中国乳业 ›› 2026, Vol. 0 ›› Issue (4): 71-79.doi: 10.12377/1671-4393.26.04.12

• 质量安全 • 上一篇    下一篇

乳品中乙型副伤寒沙门氏菌qPCR方法的构建与评价

徐佳薇1, 刘灵帆1, 蔡国屹1, 薛瑞奇2, 于玺2, 张业尼1,*   

  1. 1.天津农学院食品科学与生物工程学院,天津 300384;
    2.山东德正乳业股份有限公司,山东威海 264400
  • 出版日期:2026-04-25 发布日期:2026-05-22
  • 通讯作者: *张业尼（1984-）,女,山东济南人,博士,研究方向为病原微生物检测。
  • 作者简介:徐佳薇（2001-）,女,辽宁抚顺人,硕士,研究方向为食品加工与安全;刘灵帆（2004-）,男,河南洛阳人,本科,研究方向为微生物检测;蔡国屹（2006-）,男,贵州黔西南人,本科,研究方向为微生物检测;薛瑞奇（2001-）,男,山西怀仁人,本科,研究方向为乳源功能物质开发;于 玺（1982-）,男,山东威海人,本科,研究方向为功能性乳制品开发。
  • 基金资助:
    天津农学院校企合作项目—高蛋白健康乳粉深加工关键技术及产业化（TNHXKJ2025016）

Construction and Evaluation of qPCR Method for Salmonella Paratyphi B in Dairy Products

XU Jiawei1, LIU Lingfan1, CAI Guoyi1, XUE Ruiqi2, YU Xi2, ZHANG Yeni1,*   

  1. 1. College of Food Science and Bioengineering,Tianjin Agricultural University,Tianjin 300384;
    2. Shandong Dezheng Dairy Co.,Ltd.,Weihai Shandong 264400
  • Online:2026-04-25 Published:2026-05-22

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摘要： [目的]构建并评价乳品中乙型副伤寒沙门氏菌的实时荧光定量PCR（qPCR）检测方法。[方法]以乙型副伤寒沙门氏菌的特异性invA基因为靶标设计引物,优化SYBR Green Ⅰ染料法的qPCR反应体系与条件后,对梯度稀释的质粒标准品进行扩增以建立标准曲线,并系统验证该方法的灵敏度、特异性与重复性,同时与传统培养法进行检测性能对比。[结果]所建立的标准曲线方程为y=-3.186 6x+13.172,相关系数（R²）达0.994 9,扩增效率（E%）为105.9%,线性关系良好;该qPCR方法检测乙型副伤寒沙门氏菌的灵敏度可达17.9 拷贝数/μL;特异性验证中,仅乙型副伤寒沙门氏菌呈阳性扩增,大肠杆菌、金黄色葡萄球菌等非沙门氏菌均为阴性,无交叉反应;重复性试验表明,组内变异系数为0.2%~0.69%,组间变异系数为0.49%~1.54%,均小于3%,稳定性优异。与传统培养法（需5~7 d）相比,该qPCR方法可将检测时间缩短至6 h内,显著提升检测效率。[结论]本研究建立的SYBR Green Ⅰ qPCR方法具有灵敏度高、特异性强、重复性好、操作简便及检测快速的优势,为乳品中乙型副伤寒沙门氏菌的快速筛查、精准定量及食品安全风险防控提供了可靠技术支撑。

关键词: qPCR, 乙型副伤寒沙门氏菌, invA基因

Abstract: [Objective] To establish and evaluate a real-time fluorescent quantitative PCR（qPCR）method for detecting Salmonella paratyphi B in dairy products. [Method] Specific invA gene of Salmonella paratyphi B was used as the target to design primers. After optimizing the qPCR reaction system and conditions with SYBR Green Ⅰ dye,amplification was performed on gradient diluted plasmid standards to establish a standard curve. The sensitivity,specificity and repeatability of the method were systematically verified,and the detection performance was compared with the traditional culture method. [Result] The standard curve equation was y=-3.186 6 +13.172,with a correlation coefficient（R²）of 0.9949. The amplification efficiency（E%）was 105.9%. The linear relationship was good. The qPCR method for detecting Salmonella paratyphi B had a sensitivity of 17.9 copies/μL. In the specificity verification,only Salmonella paratyphi B showed positive amplification,while Escherichia coli,Staphylococcus aureus, etc. were negative. There was no cross-reaction. The repeatability test showed that the intra-group coefficient of variation was 0.2%~0.69%,and the inter-group coefficient of variation was 0.49%~1.54%,all of which were less than 3%,indicating excellent stability. Compared with the traditional culture method（requiring 5~7 days）,this qPCR method can shorten the detection time to within 6 hours,significantly improving the detection efficiency. [Conclusion] The SYBR Green Ⅰ qPCR method established in this study has the advantages of high sensitivity,strong specificity,good repeatability,simple operation and rapid detection,providing reliable technical support for the rapid screening,precise quantification and food safety risk prevention and control of Salmonella paratyphi B in dairy products.

Key words: qPCR, Salmonella Paratyphi B, invA gene

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