Bacillus subtilis var. natto as a naturally optimized biotin biosensor

Bacillus subtilis var. natto is a naturally occurring biotin auxotroph widely used in traditional soybean fermentation. In this study, we evaluated its potential as a microbial biotin biosensor without genetic modification. Using a two-layer agar diffusion assay, growth responses were examined across biotin concentrations ranging from 0.001 to 200 mg/L. Halo diameters increased proportionally with biotin concentrations. The strain exhibited detectable responses over more than five orders of magnitude of biotin concentration without engineering of biotin transport or biosynthetic pathways. These results demonstrate that B. subtilis var. natto can serve as a simple, safe, and cost-effective biotin bioassay platform and highlight the utility of naturally evolved auxotrophic microorganisms as biosensors.