Sarah Borg (1), Benjamin Junge (1); (1) BIOTECON Diagnostics, Potsdam, Germany
In general beer is a hostile environment for most microorganisms. Only a few kinds of bacteria are able to grow under such inhospitable conditions like the ethanol concentration, the relatively low pH and the dissolved carbon dioxide concentration and are able to spoil beer. The detection and identification of such beer-spoilage bacteria by conventional methods in a routine lab of a brewery is a very time-consuming and laborious task. This study attempts to verify that real-time PCR provides an easy, fast, and reliable alternative testing solution. For the existing food-proof Beer Screening Kit BIOTECON Diagnostics uses hybridization probes (LightCycler technology). This kit allows the detection of 30 beer-spoilage bacteria, including identification of 12 species, in a single reaction within 24-48 hr. The range of detected organisms includes the most beer-relevant species of the genera Lactobacillus, Pediococcus, Pectinatus and Megasphaera. The method is adjusted to the routine lab, permitting a throughput of up to 94 samples per PCR run. With this kit real-time PCR is performed on a LightCycler instrument from Roche and uses FRET (fluorescence resonance energy transfer) to detect the DNA. It will be demonstrated that after a PCR run, the absence or presence of beer spoilers can be detected immediately. Subsequent melting curve analysis allows the identification of the most important spoilage bacteria like Lactobacillus brevis, L. lindneri or Pediococcus inopinatus from a positive result without any further testing. Differences such as length, G-C-content, and base sequence, make the signal obtained by melting curve investigation distinct for nearly every probe-DNA combination. Within the target organisms another beer-spoiling bacteria, Lactobacillus acetotolerans, will be included as number 31. A second food-proof Beer Screening Kit for almost all the other PCR instruments is currently being developed. The kit is based on 5′ nuclease technology (TaqMan). It also allows the detection of the above mentioned 31 different beer-spoilage bacteria in one test. In addition it will be shown that the detection of hop tolerance genes in a separate fluorescence channel is possible. Lactobacillus brevis, the most important bacterial beer spoiler, can be identified in a third fluorescence channel. Beer-spoilage organisms also include several so-called wild yeasts, of which Saccharomyces species are generally considered the most important. Wild yeast can be defined as any yeast that a brewer did not intentionally introduce into a beer. Similar to beer-spoiling bacteria, wild yeasts can produce a wide variety of undesired flavors in finished beer. To detect the most important spoilage yeasts, BIOTECON Diagnostics is currently developing a real-time PCR kit that detects the genera Saccharomyces spp., Zygosaccharomyces spp. and Dekkera/Brettanomyces spp. in one single test. Both new kits are planned to be launched by the end of 2016. Validation data for all three methods, including sensitivity, species inclusivity and exclusivity, accuracy, matrix compatibility and others will be demonstrated in the presentation.
Sarah Borg is key account manager for North America at BIOTECON Diagnostics. She graduated from the Technical University of Braunschweig in biotechnology and wrote her master’s thesis at the Institute of Chemistry at the University of Waterloo in Canada under Prof. John Honek. She performed her Ph.D. thesis at the Ludwig-Maximilian University of Munich. The topic of her Ph.D. was the functionalization of bacterial nanoparticles. At BIOTECON Diagnostics she specializes in the craft beer market and the detection of beer spoilers.