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.

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.

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.

Uncertainty in Widmark calculations: ABV variation in packaged versions of the most popular beers in the UK.

Forensic practitioners regularly use the Widmark equation to determine theoretical blood alcohol concentrations for use in cases involving alcohol. It is important with these calculations to determine the uncertainty associated with any result. Previous work has investigated the uncertainty in percent alcohol by volume (%ABV) from beers produced by small independent breweries in the UK but did not study the top selling beers in the UK. The top selling lagers and ales/bitters in the UK were identified by sales volume and the %ABV determined. These data was then used to determine the percent coefficient of variation (%CV) that should be used by forensic practitioners when constructing alcohol technical defence reports for use in forensic cases. These samples, from what may be described as 'big' brewers, were determined to have a smaller root mean square error (RMSE) (±0.1%v/v, n = 35), and %CV than those previously reported for beers produced by small, independent breweries in the UK. The results from this study shows that different RMSE's should be used for %ABV when determining the uncertainty of results from Widmark calculations depending if the drinks consumed have been from either 'big' brewers or small, independent breweries.

The influence of alcohol content variation in UK packaged beers on the uncertainty of calculations using the Widmark equation.

It is common for forensic practitioners to calculate an individual's likely blood alcohol concentration following the consumption of alcoholic beverage(s) for legal purposes, such as in driving under the influence (DUI) cases. It is important in these cases to be able to give the uncertainty of measurement on any calculated result, for this reason uncertainty data for the variables used for any calculation are required. In order to determine the uncertainty associated with the alcohol concentration of beer in the UK the alcohol concentration (%v/v) of 218 packaged beers (112 with an alcohol concentration of ≤5.5%v/v and 106 with an alcohol concentration of >5.5%v/v) were tested using an industry standard near infra-red (NIR) analyser. The range of labelled beer alcohol by volume (ABV's) tested was 3.4%v/v - 14%v/v. The beers were obtained from a range of outlets throughout the UK over a period of 12 months. The root mean square error (RMSE) was found to be ±0.43%v/v (beers with declared %ABV of ≤5.5%v/v) and ±0.53%v/v (beers with declared %ABV of >5.5%v/v) the RMSE for all beers was ±0.48%v/v. The standard deviation from the declared %ABV is larger than those previously utilised for uncertainty calculations and illustrates the importance of appropriate experimental data for use in the determination of uncertainty in forensic calculations.

Improving uncertainty in Widmark equation calculations: Alcohol volume, strength and density.

The Widmark equation is probably the most commonly used calculation for medicolegal purposes. Recently the National Research Council (USA) and the Forensic Science Regulator (UK) have called for the uncertainty of all results to be given with all forensic measurements and calculations. To improve the uncertainty of measurement of results from Widmark calculations we have concentrated on the uncertainties of measurement involved in the calculation of amount of alcohol, that of the volume of alcohol, the concentration of alcohol and the density of alcohol as previous studies have investigated some of the other factors involved. Using experimental studies, the scientific literature and legal statutes, we have determined revised and improved uncertainties of the concentration of ethanol for Widmark calculations for both the USA and UK. Based on the calculations that we have performed we recommend the use of Monte Carlo Simulation for the determination of uncertainty of measurement for Widmark Calculations.

Chronological and replicative lifespan of polyploid Saccharomyces cerevisiae (syn. S. pastorianus).

Chronological lifespan may be defined as the result of accumulation of irreversible damage to intracellular components during extended stationary phase, compromising cellular integrity and leading to death and autolysis. In contrast, replicative lifespan relates to the number of divisions an individual cell has undertaken before entering a non-replicative state termed senescence, leading to cell death and autolysis. Both forms of lifespan have been considered to represent models of ageing in higher eukaryotes, yet the relation between chronologically and replicatively aged populations has not been investigated. In this study both forms of lifespan have been investigated in Saccharomyces cerevisiae (Syn. S. pastorianus) to establish the relationship between chronological and replicative ageing.