Effect of plasmid loss on the beer-spoiling phenotype of Pediococcus claussenii ATCC BAA-344T

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

ABSTRACT: Pediococcus claussenii is a species known to spoil beer. The P. claussenii ATCC BAA-344T (Pc344) genome was recently sequenced; however, many of the genes that permit this organism to proliferate amid the harsh conditions in beer have yet to be identified. It has been determined that Pc344 carries eight plasmids, six of which have coding capacity and two of which are cryptic. One of these plasmids has previously been shown to harbor horA, a well-established beer-spoilage associated gene involved in hops resistance. To further examine the role that plasmids may play in the ability of Pc344 to spoil beer, we obtained a collection of isolates with altered plasmid profiles and put these isolates through a series of phenotypic analyses designed to mimic the harsh conditions present in beer. Plasmid-minus Pc344 variants were generated either by repeatedly culturing in a non-beer medium or by incubating in a non-beer medium with a sub-lethal concentration of the antimicrobial compound novobiocin. Single colonies were selected by spreading onto MRS agar plates and then screened for an altered plasmid profile using multiplex polymerase chain reactions with primer sets designed to specific regions of each of the six plasmids with coding capacity. The plasmid-minus isolates obtained were then subjected to comparative phenotypic analyses, including monitoring growth in the presence of varying levels of hops using gradient agar plates, as well as relative growth rates in beer. Variability in beer-growth rate was noted among the plasmid-minus variants compared to the typical growth pattern of the parental Pc344 isolate possessing a complete set of plasmids. Variability of growth was also observed when analyzing for hops resistance. The correlation between plasmid elimination and altered phenotype confirms that the methodology used here is an appropriate starting point for investigating the role that plasmid genes play in the beer-spoiling capability of Pc344. Being able to attribute an organism’s beer-spoiling capacity in part to the presence of specific plasmids provides a clear direction for a more refined search of beer-spoilage associated genes.

Barry Ziola received a B.S. degree (with honors) in botany from McGill University, Montreal, in 1970. After completing a Ph.D. degree in biochemistry at the University of Alberta, Edmonton, in 1975, he undertook a three-year post-doctoral stint at the University of Turku, Turku, Finland. He has been at the University of Saskatchewan, Saskatoon, since 1978, with promotion to professor coming in 1986. His interest and continuing research in brewing spoilage bacteria dates to the mid-1980s.