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The modes through which individuals disperse prior to reproduction has important consequences for gene flow in populations. In honey bees (Apis sp.), drones (males) reproduce within a short flight range of their natal nest, leaving and returning each afternoon within a narrow mating window. Drones are assumed to return to their natal nests as they depend on workers to feed them. However, in apiaries, drones are reported to regularly make navigation errors and return to a non-natal nest, where they are accepted and fed by unrelated workers. If such a “drone drift” occurred in wild populations, it could facilitate some further degree of dispersal for males, particularly if drones drift into host nests some distance away from their natal nest. Here, we investigated whether drone drift occurs in an invasive population of the Asian honey bee (Apis cerana). Based on the genotypes of 1462 drones from 19 colonies, we found only a single drone that could be considered a candidate drifter (~0.07%). In three other colonies, drones whose genotypes differed from the inferred queen were best explained by recent queen turnover or worker-laying. We concluded that drone drift in this population is low at best, and A. cerana drones either rarely make navigation errors in wild populations or are not accepted into foreign nests when they do so. We therefore confirm that drone dispersal distance is limited to the distance of daily drone flights from natal nests, a key assumption of both colony density estimates based on sampling of drone congregation areas and population genetic models of gene flow in honey bees.

Extracellular vesicles (EVs) are membranous structures containing bioactive molecules, secreted by most cells into the extracellular environment. EVs are classified by their biogenesis mechanisms into two major subtypes: ectosomes (enriched in large EVs; lEVs), budding directly from the plasma membrane, which is common in both prokaryotes and eukaryotes, and exosomes (enriched in small EVs; sEVs) generated through the multivesicular bodies via the endomembrane system, which is unique to eukaryotes. Even though recent proteomic analyses have identified key proteins associated with EV subtypes, there has been no systematic analysis, thus far, to support the general validity and utility of current EV subtype separation methods, still largely dependent on physical properties, such as vesicular size and sedimentation. Here, we classified human EV proteomic datasets into two main categories based on distinct centrifugation protocols commonly used for isolating sEV or lEV fractions. We found characteristic, evolutionarily conserved profiles of sEV and lEV proteins linked to their respective biogenetic origins. This may suggest that the evolutionary trajectory of vesicular proteins may result in a membership bias toward specific EV subtypes. Protein–protein interaction (PPI) network analysis showed that vesicular proteins formed distinct clusters with proteins in the same EV fraction, providing evidence for the existence of EV subtype-specific protein recruiters. Moreover, we identified functional modules enriched in each fraction, including multivesicular body sorting for sEV, and mitochondria cellular respiration for lEV proteins. Our analysis successfully captured novel features of EVs embedded in heterogeneous proteomics studies and suggests specific protein markers and signatures to be used as quality controllers in the isolation procedure for subtype-enriched EV fractions.

The subspecies of honeybee indigenous to the Cape region of South Africa, Apis mellifera capensis, is unique because a high proportion of unmated workers can lay eggs that develop into females via thelytokous parthenogenesis involving central fusion of meiotic products. This ability allows pseudoclonal lineages of workers to establish, which are presently widespread as reproductive parasites within the honeybee populations of South Africa. Successful long-term propagation of a parthenogen requires the maintenance of heterozygosity at the sex locus, which in honeybees must be heterozygous for the expression of female traits. Thus, in successful lineages of parasitic workers, recombination events are reduced by an order of magnitude relative to meiosis in queens of other honeybee subspecies. Here we show that in unmated A. m. capensis queens treated to induce oviposition, no such reduction in recombination occurs, indicating that thelytoky and reduced recombination are not controlled by the same gene. Our virgin queens were able to lay both arrhenotokous male-producing haploid eggs and thelytokous female-producing diploid eggs at the same time, with evidence that they have some voluntary control over which kind of egg was laid. If so, they are able to influence the kind of second-division meiosis that occurs in their eggs post partum.

In honey bees, drones disperse to specific locations in order to mate before returning to their natal nests. However, in apiaries of Apis mellifera, drones have been witnessed returning to non-natal nests; this behavior may also sometimes occur at natural nests. This behavior could be due to the unnaturally high density of nests in apiaries. In this study, we genotyped colonies of the Apis cerana honey bee collected in its invasive range of Far North Queensland, Australia, to determine whether there is evidence of drone drift between colonies. We found no such evidence of drone drift. Some drones that do not match the resident queen's genotype are instead likely due to queen turnover or worker reproduction. The modes through which individuals disperse prior to reproduction has important consequences for gene flow in populations. In honey bees (Apis sp.), drones (males) reproduce within a short flight range of their natal nest, leaving and returning each afternoon within a narrow mating window. Drones are assumed to return to their natal nests as they depend on workers to feed them. However, in apiaries, drones are reported to regularly make navigation errors and return to a non-natal nest, where they are accepted and fed by unrelated workers. If such a \"drone drift\" occurred in wild populations, it could facilitate some further degree of dispersal for males, particularly if drones drift into host nests some distance away from their natal nest. Here, we investigated whether drone drift occurs in an invasive population of the Asian honey bee (Apis cerana). Based on the genotypes of 1462 drones from 19 colonies, we found only a single drone that could be considered a candidate drifter (~0.07%). In three other colonies, drones whose genotypes differed from the inferred queen were best explained by recent queen turnover or worker-laying. We concluded that drone drift in this population is low at best, and A. cerana drones either rarely make navigation errors in wild populations or are not accepted into foreign nests when they do so. We therefore confirm that drone dispersal distance is limited to the distance of daily drone flights from natal nests, a key assumption of both colony density estimates based on sampling of drone congregation areas and population genetic models of gene flow in honey bees.

Honey bees ( Apis spp.) are important pollinators in many natural and agro-ecosystems across the world. Effective means of surveying wild populations are therefore key to their conservation and management. One available survey method infers honey bee colony density from the genotype of drones (males) sampled from sites known as Drone Congregation Areas (DCAs). While this approach has been commonly used for the Western honey bee ( A. mellifera ), its feasibility for other Apis species is unknown. Here, we investigate drone congregation behaviour in the Asian honey bee Apis cerana in north-east Australia and its suitability for inferring colony density. As this A. cerana population is invasive, surveys in this case can aid in monitoring the population’s growth and spread. Over 5 years, we identified 30 DCAs, many of which were stable across time. DCAs were sheltered areas beside tree-lines or openings in the forest canopy. A. cerana drones joined DCAs during 1–2-h afternoon intervals and could be sampled at heights of 2–24 m via adhesive lines attached to helium balloons carrying lures coated in queen pheromone. Drones were more likely to be present at a DCA as temperature increased, though abiotic factors did not predict overall drone abundance. Drones could be sampled in low numbers even where colony density was extremely low. Based on the genotyping and inferred sibship of drones sampled at DCAs between 2016 and 2021, we estimate population density in Australia’s A. cerana to be in the range 1.1–8.1 colonies/km 2 . This extrapolates to a total population size in the range 11,000–83,000 colonies, with more refined estimates requiring better knowledge of drone flight distance and the effect of habitat on colony density. We conclude that population surveys based on drones from DCAs are possible for A. cerana and propose that this approach be part of a toolkit of methods used to monitor Asian honey bee populations in both their native and invasive ranges.

AbstractHoney bees (Apis spp.) are important pollinators in many natural and agro-ecosystems across the world. Effective means of surveying wild populations are therefore key to their conservation and management. One available survey method infers honey bee colony density from the genotype of drones (males) sampled from sites known as Drone Congregation Areas (DCAs). While this approach has been commonly used for the Western honey bee (A. mellifera), its feasibility for other Apis species is unknown. Here, we investigate drone congregation behaviour in the Asian honey bee Apis cerana in north-east Australia and its suitability for inferring colony density. As this A. cerana population is invasive, surveys in this case can aid in monitoring the population’s growth and spread. Over 5 years, we identified 30 DCAs, many of which were stable across time. DCAs were sheltered areas beside tree-lines or openings in the forest canopy. A. cerana drones joined DCAs during 1–2-h afternoon intervals and could be sampled at heights of 2–24 m via adhesive lines attached to helium balloons carrying lures coated in queen pheromone. Drones were more likely to be present at a DCA as temperature increased, though abiotic factors did not predict overall drone abundance. Drones could be sampled in low numbers even where colony density was extremely low. Based on the genotyping and inferred sibship of drones sampled at DCAs between 2016 and 2021, we estimate population density in Australia’s A. cerana to be in the range 1.1–8.1 colonies/km2. This extrapolates to a total population size in the range 11,000–83,000 colonies, with more refined estimates requiring better knowledge of drone flight distance and the effect of habitat on colony density. We conclude that population surveys based on drones from DCAs are possible for A. cerana and propose that this approach be part of a toolkit of methods used to monitor Asian honey bee populations in both their native and invasive ranges.

Human-induced shifts in species’ ranges can increase contact between closely related species and lead to reproductive interference. In Australia, climate change and trade in stingless bee colonies is increasing the range overlap of two cryptic species: Tetragonula carbonaria and T. hockingsi. To investigate reproductive interactions between these species, we validated a diagnostic-PCR test based on the mitochondrial gene COI to ID field specimens to species. We then assessed the likelihood of reproductive interference in four ways. First, we imaged the male genitalia of each species and found no evidence of reproductive character displacement. Second, we assessed species composition of mating aggregations in an area of sympatry (Southeast Queensland) and confirmed that some males join the mating aggregations of interspecific colonies. Third, we translocated T. hockingsi colonies into the southern range of T. carbonaria (Sydney) and tracked their ability to requeen. These translocated colonies attracted mating aggregations comprised almost entirely of interspecific males, but never formed hybrid colonies; instead, queens either mated with their brothers, or the colony failed to requeen at all. Finally, we presented T. carbonaria males with either conspecific or interspecific virgin queens and found that males attempted to mate only with their own species’ queens. In all, we conclude that reproductive barriers between these species are complete with respect to “short-range” mating cues, but not for “long-range” mate attraction cues. Our study highlights that hive movements can increase some forms of pre-mating reproductive interference between managed bee species, even where the species do not actually mate or hybridize.

Honey bee ( Apis  mellifera ) breeding has intensified as populations experience increasing stress and pollination demand increases. Breeding programmes risk genetic diversity losses as mating is focused on a small group of individuals. Loss of diversity at the complementary sex determiner ( csd ) locus results in decreased viability and reduced honey production. Bees that are homozygous at csd become inviable males rather than workers. We examined csd diversity in four Australian breeding populations: a queen bee breeder from New South Wales, another from Queensland, a Western Australian breeding programme involving 11 bee breeders, and a research population at the NSW Department of Primary Industries. We found 82 unique csd alleles across the four populations, 16 of which have not been previously reported. This study provides a snapshot of csd diversity in Australia which will be useful for the national honey bee genetic improvement programme (Plan Bee).

Generating reference maps of interactome networks illuminates genetic studies by providing a protein-centric approach to finding new components of existing pathways, complexes, and processes. We apply state-of-the-art methods to identify binary protein-protein interactions (PPIs) for Drosophila melanogaster . Four all-by-all yeast two-hybrid (Y2H) screens of > 10,000 Drosophila proteins result in the ‘FlyBi’ dataset of 8723 PPIs among 2939 proteins. Testing subsets of data from FlyBi and previous PPI studies using an orthogonal assay allows for normalization of data quality; subsequent integration of FlyBi and previous data results in an expanded binary Drosophila reference interaction network, DroRI, comprising 17,232 interactions among 6511 proteins. We use FlyBi data to generate an autophagy network, then validate in vivo using autophagy-related assays. The deformed wings ( dwg ) gene encodes a protein that is both a regulator and a target of autophagy. Altogether, these resources provide a foundation for building new hypotheses regarding protein networks and function. Maps of protein-protein interactions (PPIs) help identify new components of pathways, complexes, and processes. In this work, state-of-the-art methods are used to identify binary Drosophila PPIs, generating broadly useful physical and data resources.

Honey bee ( Apis  mellifera ) breeding has intensified as populations experience increasing stress and pollination demand increases. Breeding programmes risk genetic diversity losses as mating is focused on a small group of individuals. Loss of diversity at the complementary sex determiner ( csd ) locus results in decreased viability and reduced honey production. Bees that are homozygous at csd become inviable males rather than workers. We examined csd diversity in four Australian breeding populations: a queen bee breeder from New South Wales, another from Queensland, a Western Australian breeding programme involving 11 bee breeders, and a research population at the NSW Department of Primary Industries. We found 82 unique csd alleles across the four populations, 16 of which have not been previously reported. This study provides a snapshot of csd diversity in Australia which will be useful for the national honey bee genetic improvement programme (Plan Bee).