Alexandros Papachristoforou

Defensive behaviour of Apis mellifera against Vespa velutina in France: Testing whether European honeybees can develop an effective collective defence against a new predator

We investigated the prey-predator interactions between the European honeybee, Apis mellifera, and the invasive yellow-legged hornet, Vespa velutina, which first invaded France in 2004 and thereafter spread to neighbouring European countries (Spain, Portugal and Italy). Our goal was to determine how successfully honeybees are able to defend their colonies against their new predator in Europe. Experiments were conducted in the southwest of France-the point of entry of the hornet in Europe-under natural and semi-controlled field conditions. We investigated a total of eight apiaries and 95 colonies subjected to either low or high levels of predation. We analyzed hornet predatory behaviour and collective response of colonies under attack. The results showed that A. mellifera in France exhibit an inefficient and unorganized defence against V. velutina, unlike in other regions of Europe and other areas around the globe where honeybees have co-evolved with their natural Vespa predators.

The bite of the honeybee: 2-heptanone secreted from honeybee mandibles during a bite acts as a local anaesthetic in insects and mammals.

Honeybees secrete 2-heptanone (2-H) from their mandibular glands when they bite. Researchers have identified several possible functions: 2-H could act as an alarm pheromone to recruit guards and soldiers, it could act as a chemical marker, or it could have some other function. The actual role of 2-H in honeybee behaviour remains unresolved. In this study, we show that 2-H acts as an anaesthetic in small arthropods, such as wax moth larva (WML) and Varroa mites, which are paralysed after a honeybee bite. We demonstrated that honeybee mandibles can penetrate the cuticle of WML, introducing less than one nanolitre of 2-H into the WML open circulatory system and causing instantaneous anaesthetization that lasts for a few minutes. The first indication that 2-H acts as a local anaesthetic was that its effect on larval response, inhibition and recovery is very similar to that of lidocaine. We compared the inhibitory effects of 2-H and lidocaine on voltage-gated sodium channels. Although both compounds blocked the hNav1.6 and hNav1.2 channels, lidocaine was slightly more effective, 2.82 times, on hNav.6. In contrast, when the two compounds were tested using an ex vivo preparation–the isolated rat sciatic nerve–the function of the two compounds was so similar that we were able to definitively classify 2-H as a local anaesthetic. Using the same method, we showed that 2-H has the fastest inhibitory effect of all alkyl-ketones tested, including the isomers 3- and 4-heptanone. This suggests that natural selection may have favoured 2-H over other, similar compounds because of the associated fitness advantages it confers. Our results reveal a previously unknown role of 2-H in honeybee defensive behaviour and due to its minor neurotoxicity show potential for developing a new local anaesthetic from a natural product, which could be used in human and veterinary medicine.

Attack or retreat: Contrasted defensive tactics used by Cyprian honeybee colonies under attack from hornets

This study describes the tactics used by Cyprian honeybees (Apis mellifera cypria) to defend their colonies against hornet (Vespa orientalis orientalis) attacks. We use simulated hornet attacks and a combination of video recordings and image analysis to reveal, for the first time, contrasted intra-subspecies defensive tactics that operate at the colony level during predation. In some colonies, when attacked, the numbers of guards at the hive entrance increases rapidly to attack, engulf, and kill invading hornets. In other colonies, guards avoid conflicts with hornets by retreating gradually and by forming a defensive line of honeybees at the hive entrance. Retreater colonies have propolis walls at the hive entrances with small apertures that are too narrow to allow the hornet to access the hive and that therefore reinforces entrance protection. On the contrary, attacker colonies have propolis walls with large openings through which the hornet can pass; these bees block the hornets access by intensively guarding the hive entrance. We experimentally destroy propolis walls to test whether colonies consistently rebuild walls with the same intrinsic characteristics and we also monitor the survival rate of each anti-predator tactic after massive natural predation by hornets.

High frequency sounds produced by Cyprian honeybees Apis mellifera cypria when confronting their predator, the Oriental hornet Vespa orientalis

Honeybees face several predators and their ability to express collective defence behaviour is one of their major life traits that promote colony survival. We discovered that, while confronting attacks by the Oriental hornet Vespa orientalis, Apis mellifera cypria honeybees engage in a distinct acoustic behaviour: they produce a characteristic hissing sound of unexpectedly high frequency. When recording and analysing these hissing sounds during an extended sample of artificial attacks by hornets, we found that honeybees can produce sounds covering a wide frequency spectrum with a dominant frequency around 6 kHz. Notably, these acoustic emissions are distinct from the background noise of neighbouring flying bees. These results provide a detailed description of the sounds generated by A. m. cypria when defending their nest against hornets, and they could be used for future research to better understand the biological function of the acoustic behaviour in honeybees’ colony defence.ZusammenfassungWährend der Abwehr von Angriffen durch die orientalische Hornisse Vespa orientalis zeigen die zyprischen Honigbienen Apis mellifera cypria ein spezifisches akustisches Verhalten, bei dem sie einen charakteristischen Zischlaut von unerwartet hoher Frequenz erzeugen. Dieser charakteristische Ton wird während der Annäherung des Räubers und während der Auseinandersetzung mit verteidigenden Bienen am Eingang der Kolonie hervorgebracht. Wir nahmen diese Zischlaute während einer ausgedehnten Serie künstlicher Angriffe von Hornissen auf und analysierten sie. Für die Aufnahmen verwendeten wir ein mit einem Marantz PMD 670 Digitalrekorder verbundenes Sennheiser ME64 Mikrofon. Die Aufnahmen wurden unter Verwendung der Avisoft und Seewave Software untersucht. Wir ermittelten, dass die Bienen ein weites Frequenzspektrum mit einer dominanten Frequenz um 6 kHz erzeugen können. Die Zischlaute zeigten ein bis zu 16–17 kHz ausgedehntes Spektrum, das bis zu 30–35 harmonische Bänder einschloss. Die Untersuchung von 93 an 9 Kolonien aufgenommenen verschiedenen Zischlauten ergab keine Hinweise auf akustische Kennzeichen der unterschiedlichen Kolonien. Insbesondere waren die akustischen Emissionen deutlich von den durch benachbarte fliegende Bienen hervorgebrachten Hintergrundgeräuschen abgesetzt. Diese Ergebnisse beinhalten eine detaillierte Beschreibung der von A. m. cypria bei der Verteidigung gegen Hornissen hervorgebrachten Töne und können in zukünftigen Untersuchungen zu einem besseren Verständnis der biologischen Bedeutung des akustischen Verhaltens bei der Nestverteidigung der Honigbienen beitragen.

Genetic characterization of the cyprian honey bee (Apis mellifera cypria) based on microsatell ites and mitochondrial DNA polymorphisms

Abstract Honey bee populations from the island of Cyprus were analyzed using microsatellite and mitochondrial DNA markers. A total of 268 colonies were sampled in Cyprus, at six different locations-Kyrenia, Katydata, Flassou, Alabra, Troulloi, and Alassa-covering a wide area of the island. Results showed that the Cyprian honey bee Apis mellifera cypria could be distinguished from other Apis mellifera subspecies based on a „double pattern“ of mitochondrial DNA belonging to the C1 lineage and microsatellite DNA belonging to the O lineage. All populations were homogeneous, except the population from Kyrenia, probably due to the introduction of queens or colonies belonging to the C2, C6, and M7 lineages.

Attraction and direct establishment of primary and secondary honey bee swarms using swarm-tissue sachets

Summary In an effort to attract swarms within an apiary and to induce their direct establishment in hive-traps, we investigated the use of a new honey bee swarm attractant in the form of a small sachet containing a wet-wipe tissue, including geranic acid, citral, geraniol and rose oil as attractants. When swarm-tissues sachets were placed in hive traps at a distance of either 5 or 30m from the colonies, 60% of the swarms were established directly and permanently inside hive traps. All established swarms were secondary, headed by virgin queen(s). No primary swarm was attracted. When swarm-tissue sachets were placed in hive traps and were also hung in nearby tree branches, 90.9% of the swarms were attracted. Nine primary swarms formed clusters around the swarm attractant sachets while one secondary swarm was attracted and established directly in a hive trap.

The application of highly centrifuged honey as an improved diet for experimentally caged honey bees

Summary Whilst sucrose syrup appears to be the most commonly used diet applied during honey bee cage experiments, dead honey bees may occur within the first week of feeding. However, sucrose syrup remains the dominant food supplement for cage experiments since past research has proved that all alternative hydrocarbon diets, including honey, decrease the lifespan of honey bees when compared to syrup. In this study we tested feeding caged honey bees with honey from which all pollen grains and possibly all pathogenic spores were removed through high speed centrifugation. This preparation significantly extended the lifespan of honey bees in cages compared with sucrose syrup and honey. The analysis of the glycoside content of centrifuged honey and of the honey from which it was produced showed that the two products were identical. The 1H NMR spectra of the two samples displayed an identical resonance pattern. This refers to the qualitative content, as well as the ratio of the constituent sugars of the two samples. This study showed that highly centrifuged honey could be applied as an improved reliable diet for experimentally caged honey bees.