Due to its multifunctional and complex role in the honey bee colony functioning and health (construction material allowing food storage, brood rearing, thermoregulation, mediation in chemical and mechanical communication, substrate for pathogens, toxins and waste), Apis mellifera beeswax has been widely studied over the last five decades. This is supported by a comprehensive set of scientific reports covering different aspects of beeswax research. In this article, we present an overview of the methods for studying chemical, biological, constructional, and quality aspects of beeswax. We provide a detailed description of the methods for investigating wax scales, comb construction and growth pattern, cell properties, chemical composition of beeswax using different analytical tools, as well as the analytical procedures for provenancing beeswax and beeswax-derived compounds based on the hydrogen isotope ratio (IRMS). Along with classical physico-chemical and sensory analysis, we describe more precise and accurate methods for detection of adulterants in beeswax (GC-MS and FTIR-ATR). Moreover, we present methods for studying the influence of beeswax (comb foundation) adulteration on comb construction. Analytical protocols for determining the pesticide residues using different chromatographic and spectroscopic techniques are also described. As beeswax is an agent of high risk for the transmission of bee diseases, we present methods for detection of pathogens in beeswax. To ensure the reproducibility of experiments and results, we present best practice approaches and detailed protocols for all methods described, as well as their advantages and disadvantages.

BackgroundPropolis is a bee product widely used in folk medicine and possessing many pharmacological properties. In this study we aimed to investigate: i) the antiviral activities of Hatay propolis samples against HSV-1 and HSV-2 in HEp-2 cell line, and ii) the presence of the synergistic effects of propolis with acyclovir against these viruses.Material/MethodsAll experiments were carried out in HEp-2 cell cultures. Proliferation assays were performed in 24-well flat bottom microplates. We inoculated 1×105 cells per ml and RPMI 1640 medium with 10% fetal calf serum into each well. Studies to determine cytotoxic effect were performed. To investigate the presence of antiviral activity of propolis samples, different concentrations of propolis (3200, 1600, 800, 400, 200, 100, 75, 50, and 25 μg/mL) were added into the culture medium. The amplifications of HSV-1 and HSV-2 DNA were performed by real-time PCR method. Acyclovir (Sigma, USA) was chosen as a positive control. Cell morphology was evaluated by scanning electron microscopy (SEM).ResultsThe replication of HSV-1 and HSV-2 was significantly suppressed in the presence of 25, 50, and 100 μg/mL of Hatay propolis. We found that propolis began to inhibit HSV-1 replication after 24 h of incubation and propolis activity against HSV-2 was found to start at 48 h following incubation. The activity of propolis against both HSV-1 and HSV-2 was confirmed by a significant decrease in the number of viral copies.ConclusionsWe determined that Hatay propolis samples have important antiviral effects compared with acyclovir. In particular, the synergy produced by antiviral activity of propolis and acyclovir combined had a stronger effect against HSV-1 and HSV-2 than acyclovir alone.

The aim of this study was to improve cage systems for maintaining adult honey bee (Apis mellifera L.) workers under in vitro laboratory conditions. To achieve this goal, we experimentally evaluated the impact of different cages, developed by scientists of the international research network COLOSS (Prevention of honey bee COlony LOSSes), on the physiology and survival of honey bees. We identified three cages that promoted good survival of honey bees. The bees from cages that exhibited greater survival had relatively lower titers of deformed wing virus, suggesting that deformed wing virus is a significant marker reflecting stress level and health status of the host. We also determined that a leak- and drip-proof feeder was an integral part of a cage system and a feeder modified from a 20-ml plastic syringe displayed the best result in providing steady food supply to bees. Finally, we also demonstrated that the addition of protein to the bees' diet could significantly increase the level ofvitellogenin gene expression and improve bees' survival. This international collaborative study represents a critical step toward improvement of cage designs and feeding regimes for honey bee laboratory experiments.

Propolis is a honeybee product with a very complex chemical composition and various pharmacological properties. This study was aimed to investigate antileishmanial activities of "Bursa" and "Hatay" propolis samples against Leishmania infantum and Leishmania tropica strains. Propolis samples were analysed with the gas chromatography-mass spectrometry technique. Promastigotes were incubated in Roswell Park Memorial Institute culture medium in the absence and presence of several concentrations (50, 100, 250, 500, 750, and 1,000 μg/mL) of each propolis sample. The viability and cell morphology of promastigotes in each concentration were examined after 24, 48, 72, and 96 h of incubation. The growth of leishmania parasites was significantly suppressed in the presence of 500, 750, and 1,000 μg/mL of Hatay propolis. Bursa propolis was found to be efficient in inhibiting the growth of leishmania promastigotes in culture media at these concentrations, 250, 500, 750, and 1,000 μg/mL. Thus, the in vitro results showed that the Hatay and Bursa propolis samples decreased significantly the proliferation of L. infantum and L. tropica parasites (p < 0.001); however, Bursa propolis was found to be more effective than Hatay propolis against leishmania promastigotes. These two natural products may be useful agents in the prevention of leishmanial infections.

Modern European beekeeping is facing numerous challenges due to a variety of factors, mainly related to globalisation, agrochemical pollution and environmental changes. In addition to this, new pathogens threaten the health of European honeybees. In that context, correct colony management should encompass a wider vision, where productivity aspects are linked to a One Health approach in order to protect honeybees, humans and the environment. This paper describes a novel tool to be applied in beekeeping operations: good beekeeping practices (GBPs). The authors ranked a list of GBPs scored against their importance and validated by an international team, including researchers, national animal health authorities and international beekeepers' associations. These activities were carried out in the project 'BPRACTICES', approved within the transnational call of the European Research Area Network on Sustainable Animal Production Rev. Sci. Tech. Off. Int. Epiz., 38 (3) 3 3/27 (ERA-NET SusAn) in the Horizon 2020 research and innovation programme of the European Union. This study, created through an international collaboration, aims to present an innovative and implementable approach, similar to applications already adopted in other livestock production systems.

This study investigated the geographical distribution and molecular analysis of Culicoides species in the Southern and Southeastern Turkey during the 2012 outbreak of bovine ephemeral fever (BEF). The midge specimens caught by Onderstepoort-type light traps from livestock farms were tested for molecular evidence of existence of viral genome. Blood specimens were collected from clinically BEF-suspected acute febrile cattle. Total nucleic acid samples obtained from field specimens were checked against the BEF virus G gene and Culicoides internal transcribed spacer 1 (ITS-1) gene. A total of 20,845 Culicoides specimens (20,569 ♀♀, 276 ♂♂) comprising 11 species (Culicoides badooshensis, Culicoides circumscriptus, Culicoides gejgelensis, Culicoides imicola, Culicoides kibunensis, Culicoides longipennis, Culicoides newsteadi, Culicoides nubeculosus, Culicoides odiatus, Culicoides punctatus, Culicoides schultzei, Culicoides spp.) were collected. C. schultzei (18,032) was found as the dominant species and followed by C. imicola (1,857), C. nubeculosus complex (545), and C. circumscriptus (259), respectively. C. kibunensis was identified as new species for this region. PCR positivity of BEF was found 37.14% (13/35) in blood samples whereas no viral genome was obtained from Culicoides specimens. Culicoides spp. ITS-1 gene sequences were analyzed phylogenetically with GenBank ITS-1 sequences. Molecular homology of Culicoides ITS-1 gene was ranged between 62.74 and 71.39%. The results described first molecular detection and phylogenetic analysis of Culicoides ITS-1 gene with reference to the 2012 BEF outbreak in Turkey.

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