Wendy G. Box

Carbohydrate utilization and the lager yeast transcriptome during brewery fermentation

The fermentable carbohydrate composition of wort and the manner in which it is utilized by yeast during brewery fermentation have a direct influence on fermentation efficiency and quality of the final product. In this study the response of a brewing yeast strain to changes in wort fermentable carbohydrate concentration and composition during full‐scale (3275 hl) brewery fermentation was investigated by measuring transcriptome changes with the aid of oligonucleotide‐based DNA arrays. Up to 74% of the detectable genes showed a significant (p⩽0.01) differential expression pattern during fermentation and the majority of these genes showed transient or prolonged peaks in expression following the exhaustion of the monosaccharides from the wort. Transcriptional activity of many genes was consistent with their known responses to glucose de/repression under laboratory conditions, despite the presence of di‐ and trisaccharide sugars in the wort. In a number of cases the transcriptional response of genes was not consistent with their known responses to glucose, suggesting a degree of complexity during brewery fermentation which cannot be replicated in small‐scale wort fermentations or in laboratory experiments involving defined media. Copyright

The oxidative stress response of a lager brewing yeast strain during industrial propagation and fermentation

Commercial brewing yeast strains are exposed to a number of potential stresses including oxidative stress. The aim of this investigation was to measure the physiological and transcriptional changes of yeast cells during full-scale industrial brewing processes with a view to determining the environmental factors influencing the cells oxidative stress response. Cellular antioxidant levels and genome-wide transcriptional changes were monitored throughout an industrial propagation and fermentation. The greatest increase in cellular antioxidants and transcription of antioxidant-encoding genes occurred as the rapidly fermentable sugars glucose and fructose were depleted from the growth medium (wort) and the cell population entered the stationary phase. The data suggest that, contrary to expectation, the oxidative stress response is not influenced by changes in the dissolved oxygen concentration of wort but is initiated as part of a general stress response to growth-limiting conditions, even in the absence of oxygen. A mechanism is proposed to explain the changes in antioxidant response observed in yeast during anaerobic fermentation. The available data suggest that the yeast cell does not experience oxidative stress during industrial brewery handling. This information may be taken into consideration when setting parameters for industrial brewery fermentation.

Evaluation of ITS PCR and RFLP for Differentiation and Identification of Brewing Yeast and Brewery ‘Wild’ Yeast Contaminants

ABSTRACT A reference library of ITS PCR/RFLP profiles was collated and augmented to evaluate its potential for routine identification of domestic brewing yeast and known ‘wild’ yeast contaminants associated with wort, beer and brewing processes. This library contains information on band sizes generated by restriction digestion of the ribosomal RNA‐encoding DNA (rDNA) internal transcribed spacer (ITS) region consisting of the 5.8 rRNA gene and two flanking regions (ITS1 and ITS2) with the endonucleases CfoI, HaeIII, HinfI and includes strains from 39 non‐Saccharomyces yeast species as well as for brewing and non‐brewing strains of Saccharomyces. The efficacy of the technique was assessed by isolation of 59 wild yeasts from industrial fermentation vessels and conditioning tanks and by matching their ITS amplicon sizes and RFLP profiles with those of the constructed library. Five separate, non‐introduced yeast taxa were putatively identified. These included Pichia species, which were associated with conditioning tanks and Saccharomyces species isolated from fermentation vessels. Strains of the lager yeast S. pastorianus could be reliably identified as belonging to either the Saaz or Frohberg hybrid group by restriction digestion of the ITS amplicon with the enzyme HaeIII. Frohberg group strains could be further sub‐grouped depending on restriction profiles generated with HinfI.

Amino Acid Uptake and Yeast Gene Transcription During Industrial Brewery Fermentation

Amino acid utilization by yeast during brewers wort fermentation influences both the fermentation performance and flavor profile of the finished product. To better understand the relationship between the yeast cell and wort amino acid composition, oligonucleotide microarrays were employed to measure the changes in transcription of genes associated with amino acid uptake and utilization during industrial-scale fermentation. Amino acid permeases with narrow specificity for amino acids had the lowest transcription values at the beginning of fermentation (when amino acids were replete), suggesting nitrogen catabolite repression, whereas relatively high transcription at the beginning of fermentation was confined to genes encoding permeases with broad substrate specificity. Nine genes involved in amino acid catabolism demonstrated significant changes in transcription, with most having the highest activity at 60 hr. Exceptions were PUT1, encoding proline oxidase, and CHA1, encoding L-serine deaminase, both with peak transcription at the beginning of fermentation. The only gene demonstrating increased activity in the final stages of fermentation was the threonine aldolase-encoding gene GLY1—a result that could explain the atypical low uptake of threonine. The majority of genes involved in amino acid biosynthesis had maximal expression at low amino acid concentrations, with notable exceptions being genes involved in central nitrogen metabolism and synthesis of glutamine (GLN1 and LYS9) and glutamate (GDH1)—a result that was consistent with the early depletion of glutamine from the wort and supported by the fact that these genes are regulated by nitrogen catabolite response-related transcription factors.

Does Generation Number Matter? The Impact of Repitching on Wort Utilization

Serial repitching is an accepted practice in brewing. The number of completed fermentations (generation number) is the metric applied to brewing yeast fitness for fermentation. Despite this, fermentations that use freshly propagated (generation 0) yeast are deemed to be inefficient when compared to yeast that has been repitched one or more times. The aim of this study was to investigate the impact of yeast generation number (equating to the number of fermentations completed) on wort utilization and fermentation progression. Wort composition and pH downshift were used to assess “fitness to ferment” of yeast generations within laboratory scale fermentations. Differences in pH, FAN, and amino acid assimilation were observed.