Specific isolates of lactic acid bacteria (LAB) can grow in the harsh beer environment, thus posing a threat to brew quality and the economic success of breweries worldwide. Plasmid-localized genes, such as horA, horC, and hitA, have been suggested to confer hop tolerance, a trait required for LAB survival in beer. The presence and expression of these genes among LAB, however, do not universally correlate with the ability to grow in beer. Genome sequencing of the virulent beer spoilage organism Lactobacillus brevis BSO 464 revealed the presence of eight plasmids, with plasmids 1, 2, and 3 containing horA, horC, and hitA, respectively. To investigate the roles that these and the other five plasmids play in L. brevis BSO 464 growth in beer, plasmid curing with novobiocin was used to derive 10 plasmid variants. Multiplex PCRs were utilized to determine the presence or absence of each plasmid, and how plasmid loss affected hop tolerance and growth in degassed (noncarbonated) beer was assessed. Loss of three of the eight plasmids was found to affect hop tolerance and growth in beer. Loss of plasmid 2 (horC and 28 other genes) had the most dramatic effect, with loss of plasmid 4 (120 genes) and plasmid 8 (47 genes) having significant, but smaller, impacts. These results support the contention that genes on mobile genetic elements are essential for bacterial growth in beer and that beer spoilage ability is not dependent solely on the three previously described hop tolerance genes or on the chromosome of a beer spoilage LAB isolate.
Beer is a very harsh environment for bacterial growth due to unique physiochemical attributes. Specifically, the presence of antimicrobial hop compounds (15 to 55 ppm iso-␣-acids) and ethanol (0.5 to 14% [vol/vol]) (1), coupled with the low availability of nutrients, a microaerophilic environment, low pH, and, finally, pressure in the packaged product due to CO 2 content, together mean that contaminating microorganisms must employ compensatory mechanisms to survive in the milieu. When successful microbial growth in beer does occur, the result is a spoiled product with unappealing off flavors and an altered sensory profile, ultimately leading to issues with brand confidence and economic loss for brewers globally (2).Isolated beer-spoiling organisms (BSOs) frequently belong to the genus Lactobacillus or Pediococcus, both of which are classified as lactic acid bacteria (LAB) (3, 4). The ability to spoil beer, however, is not intrinsic to all members of a given species in each genus and is instead isolate specific. This indicates that BSOs have undergone a degree of genetic specialization relative to non-BSO LAB isolates of the same genus and species. LAB are a diverse group of organisms, and a given species can be isolated from a number of environments. For example, Lactobacillus plantarum isolates can be found, not only in spoiled beer, but also in spoiled meats and the human intestinal microbiome (5). Due to the varied nature of these environments and the frequency with which specific bacteri...