Yeasts associated with nests of Australian stingless bees (Meliponini)

Australian eusocial stingless bees of the tribe Meliponini raise young on honey and pollen stored in pots made of resin-beeswax. These provisions harbour a diverse microbiome, including bacteria and fungi, that can contribute to improved nutritional value and preservation and thus to the overall health of the bee colony. Chemical analyses of Australian pot-pollen have suggested that some bioactive volatiles might originate from microbial fermentation. The gut microbiota of Australian Meliponini has been studied, but the microbiota associated with their nests remains unexplored. In this study, fungi and yeasts were isolated using selective high-osmolarity media and antibacterial agents from pot-pollen, pot-honey, brood cell walls (‘cerumen’) and food provisions from nine colonies of three Australian Meliponini species (Tetragonula carbonaria, T. hockingsi and Austroplebeia australis) located at the same site in South East Queensland. Bees from the 9 colonies were assumed to be foraging within the same radius. Fungi and yeasts were characterised by microscopy, sequencing of ITS and LSU ‘barcodes’ and subsequent BLAST search against GenBank, phylogenetic tree construction and amplicon concatenation. The relative abundance of bacteria in pot-pollen was estimated from 16S ‘barcodes’ by PCR amplification and Next Generation Sequencing using the MiSeq platform. Organisms identified include Ascomycota, Basidiomycota, Firmicutes and Proteobacteria previously unreported in nest materials of Australian Meliponini. The distribution of the microorganisms across the three bee species and their nest materials are described. Further analysis of yeast-derived volatiles is on-going as part of a larger project on the relationships between Australian Meliponini bees and yeast symbionts.