Insecticidal activities of three recombinant venom proteins of the predatory stink bug, Arma custos.

BACKGROUND: Widespread resistance of insect pests to insecticides and transgenic crops in the field is a significant challenge for sustainable agriculture, and calls for the development of novel alternative strategies to control insect pests. One potential resource for the discovery of novel insecticidal molecules is natural toxins, particularly those derived from the venoms of insect predators. RESULTS: In this study, we identified three insecticidal proteinaceous toxins from the venom glands (VGs) of the predatory stink bug, Arma custos (Hemiptera: Asopinae). Transcriptomic analysis of A. custos VGs revealed 151 potentially secreted VG-rich venom proteins. Three VG-rich venom proteins (designated AcVP1 ~ 3) were produced by overexpression in Escherichia coli. Injection of the recombinant proteins into tobacco cutworm (Spodoptera litura) larvae showed that all of the three recombinant proteins caused paralysis, liquefaction and death. Injection of recombinant proteins into rice brown planthopper (Nilaparvata lugens) nymphs showed higher insecticidal activities, among which a trypsin (AcVP2) caused 100% mortality postinjection at 1.27 pmol mg-1 body weight. CONCLUSION: A natural toolkit for the discovery of insecticidal toxins from predatory insects has been revealed by the present study. © 2024 Society of Chemical Industry.

Combined effects of four pesticides and heavy metal chromium (â ¥) on the earthworm using avoidance behavior as an endpoint.

In natural ecosystems, organisms are commonly exposed to chemical mixtures rather than individual compounds. However, environmental risk is traditionally assessed based on data of individual compounds. In the present study, we aimed to investigate the individual and combined effects of four pesticides [fenobucarb (FEN), chlorpyrifos (CPF), clothianidin (CLO), acetochlor (ACE)] and one heavy metal chromium [Cr(Ⅵ)] on the earthworm (Eisenia fetida) using avoidance behavior as an endpoint. Our results indicated that CLO had the highest toxicity to E. fetida, followed by Cr(Ⅵ), while FEN showed the least toxicity. Two mixtures of CPF+CLO and Cr(Ⅵ)+CPF+CLO+ACE exhibited synergistic effects on the earthworms. The other two quaternary mixtures of CLO+FEN+ACE+Cr(Ⅵ) and Cr(Ⅵ)+FEN+CPF+ACE at low concentrations also displayed synergistic effects on the earthworms. In contrast, the mixture of Cr(Ⅵ)+FEN had the strongest antagonistic effects on E. fetida. Besides, the quinquenary mixture of Cr(Ⅵ)+FEN+CPF+CLO+ACE also exerted antagonistic effects. These findings highlighted the importance to evaluate chemical mixtures. Moreover, our data strongly pointed out that the avoidance tests could be used to assess the effects of combined effects.

Interaction of proteinase inhibitors with Cry1Ac toxicity and the presence of 15 chymotrypsin cDNAs in the midgut of the tobacco budworm, Heliothis virescens (F.) (Lepidoptera: Noctuidae).

BACKGROUND: The potential development of resistance to Bacillus thuringiensis (Bt) cotton and surging of non-targeted insects is a major risk in the durability of Bt plant technology. Midgut proteinases are involved in Bt activation and degradation. Proteinase inhibitors may be used to control a wide range of insects and delay Bt resistance development. Proactive action to examine proteinase inhibitors for synergistic interaction with Bt toxin and cloning of proteinase cDNAs for RNAi is necessary to make transgenic cotton more versatile and durable. RESULTS: A sublethal dose (15 ppb) of Cry1Ac, 0.5% benzamidine and 0.02% phenylmethylsulfonyl fluoride significantly suppressed midgut azocaseinase, tryptic and chymotryptic activities, and resulted in reductions in larval and pupal length and mass of Heliothis virescens. The combination of proteinase inhibitor and Bt suppressed 20-37% more larval body mass and 26-80% more enzymatic activities than the inhibitor only or Bt only. To facilitate knockdown-resistance-related proteinase genes, 15 midgut chymotrypsin cDNAs were sequenced. Most predicted chymotrypsins contained the conserved N-termini IVGG, three catalytic center residues (His, Asp and Ser), substrate specificity determinant (Ser or Gly) and cysteines for disulfide bridges. These putative chymotrypsins were separated into three distinct groups, indicating the diverse proteinases evolved in this polyphagous insect. CONCLUSION: H. virescens has evolved diverse midgut proteinase genes. Proteinase inhibitors have potential insecticidal activity, and the interaction of Bt with proteinase inhibitors is desirable for enhancing Bt toxicity and delaying resistance development. Intensive sequencing of chymotrypsin cDNAs will facilitate future functional examinations of individual roles in Bt toxicity and resistance development and facilitate targeted control using RNAi and/or proteinase inhibitors.