Detection of Spoilage-Associated Acetic Acid Levels Using a Transcription-Based Whole-Cell Biosensor

Monitoring acetic acid (AC) in fermentation processes is essential as excessive AC accumulation, particularly during alcoholic fermentation, can disrupt fermentation and lead to spoilage. However, conventional detection methods such as steam distillation, GC–MS, and HPLC are costly, time-consuming, and require liquid-phase samples, limiting their use for real-time monitoring and early identification of AC buildup. Here, we present an alternative tool for AC detection using a whole-cell bacterial biosensor, which utilises the YwbIR transcriptional regulator from Bacillus subtilis. The designed biosensor exhibits high sensitivity, manifesting a linear response with (R² = 0.97) from 0 to 1.0 g/L and a 5–8 fold induction at wine spoilage-relevant concentrations. It retains functionality in ethanol-rich matrices (up to 14.5% v/v) and enables headspace detection. Specificity assays and molecular docking analyses confirm high affinity for AC over other volatile fatty acids. This biosensor offers a low-cost solution for real-time AC monitoring, allowing timely intervention before spoilage occurs and supporting improved quality assurance in fermentation-driven food and beverage production.