基于流式细胞仪提高乳制品商业无菌检验时效性的研究

Research on improving the timeliness of commercial sterile testing of dairy products based on flow cytometry

中文关键词:  商业无菌  地衣芽孢杆菌  蜡样芽胞杆菌  乳制品  流式细胞仪

英文关键词:Commercial sterility  Bacillus licheniformis  Bacillus cereus  Dairy products  Flow cytometry

中文摘要:

      目的 基于流式细胞仪探究提高乳制品商业无菌检验项目时效性。方法 以地衣芽孢杆菌CMCC(B) 63552和蜡样芽孢杆菌CMCC(B) 63303菌株制备定量参考品。以4种灭菌乳为基质，制备染菌浓度分别为800、8000、80000 CFU/mL的菌悬液，测试流式细胞仪法灵敏度。制备染菌浓度为8 CFU/样品和80 CFU/样品的灭菌乳和调制乳样品，经(36±1) ℃保温(24±2) h后，按照GB 4789.26法和流式细胞仪法进行检测，并分析比较两种方法检测结果。结果 在灭菌乳基质中，染菌浓度为800 CFU/mL时，流式细胞仪法对人工污染地衣芽孢杆菌样品的阳性检出率为75%，而对人工污染蜡样芽孢杆菌的样品均未检出阳性；染菌浓度为8000 CFU/mL和80000 CFU/mL时，流式细胞仪法对地衣芽孢杆菌和蜡样芽孢杆菌污染样品的阳性检出率均为100%。对人工污染8 CFU和80 CFU地衣芽孢杆菌和蜡样芽孢杆菌的灭菌乳和调制乳样品，经保温处理后，流式细胞仪法检出率为100%，GB 4789.26法中未见样品膨胀，但是镜检和培养结果检出率为100%。结论 当样品中染菌浓度≥8000 CFU/mL时，流式细胞仪法可稳定检出。人工污染地衣芽孢杆菌和蜡样芽孢杆菌的灭菌乳和调制乳样品经保温处理(24±2) h后，流式细胞仪法和GB 4789.26法检出率均为100%。

英文摘要:

      Objective To improve the timeliness of commercial sterile testing projects for dairy products based on flow cytometry. Methods Prepare quantitative reference materials with Bacillus licheniformis CMCC (B) 63552 and Bacillus cereus CMCC (B) 63303. Prepare bacterial suspensions with concentrations of 800, 8000 80000 CFU/mL using four types of sterilized milk as substrates, and test the sensitivity of flow cytometry method. Make up sterilized milk and modified milk samples with bacterial concentrations of 8 CFU/sample and 80 CFU/sample, respectively. After being kept at (36±1) ℃ for (24±2) hours, detect them according to GB 4789.26 and flow cytometry methods, and compare the detection results of the two methods. Results In the sterilized milk matrix, when the concentration was 800 CFU/mL, the positive detection rate of flow cytometry for artificially contaminated Bacillus licheniformis samples was 75%, while no positive results were detected for artificially contaminated Bacillus cereus samples. When the concentration are 8000 CFU/mL and 80000 CFU/mL, the positive detection rates of flow cytometry for contaminated samples of Bacillus licheniformis and Bacillus cereus were 100%. In the insulation test, the detection rates of sterilized and modified milk samples artificially contaminated with 8 CFU and 80 CFU of Bacillus licheniformis and Bacillus cereus were 100% using flow cytometry. The GB 4789.26 method showed no expansion in the samples, but positive rate was 100% in microscopic detection. Conclusion When the concentration was ≥8000 CFU/mL, the flow cytometry method could show positive results stably. For the samples contaminated with 8 CFU and 80 CFU of Bacillus licheniformis and Bacillus cereus, the detection rates of GB 4789.26 and flow cytometry methods were 100% after insulation (24±2 h).