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16. The lifestyle of beer-spoiling lactic acid bacteria is based on digging in the plasmid pool

Rudi F. Vogel (1), Jürgen Behr (1), Andreas Geißler (1); (1) Technische Universität München, Freising, Germany

Technical Session 5: Lactic Acid Bacteria
Sunday, August 14  •  2:00–3:15 p.m.
Plaza Building, Concourse Level, Governor’s Square 15

Lactic acid bacteria (LAB) are considered the most important and hazardous spoilage microorganisms in breweries. Typically, one species of LAB contains non-spoiling and also beer-spoiling strains. Their ability to grow in beer is often connected to the presence of plasmid-encoded lifestyle genes like horA or horC. These hop tolerance genes not only contribute to improved growth and adaption to beer, but also represent important species-independent diagnostic marker genes (DMGs) for the identification of beer spoiling LAB. However, previous genome sequencing projects with brewery-isolated LAB have revealed the prevalence of high numbers of plasmids, while consecutive experiments could demonstrate that plasmids lacking known hop resistance genes also support LAB adaption to and growth in beer. We sequenced 20 and analyzed the genomes of 22 strains with varying beer spoilage ability, comprising the important beer spoiling species Lactobacillus brevis, L. lindneri, L. backii, L. paracollinoides, Pediococcus claussenii and P. damnosus. The investigated genomes are characterized by up to 10 plasmids, and comparative analysis revealed the presence of a shared, plasmid-encoded genetic pool. Of 3,209 brewery plasmid-encoded genes, 589 were found to be present in at least two genomes, while 59% of these genes were found within two or more species. This is not only due to the presence of identical plasmids (25%), but also a consequence of shared pieces of homologous DNA integrated into different plasmids. Besides hop tolerance, these genes encode various functions, including carbohydrate metabolism, cell envelope modification or ion homeostasis. While some genes were found to be restricted to one or two species, others are spread within all six species, suggesting that these bacteria only take/keep what they need to survive and grow in beer and do not accumulate “brewery DNA” in an arbitrary way. As an example, we found a plasmid-encoded complete fatty acid biosynthesis (FAS) cluster shared exclusively by beer-spoiling strains of P. damnosus and L. backii, while both species lack a chromosomal FAS. In P. damnosus, the ability to produce acetoin and butandiol from pyruvate is plasmid mediated, leading to the subsequent formation of the known off-flavor diacetyl. This is advantageous for their growth in beer, as it enables the cell to avoid acid stress by formation of non-acidic end products instead of lactate. Apparently, brewery LAB dig in the plasmid pool and keep what they need, even in inter-species transfers. While the adopted abilities contribute to their lifestyle in this habitat, they also allow us the deduction of novel DMGs for the identification of beer-spoiling strains. FabZ, part of the abovementioned FAS cluster, allows a 100% correct discrimination of beer spoiling and non-spoiling strains of P. damnosus, which was validated for 20 characterized strains using PCR.

Rudi F. Vogel is a biochemist interested in food microbiology and biotechnology. As head of Technische Mikrobiologie at the Technische Universität München, Germany, he conducts research on starter culture development, high pressure in food and biosciences, as well as control of unwanted microbes in food. A clear focus is on lactic acid bacteria, their metabolism and genetics, mechanisms of stress response and adaptation, as well as interaction with other microbes. In this context beer spoiling bacteria are a model for the study of bacterial stress response, adaptation, and genome plasticity.