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​35. Pediococcus claussenii genetic expression during growth in beer assessed by transcriptome sequencing (RNA-seq)

Vanessa Pittet, University of Saskatchewan, Saskatoon, Saskatchewan, Canada

 

ABSTRACT: The beer environment is very inhospitable for microbes as it typically contains high levels of CO₂, hops, and ethanol and has low pH, oxygen, and available nutrients. As such, most microbial growth is inhibited in beer. However, specific organisms have adapted to overcome the stresses found in beer and, therefore, can grow in and spoil the product. The most common beer-spoilage bacteria are lactobacilli and pediococci, and their presence in a brewery can have a major economic impact due to product spoilage. To elucidate the mechanisms that these organisms use to grow in a beer environment, we sequenced the genome of the brewery isolate Pediococcus claussenii ATCC BAA-344T. We found that this organism has the means for genetic diversity, particularly via the eight plasmids that are present. To gain a better understanding of the role that various genes play, we performed whole transcriptome sequencing (RNA-seq) of P. claussenii during growth in beer and a non-beer medium (MRS). RNA was extracted during mid-logarithmic growth from both MRS broth and a Canadian-brewed beer (5% [v/v] ethanol, pH 4.2, and roughly 11 BU). Transcriptome sequencing was done via illumina technology, and bioinformatic analyses were performed to determine gene expression in both growth conditions. We found that the majority of genes are expressed in both environments, indicating that most of the coding capacity of this isolate is used (i.e., at least basally expressed). Roughly half of the genes did not show differential expression in either medium. However, of those showing significant differential expression, half of the genes had increased expression during growth in beer, while half showed decreased expression. Several operons were very highly expressed during growth in beer but not during growth in MRS, suggesting that these genes play a major role in the ability of P. claussenii to grow in beer. As expected, the previously described hop-resistance gene horA had increased expression during growth in beer. Interestingly, a number of the genes with increased expression in beer played a role in nutrient acquisition and hop resistance, leading to the conclusion that the other stress factors found in beer (e.g., ethanol) may not play a large role in determining if growth of the isolate will be inhibited in beer or not. This is most likely due to the intrinsic resistance that most lactic acid bacteria have to a range of stressors, including ethanol and low pH. This study provides a better understanding of the genetic mechanisms that bacteria can use to overcome the inhospitable environment of beer. This information can be used as the basis for further studies into potential targets for detection of beer-spoilage bacteria in a brewery setting.