Tests of hybridisation in Tetragonula stingless bees using multiple genetic markers

Discrepancies in mitochondrial and nuclear genetic data are often interpreted as evidence of hybridisation. We re-examined reports of hybridisation in three cryptic stingless bee species in the genus in South East Queensland, Australia (, , and ). Previous studies on this group using microsatellites proposed that rare hybrids do occur. In contrast, we find that allele frequencies at neutral regions of the nuclear genome (both microsatellites and random ) reliably separated the three species and revealed no evidence of hybridisation. We found no inter-species variation in the nuclear gene , but low and moderate species-specific polymorphisms in the nuclear gene and the mitochondrial respectively, with no cases of mito-nuclear discord at these genes. We confirm that nuclear divergence between these species is low, based on 10-26kb of non-coding sequence flanking and (0.7-1% pairwise difference between species). However, we find mitogenomes to be far more diverged than nuclear genomes (21.6-23.6% pairwise difference between species). Based on these comprehensive analyses of multiple marker types, we conclude that there is no ongoing gene flow in the species of South East Queensland, despite their high morphological similarity and low nuclear divergence. The mitochondrial genomes indicate that divergence could be deep between these lineages but the low divergence in the nuclear genomes means that further investigation is required before divergence times can be estimated accurately. The mitogenomes, and draft nuclear genomes provided for these species will be a resource for further molecular studies on this group.