In this study, a PMA-qPCR assay was developed for the enumeration of Bifidobacterium animalis subsp. lactis BB-12 viable cells in a non-dairy probiotic beverage. Probiotic viability was monitored in three formulations of probiotic passion fruit juice microencapsulated by spray drying, during 30 days of storage at 4 °C. Viable cells were quantified using qPCR and PMA-qPCR assays targeting tuf gene and by plate counting method. The limit of detection for all samples was 103 genome copies, corresponding to 21.3 pg of DNA. Higher CFU values were obtained for B. lactis BB-12 enumeration by qPCR, when compared to those obtained by PMA-qPCR and plate count, for all probiotic juice microcapsules. Similar quantification values were obtained by PMA-qPCR and plate counting for all samples and remained above 8 log CFU/g during the storage period. These results demonstrated that the PMA-qPCR technique is a promising approach for B. lactis BB-12 viable cell enumeration in complex matrices such as passion fruit juice microcapsules. This PMA-qPCR assay allowed the achievement of reliable results faster than with the traditional plate counting method.

中文翻译：

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单叠氮化丙锭与定量PCR结合应用评估动物双歧杆菌亚种的细胞活力 球菌在非乳制饮料益生菌

在这项研究中，开发了一种用于动物双歧杆菌亚种枚举的PMA-qPCR测定法。非乳益生菌饮料中的乳酸菌BB-12活细胞。在4°C下储存30天期间，通过喷雾干燥微胶囊化的三种益生百香果汁的配方中监测了益生菌的生存能力。使用靶向tuf基因的qPCR和PMA-qPCR分析方法以及平板计数法对活细胞进行定量。所有样品的检出限为103个基因组拷贝，相当于21.3 pg DNA。对于所有益生菌汁微胶囊，与通过PMA-qPCR和平板计数获得的乳酸菌相比，通过qPCR获得的乳酸双歧杆菌BB-12的CFU值更高。通过PMA-qPCR和所有样品的平板计数获得相似的定量值，并且在存储期间保持在8 log CFU / g以上。这些结果表明，PMA-qPCR技术是一种在复杂基质（如百香果汁微胶囊）中进行乳芽孢杆菌BB-12活细胞计数的有前途的方法。与传统的板计数方法相比，这种PMA-qPCR分析可以更快地获得可靠的结果。