Rapid Detection of Yeast and Mold in Filterable Beverage Using PCR Assay

Introduction: Fungal spoilage of food accounts for 5% - 10% of all losses in global food production. This spoilage is not only a quality issue, but also food safety concern since many common mold species are dangerously toxigenic. Due to this concern, it is imperative that manufacturers closely monitor their processes to prevent such contamination from occurring. While traditional colony count methods for yeasts and molds require 5-7 days to achieve a result, the Yeast and Mold PCR assay provides same day results for food samples containing > 500 CFU/g (direct method), or in just 2 days for food samples with 50-500 CFU/g (enrichment method). This study was to validate Yeast and Mold PCR Assay in filterable beverage for the detection of yeast and mold at very low level.

Purpose: The objectives of this study were to validate the PCR assay for detecting low levels (1 CFU/5ml) of yeast and mold in filterable beverage and to compare the efficiency of sample processing through both filtration and centrifugation to collect yeast and mold cells.

Methods: Two master samples of soda were spiked with Saccharomyces cerevisiae and Aspergillus niger at a level targeting 1 CFU/5ml. Samples were either filtered using in-line filter units or subjected to centrifugation, then all samples were incubated at 25 °C for 44 hrs. Samples were prepared for yeast and mold detection and a full process was run on the BAX® System instrument according to the procedures described in the BAX® System User Guide.

Results: All spiked samples regardless of the sample processing method used, returned positive results with the PCR assay for the target organism, and all non-spiked samples returned negative results. Actual spiking levels for inoculated samples was determined to be 0.6 CFU/ml for S. cerevisiae and 0.14 CFU/ml for A. niger after the 44-hour incubation.

Significance: This study demonstrates that the PCR assay can detect yeast and mold in soda within two days at only 1-3 CFU/5 ml. Sample processing using filtration is equivalent to centrifugation to collect cells from filterable beverages. This study also demonstrates that the PCR assay can detect yeast and mold in filterable beverages, provided that at least 1 CFU is retained by the filter.