Standardized methods to rear high-quality Galleria mellonella larvae for the study of fungal pathogens

The greater wax moth, Galleria mellonella (L.) (Lepidoptera: Pyralidae), has gathered widespread interest in the biomedical research community as an alternative non-mammalian model to study bacterial and fungal infections in humans. Nevertheless, most laboratories rely on commercial breeding companies that grow these larvae in bulk. Variation in rearing conditions between these companies and shipping of the larvae affects the response of G. mellonella to infection, both decreasing the reproducibility and comparability of infection experiments across laboratories. To unlock the full potential of G. mellonella as an infection model, standardized rearing is essential to produce healthy larvae of uniform quality. To this end, we developed an optimized protocol using a high-nutritional artificial diet and cost-effective housing solutions to ensure optimal conditions during each stage of the G. mellonella life cycle. This resulted in the rapid generation of larvae with a short larval phase, large size, and increased weight compared to other protocols and commercial larvae. A survival analysis comparing the standardized larvae with commercially obtained larvae showed significantly higher survival of standardized reared larvae after infection with Candida albicans (CP Robin) Berkhout (Saccharomycetales), proving the higher quality of standardized reared larvae.