Liesel Brenda Gende

Antimicrobial activity of some Argentinean wild plant essential oils against Paenibacillus larvae larvae, causal agent of American foulbrood (AFB)

Summary Five essential oils from Argentinan wild plants were tested in vitro for antimicrobial activity against different strains of Paenibacillus larvae larvae, causal agent of American foulbrood in honey bees, Apis mellifera L. The species selected from which to extract the essential oils were aguaribay (Schinus molle var. areira L), Andean thyme (Acantholippia seriphioides A. Gray), peperine (Mintosthachys mollis H.B.K. Gris), camomile (Tagetes minuta L.) and pennyroyal (Lippia turbinata Griseb.). Oils were extracted from different parts of the plants by steam distillation. Minimal inhibitory concentration (MIC) and minimal bactericide concentration (MBC) were evaluated on Mueller-Hinton broth and on MYPGP agar respectively, by the tube dilution method. MIC and MBC minimum and maximum values obtained were 200–250 mgL−1 and 200–300 mgL−1 for Andean thyme and 800–1000 mgL−1 and 850–1100 mgL−1 for pennyroyal, respectively. The other oils had intermediate MIC and MBC values between the formers mentioned. Highly significant differences between MIC and MBC values for the essential oils tested were found. Tukeys mean separation test indicated that Andean thyme (A. seriphioides) essential oil was different from the other four essential oils at α<0.01. Andean thyme proved to be the most effective in vitro essential oil against P. larvae larvae. Therefore, it may be a promising natural alternative to traditional antibiotics, meriting further studies on its field applicability for AFB control.

Antimicrobial activity of Pimpinella anisum and Foeniculum vulgare essential oils against Paenibacillus larvae.

Abstract The essential oil obtained by hydrodistillation from the fruits of Pimpinella anisum L. (green anise) and Foeniculum vulgare Miller (fennel) were analyzed by GC and GC/MS and physicochemical properties. The oils of P. anisum and F. vulgare were found to be especially rich in (E)-anethole, 96.3% and 92.7%, respectively. The MICs were determined by the tube dilution method against Paenibacillus larvae. The oils showed MICs values were 300 µg/mL and 250 μg/mL for P. anisum and F. vulgare, respectively. Both oils presented great similarity in physiochemical properties values and antimicrobial activity.

LC50 baseline levels of amitraz, coumaphos, fluvalinate and flumethrin in populations of Varroa destructor from Buenos Aires Province, Argentina

Summary This study estimates the LC50 baseline levels for amitraz, coumaphos, fluvalinate, and flumethrin in susceptible Varroa destructor populations in Argentina. Concentration response bioassays were conducted with each acaricide. Laboratory results of lethal concentrations (LC50) were: 0.1 μg/dish for amitraz; 0.29 μg/dish for fluvalinate; 0.34 μg/dish for flumethrin; and 0.57 μg/dish for coumaphos, respectively. All tests guaranteed 100 % bee survival. LC50 references for mite populations susceptible to the most commonly used acaricides were thus established for Argentina, which will aid the establishment of integrated pest management programmes for V. destructor.

Efficacy of Essential Oils of Thymus vulgaris and Origanum vulgare on Echinococcus granulosus

The aim of the present work was to determine the in vitro effect of T. vulgaris and O. vulgare essential oils against E. granulosus protoscoleces and cysts. Essential oils were added to the medium resulting in thymol final concentrations of 10 μg/mL. The essential oils had a time-dependent effect provoking the complete loss of protoscolex viability after 72 days of postincubation. The results were confirmed at the ultrastructure level. Loss of infectivity in protoscoleces incubated with O. vulgare after 60 days was observed. On the other hand, the weight of cysts recorded in mice inoculated with T. vulgaris treated protoscoleces was significantly lower than that obtained in control group. Gamma-glutamyl-transpeptidase activity was readily detected in the culture supernatant of protoscoleces treated either with the essential oils or thymol. T. vulgaris and O. vulgare essential oils and thymol can induce cell apoptosis of protoscoleces after short incubation times. The efficacy of T. vulgaris and O. vulgare essential oils was also demonstrated in vitro on E. granulosus murine cysts. Our data suggest that essential oils of T. vulgaris and O. vulgare have anthelmintic effect against protoscoleces and cysts of E. granulosus.

Bioactivity of Rosmarinus officinalis essential oils against Apis mellifera, Varroa destructor and Paenibacillus larvae related to the drying treatment of the plant material

In this study, chemical composition, physicochemical properties and bioactivity of two essential oils of Rosmarinus officinalis extracted from plant material with different drying treatments against Apis mellifera, Varroa destructor and Paenibacillus larvae were assessed. The lethal concentration 50 (LC50) for mites and bees was estimated using a complete exposure method test. The broth microdilution method was followed in order to determine the minimum inhibitory concentrations (MICs) of the essential oils against P. larvae. Physicochemical properties were similar in both the essential oils, but the percentage of components showed certain differences according to their drying treatment. β-Myrcene and 1,8-cineole were the main constituents in the oils. The LC50 for complete exposure method at 24, 48 and 72 h was minor for mites exposed to R. officinalis essential oil dried in oven conditions. MIC values were 700–800 µg mL−1 and 1200 µg mL−1 for R. officinalis dried in air and oven conditions, respectively. The results reported in this research show that oil toxicity against V. destructor and P. larvae differed depending on the drying treatment of the plant material before the distillation of essential oil.

Evaluation of the toxicity of a propolis extract on Varroa destructor (Acari: Varroidae) and Apis mellifera (Hymenoptera: Apidae)

Summary The effects of a propolis extract on Varroa destructor and Apis mellifera were evaluated by three different application methods: topical, spraying and oral. A propolis sample was extracted and its organoleptic and physic-chemically traits characterized. These analyses showed that it was a typical propolis from the Pampean region in Argentina, with elevated contents of biologically active compounds. Topical application was carried out by subjecting mites to contact with various propolis concentrations for different periods of time, which resulted in mortality and narcosis. Acaricidal effects were stronger with increasing concentrations of the propolis extracts. Spraying infested bees with a 10% propolis solution was harmless for bees but killed 78% of mites. Feeding infested bees with propolis extract in sugar syrup was not toxic to the mites but caused the death of bees treated with the highest concentration. Our results suggest that the propolis extracts from the Pampean Region could be incorporated into bee colonies by spraying, although the appropriate doses and concentrations to be administered, and the mechanism of action of the extracts on the mites are still to be elucidated.

Assessing in Vitro Acaricidal Effect and Joint Action of a Binary Mixture Between Essential Oil Compounds (Thymol, Phellandrene, Eucalyptol, Cinnamaldehyde, Myrcene, Carvacrol) Over Ectoparasitic Mite Varroa Destructor (Acari: Varroidae)

Abstract Varroa destructor (Anderson & Trueman, 2000) causes the most important parasitosis of beekeeping in the world. For this reason, prevention is needed to avoid colony death. The most typical treatments involve synthetic acaricides. However, the use of these acaricides results in the emergence of resistant populations of mites to these products and in the appearances of drug residues in products of the hives. Compounds of essential oils have emerged as an alternative to traditional acaricides; however the toxicity produced by these mixtures is currently poorly explored. The aim of this work was to assess, by means of in vitro tests with adult bees, how acaricidal action and toxic interactions in a binary mixture of essential oil compounds (Thymol, Phellandrene, Eucalyptol, Cinnamaldehyde, Myrcene, and Carvacrol) affect V. destructor. Calculations of LC50 ’s of the individual compounds on A. mellifera and V. destructor made clear that the toxic effect of each compound is different for both species. Thymol and Phellandrene turned out to be lethal for mites at lower concentrations than for bees. The binary mixture of these two substances presented a different toxicity than one produced by each pure compound, as it was highly selective for mites in bioassays at 24 hours through complete exposure to both A. mellifera and V. destructor. These results make such formulations optimal substances to be considered as alternative controls for the parasitosis.

Effect of Propolis Oral Intake on Physiological Condition of Young Worker Honey Bees, Apis Mellifera L.

Abstract Honey bees collect resin from various plant species and transform it into propolis that is incorporated into the nest. The role of resins in the bee health field is poorly understood. The aim was to evaluate the effects of forced consumption of propolis on the physiological condition and short-term survival of Apis mellifera worker bees. It was tested if the number of circulating hemocytes in hemolymph, the abdominal fat bodies and the hypopharyngeal glands development were affected by the feeding with propolis extracts in laboratory conditions during the warm and the cold seasons. Propolis added to sugar candy was consumed by workers for fourteen days without affecting the bee survival. The number of circulating hemocytes in hemolymph remained constant despite the differential diet during the experiment. However, the development of fat bodies and hypopharyngeal glands was altered by propolis ingestion. The abdominal fat body development in winter bees diminished after fourteen days of propolis consumption, while it increased in summer bees. The hypopharyngeal gland development decreased for the assayed period in workers from both seasons. Our results encourage us to continue exploring this research field and learn how long-term forced ingestion of a plant-derived compound, a non-nutritive substance, can modify physiological bee parameters. A broader understanding of the multiple roles of propolis in the health of the honey bee colonies could be obtained by studying the ways in which it is processed and metabolized and the effect that generates in another physiological responses.

Advances in Paenibacillus larvae and American foulbrood monitoring in honey bee colonies from Argentinean apiaries

Advances in Paenibacillus larvae and American foulbrood monitoring in honey bee colonies from Argentinean apiaries Natalia Jorgelina Fernández, Liesel Brenda Gende and Martín Javier Eguaras Arthropods Laboratory, School of Natural and Exact Sciences, Universidad Nacional de Mar del Plata, Funes, 3350. 7600, Mar del Plata, Argentina. CONICET [National Council for Scientific and Technical Research], Rivadavia 1917. C1033AJ, Buenos Aires, Argentina.

Laurus nobilis L . Extracts against Paenibacillus larvae : Antimicrobial activity, antioxidant capacity, hygienic behavior and colony strength

The aim of this work was to compare the antimicrobial activity against Paenibacillus larvae and the antioxidant capacity of two Laurus nobilis L. extracts obtained by different extraction methods. The hydroalcoholic extract was moreover added as supplementary diet to bees in field conditions to test behavioural effects and colony strength. Both laurel extracts were subjected to different phytochemical analysis to identify their bioactive compounds. Antimicrobial activity was analyzed by the minimal inhibitory concentration (MIC) determination by means the agar dilution method. The hydroalcoholic extract (HE) was able to inhibit the bacterial growth of all P. larvae strains, with 580 µg/mL mean value. This better antibacterial activity in relation to the essential oil (EO) could be explained by the presence of some phenolic compounds, such as flavonoids, evidenced by characteristic bands resulting from the Fourier Transform Infrared Spectroscopy (FTIR) analysis. Antioxidant activities of the extracts were evaluated by 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical-scavenging ability and ferric reducing antioxidant power (FRAP) assays. The HE showed the highest antioxidant activity as measured by DPPH, with IC50 values of 257 ± 12 μg/mL. The FRAP assay method showed that the HE was 3-fold more effective reducing agent than the EO. When the bee colonies were supplied with laurel HE in sugar paste an improvement in their general condition was noticed, although neither the hygienic behavior nor the proportions of the breeding cells varied statistically due to the treatment. In conclusion, the inhibition power against P. larvae attributable to the phenolic compounds, the antioxidant capacity of the HE, and the non-lethal effects on adult honey bees on field trials suggest the HE of laurel as a promising substance for control American foulbrood disease.