The Bacillus thuringiensis δ-endotoxins (Bt toxins) are widely used insecticidal proteins in engineered crops that provide agricultural, economic, and environmental benefits. The development of insect resistance to Bt toxins endangers their long-term effectiveness. Here we have developed a phage-assisted continuous evolution selection that rapidly evolves high-affinity protein-protein interactions, and applied this system to evolve variants of the Bt toxin Cry1Ac that bind a cadherin-like receptor from the insect pest Trichoplusia ni (TnCAD) that is not natively bound by wild-type Cry1Ac. The resulting evolved Cry1Ac variants bind TnCAD with high affinity (dissociation constant Kd = 11-41 nM), kill TnCAD-expressing insect cells that are not susceptible to wild-type Cry1Ac, and kill Cry1Ac-resistant T. ni insects up to 335-fold more potently than wild-type Cry1Ac. Our findings establish that the evolution of Bt toxins with novel insect cell receptor affinity can overcome insect Bt toxin resistance and confer lethality approaching that of the wild-type Bt toxin against non-resistant insects.

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Continuous evolution of Bacillus thuringiensis toxins overcomes insect resistance

The sap sucking insects (Hemiptera), which include aphids, whiteflies, plant bugs and stink bugs, have emerged as major agricultural pests. The Hemiptera cause direct damage by feeding on crops, and in some cases indirect damage by transmission of plant viruses. Current management relies almost exclusively on application of classical chemical insecticides. While the development of transgenic crops expressing toxins derived from the bacterium Bacillus thuringiensis (Bt) has provided effective plant protection against some insect pests, Bt toxins exhibit little toxicity against sap sucking insects. Indeed, the pest status of some Hemiptera on Bt-transgenic plants has increased in the absence of pesticide application. The increased pest status of numerous hemipteran species, combined with increased prevalence of resistance to chemical insecticides, provides impetus for the development of biologically based, alternative management strategies. Here, we provide an overview of approaches toward transgenic resistance to hemipteran pests.

https://www.mdpi.com/2072-6651/4/6/405/pdf

Toxins for transgenic resistance to hemipteran pests.

Genetically engineered crops help support conservation biological control

Cross-order and cross-phylum activity of Bacillus thuringiensis pesticidal proteins.

Although transgenic crops expressing Bacillus thuringiensis (Bt) toxins have been used successfully for management of lepidopteran and coleopteran pest species, the sap-sucking insects (Hemiptera) are not particularly susceptible to Bt toxins. To overcome this limitation, we demonstrate that addition of a short peptide sequence selected for binding to the gut of the targeted pest species serves to increase toxicity against said pest. Insertion of a 12-aa pea aphid gut-binding peptide by adding to or replacing amino acids in one of three loops of the Bt cytolytic toxin, Cyt2Aa, resulted in enhanced binding and toxicity against both the pea aphid, Acyrthosiphon pisum, and the green peach aphid, Myzus persicae. This strategy may allow for transgenic plant-mediated suppression of other hemipteran pests, which include some of the most important pests of global agriculture.

https://www.pnas.org/content/pnas/110/21/8465.full.pdf

Retargeting of the Bacillus thuringiensis toxin Cyt2Aa against hemipteran insect pests

The plant bugs Lygus hesperus Knight (Hemiptera: Miridae) and L. lineolaris (Palisot de Beauvois) have emerged as economic pests of cotton in the United States. These hemipteran species are refractory to the insect control traits found in genetically modified commercial varieties of cotton. In this article, we report the isolation and characterization of a 35 kDa crystal protein from Bacillus thuringiensis, designated TIC807, which causes reduced mass gain and mortality of L. hesperus and L. lineolaris nymphs when presented in an artificial diet feeding assay. Cotton plants expressing the TIC807 protein were observed to impact the survival and development of L. hesperus nymphs in a concentration-dependent manner. These results, demonstrating in planta activity of a Lygus insecticidal protein, represent an important milestone in the development of cotton varieties protected from Lygus feeding damage.

https://bioone.org/journals/Journal-of-Economic-Entomology/volume-105/issue-2/EC11207/Cotton-Plants-Expressing-a-Hemipteran-Active-Bacillus-thuringiensis-Crystal-Protein/10.1603/EC11207.pdf

Cotton plants expressing a hemipteran-active Bacillus thuringiensis crystal protein impact the development and survival of Lygus hesperus (Hemiptera: Miridae) nymphs.

A novel Bacillus thuringiensis crystal protein exhibiting insect inhibitory activity is disclosed. Growth of Lygus insects is significantly inhibited by providing the novel crystal protein in Lygus insect diet. Polynucleotides encoding the crystal protein, transgenic plants and microorganisms that contain the polynucleotides, isolated peptides derived from the crystal protein, and antibodies directed against the crystal protein are also provided. Methods of using the crystal protein and polynucleotides encoding the crystal protein to control Hemipteran insects are also disclosed.

Hemipteran and coleopteran active toxin proteins from Bacillus thuringiensis

The present invention discloses a genus of insect inhibitory proteins that exhibit properties directed to controlling Lepidopteran and/or Hemipteran crop pests, methods of using such proteins, nucleotide sequences encoding such proteins, methods of detecting and isolating such proteins, and their use in agricultural systems.

Insect inhibitory toxin family active against hemipteran and/or lepidopteran insects

The invention provides compositions comprising polynucleotide molecules encoding certain pesticidal polypeptides which exhibit plant parasitic nematode and/or insect control properties, and are particularly directed to controlling plant parasitic pest species of nematodes and insects known to infest crop plant species. Methods for controlling pests are disclosed in which the toxic proteins are provided in the diet of the targeted plant pests. The invention also provides compositions such as nucleic acids, proteins, and plant and bacterial cells, plants, and seeds containing the nucleic acid and protein compositions, as well as methods and kits for identifying, detecting, and isolating the compositions of the present invention. The invention further provides a method of producing crops from recombinant seeds which contain the polynucleotide molecules encoding the pesticidal polypeptides of the present invention.

Pesticidal nucleic acids and proteins and uses thereof

La presente invention concerne une nouvelle proteine cristalline de Bacillus thuringiensis presentant une activite inhibitrice contre les insectes. Le developpement des insectes Lygus est significativement inhibe par l’incorporation de la nouvelle proteine cristalline dans l’alimentation de l’insecte Lygus . L’invention concerne egalement des polynucleotides codant pour la proteine cristalline, des plantes et des microorganismes transgeniques qui contiennent les polynucleotides, des peptides isoles derives de la proteine cristalline, et des anticorps diriges contre la proteine cristalline. L’invention concerne egalement des procedes d’utilisation de la proteine cristalline et des polynucleotides codant pour la proteine cristalline afin de lutter contre les insectes hemipteres.

Uma Rao Sukuru

Papers

Cotton plants expressing a hemipteran-active Bacillus thuringiensis crystal protein impact the development and survival of Lygus hesperus (Hemiptera: Miridae) nymphs.

Hemipteran and coleopteran active toxin proteins from Bacillus thuringiensis

Insect inhibitory toxin family active against hemipteran and/or lepidopteran insects

Pesticidal nucleic acids and proteins and uses thereof

Nouvelles protéines toxiques actives contre les hémiptères et les coléoptères issues de bacillus thuringiensis