Engineered symbiont biosensor maps micron-scale sugar gradients in the honeybee gut

The honeybee gut microbiota plays a key role in shaping host health and susceptibility to disease. Yet, the nutrient environment it experiences within the gut remains poorly characterized. In particular, little is known about the spatial distribution of nutrients across the microbial community, as resolving such fine gradients in vivo has been technically challenging. Here, we engineer the native honeybee symbiont Snodgrassella alvi as a living biosensor to quantify the bioavailability of the dietary sugar arabinose within the gut. By expanding the genetic toolkit for S. alvi through chromosomal integration of high-burden genes and a suite of low-strength promoters, we achieve stable multi-gene expression without compromising host colonization. The resulting biosensor generates a specific, dose-dependent fluorescent response to arabinose in the living host, enabling visualization of sugar gradients across gut-associated bacterial biofilms at micron-scale resolution. Upon co-colonization with distinct Gilliamella species that differ in arabinose metabolism, the biosensor reported differential in vivo arabinose consumption, directly validating species-specific metabolic specialization within the host. Feeding bees with pollen further uncovered pronounced radial heterogeneity in the distribution of pollen-derived arabinose. These findings demonstrate how diet composition and microbial specialization generate fine-scale microenvironments within the gut. More broadly, this work establishes S. alvi as a genetically tractable platform for in situ biosensing, opening new avenues for dissecting metabolic interactions and nutrient distribution within living hosts.