Janagama Harish K, Mai Tam, Han Sukkhyun, Nadala Lourdes, Nadala Cesar, Samadpour Mansour
Molecular Epidemiology, Inc., Lake Forest Park, WA 98115.
J AOAC Int. 2019 Mar 1;102(2):490-496. doi: 10.5740/jaoacint.18-0144. Epub 2018 Aug 20.
The presence of microbial contaminants such as in wine can lead to undesirable wine. Therefore, monitoring for the presence of these spoilage organisms is critical for winemakers to ensure the quality of their end product. 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. 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 used as-is without spiking for assessing the performance characteristics 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. 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 h postenrichment in WSE, except for a few strains of that required longer incubation times. The wine-spoilage organism detection kit has a detection limit of 10 cells/mL. The kit can be used at different stages of the wine-making process to detect multiple spoilage-causing microorganisms in a single assay, thus offering a convenient test system for winemakers interested in monitoring the quality of their product.
葡萄酒中存在如 等微生物污染物会导致葡萄酒品质不佳。因此，监测这些腐败微生物的存在对于酿酒师确保最终产品质量至关重要。为解决这一问题，分子流行病学公司（位于华盛顿州西雅图市的MEI）开发了一种葡萄酒腐败生物检测试剂盒，该试剂盒由一种多重PCR DNA试纸条组成，可同时检测这些生物。从当地酒庄获取的经常规微生物培养检测为阴性的葡萄酒样品接种目标微生物，而酒庄认定为变质的样品则直接使用，不进行接种，以评估DNA试纸条检测方法的性能特征。微生物计数按照标准微生物平板培养方法进行。接种低细胞数（每100 mL少于5个细胞）的样品在使用DNA试纸条检测前，先用葡萄酒富集培养基（WSE；试剂盒的可选成分）进行富集。将WSE培养基支持葡萄酒腐败微生物生长的适用性与标准微生物培养基进行了比较。对酒庄和食品生产中常见的92种不同细菌和酵母菌株以及从酒庄分离出的50种不同腐败生物菌株进行测试表明，试纸条检测方法能够专门检测目标葡萄酒腐败微生物。在WSE中富集48小时后，通过试纸条检测法可检测出含有低细胞数（每100 mL少于5个细胞）样品中的所有目标腐败生物，除了少数几株 需要更长培养时间的菌株。该葡萄酒腐败生物检测试剂盒的检测限为10个细胞/mL。该试剂盒可用于葡萄酒酿造过程的不同阶段，在一次检测中检测多种引起腐败的微生物，从而为有兴趣监测其产品质量的酿酒师提供了一种便捷的检测系统。
