Evaluation of kaolinite-based particle films to control Tribolium species (Coleoptera: Tenebrionidae)

Adults of Tribolium castaneum (Herbst), the red flour beetle, and Tribolium confusum (du Val), the confused flour beetle, were exposed to kaolinite-based particle film dusts. When beetles were continuously exposed to the hydrophobic particle film M-96-018 at the rate of 0.1-0.5 mg/cm², all the T. castaneum at 0.1 mg/cm² were dead after 3 days, but 40±13.8% of the exposed T. confusum were still alive after 7 days. At higher concentrations, all the T. castaneum were dead after 2 days, but 5-6 days of exposure were needed to kill all T. confusum. In a subsequent test, adults of both species were exposed for 8-72 h to 0.5 mg/cm² of the particle film M-96-018, removed, then held without food for 1 week. No T. castaneum survived, while survival of the T. confusum ranged from 0 to 55±17.3%, depending on the exposure interval. In a test conducted at controlled conditions of 40%, 57% and 75% r.h., 27°C, T. confusum were exposed for 8-72 h to the particle film M-96-018 and a hydrophilic particle film M-97-009 at the rate of 0.5 mg/cm², then removed and held either with or without wheat flour for 1 week. All the T. confusum exposed to the particle film M-97-009 usually survived, while survival of the T. confusum exposed to the particle film M-97-018 after the 1-week holding period increased with increasing relative humidity and with the presence of food. The particle film M-96-018 was effective against both the Tribolium species, and appears to have a potential for use in management programs to control beetles within storage facilities.     

Differential susceptibilities of two closely-related stored product pests,the red flour beetle (Tribolium castaneum) and the confused flour beetle (Tribolium confusum), to five selected insecticides

The red flour beetle (Tribolium castaneum Herbst) and the confused flour beetle (Tribolium confusum Jacquelin du Val) are among the most commonly encountered insects infesting stored food, but their susceptibilities to different insecticides often differ significantly, which complicates efforts to manage populations in milling and processing facilities. In this study, the susceptibilities of T. castaneum and T. confusum late-stage larvae to five selected insecticides, esfenvalerate, pyrethrins, dichlorvos, methoprene and pyriproxyfen, were assessed with and without synergists using topical applications. In four-day bioassays (without flour) with dichlorvos, esfenvalerate and pyrethrins, T. castaneum larvae were less susceptible (low larval mortality) to dichlorvos and esfenvalerate than T. confusum, whereas the reverse was true for treatment with pyrethrins. Pre-treatment with one of three synergists, piperonyl butoxide (PBO, cytochrome P450 monooxygenase inhibitor), S,S,S-tributyl phosphorotrithioate (DEF, esterase inhibitor), or diethyl maleate (DEM, glutathione S-transferase inhibitor), suggested involvement of esterases in the detoxification of dichlorvos and pyrethrins, and cytochrome P450 monooxygenases in the detoxification of esfenvalerate in both species. Interestingly, pre-treatment with some synergists increased the toxicity of insecticides in only one species: DEM and DEF increased the toxicity of dichlorvos to T. castaneum, whereas only DEF increased dichlorvos toxicity to T. confusum. In 28-day bioassays with larvae treated with each of two insect growth regulators (IGRs), methoprene and pyriproxifen, T. confusum was the more tolerant species. All T. castaneum died in either the larval or pupal stages with either IGR. In contrast, only pyriproxyfen caused complete mortality in T. confusum larvae, and even the highest dose of methoprene allowed nearly 70% of T. confusum larvae to pupate, and 4.5% of adults to emerge. Our results show that although these two species are closely related, they display very different susceptibilities to different insecticides, and different metabolic detoxification mechanisms may contribute to their differential insecticide susceptibilities.     

Olfactory host location and host preference of Holepyris sylvanidis (Brèthes) (Hymenoptera: Bethylidae), a parasitoid of Tribolium confusum Jacquelin du Val and T. castaneum (Herbst) (Coleoptera: Tenebrionidae)

Parasitoids can suppress populations of their host and thus play a primary role in Integrated Pest Management. We studied foraging cues in Holepyris sylvanidis (Hymenoptera: Bethylidae), a larval parasitoid of Tribolium species, in a four-chamber olfactometer. H. sylvanidis is reported as a cosmopolitan parasitoid of Coleopteran including two major pests of stored products, the confused flour beetle Tribolium confusum and the red flour beetle T. castaneum. Our study reveals that the host complexes of both Tribolium species and different living host stages attract naive H. sylvanidis females, whereas no reaction was observed to uninfested substrates. Our findings may contribute to the development of biological control strategies of T. castaneum and T. confusum with parasitoids.     

Residual efficacy of synergized pyrethrin + methoprene aerosol against larvae of Tribolium castaneum and Tribolium confusum (Coleoptera: Tenebrionidae)

Wheat flour and different packaging surfaces (cardboard, flour bag, muslin bag, paper bag, pallet wrap, plastic overwrap, polyethylene) were exposed to aerosol formulations of either 1% active ingredient (AI) pyrethrin (synergized with piperonyl butoxide)+ 33.6% (AI) methoprene or 3% (AI) pyrethrin + 33.6% AI methoprene. The mixture was formulated as specified on the insecticide labels to give a 100 to 1 ratio of active ingredient pyrethrin to methoprene. Residual bioassays were conducted every two weeks for 16 weeks post-exposure to the aerosol by placing four-week-old larvae of the red flour beetle, Tribolium castaneum (Herbst), or the confused flour beetle, Tribolium confusum (Jacqueline duVal), on treated flour or a treated packaging surface with untreated flour added. T. castaneum was clearly the more susceptible of the two species. Less than 2% of T. castaneum larvae exposed to aerosol-treated flour or packaging surfaces emerged as normal adults, regardless of the pyrethrin concentration. Most of the T. castaneum larvae on treated flour did not advance to the pupal stage because they were either developmentally arrested or died as larvae. They were able to develop further on the treated packaging surfaces, but still could not emerge as adults. T. confusum larvae exposed to aerosol-treated flour or packaging surfaces were able to develop to the pupal or adult stage. Emergence of normal-appearing adults from T. confusum larvae exposed on the packaging surfaces treated with 1% pyrethrin + methoprene gradually increased (range of 29.7 ± 2.9 to 49.0 ± 6.7%, depending on the surface), whereas adult emergence of larvae exposed to treated flour peaked at 10 weeks post-exposure. However, when T. confusum was exposed to 3% pyrethrin + methoprene treated flour or packaging surfaces, adult emergence was reduced. Overall there were few significant differences attributable to the individual packaging surfaces.     

Impact of physical and biological factors on susceptibility of Tribolium castaneum and Tribolium confusum (Coleoptera: Tenebrionidae) to new formulations of hydroprene

Three separate experiments were conducted to: (1) evaluate two new commercial formulations (202-080 and 202-084) of the insect growth regulator hydroprene, (2) determine residual efficacy of hydroprene-treated concrete stored at different environmental conditions, and (3) assess the impact of accumulated flour on residual efficacy. In the first test, late instars of Tribolium castaneum (Herbst), the red flour beetle, and Tribolium confusum (du Val), the confused flour beetle, were exposed on concrete treated with hydroprene. At 40% relative humidity (r.h.), there was no difference between species regarding the percentage of individuals that stopped development in the larval stage (arrested larvae), but at 75% r.h. there were more arrested T. castaneum than T. confusum in all treatments except the low rate of formulation 202-084. No adult T. castaneum lived after emergence (live adults) at either relative humidity, but the percentage of live adult T. confusum ranged from 1.0% to 41.0%, depending on treatment. In the second test, late instars of T. confusum were exposed at 6 and 12 weeks post-treatment on concrete treated with the two experimental formulations and stored under different environmental conditions. At 6 weeks there was no difference between formulations. At 12 weeks, fewer live adults and more dead emerged adults with gross morphological deformities were found on concrete treated with formulation 202-084 and stored at 32°C, 75% r.h. compared to other treatment combinations. In the final experiment, wheat flour was added to treated concrete for 5 weeks before the bioassays were conducted with late-instar T. confusum. There were few live adults produced in the initial bioassays, and dead adults with gross morphological deformities ranged from 83.1% to 97.6%. However, in bioassays conducted with late-instar larvae at 6 weeks, most adults eventually emerged with few deformities. The presence of the flour apparently compromised residual control and the hydroprene was no longer effective. In summary, the new hydroprene formulations were equivalent to the registered product Gentrol^®. Tribolium confusum was less susceptible than T. castaneum, and residual control of T. confusum on a clean surface without flour lasted about 6-12 weeks.     

Efficacy of a volatile formulation of hydroprene (Pointsource™) to control Tribolium castaneum and Tribolium confusum (Coleoptera: Tenebrionidae)

Three- and 4-week-old Tribolium castaneum (Herbst), the red flour beetle, and Tribolium confusum (du Val), the confused flour beetle, were exposed at five different temperature-relative humidity (r.h.) combinations to a volatile formulation of the insect growth regulator (IGR) hydroprene (called Pointsource™). Typical effects associated with IGR exposure, such as arrested larval growth, morphological deformities in adults, twisted and deformed wings, and incomplete adult emergence were produced in both species. Tribolium castaneum and T. confusum were susceptible to Pointsource™, but T. castaneum appeared to be the more susceptible species. More 3-week-old larvae of both species were arrested in that stage compared to the 4-week-old larvae. Nearly all of the 3- and 4-week-old T. castaneum larvae that were able to complete development to the adult stage quickly died after they emerged and were grossly morphologically deformed. In contrast, some emerged adult T. confusum remained alive after they emerged and were not deformed in any manner or had only twisted and incomplete wings. A greater percentage of larvae of both species were arrested in the larval stage and more adults died after they emerged in exposure studies conducted at 32°C, 75% r.h. as compared with 32°C, 30% r.h., but the reverse was true for exposures conducted at 27°C. Pointsource™ appears to have excellent potential for use in controlling Tribolium species within indoor facilities.     

Tanacetum vulgare essential oil as grain protectant against adults and larvae of four major stored-product insect pests

In this study the adulticidal and larvicidal effect of tansy, Tanacetum vulgare L. (Asteraceae) essential oil (EO) was estimated against four noxious stored-product insect species; Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae), Tribolium confusum Jacquelin du Val (Coleoptera: Tenebrionidae), Tenebrio molitor L. (Coleoptera: Tenebrionidae) and Oryzaephilus surinamensis (L.) (Coleoptera: Silvanidae). The EO chemical composition, as determined by GC-MS, was dominated by oxygenated monoterpenes (68.2%), with borneol (13.6%), umbellulone (11.7%), artemisia ketone (9.3%), cis-chrysanthenol (6.9%), camphor (5.9%), and terpinen-4-ol (5.5%) as the major constituents. This profile was quite different from those previously reported from other T. vulgare European accessions which are characterized by high content of the toxic trans-thujone. The T. vulgare EO was applied as wheat protectant at two concentrations of 500 and 1000 ppm. Adult and larval mortality levels were estimated after 4, 8 and 16 h and 1, 2, 3, 4, 5, 6 and 7 days. Tanacetum vulgare EO caused complete mortality (100%) to T. castaneum larvae, but only 25.6% to adults at 1000 ppm 6 and 7 days post-exposure, respectively. The mortality rates of T. confusum larvae and adults were 56.7 and 8.9% on wheat treated with 1000 ppm EO, respectively after 7 days of exposure. The ΕΟ caused moderate mortality to T. molitor adults (52.2%), and low mortality to larvae (8.9%), at 1000 ppm at the end of the experimental period. High mortality of O. surinamensis larvae (93.3%) was noticed at 1000 ppm after 7 days of exposure, while only 13.3% of the exposed adults were dead at the same concentration and exposure interval. Overall, the T. vulgare EO could be considered as a potential wheat protectant for the management of important stored-product insects. However, its efficacy depends on the species and the life stage of the target pest.     

Efficacy of diatomaceous earth formulations admixed with grain against populations of Tribolium castaneum

The efficacy of diatomaceous earth (DE) to control stored-products Coleoptera on stored grain was examined against several populations of the red flour beetle, Tribolium castaneum (Herbst). Four commercially available DE formulations were tested: INSECTO^®, Perma-Guard™, Protect-It^® and the diatomite used for the production of Dryacide^®, each at six concentrations (100-1000 ppm). A great variation of efficacy was observed among the DE formulations tested. Protect-It at concentrations up to 400 ppm was found to be the most effective formulation to control red flour beetle populations. However, a concentration of 1000 ppm of Protect-It was necessary to control all adults of all populations. Most T. castaneum populations, except one from Ivory Coast (Asm), were more than 90% controlled with INSECTO and Dryacide DE at 600 ppm. At this concentration, about 88% and 22% Asm adults died with INSECTO and Dryacide DE, respectively. Perma-Guard was the least efficient DE formulation to control T. castaneum adults with three populations exhibiting some survival at 1000 ppm. Reduced susceptibility to DE was observed in two populations, Asm and Lab susceptible from Kansas (Lab-S). As neither population had been previously exposed to DE, it is suggested that red flour beetles may naturally vary in susceptibility to DE. In addition, it was found that some populations can be satisfactorily controlled with some DE formulations but not with others.     

Influence of temperature and relative humidity on the efficacy of thiamethoxam for the control of three stored product beetle species

Laboratory bioassays were carried out in order to evaluate the effectiveness of thiamethoxam in different levels of temperature and relative humidity (r.h.) against three major stored-grain beetle species, the lesser grain borer Rhyzopertha dominica (F.), the rice weevil, Sitophilus oryzae (L.) and the confused flour beetle, Tribolium confusum Jacquelin du Val. Adults of the three species were exposed on wheat treated with thiamethoxam at 0.1, 1 and 5 ppm and under six different combinations of temperature (20, 25 and 30 °C) and r.h. levels (55 and 75%). Our study showed that the increase in temperature increased mortality in all insect species, particularly at the lowest concentration. For some of the combinations tested, the low r.h. seemed to play a significant role in thiamethoxam efficacy in the case of R. dominica. Moreover, of the species tested, T. confusum was found to be the least susceptible, while R. dominica the most susceptible. In general, thiamethoxam was found to be effective against all three species, at concentrations that are comparable with most commercially available grain protectants.     

Insecticidal effect of six entomopathogenic nematode strains against Lasioderma serricorne (F.) (Coleoptera: Anobiidae) and Tribolium confusum Jacquelin du Val (Coleoptera: Tenebrionidae)

The insecticidal effect of Heterorhabditis bacteriophora Poinar (Nematoda: Heterorhabditidae), Heterorhabditis megidis Poinar, Jackson and Klein (Nematoda: Heterorhabditidae) and two strains of Steinernema carpocapsae Weiser (Nematoda: Steinernematidae) and Steinernema feltiae Filipjev (Nematoda: Steinernematidae) against the cigarette beetle, Lasioderma serricorne (Fabricius) (Coleoptera: Anobiidae) and the confused flour beetle, Tribolium confusum Jacquelin du Val (Coleoptera: Tenebrionidae) was examined under laboratory conditions. The nematodes were applied at the following doses: 0 (control), 100, 1000, 5000, 10,000 and 20,000 infective juveniles (IJs) per ml, corresponding to 0, 10, 100, 500, 1000, and 2000 IJs per insect, and their infectivity was tested at 27 °C and 70% r.h. after 4 and 8 days of exposure. Adult emergence and progeny production was evaluated 4 and 8 weeks after the initiation of the bioassays. In the case of L. serricorne adults, one strain of S. carpocapsae caused 15.6 and 58.9% mortality after 4 and 8 days exposure, respectively, in semolina treated with 20,000 IJs per ml. On the other hand, larval mortality of L. serricorne did not exceed 19% in all treatments tested. Similarly, larval mortality of T. confusum was low, reaching 15.2 and 22.4% after 4 and 8 days exposure, respectively, at the highest dose tested. This is the first report on the effect of entomopathogenic nematodes against the cigarette beetle L. serricorne. Further experimental work is required to optimize the conditions under which these entomopathogenic nematodes could be used for the effective control of stored-product pests.     

From immobilization to recovery: Towards the development of a rapid diagnostic indicator for phosphine resistance

The aim of this work was to evaluate the insect mobility patterns of phosphine-resistant and -susceptible adults of the red flour beetle, Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae) after exposure to phosphine. Exposure bioassays were carried out at two concentrations, 1000 and 3000 ppm, while adults were observed every 15 min, for a total period of 90 min. During this observation interval, adults were visually classified as active (able to walk normally), partially immobilized (not able to walk, but showing a minimal movement), or completely immobilized (no visible movement). After the observation period, all adults were placed in a phosphine-free environment, and they again were classified as active, partially immobilized or completely immobilized. At 1000 ppm, the majority of adults of the susceptible T. castaneum population were quickly immobilized after a 15 min observation period, while in contrast, the majority of adults of the resistant T. castaneum population were still active after the termination of the 90 min interval. At 3000 ppm, the percentage of immobilized susceptible adults was increased at the 15 min observation period, while the majority of resistant adults were immobilized only after 90 min. In the post-exposure period, the vast majority of the susceptible adults were dead. In contrast, most resistant adults recovered, regardless of the concentration that had been exposed. The results of this study delineate major differences in movement in phosphine-resistant and -susceptible T. castaneum strains, and can be applied as a quick diagnostic bioassay for the evaluation of resistance to phosphine in stored product insects.     

Physical and chemical properties of flour products affect the development of Tribolium castaneum

The rust-red flour beetle, Tribolium castaneum, is a polyphagous pest of cereal grains and many other stored products. The presence of this pest causes contamination and economic losses by reducing the quality and quantity of commodities in storage. This research was aimed to study the development of T. castaneum on different flour products and investigate the factors affecting the survivorship and development time of T. castaneum. We used 16 flour products differentiated by production process (commercial vs. non-commercial) and commodity types. We conducted experiments on the development of T. castaneum in the laboratory by placing 15 mating pairs of T. castaneum into a treatment jar and allowing them to feed on different flour products. The results showed that T. castaneum adults laid eggs in all flour products. The larvae could live and survive, although the population density was found to be significantly lower in the commercial products. The pupae and adult (F₁ progeny) were only found in all non-commercial products and three commercial products. In addition, the total development time of T. castaneum was shown to be significantly faster in commercial products and did not differ among commodity types. We found that particle size variation and protein content of flour products affected the survivorship and development time of T. castaneum. In conclusion, understanding the particle size and chemical composition of flour products is a pivotal step in developing a pest management strategy for T. castaneum.     

Initial and delayed mortality of late-instar larvae, pupae, and adults of Tribolium castaneum and Tribolium confusum (Coleoptera: Tenebrionidae) exposed at variable temperatures and time intervals

Late-instar larvae, pupae, and adults of Tribolium castaneum (Herbst), the red flour beetle, or Tribolium confusum (DuVal), the confused flour beetle, were exposed for variable durations at 36-54 °C. Beetles were placed in laboratory ovens set at a baseline of 27 °C, the temperature was increased by 0.1 °C per minute until the target temperature was achieved, and beetles were then held for specified exposure durations. There was no mortality after initial exposure or after a 1-week holding period of any life stage of T. castaneum or T. confusum exposed for 32 h to 36, 39, or 42 °C. At 45 °C, there was no initial mortality of either species exposed for different time intervals except for those exposed for 28 h. However, there was a significant increase in mortality after the 1-week holding period of those beetles exposed initially for at least 16 h to 45 °C. There was a sharp increase in mortality after the initial exposures of 4 h at 48 °C; mortality of T. confusum larvae was 90.0±5.7% but was only 10.0±10.0% for larvae of T. castaneum, and no pupae of either species were dead. All life stages of both species were killed after the initial exposure of 12 h, and 1-week mortality of beetles exposed for 4 and 8 h was generally greater than initial mortality. At 51 and 54 °C, 2- and 1-h exposures, respectively, killed all life stages of each species. Mortality in conditions of gradual temperature increase was less than previous studies with sudden temperature increases.     

Effectiveness of diatomaceous earth for control of Sitophilus zeamais (Coleoptera: Curculionidae), Tribolium castaneum and Palorus subdepressus (Coleoptera: Tenebrionidae)

The effectiveness of diatomaceous earth (DE) or diatomite has been assessed against three major beetle pests of stored maize: Sitophilus zeamais (the maize weevil), Tribolium castaneum (the red flour beetle) and Palorus subdepressus (the depressed flour beetle). Maize has been treated with four doses of DE (1.5, 3, 4.5 and 6 g/kg) and four doses of Actellic Super™ Dust a chemical insecticide used as a reference (0.25, 0.5, 0.75 and 1 g/kg). The effective mortality was measured for each treatment after 1, 2, 4, 7 and 14 d of exposure. The corrected mortality rates were calculated by considering the mortality in the control group. Diatomaceous earth was as effective as Actellic Super™ Dust but required higher doses. Diatomaceous earth acted faster on S. zeamais and P. subdepressus compared to T. castaneum. The highest dose tested for DE caused the same mortality in T. castaneum as the recommended dose of Actellic Super™ Dust. Diatomaceous earth is a good alternative for the control of these three devastating insect pests of maize stocks and can readily be incorporated into integrated stored products pest management programs.     

Characterization of volatile organic compounds released by granivorous insects in stored wheat

Identification of the volatile organic compounds released by insects can be used to detect insects in stored grains. An attempt was made to identify the volatile organic compounds released by Tribolium castaneum (red flour beetle) and Cryptolestes ferrugineus (rusty grain beetle) by headspace analysis. Feasibility of the automatic headspace sampler in headspace analysis was found to be positive. The amount of volatiles produced by T. castaneum adults increased with an increase in insect density. The concentration of methyl-1,4-benzoquinone; ethyl-1,4-benzoquinone; and 1-tridecene released by ten adult insects were: 8.5, 9.1 and 10.6 μg/100 μL compared to 7, 8 and 4.2 μg/100 μL for five adult insects. Extreme high or low temperature leading to death produced very high amounts of volatiles compared to insects kept at 35 °C. The larvae of the T. castaneum insects did not produce any volatiles at ambient condition or at extreme cold or hot condition. The C. ferrugineus adults did not produce any detectable amount of volatiles even at very high insect density after up to 3 days. The results of the combination of T. castaneum and C. ferrugineus insects gave the same volatile compounds as produced by T. castaneum insects alone.     

The effect of a new chitin synthesis inhibitor, novaluron, on various developmental stages of Tribolium castaneum (Herbst)

Novaluron, a novel chitin synthesis inhibitor (CSI), was tested against the various developmental stages of Tribolium castaneum (Herbst), at concentrations ranging from 0.1 to 100 ppm. It did not kill T. castaneum adults at these concentrations, but at 1.0 ppm it caused total mortality of third-instar larvae. Novaluron did not affect the number of eggs laid by T. castaneum adults that were exposed to treated wheat flour, but it totally inhibited their hatching after the third day of exposure at a concentration of 1 ppm. On the first day after infestation novaluron did not totally prevent hatching, even at the highest tested concentration of 100 ppm. The time needed to restore egg hatchability after adults were transferred to untreated flour depended on the concentration of novaluron used in the treatment. When the adults of T. castaneum were exposed to novaluron-treated whole wheat grains (at 1 ppm), similar effects to those of the treated flour at the same concentration were observed: egg hatching was drastically reduced. The effect of uptake via contact of adults with novaluron-treated surfaces was examined by exposing them to a mixture of untreated flour and 10% novaluron-treated sand at a concentration of 10 ppm, and only 1% of the eggs hatched after 18 days of exposure. When adults were removed from treated surfaces of Petri dishes to untreated flour, the hatching rate was again reduced by 100%. It was concluded that the contact penetration of novaluron into T. castaneum adults prevents the hatching of eggs subsequently laid.     

Toxicity of coumaran to stored products beetles

The fumigant activity of coumaran was tested against mixed-age cultures of the rice weevil (Sitophilus oryzae L.), the lesser grain borer (Rhyzopertha dominica F.), the rust-red flour beetle (Tribolium castaneum Herbst), saw-toothed grain beetle (Oryzaephilus surinamensis L.) and flat grain beetle (Cryptolestes ferrugineus Stephens). T. castaneum was tolerant to coumaran, for total population extinction was achieved in 72 h exposure at 50 μg/L dose only. In contrast all developmental stages of O. surinamensis, C. ferrugineus and R. dominica were quite susceptible 100% mortality was attained at < 50 μg/L doses in 24 or 72 h exposure. Further results showed that T. castaneum, generally tolerant pest to fumigants and R. dominica, a predominant stored grain pest in India, could be controlled by coumaran. The findings indicate that, coumaran is good source for alternate to methylbromide for effective control of stored grain insect pests.     

Feasibility of protein fingerprinting technology for detecting Tribolium castaneum (Herbst) insect fragments in wheat flour

The red flour beetle, Tribolium castaneum (Herbst), is one of the predominant insects that cause serious decrease in quality of wheat flour. Protein mass fingerprinting has the potential to accurately detect insect parts in wheat flour, based on special peptides of T. castaneum. For determining the capability of protein fingerprinting for detecting insect parts in wheat flour, T. castaneum, wheat flour, and mixtures of T. castaneum and wheat flour were tested in this study. Three protein extraction buffers: 50% acetonitrile; sterilized water; and the buffer of 10 mM imidazole pH 7.0, 1 mM phenylthiourea, 1% Triton X-100 and protease inhibitor cocktail (PTU buffer), were compared for insect protein detection. An Agilent 1100 reversed phase high performance liquid chromatography (RP-HPLC) instrument was used for peptide purification and fractionation. Matrix-assisted laser desorption/ionization (MALDI) was used to read the RP-HPLC fractions, and the results were analyzed using sMART Engine, details given later. Fifty percent acetonitrile was adopted as protein extract buffer. Since very limited information about T. castaneum protein can be found in the database of sMART and National Center for Biotechnology Information (NCBI), peptide mapping method was used for identifying insect from the wheat flour. Peptide mapping method, which could set up biomarkers for each type of insect, was proven to be a more effective and practical way to detect insect contamination. This peptide mapping technique could obtain 78 biomarkers of T. castaneum, when T. castaneum was 0.1% in wheat flour by mass. This sensitivity needs to be increased further during future research.     

Toxicity and repellent action of Coffea arabica against Tribolium castaneum (Herbst) adults under laboratory conditions

Coffea arabica parchment extracts and caffeine isolated from the plant were evaluated randomly against 1-week-old adults of red flour beetle, Tribolium castaneum for fumigant toxicity and repellent action under laboratory conditions. The effects on detoxification enzymes and neuroenzyme was also determined. Among the various extracts prepared sequentially using hexane, dichloromethane, ethyl acetate and methanol as solvents, the dichloromethane extract did repel insects on contact (EC₅₀ = 4380.65 ppm). The repellence was more prominent when an olfactory response was measured (EC₅₀ = 2571 ppm). The active ingredient isolated from dichloromethane extract was identified as caffeine which showed very strong repellency as compared to the extract. In terms of toxicity of this extract, a significant mortality was recorded in fumigation assay (LC₅₀ = 5555 and 791 ppm, 24 and 48 h post-treatment, respectively). However, caffeine did not induce similar toxicity as the dichloromethane extract. The studies on the impact on detoxification enzymes of T. castaneum showed that dichloromethane extract inhibited carboxyl esterase activity, which possibly led to high toxicity. However, caffeine inhibited glutathione-s-transferase and induced carboxylesterase enzymes. It was, therefore, obvious that C. arabica parchment crude extracts have dual effects against T. castaneum adults, i,e, fumigant toxicity and repellent effects. However, the active compounds responsible for the two activities are surely different as caffeine could only induce repellent action against the beetles and the toxic compound needs to be identified, which is presently being investigated.     

Performance of diatomaceous earth and imidacloprid as wheat,rice and maize protectants against four stored-grain insect pests

Laboratory experiments were conducted in Pakistan to investigate the effectiveness of the diatomaceous earth (DE) formulation, Protect-It, at 150 ppm and imidacloprid, at 1.25, 2.5 and 5.0 ppm, alone or in combination, against the rusty grain beetle, Cryptolestes ferrugineus Stephens (Coleoptera: Laemophloeidae), the lesser grain borer, Rhyzopertha dominica (F.) (Coleoptera: Bostrychidae), the red flour beetle, Tribolium castaneum (Herbst) (Coleoptera: Tenebrionidae) and the psocid, Liposcelis paeta Pearman (Psocoptera: Liposcelididae). The bioassays were conducted at 25 °C, 65% relative humidity (r.h.) and continuous darkness by using adults of each insect pest that were exposed on treated or untreated wheat, rice and maize. Adult mortality counts were performed 1, 2, 4 and 7 days post-exposure. Progeny production was also evaluated 62 and 30 days after the last mortality count for beetles and psocid respectively. Results showed that mortality was increased with the increase of dose and exposure interval. Imidacloprid was more effective at the highest dose than the DE alone. The combination of DE plus imidacloprid resulted to higher mortalities than DE or imidacloprid alone at almost all exposure intervals. There was higher mortality on wheat than on rice or maize. In general, T. castaneum was the most tolerant insect species to any treatment while L. paeta was the least tolerant. The highest progeny production was observed for T. castaneum (i.e., 33.1 individuals per vial) while the lowest for L. paeta (i.e., 11.1 individuals per vial) on maize treated with Protect-It alone. The findings of the current study indicate that the combined use of DE and imidacloprid, as wheat, rice and maize protectants, may provide adequate level of management to several important stored-product insects.