Establishment of a rapid counting method for lactic acid bacteria and yeast in dairy products

Abstract

The plate count agar (PCA) method is a common technique used to count microorganisms in dairy products, but this method is time-consuming and highly selective by the medium. This study adopted Lab158 and PF2 as the universal probes for hybridising lactic acid bacteria and yeasts, respectively, and investigated the effects of fixation time, hybridisation time, hybridisation temperature, probe concentration and propidium iodide (PI) concentration on hybridisation efficiency in the samples. The results showed that the highest hybridisation efficiency for lactic acid bacteria and yeast, along with the best double staining results, was observed under the following conditions: a fixation time of 80 min, a hybridisation time of 180 min, a hybridisation temperature of 48°C, a probe volume of 5 μL for lactic acid bacteria and 3 μL for yeast, and a PI volume of 5 μL for lactic acid bacteria and 3 μL for yeast. The fluorescence in situ hybridisation-flow cytometry (FISH-FCM) and PCA counting results showed a significant positive correlation (r > 0.9). The FISH-FCM and PCA counting results showed a significant positive correlation (r > 0.9). The FISH-FCM counting method can serve as a quick and accurate method for the counting of lactic acid bacteria and yeast in dairy products.