Monitoring of Biofilm Development in Bee Hemolymph Through In-Fiber Interferometers

This work presents a wavelength-encoded fiber optic interferometric platform for real-time biofilm detection and monitoring. The sensors were realized through a simple and cost-effective fabrication procedure based on overlap fusion splicing of standard single-mode optical fibers. Numerical simulations were conducted to provide design guidelines and achieve high sensitivity. The fabricated devices exhibited a sensitivity to the surrounding medium refractive index (SRI) of approximately 200 nm/RIU in aqueous solutions. The platform was first validated by monitoring Pseudomonas alcaligenes biofilm formation over 70 h through the wavelength shifts of the interference minima. Most importantly, it was subsequently applied to in situ and real-time monitoring of hemolymph-derived biofilm directly inside living bee larvae over a one-day period, representing, to the best of our knowledge, the first demonstration of this application. Atomic force microscopy (AFM) characterized the biological layers formed on the optical fiber surface, revealing thicknesses up to approximately 2 μm and enabling their development to be correlated with the optical response. These results demonstrate the potential of compact, low-cost in-fiber interferometers for wavelength-encoded biofilm monitoring in complex biological environments.