Abstract
Initial microbiological species and quantity of raw milk significantly affect dairy product quality. In particular, some psychrotrophic bacteria in raw milk can produce heat-stable hydrolases that reduce the stability of dairy products during their shelf life. The purpose of this study was to establish a multiplex real-time fluorescence quantitative polymerase chain reaction (qPCR) rapid detection method for Pseudomonas fluorescens and Lactococcus lactis subsp. lactis, which generally produce protease in raw milk. The primers and probes with good specificity, universality and high sensitivity to the target bacteria were screened by ordinary PCR and qPCR experiments. The quantitative linear relationship and sensitivity of the method for detecting pure DNA and artificially contaminated simulated milk samples were verified. The results showed that the limit of detection (LOD) of this method for pure DNA could reach 3.16 × 10−5 ng/μL. The detection of artificially contaminated samples showed that the LOD for P. fluorescens and L. lactis subsp. lactis could reach 250 and 22 CFU/mL, respectively. The multiplex qPCR method established in this study has good specificity and high sensitivity, which can be used to rapidly detect psychrotrophic bacteria in raw milk. It provides an effective reference strategy for the dairy industry to ensure the microbiological quality of raw milk from the source.
Published Version
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