Toxicity and Repellency Efficacy of Benzyl Alcohol and Benzyl Benzoate as Eco-Friendly Choices to Control the Red Flour Beetle Tribolium castaneum (Herbst. 1797).

Tribolium castaneum is a damaging pest of stored grains, causing significant losses and secreting lethal quinones, which render the grains unfit for human consumption. Chemical insecticides are the most commonly used approach for control; however, they create insecticide resistance and affect the health of humans, animals, and the environment. As a result, it is critical to find an environmentally friendly pest-management strategy. In this study, two naturally occurring chemicals, benzyl alcohol (BA) and benzoyl benzoate (BB), were investigated for insecticidal activity against T. castaneum using different assays (impregnated-paper, contact toxicity, fumigant, and repellency assays). The results showed that BA had a significant insecticidal effect, with the LC50 achieved at a lower concentration in the direct-contact toxicity test (1.77%) than in the impregnated-paper assay (2.63%). BB showed significant effects in the direct-contact toxicity test, with an LC50 of 3.114%, and a lower toxicity in the impregnated-paper assay, with an LC50 of 11.75%. Furthermore, BA exhibited significant fumigant toxicity against T. castaneum, with an LC50 of 6.72 µL/L, whereas BB exhibited modest fumigant toxicity, with an LC50 of 464 µL/L. Additionally, at different concentrations (0.18, 0.09, 0.045, and 0.0225 µL/cm2), BA and BB both showed a notable and potent repelling effect. BA and BB significantly inhibited acetylcholinesterase, reduced glutathione (GSH), and increased malondialdehyde (MDA) in treated T. castaneum. This is the first report of BA insecticidal activity against the red flour beetle. Also, the outcomes of various assays demonstrated that the application of BA induces a potent bio-insecticidal effect. BA may be a promising eco-friendly alternative to control T. castaneum due to its safety and authorization by the EFSA (European Food Safety Authority).

Biotoxicity comparison of Bacillus thuringiensis to control vector borne diseases against mosquito fauna.

The current study was designed to evaluate the biotoxicity of screened echo-friendly Bacillus thuringiensis strains from different areas of Pakistan. Out of 50 samples, 36% Bt. isolates were quarantined from soil containing cattle waste after morphological, biochemical, and molecular characterization. The toxicity bioassays with Bt. spores and protein diet proved that 11 Bt. isolates were utmost noxious to 3rd instar larvae of mosquitoes Aedes aegypti, Anopheles stephensi, and Culex pipiens. The entopathogenic activity of first 4 Bt. toxins against A. aegypti was highly lethal as compared to the other dipteran larvae. The toxicity (LC50) of spore diet of Bt. strains GCU-DAB-NF4 (442.730 ± 0.38 µg/ml), NF6 (460.845 ± 0.29 µg/ml), NF3 (470.129 ± 0.28 µg/ml), and NF7 (493.637 ± 0.70 µg/ml) was quite high against A. aegypti as compared to the C. pipiens after 24 h of incubation. The highest toxicity of total cell protein was shown by GCU-DAB-NF4 (LC50 = 84.10 ± 50 µg/ml), NF6 (95.122 ± 0.40 µg/ml), NF3 (100.715 ± 06 µg/ml), and NF5 (103.40 ± 07 µg/ml) against A. aegypti after 24 h. So, these strains a have great potential to be used as biological control especially against A. aegypti as compared to the C. pipiens.

Efficacy of Mentha aquatica L. Essential Oil (Linalool/Linalool Acetate Chemotype) against Insect Vectors and Agricultural Pests.

In recent years, agrochemical industries have been focused on the development of essential oil (EO)-based biopesticides, which can be considered valuable alternatives to traditional chemical products. The genus Mentha (Lamiaceae) comprises 30 species characterized by a wide range of biological activities, and some of their EOs showed good potential as pesticidal agents. In this regard, the aim of this study was to evaluate the insecticidal activity of the EO obtained from a rare linalool/linalool acetate chemotype of Mentha aquatica L. The EO was found to be highly effective against Culex quinquefasciatus (Say) 2nd instar larvae, Metopolophium dirhodum (Walker) adults, Spodoptera littoralis (Boisduval) 2nd instar larvae, and Tetranychus urticae (Koch) adults, showing lethal concentrations (LC50) or doses (LD50) of 31.5 ± 2.2 µL L-1, 4.9 ± 0.8 mL L-1, 18.5 ± 2.1 µg larvae-1, and 3.3 ± 0.5 mL L-1, respectively. On the contrary, Musca domestica L. adults and 3rd instar larvae of C. quinquefasciatus and S. littoralis were moderately affected by the treatment (LC50 or LD50: 71.4 ± 7.2 µg adult-1, 79.4 ± 5.2 µL L-1, 44.2 ± 5.8 µg larvae-1, respectively). The results obtained in this work demonstrated that various insects and pests could be differently sensible to the same EO and may lead to the exploitation of this plant or its major volatile compounds as novel ingredients of botanical insecticides and pesticides.

The bio-interactions between plants, insecticides and fertilizers: an innovative approach for the research of xenobiotic substances.

In this experiment carried out on Caribbean chili pepper plants (Capsicum chinensis), the bio-insecticide azadirachtin in combination with an NPK fertilizer proved to have a greater lethal impact on the larvae of Aedes albopictus than each substance on its own. This synergistic effect is noticeably important when both inputs are sprayed directly on the leaves of the plant (foliar application). While the plants treated with azadirachtin or NPK alone cause a 33.6% and 36.4% mortality respectively of the Ae. albopictus larvae, the combination of the two inputs induces a 74.4% mortality on the mosquito larvae. To account for this synergistic effect phenomenon inside the plant, the azadirachtin + NPK combination most likely interacts with the capsaicinoid compounds naturally produced by the plant. Not only does this study carried out on azadirachtin reveal major results but the methodology itself offers a most interesting approach on how to boost the agricultural inputs within the plants. As a matter of fact, this research axis demands developing since the control of pests harmful to men has been dramatically lacking insecticide molecules acting on new targets over the past three decades.

Natural insecticides for the control of urticating ant, Tetramorium aculeatum Mayr (Hymenoptera: Formicidae) in a coffee plantation of Southwestern Ethiopia.

Coffee, Coffea arabica L. is a vital crop in Ethiopia's social, cultural, and national economy. Despite its dominant role in the country, the crop is challenged by various insect pests, which reduce coffee yield and quality. The urticating ant or biting ant, Tetramorium aculeatum Mayr (Hymenoptera: Formicidae), is an important pest in coffee that affects farming activities in Southwestern Ethiopia. The objective of this study was to evaluate the efficacy of some natural insecticides for the control of biting ants. Nine insecticide treatments were evaluated both under ex-situ and in-situ conditions in CRD and RCBD designs, respectively. Under the ex-situ evaluation, all the insecticide treatments caused significantly higher mortality of biting ants and caused complete mortality within 24 h. Under field conditions, the insecticides showed significant variations in the percent of evacuated nests, and the number of newly constructed nests. In both efficacy tests of ex-situ and in-situ experiments, the mortality of biting ants and percent of evacuated nests in the coffee trees treated with oxymatrine and nimbicidine were found to be comparable and effective as the deltamethrin 2.5% EC. All the insecticide treatments significantly reduced the number of newly constructed nests compared to the control plot. On the other hand, deltamethrin 2.5% EC significantly decreased newly constructed nests after 30 days of application compared to the other insecticide treatments. Generally, the natural insecticides effectively minimized the biting ant infestation, but further studies are essential on the frequency of spray to use them for sustainable management approaches of biting ant in the coffee plantation.

Proteomic Analysis of the Venom from the Ruby Ant Myrmica rubra and the Isolation of a Novel Insecticidal Decapeptide.

Ants are a biodiverse group of insects that have evolved toxic venom containing many undiscovered bioactive molecules. In this study, we found that the venom of the ruby ant Myrmica rubra is a rich source of peptides. LC-MS analysis revealed the presence of 142 different peptides varying in molecular weight, sequence length, and hydrophobicity. One of the most abundant peaks was selected for further biochemical and functional characterization. Combined Edman degradation and de novo peptide sequencing revealed the presence of a novel decapeptide (myrmicitoxin) with the amino acid sequence NH2-IDPKLLESLA-CONH2. The decapeptide was named U-MYRTX-MRArub1 and verified against a synthetic standard. The amidated peptide was tested in a synthetic form to determine the antimicrobial activity towards the bacterial pathogens and insecticidal potential against pea aphids (Acyrthosiphon pisum). This peptide did not show antimicrobial activity but it significantly reduced the survival of aphids. It also increased the sensitivity of the aphids to two commonly used chemical insecticides (imidacloprid and methomyl). Since ant venom research is still in its infancy, the findings of this first study on venom peptides derived from M. rubra highlight these insects as an important and rich source for discovery of novel lead structures with potential application in pest control.

Conservation of stored food using plant's extracts. Effect of oregano (Origanum vulgaris) essential oils on the reproduction and development of flour moth (Ephestia kuehniella).

In the search for effective methods of biological control of stored products insect-pests, the essential oil of Origanum vulgaris was extracted using Clevenger apparatus and tested on reproduction and mortality of flour moth Ephestia kuehniella. Bio-insecticide works with a double action mechanism; administered by inhalation in pupae, essential oil affects their pupal development. It also disrupts the reproduction of exuviated adults by extending the preoviposition period and reducing the period of egg laying and fecundity because fecundated females cannot live more than four days compared to control group. Moreover, the essential oil administered by inhalation in adults causes a significant mortality rate compared to control group by reducing their lifespan. A repellent effect against larvae and adults of this pest was noticed in this work. This allowed us ranking this oil as "moderately repulsive".

Bio-insecticidal effects of Oleaster leaves aqueous extracts against Psylla larvae (Euphyllura olivina (Costa)), a primary pest of Olea europaea L.

Many plant species produce phenolic compounds in their various organs and their use in crop protection. These plant secondary metabolites may serve as toxins against the insect pests. The objective of this study was to evaluate in vitro the bio-insecticidal effect of an aqueous extract of wild Olive leaves on Psylla larvae (Euphyllura olivina), a primary pest of the cultivated Olive tree (Olea europaea L. subsp sativa). Two concentrations of 0.05g/ml and 0.1g/ml leaves grinding powder in distilled water were sprayed on branches infested with Psylla larvae. The obtained results revealed a very significant mortality rate of the larvae 24 hours after spraying. The chemical composition of Oleaster leaves aqueous extracts is determined by HPLC-DAD. The results show in majority the presence of phenolic compounds represented by oleuropein and its metabolite hydroxytyrosol. The phenolic compounds of the crude extract were at the origin of this mortality. The Analysis of Variance revealed highly significant results both between the sampled trees and between the tested concentrations. The Principal Component Analysis (PCA) revealed a close relation between the physiological state of the studied trees and the degree of their infestation by the phytophagus. Taking into account, the physical and chemical characteristics of the sampled soils, data analysis showed that trees growing on nitrogen-rich soils were more infested than those growing on soils rich in organic carbon (Corg) and phosphorus (Porg).

Evaluation of Bacillus vallismortis (Bacillales: Bacillaceae) R2 as insecticidal agent against polyphagous pest Spodoptera litura (Lepidoptera: Noctuidae).

The insecticidal potential of cells and acid-precipitated biomolecules (APB) of Bacillus vallismortis (Roberts) (Bacillales: Bacillaceae) R2 was evaluated against polyphagous pest Spodoptera litura. The intact cells of isolate R2 and its APB preparation significantly increased larval mortality. Both cells and APB significantly delayed the development and reduced adult emergence of S. litura. The toxicity of isolate R2 was evident from the emergence of morphologically deformed adults with crumpled and underdeveloped wings. The nutritional physiology of larvae fed on APB-supplemented diet was also adversely affected resulting in significant reduction of relative growth and consumption rate as well as efficiency of conversion of ingested and digested food. Thus, the intact viable cells and APB of B. vallismortis R2 may serve as environmental-friendly alternatives to chemical insecticides.

Essential Oils of Hyptis pectinata Chemotypes: Isolation, Binary Mixtures and Acute Toxicity on Leaf-Cutting Ants.

Leaf-cutting ants are pests of great economic importance due to the damage they cause to agricultural and forest crops. The use of organosynthetic insecticides is the main form of control of these insects. In order to develop safer technology, the objective of this work was to evaluate the formicidal activity of the essential oils of two Hyptis pectinata genotypes (chemotypes) and their major compounds on the leaf-cutting ants Acromyrmex balzani Emery and Atta sexdens rubropilosa Forel. Bioassays of exposure pathways (contact and fumigation) and binary mixtures of the major compounds were performed. The major compounds identified in the essential oils of H. pectinata were ß-caryophyllene, caryophyllene oxide and calamusenone. The essential oils of H. pectinata were toxic to the ants in both exposure pathways. Essential oils were more toxic than their major compounds alone. The chemotype calamusenone was more toxic to A. balzani in both exposure pathways. A. sexdens rubropilosa was more susceptible to the essential oil of the chemotype ß-caryophyllene in both exposure pathways. In general, the binary mixtures of the major compounds resulted in additive effect of toxicity. The essential oils of H. pectinata is a raw material of great potential for the development of new insecticides.

Assessment of the Toxicity of Sub-chronic Low and High Doses of the Bio-insecticide Spinosad on the Liver, Kidney and the Cerebellum in Male Albino Mice

ABSTRACT Spinosad (SPD) is a highly selective insect control product. However, it was reported that SPD has toxicity toward other non-target organisms. This study was conducted to address the toxic effect of two sub-chronic low and high doses; 35 and 350 mg/kg SPD on some biochemical, histological and immunohistochemical parameters of the liver, kidney and cerebellum. Thirty-six male Swiss mice were divided into three groups of 12 mice each; first group (G1) served as a control, second group (G2) received a low sub-chronic dose of SPD that is equal to 35 mg/kg, and third group (G3) received a high sub-chronic dose of SPD that is equal to 350 mg/kg. The results showed that mice which were received 350 mg/kg SPD showed a significant decrease in the body weight and a significant increase in their relative kidney and spleen weights. They also showed a significant increase in alanine aminotransferase (ALT), triglycerides and urea levels. Histopathological examination showed cytoplasmic degeneration and cell necrosis in the liver and kidney. Immunohistochemical examination showed that cerebellum illustrated several neurodegenerative changes and a down-regulation of synaptophysin-Syp. In conclusion, exposure to a high dose of SPD that is equal to 350 mg/kg could cause a marked toxicity on the liver, kidney and cerebellum in male albino mice.

Receptors are affected by selection with each Bacillus thuringiensis israelensis Cry toxin but not with the full Bti mixture in Aedes aegypti.

Bacillus thuringiensis israelensis (Bti) toxins are increasingly used for mosquito control, but little is known about the precise mode of action of each of these toxins, and how they interact to kill mosquito larvae. By using RNA sequencing, we investigated change in gene transcription level and polymorphism variations associated with resistance to each Bti Cry toxin and to the full Bti toxin mixture in the dengue vector Aedes aegypti. The up-regulation of genes related to chitin metabolism in all selected strain suggests a generalist, non-toxin-specific response to Bti selection in Aedes aegypti. Changes in the transcription level and/or protein sequences of several putative Cry toxin receptors (APNs, ALPs, α-amylases, glucoside hydrolases, ABC transporters) were specific to each Cry toxin. Selective sweeps associated with Cry4Aa resistance were detected in 2 ALP and 1 APN genes. The lack of selection of toxin-specific receptors in the Bti-selected strain supports the hypothesis that Cyt toxin acts as a receptor for Cry toxins in mosquitoes.

Regulation analysis of AcMNPV-mediated expression of a Chinese scorpion neurotoxin under the IE1, P10 and PH promoter in vivo and its use as a potential bio-insecticide.

OBJECTIVE: To analyze the regulation mechanism of AcMNPV (Autographa californica multicapsid nucleopolyhedrovirus)-mediated expression of BmK IT under IE1, P10 and PH promoters in the larva of Heliothis armigera.. RESULTS: The transcription level of BmK IT gene in midgut and epidermal tissue was analyzed by quantitative PCR. The start time of transcription of recombinant BmK IT gene was early under the regulation of IE promoter, whereas transcription of BmK IT was high under the regulation of P10 promoter in the midgut tissue of infected larvae. TdT-UTP nick-end labeling (TUNEL) assay showed the degree of apoptotic cell death in the midgut tissue of AcMNPV-BmK IT-transfected insect larvae was higher than that in the AcMNPV treatment group at 8 h post-infection. The time-effect relationship between the insect's humoral immunity and regulation of promoters was confirmed in the phenoloxidase activity assay. CONCLUSION: The anti-insect mechanism and regulation of different promoters in AcMNPV-BmK IT at molecular and cellular levels provide an experimental basis for the development of recombinant baculovirus biopesticides.

Pathogenicity of bacteria isolated from gut of Spodoptera litura (Lepidoptera: Noctuidae) and fitness costs of insect associated with consumption of bacteria.

Gut microbes contribute to the health of insects and perturbations in the composition or location of gut microbiota can lead to pathological states and host mortality. We explored the culturable bacterial community in the gut of Spodoptera litura (Fab.) larvae, which is a polyphagous pest. Bacterial isolates were identified as Microbacterium arborescens (SL6), Enterococcus casseliflavus (SL10) and Enterobacter cloacae (SL11) by using culture dependent technique based on 16S rRNA gene sequencing. Screening of these three isolates for insecticidal potential against the same host i.e. S. litura indicated the highest larval mortality in E. cloacae (73.33%). Further, we assessed the effect of E. cloacae (SL11) infection on growth and development of S. litura. A significant effect of E. cloacae was observed on various biological parameters viz. larval and pupal period, total development period and reproductive potential of S. litura. E. cloacae significantly influenced the immune response of S. litura. A marked decrease in total hemocyte count was observed in larvae infected with E. cloacae whereas lysozyme and phenoloxidase activity increased initially followed by a decline. The gut microbial diversity in larvae infected with E. cloacae differed from control larvae. The population of E. cloacae in the gut of infected larvae exceeded over the other two microbes and resulted in pathogenicity and death of S. litura larvae. This indicates that E. cloacae can have the potential to be used as a promising biological control agent.

Insecticidal and genotoxic potential of two semi-synthetic derivatives of dillapiole for the control of Aedes (Stegomyia) aegypti (Diptera: Culicidae).

The effects of two semi-synthetic dillapiole derivatives, ethyl-ether dillapiole and n-butyl ether dillapiole, on eggs and larvae of Aedes aegypti were studied in view of the need for expansion and renovation of strategic action to control this mosquito - the vector of Dengue virus -, which currently shows a high resistance to chemical insecticides. Eggs and third-instar larvae of A. aegypti that had been exposed to different concentrations of these two compounds showed toxicity and susceptibility, with 100% mortality. Classical cytogenetic assays showed genotoxicity caused by the two compounds in A. aegypti from the cumulative effect of nuclear abnormalities, indicating that these derivatives may be potential alternatives to control A. aegypti.