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In 2010, bottles of beer containing viable bacteria of the common beer-spoilage species Lactobacillus backii and Pediococcus damnosus were recovered from a shipwreck near the Åland Islands, Finland. The 170-year quiescent state maintained by the shipwreck bacteria presented a unique opportunity to study lactic acid bacteria (LAB) evolution vis-a-vis growth and survival in the beer environment. Three shipwreck bacteria (one L. backii strain and two P. damnosus strains) and modern-day beer-spoilage isolates of the same two species were genome sequenced, characterized for hop iso-α-acid tolerance, and growth in degassed lager and wheat beer. In addition, plasmid variants of the modern-day P. damnosus strain were analyzed for the effect of plasmid-encoded genes on growth in lager beer. Coding content on two plasmids was identified as essential for LAB growth in modern lager beer. Three chromosomal regions containing genes related to sugar transport and cell wall polysaccharides were shared by pediococci able to grow in beer. Our results show that the three shipwreck bacteria lack the necessary plasmid-located genetic content to grow in modern lager beer, but carry additional genes related to acid tolerance and biofilm formation compared to their modern counterparts.

The food industry, including the meat industry, is currently looking for natural preservatives to prevent the growth of harmful microbes in foods. The potential of plant-derived antimicrobial extracts to increase the shelf life and to delay the microbiological spoilage of marinated broiler chicken cuts in modified atmosphere packages during cold storage was investigated in this study. We evaluated the impact of aqueous ethanolic extracts of Finnish sea buckthorn berries and lingonberries and supercritical CO2-extracted herbal extracts from an antimicrobial blend and oregano leaves on the shelf life of broiler meat. The commercial antimicrobial blend extract and the oregano extract inhibited the growth of lactic acid bacteria (LAB) and Brochothrix thermosphacta in the marinated samples. The antimicrobial blend extract also reduced the growth of psychrotrophic aerobic bacteria, whereas the sea buckthorn and lingonberry extracts did not. Only minor antimicrobial activity against Enterobacteriaceae by all the extracts was observed. Plate count analysis, denaturing gradient gel electrophoresis, and quantitative real-time PCR indicated that LAB, which are the major spoilage group in marinated modified atmosphere–packaged poultry products, were not significantly affected by the berry extracts studied. During this shelf-life study, LAB isolates of Lactobacillus and Leuconostoc were identified in the marinated samples. Antimicrobial blends and oregano leaf extracts can act as antimicrobial agents in marinade blends, although tailoring of the dose is needed because of their strong taste. Further studies for exploiting synergistic effects of plant extracts could contribute to the development of potential and more effective antimicrobial blends. Studies are needed in meat matrices and in product applications to demonstrate the efficacy of these compounds.

The aim of this study was to find out whether the first bacterial colonizers of brewery process surfaces after washing produce acyl homoserine lactone (AHL) quorum sensing signalling molecules. Microbial attachment and biofilm formation was studied by mounting sterile uncoated and coated stainless steel coupons onto critical places of the filling machines in three breweries. In the present study 26 previously deposited bacterial strains, as well as approximately 2,300 colonies from 76 process surface samples, were screened using reporter bacteria Chromobacterium violaceum CV026 and Agrobacterium tumefaciens NTLR4. AHL‐producing bacteria were detected both from process samples and among the previous isolates. From the process samples, up to 15% of the screened colonies produced AHL molecules. Production of long chain AHLs was more common than short chain AHLs. The identified AHL‐producing isolates belonged to the genera Pseudomonas, Serratia, Hafnia, Rahnella, Enterobacter and Aeromonas, which all belong to commonly found primary colonizers of brewery process surfaces. The same microbial genera producing AHL molecules were found from different breweries. Brewery filling equipment is susceptible to microbial attachment and accumulation on surfaces. In the future, inhibition of quorum sensing could be one additional way of controlling biofilm formation.

Beer is generally a hostile environment for most microorganisms rendering it safe in terms of food-borne illnesses. However, species adapted to the brewery environment can cause product spoilage. In this chapter, microorganisms associated with brewing and the production of beer as well as with beer dispensing are briefly described. Keeping the beer free from contaminating organisms is important for image reasons and, additionally, production plant hygiene has both economic and environmental motivation. In the effort of improving brewery hygiene, studies regarding brewery biofilms have lately been carried out. This chapter aims at reviewing the current knowledge of biofilms occurring in the brewery. It also summarises the means available for prevention of biofilm formation with special emphasis on emerging technologies and future trends in biofilm combat and their application in breweries.

Publisher Summary The presence of inhibitors, such as hop compounds, alcohol, carbon dioxide, and sulfur dioxide, as well as the shortage of nutrients and oxygen and the low pH make beer resistant to microbial contamination. Moreover, processes such as filtration, storage at low temperatures and possible pasteurization reduce contamination. The special environment in the brewing process restricts the range of microorganisms likely to be encountered to relatively few species. Although the contaminants found may cause quality defects, pathogens have not to knowledge been reported to grow in standard beer products. Ensuring the well-being of the production, yeast strains is a fundamental part of brewing as in all processes based on fermentation technology. Thus, monitoring yeast quality and quantity is also an important part of the microbiological control carried out in breweries. This chapter presents a summary of the microorganisms likely to be encountered in breweries and the different possibilities to detect and identify them. It also describes the different possibilities to quantify yeast mass and estimate the brewing performance as well as differentiate between yeast strains and discusses both methods currently in use and emerging technologies. The chapter reviews those methods that show most potential for brewery applications till date.

Contamination of brewing process by spoilage organisms may lead to production problems and quality defects in final product. Present quality control methods based on cultivation reveal the possible contamination only after several days or weeks of delay. Moreover, they do not discriminate between spoilage and non-spoilage microbes nor allow the detection of fastidious or injured organisms. In recent years, several promising molecular biological applications have been described for the detection, characterization and identification of brewery contaminants. Group, genus and species specific PCR tests have been developed for contaminants present in brewing process, i.e. for lactic acid bacteria (Lactobacillus, Pediococcus), strictly anaerobic bacteria (Pectinatus, Megasphaera, Selenomonas, Zymophilus), enterobacteria, acetic acid bacteria and wild yeasts. Various PCR detection formats (PCR-ELISA, LightCyclerTM PCR, standard PCR) have been set-up for most of the beer spoilage organisms. Pre-PCR treatment methods have been developed for filterable samples, such as bright beer and process water, and for unfilterable samples, such as pitching yeast, fermenting wort and unfilterable beer. Due to the very low amounts of microbes in brewing samples, they need to be enriched in a suitable medium prior to the PCR. Practical DNA extraction methods and removal of PCR inhibitors have been developed for different sample types. For example, a commercial system containing magnetic particles for DNA collection and purification can be used for this purpose. Modular PCR kits for standard PCR and real-time PCR are available from several companies. The developed PCR applications are, despite of the enrichment step, more rapid and specific than conventional methods. Without prior enrichment, sensitivity of PCR methods is lower and they are at the moment expensive to use compared to cultivation methods. An alternative to PCR, a commercial fluorescent in situ hydridization (FISH) method has been developed for beer-spoilage bacteria. It is based on the use of fluorescently labelled gene probes and epifluorescence microscope. The automated ribotyping system (RiboPrinter®System DuPont Qualicon,USA) is rapid molecular biological method for identification and characterization of bacteria to species or even below species level. The comprehensive identification database using three different restriction enzymes has been created to the brewery contaminants including Lactobacillus spp., Pediococcus spp, Pectinatus spp., Megasphaera cerevisiae and Obesumbacterium proteus. In addition to identification purposes and renaming of bacteria, the database has been applied in tracing contamination sources and in detection of troublesome "house flora" in industrial processes.

Immobilized primary fermentation bioreactors were deliberately contaminated with an enterobacterium, Klebsiella terrigena, and changes were monitored in yeast performance and viability and in chemical composition of the green beers. In addition, the formation of biogenic amines, possible carcinogenic substances, was monitored. K. terrigena caused elevated concentrations of dimethyl sulfide in beer but did not cause significant changes in the concentrations of the analyzed higher alcohols or esters. On the other hand, the ratio between diacetyl and pentanedione was increased. The yeast viability was lowered without observed changes in fermentation capacity. The increased concentration of biogenic amines was mainly due to changes in the cadaverine concentration.