Evaluation of Beauveria bassiana and Metarhizium anisopliae for Controlling Chilo partellus (Lepidoptera: Crambidae) in Maize

Maize (variety Katumani) was planted in a greenhouse and plants were infested with 20 Chilo partellus second instar larvae 3 and 4 weeks after plant emergence. One isolate of Beauveria bassiana (BB-01) and four isolates of Metarhizium anisopliae (PPRC-4, PPRC-19, PPRC-61 and EE-01) were sprayed onto the leaf whorl at 2×10⁸ conidia/mL 24 h after infestation. Leaf damage by the larvae was greatly reduced by the treatments. The mean daily temperature and relative humidity in the greenhouse ranged from 10 to 35°C and 30 to 90%, respectively. The growth of infested unsprayed plants was less than that of fungi treated plants. Stem tunneling (1-5%), deadheart (0-33%), number of attacked nodes (0.3-2.5) and holes (0.2-3.3) were also reduced in plants sprayed with conidial suspensions. Isolates PPRC-4, PPRC-19 and PPRC-61 seemed to be the best candidates for further development.     

Effects of Fluctuating Moisture and Temperature Regimes on Sporulation of Beauveria bassiana on Cadavers of Rhodnius prolixus

The effects of both moisture and temperature on the sporulation of an isolate of Beauveria bassiana , highly virulent to the Chagas' disease vector, Rhodnius prolixus , were studied in the laboratory under fluctuating regimes. Quantitative assays for conidial production from funguskilled cadavers from 10-day-old tests showed that recycling was highly dependent on the duration of exposure to high relative humidity (RH). On first-instar larval cadavers, sporulation reached more than 10 5 conidia/nymph under most regimes with a favourable phase combining high humidity (97% RH) and temperatures of 20 or 25 C for at least 12 h/day. Sporulation on third- and fifth-instar nymphs required a longer period (16 h/day or more) of favourable conditions (97% RH and 25 C) per day. The intensity of the conidial production from Rhodnius cadavers was enhanced when diurnal temperatures were high (28 and 35 C). From a vector control standpoint, daily high humidity requirements appear to be a crucial constraint. The recycling ability of B. bassiana on fungus-killed R. prolixus is most likely to contribute to the regulation of this triatomine vector if applications are made during the rainy seasons and in the most favourable habitats.     

Infection of the Coffee Berry Borer Hypothenemus hampei (Coleoptera: Scolytidae) by Brazilian Isolates of the Entomopathogenic Fungi Beauveria bassiana and Metarhizium anisopliae (Deuteromycotina: Hyphomycetes)

The virulence of four fungal isolates (three Beauveria bassiana and one Metarhizium anisopliae ) against adult female coffee berry borers (CBB) was investigated. The most virulent isolate from initial bioassays, B. bassiana LPP1, with a LT 50 of 3.4 days, was further investigated by application to berries prior to infestation and to berries already infested with CBB. At the highest concentration applied to berries (1 ×10 7 conidia mL -1 ), CBB mortality was 83% (berries inoculated prior to infestation) and 62% (berries inoculated after infestation).     

Laboratory and semi-field evaluation of Beauveria bassiana (Ascomycota: Hypocreales) against the lettuce aphid,Nasonovia ribisnigri (Hemiptera: Aphididae)

The lettuce aphid, Nasonovia ribisnigri (Mosley), is an economically important pest of lettuce worldwide. The entomopathogenic fungus Beauveria bassiana strain GHA has recently been reported as a potential biocontrol candidate for use against the lettuce aphid. This study provides information on the mortality inflicted by B. bassiana when applied against different life stages of the lettuce aphid under laboratory conditions and how fungus infection affects the aphid fecundity. In addition, temporal changes in persistence of fungus inoculum applied to foliage of young lettuce plants under semi-field conditions was analysed. Immature life stages were generally the least susceptible to fungal infection and the susceptibility of all stages was dose-dependent, with the highest mortality occurring at the highest dose. B. bassiana significantly affected the rate of nymph production by the lettuce aphid, with the highest effect seen when the alatoid fourth instar of N. ribisnigri was inoculated with B. bassiana. The persistence of B. bassiana conidia on lettuce foliage was not influenced by leaf position. Within 5 days, the cumulative percentage decline in the conidial population was 38% which declined further to 92% and 99% on day 11 and 20 post-spraying, respectively. In accordance, the infectivity to second instar lettuce aphid nymphs of B. bassiana conidia deposited on leaves declined according to an exponential decay model predicting an intercept of 0.59 ± 0.03 (S.E), a reduction in aphid mortality at a rate of 11% with each increasing day after fungal application and a fungus half-life of 6.34 ± 0.69 days.     

Laboratory Evaluation of the Pathogenicity of Three Isolates of the Entomopathogenic Fungus Beauveria bassiana (Bals.) Vuillemin on the American Cockroach (Periplaneta americana)

Mycopesticides can be ideal for the biocontrol of cockroaches because the habitat of these insects promotes initial fungal infection and its subsequent spread. The pathogenicity of three isolates of Beauveria bassiana to the American cockroach was tested. The insects were treated in three different ways, by direct contact with spore mass, a spore-wheat flour mixture and a spray of an aqueous spore suspension. A mortality of 100% in the first treatment, 67-100% in the second treatment and 17-75% in the third treatment was observed. These results suggest that B. bassiana spore formulations in food baits can be developed for cockroaches.     

Potential of Metarhizium anisopliae and Beauveria bassiana to control Dinoderus porcellus (Coleoptera: Bostrychidae) infesting yam chips

The beetle Dinoderus porcellus Lesne is a serious storage insect pest that causes important losses by destroying stocks of yam chips. In the aim to found an alternative control method to the use of synthetic insecticides for its management, the virulence of the entomopathogenic fungi Beauveria bassiana (Balsamo) Vuillemin (isolate Bb115) and Metarhizium anisopliae (Metschnikoff) Sorokin (isolate Met 31) against adults of D. porcellus was evaluated under laboratory conditions (25 ± 2 °C and 70 ± 5% RH). Then, the effectiveness of the most virulent entomopathogenic fungus as biological agent against D. porcellus was assessed under farmer storage conditions. For each entomopathogenic fungus isolate, four conidial concentration (0, 10⁵, 10⁷, and 10⁹ conidia/mL) at the dose of 1 µL were inoculated topically on D. porcellus adults (3–5 days old). Observations focused on insect mortality, cadaver sporulation and weight loss of yam chips. Lethal dose and lethal time values were estimated using probit analysis. Both fungal isolates at all conidial dose caused more than 50% mortality on day 7, with the highest mortality (94.44%) achieved using B. bassiana at the 10⁹ conidia/mL. LT₅₀ values for B. bassiana and M. anisopliae isolates were 2.63 and 3.35 days, respectively, while their LT₉₀ values were 6.15 and 9.87 days, respectively. Yielding the lower LD₉₀ values and the highest rates of cadaver sporulation, B. bassiana isolate appeared as the most virulent against D. porcellus. After 3 months of storage, comparatively to the control, the B. bassiana isolate at the highest conidial dose (10⁹ conidia/mL) significantly reduced D. porcellus populations, and weight loss of yam chips. This study revealed the potential of B. bassiana and M. anisoplae isolates as biological control agent against D. porcellus for yam chips protection.     

Generation of Benomyl Resistant Beauveria bassiana Strains and Their Infectivity Against Helicoverpa armigera

Beauveria bassiana transformants were obtained by conventional protoplasting and transformed by eletroporation and polyethylene glycol (PEG) treatment. These displayed mitotic stability in Beauveria bassiana . Strains transformed with pSV50 harbouring the β-tubulin gene of Neurospora crassa grew well on benomyl concentrations of 10 μg ml -1 unlike the recipient strain. The transformants were mitotically stable on either selective or non-selective medium. The efficiency of transformation by linear and circular pSV50 cosmid was 8 and 10 transformants per mug DNA per ml viable protoplast by electroporation, respectively, and 4 and 6 by the protoplast PEG method, respectively. Southern blot and hybridization of undigested fungal DNA of wild type and four transformants, probed with β-tubulin sequence of pSV50, showed hybridization at high M r region of genomic DNA in four transformants, whereas in wild type genomic DNA, no homology of the sequence was observed. Digested genomic DNA, of four transformants gave a complex hybridization pattern. Virulence tests of the transformants showed that there was no significant loss in the pathogenicity toward Helicoverpa armigera third instar larvae. This method of transformation should prove useful with entomopathogenic fungal species in which a genetic transformation system has not yet been established.     

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.     

Selection of a Highly Virulent Fungal Isolate,Metarhizium anisopliae Ma TB 101, for Control of Taro Beetle,Papuana uninodis (Coleoptera: Scarabaeidae)

Fungal isolates (31 Metarhizium anisopliae var. anisopliae , five M. anisopliae var majus , three Beauveria bassiana and four B. brongniartii ) originating from a wide range of geographical locations, insect species and soil types were tested against Papuana uninodis (Coleoptera: Scarabaeidae). In a first test series, spores were applied topically to third-instar larvae and adults. The most effective strain against P. uninodis larvae and adults was Ma TB 101, a M. anisopliae isolate from adult P. woodlarkiana . For adults, strain Ma FI 384, a M. anisopliae from Popillia japonica , was almost equally effective. The 11 most effective isolates (nine M. anisopliae var. anisopliae , one B. brongniartii and one B. bassiana ) with LT values of less than 6 weeks in 50 adults and/or less than 4 weeks in larvae were tested for their efficacy against adults and larvae of P. uninodis by application of spores to soil (107 spores/g). Ma TB 101 was significantly more effective against both adults and larvae (LT ca. 10 days) than all other isolates (LT > 50 50 3 weeks). Two other M. anisopliae strains, Ma F 248 from soil and Ma FI 384 from P.japonica , were more effective than most isolates in adults. The latter three M. anisopliae isolates were tested in a concentration series against third-instar larvae and adults. Mortality was concentration related. Isolates Ma F 248 and Ma FI 384 did not achieve 50% mortality within the test period at concentrations lower than 107 spores/g of soil or feed. For concentrations of Ma TB 101 from 1 107 to 2 105 spores/g the LT ranged from 13 to 30 days in adults and 12 to 24 50 days in third-instar larvae. Accordingly, concentrations causing 50% mortality (LC ) for Ma 50 TB 101 were significantly lower than for the two other M. anisopliae isolates tested.     

Effects of Temperature, Humidity and Inoculation Method on Susceptibility of Schistocerca gregaria to Metarhizium flavoviride

The effects of temperature on conidial germination and susceptibility of adults of the desert locust , Schistocerca gregaria, to four isolates of Metarhizium flavoviride were determined . In addition , the effects of inoculation method (topical or spray) , spore carrier (oil or water) and ambient relative humidity (RH) on susceptibility of the locust to the most thermotolerant isolate (Mf324) were investigated . There were differences among the isolates in the effects of temperature on germination of conidia after a 24 - h incubation period . Over 90% of conidia of all isolates germinated after 24 h at 30 o C . In contrast , at 40 o C , none of the isolates germinated for up to 72 h . However , there were differences in germination between the isolates at 35 o C . Locust mortality and disease progression were significantly affected by temperature . At both 25 and 30 o C , all isolates induced 98 - 100% mortality within 8 days; however , there were differences between isolates at 35 o C . None of the isolates caused significant mortality at 40 o C . Humidity and inoculation method had no influence on levels of mortaility in fungus treated locusts . In contrast , carrier type significantly affected cumulative mortality . Topical oil treatment resulted in higher overall mycosis than the three other treatments . Control mortality on the other hand was strongly affected by inoculation method and to a lesser extent by humidity and carrier . In these conditions , application of oil by spray was generally toxic at all humidities whereas topical application of water was most toxic at near saturation . The results of these laboratory studies demonstrate the importance of strain selection , formulation and application method in the development of a microbial control agent against the desert locust . Low RH should not impede use of this fungus under dry conditions .     

Sporulation of Metarhizium anisopliae and Beauveria bassiana on Coptotermes formosanus and in vitro

The sporulation of 22 total isolates of Metarhizium anisopliae and Beauveria bassiana was quantified on cadavers of the Formosan subterranean termite, Coptotermes formosanus. Conidial production increased significantly over 11 days post-death. Effects of isolates of M. anisopliae and B. bassiana on in vivo sporulation were significant. Although the overall effects of fungal species on in vivo sporulation were not significant, the interactions between fungal species and certain times post-death were significant, indicating different sporulation patterns between the two fungal species. B. bassiana isolates could be categorized into a group with high total sporulation (day 11) and low quick sporulation (on days 2 and 3), while M. anisopliae isolates fell into another group with high quick sporulation and low total sporulation. This could give M. anisopliae an advantage over B. bassiana in termite microbial control due to termite defensive social behaviors. Conidial production was significantly higher in vitro than in vivo. In vitro and in vivo sporulation differed by as much as 89x and 232x among the selected isolates of B. bassiana and M. anisopliae, respectively. Correlation between in vivo and in vitro conidial production was positive and significant. This may allow preliminary in vitro screening of a large number of isolates for high in vivo sporulation.     

白僵菌、绿僵菌对雷公藤丽长角巢蛾的侵染力测定

经室内侵染力测定,球孢白僵菌Beauveria bassiana和金龟子绿僵菌Metarhizium anisopliae皆能侵染雷公藤丽长角巢蛾Xyrosaria lichneuta幼虫,不同菌株的侵染力存在显著差异。3株白僵菌中,Bb27菌株毒力最强,菌液为1×10^5～1×10^9个·mL^-1五个不同浓度梯度的孢子悬浮液时,害虫15d校正死亡率在58．3％-100％之间;LT50分别为9．36d,6．26d,3．89d、2．54d．2．10d;LC50为4．87×10^4个·ML^-1。3株绿僵菌中,MaZPTP01菌株毒力最强,5种孢子浓度15d校正死亡率分别为52．4％、69．0％、76．2、92．9％、95．2％;LT50分别为11．47d、6．89d、4．36d、2．68d、2．13d;LC50为7．39×10^4个·mL^-1。Bb27和MaZFIT01菌株可应用于林间作进一步防治试验。     

Diversity of indigenous Beauveria and Metarhizium spp. in a commercial banana field and their virulence toward Cosmopolites sordidus (Coleoptera: Curculionidae)

Metarhizium anisopliae and Beauveria bassiana sensu lato were isolated, from 7 and 41 % of soil samples from a commercial banana field, with average fungal density of 4.3 × 10³ and 8.2 × 10³ CFU g^?1 soil, respectively. Twenty-one morphologically distinct B. bassiana and four M. anisopliae sensu lato isolates from different plots within the field were further characterized. ISSR fingerprinting revealed six different clusters for B. bassiana, whereas gene sequencing revealed three M. anisopliae sensu stricto and one unclassified Metarhizium sp. Bioassays with one or more representative isolates from each Metarhizium species and B. bassiana cluster showed that all indigenous isolates had lower virulence and significantly greater ST₅₀s than reference (exotic) isolates. The data suggest that the low virulence of most indigenous isolates toward Cosmopolites sordidus adults and their relatively low density in soil samples, may help explain the low occurrence of epizootics caused by entomopathogenic fungi in populations of this pest, also known to burrow under the soil surface in banana plantations.     

Susceptibility of the Hymenopteran Parasitoids Apoanagyrus ( = Epidinocarsis ) lopezi (Encyrtidae) and Phanerotoma sp. (Braconidae) to the Entomopathogenic Fungus Metarhizium anisopliae var. acridum (Deuteromycotina: Hyphomycetes)

Laboratory and so-called extended laboratory bioassays were conducted in Benin, West Africa, to investigate the pathogenicity and virulence of the entomopathogenic fungus Metarhizium anisopliae var. acridum , a biocontrol agent against locusts and grasshoppers, to two hymenopteran parasitoids, Apoanagyrus ( = Epidinocarsis ) lopezi and Phanerotoma sp. Treatments were carried out under simulated field conditions at standard field dose rates of 2.5 and 5.0 ×10 12 conidia ha -1 . Test organisms were continuously (3 weeks) exposed to spray residues in artificial or simulated natural environments. The standard strain IMI 330 189 of the mycopesticide Green Muscle caused a significant reduction of 24% in the longevity ( = average survival time, AST) of A. lopezi , relative to the untreated control. Mycosis was confirmed in 16% of all cadavers. AST was shorter under low relative humidity (RH) conditions, and these conditions seemed to enhance susceptibility to fungal infection. However, this effect was only marginally significant. In contrast, average longevity of untreated A. lopezi was slightly yet significantly shorter at low RH (50-60%) than at high RH (80-90%). In the extended laboratory assay, the same isolate had no significant effect on mortality, parasitoid emergence ( = beneficial capacity) and sex ratio. In a further screening test with isolates IIBC I91 609, IIBC I93 833 and IMI 330 189 (reference), no infection of A. lopezi was confirmed. Similarly, Phanerotoma sp. was not susceptible to IMI 330 189. It is concluded that mycopesticides based on the three strains tested pose a low risk to parasitic hymenopterans under field conditions.     

Laboratory and Field Trials of Bait Formulations of the Fungal Pathogen, Metarhizium flavoviride, Against a Tropical Grasshopper and Locust

Extrusion technology was used to produce maize-starch 'contact bait' carriers, and vegetable-oil suspensions of Metarhizium flavoviride were incorporated into the baits after extrusion. Laboratory bioassays using the locust, Schistocerca gregaria, demonstrated mortality and reduction of 8 1 feeding after exposure to the baits, and gave an LC 50 of 1.5 10 spores l slope 0.63 for fifth instar nymphs. In laboratory trials, maize-starch contact baits showed improved efficacy relative to wheat-bran edible bait formulations of M. flavoviride. Field cage experiments, undertaken in Mali, using fourth and fifth instar nymphs of the grasshopper, Hieroglyphus dagenensis, demonstrated significant mortality with both fresh and weathered maize-starch contact bait formulations of M. flavoviride, relative to untreated controls.     

Comparative impact of artificial selection for fungicide resistance on Beauveria bassiana and Metarhizium brunneum

Hypocreales fungi such as Beauveria bassiana (Balsamo) Vuillemin and Metarhizium brunneum Petch can be negatively affected by fungicides thereby reducing their biocontrol potential. In a previous study, we demonstrated enhanced fungicide resistance in B. bassiana through artificial selection. However, it is not clear if the enhanced resistance was because of improved germination, vegetative growth, or both. Additionally, the enhanced fungicide resistance has only been demonstrated in B. bassiana, and therefore it is of interest to investigate the potential to enhance resistance in other fungi. Thus, the objectives in this study were to determine the potential to enhance fungicide resistance in M. brunneum through artificial selection, and investigate if selection is based on germination, vegetative growth, or both in B. bassiana and M. brunneum. Selection for resistance to fenbuconazole, and triphenyltin hydroxide was assessed through inhibition evaluations on solid media, and germination and mycelial growth in liquid media. Increased resistance after selection was observed for all fungicide-fungus combinations on solid and or liquid media. Selection resulted in increased resistance to fenbuconazole in both fungi in solid and liquid media; in liquid culture fungicide resistance in B. bassiana was manifested by increased germination and mycelial growth, whereas in M. brunneum fungicide resistance concerned only mycelial growth. Selection for resistance to triphenyltin hydroxide varied in the different media. For B. bassiana, triphenyltin hydroxide resistance was enhanced on solid media but not in liquid, whereas enhanced resistance of M. brunneum was detected in both media. Fungicide sensitivity and selection potential differs based on the medium and fungal species. Selection for fungicide resistance, had negative effects on other beneficial traits when fungicide pressure was removed, for example, some selected populations showed decreased germination or growth, relative to their nonselected control populations. Additionally, reduced virulence to the greater wax moth, Galleria mellonella (L.), was observed in all fungal populations that were exposed to fungicide resistance regimes. We conclude that it is possible to use genetic selection to enhance fungicide resistance in B. bassiana and M. brunneum, but before use the resulting populations should be screened for inadvertent negative impacts on beneficial traits.     

寄主和分布区系不同的球孢白僵菌菌株近缘关系的研究

观察、测定了来源不同的球孢白僵菌 1 9个菌株的培养性状、产孢量、萌发中时、水分活性、紫外照射活率、水浴活率等指标 ,将培养性状编码后 ,由所得的 8个指标对供试菌株求相关系数矩阵 .分析表明 ,不同菌株近缘关系的远近主要由菌株原始分布区系距离远近所决定 ,受寄主不同的影响次之 .来源于相同 (或相近 )地区的菌株相关系数可达极显著水平 ,而从相同寄主角度分析 ,得到菌株间的相关系数有高有低 ,多表现为相关不显著     

Field Treatment of Desert Locust ( Schistocerca gregaria Forskal) Hoppers in Mauritania Using an Oil Formulation of the Entomopathogenic Fungus Metarhizium flavoviride

A field trial was conducted to test the efficacy of an oil formulation of the entomopathogenic fungus , Metarhizium flavoviride Gams & Rozsypal (Deuteromycotina: Hyphomycetes) , as a control agent for the desert locust , Schistocerca gregaria Forskal (Orthoptera: Acrididae) , in Mauritania . The treatment was applied to hopper bands in the field . In caged samples of the treated insects , 99% mortality was observed in 15 days . On average during application only 51% of the hoppers received spray droplets directly , the additional mortality being due to pick - up of spores from the spray residue . This mortality was assessed by exposing insects to a treated plot at intervals , and monitoring disease levels during subsequent incubation in cages . The spray residue remained highly infective for a period of 6 days after application . The number of nymphs in the hopper bands was estimated using a computer - based photographic image - processing technique . Unfortunately , most of the hopper bands dispersed into small groups , but a population reduction could be demonstrated for one hopper band . Maximal daily mortality in the field was observed 10 - 11 days after application , compared with 6 - 10 days in cages .     

Effect of temperature on virulence of Beauveria bassiana and Metarhizium anisopliae isolates to Tetranychus evansi

The virulence of three isolates of Beauveria bassiana (Bals.) Vuill. and 23 isolates of Metarhizium anisopliae (Metschnik.) Sorok. (Ascomycota: Hypocreales) against the tomato spider mite, Tetranychus evansi Baker and Pritchard (Acari: Tetranychidae), was assessed in the laboratory. The effect of temperature on germination, radial growth and virulence of selected isolates (two isolates of B. bassiana and nine of M. anisopliae) on T. evansi was also investigated in the laboratory. All the fungal isolates tested were pathogenic to the adult females of T. evansi, and there were significant differences in mortality between fungal isolates. The lethal time to 50% mortality (LT(50)) values ranged from 4.2 to 8.1 days and the LT(90) values from 5.6 to 15.1 days. Temperature had significant effects on germination, radial growth and virulence of the various isolates. The best fungal germination was observed at 25 and 30 degrees C, while for the fungal radial growth it was 30 degrees C. All the isolates germinated and grew at all temperatures, but germination and radial growth varied with isolate and temperature. The selected isolates were all virulent to T. evansi, but virulence varied also with isolate and temperature.