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Tribolium castaneum is a challenging pest of stored products, causing significant economic losses. The present study explored the efficacy of Coridothymus capitatus essential oil and its primary constituent, carvacrol, as eco-friendly alternatives for managing this pest. To evaluate their insecticidal potential, repellency, fumigant toxicity, and antifeedant properties, progeny inhibition assays were performed. Carvacrol exhibited superior repellency compared to the essential oil, achieving a 92% repellency rate at 2 mg/cm 2 . Both compounds demonstrated significant fumigant toxicity against T. castaneum , with LC 50 values of 168.47 and 106.5 μL/L for the essential oil and carvacrol, respectively, after 24 h. Carvacrol also outperformed the essential oil in antifeedant activity, inducing an 80.7% feeding deterrence at 1.17 mg/g. Moreover, both treatments effectively suppressed the development of the pest's progeny. These results collectively underscore the potent insecticidal properties of C. capitatus essential oil and carvacrol, particularly carvacrol, as promising candidates for the sustainable management of T. castaneum in stored product protection.

Encapsulation of essential oils (EOs) is an important strategy that can be applied to intensify the stability and efficiency of these compounds in integrated pest management. The present study aimed to investigate the sub-lethal activity of polymer-based EOs nanoparticles against red flour beetle, Tribolium castaneum adults as an important critical pest of stored products. Chitosan nanoparticles (CSNPs) containing garlic and cinnamon essential oils (GEO and CEO) prepared using the ionic cross-link technique. Stability of nano-formulations evaluated over temperature and storage time. The fumigant effect (LC 10 , LC 20 , LC 30 ) and contact toxicity (LC 10 , LC 15 , LC 25 ) determined. In addition, the contact toxicities of EOs and their nanoparticles on nutritional indices evaluated. An olfactometer used to assess the repellent activity of EOs and EOs loaded in CSNPs (EOs@CSNPs) in sub-lethal fumigant concentrations. Characterization results showed GEO loaded in CSNPs has particle size of 231.14 ± 7.55 nm, polydispersity index (PDI) value of 0.15 ± 0.02, encapsulation efficiency (EE) percentage of 76.77 ± 0.20 and zeta potential of - 18.82 ± 0.90 mV, in which these values for the CEO loaded in CSNPs (CEO@CSNPs) changed to 303.46 ± 0.00 nm, 0.20 ± 0.05, 86.81 ± 0.00% and - 20.16 ± 0.35 mV, respectively. A lower PDI value for both CSNPs showed an appropriate NPs size distribution. Furthermore, NPs size and encapsulation efficiency did not change in various temperatures and during four months which confirm good stability of the EOs@CSNPs. In LC 30 of GEO@CSNPs, the maximum repellency was determined as 66.66 ± 3.33. Among nutritional indices, in LC 25 of GEO@CSNPs, the relative growth rate (RGR) (0.011 ± 0.003 mg.mg -1 .day -1 ), relative consumption rate (RCR) (0.075 ± 0.004 mg.mg -1 .day -1 ) and feeding deterrence index (FDI) (54.662 ± 1.616%) were more affected, so GEO@CSNPs was more effective than CEO@CSNPs. The results of repellent and anti-dietary activities of EOs and EOs@CSNPs confirmed the higher repellency and adverse effectivity on nutritional indices of Tribolium castaneum pest treated with EOs@CSNPs compared to free EOs. In conclusion, the NPs form of GEO and CEO can be a novel and efficient carrier for improving the repellent and anti-nutritional activities of EOs., (© 2024. The Author(s).)

Background: Wheat grain containers or silos can be perfect habitats for insects, which generate large economic losses to grain production. Natural alternatives to synthetic insecticides have grown in popularity because of health, economic and ecological issues. Diatomaceous earth is a natural compound that has an insecticide effect by enhancing an insect's dehydration with no toxicity on mammals including humans. The aim of this study is to confirm the effect of diatomaceous earth as an insecticide for the wheat grain pest, the red flour beetle Tribolium castaneum (Coleoptera: Tenebrionidae) and demonstrate its underlying mechanisms as an insecticide by open-flow respirometry and scanning electron microscopy., Results: Survival bioassays of T. castaneum revealed a dose-dependent insecticide effect of diatomaceous earth. Gravimetric measurements showed that 2 days exposure to diatomaceous earth produces a significant increase of mass loss. Open-flow respirometry measurements showed an increase of total water emission rate on insects due to an increase of both, respiratory and cuticular water loss. Our study revealed that diatomaceous earth produces an increase of insect's cuticle permeability, which is responsible for elevated cuticular water loss. Scanning electron microscopy images provided visual evidence of the lipid absorbent properties of diatomaceous earth particles, and showed a tendency for higher, although not significant, damaged area of the cuticle's surface from diatomaceous earth treated insects compared to control ones., Conclusion: With state-of-the art techniques like open-flow respirometry and scanning electron microscopy, we demonstrated the underlying mechanism of diatomaceous earth as an insecticide and provided new cues for understanding the properties of the cuticle and its ecological importance. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

Tribolium castaneum, also known as the red flour beetle, is a polyphagous pest that seriously damages agricultural products, including stored and processed grains. Researchers have aimed to discover alternative pest control mechanisms that are less harmful to the ecosystem than those currently used. We conduct the purification and characterization of a protease inhibitor from C. plumieri seeds and an in vitro evaluation of its insecticidal potential against the insect pest T. castaneum. The trypsin inhibitor was isolated from C. plumieri seeds in a single-step DEAE-Sepharose column chromatography and had a molecular mass of 50 kDA. When analyzed for interaction with different proteolytic enzymes, the inhibitor exhibited specificity against trypsin and no activity against other serine proteases such as chymotrypsin and elastase-2. The isolated inhibitor was able to inhibit digestive enzymes of T. castaneum from extracts of the intestine of this insect. Therefore, we conclude that the new protease inhibitor, specific in tryptic inhibition, of protein nature from the seeds of C. plumieri was effective in inhibiting the digestive enzymes of T. castaneum and is a promising candidate in the ecological control of pests., Competing Interests: Declaration of competing interest The authors whose names are listed immediately below certify that they have NO affiliations with or involvement in any organization or entity with any financial interest (such as honoraria; educational grants; participation in speakers’ bureaus; membership, employment, consultancies, stock ownership, or other equity interest; and expert testimony or patent-licensing arrangements), or non-financial interest (such as personal or professional relationships, affiliations, knowledge or beliefs) in the subject matter or materials discussed in this manuscript., (Copyright © 2024 Elsevier Inc. All rights reserved.)

The thrust of the study was to determine the chemical composition of the essential oils extracted from Thymus pallescens de Noé and Cymbogon citratus Stapf. as well as to evaluate their efficacy in controlling Sitophilus zeamais Motschulsky and Tribolium castaneum (Herbst) in either single or combined populations. Carvacrol (56.04%) and geraniol (20.86%) were identified as the major constituents of T. pallescens and C. citratus respectively. The tested essential oils showed pronounced insecticidal activity against the pest species in relation with the applied doses. T. pallescens EO had the highest efficacy and S. zeamais was found to be more susceptible to both individual and combined treatments. With reference to the contact and fumigation assessments, T. pallescens EO effectuated corrected mortality rates ranging from 42.5-100% to 25-100% in S. zeamais with corresponding lethal concentration (LC 50 ) values of 17.7 µl/ml and 15µL/L air respectively. Whereas, the T. pallescens EO exhibited corrected mortality rates of 42.5-100% and 20-100% with corresponding LC 50 values of 18.1 µl/ml and 15.5 µL/L air against T. castaneum in contact and fumigation assessments, respectively. The corrected mortality rates increased for both insect species when using combination treatments, with significant increases in the LC 50 values, ranging from 8.59 to 49.9% for both pest species. Analysis of energy biomarkers in the treated insects indicate significantly increased protein and carbohydrate contents and decreased lipids levels. The study therefore demonstrated the bio-insecticidal toxicity of the EOs from T. pallescens and C. citratus against two important maize post-harvest pests, concurrently revealing significant positive and negative insecticidal activity gradients in relation to single or combined populations., (© 2024. The Author(s).)

Background: Reduced glutathione (GSH) synthesis is vital for redox homeostasis, cell-cycle regulation and apoptosis, and immune function. The glutamate-cysteine ligase catalytic subunit (Gclc) is the first and rate-limiting enzyme in GSH synthesis, suggesting the potential use of Gclc as a pesticide target. However, the functional characterization of Gclc, especially its contribution in metamorphosis, antioxidant status and insecticide resistance, is unclear in Tribolium castaneum., Results: In this study, we identified and cloned Gclc from T. castaneum (TcGclc) and found that its expression began to increase significantly from the late larvae (LL) stage (3.491 ± 0.490-fold). Furthermore, RNA interference-mediated knockdown of TcGclc resulted in three types of aberration (100% total aberration rate) caused by the downregulation of genes related to the 20-hydroxyecdysone (20E) pathway. This deficiency was partially rescued by exogenous 20E treatment (53.1% ± 3.2%), but not by antioxidant. Moreover, in the TcGclc knockdown group, GSH content was decreased to 62.3%, and total antioxidant capacity, glutathione peroxidase and total superoxide dismutase activities were reduced by 14.6%, 83.6%, and 82.3%, respectively. In addition, treatment with different insecticides upregulated expression of TcGclc significantly compared with a control group during the late larval stage (P < 0.01)., Conclusion: Our results indicate that TcGclc has an extensive role in metamorphosis, antioxidant function and insecticide resistance in T. castaneum, thereby expanding our understanding of GSH functions and providing a scientific basis for pest control. © 2024 Society of Chemical Industry., (© 2024 Society of Chemical Industry.)

This study aimed to develop a relatively natural and safe botanical insecticide for controlling the storage pest Tribolium castaneum in the egg and pupal stages. It examined how Elsholtzia densa Benth. essential oil (EO) and its primary components, β-caryophyllene and limonene, affected T. castaneum eggs and pupae through contact and fumigation. Among th, the contact activities of β-caryophyllene against T. castaneum eggs and pupae are LD 50 (median lethal dose, 50%) = 0.156 mg/cm 2 and ED 50 (median effective dose, 50%) = 16.35 mg/pupa respectively. The study also investigated the effect of β-caryophyllene and limonene on T. castaneum eggs and pupae through synergistic contact and fumigation. When the mixing ratio of β-caryophyllene and limonene was 7:1, the LD 50 value of contact activity against T. castaneum eggs was reduced to 0.100 mg/cm 2 , displaying an obvious synergistic effect. Experiments were conducted to investigate the antitoxic effect of β-caryophyllene on T. castaneum eggs and pupae, as well as its effects on the enzymatic activity of acetylcholinesterase, succinate dehydrogenase, glutathione S-transferase and carboxylesterase in T. castaneum pupae. Finally, the molecular docking techniques were employed to confirm the aforementioned effects on enzyme function. The findings of this study might help improve storage pest control with T. castaneum and create eco-friendly insecticides using E. densa EO, β-caryophyllene, and limonene., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier Inc.)

Background: Phosphine resistance in Tribolium castaneum challenges grain storage. This study investigates the impact of cytochrome P450 (CYP) enzymes and CYP346 family genes on phosphine resistance in Indian Tribolium castaneum populations., Methods: Seven field populations of T. castaneum were compared with Lab- susceptible population for their resistance to phosphine. The levels of cytochrome P450 enzyme and expression of certain CYP346 family genes were tracked in these populations., Results: The highly resistant Patiala population showed significantly increased CYP450 activity (11.26 ± 0.14 nmol/min/mg protein, 7.41-fold higher) compared to the lab-susceptible population (1.52 ± 0.09 nmol/min/mg protein) when assayed using 8 mM p-nitroanisole as the substrate. The mRNA expression was measured relative to the standard gene RPS18 and revealed significant upregulation of CYP346B1 and CYP346B3 in highly resistant populations Moga and Patiala ( CYP346B1 : 12.09 ± 2.19 to 21.74 ± 3.82; CYP346B3 : 59.097 ± 10.265 to 50.148 ± 8.272). Patiala's CYP346B1 exhibited an impressive 685.76-fold change, and Moga's CYP346B3 showed a 361.893-fold change compared to lab-susceptible. Linear regression confirmed robust fits for each gene (R2: 0.693 to 0.756). Principal component analysis (PCA) demonstrated a strong positive correlation between CYP346 genes expression; and cytochrome P450 activity. Patiala, Moga, and Hapur populations showed conformity, associating higher resistance with increased P450 activity and CYP346 gene expression. Cluster analysis highlighted a potential correlation between CYP346B1 , CYP346B2 , and CYP346B3 and P450 activity, with Patiala and Moga clustering together., Conclusions: Variability in CYP346B1 and CYP346B3 in strong resistance populations may contribute to adaptation and resistance mechanisms. The study provides insights into specific CYP346 family genes associated with phosphine resistance, emphasizing the intricate interaction between CYP450 detoxifying enzymes, CYP346 family genes, and resistance mechanisms. The upregulation of CYP346 genes suggests a survival advantage for T. castaneum against phosphine, diminishing phosphine's efficacy as a pest control measure., Competing Interests: The authors declare no conflict of interest., (© 2024 The Author(s). Published by IMR Press.)

Tribolium castaneum is a worldwide pest of stored grain that mainly damages flour, and not only causes serious loss of flour quality but also leads to deterioration of flour quality. Chemical detection plays a key role in insect behavior, and the role of odorant-binding proteins (OBPs) in insect chemical detection has been widely studied. OBPs can interact with small molecule compounds and thereby modulate variation in insecticide susceptibility in insects. In this study, a total of 65 small molecule compounds are selected to investigate the bound effect with TcOBP C12. The molecular docking results showed that β-caryophyllene, (-)-catechin, butylated hydroxytoluene, diphenyl phthalate and quercetin were the top five compounds, with docking binding energies of -6.11, -5.25, -5.09, -5.05, and - 5.03 Kcal/mol, respectively. Molecular dynamics analysis indicated that odorant binding protein C12 (TcOBP C12) exhibited high binding affinity to all five tested chemical ligands, evidenced by fluorescence quenching assay in vitro. In addition, the contact toxicity assay results suggested that these chemical agents caused a dose-dependent increase in mortality rate for T. castaneum adults. The TcOBP C12 gene was upregulated >2 times after a 24-h exposure, indicating that OBP C12 may play an important role for T. castaneum in response to these chemical agents. In conclusion, our results provide a theoretical basis for future insecticide experiments and pest management., Competing Interests: Declaration of competing interest The authors have declared that no conflict of interest., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Plant volatilomics such as essential oils (EOs) and volatile phytochemicals (PCs) are known as potential natural sources for the development of biofumigants as an alternative to conventional fumigant pesticides. This present work was aimed to evaluate the fumigant toxic effect of five selected EOs (cinnamon, garlic, lemon, orange, and peppermint) and PCs (citronellol, limonene, linalool, piperitone, and terpineol) against the Callosobruchus maculatus, Sitophilus oryzae, and Tribolium castaneum adults. Furthermore, for the estimation of the relationship between molecular descriptors and fumigant toxicity of plant volatiles, quantitative structural activity relationship (QSAR) models were developed using principal component analysis and multiple linear regression. Amongst the tested EOs, garlic EO was found to be the most toxic fumigant. The PCs toxicity analysis revealed that terpineol, limonene, linalool, and piperitone as potential fumigants to C. maculatus (< 20 µL/L air of LC 50 ), limonene and piperitone as potential fumigants to T. castaneum (14.35 and 154.11 µL/L air of LC 50 , respectively), and linalool and piperitone as potential fumigants to S. oryzae (192.27 and 69.10 µL/L air of LC 50 , respectively). QSAR analysis demonstrated the role of various molecular descriptors of EOs and PCs on the fumigant toxicity in insect pest species. In specific, dipole and Randic index influence the toxicity in C. maculatus, molecular weight and maximal projection area influence the toxicity in S. oryzae, and boiling point and Dreiding energy influence the toxicity in T. castaneum. The present findings may provide insight of a new strategy to select effective EOs and/or PCs against stored product insect pests., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Volatile secondary metabolites of plants interact with environments heavily. In this work, characteristic components of Michelia yunnanensis essential oils (EOs) were isolated, purified and identified by column chromatography, GC-MS and NMR. Leaves of M. yunnanensis were collected monthly and extracted for EOs to investigate chemical and insecticidal activity variations as well as potential influencing environments. Different organs were employed to reveal distribution strategies of characteristic components. Results of insecticidal activities showed that all EOs samples exerted stronger contact activity to Lasioderma serricorne, but repellent effect was more efficient on Tribolium castaneum. One oxygenated sesquiterpene was isolated from EOs, basically it could be confirmed as (+)-cyclocolorenone (1). It exerted contact toxicity to L. serricorne (LD 50 = 28.8 μg/adult). Chemical analysis showed that M. yunnanensis leaves in reproductive period would produce and accumulate more 1 than in vegetative period. Moreover, reproductive organs (flowers and fruits) contained more 1 than vegetative organs (leaves and twigs). Partial correlation analysis indicated that temperature-related elements positively correlated with the relative content of 1.

Preventing insect infestations is a critical component for establishing a pest management plan for stored-product insects. Long-lasting insecticide-treated netting (LLIN) is a potential tool to reduce insect movement by providing a chemical barrier, where insects may be able to physically pass through but ultimately die after exposure to the netting. Sublethal effects, such as reduced movement immediately after exposure and reduced ability to colonize, have been reported. Here we examine the sublethal effects of exposure to LLIN on two beetle species, Trogoderma variabile Ballion, warehouse beetle, and Tribolium castaneum Herbst, red flour beetle. We found that both female and male T. castaneum exposed to LLIN produced significantly less adult progeny than those exposed to untreated netting. Adult progeny output did not differ for T. variabile, but survivorship increased in T. variabile females exposed to LLIN. Importantly, the overall net reproductive rate was significantly decreased for both T. variabile and T. castaneum. The number of copulation attempts did not differ between males or females exposed to LLIN compared to untreated netting, but males exposed to LLIN showed increased durations of attempted and successful copulation events. This research demonstrates that the implications of LLIN exposure extend past direct mortality, with sublethal effects on reproductive output potentially increasing the effectiveness of this tool for preventing insect infestations., (Published by Oxford University Press on behalf of Entomological Society of America 2021.)

Chemical communication via pheromones is an integral component in insect behavior, particularly for mate searching and reproduction. Aggregation pheromones, that attract conspecifics of both sexes, are particularly common and have been identified for hundreds of species. These pheromones are among the most ecologically selective pest suppression agents. In this study, we identified an activating effect of the aggregation pheromone of the red flour beetle, Tribolium castaneum (Herbst) (Coleoptera: Tenibroidae) on a highly conserved circadian clock gene (Tctimeless). Tribolium castaneum is one of the most damaging cosmopolitan pest of flour and other stored food products. Its male produced aggregation pheromone, 4,8-dimethyldecanal (DMD), attracts both conspecific males and females and is used for pest management via monitoring and mating disruption. The Tctimeless gene is an essential component for daily expression patterns of the circadian clock and plays vital roles in eclosion, egg production, and embryonic development. In this study, we demonstrate that constant exposure to the species-specific aggregation pheromone led to Tctimeless up-regulation and a different pattern of rhythmic locomotive behavior. We propose that changing the well-adapted "alarm clock", using DMD is liable to reduce fitness and can be highly useful for pest management., (© 2021. The Author(s).)

In insects, the cytochrome P450 CYP6B family plays key roles in the detoxification of toxic plant substances. However, the function of CYP6 family genes in degrading plant toxicants in Tribolium castaneum, an extremely destructive global storage pest, have yet to be elucidated. In this study, a T. castaneum CYP gene, TcCYP6BQ7, was characterized. TcCYP6BQ7 expression was significantly induced after exposure to essential oil of the plant Artemisia vulgaris (EOAV). Spatiotemporal expression profiling revealed that TcCYP6BQ7 expression was higher in larval and adult stages of T. castaneum than in other developmental stages, and that TcCYP6BQ7 was predominantly expressed in the brain and hemolymph from the late larval stage. TcCYP6BQ7 silencing by RNA interference increased larvae mortality in response to EOAV from 49.67% to 71.67%, suggesting that this gene is associated with plant toxicant detoxification. Combined results from this study indicate that the CYP6 family gene TcCYP6BQ7 likely plays a pivotal role in influencing the susceptibility of T. castaneum to plant toxicants. These findings may have implications for the development of novel therapeutics to control this agriculturally important pest., Competing Interests: Declaration of competing interest The authors declared they do not have anything to disclose regarding conflict of interest with respect to this manuscript., (Copyright © 2018 Elsevier B.V. All rights reserved.)

As a medicinal plant, Artemisia annua is widely distributed in China. The purpose of this work was to analyze the chemical composition of essential oil from A. annua aerial portions, as well as to assess its repellent activity against Lasioderma serricorne and Tribolium castaneum adults. GC-FID and GC-MS analyses enabled the identification of 15 components representing 90.1% of the essential oil. The main components included artemisia ketone (70.6%), α -caryophyllene (5.1%) and germacrene D (3.8%). The essential oil was found to possess considerable ability to repel the two storage pests. This paper provided some evidence for the exploitation and utilization of A. annua resources as a natural repellent.

Tribolium castaneum (Herbst) is an important pest of stored grain, and benzoquinones secreted by this pest are harmful to humans. T. castaneum has developed strong resistance to fumigants, and an ecofriendly alternative for managing T. castaneum is urgently needed. 1-Octen-3-ol is a major volatile compound present in many mushrooms and fungi. In the current study, the direct toxicity and sublethal and transgenerational effects of 1-octen-3-ol on T. castaneum were investigated. Our results showed that 1-octen-3-ol had strong insecticidal activity against all developmental stages of T. castaneum and repelled T. castaneum adults. 1-Octen-3-ol showed negative effects on the development and reproduction of parental T. castaneum and the subsequent generation: LC 30 and LC 50 treatments significantly decreased the pupa and adult weights, pupation and emergence rates and fecundity of the parental generation. In addition, LC 50 treatment shortened the larval and pupal periods. In the unexposed progeny (F 1 ) of 1-octen-3-ol-exposed parents, decreased survival and pupation rates as well as reduced pupa and adult weights were observed under LC 30 and LC 50 treatments. In addition, a model food-system experiment showed that 1-octen-3-ol at 98 μL/L exhibited an efficacy of 100% after 7 days of fumigation and completely eliminated T. castaneum offspring. Although a higher concentration of 1-octen-3-ol was needed to achieve an efficacy equal to that of the positive control, dichlorvos (DDVP), 1-octen-3-ol promoted the seedling growth of wheat seeds, suggesting that the concentration used was not only acceptable but also beneficial for wheat seeds. Overall, 1-octen-3-ol seems to be a promising candidate for use as a fumigant and repellent against T. castaneum as well as a seed protectant., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Knockdown and mortality of adults of the red flour beetle, Tribolium castaneum (Herbst) and the confused flour beetle, Tribolium confusum Jacquelin du Val, were assessed after exposure to two contact insecticides, chlorfenapyr and cyfluthrin, on a concrete surface. Individuals were rated on a scale for knockdown of exposed adults according to their mobility from 1, representing immobilized adults to 5, representing normally moving (similar to the controls). Only cyfluthrin gave immediate knockdown. Adults were rated at 1, 3 and 7 days post-exposure. After the final assessment, adults were discarded and the same procedure was repeated for 5 consecutive weeks with new adults exposed on the same treated surfaces. Despite initial knockdown, many individuals did not eventually die after exposure to cyfluthrin. In contrast, adults exposed to chlorfenapyr were not initially knocked down after exposure but most died after 7 days. These trends were similar during the entire 5-week residual testing period. The storage of the treated dishes in illuminated or non-illuminated conditions did not affect the insecticidal effect of either insecticide. The results of the present study can be further implemented towards the design of a "lethality index" that can serve as a quick indicator of knockdown and mortality rates caused after exposure to insecticides.

In this work, we investigated the bioactivities of the essential oil (EO) extracted from the Rhododendron thymifolium and its principal germacrone against Lasioderma serricorne and Tribolium castaneum . The EO was obtained by steam distillation. Germacrone was obtained by cryogenic crystallization. The bioactivity of EO and germacrone was tested via contact and repellent activity assays. The results showed that EO and germacrone possessed contact and repellent activities against two species of insects. EO exhibited obvious contact activity against the L. serricorn adults, larvae and T. castaneum larvae with LD 50 values of 29.15 µg/adult, 42.73 µg/larva, 19.65 µg/larva respectively. Germacrone exhibited excellent contact activity against the L. serricorne adults, larvae and the T. castaneum larvae with LD 50 values of 17.18 µg/adult, 20.94 µg/larva, 20.93 µg/larva respectively. And at the highest testing concentrations (78.63 and 15.73 nL/cm 2 ), the repellent activity of EO and germacrone on two target insects was comparable to that of the positive control (DEET) after 30 h exposure. In especially, in the treatment of the 120 h after the repellent activity of EO and germacrone against T.castaneum adults and larvae were still very significant and showed the same level percentage repellency as DEET. Meanwhile, germacrone exhibited inhibition of acetylcholinesterase activity with IC 50 values of 3%. The results indicated that the EO of R . thymifolium and germacrone had the potential to be developed as natural insecticides and repellents for the control of T. castaneum and L. serricorne .

The insecticidal activity is the result of a series of complex interactions between toxic substances as ligands and insect's enzymes as targets. Actually, synthetic insecticides used in pest control programs are harmful to the environment and may affect non-target organisms; thus, the use of natural products as pest control agents can be very attractive. In the present work, the toxic effect of aniseed ( Pimpinella anisum L.) essential oil (EO) and its nanoemulsion (NE) against the red flour beetle Tribolium castaneum , has been evaluated. To assess the EO mode of action, the impact of sub-lethal concentrations of aniseed EO and NE was evaluated on enzymatic and macromolecular parameters of the beetles, including aspartate aminotransferase (AST), alanine aminotransferase (ALT), total protein, total lipids and glucose. Finally, a molecular docking study was conducted to predict the mode of action of the major EO and NE components namely E -anethole, Limonene, alpha-himalachalene, trans-Verbenol and Linalool at binding site of the enzymes AST and ALT. Herein, the binding location of the main compounds in both proteins are discussed suggesting the possible interactions between the considered enzymes and ligands. The obtained results open new horizons to understand the evolution and response of insect-plant compounds interactions and their effect predicted at the molecular levels and side effects of both animal and human.

Resistance to phosphine fumigation has been frequently reported in insect pests of stored products and remains one of the obstacles in controlling these pests, including Tribolium castaneum. In this study, six field populations of T. castaneum were collected from different localities in China. Bioassay data showed that SZ population was strongly resistant to phosphine, followed by moderate-resistance populations WL and SF and three susceptible populations JX, YN, and ML. In addition, synergism assays showed that piperonyl butoxide significantly increased the toxicity of phosphine in resistant population SZ. Furthermore, CYP346B subfamily genes, CYP346B1, CYP346B2, and CYP346B3, were significantly overexpressed in resistant populations. Expression of CYP346B1, CYP346B2, and CYP346B3 were significantly upregulated following exposure to phosphine. RNAi assays showed that depletions on the expression levels of CYP346B1, CYP346B2, and CYP346B3 resulted in an increase of susceptibility to phosphine in T. castaneum, respectively. Our data demonstrated that CYP346B subfamily genes in T. castaneum were associated with the resistance of phosphine. Moreover, the study also increased our understanding of the molecular basis of phosphine resistance in stored pest insects., Competing Interests: Declaration of Competing Interest The authors declare that they have no conflict of interest., (Copyright © 2020 Elsevier Inc. All rights reserved.)

One new sesquiterpene (α-iso-cubebenol acetate, 8 ), together with 9 known compounds (1-7, 9, 10) were isolated from the stems of Schisandra chinensis (Turcz.) Baill. by repeated silica gel column chromatography. Based on the results of MS, NMR spectra and comparing with literature data, the six dibenzocyclooctadiene lignans were identified as schizandrin A to C (1-3), schizandrin (4), schisantherin A (5) and gomisin J (6), the two sesquiterpenes were identified as α-iso-cubebenol (7) and α-iso-cubebenol acetate (8), while the two triterpenic acids were identified as ganwuweizic acid (9) and kadsuric acid (10). The antifeedant activity of the 10 compounds against Tribolium castaneum adults was tested. Gomisin J (6) exhibited activity at 1500 ppm concentration with 40.3% antifeeding index percentages. As for the dibenzocyclooctene lignans (compounds 1-3 , 6 ), the number of methylenedioxies and the position of hydroxyl groups were the main factors to affect their antifeedant activities.

Terpinen-4-ol has high fumigating activity to stored-grain pests including Tribolium confusum. To understand the detoxification of terpinen-4-ol in insects, proteomic analysis was performed to identify related proteins and pathways in response to terpinen-4-ol fumigation in T. confusum. By using isobaric tags for relative and absolute quantitation (iTRAQ)-based strategy, 4,618 proteins were obtained from T. confusum adults in the present study. Comparative proteomic analysis showed that 148 proteins were upregulated and 137 proteins were downregulated in beetles under the LC 50 of terpinen-4-ol treatment for 24 hr. According to functional classifications, differentially expressed proteins (DEPs) were enriched in xenobiotic metabolism pathways. In the detoxification pathway, the levels of 25 cytochrome P450s, 5 glutathione S-transferases, and 2 uridine diphosphate (UDP)-glucuronosyltransferases were changed, most of which were upregulated in T. confusum exposed to terpinen-4-ol. The results indicated that terpinen-4-ol was potentially metabolized and detoxified by enzymes like P450s in T. confusum., (© 2019 Wiley Periodicals, Inc.)

Phosphine resistance is a worldwide issue threatening the grain industry. The cuticles of insects are covered with a layer of lipids, which protect insect bodies from the harmful effects of pesticides. The main components of the cuticular lipids are hydrocarbon compounds. In this research, phosphine-resistant and -susceptible strains of two main stored-grain insects, T. castaneum and R. dominica , were tested to determine the possible role of their cuticular hydrocarbons in phosphine resistance. Direct immersion solid-phase microextraction followed by gas chromatography-mass spectrometry (GC-MS) was applied to extract and analyze the cuticular hydrocarbons. The results showed significant differences between the resistant and susceptible strains regarding the cuticular hydrocarbons that were investigated. The resistant insects of both species contained higher amounts than the susceptible insects for the majority of the hydrocarbons, sixteen from cuticular extraction and nineteen from the homogenized body extraction for T. castaneum and eighteen from cuticular extraction and twenty-one from the homogenized body extraction for R. dominica . 3-methylnonacosane and 2-methylheptacosane had the highest significant difference between the susceptible and resistant strains of T. castaneum from the cuticle and the homogenized body, respectively. Unknown5 from the cuticle and 3-methylhentriacontane from the homogenized body recorded the highest significant differences in R. dominica . The higher hydrocarbon content is a key factor in eliminating phosphine from entering resistant insect bodies, acting as a barrier between insects and the surrounding phosphine environment., Competing Interests: The authors declare no conflict of interest.

Background: Citrus sinensis, Allium sativum, Ocimum basilicum, Mentha spicata, Curcuma longa, Carica papaya, Andrographis paniculata, Azadirachta indica, Cymbopogon citratus and Acorus calamus were compared in the form of extracts of chloroform, hexane and aqueous media for their effectiveness in repelling Tribolium castaneum. Filter paper of 9 cm in diameter was cut into halves. Using a micro-pipette, each extract was applied to one half of the filter paper. The volume applied was kept constant at 0.6 mL per 30 cm 2 . The filter paper was dried and reattached leaving a 0.5 cm gap between, in a Petri-plate. Ten adults of T. castaneum were introduced and the repellency was observed after 30, 60 and 180 min., Results: The aqueous extract of C. papaya exhibited the highest repellency (82-97%) followed by that of garlic bulbs (aqueous extract) with 86.67% repellency. Further, gunny bags were sprayed separately using bottle sprayers with the two most effective extracts at a constant volume of 42 mL per 575 cm 2 . These treated gunny bags were compared with malathion-treated gunny bags in the insect management unit. The observations were taken after 2, 8 and 24 h. Papaya leaf extract-treated gunny bags showed relative repellent activity similar to that of malathion-treated gunny bags after 24 h of treatment. Thus, gas chromatography-mass spectrometry analysis of aqueous extract of papaya leaves was performed., Conclusions: It can be concluded that extract of papaya leaves containing 2-methoxy-4-vinylphenol (8.47% peak area) can be used as a repellent biopesticide. © 2019 Society of Chemical Industry., (© 2019 Society of Chemical Industry.)

Several plant essential oils have been used against diverse insect pests since, unlike conventional pesticides, they pose almost no risk to humans and the environment. For this reason, the essential oil (EO) isolated from the fresh leaves of Crithmum maritimum L. and its fractions (F 1 -F 5 ) obtained by chromatographic simplification were investigated for their chemical profile, as well as for their toxicity and repellency effects against Tribolium castaneum (Herbst) adults. The analysis by GC/MS allowed the identification of 92.8-99.1 % of the compositions of the total oil (EO) and of its fractions (F 1 -F 5 ). The EO and its fractions F 3 -F 5 were characterized by the presence of a high amount of phenylpropanoids (94.4, 94.8, 93.6, and 88.7 %, respectively): in all the samples, dill apiole was the most abundant component (EO: 94.1 %, F 3 : 94.6 %, F 4 : 93.4 %, and F 5 : 83.3 %). In addition, the repellency assay results showed that the volatile fraction F 5 and the complete EO exhibited a higher repellency towards T. castaneum (97 % and 93 %, respectively) after 2 h of exposure at the dose of 0.04 μL/cm 2 . The median lethal dose of the topical application of the EO was 9 %. Furthermore, the fraction F 1 possessed interesting contact toxicity against T. castaneum (80 % of mortality) at the concentration of 10 %. These results suggested that the essential oil of C. maritimum leaves might be used as an alternative to synthetic insecticides in order to prevent insects from damaging the stored products., (© 2020 Wiley-VHCA AG, Zurich, Switzerland.)

Storing grains remain vulnerable to insect pest attack. The present study developed a biopesticide using biomolecules and their encapsulation in nanoparticles. A 25 kDa cysteine protease extracted from seeds of Albizia procera (ApCP) was encapsulated in graphene quantum dots (GQDs). The insecticidal activity of ApCP, with or without GQDs, against two stored grain insect pests, Tribolium castaneum (Herbst) and Rhyzopertha dominica (Fabricius) was explored. Insects were exposed to three concentrations 7.0, 3.5 and 1.7 mg of ApCP per a gram of wheat flour and grains. The insecticidal activity of ApCP encapsulated with GQDs was improved compared to that of ApCP without GQDs for both insect pests. The number of eggs and larvae of T. castaneum was reduced by 49% and 86%, respectively. Larval mortality was increased to 72%, and adult eclosion of T. castaneum was reduced by 98% at a 7.0 mg/g concentration of ApCP with GQDs compared to that of ApCP without GQDs. Exposure to 7.0 mg/g ApCP with GQDs, the number of R. dominica eggs and larvae was reduced by 72% and 92% respectively, larval mortality was increased by 90%, and eclosion was reduced by 97%. The extraction, purification, characterization, quantification and encapsulation of ApCP with GQDs were also studied. Cysteine protease nanocarriers have the potential to control stored grain insect pests.

The essential oils (EOs) from aerial parts of Saussurea amara (L.) DC. (SAEO) and Sigesbeckia pubescens Makino (SPEO) were analyzed for their chemical composition by GC-MS, and their repellent activities against adults of the red flour beetle, Tribolium castaneum Herbst, as well as the booklouse, Liposcelis bostrychophila Badonnel, were evaluated for the first time. Results of GC-MS analysis indicated that both SAEO and SPEO were characterized by high content of sesquiterpenoids (relative content > 70%) including oxygenated sesquiterpenoids. The two oil samples and their major component caryophyllene oxide exerted beneficial repellent effects on T. castaneum and L. bostrychophila at 2 and 4 h post-exposure. At 4 h post-exposure, the PR value of caryophyllene oxide could still reach 92% (class V) against T. castaneum at minimum testing concentration of 3.15 nL/cm 2 , and this compound was observed to result in the greatest repellency (PR = 100%) against L. bostrychophila at 12.63 nL/cm 2 . This work confirmed the potent repellent efficacy of SAEO and SPEO for controlling pest damage and suggested their potential to be developed into botanical repellents.

Background: Many insect cytochrome P450 proteins (CYPs) are involved in the metabolic detoxification of exogenous compounds such as plant toxins and insecticides. Tribolium castaneum, the red flour beetle, is a major agricultural pest that damages stored grains and cereal products. With the completion of the sequencing of its genome, two T. castaneum species-specific CYP genes, CYP4BN6, and CYP6BQ11, were identified. However, it is unknown whether the functions of most CYPs are shared by TcCYP4BN6 and TcCYP6BQ11, and the upstream regulatory mechanism of these two CYPs remains elusive., Results: QRT-PCR analysis indicated that TcCYP4BN6 and TcCYP6BQ11 were both most highly expressed at the late pupal stage and were mainly observed in the head and gut, respectively, of adults. Moreover, the transcripts of these two CYPs were significantly induced by dichlorvos and carbofuran, and RNA interference (RNAi) targeting of each of them enhanced the susceptibility of beetles to these two insecticides. Intriguingly, knockdown of the latrophilin (lph) gene, which has been reported to be related to the insecticide susceptibility, reduced the expression of TcCYP4BN6 and TcCYP6BQ11 after insecticide treatment, suggesting that these two CYP genes are regulated by lph to participate in insecticide susceptibility in T. castaneum., Conclusion: These results shed new light on the function and mechanism of CYP genes associated with insecticide susceptibility and could facilitate research on appropriate and sustainable pest control management. © 2019 Society of Chemical Industry., (© 2019 Society of Chemical Industry.)

Several pyrrole derivatives exhibit insecticidal activity and can be effective as grain protectants. In the present study, we evaluate the insecticidal efficacy of six novel pyrrole derivatives, namely methyl 3-(methylthio)-4,6-dioxo-5-phenyl-1,3a,4,5,6,6a-hexahydropyrrolo[3,4-c]pyrrole-1 carboxylate (compound syn) (2a-syn), methyl 3-(methylthio)-4,6-dioxo-5-phenyl-1,3a,4,5,6,6a-hexahydropyrrolo[3,4-c]pyrrole-1-carboxylate (compound anti) (2a-anti), methyl 3-(benzylthio)-4,6-dioxo-5-phenyl-1,3a,4,5,6,6a-hexahydropyrrolo[3,4-c]pyrrole-1-carboxylate (compound syn) (2f-syn), methyl 3-(benzylthio)-4,6-dioxo-5-phenyl-1,3a,4,5,6,6a-hexahydropyrrolo[3,4-c]pyrrole-1-carboxylate (compound anti) (2f-anti), methyl 3-(butylthio)-4,6-dioxo-5-phenyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrrole-1-carboxylate (3e), and methyl 2-benzyl-3-(methylthio)-4,6-dioxo-5-phenyl-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrrole-1-carboxylate (0665), against four important species infesting stored products, the rice weevil, Sitophilus oryzae (Coleoptera: Curculionidae), the lesser grain borer, Rhyzopertha dominica (Coleoptera: Bostrychidae), the confused flour beetle, Tribolium confusum (Coleoptera: Tenebrionidae), and the Mediterranean flour moth, Ephestia kuehniella (Lepidoptera: Pyralidae). The six pyrrole derivatives were evaluated on wheat at different doses (0.1, 1, and 10 ppm) and exposure intervals (7, 14, and 21 days). For S. oryzae adults, the highest mortality was recorded at 10 ppm of 2a-syn (36.7%) followed by 2f-syn (32.2%) and 2f-anti (27.8%) after 21 days of exposure. Regarding progeny production, the application of the six pyrrole derivatives significantly reduced offspring emergence if compared with the controls. After 21 days, mortality of R. dominica reached 50% testing 10 ppm of 2f-syn, followed by 2a-syn (46.7%), 2f-anti (41.1%), and 2a-anti (33.3%), while for 3e and 0665, mortality remained low, not exceeding 17.8%. Mortality of T. confusum adults was very low, ranging from 0 to 16.7% after 21 days of exposure. Progeny production was low (< 1.7 individuals per vial) for all doses of the tested pyrrole derivatives, including control vials. For 2a-syn, 2a-anti, 2f-anti, and 0665, no progeny production was recorded testing 1 and 10 ppm, while for 2f-syn and 3e, no offspring emergence was noted testing 10 ppm. For T. confusum larvae, after 21 days of exposure, mortality reached 62.2% testing 10 ppm of 3e followed by 0665 (55.6%) and 2a-anti (42.2%). For E. kuehniella larvae, mortality reached 57.8% at 10 ppm of 2a-syn, followed by the pyrrole derivative 2f-anti (43.3%) after 21 days of exposure. Overall, these results show that the efficacy of pyrrole derivatives strongly varied according to the exposure interval, tested dose, treated insect species and developmental instar. The tested pyrrole derivatives, with special reference to 2a-syn, 2a-anti, 2f-syn, 2f-anti and 0665, are slow-acting compounds exerting relevant toxicity on key stored-product pests over time. They can be considered further for assays with selected blends aiming to develop novel control tools against stored-product pests in real-world conditions.

The essential oil extracted from roots and rhizomes of Ligusticum jeholense Nakai et Kitagawa was investigated for its chemical composition by GC-MS analysis, and evaluated for its contact toxicity and repellency against Tribolium castaneum and Lasioderma serricorne, along with some of its individual components. The essential oil was rich in aromatics (65.34%) with low molecular weight. Major components included sedanolide (33.95%), 3-butylidenephthalide (18.76%), spathulenol (8.90%) and myristicin (6.76%). The results of bioassays indicated that the essential oil of L. jeholense and 3-butylidenephthalide possessed significant repellent activities against T. castaneum at 2 and 4 h post-exposure. Meanwhile, 3-butylidenephthalide had potent contact toxicity against L. serricorne (LD 50 = 13.64 µg/adult). The minor component n-butylbenzene in the oil was highly toxic to T. castaneum (LD 50 = 23.99 µg/adult) and L. serricorne (LD 50 = 7.86 µg/adult) in contact assays, but failed to repel these beetles at all testing concentrations. Spathulenol and myristicin exerted good insecticidal and repellent effects on the two target insects. This work suggests that the essential oil of L. jeholense has promising potential for development as natural insecticide or repellent to control pest damage in warehouses.

Latrophilin (LPH) is an adhesion G protein-coupled receptor (aGPCR) that participates in multiple essential physiological processes. Our previous studies have shown that lph is not only indispensable for the development and reproduction of red flour beetles (Tribolium castaneum), but also for their resistance against dichlorvos or carbofuran insecticides. However, the regulatory mechanism of lph-mediated insecticide susceptibility remains unclear. Here, we revealed that knockdown of lph in beetles resulted in opposing changes in two chemoreception genes, chemosensory protein 10 (CSP10) and odorant-binding protein C01 (OBPC01), in which the expression of TcCSP10 was downregulated, whereas the expression of TcOBPC01 was upregulated. TcCSP10 and TcOBPC01 were expressed at the highest levels in early pupal and late larval stages, respectively. High levels of expression of both these genes were observed in the heads (without antennae) of adults. TcCSP10 and TcOBPC01 were significantly induced by dichlorvos or carbofuran between 12 and 72 h (hrs) after exposure, suggesting that they are likely associated with increasing the binding affinity of insecticides, leading to a decrease in sensitivity to the insecticides. Moreover, once these two genes were knocked down, the susceptibility of the beetles to dichlorvos or carbofuran was enhanced. Additionally, RNA interference (RNAi) targeting of lph followed by exposure to dichlorvos or carbofuran also caused the opposing expression levels of TcCSP10 and TcOBPC01 compared to the expression levels of wild-type larvae treated with insecticides alone. All these results indicate that lph is involved in insecticide susceptibility through positively regulating TcCSP10; and the susceptibility could also further partially compensated for through the negative regulation of TcOBPC01 when lph was knockdown in the red flour beetle. Our studies shed new light on the molecular regulatory mechanisms of lph related to insecticide susceptibility., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Plants present a delimited reservoir of biologically active compounds. Many plants synthesize several compounds of secondary metabolism, such as alkaloids, terpenoids, phenolics, steroids, etc. Such compounds are generally thought to be involved in plant-insect interactions. Phytoecdysteroids are a class of chemicals that plants synthesize; these compounds are analogues of molting hormones produced by insects. In this work, the effect of the 20-hydroxyecdysone, which is a molecule that belongs to the family of phytoecdysteroids, was tested on an insect pest, Tribolium castaneum (Herbst). Firstly, the effect of this molecule on post-embryonic development parameters was tested after ingestion at 300, 600, 900, and 1,200 ppm. Secondly, the effect of the 20-hydroxyecdysone was also tested on the biological parameters (proteins, alpha-amylase, detoxification enzymes). The results of the post-embryonic parameters test showed an important induction of larval mortality and a significant reduction of pupation and adult emergence rates. On the other hand, the test on the biological parameters showed that the 20-hydroxyecdysone caused a significant decrease in the levels of soluble proteins in treated larvae. In addition, the alpha-amylase activity was significantly inhibited by the ingestion of the phytoecdysteroid. And there was also a disruption of detoxification enzymes. The whole of the disturbances recorded in this work prove that phytoecdysteroids are thought to have potential value on T. castaneum control., (© The Author(s) 2019. Published by Oxford University Press on behalf of Entomological Society of America.)

Background: Potassium ion channels play a critical role in the generation of electrical signals and thus provide potential targets for control of insect pests by RNA interference., Results: Genes encoding the small conductance calcium-activated potassium channel (SK) and the voltage-gated potassium channel (SH) were knocked down in Tribolium castaneum by injection and oral delivery of dsRNA (dsTcSK and dsTcSH, respectively). Irrespective of the delivery mechanism a dose-dependent effect was observed for knockdown (KD) of gene expression and insect mortality for both genes. Larvae fed a 400 ng dsRNA mg -1 diet showed significant gene (P < 0.05) knockdown (98% and 83%) for SK and SH, respectively, with corresponding mortalities of 100% and 98% after 7 days. When injected (248.4 ng larva -1 ), gene KD was 99% and 98% for SK and SH, causing 100% and 73.4% mortality, respectively. All developmental stages tested (larvae, early- and late-stage pupae and adults) showed an RNAi-sensitive response for both genes. LC50 values were lower for SK than SH, irrespective of delivery method, demonstrating that the knockdown of SK had a greater effect on larval mortality. Biosafety studies using adult honeybee Apis mellifera showed that there were no significant differences either in expression levels or mortality of honeybees orally dosed with dsTcSK and dsTcSH compared to control-fed bees. Similarly, there was no significant difference in the titre of deformed wing virus, used as a measure of immune suppression, between experimental and control bees., Conclusion: This study demonstrates the potential of using RNAi targeting neural receptors as a technology for the control of T. castaneum. © 2019 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry., (© 2019 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.)

Confused flour beetles are serious pests of stored grain products, and therefore, it is important to efficiently monitor and control their populations. Aggregation pheromones are commercially used for monitoring this beetle but their efficacy has been questioned and they may be inadequate for practical use. Food attractants as well as pheromones are commonly used for monitoring stored-product insects. However, food attractants may not be effective in the case of food handling facilities, which are already filled with food odours. The ancestors of flour beetles may have been associated with dead or decomposing woody vegetation, so we investigated the attractiveness of several wood odours to beetles using a pitfall olfactometer. The beetles were strongly attracted to all wood odours tested: Castanea crenata, Magnolia obovata, Paulownia tomentosa, Prunus jamasakura, and Zelkova serrata. The attractiveness of these wood odours was also stronger than that of the odours of the usual food of these beetles. Supercritical CO 2 extracts of these species of wood were also attractive to the beetles. The Z. serrata extract was the most attractive among these extracts, and was further analysed by gas chromatography mass spectrometry. One major compound, (-)-mellein, was detected in the extract. Synthetic (±)-mellein attracted the beetles.

Latrophilin (LPH) is known as an adhesion G-protein-coupled receptor which involved in multiple physiological processes in organisms. Previous studies showed that lph not only involved the susceptibility to anticholinesterase insecticides but also affected fecundity in Tribolium castaneum. However, its regulatory mechanisms in these biological processes are still not clear. Here, we identified two potential downstream carboxylesterase (cce) genes of Tclph, esterase4 and esterase6, and further characterized their interactions with Tclph. After treatment of T. castaneum larvae with carbofuran or dichlorvos insecticides, the transcript levels of Tcest4 and Tcest6 were significantly induced from 12 to 72 h. RNAi against Tcest4 or Tcest6 led to the higher mortality compared with the controls after the insecticides treatment, suggesting that these two genes play a vital role in detoxification of insecticides in T. castaneum. Furthermore, with insecticides exposure to Tclph knockdown beetles, the expression of Tcest4 was upregulated but Tcest6 was downregulated, indicating that beetles existed a compensatory response against the insecticides. Additionally, RNAi of Tcest6 resulted in 43% reductions in female egg laying and completely inhibited egg hatching, which showed the similar phenotype as that of Tclph knockdown. These results indicated that Tclph affected fecundity by positively regulating Tcest6 expression. Our findings will provide a new insight into the molecular mechanisms of Tclph involved in physiological functions in T. castaneum.

The essential oil was extracted from the roots of Valeriana officinalis L. by hydrodistillation. The qualitative and quantitative analysis of its chemical constituents was conducted on GC-MS and GC-FID in this study. Seventeen compounds were detected and the major constituents included bornyl acetate (48.2%) and camphene (13.8%). The toxic and repellent effects of the essential oil and its two major constituents were evaluated on Liposcelis bostrychophila and Tribolium castaneum. The results of bioassays indicated that the essential oil showed the promising fumigant and contact toxicity against L. bostrychophila (LC 50  = 2.8 mg/L air and LD 50  = 50.9 μg/cm 2 , respectively) and the notable contact effect on T. castaneum (LD 50  = 10.0 μg/adult). Meanwhile, the essential oil showed comparable repellent effect on T. castaneum at all testing concentrations. Bornyl acetate and camphene also exhibited strong fumigant and contact toxicity against both species of pests (LC 50  = 1.1, 10.1 mg/L air and LD 50  = 32.9, 701.3 μg/cm 2 for L. bostrychophila; > 126.3, 4.1 mg/L air, and 66.0, 21.6 μg/adult for T. castaneum). Bornyl acetate and camphene showed moderate repellent effect on T. castaneum and conversely showed attractant effect on L. bostrychophila. This work highlights the insecticidal potential of V. officinalis, which has been noted as a traditional medicinal plant.

The study reports efficacy of Melissa officinalis L. essential oil (MOEO) as a safe plant-based insecticide against Tribolium castaneum Herbst (TC) by induction of oxidative stress. MOEO nanoencapsulation in chitosan matrix was performed to enhance its bioefficacy. GC-MS analysis of MOEO depicted geranial (31.54%), neral (31.08%), and β-caryophyllene (12.42%) as the major components. MOEO showed excellent insecticidal potential in contact (100% mortality at 0.157 μL/cm 2 ) and fumigant bioassays (LC 50  = 0.071 μL/mL air) and 100% repellency at concentration ≤ 0.028 μL/cm 2 . Increased reactive oxygen species (ROS), superoxide dismutase (SOD), catalase (CAT), and decreased ratio of reduced glutathione (GSH) to oxidized glutathione (GSSG) at the LC 50 dose suggested significant oxidative stress on TC in MOEO treatment sets. The encapsulated MOEO exhibited enhanced activity as fumigant (LC 50  = 0.048 μL/mL air) and showed significant antifeedant activity in situ (EC 50  = 0.043 μL/mL). High LD 50 value (13,956.87 μL/kg body weight of mice) confirmed favorable toxicological profile for non-target mammals. The findings depict potential of nanoencapsulated MOEO as an eco-friendly green pesticide against infestation of stored food by TC.

Thinner is a solvent that contains toluene, xylene, and other substances. The aim of this study was to examine the toxicity of thinner, xylene, and toluene on adult red flour beetles, Tribolium castaneum (Herbst), as well as the effects of thinner and toluene on gene expression. The major compounds identified in the thinner with peak area >2% were p-xylene (6%), toluene (4%), 2,4-dimethylheptane (3%), methylcyclohexane (2.75%), 2-methylheptane (2%), cyclohexanone (2.6%), and nonane (2.1%). Insects were exposed to solvents at 0-240 µl/L air for 4-48 hr. Animals that survived after a 4 hr exposure to 80 µl/L air were employed to determine mRNA expression using real-time PCR. Xylene was highly toxic, and the majority of the beetles did not survive 4 hr exposure at 40 µl/L air. The lethal concentration 50 (LC 50 ) values for toluene, xylene and thinner at 48 hr exposure were 97.7, <40 and 99.8 µl/L air, respectively. Thinner and toluene induced lethargy and sluggish movement in treated insects, while with xylene these effects were not observed. Glutathione-S-transferase (Gst) gene expression increased after thinner and toluene exposure. Superoxide dismutase (SOD) gene expression rose after toluene exposure. These results suggest that T. castaneum provides an alternative model for the study of toxicological effects of volatile aromatic compounds.

The natural and ecologically safe control of stored product insects has gained considerable attention in modern society. In this study of further searching for botanical pesticides from wild-growing plant, the contact toxicity and repellency towards Tribolium castaneum and Liposcelis bostrychophila were assessed for the essential oil (EO) from Ostericum viridiflorum. The EO was distilled from aboveground parts of O. viridiflorum and checked by gas chromatography-mass spectrometry. Twenty-two compounds were identified and the main components were β-caryophyllene (24.3%), α-humulene (21.0%), apiol (10.2%), and carotol (2.5%). For bioactivity tests, results indicated that the EO and its two main compounds (β-caryophyllene and α-humulene) all showed potent contact toxicity towards L. bostrychophila with LD 50 values of 44.52 μg/cm 2 , 74.11 μg/cm 2 , and 118.56 μg/cm 2 , respectively. The EO and the two main compounds also exhibited comparable repellency towards T. castaneum and L. bostrychophila. The results evidenced the EO of O. viridiflorum aboveground parts and its major compounds could be considered for the development of eco-friendly botanical insecticides and repellents in controlling stored product insects.

Background: Epigenetic modifications including DNA methylation and post-translational modifications of histones are known to regulate gene expression. Antagonistic activities of histone acetyltransferases (HATs) and histone deacetylases (HDACs) mediate transcriptional reprogramming during insect development as shown in Drosophila melanogaster and other insects. Juvenile hormones (JH) play vital roles in the regulation of growth, development, metamorphosis, reproduction and other physiological processes. However, our current understanding of epigenetic regulation of JH action is still limited. Hence, we studied the role of CREB binding protein (CBP, contains HAT domain) and Trichostatin A (TSA, HDAC inhibitor) on JH action., Results: Exposure of Tribolium castaneum cells (TcA cells) to JH or TSA caused an increase in expression of Kr-h1 (a known JH-response gene) and 31 or 698 other genes respectively. Knockdown of the gene coding for CBP caused a decrease in the expression of 456 genes including Kr-h1. Interestingly, the expression of several genes coding for transcription factors, nuclear receptors, P450 and fatty acid synthase family members that are known to mediate JH action were affected by CBP knockdown or TSA treatment., Conclusions: These data suggest that acetylation and deacetylation mediated by HATs and HDACs play an important role in JH action.

In this work, we investigated insecticidal and repellent activities of the essential oils extracted from Ajania nitida and Ajania nematoloba against Tribolium castaneum and Lasioderma serricorne adults. The components of essential oils were analyzed by GC-MS. The main components of A. nitida oil were camphor (20.76%), thujone (18.64%), eucalyptol (13.42%), borneol (8.32%) and those of A. nematoloba oil were β-pinene (34.72%), eucalyptol (24.97%) and verbenol (20.39%). The results showed that the two essential oils possessed insecticidal and repellent activities against two species of insects. A. nitida oil possessed contact and fumigant toxicity against T. castaneum (LD 50 = 30.10 µg/adult and LC 50 = 21.07 mg/L air) and L.serricorne (LD 50 = 17.51 µg/adult and LC 50 = 11.23 mg/L air). A.nematoloba oil showed contact and fumigant toxicity against T. castaneum (LD 50 = 102.29µg/adult and LC 50 = 69.45 mg/L air) and contact toxicity against L.serricorne (LD 50 = 53.43 µg/adult), but no obvious fumigant effect was observed against L.serricorne. Both of essential oils possessed strong repellent activity against T. castaneum and certain repellent activity against L.serricorne. Especially, A. nematoloba oil showed the same level percentage repellency as DEET(the positve control) against T. castaneum. The results indicated that the essential oils of A. nitida and A. nematoloba had the potential to be developed as natural insecticides and repellents for the control of T. castaneum and L.serricorne.

Stored products represent an enormous economic output, but insects regularly immigrate into stored products from the surrounding landscape throughout the postharvest supply chain. Long-lasting insecticide-incorporatednetting (LLIN), which usually contains an incorporated pyrethroid, has been used as part of a strategy to reduce the spread of malaria in tropical regions since the 1990s and has only recently been considered for its application in pre- and postharvest agricultural contexts. The goal of this study was to determine how short-term exposure to LLINs in the laboratory impacts the locomotory behavior and mortality for adult Tribolium castaneum (Herbst), (Coleoptera: Tenebrionidae) red flour beetle, and Rhyzopertha dominica (Fauvel), (Coleoptera: Bostrichidae) lesser grain borer, at different periods of time after exposure. Exposure to LLINs resulted in multiple-fold reductions in the distance moved and elevated angular velocity in both species that quickly took effect and persisted even after 168 h compared with adults exposed to control netting. R. dominica was somewhat more susceptible than T. castaneum to LLINs. Finally, the dispersal capacity of both species, measured as ability to move to a remote resource patch, was significantly impaired or absent after exposure to LLINs compared to adults exposed to control netting. Our results demonstrate that LLINs are a promising new technology for reducing infestation by stored product insects since even short exposures limit movement and ultimately lead to knockdown and death.

To maximize the extraction of antioxidants from Chenopodium formosanum seeds, the process factors, such as the ethanol concentration (0⁻100%), extraction time (30⁻180 min) and temperature (30⁻70 °C), for the extraction of the bioactive contents as well as the antioxidant capacity are evaluated using response surface methodology (RSM). The experimental results fit well with quadratic models. The extract was identified by GC/MS, and it was found that some active compounds had antioxidant, repellency and insecticidal activities. Various concentrations of the extract were prepared for the evaluation of the insecticidal activity against Tribolium castaneum , and the toxicity test results indicated that the extract was toxic to Tribolium castaneum , with an LC 50 value of 354.61 ppm. The oxidative stability of the olive oil determined according to the radical scavenging activity and p -anisidine test demonstrates that the extract obtained from the Chenopodium formosanum seeds can retard lipid oxidation.

The essential oils (EOs) extracted from Evodia lenticellata Huang and Evodia rutaecarpa (Juss.) Benth. leaves are screened to evaluate their contact toxicity and repellency towards Tribolium castaneum (Coleoptera: Tenebrionidae), Lasioderma serricorne (Coleoptera: Anobiidae) and Liposcelis bostrychophila (Psocoptera: Liposcelididae) adults. The EOs are obtained by hydrodistillation and analyzed by gas chromatography-mass spectrometry (GC-MS). The principal components in the E. lenticellata EO are identified to be caryophyllene oxide (28.5%), β-caryophyllene (23.1%), β-elemene (14.5%), and β-cubebene (4.7%), while the main components of the E. rutaecarpa EO are α-pinene (39.4%), β-elemene (13.5%), α-ocimene (7.6%), and α-selinene (4.0%). These two kinds of EOs and their individual compounds all showed different levels of contact toxicity and repellent activity against three stored-product insects.

Maize flour was infested with fresh emerged Tribolium castaneum adults, and its nutritional composition was evaluated after 0, 45, and 90 days of the infestation. Furthermore, 99% carbon dioxide was applied to different developmental stages of T. castaneum for its management. There were six treatments and five replicates; for each replicate, maize flour (150 g) was taken in a 250-ml plastic jar container, insects were released in the flour, and jars were placed into the incubator at a temperature of 28 ± 1 °C, 60 ± 5% RH. Similarly, the application of 99% CO 2 was done with four exposure times of 12, 24, 48, and 72 h, at 35 °C 65 ± 5% RH. Present results showed major fluctuations in the nutritive composition of maize flour. Increase in the moisture, fat, fiber, protein, and ash contents was directly proportional to the increase in infestation level and time, contrary to the depletion of carbohydrates and total weight loss. The results of CO 2 treatment indicated that pupa was the most resilient stage as compared with larvae and adult stage. The observed susceptibility order was as follows: adult > larvae > pupae. The maximum mortality of adult, larval, and pupal stages was recorded after 24, 48, and 72 h of CO 2 application, respectively. These findings might be helpful to develop an ecofriendly technique to manage this crucial pest.

There has been a considerable growth in interest to use RNA interference (RNAi) as a novel insect pest management strategy in the past 10 years. However, there has been virtually no information on insect population variations in response to double-stranded RNA (dsRNA) molecules. The objective of this study was to generate baseline susceptibilities of the red flour beetle (Tribolium castaneum) to an insecticidal dsRNA targeting vacuolar H + -ATPase subunit E gene (dsvATPaseE), and correlate the susceptibility data with sequence and expression variations of the target gene (vATPaseE), expression variations of the RNAi core genes, and overall genetic differences among a laboratory strain and seven geographical field populations of T. castaneum collected in China. Our results showed limited variations in the LD 50 values of dsvATPaseE, which ranged from 0.10 to 0.29 ng/larva among the laboratory strain and the seven field populations. Considering the overlapping of the 95% confidence intervals of their LD 50 values, there were no significant differences among the laboratory strain and field populations. We also found limited sequence polymorphisms and low frequencies of the polymorphisms of vATPaseE, and limited variations (<2-fold) of the endogenous expression of vATPaseE among the laboratory strain and field populations. However, we found considerable genetic variations among the individuals within each field population for most of eight loci and moderate to large genetic variations among the field populations. These results demonstrated that although the genetic variabilities were considerable among these field populations, the efficiency of RNAi targeting vATPaseE was highly consistent in T. castaneum. Our study provides work frames of resistance risk assessment for RNAi-based insect pest management programs., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Toxicity and repellency activity of Cleistanthus collinus (Roxb.) (CC) leaf extracts were evaluated against rice weevil, Sitophilus oryzae (L.) and red flour beetle, Tribolium castaneum (Herbst) under laboratory condition. Five concentration(s) (1%, 1.5%, 2%, 2.5% and 4%) with two controls (acetone and water) treatments along with deltamethrin were used for direct and contact residual toxicity. The insect that survived after CC exposure were transferred to an untreated feeding substrate and the population buildup of subsequent two generation were recorded after 30 (F 1 ) and 60 days (F 2 ). In the contact residual toxicity, highest CC concentration (4%) produced 75% mortality in S. oryzae and 62.5% mortality in T. castaneum during 7 days of exposure, whereas in direct toxicity the mortality were 81% and 58% respectively, for S. oryzae and T. castaneum. The long term effect of CC was apparent in both the insect species, where F 2 populations were significantly decreased in the CC treatments. CC treatment at 4% produced similar adult mortality in comparison to deltamethrin at 1%. In addition, repellent activity of CC extracts was observed against both S. oryzae and T. castaneum. This is the first step towards assessing the scientific basis for the understanding the effectiveness of CC extracts against stored grain pests and it could be a viable eco-friendly option for stored grain insect pest management., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Latrophilin of Tribolium castaneum (Tclph) has been reported to play crucial roles in growth, development and reproduction. However, the regulatory mechanism of Tclph associated with these physiology processes is unknown. Thus, the global transcriptome profiles between RNAi treated (ds-Tclph) and control larvae of T. castaneum were analyzed by RNA-sequencing. Totally, 274 differentially expressed genes (DEGs) were identified between the ds-Tclph and control samples. These DEGs were classified into 42 GO functional groups, including developmental process, reproduction and stress response. The results indicated that knockdown of Tclph disturbed the antioxidant activity process, and partially inhibited the serine protease (SP) and lipase signaling pathways to regulate the development and reproduction as well as the decreasing of the stress response in T. castaneum. Additionally, knockdown of Tclph suppressed IMD immunity pathways which likely modulated the effects of Tclph on stress response. Interestingly, CSPs, ESTs, CYPs, AOXs and BGs were significantly down-regulated in ds-Tclph larvae, implying that they cooperated with Tclph to reduce the activity of cellular metabolism system. FMOs was up-regulated in ds-Tclph insects suggested it may be involved in detoxifying alkaloid of insect metabolism system. These results implied that Tclph participated in phase 0, I and II cellular detoxification. Furthermore, RNAi against Tclph increased larval susceptibility to carbamates and organophosphates insecticides, supporting that Tclph was indeed involved into the insecticide susceptibility in T. castaneum.

The major chemical constituents of the essential oil extracted from Artemisia dubia wall. ex Bess. (Family: Asteraceae) were found as terpinolene (19.02%), limonene (17.40%), 2,5-etheno[4.2.2]propella-3,7,9-triene (11.29%), isoelemicin (11.05%) and p-cymene-8-ol (5.93%). Terpinolene and limonene were separated as main components from the essential oil. The essential oil showed fumigant toxicity against Tribolium castaneum and Liposcelis bostrychophila with LC 50 values of 49.54 and 0.74 mg/L, respectively. The essential oil and isolated compounds of A. dubia showed repellency activities against both insects. Terpinolene and limonene showed the fumigant toxicity against T. castaneum. Terpinolene showed obvious fumigant toxicity against L. bostrychophila. The results indicated that the essential oil of A. dubia had potential to be developed into natural insecticides for controlling stored product pests.

The speciation of a methanolic extract of Zanthoxylum armatum stem bark has enabled the isolation and characterization of 11 known lignans. Among them, five compounds ( 6 , 8 - 11 ) are reported in this plant for the first time. All of the chemical structures were elucidated on the basis of NMR spectral analysis. Additionally, their antifeedant activities against Tribolium castaneum were evaluated scientifically. Among them, asarinin ( 1 ), with an EC 50 of 25.64 ppm, exhibited a much stronger antifeedant activity than the positive control, toosendanin (EC 50 = 71.69 ppm). Moreover, fargesin ( 2 ), horsfieldin ( 3 ), and magnolone ( 10 ), with EC 50 values of 63.24, 68.39, and 78.37 ppm, showed almost the same antifeedant activity as the positive control. From the perspective of structure-effectiveness relationship, compounds with the chemical group of methylenedioxy exhibited higher antifeedant activities and have potential to be developed into novel antifeedants or potential lead compounds to protect food and crops in storage., Competing Interests: The authors declare no conflict of interest.