Abstract The greater wax moth (GWM), Galleria mellonella (Lepidoptera: Pyralidae), is a major bee pest that causes significant damage to beehives and results in economic losses. Bacillus thuringiensis (Bt) appears as a potential sustainable solution to control this pest. Here, we develop a novel Bt strain (designated BiotGm) that exhibits insecticidal activity against GWM larvae with a LC 50 value lower than 2 μg/g, and low toxicity levels to honey bee with a LC 50  = 20598.78 μg/mL for larvae and no observed adverse effect concentration = 100 μg/mL for adults. We design an entrapment method consisting of a lure for GWM larvae, BiotGm, and a trapping device that prevents bees from contacting the lure. We find that this method reduces the population of GWM larvae in both laboratory and field trials. Overall, these results provide a promising direction for the application of Bt-based biological control of GWM in beehives, although further optimization remain necessary.