Microbiology Session
Barry Ziola, University of Saskatchewan, Saskatoon, Saskatchewan, Canada
Co-author(s): Barry Bushell and Vanessa Pittet, University of Saskatchewan, Saskatoon, SK, Canada
ABSTRACT: Important to the brewing industry is the ability
to determine if bacteria present in post-filtered beer are capable of
growing in (i.e., spoiling) the product. A major factor for bacterial
growth in beer is hop resistance, which we have previously shown can be
assessed with hop extract (iso-alpha-acids) infused agar (HGA) plates
with a bitterness gradient ranging from 0 to 29 BU. To make hop agar
gradient plates more applicable to a brewery setting, we assessed
various parameters, including the steepness of the BU gradient, effect
of the presence of ethanol, and stability of hop extract over time. HGA
plating consists of pouring a dual layer agar plate such that the bottom
layer containing the maximum BU concentration forms a 10.5 degree wedge
of hop-infused agar, which is then covered by a top layer of hop-free
agar. Diffusion from the bottom agar layer creates a hops gradient at
the agar surface. Lines of bacterial cultures are stamped upon the
surface, and the distance of growth indicates the BU level at which
growth is inhibited. The effect of ethanol on hop resistance can be
assessed by including ethanol in both agar layers. Gradient plates
containing four different ranges of hop concentrations were tested (0–29
BU, 1×; 0–58 BU, 2×; 0–87 BU, 3×; 0–116 BU, 4×). In addition, to assess
hop stability, two samples of the same commercial hop preparation
separated by 5 years in storage at 4°C were tested. In this study, 26
isolates (22 Lactobacillus and 4 Pediococcus) were
assessed. Of these, 10 grew completely through the 1× gradient plates,
and hop resistance could not be scored. With 2× hop-gradient plates only
four isolates could not be scored. Ultimately, all four could be scored
(grew only partially along the hop gradient) using a 4× plate. With 2×
plates containing 5% ethanol, three isolates showed increased hop
resistance—two of which we had previously described as having
ethanol-enhanced hop resistance. This increased hop resistance in the
presence of ethanol was verified for one of the three isolates by growth
trials in liquid medium. Lastly, no difference in antimicrobial
efficacy of the two hop extracts was found. Based on these results, the
use of 2× hop gradient agar plates (0–58 BU) is recommended for
routinely assessing bacterial hop resistance, with highly hop-resistant
isolates requiring a 4× gradient plate (0–116 BU). As well, properly
stored hop extracts can be used over extended periods of time in hop
gradient plates.
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.
VIEW PRESENTATION 174
