Liviu A. Mărghitaş

Pathogen-associated self-medication behavior in the honeybee Apis mellifera

Honeybees, Apis mellifera, have several prophylactic disease defense strategies, including the foraging of antibiotic, antifungal, and antiviral compounds of plant products. Hence, honey and pollen contain many compounds that prevent fungal and bacterial growth and inhibit viral replication. Since these compounds are also fed to the larvae by nurse bees, they play a central role for colony health inside the hive. Here, we show that honeybee nurse bees, infected with the microsporidian gut parasite Nosema ceranae, show different preferences for various types of honeys in a simultaneous choice test. Infected workers preferred honeys with a higher antibiotic activity that reduced the microsporidian infection after the consumption of the honey. Since nurse bees feed not only the larvae but also other colony members, this behavior might be a highly adaptive form of therapeutic medication at both the individual and the colony level.

What Is the Main Driver of Ageing in Long-Lived Winter Honeybees: Antioxidant Enzymes, Innate Immunity, or Vitellogenin?

To date five different theories compete in explaining the biological mechanisms of senescence or ageing in invertebrates. Physiological, genetical, and environmental mechanisms form the image of ageing in individuals and groups. Social insects, especially the honeybee Apis mellifera, present exceptional model systems to study developmentally related ageing. The extremely high phenotypic plasticity for life expectancy resulting from the female caste system provides a most useful system to study open questions with respect to ageing. Here, we used long-lived winter worker honeybees and measured transcriptional changes of 14 antioxidative enzyme, immunity, and ageing-related (insulin/insulin-like growth factor signaling pathway) genes at two time points during hibernation. Additionally, worker bees were challenged with a bacterial infection to test ageing- and infection-associated immunity changes. Gene expression levels for each group of target genes revealed that ageing had a much higher impact than the bacterial challenge, notably for immunity-related genes. Antimicrobial peptide and antioxidative enzyme genes were significantly upregulated in aged worker honeybees independent of bacterial infections. The known ageing markers vitellogenin and IlP-1 were opposed regulated with decreasing vitellogenin levels during ageing. The increased antioxidative enzyme and antimicrobial peptide gene expression may contribute to a retardation of senescence in long-lived hibernating worker honeybees.

Botanical Origin Causes Changes in Nutritional Profile and Antioxidant Activity of Fermented Products Obtained from Honey

Honey as rich source of enzymatic and nonenzymatic antioxidants serves as health-promoting nutrient in the human body. Here, we present the first time a comparative study of nutritional profiles (e.g., acidities, sugar, organic acid profile, total polyphenolic, flavonoid content) for different unifloral, multifloral honeys and their fermented products, in correlation with their antioxidant activity. Additionally, an optimized method for HPLC separation of organic acids from honey was established. The total phenolic content of honey samples varied widely among the honey types compared to fermented products. High amounts of total flavonoids were quantified in heather honey, followed by raspberry, multifloral, black locust, and linden honey. A positive correlation between the content of polyphenols, flavonoids, and antioxidant activity was observed in honey samples. After fermentation, the flavonoid content of dark honey fermented products decreased significantly. Black locust and linden honeys are more suitable for fermentation because the decrease in antioxidant substances is less pronounced.

Climate rather than geography separates two European honeybee subspecies

Both climatic and geographical factors play an important role for the biogeographical distribution of species. The Carpathian mountain ridge has been suggested as a natural geographical divide between the two honeybee subspecies Apis mellifera carnica and A. m. macedonica. We sampled one worker from one colony each at 138 traditional apiaries located across the Carpathians spanning from the Hungarian plains to the Danube delta. All samples were sequenced at the mitochondrial tRNALeu‐cox2 intergenic region and genotyped at twelve microsatellite loci. The Carpathians had only limited impact on the biogeography because both subspecies were abundant on either side of the mountain ridge. In contrast, subspecies differentiation strongly correlated with the various temperature zones in Romania. A. m. carnica is more abundant in regions with the mean average temperature below 9 °C, whereas A. m. macedonica honeybees are more frequent in regions with mean temperatures above 9 °C. This range selection may have impact on the future biogeography in the light of anticipated global climatic changes.

Comparison between local and commercial royal jelly—use of antioxidant activity and 10-hydroxy-2-decenoic acid as quality parameter

Summary Royal jelly (RJ) is secreted by worker bees of Apis mellifera to feed the queen larvae. Believed to be the longevity factor of queen honey bees and due to its variety of health promoting ingredients, RJ is used worldwide as dietary supplement, health product and in cosmetics. To reach best possible effects, application of high quality RJ is absolutely required. Here we show, based on physicochemical properties and antioxidant activity, a comparison between fresh, local RJ samples obtained directly from beekeepers and commercial ones. Analyses revealed a higher content of 10-hydroxy-2-decenoic acid, a quality parameter of RJ, for local compared to commercial RJ. Furthermore, determined ferric reducing antioxidant power (FRAP) correlated with radical scavenging activity (DPPH) and total polyphenols was significantly higher in local RJ samples. Therefore, our results indicate a higher quality for local RJ compared to commercial.

Influence of geographic origin, plant source and polyphenolic substances on antimicrobial properties of propolis against human and honey bee pathogens

Natural honey bee products, especially propolis are well known for their biological properties. However, less well known are the impact of propolis’ single phenolic substances and variation across different landscapes, geographic locations and botanical origins. Here, we determined the chemical composition of propolis samples collected in different geographic areas of Romania with a continental climate. Main groups of polyphenols, as well as individual phenolics (phenolic acids and flavonoids), were identified and quantified. The antimicrobial activity of propolis ethanolic extracts and some of the most important propolis phenolics (caffeic and ferulic acid, pinocembrin, chrysin and galangin) were evaluated against different bacterial species. Analyzing the interaction of bacterial growth inhibition and different chemical parameters, revealed that flavone/flavonol, flavanone/dihydroflavonol, chrysin and galangin are largely responsible for the propolis’ antimicrobial activity against Gram-positive bacteria. Gram-negative bacteria were mostly inhibited by galangin, caffeic acid and naringenin. Chemical composition and antimicrobial activity of the propolis samples might be explained by the botanical sources present in the specific foraging areas. The highest amounts of polyphenols and flavonoids were measured in regions where deciduous forests were dominant to coniferous, or where large areas of willow could be observed.

Genetic Characterization of Bombyx mori (Lepidoptera: Bombycidae) Breeding and Hybrid Lines With Different Geographic Origins

Abstract The domesticated silkworm Bombyx mori L. comprises a large number of geographical breeds and hybrid lines. Knowing the genetic structure of those may provide information to improve the conservation of commercial lines by estimating inbreeding over generations and the consequences of excessive use of those lineages. Here, we analyzed the genetic diversity of seven breeds and eight hybrid lines from Eastern Europe and Asia using highly polymorphic microsatellites markers to determine its genetical impact on their use in global breeding programs. No consistent pattern of deviation from Hardy–Weinberg equilibrium was found for most breed and hybrids; and the absence of a linkage disequilibrium also suggests that the strains are in equilibrium. A principal coordinate analysis revealed a clear separation of two silkworm breeds from the rest: one (IBV) originated from India and the other one (RG 90 ) from Romania/Japan. The tendency of the other breeds from different geographic origins to cluster together in a general mix might be due to similar selection pressures (climate and anthropogenic factors) in different geographic locations. Phylogenetic analyses grouped the different silkworm breeds but not the hybrids according to their geographic origin and confirmed the pattern found in the principal coordinate analysis.

Antiproliferative activity and apoptotic effects of Filipendula ulmaria pollen against C26 mice colon tumour cells

Abstract Honeybee collected pollen exhibits high nutritional and pharmaceutical benefits for the human diet and medicine. Pollen’s antioxidant, anti-ageing, anti-inflammatory, anti-atherosclerosis, and cardioprotective activity, depending on the floral origin, are well known. Recent studies proposed that pollen may also be an excellent cancer-fighting candidate, as pollen harbours high amounts of phenolic substances. In our study, Filipendula ulmaria pollen (bee collected) was methanol-water extracted and used to verify its in vitro pharmacological activities on C26 mice cancer tumour cells. Three different concentrations of the extract were tested in antitumour assays. Monitoring was done after 6, 12, 24, and 48 hours. Promising results were obtained for antiproliferative and apoptotic activity of the pollen extracts, with high efficiency for the highest concentration (1 mg/mL). For both activities, time and concentration-dependent effects were observed. Pollen extracts or bee collected pollen has a high potential as an antitumour agent for use in human medicine, because they are both rich in bioactive compounds.

Bay laurel (Laurus nobilis) as potential antiviral treatment in naturally BQCV infected honeybees

Viral diseases are one of the multiple factors associated with honeybee colony losses. Apart from their innate immune system, including the RNAi machinery, honeybees can use secondary plant metabolites to reduce or fully cure pathogen infections. Here, we tested the antiviral potential of Laurus nobilis leaf ethanolic extracts on forager honeybees naturally infected with BQCV (Black queen cell virus). Total viral loads were reduced even at the lowest concentration tested (1mg/ml). Higher extract concentrations (≥5mg/ml) significantly reduced virus replication. Measuring vitellogenin gene expression as an indicator for transcript homeostasis revealed constant RNA levels before and after treatment, suggesting that its expression was not impacted by the L. nobilis treatment. In conclusion, plant secondary metabolites can reduce virus loads and virus replication in naturally infected honeybees.