Effect of Entomopathogenic Fungi to Eggs and Nymphs Survival of Halyomorpha halys (Hemiptera: Pentatomidae) Under Laboratory Conditions.

Halyomorpha halys (Stål) (Hemiptera: Pentatomidae) is becoming a major pest of Greece's agricultural production, including tree fruit. Until now, the control of this pest has been based on conventional methods, with the application of chemical insecticides being the primary option. However, the wide distribution of H. halys, in combination with the large area of tree fruit production in Greece, raise the need for alternative control methods. In this study, we investigated the possibility of implementing microbial control agents for managing H. halys. Eggs and nymphs of H. halys were treated with 15 native entomopathogenic fungal isolates and their virulence was evaluated in the laboratory. After treatment, egg hatching time was recorded for 7 days and ranged from 4.5 to 7.0 days. Nymphal survival was recorded daily for 7 days following application and ranged from 2.1 to 6.6 days for second instar nymphs and 3.7 to 6.8 days for fourth instar ones. Beauveria varroae isolate displayed the highest toxicity to all H. halys stages that were tested and could be considered a promising biocontrol agent of this insect. Please check and confirm that the authors and their respective affiliations have been correctly identified and amend if necessary.οκ.

Entomopathogenic fungi tested in planta on pepper and in field on sorghum, to control commercially important species of aphids.

Given the aphids high reproductive capacity, assessing their biocontrol by using entomopathogenic fungi is crucial; to determine their potential, fungi were tested in planta and in field conditions. Significant decrease of Myzus persicae (Sulzer) population was observed in planta after applying Beauveria bassiana (strain 7R), Trichoderma gamsii (strain Z) or Metarhizium brunneum (strain Meta Br1) at 1 × 107 or 1 × 108 conidia/mL on pepper plants. Significant differences of aphids' populations were detected between fungus concentration and control (F = 68.743, df = 6.980, P < 0.001), where M. brunneum at 1 × 108 conidia/mL reduced aphids population close to zero. At 20 °C, dead aphids' mycosis by B. bassiana and T. gamsii was 78% and 84%; at 25 °C was 83% and 88%; and at 30 °C was 75% and 79%, respectively. In field conditions, Mexican PTG4 and commercial GHA B. bassiana strains were tested [(1 × 106 conidia/mL + corn starch) seed treatments] against the Melanaphis sacchari (Zehntner) aphid populations, on naturally infested sorghum plants. Results showed that plant germination and emergence were not affected, whereas yield (grams of sugar/plant) was significantly higher among treated compared with untreated plants. The aphid population decreased in plants from PTG4 treated seeds; indeed, this treatment had a significant positive effect on the flowering index, whereas the stem fresh weight and juice volume was significantly increased among plants from GHA treated seeds. Taken together, tested strains can be used as a tool to control aphids' population on several crops such as pepper and even increase the yield in sorghum.

A quantitative western blot technique using TMB: Comparison with the conventional technique.

Numerous molecular biological experiments performed throughout the world require the detection or quantification of a protein of interest. Western blotting is one of the most popular techniques used for this purpose and offers quantitative information with the aid of specialized software. However, its dependence on the picture that is captured, and the background and the absence of a common protocol prevent the technique from being completely quantitative. To overcome these obstacles, we present a simple and reliable assay that is similar to the regular technique, with the exception of the last stage of band visualization and quantification. We propose that small pieces of the blot that include the protein of interest can be cut and dipped in a small volume of 3,3',5,5'-tetramethylbenzidine solution, giving a colorimetric signal with linear dependence on the quantity of the protein. The reaction is stopped with H2 SO4 , and the signal is measured in a plate reader. This modification shows high linearity without additional costs and can be applied for both purified proteins and proteins found in a lysate. The results obtained with our proposed technique were compared with those obtained by the conventional method and proved to be more reliable.

Evaluation of Various Inoculation Methods on the Effect of Beauveria bassiana on the Plant Growth of Kiwi and on Halyomorpha halys Infestation: A Two-Year Field Study.

In this study, the bioinsecticidal action of a commercial formulation with Beauveria bassiana was evaluated on the new sucking pest in Greece: Halyomorpha halys, of the kiwifruit. Additionally, the biostimulant potential of the same formulation was studied on kiwi growth. The application was performed in three different ways in a commercial field of kiwi crop A. deliciosa "Hayward" field in Arta, Greece: (i) trunk spray, (ii) root injection, and (iii) trunk inoculation. During the 2 years seasons of the experiment, weekly measurements of the H. halys population were determined. The insect is sucking plants nutrients; therefore, the total chlorophyll content in the leaves of the treatments was recorded weekly. In addition, the percentage of infested kiwifruits was estimated at the end of the experiment. Moreover, to study the biostimulant potential of the formulation, growth measurements on stems and leaves were performed during the experiment. Finally, at the kiwi harvest point, the fruit biomass, dimensions, and weight were obtained, and the leaves' proline content was evaluated. The results encourage us to further study this EPF formulation as the bioinsecticidal effect was noted by the reduction in H. halys population, and biostimulant action was perceived by the higher plant biomass.

Entomopathogenic Action of Wild Fungal Strains against Stored Product Beetle Pests.

There is ample evidence that entomopathogenic fungi can be used as alternative biological control agents for the management of insect pests in storage facilities. As the market demands more environmentally friendly methods and chemical insecticides become increasingly obsolete, more studies are being conducted to evaluate new strains of entomopathogenic fungi for their efficacy in storage facilities. In this context, we tested ten species of fungi isolated from soil, belonging to the genera Cladosporium, Condenascus, Lecanicillium, and Penicillium, for their long-term effects on economically important beetle species. Whole wheat was directly sprayed with a conidial suspension of 108 spores/Ml of each of the tested fungi and then adults of Sitophilus granarius, S. oryzae, S. zeamais, Rhyzopertha dominica, and Trogoderma granarium were placed on the sprayed medium to study the mortality effects. Significantly higher mortality than the control was observed in all treatments. The lowest LT50 (9.164 days) was observed in T. granarium infected with Penicillium goetzii. The isolate with the strongest results was L. dimorphum, which recorded remarkably low LT50 values in S. oryzae (~11 days), R. dominica (~12 days), T. granarium (~10 days), and S. granarius (~13 days). However, for S. zeamais, it was more than 16 days. Our results confirm the existing literature on the efficacy of EPF on storage beetles, suggest the possible virulence of wild untested strains, and also highlight the importance of EPF specificity.

Biostimulant and Bioinsecticidal Effect of Coating Cotton Seeds with Endophytic Beauveria bassiana in Semi-Field Conditions.

Increasing commercial demands from the textile and food industries are putting strong pressure on the cultivation of cotton and its derivatives to produce high yields. At the same time, cotton has high nutrient and irrigation requirements and is highly susceptible to insect pests. Coating cotton seeds with beneficial fungi could address these problems. The aim of this study was to investigate the growth of cotton using (A) conventional seeds and (B) seeds coated with the entomopathogenic fungus Beauveria bassiana (Hypocreales: Cordycipitaceae). The experiment was conducted in a greenhouse of the Department of Agriculture of the University of Ioannina, in a completely randomized design. The growth characteristics of cotton plants were recorded weekly while the fresh weight and dry matter of the leaves, shoots and roots of the developed cotton plants were calculated at the end of the experiment. Weekly determination of total chlorophyll content (TCHL) was used as an indicator of plant robustness during the 80-day experiment. Many cotton growth parameters of treated plants, like number of leaves, shoots and apical buds, plant height, stem diameter, fresh and dried biomass and TCHL, were significantly higher than those of the untreated ones. Apart from plant growth, naturally occurring by Aphis gossypii (Hemiptera: Aphididae) infestation which also monitored for six weeks. A significantly lower aphid population was recorded for inoculated plants after the fifth week compared to the control. The overall evaluation revealed that B. bassiana coating treatments appear to have a significant biostimulatory and bioinsecticidal effect. Our results could represent responsive applications to the demands of intensive cotton growing conditions.

Dual Action of Beauveria bassiana (Hypocreales; Cordycipitaceae) Endophytic Stains as Biocontrol Agents against Sucking Pests and Plant Growth Biostimulants on Melon and Strawberry Field Plants.

Entomopathogenic fungi (EPF) can colonize plant tissues and serve crops not only as biopesticides but also as biostimulants that promote plant growth and trigger defense mechanisms. In this context, field trials were conducted evaluating two commercial strains of the entomopathogen Beauveria bassiana (Hypocreales: Cordycipitaceae), GHA (Botanigard) and PPRI 5339 (Velifer® ES) and a wild strain (AP0101) isolated from Achaia, Greece. The three strains were investigated in the field for their endophytic effects on melon Cucumis melo (Cucurbitales: Cucurbitaceae) and strawberry Fragaria sp. (Rosales: Rosaceae) plants and in particular for their ability to colonize plant tissues, control infestations of sucking insects Aphis gossypii (Hemiptera: Aphididae), Chaetosiphon fragaefolii (Hemiptera: Aphididae) and Frankliniella occidentalis (Thysanoptera: Thripidae), and improve plant growth parameters (plant height, number of flowers and fruits). All experimental fungal strains successfully colonized both plants. A significant decrease in the aphid and thrip populations was observed in the treated plants compared to the untreated control. As for plant growth, the number of flowers and fruits was significantly increased in plants treated with B. bassiana strains AP0101 and PPRI 5339. Our results clearly indicate that fungal endophytes can efficiently act as dual action agents demonstrating both insecticidal and growth-promoting effects.

Beauveria bassiana Endophytic Strain as Plant Growth Promoter: The Case of the Grape Vine Vitis vinifera.

The common grape vine, Vitis vinifera, is a widely known plant with commercial and pharmacological value. The plant hosts a variety of microorganisms known as endophytes that can live within the tissues of the plant for a considerable time period, or even their whole life cycle. The fungus Beauveria bassiana is a well-studied endophyte which can colonize a variety of plants in many ways and in different parts of the plant. In this study, we examined the effect of the endophytic fungus B. bassiana on the growth of V. vinifera. The results demonstrated not only a successful colonization of the endophyte, but also a noteworthy impact on the growth of the V. vinifera root without harming the plant in any way. The fungus was also re-isolated from the parts of the plant using inst bait method. Overall, the study demonstrates the capability of B. bassiana to colonize V. vinifera plants, adding to the already existing knowledge of its endophytic activity, and highlighting its beneficial impact on the root growth.

Larvicidal Action of Cannabidiol Oil and Neem Oil against Three Stored Product Insect Pests: Effect on Survival Time and in Progeny.

SStored product pests can be detrimental to agricultural produce. As much as chemical pesticides are effective control agents, they involve several environmental and health risks. Within the framework of studies on alternative pest management methods, interest has focused on a plethora of plants whose extracts have demonstrated promising action as insecticides. Azadirachta indica and the derived neem oil have been extensively tested against many harmful insect species. In contrast, Cannabis sativa L. and its main compound, CBD, a highly concentrated cannabinoid, have not been investigated much. The present study examined the potential insecticidal activity of CBD and neem oils against 4th instar larvae of Tribolium confusum, Oryzaephilus surinamensis and Plodia interpunctella on wheat, rice and corn seeds. Treatment efficacy was expressed in terms of larval mortality. Mortality was observed in relation to dosage, time exposure intervals and product types. The results showed clear pesticidal activity for both oils, which at high doses induced significant mortality. The treatments produced significantly fewer offspring in the insect species tested than the control. The efficacy of treatment in progeny suppression was, as expected, dose dependent.

Leguminous Seeds Powder Diet Reduces the Survival and Development of the Khapra Beetle, Trogoderma granarium Everts (Coleoptera: Dermestidae).

Chemical storage pest control is interlinked with many challenges such as environmental pollution and toxicity to humans and animals. Alternative tools are thus being increasingly researched and applied to supplement and/or substitute old-fashioned chemical means. Entomotoxic proteins, such as the lectins of leguminous seeds, have been shown to be effective alternative control agents against many serious insect pests. The objective of this work was to evaluate the effect of the flour of three leguminous seeds, Phaseolus vulgaris L. (Fabaceae) (the common bean), Vicia faba L. (Fabaceae) (the broad bean) and Glycine max L. (Fabaceae) (the soya bean), against 4th instar larvae of Trogoderma granarium Everts (Coleoptera: Curculionidae). The flours were tested at different concentrations. They all demonstrated significant effects on larval mortality, as well as they all induced a decrease in the number of larvae reaching the pupal stage. The flours of P. vulgaris and V. faba were highly insecticidal against T. granarium larvae, especially at the highest concentrations (86.7% for PV100 and 90% for VF100). Our results enrich previous findings on the entomotoxic effect of leguminous plant lectins and highlight P. vulgaris and V. faba lectins as potential alternative control agents against T. granarium.

Combined Toxicity of Cannabidiol Oil with Three Bio-Pesticides against Adults of Sitophilus Zeamais, Rhyzopertha Dominica, Prostephanus Truncatus and Trogoderma Granarium.

The present study investigates the interaction between cannabidiol (CBD) oil and three biopesticides: Azatin and two baculovirus formulations (Madex and Helicovex), both separately and in combination, in order to investigate their interaction against adults of four major coleopteran stored-product pests: Sitophilus zeamais (Coleoptera: Curculionidae), Rhyzopertha dominica (Coleoptera: Bostrichidae), Prostephanus truncatus (Coleoptera: Bostrichidae) and Trogoderma granarium (Coleoptera: Dermestidae). CBD, which has been understudied for its insecticidal properties, was applied at three different doses (500, 1500 and 3000 ppm). The biopesticides were administered at 1500 ppm. Interactions in the combined treatments were mathematically estimated as not synergistic and mostly competitive except for the combined treatments of CBD (1500 and 3000 ppm) with Azatin (1500 ppm) which were marked by an additive interaction. In its individual application, CBD oil generated the highest insect mortality while its effect was clearly dose-dependent. The findings reveal a promising effect of CBD oil against these coleopterans which had not been previously tested together.

Ιnteractions between Beauveria bassiana and Isaria fumosorosea and Their Hosts Sitophilus granarius (L.) and Sitophilus oryzae (L.) (Coleoptera: Curculionidae).

The interactions between the entomopathogenic fungus Beauveria bassiana Balsamo (Vuillemin) (Hypocreales: Cordycipitaceae) and the entomopathogenic fungus Isaria fumosorosea (Wize) Brown and Smith (Hypocreales: Clavicipitaceae) were examined on young adults of Sitophilus granarius (L.) (Coleoptera: Curculionidae) and S. oryzae (L.) (Coleoptera: Curculionidae). Conidial suspensions of these entomopathogenic fungi were applied both separately and in combination, at three dosages, 104, 106, and 108 conidia/mL. Mortality of experimental adults was recorded daily for 15 days. An overall positive interaction between the pathogenic microorganisms was observed. Mean weevil mortality caused by the separate acting fungi, B. bassiana, ranged from 26.7% to 53.3% and from 36.6% to 63.3% for S. granarius and S. oryzae, respectively. The respective values for I. fumosorosea were 20.0%-53.3% and 46.7%-66.7%. The combined treatments showed a distinct interaction between the pathogens; for S. granarius, the interaction between the pathogens was additive in all combinations, whereas, for S. oryzae, the interaction was additive in seven and competitive in two of the combinations. Applying both entomopathogenic microorganisms may offer a method for weevil control that could be more effective than using each pathogen alone.