Group Movement in Entomopathogenic Nematodes: Aggregation Levels Vary Based on Context.

Maintenance of an aggregated population structure implies within-species communication. In mixed-species environments, species-specific aggregations may reduce interspecific competition and promote coexistence. We studied whether movement and aggregation behavior of three entomopathogenic nematode species changed when isolated, as compared to mixed-species arenas. Movement and aggregation of Steinernema carpocapsae, S. feltiae and S. glaseri were assessed in sand. Each species demonstrated significant aggregation when alone. Mixed-species trials involved adding two species of nematodes, either combined in the center of the arena or at separate corners. While individual species became less aggregated than in single-species conditions when co-applied in the same location, they became more aggregated when applied in separate corners. This increased aggregation in separate-corner trials occurred even though the nematodes moved just as far when mixed together as they did when alone. These findings suggest that maintenance of multiple species within the same habitat is driven, at least in part, by species-specific signals that promote conspecific aggregation, and when the species are mixed (as occurs in some commercial formulations involving multiple EPN species), these signaling mechanisms are muddled.

A winning formula: sustainable control of three stored-product insects through paired combinations of entomopathogenic fungus, diatomaceous earth, and lambda-cyhalothrin.

This research aimed to assess the effectiveness of Metarhizium robertsii, diatomaceous earth (Protect-It), and lambda-cyhalothrin, for the long-term protection of stored wheat against three destructive grain insect pests, Rhyzopertha dominica, Tribolium castaneum, and Trogoderma granarium. Different treatments were applied, both alone and in paired combinations in laboratory and persistence trials. Single treatments exhibited significantly lower mortality rates in comparison to the paired treatments for all tested insect species. Among the single treatments, lambda-cyhalothrin (Lamb) resulted in significantly higher mortality rates in laboratory trials, followed by diatomaceous earth (DE) and M. robertsii (Mr), with insignificant differences between Mr and DE. Evidently, DE exhibited the highest persistence after 120 days of storage for all insect species and initial exposures, although variations in mortality rates among treatments were mostly insignificant. Overall, the most effective treatment in terms of mortality in laboratory, and persistence trials, and progeny production was DE + Lamb, followed by Mr + Lamb, and Mr + DE for all tested insect species. In general, the most susceptible insect species was R. dominica, followed by T. castaneum and T. granarium. This research highlights the effectiveness of M. robertsii, DE, and lambda-cyhalothrin in providing prolonged protection of stored wheat against all the examined grain insect species.

Evaluating the Compatibility of Spinosad and Alpha-Cypermethrin for Controlling Six Insect Pests Infesting Stored Wheat.

The deterioration of stored wheat due to pest infestations is a significant concern, with pests like Rhyzopertha dominica, Tribolium castaneum, Cryptolestes ferrugineus, Sitophilus oryzae, Oryzaephilus surinamensis, and Trogoderma granarium being major contributors. This study examined the efficacy of spinosad and alpha-cypermethrin, individually and in combination, against these pests under laboratory conditions. Spinosad was tested at two concentrations (0.05 and 0.1 mg/kg), while alpha-cypermethrin was applied at 0.05 mg/kg. The combined application of both insecticides led to significantly higher pest mortality compared to single treatments. Importantly, all treatments caused substantial pest mortality and exhibited the ability to suppress pest progeny production over time, as observed in both laboratory and persistence trials. Among the various treatment combinations, the joint application of 0.1 mg/kg spinosad and 0.05 mg/kg alpha-cypermethrin emerged as the most effective, resulting in elevated mortality and a marked reduction in pest progeny. Rhyzopertha dominica exhibited the highest susceptibility among the pests, followed by S. oryzae, T. castaneum, C. ferrugineus, O. surinamensis, and T. granarium. The remarkable performance of the joint action of alpha-cypermethrin and spinosad at low doses highlights this combination as an efficacious approach for safeguarding stored grain against these destructive insect pests, warranting further exploration.

Natural Warriors against Stored-Grain Pests: The Joint Action of Beauveria bassiana and Steinernema carpocapsae.

Tribolium castaneum, Trogoderma granarium, Oryzaephilus surinamensis, Sitophilus oryzae, Rhyzopertha dominica, and Cryptolestes ferrugineus are all major pests of stored grains. In this study, the efficiency of single and joint applications of the entomopathogenic nematode (EPN) Steinernema carpocapsae at two different doses (50 and 100 IJs cm-2) and the entomopathogenic fungus (EPF) Beauveria bassiana for the management of the aforementioned pests was estimated. At single treatments, both doses of S. carpocapsae caused higher mortality rates to all six pest species compared to B. bassiana. The combined treatment of EPF and EPN resulted in higher mortality compared to single treatments. Mortality was strongly influenced by the exposure interval and the application dose of the EPN at both single and combined treatments. Maximum mortality was observed for the application of the combined treatment at the high dose of S. carpocapsae and B. bassiana. Among the different insect species tested, the maximum mortality rate was observed for R. dominica (96.62%), followed by S. oryzae (90.48%), T. castaneum (87.23%), C. ferrugineus (76.05%), O. surinamensis (70.74%), and T. granarium (57.71%). The outcomes of this study demonstrate the potential of utilizing specific combinations of EPF and EPN as effective natural enemies against stored-grain pests.

Treatment of Four Stored-Grain Pests with Thiamethoxam plus Chlorantraniliprole: Enhanced Impact on Different Types of Grain Commodities and Surfaces.

An insecticide containing the neonicotinoid thiamethoxam + the diamide chlorantraniliprole was evaluated against adults of Rhyzopertha dominica, Tribolium castaneum, Trogoderma granarium, and Sitophilus oryzae under laboratory bioassays both on freshly treated grain as well as on treated grain stored over 90 days for its persistence in efficacy. In laboratory bioassays, the insecticide was applied on wheat, maize, or rice at four doses, while in persistence bioassays on wheat at the same doses. Mortality and progeny were assessed in both laboratory and persistence bioassays. After 14 days of exposure, S. oryzae exhibited 100% mortality on all three commodities at the highest dose, while R. dominica showed complete mortality on wheat or rice and T. castaneum on wheat. For a period of 90 days, S. oryzae exhibited 42.69% mortality, followed by R. dominica (35.26%), T. castaneum (27.08%), and T. granarium (18.63%) at the highest dose. Progeny was successfully suppressed in all cases of complete mortality in laboratory bioassays and for S. oryzae for 90 days in persistence bioassays. Laboratory trials were also performed on plywood, concrete, ceramic tile, and steel at one dose. The highest mortality was observed on steel, followed by concrete, ceramic tile, and plywood for all insect species tested. This study demonstrates that thiamethoxam + chlorantraniliprole is effective against the tested species depending on exposure, storage period, surface, commodity, and dose.

The Potential of Two Entomopathogenic Fungi and Enhanced Diatomaceous Earth Mixed with Abamectin: A Comprehensive Study on Mortality, Progeny Production, Application Method, and Surface Application against Tribolium castaneum.

This study determined the efficacy of Beauveria bassiana (Bals. -Criv.) Vuill., Metarhizium anisopliae (Metchnikoff) Sorokin, and diatomaceous earth mixed with abamectin (DEA) alone and in their combinations for the integrated management of larvae and adults of Tribolium castaneum (Herbst) from three field populations of Pakistan (Multan, Rawalpindi, and Rahim Yar Khan) and one laboratory population (Faisalabad). Treatments were applied on three surfaces, namely, viz. steel, concrete, and jute bags, implementing two application methods, dusting and spraying. The combined treatments were more effective in comparison with single treatments for both larvae and adults. Overall, the highest mortality rates were recorded in the Faisalabad population, followed by the Rehaim Yar Khan, Rawalpindi, and Multan populations. Progeny production was suspended 21 days after exposure to the combined treatment of DEA and both fungi in all populations except Rawalpindi. Larvae were found to be more susceptible than adults in all treatments and intervals. Dusting was more efficient than spraying for both larvae and adults and for all the populations studied. The present study provides a wholistic understanding of the impact of different factors on the success of the combined treatments using DEA and entomopathogenic fungi, supporting their use as surface treatments.

Efficacy of entomopathogenic fungi, nematodes and spinetoram combinations for integrated management of Thrips tabaci.

BACKGROUND: Two consecutive field trials using a blend of entomopathogens in combination with a new chemistry insecticide were conducted to determine treatment effects on onion thrips (Thrips tabaci Lindeman) populations, crop damage, plant development, crop yield and impact on natural enemies. Products were tested in an onion cropping system and included the insect pathogenic fungus Beauveria bassiana (isolate WG-11), an entomopathogenic nematode Heterorhabditis bacteriophora (strain VS) and the new-chemistry chemical insecticide spinetoram. RESULTS: In all treatments, a significant decrease in thrips per plant population was detected in both trials. Overall, dual application of entomopathogens and insecticide was more effective than singly applied treatments. The lowest number of thrips larvae (1.96 and 3.85) and adults (0.00 and 0.00) were recorded when treated with dual application of B. bassiana and spinetoram at 7 days post application (DPA) after the second spray application in 2017and 2018, respectively. Damage on onion plants was considerably decreased in all treatments relative to the control. The lowest damage was observed on onion plants treated with B. bassiana + spinetoram at 7 DPA after the second spray application during both years. A significant decrease in the number of natural enemies (beetles, spiders, mites, lacewings, ants and bugs) on onion plants was recorded during both years. Insect pathogens when applied alone and in combination with each other considerably protected arthropod natural enemies compared to insecticide application applied alone. Significant increase in plant agronomic traits was observed compared to the control. Among all the treatments, B. bassiana + spinetoram produced maximum leaf length, leaf weight, total leaves, neck diameter, bulb diameter, number of rings per bulb, bulb weight, dry matter and plant yield following the 2017and 2018 applications, respectively. CONCLUSION: The findings of the study reveal the potential of using insect pathogens and insecticide for control of T. tabaci. However, combinations containing spinetoram are harmful to nontarget organisms, whereas biological control agents help in protecting biodiversity in onion agroecosystems. © 2023 Society of Chemical Industry.

Development of Insecticide Resistance in Field Populations of Onion Thrips, Thrips tabaci (Thysanoptera: Thripidae).

The present study evaluated insecticide resistance in field populations of onion thrips, Thrips tabaci Lindeman (Thysanoptera: Thripidae), collected from eight different onion-growing regions of Punjab, Pakistan. These field-collected populations were assessed for resistance development against eight commonly used active ingredients including deltamethrin, lambda-cyhalothrin, imidacloprid, acetamiprid, spinosad, spinetoram, cypermethrin, and abamectin. In leaf dip bioassays, T. tabaci adults showed varied levels of resistance towards different insecticides. Moderate or high levels of resistance to deltamethrin (58-86 fold), lambda-cyhalothrin (20-63 fold), and cypermethrin (22-54 fold) were observed in T. tabaci field populations. There were very low to moderate resistance levels to imidacloprid (10-38 fold), acetamiprid (5-29 fold), and abamectin (10-30 fold). The lowest levels of resistance were detected in thrips exposed to spinosad (3-13 fold) and spinetoram (3-8 fold). Insecticide resistance levels varied among populations collected from various geographic locations, but all populations exhibited elevated levels of resistance to deltamethrin. Thrips tabaci populations with higher resistance levels were most commonly found from the southern part of Punjab, Pakistan. Our findings revealed that spinosyns could be used as alternatives to conventional insecticides for the successful management of T. tabaci in onion fields.

Two are better than one: the combinations of Beauveria bassiana, diatomaceous earth, and indoxacarb as effective wheat protectants.

The current study evaluates the efficacy of the entomopathogenic fungus Beauveria bassiana (Balsamo-Crivelli) Vuillemin (Hypocreales: Cordycipitaceae), diatomaceous earth (DE) (Protect-It), and the oxadiazine indoxacarb, at single or combined applications on wheat kernels, for the management of the rusty grain beetle, Cryptolestes ferrugineus (Stephens) (Coleoptera: Laemophloeidae), the red flour beetle, Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae), the khapra beetle, Trogoderma granarium Everts (Coleoptera: Dermestidae), and the lesser grain borer, Rhyzopertha dominica (F.) (Coleoptera: Bostrychidae). The study was conducted between November 2020 and August 2021 in Faisalabad under a complete randomized block design. The combination of DE + indoxacarb was the most efficient as it caused higher overall mortalities ranging between 59.34 and 100%, and lower overall progeny production ranging between 8.35 and 33.70 individuals per vial, than all other treatments. Beauveria bassiana alone exhibited the lowest mortality rates ranging between 22.33 and 47.76%, and the highest offspring emergence, ranging between 51.33 and 78.55 individuals per vial. Similar pattern was observed when persistence bioassays were conducted. For a period of 120 days, the DE + indoxacarb was the most powerful combination against all tested species, providing overall mortality rates between 17.06 and 63.80%. The overall progeny production was lower for the insect individuals exposed on wheat treated with the DE + indoxacarb combination, ranging between 13.66 and 52.23 individuals per vial, and higher for those exposed to B. bassiana alone, ranging between 44.03 and 107.67 individuals per vial, for the entire duration of storage. However, the efficacy of all treatments decreased gradually during the course of storage. The findings of the current study indicate that the combinations of entomopathogenic fungi, DE, and indoxacarb can be used for the prolonged protection of stored wheat from the tested noxious insect species of stored products. Further research, which will include other inert dusts in combination with entomopathogenic fungi and indoxacarb, may provide additional knowledge towards an effective management of noxious species occurring in storages.

Combinations of Beauveria bassiana and spinetoram for the management of four important stored-product pests: laboratory and field trials.

The current study examines the efficacy of the semi-synthetic insecticide spinetoram and entomopathogenic fungi Beauveria bassiana (Balsamo-Crivelli) Vuillemin (Hypocreales: Cordycipitaceae) as wheat protectants against the lesser grain borer, Rhyzopertha dominica (F.) (Coleoptera: Bostrychidae), the red flour beetle, Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae), the granary weevil, Sitophilus granarius (L.) (Coleoptera: Curculionidae), and the khapra beetle, Trogoderma granarium Everts (Coleoptera: Dermestidae), under laboratory and field trials. One dose of B. bassiana, i.e., 1 × 107 conidia/kg wheat, two doses of spinetoram, i.e., spine1: 0.05 ppm (mg/kg wheat), spine2: 0.1 ppm, and their combinations (Bb + spine1, Bb + spine2) were evaluated at 20, 25, and 30 °C. All treatments provided significantly higher mortality at 30 °C compared with the other two temperatures. Maximum mortality levels were observed in the treatments where B. bassiana was combined with the higher dose of spinetoram (0.1 ppm). All treatments reduced progeny production in comparison with the control groups. Maximum progeny reduction was observed at 30 °C, on wheat treated with the Bb + spine2 combination. The combination Bb + spine2 also provided elevated mortality rates in both laboratory and field persistence trials, but at 180 days caused moderate mortality to all tested insect species. Concerning progeny, at laboratory persistence trials, the combination Bb + spine2 exhibited the lowest offspring emergence to all tested species compared to the other treatments and control. Overall, our study showed that R. dominica was the most susceptible species followed by S. granarius, T. castaneum, and T. granarium. Our findings revealed that the combination of B. bassiana and spinetoram may be a useful tool for efficient and advanced integrated pest management strategies for long storage periods under multiple temperatures.

Combined application of entomopathogenic nematodes and fungi against fruit flies, Bactrocera zonata and B. dorsalis (Diptera: Tephritidae): laboratory cups to field study.

BACKGROUND: The peach fruit fly, Bactrocera zonata, and the Oriental fruit fly, B. dorsalis (Diptera: Tephritidae), are economically important fruit fly species in various regions of the world. We evaluated the effects of separate and combined applications of the entomopathogenic fungi (EPF) Beauveria bassiana (WG-18) and Metarhizium anisopliae (WG-02), and the entomopathogenic nematodes (EPNs) Heterorhabditis bacteriophora (VS strain) and Steinernema carpocapsae (ALL strain) against larvae, pupae and pharate adults, of B. zonata and B. dorsalis under laboratory, glasshouse and field cage conditions. RESULTS: Combined applications of EPF and EPNs produced greater mortality than individual treatments under all conditions. Against both species, the combination of B. bassiana and H. bacteriophora consistently exerted strong effects that were similar to the combined application of B. bassiana and S. carpocapsae whereas M. anisopliae applied with S. carpocapsae was least effective in all combinations. In a laboratory bioassay, synergistic interactions were detected between B. bassiana and H. bacteriophora applied against larvae and pharate adults of both fly species, between B. bassiana and S. carpocapsae against larvae of both species and pharate adults of B. zonata, and between M. anisopliae and H. bacteriophora against B. zonata larvae. Other combined treatments resulted in additive effects, especially against fly pupae. In a potted soil bioassay, there were only additive interactions in all combinations against different stages of both flies. The 3rd instar of both flies was more susceptible than pharate adult and pupal stages. Additive interactions between EPNs and EPF were detected in the glasshouse against 3rd instars and pupae, and under field conditions against 3rd instars of both fly species. CONCLUSION: These results indicate how particular combinations of entomopathogenic fungi and nematodes could be deployed in integrated pest management of tephritid fruit flies in orchard agro-ecosystems. © 2022 Society of Chemical Industry.

Laboratory and field studies on the combined application of Beauveria bassiana and fipronil against four major stored-product coleopteran insect pests.

In the current study we have tested the application of Beauveria bassiana (Hypocreales: Cordycipitaceae) alone and in combination with fipronil at two doses against Tribolium castaneum (Coleoptera: Tenebrionidae), Rhyzopertha dominica (Coleoptera: Bostrychidae), Sitophilus granarius (Coleoptera: Curculionidae), and Trogoderma granarium (Coleoptera: Dermestidae) under laboratory and field conditions. At laboratory conditions, the combination of B. bassiana with the highest dose of fipronil produced the highest mortality. At different temperatures, mortality was increased with the increase in temperature. Maximum mortality was observed at 30 °C, followed by 25 °C and 20 °C for all tested species. Different treatments significantly reduced the progeny number in comparison to control groups for all tested species at all temperatures. In the persistence trial, all treatments that included the combinations of B. bassiana with fipronil produced significantly higher mortalities than the single treatments for all tested species over a period of 6 months. Furthermore, all treatments significantly reduced the number of progenies of all insect species in comparison with the control groups over the same storage period. In field trials, mortalities of all tested insect species were significantly higher on wheat treated with B. bassiana, fipronil, or their combinations than on controls for an entire storage period of 180 days. Overall, R. dominica was found the most susceptible species followed by S. granarius, T. castaneum, and T. granarium. The findings of the current study suggest that the use of B. bassiana and fipronil as grain protectants may provide elevated control against major stored-grain insect species during a prolonged period of storage.

Evaluation of Locally Isolated Entomopathogenic Fungi against Multiple Life Stages of Bactrocera zonata and Bactrocera dorsalis (Diptera: Tephritidae): Laboratory and Field Study.

Fruit flies including Bactrocera zonata and B. dorsalis (Diptera: Tephritidae) are considered major pests of orchard systems in Pakistan. This study evaluated the laboratory virulence, sub-lethal effects, horizontal transmission, greenhouse, and field-cage efficacy of locally isolated entomopathogenic fungi (EPF) against B. zonata and B. dorsalis. In virulence assays against third instars and adults, all 21 EPF isolates (Beauveria bassiana and Metarhizium anisopliae) tested were pathogenic and caused varying levels of mortality to the fruit flies. Based on the initial screening, four isolates (B. bassiana WG-21 and WG-18 and M. anisopliae WG-07 and WG-02) were selected for further study. The isolate WG-18 was the most virulent against larvae and adults of B. zonata and B. dorsalis followed by WG-21, WG-02, and WG-07. In both species, adults were more susceptible than larvae to all isolates, and pupae were the least susceptible. Isolates WG-18 and WG-21 strongly decreased female fecundity and fertility, the highest adult and larval mortality, and longest developmental time of larvae and pupae. Fungal conidia were disseminated passively from infected to healthy adults and induced significant mortality, particularly from infected males to non-infected females. In greenhouse and field-cage experiments, WG-18 and WG-21 were the most effective isolates in reducing adult emergence when applied to larvae and pupae of both fruit fly species. Our results indicate that B. bassiana isolates WG-18 and WG-21 were the most virulent against multiple life stages of B. zonata and B. dorsalis, and also exerted the strongest sub-lethal effects.

Virulence of Entomopathogenic Nematodes to Pupae of Frankliniella fusca (Thysanoptera: Thripidae).

Tobacco thrips, Frankliniella fusca (Hinds) is an economically significant pest. Entomopathogenic nematodes (EPNs) have shown promise as biocontrol agents against certain thrips species, but they have not been explored for suppression of F. fusca. We investigated the potential of EPNs to manage F. fusca by conducting three different bioassays: 1) a small cup dose-response bioassay (25, 50, and 100 IJs cm-2) with four EPN species, 2) a broad virulence bioassay with eight EPN species at 100 IJs cm-2, and 3) a potted soil bioassay testing with four EPN species (100 IJs cm-2). In the dose-response bioassay, all treatments showed relatively lower adult emergence when compared with the control group, but the minimum adult emergence (30%) was observed at 7 d post-treatment when Heterorhabditis bacteriophora (FL1-1) was applied at the highest rate (100 IJs cm-2). In the broad virulence study, all EPN treatments caused significant reductions in F. fusca adult emergence (18.3-75.0%) in comparison with the control. H. bacteriophora (Fl1-1) was more virulent than other nematode treatments but statistically not different from Steinernema feltiae and Steinernema riobrave, while Steinernema rarum was the least virulent. In the potted soil bioassay, the lowest emergence (10.6%) was observed in H. bacteriophora (Fl1-1) treatment, followed by S. feltiae (SN), S. riobrave (355), and Heterorhabditis indica (HOM1) treatments. These results indicate that EPNs have the ability to suppress the soil dwelling stage of F. fusca and should be explored further under greenhouse and field conditions for biocontrol potential within an integrated pest management (IPM) context.

Combined Effect of Entomopathogens against Thrips tabaci Lindeman (Thysanoptera: Thripidae): Laboratory, Greenhouse and Field Trials.

Onion thrips, Thrips tabaci Lindeman (Thysanoptera: Thripidae) is one of the most damaging insect pests of onions, Allium cepa L., which is an economically important agricultural crop cultivated worldwide. In this study, the combined application of entomopathogenic nematodes with entomopathogenic fungi against different soil dwelling stages of T. tabaci was evaluated. The nematodes included Heterorhabditis bacteriophora (VS strain) and Steinernema feltiae (SN strain), and fungi included Beauveria bassiana (WG-11) and Metarhizium anisopliae (WG-02); all four paired combinations (nematode + fungus) were included. In a small cup bioassay, only the combined application of H. bacteriophora and B. bassiana (WG-11) caused a synergistic interaction against pre-pupae, while all other combinations were compatible in an additive manner against pupae and late second instars. In a larger arena, a potted soil bioassay, again, combined applications of both pathogens produced greater mortality compared to single applications of each pathogen; all the combinations exhibited additive interactions, with the highest mortality observed in pre-pupae, followed by pupae and late second instar larvae using H. bacteriophora and B. bassiana (WG-11). Additionally, in the potted plant bioassay, lower adult emergence was observed from treated groups compared to control groups. Under field conditions, lower numbers of adults and larvae were found in treated groups relative to controls. Overall, the pre-pupal stage was more susceptible to the pathogen treatments, followed by pupae and late second instar larvae, and also combined applications of both pathogens suppressed the adult population. Combined application of entomopathogenic nematodes and fungi could be used for integrated pest management (IPM) of T. tabaci in onion production systems.

Detection of Phosphine Resistance in Field Populations of Four Key Stored-Grain Insect Pests in Pakistan.

In Pakistan, the control of stored-product insect pests mainly relies on the use of phosphine gas along with other control tactics. The aim of this study was to determine the level of phosphine resistance among ten differently located populations of the lesser grain borer, Rhyzopertha dominica (F.) (Coleoptera: Bostrychidae), the granary weevil, Sitophilus granarius (L.) (Coleoptera: Curculionidae), the red flour beetle, Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae) and the khapra beetle, Trogoderma granarium Everts (Coleoptera: Dermestidae). Laboratory-susceptible populations of all insect species were also considered in the experiments. Concentration-response bioassays were conducted for each species. All of the tested populations (10 out of 10) of each species were found to be resistant to phosphine, but varied in their level of resistance. Probit analysis estimated LC50 at 2.85, 1.90, 2.54 and 2.01 ppm for laboratory-susceptible populations of R. dominica, S. granarius, T. castaneum and T. granarium, respectively. Against R. dominica, the highest and lowest resistance levels were observed in the Rahim Yar Khan (LC50 at 360.90 ppm) and Rawalpindi (LC50 at 210.98 ppm) populations, respectively. These resistant populations were 126.67- and 74.02-fold more resistant than the laboratory population. The Multan and Lahore populations of S. granarius exhibited the maximum (LC50 at 122.81 ppm) and minimum (LC50 at 45.96 ppm) resistance levels, respectively, i.e., they were 64.63- and 24.18-fold more resistant than the laboratory population. The Layyah population of T. castaneum showed the maximum resistance level (LC50 at 305.89 ppm) while the lowest was observed in the Lahore population (LC50 at 186.52 ppm), corresponding to 120.42- and 73.43-fold more resistant than the laboratory population, respectively. Regarding T. granarium, the Layyah population showed the maximum resistance level (LC50 at 169.99 ppm) while the Lahore population showed the minimum resistance (LC50 at 74.50 ppm), i.e., they were 84.57- and 37.06-fold more resistant than the laboratory population, respectively. Overall, R. dominica presented the highest resistance level, followed by T. castaneum, T. granarium and S. granarius. The current study suggests that the application of phosphine may not be an adequate control strategy for the management of the above tested insect pests in Pakistan.

Persistence and efficacy of enhanced diatomaceous earth, imidacloprid, and Beauveria bassiana against three coleopteran and one psocid stored-grain insects.

The residual efficacy of the enhanced diatomaceous earth (DE) formulation DEBBM alone and in combination with Beauveria bassiana (Hypocreales: Cordycipitaceae) or with the neonicotinoid insecticide imidacloprid against Tribolium castaneum (Coleoptera: Tenebrionidae), Rhyzopertha dominica (Coleoptera: Bostrychidae), Cryptolestes ferrugineus (Coleoptera: Laemophloeidae), and Liposcelis paeta (Psocoptera: Liposcelididae) was investigated in the laboratory. The combination treatments were more effective compared to the single treatments against all examined species. The combinations of DEBBM and imidacloprid and imidacloprid with the highest dose rate of B. bassiana provided the highest mortality values against all tested species for 90 days of storage period. The combination of DEBBM plus B. bassiana resulted to the highest mortalities and to the lowest offspring production of all combinations tested after 180 days of storage. Mortality of adults for each test insect species was decreased over the storage period of 6 months, and the progeny production was increased with the extended storage period. Among the tested insect species, L. paeta was the most susceptible to all three grain protectants followed by C. ferrugineus, R. domina, and T. castaneum. The findings of the current study suggest that the use of DEBBM, imidacloprid, and B. bassiana as grain protectants may provide elevated control of major stored-grain insect species during a prolonged period of storage.

Interactions Between Two Invertebrate Pathogens: An Endophytic Fungus and an Externally Applied Bacterium.

The members of family Noctuidae exist in diverse environments and many species from this group are of agriculture importance, particularly Helicoverpa spp. Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae) is a major pest of many legumes and cereal crops. Due to environmental and regulatory concerns, safe alternatives to broad spectrum chemical insecticides are needed for the control of key noctuid pests such as H. armigera. A strain of Beauveria bassiana (Cordycipitaceae: Hypocreales) was evaluated for its ability to colonize endophytically in chickpea plants, and its effectiveness against second (L2) and fourth (L4) larval instars of H. armigera. B. bassiana was inoculated to chickpea plants through injection and endophytic establishment was confirmed by re-isolating the fungi from leaf samples. A detached leaf assay was used to evaluate pathogenicity. Bacillus thuringiensis was also applied to both larval stages through leaf dip method. In a novel approach, combined treatments of bacteria and endophytic fungi were compared with single-pathogen treatments. Relative to the single treatments, the combined pathogen treatments exhibited an increase in larval mortality, and decrease in pupation, adult emergence and egg eclosion. Specifically, synergistic effects on mortality were observed when larvae were exposed to simultaneous application of B. bassiana (1 × 108 conidia ml-1) with B. thuringiensis (0.75 µg ml-1). Both instars exhibited varying level of growth, development, frass production, diet consumption and fecundity when exposed to the chickpea leaves inoculated with endophytic B. bassiana and dipped with sub-lethal doses of B. thuringiensis. These findings indicate that the integrated application of endophytic colonized B. bassiana and B. thuringiensis can be effectively used against H. armigera.

Virulence of Entomopathogenic Fungi to Rhagoletis pomonella (Diptera: Tephritidae) and Interactions With Entomopathogenic Nematodes.

The objectives of this study were to quantify the virulence of four entomopathogenic fungal species to pupae of Rhagoletis pomonella (Walsh) (Diptera: Tephritidae) and to determine the potential to combine entomopathogenic fungi (EPFs) and entomopathogenic nematodes (EPNs) for biological control of this pest. The four species of EPFs included Beauveria bassiana (strain GHA), Metarhizium brunneum (strain F52), Isaria javanica (wf GA17), and Isaria fumosorosea (Apopka 97 strain). In laboratory assays, all fungi reduced adult emergence but there were no differences between fungal species. Isaria javanica and M. brunneum were examined further in a EPFs and EPNs bioassay that also included the EPNs Steinernema carpocapsae (ALL strain) and S. riobrave (355 strain). All nematodes and fungi were applied either alone or in combination (fungus + nematode). There were no differences between species within the same entomopathogen group (fungi and nematodes). However, the treatment with S. riobrave resulted in lower R. pomonella emergence than either fungal species. The combination of S. riobrave and I. javanica resulted in the lowest R. pomonella emergence (3%) at fourth-week interval, which was significantly lower than any of the single-agent applications, yet virulence of the other three combination treatments was not different from their respective nematode treatments applied alone. Additive interactions were detected for all fungus-nematode combinations. This study suggests that application of entomopathogenic nematodes and fungi could be an effective option to suppress R. pomonella populations.