Université de Bordeaux, ISVV, Bordeaux Aquitaine, INRA UMR 1219, 210 Chemin de Leysotte, CS 50008, 33882 Villenave d'Ornon Cedex, France.
Food Microbiol. 2010 Aug;27(5):559-67. doi: 10.1016/j.fm.2010.01.006. Epub 2010 Feb 1.
The complex microbial ecosystem of grape must and wine harbours a wide diversity of yeast species. Specific oligonucleotide primers for real-time quantitative PCR(QPCR) were designed to analyse several important non-Saccharomyces yeasts (Issatchenkia orientalis, Metschnikowia pulcherrima, Torulaspora delbrueckii, Candida zemplinina and Hanseniaspora spp.) and Saccharomyces spp. in fresh wine must, during fermentation and in the finished wine. The specificity of all primer couples for their target yeast species were validated and the QPCR methods developed were compared with a classic approach of colony identification by RFLP-ITS-PCR on cultured samples. Once the methods had been developed and validated, they were used to study these non-Saccharomyces yeasts in wine samples and to monitor their dynamics throughout the fermentation process. This study confirms the usefulness and the relevance of QPCR for studying non-Saccharomyces yeasts in the complex yeast ecosystem of grape must and wine.
葡萄汁和葡萄酒复杂的微生物生态系统中蕴藏着丰富多样的酵母种群。本研究设计了特定的寡核苷酸引物用于实时定量 PCR（QPCR）分析，以研究新鲜葡萄汁、发酵过程中和陈酿葡萄酒中的几种重要非酿酒酵母（东方伊萨酵母、膜毕赤酵母、德巴利氏酵母、近平滑假丝酵母和汉逊酵母属）和酿酒酵母。所有针对目标酵母种的引物对的特异性均经过验证，所开发的 QPCR 方法与传统的基于 RFLP-ITS-PCR 的培养物菌落鉴定方法进行了比较。方法建立和验证后，用于研究葡萄酒样品中的这些非酿酒酵母，并监测它们在发酵过程中的动态变化。本研究证实了 QPCR 在研究葡萄汁和葡萄酒复杂酵母生态系统中非酿酒酵母中的有用性和相关性。
