Thinking beyond Western commercial honeybee hives: towards improved conservation of honey bee diversity

A decline of wild pollinators, along with a decline of bee diversity, has been a cause of concern among academics and governmental organizations. According to IPBES, a lack of wild pollinator data contributes to difficulties in comprehensively analyzing the regional status of wild pollinators in Africa, Latin America, Asia and Oceania. It may have also contributed to the prevailing lack of awareness of the diversity of honey bees, of which the managed Apis mellifera is often considered as “the (only) honey bee,” despite the fact that there are eight other honey bee species extant in Asia. A survey of 100 journal articles published in 2016 shows that 57% of the studies still identified A. mellifera as “the honey bee.” In total, 80% of studies were conducted solely on A. mellifera. This focus on A. mellifera has also caused the honey standard of Codex Alimentarius and the European Union to be based solely on A. mellifera, causing improper evaluation of honeys from other species. We recommend adapting current standards to reflect the diversity of honey bees and in the process correct failures in the honey market and pave the way towards improved protection of honey bee species and their habitats.     

Pollen diets and niche overlap of honey bees and native bees in protected areas

The decline of both managed and wild bee populations has been extensively reported for over a decade now, with growing concerns amongst the scientific community. Also, evidence is growing that both managed and feral honey bees may exacerbate threats to wild bees. In Australia, there are over 1600 native bee species and introduced European honey bees (Apis mellifera) have established throughout most landscapes. There is a major gap in knowledge of the interactions between honey bees and native bees in Australian landscapes, especially floral resource use.Here we report on the pollen diets of wild bees in protected areas of coastal heathland, an ecosystem characterised by mass flowering in late winter and spring. We sampled bees within three sites and DNA metabarcoding was used to compare the pollen diets of honey bees and native bees. We recorded 2, 772 bees in total, with 13 genera and 18 described species identified. Apis mellifera was the most common species across all locations, accounting for 42% of all bees collected. Native bee genera included eusocial Tetragonula (stingless bees) (37%), and semi-social Exoneura and Braunsapis (19.8% combined). Metabarcoding data revealed both Tetragonula and honey bees have wide foraging patterns, and the bipartite network overall was highly generalised (H₂’ = 0.24). Individual honey bees carried pollen of 7–29 plant species, and significantly more species than all other bees. We found niche overlap in the diets of honey bees and native bees generally (0.42), and strongest overlap with stingless bees (0.70) and species of Braunsapis (0.62). A surprising finding was that many species carried pollen from Restionaceae and Cyperaceae, families generally considered to be predominantly wind-pollinated in Australia. Our study showed introduced honey bee use of resources overlaps with that of native bees in protected heathlands, but there are clear differences in their diet preferences.     

Local extinction of a rare plant pollinator in Southern Utah (USA) associated with invasion by Africanized honey bees

Twenty-five years ago, Arctomecon humilis, a pollinator-dependent, endangered poppy globally restricted to the extreme northeastern Mojave Desert in southwestern Utah, was pollinated by native bee species and the European honey bee. Follow-up studies beginning in 2012 failed to find the two most important native bee pollinator species, one of which, Perdita meconis, is a strict poppy specialist. We had four objectives: (1) confirm the status of formerly important native bee pollinators; (2) determine the role of the Africanized honey bee which reportedly invaded southern Utah in 2008; (3) examine the effect of the ostensible change in pollinator fauna on fruit set in four populations; (4) describe the pollination proficiency of species that presently visit poppy flowers. For the fourth consecutive survey, P. meconis was absent; its local extinction in Utah now seems certain. Another previously important native pollinator, Eucera quadricincta, was very rare. Also uncommon was the European honey bee, having been largely replaced by Africanized honey bees which have become, in most populations, the prevalent pollinator. Africanized bees forage early in the day and quickly strip flowers of their copious pollen leaving little for native bees. We argue that the invasion of southern Utah by Africanized bees is the most likely cause of the severe disruption of the A. humilis pollination system. The ascension of the Africanized bee is also associated with reduced fruit set in all poppy populations, especially those where plants are sparse. Arctomecon humilis now appears to depend mostly on an invasive species for pollination.     

Imidacloprid Alters Foraging and Decreases Bee Avoidance of Predators

Concern is growing over the effects of neonicotinoid pesticides, which can impair honey bee cognition. We provide the first demonstration that sublethal concentrations of imidacloprid can harm honey bee decision-making about danger by significantly increasing the probability of a bee visiting a dangerous food source. Apis cerana is a native bee that is an important pollinator of agricultural crops and native plants in Asia. When foraging on nectar containing 40 µg/L (34 ppb) imidacloprid, honey bees (Apis cerana) showed no aversion to a feeder with a hornet predator, and 1.8 fold more bees chose the dangerous feeder as compared to control bees. Control bees exhibited significant predator avoidance. We also give the first evidence that foraging by A. cerana workers can be inhibited by sublethal concentrations of the pesticide, imidacloprid, which is widely used in Asia. Compared to bees collecting uncontaminated nectar, 23% fewer foragers returned to collect the nectar with 40 µg/L imidacloprid. Bees that did return respectively collected 46% and 63% less nectar containing 20 µg/L and 40 µg/L imidacloprid. These results suggest that the effects of neonicotinoids on honey bee decision-making and other advanced cognitive functions should be explored. Moreover, research should extend beyond the classic model, the European honey bee (A. mellifera), to other important bee species.     

The genomic basis of adaptation to high‐altitude habitats in the eastern honey bee (Apis cerana)

The eastern honey bee (Apis cerana) is of central importance for agriculture in Asia. It has adapted to a wide variety of environmental conditions across its native range in southern and eastern Asia, which includes high‐altitude regions. eastern honey bees inhabiting mountains differ morphologically from neighbouring lowland populations and may also exhibit differences in physiology and behaviour. We compared the genomes of 60 eastern honey bees collected from high and low altitudes in Yunnan and Gansu provinces, China, to infer their evolutionary history and to identify candidate genes that may underlie adaptation to high altitude. Using a combination of F_ST‐based statistics, long‐range haplotype tests and population branch statistics, we identified several regions of the genome that appear to have been under positive selection. These candidate regions were strongly enriched for coding sequences and had high haplotype homozygosity and increased divergence specifically in highland bee populations, suggesting they have been subjected to recent selection in high‐altitude habitats. Candidate loci in these genomic regions included genes related to reproduction and feeding behaviour in honey bees. Functional investigation of these candidate loci is necessary to fully understand the mechanisms of adaptation to high‐altitude habitats in the eastern honey bee.     

Honey Bees Modulate Their Olfactory Learning in the Presence of Hornet Predators and Alarm Component

In Southeast Asia the native honey bee species Apis cerana is often attacked by hornets (Vespa velutina), mainly in the period from April to November. During the co-evolution of these two species honey bees have developed several strategies to defend themselves such as learning the odors of hornets and releasing alarm components to inform other mates. However, so far little is known about whether and how honey bees modulate their olfactory learning in the presence of the hornet predator and alarm components of honey bee itself. In the present study, we test for associative olfactory learning of A. cerana in the presence of predator odors, the alarm pheromone component isopentyl acetate (IPA), or a floral odor (hexanal) as a control. The results show that bees can detect live hornet odors, that there is almost no association between the innately aversive hornet odor and the appetitive stimulus sucrose, and that IPA is less well associated with an appetitive stimulus when compared with a floral odor. In order to imitate natural conditions, e.g. when bees are foraging on flowers and a predator shows up, or alarm pheromone is released by a captured mate, we tested combinations of the hornet odor and floral odor, or IPA and floral odor. Both of these combinations led to reduced learning scores. This study aims to contribute to a better understanding of the prey-predator system between A. cerana and V. velutina.     

Honey bee colony collapse disorder is possibly caused by a dietary pyrethrum deficiency

Industrialized farming relies on bee keepers transporting hives to the vicinity of large areas of mono-crops for crop pollination. Hives are typically moved multiple times per growing season to satisfy the pollination need. A phenomenon wherein colonies of honey bees collapse in large numbers has been threatening crops in North America. Honey bees are hosts to at least two pathogenic mites; Varroa destructor and Acarapis woodi (a tracheal mite). Pyrethrums are a group of flowering plants which include Chrysanthemum coccineum, Chrysanthemum cinerariifolium, Chrysanthemum marschallii, and related species. These plants produce potent insecticides, also named pyrethrums, which are powerful mite toxins. We believe that a honey bee dietary deficiency of pyrethrums and other micro-nutrients from pyrethrum producing plants allows parasitic mites to either kill the honey bees directly or reduce honey bee resistance to other pathogens. Intermittent feeding of honey bees on pyrethrum producing plants might reverse or prevent colony collapse disorder.     

Nosema ceranae Escapes Fumagillin Control in Honey Bees

Fumagillin is the only antibiotic approved for control of nosema disease in honey bees and has been extensively used in United States apiculture for more than 50 years for control of Nosema apis. It is toxic to mammals and must be applied seasonally and with caution to avoid residues in honey. Fumagillin degrades or is diluted in hives over the foraging season, exposing bees and the microsporidia to declining concentrations of the drug. We showed that spore production by Nosema ceranae, an emerging microsporidian pathogen in honey bees, increased in response to declining fumagillin concentrations, up to 100% higher than that of infected bees that have not been exposed to fumagillin. N. apis spore production was also higher, although not significantly so. Fumagillin inhibits the enzyme methionine aminopeptidase2 (MetAP2) in eukaryotic cells and interferes with protein modifications necessary for normal cell function. We sequenced the MetAP2 gene for apid Nosema species and determined that, although susceptibility to fumagillin differs among species, there are no apparent differences in fumagillin binding sites. Protein assays of uninfected bees showed that fumagillin altered structural and metabolic proteins in honey bee midgut tissues at concentrations that do not suppress microsporidia reproduction. The microsporidia, particularly N. ceranae, are apparently released from the suppressive effects of fumagillin at concentrations that continue to impact honey bee physiology. The current application protocol for fumagillin may exacerbate N. ceranae infection rather than suppress it.     

Resource overlap and possible competition between honey bees and wild bees in central Europe

Evidence for interspecific competition between honey bees and wild bees was studied on 15 calcareous grasslands with respect to: (1) foraging radius of honey bees, (2) overlap in resource use, and (3) possible honey bee effects on species richness and abundance of flower-visiting, ground-nesting and trap-nesting wild bees. The grasslands greatly differed in the number of honey bee colonies within a radius of 2 km and were surrounded by agricultural habitats. The number of flower-visiting honey bees on both potted mustard plants and small grassland patches declined with increasing distance from the nearest apiary and was almost zero at a distance of 1.5–2.0 km. Wild bees were observed visiting 57 plant species, whereas honey bees visited only 24 plant species. Percentage resource overlap between honey bees and wild bees was 45.5%, and Hurlbert’s index of niche overlap was 3.1. In total, 1849 wild bees from 98 species were recorded on the calcareous grasslands. Neither species richness nor abundance of wild bees were negatively correlated with the density of honey bee colonies (within a radius of 2 km) or the density of flower-visiting honey bees per site. Abundance of flower- visiting wild bees was correlated only with the percentage cover of flowering plants. In 240 trap nests, 1292 bee nests with 6066 brood cells were found. Neither the number of bee species nor the number of brood cells per grassland was significantly correlated with the density of honey bees. Significant correlations were found only between the number of brood cells and the percentage cover of shrubs. The number of nest entrances of ground-nesting bees per square metre was not correlated with the density of honey bees but was negatively correlated with the cover of vegetation. Interspecific competition by honey bees for food resources was not shown to be a significant factor determining abundance and species richness of wild bees. Received: 22 March 1999 / Accepted: 24 September 1999     

Red mason bees cannot compete with honey bees for floral resources in a cage experiment

Intensive beekeeping to mitigate crop pollination deficits and habitat loss may cause interspecific competition between bees. Studies show negative correlations between flower visitation of honey bees (Apis mellifera) and wild bees, but effects on the reproduction of wild bees were not proven. Likely reasons are that honey bees can hardly be excluded from controls and wild bee nests are generally difficult to detect in field experiments. The goal of this study was to investigate whether red mason bees (Osmia bicornis) compete with honey bees in cages in order to compare the reproduction of red mason bees under different honey bee densities. Three treatments were applied, each replicated in four cages of 18 m³ with 38 red mason bees in all treatments and 0, 100, and 300 honey bees per treatment with 10–20% being foragers. Within the cages, the flower visitation and interspecific displacements from flowers were observed. Niche breadths and resource overlaps of both bee species were calculated, and the reproduction of red mason bees was measured. Red mason bees visited fewer flowers when honey bees were present. Niche breadth of red mason bees decreased with increasing honey bee density while resource overlaps remained constant. The reproduction of red mason bees decreased in cages with honey bees. In conclusion, our experimental results show that in small and isolated flower patches, wild bees can temporarily suffer from competition with honey bees. Further research should aim to test for competition on small and isolated flower patches in real landscapes.     

Rural avenues as a refuge for feral honey bee population

Several honey bee (Apis mellifera) subspecies are in danger of local extinction because their feral population have almost completely disappeared. An important threat to the feral populations of bees is loss of habitat and loss of woodlands. In many places the only habitat suitable for honey bee nesting are rows of trees along roadsides. We studied a feral population of honey bees inhabiting avenues in northern Poland. We inspected 142 km of avenues and found 45 feral colonies. The estimated density of feral population inhabiting the avenues was 0.10 nest km^?2. Honey bees preferred to build their nests in trees with a thick trunk and a somewhat weak state of health. There was no strong preference of bees to any species of trees. We stress the importance of protection of existing avenues and creating new ones. This can provide suitable habitat not only for honey bees but also for other endangered species.     

Using DNA Metabarcoding to Identify the Floral Composition of Honey: A New Tool for Investigating Honey Bee Foraging Preferences

Identifying the floral composition of honey provides a method for investigating the plants that honey bees visit. We compared melissopalynology, where pollen grains retrieved from honey are identified morphologically, with a DNA metabarcoding approach using the rbcL DNA barcode marker and 454-pyrosequencing. We compared nine honeys supplied by beekeepers in the UK. DNA metabarcoding and melissopalynology were able to detect the most abundant floral components of honey. There was 92% correspondence for the plant taxa that had an abundance of over 20%. However, the level of similarity when all taxa were compared was lower, ranging from 22–45%, and there was little correspondence between the relative abundance of taxa found using the two techniques. DNA metabarcoding provided much greater repeatability, with a 64% taxa match compared to 28% with melissopalynology. DNA metabarcoding has the advantage over melissopalynology in that it does not require a high level of taxonomic expertise, a greater sample size can be screened and it provides greater resolution for some plant families. However, it does not provide a quantitative approach and pollen present in low levels are less likely to be detected. We investigated the plants that were frequently used by honey bees by examining the results obtained from both techniques. Plants with a broad taxonomic range were detected, covering 46 families and 25 orders, but a relatively small number of plants were consistently seen across multiple honey samples. Frequently found herbaceous species were Rubus fruticosus, Filipendula ulmaria, Taraxacum officinale, Trifolium spp., Brassica spp. and the non-native, invasive, Impatiens glandulifera. Tree pollen was frequently seen belonging to Castanea sativa, Crataegus monogyna and species of Malus, Salix and Quercus. We conclude that although honey bees are considered to be supergeneralists in their foraging choices, there are certain key species or plant groups that are particularly important in the honey bees environment. The reasons for this require further investigation in order to better understand honey bee nutritional requirements. DNA metabarcoding can be easily and widely used to investigate floral visitation in honey bees and can be adapted for use with other insects. It provides a starting point for investigating how we can better provide for the insects that we rely upon for pollination.     

Twenty-five-year study of Nosema spp. in honey bees (Apis mellifera) in Serbia

A total of 7386 samples of adult honey bees from different areas of Serbia (fifteen regions and 79 municipalities) were selected for light microscopy analysis for Nosema species during 1992–2017. A selection of honey bee samples from colonies positive for microsporidian spores during 2009–2011, 2015 and 2017 were then subjected to molecular diagnosis by multiplex PCR using specific primers for a region of the 16S rRNA gene of Nosema species. The prevalence of microsporidian spore-positive bee colonies ranged between 14.4% in 2013 and 65.4% in 1992. PCR results show that Nosema ceranae is not the only Nosema species to infect honey bees in Serbia. Mixed N. apis/N. ceranae infections were detected in the two honey bee samples examined by mPCR during 2017. The beekeeping management of disease prevention, such as replacement of combs and queens and hygienic handling of colonies are useful in the prevention of Nosema infection.     

Foraging behavior affects pollen removal and deposition in Impatiens capensis (Balsaminaceae)

Flowers of most plant species are visited by a variety of animals. Some of these visitors are effective pollinators while others remove resources without transferring pollen. Studies comparing the effectiveness of different visitors as pollinators often compare taxa without considering variation in behavior within a taxon. Wilson and Thomson (Ecology 72: 1503-1507, 1991) documented the effects of honey bees and bumble bees on the pollination dynamics of Impatiens capensis. They found that pollen-collecting honey bees removed large numbers of pollen grains from anthers but deposited little of it on stigmas; bumble bees, which sought nectar, removed less pollen but deposited more of it on stigmas. It is unclear whether the low pollen transfer efficiencies of honey bees are explained by their morphology or by their pollen-collecting behavior. We repeated the work of Wilson and Thomson at a site where honey bees were foraging for nectar, not pollen. We measured the quantity of pollen remaining in anthers, the number of pollen grains deposited on stigmas, and seed production after single visits by honey bees and bumble bees. The differences between the taxa disappeared when they were foraging in a similar manner. Our results clearly demonstrate the importance of foraging behavior on the pollination effectiveness of floral visitors.     

Orchard pollination in Capitol Reef National Park,Utah, USA. Honey bees or native bees?

Capitol Reef National Park in central Utah, USA surrounds 22 managed fruit orchards started over a century ago by Mormon pioneers. Honey bees are imported for pollination, although the area in which the Park is embedded has over 700 species of native bees, many of which are potential orchard pollinators. We studied the visitation of native bees to apple, pear, apricot, and sweet cherry over 2 years. Thirty species of bees visited the flowers but, except for pear flowers, most were uncommon compared to honey bees. Evidence that honey bees prevented native bees from foraging on orchard crop flowers was equivocal: generally, honey bee and native bee visitation rates to the flowers were not negatively correlated, nor were native bee visitation rates positively correlated with distance of orchards from honey bee hives. Conversely, competition was tentatively suggested by much larger numbers of honey bees than natives on the flowers of apples, apricots and cherry; and by the large increase of native bees on pears, where honey bee numbers were low. At least one-third of the native bee species visiting the flowers are potential pollinators, including cavity-nesting species such as Osmia lignaria propinqua, currently managed for small orchard pollination in the US, plus several fossorial species, including one rosaceous flower specialist (Andrena milwaukiensis). We suggest that gradual withdrawal of honey bees from the Park would help conserve native bee populations without decreasing orchard crop productivity, and would serve as a demonstration of the commercial value of native pollinators.     

RNAi-mediated Double Gene Knockdown and Gustatory Perception Measurement in Honey Bees (Apis mellifera)

This video demonstrates novel techniques of RNA interference (RNAi) which downregulate two genes simultaneously in honey bees using double-stranded RNA (dsRNA) injections. It also presents a protocol of proboscis extension response (PER) assay for measuring gustatory perception.RNAi-mediated gene knockdown is an effective technique downregulating target gene expression. This technique is usually used for single gene manipulation, but it has limitations to detect interactions and joint effects between genes. In the first part of this video, we present two strategies to simultaneously knock down two genes (called double gene knockdown). We show both strategies are able to effectively suppress two genes, vitellogenin (vg) and ultraspiracle (usp), which are in a regulatory feedback loop. This double gene knockdown approach can be used to dissect interrelationships between genes and can be readily applied in different insect species.The second part of this video is a demonstration of proboscis extension response (PER) assay in honey bees after the treatment of double gene knockdown. The PER assay is a standard test for measuring gustatory perception in honey bees, which is a key predictor for how fast a honey bee''s behavioral maturation is. Greater gustatory perception of nest bees indicates increased behavioral development which is often associated with an earlier age at onset of foraging and foraging specialization in pollen. In addition, PER assay can be applied to identify metabolic states of satiation or hunger in honey bees. Finally, PER assay combined with pairing different odor stimuli for conditioning the bees is also widely used for learning and memory studies in honey bees.     

Mitochondrial DNA and AFLP markers support species status of <Emphasis Type="Italic">Apis nigrocincta</Emphasis>

Summary. New investigations into the biogeography of Asian honey bees are bringing to light previously unrecognized species. On the Indonesian island of Sulawesi, morphometrically distinct populations of Asian cavity-nesting honey bees occur in close sympatry. One population is the widespread species Apis cerana; the other has been proposed as a separate species, A. nigrocincta. A non-coding region of the mitochondrial genome, located between the leucine tRNA and cytochrome oxidase II genes was sequenced in bees from each colony sampled. Consistent sequence differences were noted between the two putative species. Analysis of Molecular Variance (AMOVA) of Amplified Fragment Length Polymorphisms (AFLPs) in each population indicate little or no gene flow between the two, supporting the status of A. nigrocincta as a separate species.     

The Potential Influence of Bumble Bee Visitation on Foraging Behaviors and Assemblages of Honey Bees on Squash Flowers in Highland Agricultural Ecosystems

Bee species interactions can benefit plant pollination through synergistic effects and complementary effects, or can be of detriment to plant pollination through competition effects by reducing visitation by effective pollinators. Since specific bee interactions influence the foraging performance of bees on flowers, they also act as drivers to regulate the assemblage of flower visitors. We selected squash (Cucurbita pepo L.) and its pollinators as a model system to study the foraging response of honey bees to the occurrence of bumble bees at two types of sites surrounded by a high amount of natural habitats (≥ 58% of land cover) and a low amount of natural habitats (≤ 12% of land cover) in a highland agricultural ecosystem in China. At the individual level, we measured the elapsed time from the departure of prior pollinator(s) to the arrival of another pollinator, the selection of honey bees for flowers occupied by bumble bees, and the length of time used by honey bees to explore floral resources at the two types of sites. At the community level, we explored the effect of bumble bee visitation on the distribution patterns of honey bees on squash flowers. Conclusively, bumble bee visitation caused an increase in elapsed time before flowers were visited again by a honey bee, a behavioral avoidance by a newly-arriving honey bee to select flowers occupied by bumble bees, and a shortened length of time the honey bee takes to examine and collect floral resources. The number of overall bumble bees on squash flowers was the most important factor explaining the difference in the distribution patterns of honey bees at the community level. Furthermore, decline in the number of overall bumble bees on the squash flowers resulted in an increase in the number of overall honey bees. Therefore, our study suggests that bee interactions provide an opportunity to enhance the resilience of ecosystem pollination services against the decline in pollinator diversity.     

Fungi isolated from honey bees, Apis mellifera, fed 2,4-D and Antibiotics

Eighteen species of fungi were isolated and identified from the intestines of 388 honey bees, Apis mellifera. Bees fed a combination of oxytetracycline and fumagillin contained fewer fungi than control bees or bees fed 2,4-D. New records of fungi associated with honey bees include Alternaria tenuissima, Cladosporium cladosporoides, Bipolaris sp., Curvularia brachyospora, Penicillium ochro-chloron, Penicillium urticae, and Rhizopus arrhizus.