A New Approach Integrating Brood-Associated Semiochemicals with Additional Feeding for Honey Bee (Apis mellifera) Colony Development

The aim of this study was to identify chemical formulations that stimulate <i>Apis mellifera</i> colony development by enhancing queen egg-laying under protein-supplemented conditions. Feeding trials were conducted in early spring, when natural food sources are scarce. The experiment was conducted in two Romanian apiaries and included four treatment groups. Three formulations included protein-enriched bee food: two standard variants and one supplemented with essential fatty acids. All were administered alongside behavioral stimulants (T1-T3). A fourth treatment served as a control, containing only protein-based food without brood pheromones or additional stimulants (T4). Pheromone blends were formulated based on brood-emitted volatiles identified by solid-phase microextraction-gas chromatography coupled with mass spectrometry (SPME-GC-MS). The effects of the treatments were evaluated by measuring queen egg-laying and brood area development. Results showed that treatments based on brood ester pheromones (BEP)-T1 and a fatty acid blend (FAB)-T3 significantly stimulated queen egg-laying and brood production, with comparable performance and a slight advantage for T3. In contrast, combining BEP with a fatty acid-supplemented protein diet (T2) produced a moderate effect, consistent with regulated lipid intake in honey bee colonies. These findings indicate that brood-associated semiochemicals, combined with protein supplementation, can effectively stimulate colony growth.