Strong antimicrobial activity and unique physicochemical characteristics in honey from Australian stingless bees Tetragonula carbonaria, Tetragonula hockingsi, and Austroplebeia australis.

Abstract

Natural products have evolved antimicrobial properties that can be exploited in our search for new ways of treating infectious diseases. This study evaluates the antimicrobial properties and chemical profiles of honey produced by the Australian stingless bee species Tetragonula carbonaria, Tetragonula hockingsi, and Austroplebeia australis against selected human pathogens. Using broth microdilution methods, we found that all tested honey samples had antimicrobial activity. The fungal dermatophyte species Trichophyton interdigitale displayed the highest susceptibility (average MICs: 4%-9% [wt/wt]), followed by Staphylococcus aureus (9%-11%), Escherichia coli (10%-13%), and Cryptococcus neoformans (24%-34%). T. carbonaria honey had the highest overall activity, while A. australis honey was the weakest. After heat treatment at 80°C for 30 min to remove hydrogen peroxide (H₂O₂)-based activity, T. carbonaria and T. hockingsi honey retained significant non-peroxide activity against E. coli (14% and 17%, respectively) and S. aureus (17% and 18%, respectively), although their efficacy against the fungal pathogens diminished. Chemical analysis revealed distinct differences in H₂O₂ production, color intensity, phenolic and antioxidant content, density, and pH among the honey types. The dynamic generation of H₂O₂ in stingless bee honey was remarkably prolonged, with some samples producing H₂O₂ for more than 6 days. Proteomic analysis identified diverse proteins that may contribute to antimicrobial efficacy. Phenolic extracts had antimicrobial activity, with flavonoids identified as potential contributors. T. carbonaria honey re-tested after 18 years of storage retained substantial non-peroxide-based activity. Overall, this study highlights the unique properties of stingless bee honeys and their potential as natural antimicrobial agents.IMPORTANCEAntimicrobial resistance poses a critical global health challenge. Various natural products have evolved as a defense against microbial attack and can be exploited as novel therapeutic agents. While honey from the European honey bee (Apis mellifera) is well studied, the medicinal potential of Australian stingless bee honey remains underexplored. This study demonstrates that honeys produced by the stingless bee species Tetragonula carbonaria, Tetragonula hockingsi, and Austroplebeia australis possess unique antimicrobial properties that persist after heat treatment and following long-term storage and are distinct from the antimicrobial properties of honey bee honey. Diverse bioactive compounds, including phenolics and proteins, were seen, underscoring the complexity of these honeys as antimicrobial agents. These honeys have potential as sustainable, natural solutions for combating drug-resistant infections and could extend the scope of honey-based therapeutics.