Dear brew enthusiasts,
In this episode of the Beerologist, we will talk about flocculation in yeast. More specifically, we will describe some work in lager yeasts that helped identify new genes with roles in flocculation. But before we start, let's briefly discuss flocculation.
Flocculation is when yeast cells clump together and settle out of the solution (beer). Yeast clumps are denser than beer, prompting their gravity-assisted drop to the bottom. While many people think that flocculation is a process that follows fermentation, this process is biologically meaningful. In nature, yeasts, like most other microbes, must cope with challenging environmental factors. Flocculation is one mechanism triggered by cellular stress that increases yeast survivability. The fact that flocculation is a biological and physiologically relevant process means that yeast must control and, by extension, is genetically hardwired to do so. From this, it follows that geneticists with interest in yeast flocculation should find these genetic components.
Why flocculation matters in brewing
Flocculation is one of the major factors that enable (commercial) brewers to separate beer from their yeast slurry and has enormous value. On the flip side, however, premature or delayed flocculation can negatively impact beer quality. Premature flocculation will inhibit fermentation at the final stages and leave fermentable sugars in a beer. Delayed flocculation can complicate processing after fermentation and the use of yeast slurries for additional fermentations (high yeast counts, yeast vitality etc.). Because of these reasons, brewers must use strains that have appropriate levels of flocculation at the right fermentation stage. Understanding the factors that determine flocculation may help us control that process.
Identification of genes that contribute to flocculation
There are various ways by which researchers can identify genes or mutations that underpin a phenotype. In a study by Zhou and authors (2021), researchers identified and compared the genomes of two lager yeast strains (G03-10 and G03-24) to their parental strain G03. G03-10 and G03-24 were isolated from yeast slurries obtained after serial pitching (starting with G03) in a commercial brewery. Both G03-10 and G03-24 sediment or flocculate more quickly than the parental strain, suggesting that a mutation(s) and selection during successive fermentations have changed phenotypes.
Figure 1c from Zhou et al. (2021). Flocculation ability of G03, G03-10, and G03-24 strains, assessed throughout the fermentation process.
The authors tested this hypothesis by sequencing the three strains and perform genome-wide comparisons between strains. In doing so, they were able to divide mutated genes into the following categories:
No change between parent, G03-10 and G03-24.
Mutation in either G03-10 or G03-24 when compared to the parent (G03)
Genes that are mutated in both G03-10 and G03-24 when compared with the parent.
Because both G03-10 and G03-24 have increased flocculation rates, the authors hypothesized that category Nr. 3 genes (mutated in both G03-10 and G03-24) impact flocculation. Thus, they decided to look at the genes that fall into that class, assign functions, and ask whether any functions link to flocculation.
You can see from the results (Figure 4c) that when you look at the gene classifications, many of the proteins have roles in stress adaptation. Protein processing in the endoplasmic reticulum, membrane lipid metabolism and other extensive stress responses, some of which are well-characterized in yeast. From the results, it thus appears that by following the evolution of beer yeasts throughout successive fermentations, we can learn about the biology of yeast relevant to beer production.
Finally and importantly, we always need to interpret results obtained with wholesale comparative analyses carefully. It remains possible that other selective forces acting on both strains have caused maintenance of common mutations. In this context, it is thus critical to test whether the genes identified impact flocculation. Validation is what the authors did to verify their analyses and results.
Figure 5a from Zhou et al (2021). Flocculation analyses of G03 strains, mutated in individual candidate genes. RIM101 and VPS36 were both found to have a measurable impact on flocculation in pilot fermentation experiments.
This analysis led to the results any biologist would expect to see. Disruption of some genes had a measurable impact on flocculation (e.g. RIM101 and VPS36), whereas others didn't.
Why does this matter?
The authors were able to show that by using a whole-genome approach, they were able to quickly define a set of candidate genes with roles in flocculation. This study highlights both the power and the limitation of the approach - we can quickly hone in on a smaller subset of genes. However, we must still characterize each gene separately to establish whether it is a genuine factor in flocculation.
Excitingly, we are only beginning to explore yeast adaptation in the brewery. This work is yet another proof of concept that is extendable to other yeasts, phenotypes or even processes.
If you are a brewer noticing a distinct (positive or negative) change in performance, it certainly pays to contact and collaborate with geneticists that can tease out underlying genetic causes!
I hope you enjoyed this read and have a great weekend!
Cheers!
Edgar, the Beerologist.