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The cotton bollworm Helicoverpa armigera , is one of the world’s major pest of agriculture, feeding on over 300 hosts in 68 plant families. Resistance cases to most insecticide classes have been reported for this insect. Management of this pest in agroecosystems relies on a better understanding of how it copes with phytochemical or synthetic toxins. We have used genome editing to knock out a cluster of nine P450 genes and show that this significantly reduces the survival rate of the insect when exposed to two classes of host plant chemicals and two classes of insecticides. Functional expression of all members of this gene cluster identified the P450 enzymes capable of metabolism of these xenobiotics. The CRISPR-Cas9-based reverse genetics approach in conjunction with in vitro metabolism can rapidly identify the contributions of insect P450s in xenobiotic detoxification and serve to identify candidate genes for insecticide resistance. Cotton bollworm is an important agricultural pest with widespread resistance to insecticides. Here Wang et al . identifies CYP6AEs from cotton bollworm involved in detoxifying plant toxins and chemical insecticides through the CRISPR-Cas9-based reverse genetics approach in conjunction with in vitro metabolism.

Frequent applications of synthetic insecticides might cause environmental pollution due to the high residue. In addition, increasing insecticide resistance in many insect pests requires novel pest control methods. Nanotechnology could be a promising field of modern agriculture, and is receiving considerable attention in the development of novel nano-agrochemicals, such as nanoinsectticides and nanofertilizers. This study assessed the effects of the lethal and sublethal concentrations of chlorantraniliprole, thiocyclam, and their nano-forms on the development, reproductive activity, oxidative stress enzyme activity, and DNA changes in the black cutworm, Agrotis ipsilon , at the molecular level. The results revealed that A . ipsilon larvae were more susceptible to the nano-forms than the regular forms of both nano chlorine and sulfur within the chlorantraniliprole and thiocyclam insecticides, respectively, with higher toxicities than the regular forms (ca. 3.86, and ca.2.06-fold, respectively). Significant differences in biological parameters, including developmental time and reproductive activity (fecundity and hatchability percent) were also observed. Correspondingly, increases in oxidative stress enzyme activities were observed, as were mutagenic effects on the genomic DNA of A . ipsilon after application of the LC 50 of the nano-forms of both insecticides compared to the control. These promising results could represent a crucial step toward developing efficient nanoinsecticides for sustainable control of A . ipsilon .

Melanin, a black-brown pigment found throughout all kingdoms of life, has diverse biological functions including UV protection, thermoregulation, oxidant scavenging, arthropod immunity, and microbial virulence. Given melanin’s broad roles in the biosphere, particularly in insect immune defenses, it is important to understand how exposure to ubiquitous environmental contaminants affects melanization. Glyphosate—the most widely used herbicide globally—inhibits melanin production, which could have wide-ranging implications in the health of many organisms, including insects. Here, we demonstrate that glyphosate has deleterious effects on insect health in 2 evolutionary distant species, Galleria mellonella (Lepidoptera: Pyralidae) and Anopheles gambiae (Diptera: Culicidae), suggesting a broad effect in insects. Glyphosate reduced survival of G . mellonella caterpillars following infection with the fungus Cryptococcus neoformans and decreased the size of melanized nodules formed in hemolymph, which normally help eliminate infection. Glyphosate also increased the burden of the malaria-causing parasite Plasmodium falciparum in A . gambiae mosquitoes, altered uninfected mosquito survival, and perturbed the microbial composition of adult mosquito midguts. Our results show that glyphosate’s mechanism of melanin inhibition involves antioxidant synergy and disruption of the reaction oxidation–reduction balance. Overall, these findings suggest that glyphosate’s environmental accumulation could render insects more susceptible to microbial pathogens due to melanin inhibition, immune impairment, and perturbations in microbiota composition, potentially contributing to declines in insect populations.

More than two million people worldwide are affected by life-threatening, invasive fungal infections annually. Candida species are the most common cause of nosocomial, invasive fungal infections and are associated with mortality rates above 40%. Despite the increasing incidence of drug-resistance, the development of novel antifungal formulations has been limited. Here we investigate the antifungal mode of action and therapeutic potential of positively charged, synthetic peptide mimics to combat Candida albicans infections. Our data indicates that these synthetic polymers cause endoplasmic reticulum stress and affect protein glycosylation, a mode of action distinct from currently approved antifungal drugs. The most promising polymer composition damaged the mannan layer of the cell wall, with additional membrane-disrupting activity. The synergistic combination of the polymer with caspofungin prevented infection of human epithelial cells in vitro, improved fungal clearance by human macrophages, and significantly increased host survival in a Galleria mellonella model of systemic candidiasis. Additionally, prolonged exposure of C.   albicans to the synergistic combination of polymer and caspofungin did not lead to the evolution of tolerant strains in vitro. Together, this work highlights the enormous potential of these synthetic peptide mimics to be used as novel antifungal formulations as well as adjunctive antifungal therapy. Fungal infections are severely underestimated as a cause of mortality, and alternative drugs are urgently needed. Here, Schaefer et al. show that a synthetic polymer mimicking defensins shows different, but synergistic activity with known antifungals.

The codling moth Cydia pomonella, a major invasive pest of pome fruit, has spread around the globe in the last half century. We generated a chromosome-level scaffold assembly including the Z chromosome and a portion of the W chromosome. This assembly reveals the duplication of an olfactory receptor gene (OR3), which we demonstrate enhances the ability of C. pomonella to exploit kairomones and pheromones in locating both host plants and mates. Genome-wide association studies contrasting insecticide-resistant and susceptible strains identify hundreds of single nucleotide polymorphisms (SNPs) potentially associated with insecticide resistance, including three SNPs found in the promoter of CYP6B2. RNAi knockdown of CYP6B2 increases C. pomonella sensitivity to two insecticides, deltamethrin and azinphos methyl. The high-quality genome assembly of C. pomonella informs the genetic basis of its invasiveness, suggesting the codling moth has distinctive capabilities and adaptive potential that may explain its worldwide expansion.

Insecticidal crystal toxins derived from the soil bacterium Bacillus thuringiensis (Bt) are widely used as biopesticide sprays or expressed in transgenic crops to control insect pests. However, large-scale use of Bt has led to field-evolved resistance in several lepidopteran pests. Resistance to Bt Cry1Ac toxin in the diamondback moth, Plutella xylostella (L.), was previously mapped to a multigenic resistance locus (BtR-1). Here, we assembled the 3.15 Mb BtR-1 locus and found high-level resistance to Cry1Ac and Bt biopesticide in four independent P. xylostella strains were all associated with differential expression of a midgut membrane-bound alkaline phosphatase (ALP) outside this locus and a suite of ATP-binding cassette transporter subfamily C (ABCC) genes inside this locus. The interplay between these resistance genes is controlled by a previously uncharacterized trans-regulatory mechanism via the mitogen-activated protein kinase (MAPK) signaling pathway. Molecular, biochemical, and functional analyses have established ALP as a functional Cry1Ac receptor. Phenotypic association experiments revealed that the recessive Cry1Ac resistance was tightly linked to down-regulation of ALP, ABCC2 and ABCC3, whereas it was not linked to up-regulation of ABCC1. Silencing of ABCC2 and ABCC3 in susceptible larvae reduced their susceptibility to Cry1Ac but did not affect the expression of ALP, whereas suppression of MAP4K4, a constitutively transcriptionally-activated MAPK upstream gene within the BtR-1 locus, led to a transient recovery of gene expression thereby restoring the susceptibility in resistant larvae. These results highlight a crucial role for ALP and ABCC genes in field-evolved resistance to Cry1Ac and reveal a novel trans-regulatory signaling mechanism responsible for modulating the expression of these pivotal genes in P. xylostella.

Greater wax moth Galleria mellonella is a serious pest in apiculture, invading honeybee colonies and inflicting widespread damage. Ozone gas generated by the non-thermal atmospheric pressure plasma produced with a dielectric barrier discharge (DBD) reactor is an attractive solution for insect control. The effect of ozone gas generated by cold plasma on the egg, larva, and pupae stages of G. mellonella was investigated by using two concentrations, 400 and 800 ppmv, at different exposure times of 5, 10, 20, 40, 60, and 80 min. The mortality rate of the larvae increased as the exposure time lengthened, while a decrease in adult survivorship was observed after exposure. Complete suppression of the survived larval instar percentage and pupation percentage was noted after 7 days of exposure to 40 and 20 min at 400 and 800 ppmv of ozone, respectively, compared to the control group, which had a 96.67% survivorship. Pupal exposure to 800 ppmv for 20 min was sufficient to completely inhibit adult emergence. Larvae exhibited greater tolerance to ozone gas generated by cold plasma than eggs and pupae 24 h after exposure. Additionally, ozone treatment at 400 and 800 ppmv significantly increased total hemolymph protein content after 24 h, reaching 17.55 g/L and 18.50 g/L, respectively, compared to 16.52 g/L for the control. Cold plasma ozone treatment altered the stored beeswax matrix by modifying essential hydrocarbons and enhancing fatty acid diversity, while preserving the fundamental ester structure, thus converting it into a functionally changed biomaterial. Exposure to ozone gas at 800ppmv caused remarkable abnormalities in appearance in wax moths throughout their life stages, resulting in shrunken, twisted, and crippled insects unable to complete normal development.

The diamondback moth (DBM), Plutella xylostella (Lepidoptera: Plutellidae) is a very destructive crucifer-specialized pest that has resulted in significant crop losses worldwide. DBM is well attracted to glucosinolates (which act as fingerprints and essential for herbivores in host plant recognition) containing crucifers such as wintercress, Barbarea vulgaris (Brassicaceae) despite poor larval survival on it due to high-to-low concentration of saponins and generally to other plants in the genus Barbarea. B. vulgaris build up resistance against DBM and other herbivorous insects using glucosinulates which are used in plant defense. Aside glucosinolates, Barbarea genus also contains triterpenoid saponins, which are toxic to insects and act as feeding deterrents for plant specialist herbivores (such as DBM). Previous studies have found interesting relationship between the host plant and secondary metabolite contents, which indicate that attraction or resistance to specialist herbivore DBM, is due to higher concentrations of glucosinolates and saponins in younger leaves in contrast to the older leaves of Barbarea genus. As a response to this phenomenon, herbivores as DBM has developed a strategy of defense against these plant biochemicals. Because there is a lack of full knowledge in understanding bioactive molecules (such as saponins) role in plant defense against plant herbivores. Thus, in this review, we discuss the role of secondary plant metabolites in plant defense mechanisms against the specialist herbivores. In the future, trials by plant breeders could aim at transferring these bioactive molecules against herbivore to cash crops.

Background Infectivity trials and toxicity testing in rodents are important prerequisites to the use of compounds in man. However, trials in rats and mice are expensive and there are ethical considerations. Galleria mellonella (greater wax moth) larvae are a potential alternative. We have assessed the use of these insects in infectivity trials and toxicity testing. Findings Using four bacterial species (two Gram-negative and two Gram-positive) we have assessed the efficacy of four antibiotics against infections in Galleria and compared the antibiotic susceptibility with that in humans. In general, we find a good correlation. Similarly, we have assessed 11 compounds (initially tested blind) for their toxicity in Galleria and compared this with toxicity trials in mice and rats. Again we found a good correlation between toxicity in Galleria and that in rodents. Conclusion We have found, in our hands, that G. mellonella larvae can be used in infectivity trials and toxicity testing, and that these assays represent an inexpensive and readily executable alternative to testing in rodents.

Biopesticides have emerged as a global trend to minimize the risks associated with synthetic agrochemicals. However, their stability and efficacies remain challenges for widespread application. Herein, co-assembled nanoparticles (AT NPs or AP NPs) based on azadirachtin (AZA) and tannic acid (TA) or phenylalanine (PA) are constructed in aqueous solution through self-assembly technology. The small particle size, low PDI, high ζ-potential, and related other physicochemical characteristics of nanoparticles can improve wettability, adhesiveness, rain erosion resistance, and photostability compared to the commercial AZA formulation. Importantly, co-assemblies with bidirectional pH-responsive disassembly in acidic or alkaline solutions, allow them to respond to microenvironmental stimuli of targets and enable controlled release of AZA. The nanosystems demonstrated remarkable in vitro and in vivo insecticidal activities against Ostrinia furnacalis and Aphis gossypii . This study illustrates a distinctive perspective for developing eco-friendly nanosystems, highlighting a water-based treatment method for biopesticides with improved physicochemical properties and utilization efficiency. The stability and delivery of biopesticides limits application. Here, azadirachtin is co-assembled with plant secondary metabolites for efficient drug delivery, demonstrating pest management, the activation of plant defensive capacity, while increasing the production yield of crops.