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WO2013052536A4 - Exploiting host molecules to augment the virulence of mycoinsecticides - Google Patents

Exploiting host molecules to augment the virulence of mycoinsecticides Download PDF

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Publication number
WO2013052536A4
WO2013052536A4 PCT/US2012/058543 US2012058543W WO2013052536A4 WO 2013052536 A4 WO2013052536 A4 WO 2013052536A4 US 2012058543 W US2012058543 W US 2012058543W WO 2013052536 A4 WO2013052536 A4 WO 2013052536A4
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WO
WIPO (PCT)
Prior art keywords
entomopathogenic fungus
genetically modified
fungus
insect
modified strain
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/US2012/058543
Other languages
French (fr)
Other versions
WO2013052536A2 (en
WO2013052536A3 (en
Inventor
Nematollah KEYHANI
Yanhua FAN
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
University of Florida
University of Florida Research Foundation Inc
Original Assignee
University of Florida
University of Florida Research Foundation Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by University of Florida, University of Florida Research Foundation Inc filed Critical University of Florida
Publication of WO2013052536A2 publication Critical patent/WO2013052536A2/en
Publication of WO2013052536A3 publication Critical patent/WO2013052536A3/en
Publication of WO2013052536A4 publication Critical patent/WO2013052536A4/en
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

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Classifications

    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09Recombinant DNA-technology
    • C12N15/63Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
    • C12N15/79Vectors or expression systems specially adapted for eukaryotic hosts
    • C12N15/80Vectors or expression systems specially adapted for eukaryotic hosts for fungi
    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01NPRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N63/00Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
    • A01N63/30Microbial fungi; Substances produced thereby or obtained therefrom
    • AHUMAN NECESSITIES
    • A01AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01NPRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N63/00Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
    • A01N63/50Isolated enzymes; Isolated proteins
    • CCHEMISTRY; METALLURGY
    • C07ORGANIC CHEMISTRY
    • C07KPEPTIDES
    • C07K14/00Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/43504Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from invertebrates
    • C07K14/43563Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from invertebrates from insects

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  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Zoology (AREA)
  • Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Wood Science & Technology (AREA)
  • Biotechnology (AREA)
  • Microbiology (AREA)
  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Plant Pathology (AREA)
  • Dentistry (AREA)
  • Biomedical Technology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • General Engineering & Computer Science (AREA)
  • Environmental Sciences (AREA)
  • Mycology (AREA)
  • Virology (AREA)
  • Agronomy & Crop Science (AREA)
  • Pest Control & Pesticides (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Physics & Mathematics (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Toxicology (AREA)
  • Tropical Medicine & Parasitology (AREA)
  • Insects & Arthropods (AREA)
  • Medicinal Chemistry (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Peptides Or Proteins (AREA)

Abstract

The present disclosure provides methods of insect control in which fungal expression of insect-derived molecules is exploited for target-specific augmentation of entomopathogenic fungi virulence. The major advantages that depends upon the host molecule (peptide) chosen is the increase in virulence that can be tailored to be host specific, and the development of resistance that is be minimized since the host peptides and/or hormones regulate developmental processes that are species- and tissue-specific. Any adaptive mutations that arise in the targeted insect that could compensate for the fungal-expressed product during infection will lead to developmental defects or a fitness cost far greater than developing resistance to a pesticide. The methods, fungal strains, and nucleic acid constructs of the disclosure have been shown to be effective against a variety of insect host species.

Claims

AMENDED CLAIMS received by the International Bureau on 16 April 2013 (16.04.2013) Therefore, the following is claimed:
1. A genetically modified strain of an entomopathogenic fungus comprising a heterologous recombinant nucleotide sequence encoding a peptide, polypeptide, or protein of a target insect host wherein, when said nucleotide sequence is expressed by the genetically modified entomopathogenic fungus when said fungus has infected the target insect host, the peptide, polypeptide, or protein increases the virulence of the fungus in the target insect compared to the virulence of a non-genetically modified strain of the entomopathogenic fungus in the target insect host.
2. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the heterologous recombinant nucleotide sequence is operably linked to a gene expression controlling region, wherein the gene expression controlling region directs production of a transcript from the heterologous recombinant nucleotide sequence in a recipient entomopathogenic fungus.
3. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the entomopathogenic fungus is of the class Hyphomycetes.
4. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the entomopathogenic fungus is virulent against the fire ant {Solenopsis invicta), a mosquito species, a Lepidopteran species, a Dipteran, or a Hemipteran species.
5. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the entomopathogenic fungus is virulent against the fire ant (Solenopsis invicta).
6. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the entomopathogenic fungus is virulent against a mosquito species.
7. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the entomopathogenic fungus is a strain of Metarhizium anisopliae or Beauveria bassiana.
8. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the heterologous recombinant nucleotide sequence encodes an amino acid sequence specifically inducing a biochemical or physiological reaction in a target insect, wherein the amino acid sequence is an insect neurohormone, an insect diuretic hormone, or a trypsin modulating oostatic factor.
9. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the heterologous recombinant nucleotide sequence encodes an insect neurohormone, or a bioactive fragment thereof.
10. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the insect neurohormone is a PBAN/pyrokinin or a bioactive fragment thereof.
11. The genetically modified strain of an entomopathogenic fungus of claim 9, wherein the heterologous recombinant nucleotide sequence encodes an insect neurohormone, or a bioactive fragment thereof, specifically inducing a biochemical or physiological reaction in the fire ant Solenopsis invicta.
12. The genetically modified strain of an entomopathogenic fungus of claim 11 , wherein the insect neurohormone is β-neuropeptide.
13. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the heterologous recombinant nucleotide sequence encodes an insect diuretic hormone.
14. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the heterologous recombinant nucleotide sequence encodes Manduca sexta diuretic hormone (MSDH).
15. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the heterologous recombinant nucleotide sequence encodes a trypsin modulating oostatic factor.
16. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the trypsin modulating oostatic factor is a trypsin modulating oostatic factor of Aedes aegyti or S. bullata.
17. The genetically modified strain of an entomopathogenic fungus of claim 1 , wherein the heterologous recombinant nucleotide sequence encodes a polypeptide having about 90% sequence similarity with a sequence selected from the group consisting of: SEQ ID Nos. 1 , 22, 23, and 24.
18. The genetically modified strain of an entomopathogenic fungus of claim 17, wherein the heterologous recombinant nucleotide sequence encodes a polypeptide having about 95% sequence similarity with a sequence selected from the group consisting of: SEQ ID Nos. 1 , 22, 23, and 24.
19. The genetically modified strain of an entomopathogenic fungus of claim 16, wherein the heterologous recombinant nucleotide sequence encodes a polypeptide having an amino acid sequence selected from the group consisting of: SEQ ID Nos. 1 , 22, 23, and 24.
20. The genetically modified strain of an entomopathogenic fungus of claim 2, wherein the gene controlling region has about 90% sequence similarity with a sequence encoding a β. bassiana chitinase gene signal peptide.
21. The genetically modified strain of an entomopathogenic fungus of claim 20, wherein the B. bassiana chitinase gene signal peptide has the amino acid sequence according to SEQ ID No.: 2.
22. An expression vector comprising a fungal gene expression controlling region nucleotide sequence operably linked to a nucleic acid encoding an insect-specific polypeptide wherein the gene expression controlling region directs production of a transcript, wherein when said transcript is expressed by a entomopathogenic fungus when said fungus has infected a target insect host, the insect-specific polypeptide increases the virulence of the fungus in the target insect compared to the virulence of a strain of the entomopathogenic fungus not expressing the insect specific polypeptide.
23. The expression vector of claim 22, wherein the gene controlling region has about 90% sequence similarity with a sequence encoding a B. bassiana chitinase gene signal peptide.
24. The expression vector of claim 23, wherein the B. bassiana chitinase gene signal peptide has the amino acid sequence according to SEQ ID No.: 2.
25. The expression vector of claim 22, wherein the insect-specific polypeptide is selected from the group consisting of: an insect neurohormone, an insect diuretic hormone, and a trypsin modulating oostatic factor, or a bioactive homolog or fragment thereof.
26. The expression vector of claim 25, wherein the insect-specific polypeptide is selected from the group consisting of: β-neuropeptide specific to the fire ant Solenopsis invicta, Manduca sexta diuretic hormone (MSDH), and a trypsin modulating oostatic factor of Aedes aegyti or S. bullata.
27. A method of increasing the virulence of an entomopathogenic fungus in a target insect host, comprising the steps of:
(a) obtaining a genetically modified strain of a entomopathogenic fungus according to claims 1 ; and
(b) delivering the genetically modified strain of entomopathogenic fungus, or spores thereof, to a target insect host desired to be infected by the fungus, wherein the fungus delivers a target insect-specific polypeptide or peptide to the target insect host, thereby increasing the virulence of the entomopathogenic fungus in the target insect host.
28. The method of claim 27, wherein the wherein the entomopathogenic fungus is Metarhizium anisopliae or Beauveria bassiana.
29. The method of claim 27, wherein the target insect host is the fire ant (Solenopsis invicta), a mosquito species, a Lepidopteran species, a Dipteran, or a Hemipteran species.
PCT/US2012/058543 2011-10-05 2012-10-03 Exploiting host molecules to augment the virulence of mycoinsecticides Ceased WO2013052536A2 (en)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US201161543345P 2011-10-05 2011-10-05
US61/543,345 2011-10-05

Publications (3)

Publication Number Publication Date
WO2013052536A2 WO2013052536A2 (en) 2013-04-11
WO2013052536A3 WO2013052536A3 (en) 2013-06-06
WO2013052536A4 true WO2013052536A4 (en) 2013-07-25

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Families Citing this family (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2018140496A1 (en) * 2017-01-24 2018-08-02 Flagship Pioneering, Inc. Methods and related compositions for manufacturing food and feed
CN114480300B (en) * 2021-06-28 2024-12-06 吉林省农业科学院 A fungal virus for improving the toxicity of Beauveria bassiana and a method for transmitting the virus
CN115991759A (en) * 2022-10-20 2023-04-21 曲靖师范学院 Paraphalocrocis medinalis sex pheromone binding protein gene GmelPBPs and application thereof
CN115505538B (en) * 2022-11-17 2023-08-04 中国农业科学院植物保护研究所 Metarhizium anisopliae strain CIPPMA0941, application thereof in preventing and treating solenopsis invicta and microbial inoculum
ES2978576B2 (en) * 2023-02-09 2025-03-06 Instituto Valenciano De Investig Agrarias Ivia Machine for inoculating entomopathogenic fungal conidia in powder form onto sterile males of Ceratitis capitata

Family Cites Families (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5683689A (en) * 1991-04-18 1997-11-04 University Of Florida Research Foundation, Inc. Controlling cockroaches, carpenter ants, and pharaoh ants using strains of beauveria bassiana
US5516513A (en) * 1994-07-14 1996-05-14 Troy Biosciences, Inc. Biological ovicide for control of lepidopterous insects
WO1999013727A1 (en) * 1997-09-17 1999-03-25 University Of Florida Methods and materials for control of termites

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WO2013052536A3 (en) 2013-06-06

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