Evaluation of a Highly Multiplexed, Automated Assay for the Detection of Beer Spoilage Flora

Introduction:  Organisms that can spoil beer include both yeast and bacterial strains. Culture-based isolation and identification may require lengthy procedures with skilled interpretation. Rapid, non-culture methods, such as PCR, can provide fast and accurate detection. These methods shorten the overall time to result from more than five days to a single day. The Rheonix SpoilerAlert™ assay kit is highly automated, with only a single pipetting step required to obtain PCR results from a sample. The initial commercial version of the assay amplifies 10 target genes present, individually or in combination, in various lactic acid bacteria and the wild yeast, Brettanomyces bruxellensis. The assay also detects the brewers’ yeast, Saccharomyces cerevisiae. Due to industry concern, S. cerevisiae var. diastaticus target was added.

Purpose: The purpose of this study was to characterize the performance of the updated assay for detection of spoilage flora and hop resistance genes. Inclusivity and exclusivity of the new assay was characterized and sensitivity was determined.

Methods: Studies evaluating the assay were conducted using target and non-target cells, with and without enrichment.

Results: The sensitivity of the assay was determined to be ~10⁴ CFU/ml for each of the targets in post enriched samples and <10 CFU/sample before enrichment. The additional target for S. cerevisiae var. diastaticus was detected, as expected, in this variant but was not detected in multiple S. cerevisiae strains that were not S. cerevisiae var. diastaticus.

Significance: This method for beer spoilage flora detection and characterization provides reduced processing time compared to culture and provides a genetic characterization of the bacterial genes that confer hop resistance. This allows brewers to make earlier decisions regarding spoilage flora presence in their products and processing environments. This technology can be used for other market segments, where well characterized microbiota are associated with reduced shelf life.