Simultaneous Detection of Multiple Wine-Spoilage Organisms Using a PCR-Based DNA Dipstick Assay

Author:

Janagama Harish K1,Mai Tam1,Han Sukkhyun1,Nadala Lourdes1,Nadala Cesar1,Samadpour Mansour2

Affiliation:

1. Molecular Epidemiology, Inc., Lake Forest Park, WA 98115, USA

2. IEH Laboratories and Consulting Group, Inc.,Lake Forest Park, WA 98155

Abstract

Abstract Background: The presence of microbial contaminants such as Brettanomyces in wine can lead to undesirable wine. Therefore, monitoring for the presence of these spoilageorganisms is critical for winemakers to ensure thequality of their end product. Objective: To address this problem, Molecular Epidemiology, Inc. (MEI, Seattle, WA) has developed a wine-spoilage organism detection kit consisting of a multiplex PCR DNA dipstick that simultaneously detects these organisms. Methods: Wine samples obtained from local wineries that tested negative by routine microbiological culture were spiked with the target microorganisms, while samples that were designated as spoiled by the wineries were usedas-is without spiking for assessing the performancecharacteristics of the DNA dipstick assay. Microbial enumeration was performed following standard microbiological plating methods. Samples spiked with low cell numbers (<5 cells per 100 mL) were enriched using wine enrichment media (WSE; optional component of the kit) prior to analysis using the DNA dipstick assay. Suitability of WSE medium to support the growth of wine-spoilage microorganisms was compared with standard microbiological media. Results: Testing of 92 diverse bacterial and yeast strains commonly found in winery and food operations and 50 various strains of spoilage organisms isolated from wineries indicated that the dipstick assay can exclusively detect the target wine-spoilage microorganisms. All target spoilage organisms in samples containing low cell numbers (<5 cells per 100 mL) were detected by dipstick assay 48 hpostenrichment in WSE, except for a few strains of Brettanomyces bruxellensis that required longer incubation times. Conclusions: The wine-spoilage organism detection kithas a detection limit of 10 cells/mL. Highlights: The kit can be used at different stages of the wine-making process to detect multiplespoilage-causing microorganisms in a single assay, thus offering a convenient test system for winemakers interested in monitoring the quality of their product.

Publisher

Oxford University Press (OUP)

Subject

Pharmacology,Agronomy and Crop Science,Environmental Chemistry,Food Science,Analytical Chemistry

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