Rapid monitoring of sourdough starter and contaminants using a dual-lateral flow immunochromatographic assay.

Sourdough maintenance via daily backslopping faces the persistent risk of starter lactic acid bacteria (LAB) being displaced by competing contaminants. Here, we developed a dual-lateral flow immunochromatographic assay (LFIA) system to simultaneously monitor starter health (Fructilactobacillus sanfranciscensis; Fsanf-LFIA) and contaminant presence (Pediococcus acidilactici; Ped-LFIA). Fsanf-LFIA and Ped-LFIA both exhibited detection limits of 5 log cells/test. Fsanf-LFIA was highly specific to certain strains of F. sanfranciscensis, whereas Ped-LFIA demonstrated a broader detection spectrum, cross-reacting with P. pentosaceus, other LAB (excluding F. sanfranciscensis), Bacillus, and Staphylococcus species. To evaluate practical on-site utility, sourdoughs inoculated with F. sanfranciscensis (9 log cells/g) and P. acidilactici (2 log cells/g) were maintained via daily backslopping at 23 °C, 30 °C, and 37 °C for 4 days. On day 4, conventional monitoring parameters failed to capture any change in sourdough grown at 37 °C, as pH levels dropped below 4.5 and total LAB counts remained virtually identical across all temperatures. However, while Fsanf-LFIA remained positive across all temperatures, Ped-LFIA turned positive exclusively at 37 °C. Amplicon sequencing validated these results, revealing that the abundance of F. sanfranciscensis plummeted to 6.2% at 37 °C, and Pediococcus became the dominant taxon. These findings demonstrate that the developed dual-LFIA system, which requires no specialized equipment, can quickly (within 1 h) detect early microbial displacement via visual-inspection-based binary judgment. This user-friendly system holds strong promise as a versatile quality-control tool for both industrial and artisanal sourdough-manufacturing facilities.