Effects of inoculation method on efficacy of wettable powder and oil dispersion formulations of Beauveria bassiana against Colorado potato beetle larvae under low-humidity conditions

The effects of inoculation method on efficacy of two formulations of Beauveria bassiana strain GHA against Colorado potato beetle larvae were investigated. Under dry greenhouse conditions, ca. 58% mortality was observed among second-instar larvae exposed directly to sprays of B. bassiana conidia, whereas mortality among larvae exposed to similarly treated foliage (either leaf dorsal or ventral surfaces) was <10%. Mortality was ca. 64% among larvae exposed to both direct sprays and foliar spray deposits. Equivalent rates of mortality were observed among larvae treated with a clay-based wettable powder versus an emulsifiable oil-based formulation of B. bassiana conidia; however, this was observed despite application of an approximately 40% greater dose of WP-formulated conidia, indicating greater efficacy of the emulsifiable oil formulation. These results suggest that, under dry conditions, potato beetle larvae do not readily acquire an effective dose of conidia from treated foliage and that development of improved application technologies to more effectively target the larvae may ultimately prove more beneficial than development of formulations with greater foliar persistence.     

A new microbial consortia containing entomopathogenic fungus,Beauveria bassiana and plant growth promoting rhizobacteria,Pseudomonas fluorescens for simultaneous management of leafminers and collar rot disease in groundnut

The virulence of 20 isolates of Beauveria bassiana (Balsamo) Vuillemin to larvae of the leafminer, Aproaerema modicella, was tested in the laboratory. Leafminer larvae were sprayed with a standard concentration of 1×10⁸ condia/mL of each B. bassiana isolate. All the B. bassiana isolates tested were pathogenic to A. modicella and the mortality varied between 16.7 and 68.9%. Beauveria bassiana isolate B2 was found to be the most virulent followed by isolate B4 which resulted in 59% mortality. Beauveria isolate B2 was selected for dose–response mortality studies with four different doses (1×10², 1×10⁴, 1×10⁶ and 1×10⁸ conidia/mL). Among the various doses tested, 1×10⁸ conidia/mL produced the highest mortality (70.7%). In addition, morphogenesis of the insect pest in all stages, larval, pupal and adult was greatly affected due to fungal infection. Further, B. bassiana isolate B2 and two Pseudomonas fluorescens strains, TDK1 and Pf1 were tested alone and in combination for suppression of groundnut leafminer and collar rot disease and promotion of plant growth and yield both under glasshouse and field conditions. The mixture of B. bassiana and P. fluorescens strains significantly reduced the leafminer and collar rot disease incidences when applied as talc-based formulation through seed, soil and foliar application under glasshouse and field conditions.     

Two strains of Pseudomonas fluorescens bacteria differentially affect survivorship of waxworm (Galleria mellonella) larvae exposed to an arthropod fungal pathogen,Beauveria bassiana

Two strains of Pseudomonas fluorescens were found contaminating a biopesticide used in a previous study against Varroa destructor infestations in honey bee hives. In that study, the biopesticide, a formulation of a fungal pathogen of arthropods, Beauveria bassiana, failed to have any negative impact on the mite infestation despite successful results in previous studies using uncontaminated batches of the same biopesticide. The objective of the present research was to determine whether the bacteria may have interfered with the infectivity and/or virulence of B. bassiana in a simplified system; positive results in that system would then provide a rationale for further work under more complex conditions. Galleria mellonella late instar larvae treated topically with both a bacterial suspension of 6.8 to 7.0×10⁷ cfu/ml and a fungal suspension of 2.5×10⁷ or 2.5×10⁸ B. bassiana conidia/ml showed, in the case of one of the bacterial strains, significantly increased survivorship compared to larvae treated with just the B. bassiana suspension. When larvae were immersed in a bacterial suspension prior to application of B. bassiana suspension using a spray tower, a significant positive effect of the same P. fluorescens strain on larval survivorship was observed at 2.5×10⁸ conidia/ml. Neither the bacterial suspensions alone nor blank control solutions had any effect on larval survivorship. These results show that an interaction between the bacteria and the pathogen may explain some of the results from the prior field trial.     

Evaluation of Beauveria bassiana applications against adult house fly, Musca domestica, in commercial caged-layer poultry facilities in New York state

Applications of a commercially produced Beauveria bassiana product, balEnce, were compared with pyrethrin treatments for the control of adult house flies in New York high-rise, caged-layer poultry facilities. An integrated fly management program, which included the release of house fly pupal hymenopteran parasitoids, was used at all facilities. Adult house fly populations were lower in B. bassiana-treated facilities during the spray and post-spray periods, as recorded on spot cards. Concurrently, the numbers of house fly larvae recovered in B. bassiana-treated facilities were less than one-half that of the pyrethrin-treated facilities. House fly pupal parasitism levels were low, but similar under both treatment regimes. The numbers of adult and larval Carcinops pumilio, a predatory beetle, recovered from B. bassiana-treated facilities were 43 and 66% greater than from the pyrethrin-treated facilities, respectively.     

A field study investigating the insecticidal efficacy against Diaphorina citri Kuwayama on Kinnow mandarin,Citrus reticulata Blanco trees

Asian citrus psyllid is a most damaging insect pest of citrus. In this field study, the efficacy of seven insecticides (emamectin benzoate, bifenthrin, chlorfenapyr, fipronil, imidacloprid, pyriproxyfen and thiamethoxam) was evaluated against Diaphorina citri Kuwayama in the citrus orchard of Kinnow mandarin, Citrus reticulata Blanco. The insecticides revealed a differential and substantial relative efficacy against D. citri compared to the untreated plants. The insecticidal effect attributed as percent reduction in insect population was more prominent after three days of spray: highest reduction values were recorded with thiamethoxam (50.89%), imidacloprid (44.27%) and bifenthrin (42.94%) after first spray, and thiamethoxam (83.36%), imidacloprid (73.20%) and bifenthrin (72.66%) after second spray. Thus, neonicotinoids (thiamethoxam and imidacloprid) and pyrethroid (bifenthrin) resulted as highly effective against D. citri at three days after both sprays. At seven days, imidacloprid (63.53%) and fipronil (62.47%) presented relatively higher population reduction after first spray, and thiamethoxam (92.66%) and chlorfenapyr (89.59%) after second spray. At 12 days, the insecticidal effect on insect population became significantly at par after each spray except chlorfenapyr that reflected high population reduction (93.17%) only after second spray. It is also obvious from the data that there is need of regular monitoring to suppress the psyllids population below threshold level by timely application of the second insecticidal spray.     

Efficacy of Oxamyl Against Heterodera schachtii on Cabbage

The efficacy of oxamyl in controlling Heterodera schachtii on cabbage was determined by applying various contbinations of soil drenches at 6.7 kg (a.i.)/ha and foliar sprays at 0.04 kg (a.i.)/100 liters of water to cabbage seedlings. Pretransplant drenches provided some control of H. schachtii over a 13-week period. A single foliar spray of oxamyl 1 week before transplanting apparently prevented penetration of H. schachtii larvae; post-transplant sprays were relatively ineffective. A pretransplant or transplant drench combined with a foliar application 2 weeks after transplanting provided the most effective control. The effectiveness of drenches plus post-transplant sprays is probably due to the spray augmenting the action of the drench in inhibiting the development of larvae after penelration.     

Effects of manipulating spray-application parameters on efficacy of the entomopathogenic fungus Beauveria bassiana against western flower thrips,Frankliniella occidentalis,infesting greenhouse impatiens crops

The effects of various spray application parameters on the efficacy of a clay-based wettable powder formulation of Beauveria bassiana strain GHA conidia against western flower thrips, Frankliniella occidentalis, were evaluated in a series of greenhouse tests. With the aim of optimizing spray application methods to maximize biopesticide efficacy, a series of independent experiments was conducted that varied four spray parameters: application interval, rate, volume, and spray-program timing. Impatiens crops infested with western flower thrips were treated with (1) multiple sprays at the rate of 5×10¹³ conidia in 935 L aqueous carrier ha^?1 applied at 3-, 5-, and 7-day intervals, (2) weekly sprays at rates of 5×10¹³, 1×10¹⁴, and 2.0×10¹⁴ conidia in 935 L carrier ha^?1, (3) weekly sprays at a rate of 2.0×10¹⁴ conidia in volumes of 935, 1870, and 3740 L water ha^?1, and (4) multiple sprays at the rate of 2.0×10¹⁴ conidia in 3740 L carrier ha^?1 applied at 5-day intervals in spray programs initiated before versus after the onset of flowering. Pollen-bearing impatiens flowers were sampled twice weekly to estimate thrips population density, and adult female and second-instar thrips were collected 24 h post application for determination of acquired dose (conidia/insect). Numbers of conidia inoculated onto thrips increased with increasing spray frequency and volume. Dose was unexpectedly not directly correlated with application rate when volume was held constant, suggesting that thrips avoided concentrated spray residues. Statistically significant thrips population reductions relative to controls were achieved only when three to four sprays were applied at the highest label rate in the highest volume at < 7-day intervals. Applications against thrips infesting young, preflowering impatiens crops were not consistently more effective than applications in older crops. The most effective treatment programs reduced pest populations by 30–40% compared to untreated controls; this slowed, but did not stop, the growth of pest populations. Results indicate that use of fungi for thrips management will require integration with other control agents.     

Persistence of spores of Pleistophora schubergi (Cnidospora: Microsporida) in the field,and their application in microbial control

Spores of Pleistophora schubergi, when applied to oak trees in the field at 2 × 10⁸ spores/ml with a uv protectant, “Shade,” infected 88% of Anisota senatoria larvae at 4 days after spray application. Spores without the uv protectant infected only 10% of the larvae at 4 days after application. When the spores were applied at the rate of 2 × 10⁸ and 2 × 10⁷ spores/ml in the field, 96 and 72% of the A. senatoria larvae and 100 and 100% of the Symmerista canicosta larvae were infected 14 days after spray application.     

Sensitivity of the Entomogenous Fungus Beauveria bassiana to Selected Plant Growth Regulators and Spray Additives

Mefluidide was the only one of four plant growth regulators that caused little to no significant inhibition of in vitro germination and growth of the entomogenous fungus Beauveria bassiana. Silaid, paclobutrazol, and flurprimidol significantly inhibited germination and growth. Mortality of fall armyworm, Spodoptera frugiperda, resulting from B. bassiana was significantly reduced when larvae were exposed to conidia plus soil treated with paclobutrazol. Larval mortality resulting from conidia plus soil treated with mefluidide did not differ significantly from mortality resulting from untreated conidia. Triton CS-7 was the only one of eight spray adjuvants that significantly inhibited germination of B. bassiana conidia.     

Oosporein,an abundant metabolite in Beauveria caledonica,with a feedback induction mechanism and a role in insect virulence

Oosporein was first identified from the insect pathogen Beauveria bassiana >50 y ago. Here, we investigate the insecticidal, anti-feedant and immunomodulation effects of oosporein produced by Beauveria caledonica on the forestry pest Hylobius abietis and model insect Galleria mellonella. We report a novel feedback induction mechanism regulating oosporein production in B. caledonica; exogenous oosporein induces the expression of the oosporein cluster, leading to increased abundance of oosporein biosynthetic enzymes, as shown by label-free quantitative proteomics. Oosporein did not have an anti-feedant effect on H. abietis adults – on the contrary, insects exposed to oosporein-treated food fed more than those exposed to untreated food only. Injected oosporein did not kill insect larvae but increased susceptibility of H. abietis to a subsequent infection. Oosporein did not act as a contact toxin on H. abietis adults and G. mellonella larvae at the concentrations tested. Therefore, it appears that oosporein promotes infection rather than directly killing insects; this could be mediated both by a reduction in haemocyte numbers and by alterations to the humoral immune system. This work makes a case for future research into the potential use of B. caledonica as a biocontrol agent through combinations with oosporein or with enhanced production of oosporein.     

Potential of an indigenous strain of the entomopathogenic fungus Beauveria bassiana as a biological control agent against the Red Palm Weevil, Rhynchophorus ferrugineus

The potential of a strain of Beauveria bassiana (Ascomycota: Clavicipitaceae) obtained from a naturally infected Rhynchophorus ferrugineus (Coleoptera: Curculionidae) pupa as a biological control agent against this weevil was evaluated both in the laboratory and in semi-field assays. Laboratory results indicate that this strain of B. bassiana can infect eggs, larvae and adults of R. ferrugineus (LC₅₀ from 6.3 × 10⁷ to 3.0 × 10⁹ conidia per ml). However, mortality was not the only indicator of treatment efficacy because adults of either sex inoculated with the fungus efficiently transmitted the disease to untreated adults of the opposite sex, with male-to-female and female-to-male rates of transmission of 55% and 60%, respectively. In addition, treatment with B. bassiana significantly reduced fecundity (up to 62.6%) and egg hatching (32.8%) in pairing combinations with fungus-challenged males, females or both sexes. Likewise, 30-35% increase in larval mortality was observed in larvae obtained from eggs from fungus-challenged females or from untreated females coupled with inoculated males, resulting in an overall 78% progeny reduction. Semi-field preventive assays on potted 5-year old Phoenix canariensis palms, with efficacies up to 85.7%, confirmed the potential of this strain as a biological control agent against R. ferrugineus.     

Role of cuticle-degrading enzymes of Beauveria bassiana and Metarhizium anisopliae in virulence on Plodia interpunctella (Lepidoptera,Pyralidae) larvae

Entomopathogenic fungi (EPF) are important biological control agents in pest management programs in agroecosystems against insect pests. EPF such as Beauveria bassiana (Bals.) Vuillemin and Metarhizium anisopliae (Metchn.) Sorokin produce a wide range of extracellular enzymes involved in disturbance of the first barrier in the insect cuticle comprising proteins, chitin, and lipids. Realizing relationships between the expression of these enzymes and fungal virulence might aid in development of effective mycoinsecticides. The virulence of B. bassiana (isolates TV and OZ1) and M. anisopliae (isolate CS1) were investigated on Plodia interpunctella (Hübner) larvae in this study. The third instar larvae were immersed in a suspension containing 1 × 10⁸ conidia mL^?1 of fungal conidial inoculum. The results revealed that all three fungal isolates caused mortality in larvae, but there was a considerable variation in their virulence. Total proteinase, exochitinase and lipase assays were done for these isolates. The TV isolate with the highest mortality with 41.7%, had the highest level of specific activity of exochitinase, protease, and lipase with 0.148, 0.654, and 0.190 U. mg^?1 protein, respectively. In the current study, a positive correlation was determined between the virulence of fungal isolates and the activities of protease and lipase, but this link was not significant for exochitinase. Our results demonstrated that extracellular enzymes, particularly protease and lipase, may play a crucial role in the virulence of these fungal isolates against P. interpunctella larvae.     

Comparative laboratory studies on three fungal pathogens of the elm bark beetle Scolytus scolytus: Pathogenicity of Beauveria bassiana,metarhizium anisopliae, and Paecilomyces farinosus to larvae and adults of S. scolytus

The entomogenous fungi Beauveria bassiana (nine isolates), Metarhizium anisopliae (seven isolates), and Paecilomyces farinosus (four isolates) were tested as pathogens of larvae of the elm bark beetle, Scolytus scolytus. Single isolates of B. bassiana and M. anisopliae were also tested against adult beetles. Of the 21 isolates tested as conidial suspensions against larvae, all proved pathogenic. The three most and least virulent isolates were, respectively, isolates of B. bassiana and M. anisopliae. The other isolates fell between these two extremes, with the four P. farinosus isolates all moderately virulent. Spore retention on larvae following inoculation was estimated by washing conidia off the larvae. From the results it was possible to relate larval mortality to the approximate spore dose causing infection at different spore concentrations. Thus, application of spores of the three pathogens at a concentration of 10³ spores/ml resulted in limited mortality. At this concentration, an average of only a single spore was recovered from the inoculated larva. Adult bark beetles also proved susceptible to infection by isolates of B. bassiana and M. anisopliae. They were exposed to discs of elm bark dipped in a conidial suspension. It was estimated that a dose of less than 100 spores could cause infection of beetles following feeding on the elm bark discs.     

Efficacy of entomopathogenic fungi and nematodes againstArgyresthia conjugella (Lep.: Yponomeutidae)

The efficacy ofPaecilomyces fumosoroseus (Wize) Brown & Smith,P. farinosus (Holm ex S. F. Gray) Brown & Smith,Beauveria bassiana (Balsamo) Vuillemin,Metarhizium anisopliae (Metschnikoff) Sorokin and an entomoparasitic nematode,Steinernema feltiae (Filipjev), against larvae and pupae of the apple fruit mothArgyresthia conjugella (Zell.) (Lepidoptera: Yponomeutidae) was studied in field and laboratory experiments. All species of fungi and the nematode were highly pathogenic to larvae in Petri dishes. In soil, the pathogenicity ofB. bassiana andP. farinosus for larvae decreased. Pupae inside cocoons were highly susceptible toP. fumosoroseus, but not toP. farinosus orB. bassiana. In the field experiment, treatment of soil with conidia ofP. fumosoroseus at a rate of 1×10⁷ conidia per cm² reduced the emergence of overwinteredA. conjugella adults by 70.1% compared to untreated control soil.     

Can Insects Develop Resistance to Insect Pathogenic Fungi?

Microevolutionary adaptations and mechanisms of fungal pathogen resistance were explored in a melanic population of the Greater wax moth, Galleria mellonella. Under constant selective pressure from the insect pathogenic fungus Beauveria bassiana, 25^th generation larvae exhibited significantly enhanced resistance, which was specific to this pathogen and not to another insect pathogenic fungus, Metarhizium anisopliae. Defense and stress management strategies of selected (resistant) and non-selected (susceptible) insect lines were compared to uncover mechanisms underpinning resistance, and the possible cost of those survival strategies. We hypothesize that the insects developed a transgenerationally primed resistance to the fungus B. bassiana, a costly trait that was achieved not by compromising life-history traits but rather by prioritizing and re-allocating pathogen-species-specific augmentations to integumental front-line defenses that are most likely to be encountered by invading fungi. Specifically during B. bassiana infection, systemic immune defenses are suppressed in favour of a more limited but targeted repertoire of enhanced responses in the cuticle and epidermis of the integument (e.g. expression of the fungal enzyme inhibitor IMPI, and cuticular phenoloxidase activity). A range of putative stress-management factors (e.g. antioxidants) is also activated during the specific response of selected insects to B. bassiana but not M. anisopliae. This too occurs primarily in the integument, and probably contributes to antifungal defense and/or helps ameliorate the damage inflicted by the fungus or the host’s own immune responses.     

Identification of highly pathogenic Beauveria bassiana strain against Pieris rapae larvae

Seven different strains of Beauveria bassiana were used in a bioassay on Pieris rapae larvae. The results showed that an B. bassiana strain showed relatively high pathogenicity towards P. rapae larvae. The adjusted mortality rate was 92.86 %, and the infection rate was 85.71 % in 10 days post inoculation. Molecular identification was performed to identify the unknown strain. Internal Transcribed Spacer sequence analysis showed that the polymerase chain reaction amplicon length of the unknown strain of Beauveria sp. was 573 bp, and sequence similarity to the known B. bassiana sequences in the NCBI database was 99 %. The B. bassiana strain was named Bb01. The changes of proteins and PPO of P. rapae larvae infected by B. bassiana Bb01 strain at different times was determined. The activity of PPO increased in 1–6 d and decreased in 7 d again after inoculation. The B. bassiana invaded into the insect body affected the balance of the proteins and PPO.     

Time and concentration dependent interactions of Beauveria bassiana with sublethal rates of imidacloprid against the aphid pests Macrosiphoniellasanborni and Myzus persicae

A series of laboratory bioassays with each consisting of low, medium and high concentration treatments of the fungal biocontrol agent Beauveria bassiana alone or supplemented with an increasing sublethal rate of imidacloprid were conducted to quantify the fungal and chemical interactions on chrysanthemum aphid Macrosiphoniella sanborni (0.01‐0.05 a.i. μg mL^?1) and green peach aphid Myzus persicae (0.05‐0.5 a.i. μg mL^?1). During one week after exposure to a 1 mL spray onto a 95 cm² area in a Potter Spray Tower, M. sanborni was either more susceptible to B. bassiana or more sensitive to imidacloprid than M. persicae. The time–concentration–mortality (TCM) responses of each aphid species in each of five bioassays fit well to a TCM model, indicating a strong dependence of the fungal and chemical interactions on both concentration and post‐spray time. Adding imidacloprid to B. bassiana sprays at the rates of 0.025–0.05 μg mL^?1 against M. sanborni or 0.1–0.5 μg mL^?1 against M. persicae significantly enhanced or accelerated the fungal action. Based on the LC₅₀ or LC₉₀ estimates and their variances determined by the fitted TCM relationships for each aphid species, the relative potencies of an imidacloprid‐inclusive bioassay over those with B. bassiana alone or together with a lower sublethal rate ranged from a few to hundreds of times and varied over days after spray. These results suggest an alternative tactic for practical control of the aphid pests by a combined formulation or application of B. bassiana and imidacloprid and manage aphid resistance to the chemical insecticide.     

Growth and insect assays of Beauveria bassiana with neem to test their compatibility and synergism

Beauveria bassiana is being used as a biopesticide for many insect pests. Neem oil (azadirachtin) is an eco-safe popular botanical pesticide. A biopesticde with a neem compatible isolate of B. bassiana will enable their simultaneous use in pest management. A sample of 30 isolates of B. bassiana from culture collections was screened for compatibility with a commercial formulation of neem oil (Margoside®) at the field recommended dose (0.3%, v/v). Compatibility was tested in vitro through germination and growth assays. In all isolates, conidial germination was delayed but not significantly decreased by neem. In the growth assays, 23 isolates were found compatible with neem. In the neem sensitive isolates, growth was decreased but not totally inhibited. The effect of combined treatment with B. bassiana and neem in comparison to single treatments with either of them on Spodoptera litura Fabricius was tested in laboratory bioassays. The combined treatment was found to have synergistic effect on insect mortality when a B. bassiana isolate compatible with neem was used, while, with an isolate sensitive to neem, an antagonistic effect was observed.     

Susceptibility of preimaginal western cherry fruit fly, Rhagoletis indifferens (Diptera: Tephritidae) to Beauveria bassiana (Balsamo) Vuillemin Clavicipitaceae (Hypocreales)

Last-instar larvae of the western cherry fruit fly, Rhagoletis indifferens, were subjected to Beauveria bassiana GHA incorporated into sterile sand and non-sterile orchard soil. Mycosis in the pupal stage was observed in >20% of buried R. indifferens pupae and >80% of larvae entering sand treated with either of two B. bassiana isolates. When pre-pupal larvae burrowed into conidium-treated non-sterile cherry orchard soil, the incidence of mycosis, on both the puparia and internally developing pupae, increased with dose. Internal pupal tissues were found to contain B. bassiana. Increasing the soil moisture level from 20% to 35% water holding capacity did not have an effect on the percentage of mycosed pupae. This is the first evidence that the preimaginal stages of R. indifferens are susceptible to infection by B. bassiana.