Quantitative real-time PCR analysis of putative beer-spoilage associated genes in Pediococcus claussenii and Lactobacillus brevis

Microbiology Session
Jordyn Bergsveinson, University of Saskatchewan, Saskatoon, Saskatchewan, Canada
Co-author(s): Vanessa Pittet and Barry Ziola, University of Saskatchewan, Saskatoon, SK, Canada

ABSTRACT: Although the unique chemical and physical composition of beer provides an incredibly inhospitable environment for bacterial growth, lactic acid bacteria (LAB) frequently survive in and spoil beer. The presence of contaminating bacteria therefore poses a great threat to the brew quality and economic success of a brewery. Pediococcus and Lactobacillus are common genera of LAB isolated from spoiled beer; however, not all isolates of a given species in either genus can grow in beer. This indicates there is genetic specialization in beer-spoiling organisms such as Pediococcus claussenii ATCC BAA-344T (Pc344NR; non-ropy) and Lactobacillus brevis BSO 464 (BSO 464), both of which are capable of flourishing in a beer environment. Despite the evident genetic adaptations of beer-spoilage LAB, very few genes have been found to correlate with the beer-spoiling ability of an organism. To investigate the role of several putative beer-spoilage related genes within each organism in relation to growth in beer, quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) was utilized. RNA was extracted from both Pc344NR and BSO 464 cultures at mid-logarithmic growth in both beer and non-beer environments and then converted to cDNA and subjected to qRT-PCR analysis. In order to accurately assess differential gene expression in the beer environment, appropriate internal control genes are needed to normalize the experimental setup for sample variation and experimental error. As such, expression stability (in both growth conditions) of 12 candidate normalization genes was performed and analyzed with the application geNorm. Subsequently, cDNA samples were analyzed to determine the expression levels of the putative beer-spoilage related genes hitA, horA, horB, horC, and bsrA. Using this methodology, we now have a better understanding of the role these genes play during growth in beer for our two LAB isolates. It is anticipated that this work will be extended to include qRT-PCR analysis of possible beer-spoilage related genes in a broad range of LAB with the intent to definitively identify genes that can serve as genetic markers for assessing bacterial beer-spoilage potential.

Jordyn Bergsveinson received a B.S. degree (with honors) in microbiology and immunology in 2011 from the University of Saskatchewan. She is currently working toward completing an M.S. degree in health sciences through the College of Medicine at the University of Saskatchewan, under the supervision of Barry Ziola. Her research topic and area of interest concerns the genetic and molecular analysis of beer-spoiling microorganisms.