国标现行的平板活菌计数无法实现益生菌在复杂生物体系中的菌株水平鉴别与定量,成为制约当前产业监管和产品区分的一个重要技术难题。该研究以鼠李糖乳酪杆菌MP108为研究示例,建立以菌株全基因组信息为基础、特征序列为靶点、实时荧光定量PCR等技术协同的菌株水平定量方法。结果表明,该方法能够在复杂微生物体系(粪便样品)中鉴别出MP108菌株的存在并实现单菌定量,检出限为104 细胞数/g,灵敏度为10 copies/μL DNA溶液,在复杂体系中的定量准确度与基于纯培养菌株的平板计数法对照值无显著性差异。该方法的建立有望为推动益生菌食品精细化检测提供理论参考。
The prevailing national standard for probiotic enumeration falls short in achieving strain-level identification and quantification within complex biological matrices, posing a notable technical hurdle that constrains current industry regulations and impedes product differentiation.To address this issue, this study utilized Lactobacillus rhamnosus MP108 as a model and devised a strain-specific quantitative approach leveraging whole-genome data of the strain, targeting characteristic genetic sequences, and employing advanced techniques such as real-time fluorescence quantitative PCR.The findings demonstrated that this method effectively discerned the presence of the MP108 strain and enabled precise quantification in complex microbial environments, boasting a detection threshold of 104 cells per gram of biological sample and a sensitivity of 10 copies per microliter of DNA solution.Notably, the quantitative accuracy of this approach in complex systems closely aligned with reference values obtained through conventional plate counting methods utilizing pure culture strains.Consequently, this method holds promise in furnishing foundational insights for enhancing the nuanced detection of probiotic foods.
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