Quantification of viable bacteria in fermented foods using an enzymatic cascade-based pre-amplification-free RNA assay

Abstract

Rapid measurement of viable bacteria during fermentation and within fermented food products is important to ensure their quality and safety, however, tools are still lacking. Herein, we developed an LbuCas13a–LbCas12a cascade assay that enables rapid quantification of viable bacteria in fermented foods. By leveraging the cascade reaction to amplify the signal triggered by bacterial RNAs, the assay allows to detect as low as 5 × 10³ CFU mL⁻¹ lactic acid bacteria without nucleic acid pre-amplification. Using the assay, we achieved accurate quantification of viable bacteria proportions in heterogeneous populations and successfully evaluated the viable content in fermented dairy products. This strategy provides an efficient molecular tool for quality assessment of fermented food products.