Virulence potential of some fungal isolates and their control-promise against the Egyptian cotton leaf worm,Spodoptera littoralis

A preliminary virulence test of four fungal isolates, Beauveria bassiana IMI 382302, Beauveria bassiana IMI 386701, Trichoderma harzianum T24 and Aspergillus flavus Link against larvae of Spodoptera littoralis was performed. The most effective isolates against larvae of S. littoralis were B. bassiana 302 and T. harzianum T24, which also showed the lower percentage of pupation compared with the other two isolates under the same conditions of treatments. Three concentrations (1 × 10⁶, 1 × 10⁷ and 1 × 10⁸ ml^?1) of the aqueous conidial suspension of the four tested isolates were carried out against both larval and pupal stages of S. littoralis within five days post-treatment. T. harzianum T24 showed 80% larval mortality only when applied at the highest conidial concentration, while A. flavus showed 100% pupal mortality only, at all of its conidial concentrations. However, B. bassiana IMI 382302 showed relatively high dose-dependant larval and pupal mortalities, while strain IMI 386701 of B. bassiana showed a very weak mortality against pupae at higher concentrations, and no virulence against larvae was recorded. Enzymatic and antibiosis bioassays of the four fungal isolates showed relatively high activities against Fusarium spp. for most of the tested isolates. Clear zone of enzyme activity on agar plates proportionally increased with increasing the concentration of enzyme substrate and prolongation of the incubation period. Mtabolites produced in the agar culture inhibited the growth of Fusarium spp. and the productivity differed greatly among isolates or strains of the same isolate. Volatile and non-volatile compounds produced by A. flavus Link showed a higher inhibition activity against Fusarium spp. compared with the other fungal isolates. The humoral antifungal response of insect host is relatively high compared to the anti-bacterial one. Injection of larvae with the immune sensitive bacteria Micrococcus luteus (5 × 10³ bacteria/larva) showed a detectable humoral response by 2 h, peaked around 12 h and became hardly detectable by 24 h post-injection. Injection of larvae with conidial suspension (5 × 10³ conidia/larva) from each of the fungal isolates showed humoral antifungal activity against B. bassiana IMI 386701 and A. flavus only. This activity was detectable by 12 h, peaked around 36 h and became hardly detectable by 48 h post-injection. Although the humoral antifungal response was started slowly compared to the antibacterial one, it lasted for longer and enabled larvae to withstand the infection with these immune-sensitive fungal strains. No humoral activity was detected against B. bassiana IMI 382302, although however, weak activity was detected against T. harzianum T24 only at the low conidial concentration but not at the higher one (1 × 10⁸ ml^?1). Thus, this study concludes that larvae of S. littoralis showed immune-dependant sensitivity to T. harzianum T24 and B. bassiana IMI 382302. Therefore, this study may recommend these two fungal isolates as mycoinsecticides in the battle against cotton leaf worm in Egypt. Hence, they have been selected for future comprehensive bioassays in the laboratory under conditions similar to that in the field. This, in fact, may help for developing effective mycoinsecticides against this pest. Penetration mechanims of insect cuticle by entomopathogenic fungi will be discussed.     

Entomopathogenic fungus isolates for adult Tuta absoluta (Lepidoptera: Gelechiidae) management and their compatibility with Tuta pheromone

Tomato, Solanum lycopersicum, is a widely consumed and economically important vegetable in the tropics. However, its production and productivity have been recently hampered by the invasive Tuta absoluta (Meyrick), a key pest of tomato causing extensive economic damage in Africa. Currently, the commonly used management strategy is synthetic insecticides with hazardous effects on humans and environment. The application of entomopathogenic fungal and viral-based biopesticides is promising alternatives. To identify potent strains, pathogenicity of dry conidia of 12 Metarhizium anisopliae isolates was evaluated against adult T. absoluta. Further effects of the most potent isolates on pupation through the late 4th instar larval infection, adult emergence and horizontal transmission of inoculum were assessed. Metarhizium anisopliae ICIPE 18, ICIPE 20 and ICIPE 665 outperformed the other isolates causing adult mortality of 95.0%, 87.5% and 86.25%, respectively. The three potent isolates differed significantly for LT₅₀ values of 5.13, 3.17 and 2.38 days for ICIPE 18, ICIPE 20 and ICIPE 665, respectively. Pupation and adult emergence were significantly reduced by these potent isolates, which were also compatible with Tuta pheromone (TUA-Optima^®) with ≥90% conidial germination 24 hr post-exposure. These potent isolates could be developed as effective biopesticides and used in combination with TUA-Optima^® for mass trapping and autodissemination for T. absoluta management in solanaceous crop production systems.     

Pathogenicity of Beauveria bassiana isolated from Moroccan Argan forests soil against larvae of Ceratitis capitata (Diptera: Tephritidae) in laboratory conditions

The Mediterranean fruit fly, Ceratitis capitata Wiedemann (Diptera: Tephritidae), is the major tephritid pest in Morocco. This pest survives in Moroccan forests Argania spinosa and continually invades the nearest agricultural areas. Entomopathogenic fungi are an interesting tool for fruit fly control and hold a useful alternative to conventional insecticides. However, primary selection of effective pathogens should be taken in laboratory condition prior to applying them in the field. Here, we used third late instar larvae of C. capitata to investigate the effectiveness of 15 local Beauveria bassiana isolates. Results showed that all isolates were able to infect the larval stage, producing a large mortality rate in puparia ranging from 65 to 95 % and caused significant reduction in adult emergence. The fungal treatments revealed that the mycosis occurred also in adults escaping infection as pupariating larvae. The percentage of mycosed puparia was highest in strain TAM6.2 (95 %) followed by ERS4.16 (90 %), therefore they were the most virulent. Median lethal concentration (LC₅₀) was studied for five isolates at four concentrations ranging from 10⁵ to 10⁸ conidia ml^?1. The results showed that the slopes of regression lines for B. bassiana ERS4.16 (slope = 0.386) and TAM6.2 (slope = 0.41) were the most important and had the lowest LC₅₀ values (2.85 × 10³ and 3.16 × 10³ conidia ml^?1 respectively). This investigation suggests that the soil of Argan forests contains pathogenic B. bassiana isolates and highlights for the first time their potential as biological control toward C. capitata larval stage in Morocco.     

A rapid knockdown effect of Penicillium citrinum for control of the mosquito Culex quinquefasciatus in Thailand

Twenty local isolates of entomopathogenic fungi were determined for control of the larvae and adults of Culex quinquefasciatus. In a laboratory experiment, a Penicillium sp. CM-010 caused 100 % mortality of third-instar larvae within 2 h using a conidial suspension of 1 × 10⁶ conidia ml^?1. Its LC₅₀ was 3 × 10⁵ conidia ml^?1, and the lethal time (LT₅₀) was 1.06 h. Cloning and sequencing of its internal transcribed spacer region indicated that this Penicillium species is Penicillium citrinum (100 % identity in 434 bp). Mortality of the adult was highest with Aspergillus flavus CM-011 followed with Metarhizium anisopliae CKM-048 from 1 × 10⁹ conidia ml^?1. P. citrinum CM-010 at 1 × 10⁶ conidia ml^?1 killed 100 % larvae within 2 h while Bacillus thuringiensis var. israelensis at 5 ITU ml^?1 required 24 h. This P. citrinum CM-010 also greatly reduced survival of C. quinquefasciatus larvae in an unreplicated field test. Light and transmission electron micrographs showed that the fungal conidia were ingested by the larvae and deposited in the gut. The metabolite patulin was produced by P. citrinum CM-010 instead of citrinin.     

Control of green mould of citrus by using Trichoderma harzianum,humic acid or garlic

Penicillium digitatum, an aggressive fungus causes post-harvest decay of mandarin sweet orange and Washington navel. In vitro Trichoderma harzianum or humic acid (HA) or powdered cloves of garlic caused inhibition of fungal growth of isolates P₁ and P₂. Under storage conditions, the fruit citrus is protected by using T. harzianum with standard volume 2.0?ml (9.6?×?10⁶?conidia/ml) and application 24?h before inoculation reduces disease incidence and disease severity after seven?days from inoculation with P. digitatum spore suspension (1.0?×?10⁶?spores/ml) compared to control. Spraying the fruit citrus by standard volume of 2.0?ml of either HA or powder cloves of garlic 1% on each fruit 24?h before inoculation reduces disease incidence and disease severity after seven?days from inoculation with P. digitatum (1.0?×?10⁶ spores/ml) compared to control. The lowest percentage of disease incidence and disease severity were associated with powder of cloves garlic and followed by HA and T. harzianum during two growing seasons compared with the untreated and control.     

Pine processionary moth (Thaumetopoea pityocampa,Lepidoptera: Thaumetopoeidae) larvae are highly susceptible to the entomopathogenic fungi Metarhizium brunneum and Beauveria bassiana

The larvae of the pine processionary moth (PPM), Thaumetopoea pityocampa, feed on the needles of pine and cedar. The urticating hairs of older instars pose a threat to human and animal health. Strains of the entomopathogenic fungi, Metarhizium brunneum (V275, ARSEF 4556) and Beauveria bassiana (KTU-24), were assayed against first to fourth instar T. pityocampa using doses ranging from 1?×?10⁵ to 1?×?10⁸ conidia mL^?1. The three strains differed slightly in their virulence but caused 100% mortality of all instars at the highest dose. The newly emerged or first instar larvae were extremely susceptible with 100% mortality being achieved 2–4 days post inoculation with V275 at all but the lowest dose. The fourth instar larvae appeared to be less susceptible than earlier instars. There was good horizontal transmission of conidia from treated to un-inoculated larvae. However, mortality was higher in third and fourth instars and where the ratio of inoculated versus untreated larvae was high. This we presume is due to spores being more readily trapped by the urticating hairs found on third and older instar larvae. Injection of the nests offers a simple and environmentally friendly way of controlling the pest with reduced risk to operators.     

Effect of dual inoculation with nematodes and fungal pathogens on the survival of Phyllophaga polyphylla larvae (Coleoptera: Scarabaeidae)

Phyllophaga polyphylla (Coleoptera: Scarabaeidae) is an important pest of maize and other crops in Mexico. Previous reports showed that this pest was highly resistant to fungal and nematode infection when each pathogen was inoculated separately; in this study, we evaluated whether dual inoculation of fungi and nematodes, in all possible pair-wise combinations and orders of inoculation and including an evaluation of a time separation of 73 hours between each pathogen's inoculation, would increase mortality in P. polyphylla larvae. The pathogens were two isolates of Heterorhabditis bacteriophora applied at a concentration of 50 infective juveniles (IJ) mL^–1, an isolate of Beauveria pseudobassiana and of Metarhizium pingshaense both applied at a concentration of 1 × 10⁸ conidia mL^–1. In the first experiment, the combined mortality when pathogens were dual-inoculated (13%), although significantly higher than single-inoculated treatments (8%), demonstrated that antagonistic interactions were ongoing between the pathogens, as confirmed by the χ²-test. In a separate experiment, using only the B. pseudobassiana isolate (1 × 10⁸ conidia mL^–1) and one isolate of H. bacteriophora (100 IJ mL^–1), we studied the effect of different order of inoculations but included a two-week separation between inoculation of each pathogen. Mortalities obtained were similar to the previous experiment; all interactions resulted in antagonistic effects, except when the fungal pathogen was inoculated first, which resulted in an additive interaction. Understanding the mechanisms for the interaction requires further study but, for practical biological control, we suggest that more virulent fungal and nematode isolates are necessary to achieve control of P. polyphylla.     

Isolation and virulence of entomopathogenic fungi against the great spruce bark beetle,Dendroctonus micans (Kugelann) (Coleoptera: Scolytidae)

Twelve fungal strains including Lecanicillium muscarium (Petch.) Zare and Gams, Isaria farinosa (Holmsk.) Fr., Fusarium sp., Beauveria bassiana Sensu Lato and Beauveria sp. were isolated from larvae and adults of D. micans. In addition, virulence of these isolates against this pest was determined. Conidia suspensions of 1×10⁶ conidia mL^–1 were applied to larvae and adults. The highest mortality and mycosis for larvae were obtained from isolate ARSEF 9271 (Beauveria bassiana) with 90% mortality and mycosis within 10 days. ARSEF 9271 also produced 93% mortality and mycosis in adults. On the other hand, the highest mortality and mycosis for adults were obtained with isolate ARSEF 9272 (Beauveria sp.), with 100% mortality and 80% mycosis within 10 days. These results indicate that isolates ARSEF 9271 and ARSEF 9272 seem to be the most promising potential fungal biocontrol agents against D. micans.     

Biology and performance of two indigenous larval parasitoids on Tuta absoluta (Lepidoptera: Gelechiidae) in Sudan

Tuta absoluta is an alien invasive pest in Sudan. Since it was detected in the country, the pest continues to cause major tomato losses. Alarmed by its devastating nature and the speed of spread, tomato growers resorted to the indiscriminate use of broad-spectrum insecticides. Promising indigenous parasitoid(s) may contribute to suppression of this pest. The biology and performance of the native parasitoids, Bracon nigricans Szépligeti (Braconidae) and Dolichogenidea appellator (Telenga) (Braconidae), was therefore evaluated against different immature stages of T. absoluta under laboratory conditions. A significantly higher number of fourth instar larvae was accepted for oviposition by B. nigricans. Similarly, fourth instar larvae yielded a considerably higher number of parasitoid offspring compared to third instar larvae. The male ratio of B. nigricans offspring which emerged from both larval instars was high, 0.96 and 0.66 for third and fourth instar larvae of T. absoluta, respectively. There was, however, no significant difference in the number of males and females that emerged from fourth instar larvae. Conversely, the number of hosts killed by stinging behaviour or host feeding was significantly higher on third instar larvae. B. nigricans potential fecundity varied with the age of the wasp with the lowest fecundity on the day of eclosion. The performance of D. appellator in terms of the total number of offspring produced and female progeny was similar for second and third instar larvae of T. absoluta. The preimaginal developmental time for both parasitoid species did not vary with either host stage or sex of the parasitoid. B. nigricans adult longevity was similar for both sexes, while that of D. appellator significantly varied with females living longer than males. Prospects for conservative biological control of T. absoluta are discussed in the light of the results of this study.     

Effectiveness of different release rates of Trichogramma cacoeciae (Hymenoptera: Trichogrammatidae) against Tuta absoluta (Lepidoptera: Gelechiidae) in protected and open field tomato crops in Tunisia

Tuta absoluta (Meyrick) is a key insect pest of tomato crop causing major economic losses worldwide. Biological control using Trichogramma parasitoids is considered a promising, eco-friendly management tactic of this pest. We performed inundative releases of Trichogramma cacoeciae (Marchal) (Hymenoptera: Trichogrammatidae) against T. absoluta in Takelsa (northeastern Tunisia). Three weekly releases of either (i) 10, 20 or 30 Trichogramma per plant in protected (greenhouse) tomato crops or (ii) 20 or 40 Trichogramma per plant in open field tomatoes were evaluated for their effectiveness. Results indicated that 20 Trichogramma per plant was the most effective release rate in significantly decreasing the pest’s life stage densities. Parasitism rates were significantly higher on apical tomato leaves (estimated at 58.6% for the rate 20 Trichogramma per plant in greenhouses and at about 56% for the rate 40 Trichogramma per plant in open field) compared to those observed on middle leaves (24.6% and 45.26% for rate 2 respectively under greenhouse and open field conditions). Furthermore, the overall number of T. absolua eggs and larvae was significantly reduced after releases of Trichogramma parasitoids. These results clearly indicate that T. cacoeciae could be a promising biocontrol agent of T. absoluta in the largest tomato-producing area in Tunisia.     

Evaluation of a granular formulation containing Metarhizium brunneum F52 (Hypocreales: Clavicipitaceae) microsclerotia in controlling eggs of Aedes aegypti (Diptera: Culicidae)

We evaluated the potential of a granular formulation of Metarhizium brunneum F52 containing microsclerotia (MbMSc granules) for control of Aedes aegypti by targeting eggs. MbMSc granules produced infective conidia within 14 days after application to 2.5?g moist potting soil, producing 5.9?×?10⁵, 2.08?×?10⁶ and 6.85?×?10⁶ conidia from 1, 5 and 25?mg MbMSc granules, respectively. Application of MbMSc triggered premature eclosion of eggs (EC_50?=?12?mg) with percentages as high as 31?±?2.9% and 67?±?4.3% of the eggs treated with 5 and 25?mg MbMSc granules, respectively, after 14 days on moist filter paper. Premature eclosion of eggs started at 3 days subsequent to MbMSc granule application and survival of larvae was significantly reduced for granule treated eggs (74?±?2.2%, 39?±?2.0% and 23?±?4.9% larvae survived for 1, 5 and 25?mg granule treatments, respectively, EC₅₀?=?4.9?mg). When MbMSc granules were applied in moist potting soil with mosquito eggs, rates of 1, 5 and 25?mg of MbMSc granules significantly reduced adult emergence with only 81?±?2.1%, 47?±?1.9%, and 34?±?2.1% emergence, respectively (EC₅₀?=?7?mg). Eggs treated with increasing concentrations of fungal conidia enhanced premature eclosion of eggs with an EC₅₀?=?1.6?×?10⁶ conidia/mL. Our results demonstrate that MbMSc granules are a promising candidate for control of A. aegypti and that fermentative production of Mb F52 microsclerotia as the active propagule has the potential for use for mosquito control.     

Investigation of native isolates of entomopathogenic fungi for the biological control of three citrus pests

The pathogenicity of 15 isolates of Beauveria bassiana (Balsamo) Vuillemin, five isolates of Metarhizium anisopliae (Metschnikoff) Sorokin and one isolate of M. flavoviride (Gams and Rozsypal) were tested under laboratory conditions against the subterranean life stages of the citrus pests, Ceratitis rosa Karsch, C. capitata Wiedemann (Diptera: Tephritidae) and Thaumatotibia leucotreta Meyrick. (Lepidoptera: Tortricidae). When these citrus pests were treated with a concentration of 1×10⁷ conidia mL^?1, fungal isolates had a significantly greater effect on the adults of C. rosa and C. capitata than they did on the puparia of these two fruit fly species. Further, C. rosa and C. capitata did not differ significantly in their response to entomopathogenic fungi when adult and pupal mycosis were considered. Depending on fungal isolate, the percentage of T. leucotreta adults which emerged from fungal treated sand ranged from 5 to 60% and the percentage of pupae with visible signs of mycosis ranged from 21 to 93%. The relative virulence of the four most promising fungal isolates, as well as the commercially available B. bassiana product, BroadBand® (Biological Control Products, South Africa), were compared against one another as log-probit regressions of mortality against T. leucotreta which exhibited a dose-dependent response. The estimated LC₅₀ values of the three most virulent B. bassiana isolates ranged from 6.8×10⁵ to 2.1×10⁶ conidia mL^?1, while those of the least pathogenic ranged from 1.6×10⁷ to 3.7×10⁷ conidia mL^?1.     

Isolation and pathogenicity of some South African entomopathogenic fungi (Ascomycota) against eggs and larvae of Cydia pomonella (Lepidoptera: Tortricidae)

Virulent entomopathogenic fungal strains against Cydia pomonella (Lepidoptera: Tortricidae) were isolated and identified in the Western Cape Province of South Africa. Thirty-nine isolates belonging to six species were obtained using the insect bait method. Generally, Metarhizium robertsii (Ascomycota: Hypocreales) was the most frequently encountered species representing 51% of the total number of isolates collected from the soil samples. This is the first report of M. robertsii from southern Africa. Mortality data from an immersion bioassay indicated that the 39 fungal isolates were pathogenic against fifth instar larvae of C. pomonella inducing 47–85% insect mortalities. Two M. robertsii strains, MTL151 and GW461, induced 85% larval mortality and were selected for further evaluations. The exposure of freshly laid eggs to wax papers that were pre-treated with fungal spores ranging from 10³ to 10⁸ spores ml^?1 of MTL151 and GW461 resulted in a significant reduction of egg hatchabilities, from 95 to 66% and 93 to 71%, respectively as spore concentration increased. First instar larval neonates were exposed to apples that were pre-sprayed topically with varied conidia suspensions (10³?10⁸ spores ml^?1). The mean percentage of participating apples with larvae in their cores/flesh significantly reduced from 53 to 10% (MTL151) and 76 to 10% (GW461) of 10 apples, and a concurrent decrease in the incidence of apple fruit rot was observed as conidia concentration increased. Up to 90% of apples treated with 1 × 10⁸ spores ml^?1 had no larvae present in their cores and this result compared favourably with the commercial pesticide Fruitfly®, containing the active ingredient cypermethrin (20 g/l) used at a recommended dose of 0.25 ml/250 ml of water.     

Susceptibility of Dendroctonus simplex to Hypocreales fungi: towards the development of a biological control strategy

The use of Hypocreales fungi against bark beetles is an attractive alternative strategy for biological control in forestry. However, fungal inoculation methods remain a major challenge due to the cryptic behaviour of these beetles. Experiments were conducted to evaluate the susceptibility of adult eastern larch beetles, Dendroctonus simplex, to different isolates of Hypocreales fungi and to assess the efficiency of direct inoculation of fungal conidia using an assisted autodissemination device. Among the nine fungal isolates tested, Beauveria bassiana INRS‐242 was selected as the utmost isolate causing a mortality rate of 64.4% of D. simplex adults within 10 days and having a high conidial viability of 81.7%. Using an assisted autodissemination device, a fungal‐coated pouch was a promising contamination chamber. Once it had fallen onto the fungal‐coated pouch, each D. simplex adult loaded an average lethal dose of 2.1 × 10⁹ conidia, which induced a mortality rate of 98.0%. Our results demonstrated the fungal pathogenicity of B. bassiana INRS‐242 against the eastern larch beetles and pointed out the potential of the assisted autodissemination device as an effective method of conidia inoculation. These findings represent the first step to develop a biological control strategy against D. simplex populations.     

Effects of adjuvant and conidial concentration on the efficacy of <Emphasis Type="Italic">Beauveria bassiana</Emphasis> for the control of the two spotted spider mite, <Emphasis Type="Italic">Tetranychus urticae</Emphasis>

Greenhouse experiments were conducted on various crops (cucumber, tomato, eggplant, green bean) to ascertain the effects of Break-thru^® (polyether-polymethylsiloxane-copolymer, a silicone surfactant) and an oil emulsion, on Beauveria bassiana (Balsamo) Vuillemin (Bb) applications for the control of the two spotted spider mite, Tetranychus urticae Koch. The objectives were to compare a) the efficacy of Bb control when applied in aqueous Break-thru^® or an oil emulsion; b) the effects of various concentrations of Bb conidia, as affected by each surfactant; and c) the effects of Break-thru^® on the activity of the fungus. Conidia were suspended either in an aqueous Break-thru^® or an emulsifiable formulation at different conidial concentrations (1.05 × 10⁶, 2.1 × 10⁶ and 4.2 × 10⁶ conidia ml^?1) and sprayed onto leaves 2 weeks after artificial pest inoculation. Two sprays were performed, with an interval of one week from one spray to another, and T. urticae population counts (both motile and egg stages) were made on plant leaves 7 days after each spray. Bb conidia in Break-thru^® were more efficacious than conidia in emulsifiable formulation. With the highest rate of conidia (4.2 × 10⁶ conidia ml^?1), mortality of adult mites ranged from 60 ± 4.2 (mean ± SE) to 85.7 ± 4.3% in the Break-thru^® suspension and 39.4 ± 7 to 61.3 ± 6% in the oil emulsion. Leaf damage index was also substantially reduced from 70% in the unsprayed control to 40% by the application of Bb conidia at the highest rate with Break-thru^®. Break-thru^® can be combined with Bb in the integrated management of T. urticae and Isolate R444 is a promising candidate for the control of the pest.     

Time-concentration-mortality response of potato tuber moth pupae to the biocontrol agent Cordyceps tenuipes

ABSTRACT

The potato tuber moth (PTM) Phthorimaea operculella is a critical potato pest. Larvae infest both foliage and tubers and mature larvae pupate in the soil or other safe places. Cordyceps tenuipes, an entomopathogenic fungus, infect lepidopteran pupae. To determine the effectiveness of this fungus as a biocontrol agent for PTM, we evaluated the time-concentration-mortality (TCM) response of PTM pupae to C. tenuipes using the following bioassays: (1) direct immersion in conidial suspensions, (2) incubation in sterilised or (3) unsterilised soilpremixed with conidia, and (4) incubation in unsterilised soil drenched with conidial suspensions to simulate field conditions. Fungal infection caused 100%, 83.3%, 73.3%, and 85.0% mortality of PTM pupae in assays 1–4, respectively. At 10⁸ conidia/mL or conidia/g concentration, assays 1 and 4 had short lethal times (LT₅₀) of 2.2 and 2.6 days compared with 3.7 and 4.8 days for assays 2 and 3, respectively. On day 7 after inoculation, assays 1 and 4 also had low lethal concentrations (LC₅₀) of 1.69 × 10³ conidia/mL and 1.10 × 10⁵ conidia/g compared with those of assays 2 and 3, which showed low virulence, with LC₅₀ of 3.50 × 10⁵ and 3.60 × 10⁶ conidia/mL, respectively. Our results demonstrate that C. tenuipes is a promising candidate for PTM biocontrol at the pupal stage. Drenching the soil surface with conidial suspensions may be the most effective method of field application.     

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.     

Susceptibility of Phyllophaga polyphylla and Anomala cincta larvae to Beauveria bassiana and Metarhizium anisopliae isolates, and the interaction with soil properties

The white grub species Phyllophaga polyphylla and Anomala cincta (Coleoptera: Melolonthidae) are economically important species that affect many crops in Mexico. A series of experiments to study the pathogenic interaction between isolates of Beauveria bassiana and Metarhizium anisopliae and these two insect species were undertaken. First, the susceptibility of third instar P. polyphylla larvae to each of seven isolates representing both species of fungus was evaluated by dipping the insects in 1?×?10⁸ conidia?ml^?1 suspensions. A second study examined the differences in the susceptibility of P. polyphylla and A. cincta larvae to two selected isolates for each of the fungal species. Finally, the susceptibility of A. cincta larvae to one M. anisopliae isolate when incubated in soil collected from four different sites was assessed. No significant differences in proportion of infection of P. polyphylla larvae were observed amongst the fungal isolates tested and mortality due to fungal infection was never greater than 20% after 36?days incubation. Anomala cincta larvae were more susceptible than P. polyphylla larvae, with greater than 90% infection when inoculated with isolates of M. anisopliae whereas mortalities of only 20% where achieved against P. polyphylla larvae. The soil type in which A. cincta were incubated following inoculation with M. anisopliae affected their susceptibility to infection. The results demonstrated that there is a complex interaction amongst entomopathogenic fungi, white grub larvae and soil properties, and points to the need of further investigation of this system in order to optimize the efficacy of entomopathogenic fungi against these insect species.     

Nanosilica and jasmonic acid as alternative methods for control Tuta absoluta (Meyrick) in tomato crop under field conditions

Recently, Tuta absoluta became one of the major pests that attack commercial tomato globally. Field test was done to evaluate the effect of different concentrations of nanosilica (NS) and jasmonic acid (JA), and compared them with indoxacrb (recommended insecticide) on reduction of damage rate caused by T. absoluta larvae under field conditions. Nanosilica (600 ppm) and indoxacrb (0.25 cm³/L) had the highest efficiency to reduce the rate of mines in the leaves. Jasmonic acid at rate 1.141 μM/plant showed a good reduction of number of fruits damaged. Nanosilica with 600 ppm concentration and Jasmonic acid at rate 1.141 μM/plant is used to control T. absoluta in the field.     

Responses of diamondback moth to diverse entomopathogenic fungi collected from non-agricultural habitats – Effects of dose,temperature and starvation

We evaluated the virulence of Beauveria bassiana and Metarhizium isolates from soil collected across different vegetation types in Queensland, against chlorantraniliprole-resistant and insecticide-susceptible diamondback moth (DBM) larvae. Host insecticide resistance status had no effect on susceptibility to the pathogens when conidia were topically applied to larvae in the laboratory, and one B. bassiana isolate was significantly more virulent to larvae than the others (seven days after inoculation). The influence of temperature (15, 20, 25 or 30 °C): (i) at the point of host inoculation with conidia and (ii) when the pathogens had already initiated infection and were proliferating in the host haemocoel, was determined experimentally for its influence on virulence, disease progression, and sporulation. Temperature at inoculation had a greater effect on host insect mortality than it did when the fungus was already proliferating in the host haemocoel. The rearing temperature of hosts prior to inoculation had a greater effect on host susceptibility to disease than starvation of the larvae at the time of inoculation. Our results also show that each fungal isolate has its own temperature relations and that these can vary considerably across isolates, and at different points in the pathogen life cycle (germination and cuticular penetration versus growth in the host haemocoel). Temperature also had an idiosyncratic effect, across isolates and across the variables typically used to assess the potential of fungal entomopathogens as biological control agents (time to death, mortality and sporulation rates). This study demonstrates that in addition to pathogenicity and virulence, the temperature relationships of each fungal isolate when infecting insects needs to be taken into account if we are to understand their ecology and use them effectively in pest management.