Purpose: In order to provide a tool for controlling Z. bailii in beverages, a specific real-time PCR assay with associated DNA extraction method was developed allowing to recover DNA from complex matrices like wines.
Methods: This method was validated on a comprehensive panel of inclusivity and exclusivity strains naturally present in wine matrices. The performance of the DNA extraction was tested on various wine matrices from European and US origins as well as on fruit juice matrices to assess the efficiency of method to deal with potential PCR inhibitors. Eighty beverage samples corresponding to 15 different matrices (wine or fruit juices) were spiked using 2 different strains of Z. bailii to a theoretical level comprised between 1 and 1.8 x 107cell/ml.
Results: Twelve Z. bailii strains from different origins were assayed and were adequately detected. Conversely, 46 strains representing 26 species of yeast, mold or bacteria that can be found in wine did not yield a specific signal, thereby demonstrating the selectivity of the method. A standard calibration curve was generated demonstrating PCR efficiency of 145%, thereby establishing a good correlation between Cq values and CFU/ml of the 80 beverage samples. Finally, the assay was determined to detect as little as 1 CFU/ml.
Significance: This level of sensitivity can help food manufacturers and enologists in taking appropriate decisions for the management of wine and fruit beverages to avoid fermentation after bottling that may lead to explosion of bottles or bag-in-box.