牛奶发酵过程中阪崎克罗诺杆菌可形成活的非可培养状态

该研究以保加利亚乳杆菌为牛奶发酵剂，研究了牛奶发酵过程中阪崎克罗诺杆菌与保加利亚乳杆菌的相互作用以及阪崎克罗诺杆菌存活量的变化情况，并结合16S rDNA高通量测序技术和PMA-qPCR方法分析阪崎克罗诺杆菌在牛奶发酵过程中是否形成“活的非可培养”状态。结果显示，保加利亚乳杆菌在牛奶中发酵48 h后pH值可达到3.40，酸度值可达212.00。在发酵前期接种阪崎克罗诺杆菌，48 h后pH值和酸度值分别为3.50和193.30；在发酵中期接种阪崎克罗诺杆菌，48 h后pH值和酸度值分别为3.47和214.30。发酵前期阪崎克罗诺杆菌的污染对保加利亚乳杆菌发酵结果的影响更大。阪崎克罗诺杆菌在牛奶中生长48 h后菌浓度可稳定在9.08 lg(CFU/mL)；不论在牛奶经发酵前期或中期接种阪崎克罗诺杆菌，发酵后期平板计数法均检测不到阪崎克罗诺杆菌，但16s rDNA和PMA-qPCR技术均可检测到阪崎克罗诺杆菌活菌的存在。该研究结果说明，阪崎克罗诺杆菌在牛奶发酵后进入了“活的非可培养”状态，该状态的存在会导致该菌检测时活菌数量的低估，从而引发乳及乳制品的安全隐患。

Formation of Viable but Non-culturable State of Cronobacter sakazakii during Milk Fermention

In this study, Lactobacillus bulgaricus, a commonly used milk starter, was selected for milk fermentation. The interactions between C. sakazakii and Lactobacillus bulgaricus and the survival of C. sakazakii were investigated during milk fermentation. Furthermore, 16s rDNA high-throughput sequencing technique and PMA-qPCR method were used to analyze whether C. sakazakii entered into the viable but non-culturable (VBNC) state during milk fermentation. The results showed that after 48 h-fermentation of L. bulgaricus in milk, the values of pH and acidity reached to 3.40 and 212.00, respectively. When C. sakazakii was inoculated in milk in the middle fermentation stage, the pH value and acidity value were 3.47 and 214.30 respectively. When C. sakazakii was inoculated in milk in the early fermentation stage, the values of pH and acidity were 3.50 and 193.30 respectively after 48 h fermentation; which had a greater adverse impact on the fermentation. The concentration of C. sakazakii stabilized at 9.08 lg(CFU/mL) after 48 hours of growth in milk, the growth of C. sakazakii could not be detected by plate counting in the later stage of fermentation whenever C. sakazakii was inoculated into the milk. However, the existence of viable C. sakazakii could be detected by 16s rDNA and PMA-qPCR technology, which indicated that C. sakazakii could be VBNC state during milk fermentation. The existence of VBNC state of C. sakazakii might lead to an underestimation of total viable pathogenic cells, thus causing potential health hazards of milk and dairy products.