Fungal endophytes for improved crop yields and protection from pests

• US 9,756,865 B2
• Filed: 07/15/2016
• Issued: 09/12/2017
• Est. Priority Date: 11/06/2013
• Status: Active Grant

• Current Assignee: Texas A&M University System
• Original Assignee: Texas A&M University System
• Inventors: Sword, Gregory A.
• Primary Examiner(s): Pryor, Alton
• Application Number:
  US15/212,038
• Publication Number:
  US 20160316763A1
• Time in Patent Office: 424 Days
• Field of Search: None
• CPC Class Codes: A01N 63/30 Microbial fungi; Substances produced thereby or obtained therefrom

Abstract

The invention provides a synthetic combination of a crop and at least one fungal endophyte, wherein the crop is a host plant of the endophyte. Provided are also methods and compositions for producing such synthetic combinations. The endophyte reproduces and enhances the agronomic characteristics of the crop. Methods for inoculating the host plant with the endophyte, for propagating the host-endophyte combination, and for detecting the presence of the endophyte and of its metabolites within a host plant are also described.

Claims

1. A method for improving drought tolerance in a cotton plant, the method comprising:

contacting a cotton seed with a formulation comprising purified filamentous, spore-forming, facultative fungal endophytes of at least one species, wherein the facultative fungal endophytes are of the sub-class Hypocreomycetidae, and of the Family, Incertae sedis, Nectriaceae, or Plectosphaerellaceae, wherein the facultative fungal endophytes are present in the formulation in an amount effective to improve drought tolerance of a cotton plant grown from the contacted seed compared to a reference cotton plant or a cotton plant grown from a reference cotton seed.

2. The method of claim 1, wherein the formulation contains at least 100 spores/ml or 100,000 spores/g dry weight of the fungal endophytes.

3. The method of claim 1, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to a nucleic acid sequence selected from the group consisting of:

SEQ ID NO;

7, 53, 54, 55, and 73.

4. The method of claim 1, wherein the fungal endophytes are present in the formulation in an amount effective to reduce yield loss of the cotton plant grown from the contacted seed compared to the reference cotton plant or the cotton plant grown from the reference seed.

5. The method of claim 4, wherein yield loss is reduced by at least 5%.

6. The method of claim 4, wherein the reduction in yield loss results in a yield increase of at least 5%.

7. The method of claim 1, wherein improved drought tolerance is assessed by withholding water from 7-day old seedlings of the cotton plant grown from the contacted seed in the greenhouse, wherein the seedlings have increased time to wilt as compared to a reference seedling grown from the reference seed.

8. The method of claim 1, wherein the facultative fungal endophytes are of the sub-class Hypocreomycetidae, and of the Family Incertae sedis.

9. The method of claim 1, wherein the facultative fungal endophytes are of the sub-class Hypocreomycetidae, and of the Family Nectriaceae.

10. The method of claim 1, wherein the facultative fungal endophytes are of the sub-class Hypocreomycetidae, and of the Family Plectosphaerellaceae.

11. The method of claim 1, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

7.

12. The method of claim 1, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

53.

13. The method of claim 1, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

54.

14. The method of claim 1, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

55.

15. The method of claim 1, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

73.

16. The method of claim 1, wherein the fungal endophytes comprise SEQ ID NO:

7.

17. The method of claim 1, wherein the fungal endophytes comprise SEQ ID NO:

53.

18. The method of claim 1, wherein the fungal endophytes comprise SEQ ID NO:

54.

19. The method of claim 1, wherein the fungal endophytes comprise SEQ ID NO:

55.

20. The method of claim 1, wherein the fungal endophytes comprise SEQ ID NO:

73.

29. The method of claim 1, wherein the facultative fungal endophytes are of the sub-class Hypocreomycetidae, and of the Family Plectosphaerellaceae.

21. A method of treating seeds of a cotton plant, comprising contacting a plurality of seeds with a formulation comprising fungal endophytes of the sub-class Hypocreomycetidae, and of the Family, Incertae sedis, Nectriaceae, or Plectosphaerellaceae, wherein the fungal endophytes are present in the formulation in an amount effective to improve drought tolerance of a cotton plant grown from the contacted seed compared to a reference cotton plant or a cotton plant grown from a reference cotton seed.

22. The method of claim 21, wherein the formulation contains at least 100 (10^2) spores/ml or 100,000 (10^5) spores/g dry weight of the fungal endophytes.

23. The method of claim 21, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to a nucleic acid sequence selected from the group consisting of:

SEQ ID NO;

7, 53, 54, 55, and 73.

24. The method of claim 21, wherein the fungal endophytes are present in the formulation in an amount effective to reduce yield loss of the cotton plant grown from the contacted seed compared to the reference cotton plant or the cotton plant grown from the reference seed.

25. The method of claim 21, wherein yield loss is reduced by at least 5%.

26. The method of claim 25, wherein the reduction in yield loss results in a yield increase of at least 5%.

27. The method of claim 21, wherein the facultative fungal endophytes are of the sub-class Hypocreomycetidae, and of the Family Incertae sedis.

28. The method of claim 21, wherein the facultative fungal endophytes are of the sub-class Hypocreomycetidae, and of the Family Nectriaceae.

30. The method of claim 21, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

7.

31. The method of claim 21, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

53.

32. The method of claim 21, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

54.

33. The method of claim 21, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

55.

34. The method of claim 21, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

73.

35. The method of claim 21, wherein the fungal endophytes comprise SEQ ID NO:

7.

36. The method of claim 21, wherein the fungal endophytes comprise SEQ ID NO:

53.

37. The method of claim 21, wherein the fungal endophytes comprise SEQ ID NO:

54.

38. The method of claim 21, wherein the fungal endophytes comprise SEQ ID NO:

55.

39. The method of claim 21, wherein the fungal endophytes comprise SEQ ID NO:

73.

41. The synthetic combination of claim 34, wherein the fungal endophytes are present at a concentration of at least 100(10^2) spores/seed on the surface of the seed.

42. The synthetic combination of claim 34, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to a nucleic acid sequence selected from the group consisting of:

SEQ ID NO;

7, 53, 54, 55, and 73.

40. A synthetic combination of a cotton seed and purified filamentous, spore-forming, facultative fungal endophytes of at least one species, wherein the facultative fungal endophytes are of the sub-class Hypocreomycetidae, and of the Family, Incertae sedis, Nectriaceae, or Plectosphaerellaceae, wherein the facultative fungal endophytes are present in the combination in an amount effective to improve drought tolerance of a cotton plant grown from the synthetic combination compared to a cotton plant grown from a reference cotton seed.

43. The synthetic combination of claim 40, wherein the fungal endophytes are present in the formulation in an amount effective to reduce yield loss in the cotton plant grown from the synthetic combination compared to the cotton plant grown from the reference seed.

44. The synthetic combination of claim 43, wherein the fungal endophytes are present in the formulation in an amount effective to reduce yield loss by at least 5%.

45. The synthetic combination of claim 43, wherein the fungal endophytes are present in the formulation in an amount effective to reduce yield loss resulting in a yield increase of at least 5%.

46. The synthetic combination of claim 40, wherein the effective amount of the fungal endophytes improves drought tolerance of the cotton plant as assessed by withholding water from 7-day old seedlings of the cotton plant grown from the synthetic combination in the greenhouse, wherein seedlings have increased time to wilt as compared to a reference seedling grown from the reference seed.

47. The synthetic combination of claim 40, wherein the facultative fungal endophytes are of the sub-class Hypocreomycetidae, and of the Family Incertae sedis.

48. The synthetic combination of claim 40, wherein the facultative fungal endophytes are of the sub-class Hypocreomycetidae, and of the Family Nectriaceae.

49. The synthetic combination of claim 40, wherein the facultative fungal endophytes are of the sub-class Hypocreomycetidae, and of the Family Plectosphaerellaceae.

50. The synthetic combination of claim 40, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

7.

51. The synthetic combination of claim 40, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

53.

52. The synthetic combination of claim 40, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

54.

53. The synthetic combination of claim 40, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

55.

54. The synthetic combination of claim 40, wherein the fungal endophytes comprise a nucleic acid sequence that is at least 97% identical to SEQ ID NO:

73.

55. The synthetic combination of claim 40, wherein the fungal endophytes comprise SEQ ID NO:

7.

56. The synthetic combination of claim 40, wherein the fungal endophytes comprise SEQ ID NO:

53.

57. The synthetic combination of claim 40, wherein the fungal endophytes comprise SEQ ID NO:

54.

58. The synthetic combination of claim 40, wherein the fungal endophytes comprise SEQ ID NO:

55.

59. The synthetic combination of claim 40, wherein the fungal endophytes comprise SEQ ID NO:

73.