicated by both surface energy and MATS analysis. Overall, there was a linear correlation between adhesion to HAP and the overall surface free energy of the yeasts. This is the first time that the relationship between yeast cell surface energy and adherence to a biomaterial has been described.

Characterization of Pot Ale from a Scottish Malt Whisky Distillery and Potential Applications.

Over 2.7 billion liters of pot ale is produced annually as a co-product of Scottish malt whisky, and apart from evaporation to pot ale syrup as a feed, it is primarily treated by anaerobic digestion or land/sea disposal. The aim of this study was to assess pot ale components and their potential applications. The insoluble solid fraction, mainly consisting of yeast, contained 55% protein, and as a protein feed ingredient, this could yield 32,400 tons of feed per annum, although the Cu content of this fraction would need to be monitored. The liquid fraction could yield 33,900 tons of protein per annum, and an SDS-PAGE profile of this fraction demonstrated that the proteins may be similar to those found in beer, which could extend their application as a food ingredient. This fraction also contained phosphorus, potassium, and polyphenols among other components, which could have added value. Overall, fractionation of pot ale could offer an alternative to evaporation to pot ale syrup while retaining the protein fraction in the food chain.

Bioconversion of brewer's spent grains to bioethanol.

Spent grains (SG), the residue remaining after extraction of wort, are a major by-product of brewing. This lignocelluose-rich biomass may provide a source of sugars for fuel ethanol fermentations. Dilute acid and enzyme treatments were developed to convert the hemicellulose and cellulose fractions to glucose, xylose and arabinose. Pretreatment of dried, milled grains with 0.16 N HNO(3) at 121 degrees C for 15 min was chosen as the most suitable method for solubilizing grains before enzymatic digestion with cellulase and hemicellulase preparations. Solids loading concentrations (10%, 15% and 20% w/v) were compared and reducing sugar between 40 and 48 g (100 g SG)(-1) was extracted. Hydrolysate, prepared from 20% SG, pretreated with 0.16 N HNO(3), partially neutralized to pH 5-6 and digested with enzymes for 18 h, contained 27 g L(-1) glucose, 16.7 g L(-1) xylose and 11.9 g L(-1) arabinose. Fermentation of this hydrolysate for 48 h by Pichia stipitis and Kluyveromyces marxianus resulted in 8.3 and 5.9 g L(-1) ethanol corresponding to ethanol conversion yields of 0.32 and 0.23 g ethanol (g substrate)(-1), respectively. Substrate utilization efficiency was less when compared with glucose/xylose mixtures in synthetic media, suggesting that yeast inhibitory compounds derived from SG were present in the hydrolysate.

Role of speciation in organotin toxicity to the yeast Candida maltosa.

Assessment of organometal pollution requires an understanding of the various processes that influence the bioavailability and toxicity of the contaminant. Organotins may exist as both cationic species and neutral hydroxides in aqueous solution, with the formation of chloride species in the presence of Cl-. Although these species have different chemical properties, there is very little information on the influence of speciation on organotin and microbial cell interactions. Tributyltin (TBT) and triphenyltin (TPT) interactions with the yeast Candida maltosa were investigated between pH 3.5 and 7.5 and in up to 0.5 M NaCl at pH 5.5. Toxicity increased with both pH and NaCl concentration and the mechanisms of interaction depended on the species present in solution. TBT and TPT interacted by different mechanisms, as evidenced by action on membrane fluidity. Furthermore, there was a strong correlation between toxicity and overall octanol-water distribution ratio (D(OW)) of organotin compounds. Triorganotin cations are less toxic than triorganotin hydroxides, which are in turn less toxic than triorganotin chlorides. These findings underline the importance of speciation effects on organotin interactions in the environment.