Compositions and methods for controlling insect pests

• US 10,378,012 B2
• Filed: 07/28/2015
• Issued: 08/13/2019
• Est. Priority Date: 07/29/2014
• Status: Active Grant

• Current Assignee: Monsanto Technology LLC (Bayer AG)
• Original Assignee: Monsanto Technology LLC (Bayer AG)
• Inventors: Crawford, Michael John, Eads, Brian Donovan
• Primary Examiner(s): Zheng, Li
• Application Number:
  US15/329,808
• Publication Number:
  US 20170260522A1
• Time in Patent Office: 1,477 Days
• Field of Search: 800279
• CPC Class Codes: A01N 57/16 containing heterocyclic radicals
• CPC Class Codes: A01N 63/60 Isolated nucleic acids
• CPC Class Codes: A01N 65/00 Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof containing compounds of determined constitution A01N27/00 - A01N59/00
• CPC Class Codes: C12N 15/111 General methods applicable to biologically active non-coding nucleic acids
• CPC Class Codes: C12N 15/113 Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing when used in plants C12N15/8218
• CPC Class Codes: C12N 15/8286 for insect resistance
• CPC Class Codes: C12N 2310/14 interfering N.A.
• CPC Class Codes: Y02A 40/146 Genetically Modified [GMO] plants, e.g. transgenic plants

Abstract

Disclosed herein are polynucleotides, compositions, and methods for controlling insect pests, especially flea beetles, such as Phyllotreta spp. and Psylliodes spp., particularly in plants. More specifically, polynucleotides such as double-stranded RNA triggers and methods of use thereof for modifying the expression of genes in flea beetles.

Claims

1. A method of causing mortality or stunting in an insect, comprising contacting an insect with at least one recombinant double-stranded RNA (dsRNA) comprising at least one silencing element comprising an RNA strand identical or complementary to 21 or more contiguous nucleotides of an insect target gene sequence that has a DNA sequence of SEQ ID NO:

327, and the complement thereof, wherein said insect is Phylotreta spp., and wherein ingestion of said recombinant dsRNA by said insect results in mortality or stunting in said insect.

2. The method of claim 1, wherein said at least one silencing element comprises an RNA strand having a sequence identical or complementary to 21 or more contiguous nucleotides of SEQ ID NO:

1186.

3. The method of claim 1, wherein said insect is Phyllotreta cruciferae (canola flea beetle).

4. The method of claim 1, wherein said silencing element comprises an RNA strand having a sequence of SEQ ID NO:

1186.

5. The method of claim 1, wherein said recombinant dsRNA is provided in a microbial or plant cell that expresses said recombinant dsRNA, or in a microbial fermentation product, or is chemically synthesized.

6. The method of claim 1, wherein said at least one recombinant dsRNA is provided in a composition comprising said at least one recombinant dsRNA, wherein said composition is applied to a surface of said insect or to a surface of a seed or a plant in need of protection from infestation by said insect.

7. The method of claim 6, wherein said plant is:

(a) selected from an ornamental plant or a crop plant;

(b) a plant in the family Brassicaceae;

(c) a Brassica species selected from the group consisting of B. napus, B, juncea, B. carinata, B. rapa, B. oleracea, B. rupestris, B. septiceps, B. nigra, B. narinosa, B. perviridus, B. tournefortii, and B. fructiculosa;

(d) a Brassica plant selected from the group consisting canola, rapeseed, turnips, and field mustard or turnip rape;

(e) selected from the group consisting of Glycine max, Linum usitatissimum, Zea mays, Carthamus tinctorius, Helianthus annuus, Nicotiana tabacum, Arabidopsis thaliana, Betholettia excelsa, Ricinus communis, Cocos nucifera, Coriandrum sativum, Gossypium spp., Arachis hypogaea, Simmondsia chinensis, Solanum tuberosum, Elaeis guineensis, Olea europaea, Oryza sativa, Cucurbita maxim, Hordeum vulgare, and Triticum aestivum;

or

(f) a potato plant and said insect is Epitrix cucumeris (potato flea beetle).

8. The method of claim 6, wherein said contacting comprises:

(a) oral delivery to said insect, non-oral delivery to said insect, or a combination of oral and non-oral delivery to said insect;

(b) application of a composition comprising said recombinant dsRNA to a surface of said insect or to a surface of said plant infested by said insect;

(c) providing said recombinant dsRNA in a composition that further comprises one or more components selected from the group consisting of a carrier agent, a surfactant, an organosilicone, a polynucleotide herbicidal molecule, a non-polynucleotide herbicidal molecule, a non-polynucleotide pesticide, a fungicide, a safener, a fertilizer, a micronutrient, an insect attractant, and an insect growth regulator;

(d) providing said recombinant dsRNA in a composition that further comprises at least one pesticidal agent selected from the group consisting of a patatin, a plant lectin, a phytoecdysteroid, a Bacillus thuringiensis insecticidal protein, a Xenorhabdus insecticidal protein, a Photorhabdus insecticidal protein, a Bacillus laterosporous insecticidal protein, a Bacillus sphaericus insecticidal protein, a bacterium that produces an insecticidal protein, an entomicidal bacterial species, Lysinibacillus sphaericus (Bacillus sphaericus), Brevibacillus laterosporus (Bacillus laterosporus), Chromobacterium species, Chromobacterium subtsugae, Paenibacillus species, Paenibacillus lentimorbus, and Paenibacillus popilliae;

or

(e) providing said recombinant dsRNA in a composition that is ingested by said insect.

9. An insecticidal composition comprising an insecticidally effective amount of a recombinant dsRNA molecule, wherein said recombinant dsRNA molecule comprises an RNA strand complementary to 21 or more contiguous nucleotides of a target gene of an insect that infests a plant, and the complement thereof, wherein said insect is Phylotreta spp., and wherein said target gene has a DNA sequence of SEQ ID NO:

327.

10. The insecticidal composition of claim 9, wherein said recombinant dsRNA molecule:

(a) comprises at least one RNA strand having a sequence identical or complementary to 21 or more contiguous nucleotides of SEQ ID NO;

1186;

(b) comprises dsRNA of at least 50 base pairs in length;

(c) comprises blunt-ended dsRNA;

(d) comprises dsRNA with an overhang at at least one terminus;

(e) comprises dsRNA comprising at least one stem-loop;

(f) comprises dsRNA that is chemically modified;

(g) is a dsRNA comprising an RNA strand having a sequence identical or complementary to 21 or more contiguous nucleotides of SEQ ID NO;

1186;

(h) is a dsRNA comprising an RNA strand having a sequence of SEQ ID NO;

1186;

or

(i) is provided in a microbial or plant cell that expresses said recombinant dsRNA, or in a microbial fermentation product;

or is chemically synthesized.

11. The insecticidal composition of claim 9, wherein said insecticidal composition:

(a) further comprises one or more components selected from the group consisting of a carrier agent, a surfactant, an organosilicone, a polynucleotide herbicidal molecule, a non-polynucleotide herbicidal molecule, a non-polynucleotide pesticide, a fungicide, a safener, a fertilizer, a micronutrient, an insect attractant, and an insect growth regulator;

(b) further comprises at least one pesticidal agent selected from the group consisting of a patatin, a plant lectin, a phytoecdysteroid, a Bacillus thuringiensis insecticidal protein, a Xenorhabdus insecticidal protein, a Photorhabdus insecticidal protein, a Bacillus laterosporous insecticidal protein, a Bacillus sphaericus insecticidal protein, a bacterium that produces an insecticidal protein, an entomicidal bacterial species, Lysinibacillus sphaericus (Bacillus sphaericus), Brevibacillus laterosporus (Bacillus laterosporus), Chromobacterium species, Chromobacterium subtsugae, Paenibacillus species, Paenibacillus lentimorbus, and Paenibacillus popilliae;

or

(c) is in a form selected from the group consisting of a solid, liquid, powder, suspension, emulsion, spray, encapsulation, microbeads, carrier particulates, film, matrix, soil drench, insect diet or insect bait, and seed treatment.

12. A plant treated with the insecticidal composition of claim 9, or a plant grown from seed treated with the insecticidal composition of claim 9, wherein said plant exhibits improved resistance to said insect.

13. A recombinant DNA construct comprising a heterologous promoter operably linked to DNA encoding a dsRNA comprising a sequence identical or complementary to 21 or more contiguous nucleotides of SEQ ID NO:

1186, and the complement thereof.

14. A recombinant vector, plant chromosome or plastid, or transgenic plant cell comprising the recombinant DNA construct of claim 13.

15. A method of providing a plant having improved resistance to a Phylotrera spp. insect, comprising expressing in said plant the recombinant DNA construct of claim 13.

16. A plant having improved resistance to a Phylotrera spp. insect, provided by the method of claim 15, or fruit, seed, or propagatable parts of said plant.