Isolated Complex Endophyte Compositions and Methods for Improved Plant Traits

• US 20160316760A1
• Filed: 04/29/2016
• Published: 11/03/2016
• Est. Priority Date: 05/01/2015
• Status: Active Grant

• Current Assignee: Indigo Agriculture, Inc.
• Original Assignee: Indigo Agriculture, Inc.
• Inventors: Boghigian, Brett A., Gray, Paul Andrew, Ambrose, Karen V., Djonovic, Slavica, Toledo, Gerardo V., Marquez, Luis Miguel, von Maltzahn, Geoffrey
• Granted Patent: US 10,212,940 B2
• Time in Patent Office: Days
• US Class Current: 1/1
• CPC Class Codes: A01G 18/10 Mycorrhiza; Mycorrhizal associations
• CPC Class Codes: A01G 22/00 Cultivation of specific crops or plants not otherwise provided for
• CPC Class Codes: A01G 22/05 Fruit crops, e.g. strawberries, tomatoes or cucumbers
• CPC Class Codes: A01G 22/10 Asparagus
• CPC Class Codes: A01G 22/15 Leaf crops, e.g. lettuce or spinach
• CPC Class Codes: A01G 22/20 Cereals
• CPC Class Codes: A01G 22/22 Rice
• CPC Class Codes: A01G 22/25 Root crops, e.g. potatoes, yams, beet or wasabi
• CPC Class Codes: A01G 22/35 Bulbs; Alliums, e.g. onions or leeks
• CPC Class Codes: A01G 22/40 Fabaceae, e.g. beans or peas
• CPC Class Codes: A01G 22/50 Cotton
• CPC Class Codes: A01G 22/55 Sugar cane
• CPC Class Codes: A01G 22/60 Flowers; Ornamental plants
• CPC Class Codes: A01G 24/30 based on or containing synthetic organic compounds
• CPC Class Codes: A01N 63/00 Biocides, 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 containing compounds of determined constitution A01N27/00 - A01N59/00; unicellular algae A01N65/03
• CPC Class Codes: A01N 63/20 Bacteria; Substances produced thereby or obtained therefrom
• CPC Class Codes: A01N 63/30 Microbial fungi; Substances produced thereby or obtained therefrom

Abstract

This invention relates to methods and materials for providing a benefit to a plant by associating the plant with a complex endophyte comprising a host fungus further comprising a component bacterium, including benefits to a plant derived from a seed or other plant element treated with a complex endophyte. For example, this invention provides purified complex endophytes, purified complex endophyte components such as bacteria or fungi, synthetic combinations comprising said complex endophytes and/or components, and methods of making and using the same.

Claims

1. A synthetic composition, comprising a plant element heterologously associated with a complex endophyte, wherein said complex endophyte is capable of providing a trait of agronomic importance to said plant element.

2. The synthetic composition of claim 1, wherein said trait of agronomic importance is selected from the group consisting of:

altered oil content, altered protein content, altered seed carbohydrate composition, altered seed oil composition, altered seed protein composition, increased chemical tolerance, increased cold tolerance, delayed senescence, increased disease resistance, increased drought tolerance, increased ear weight, growth improvement, health enhancement, increased heat tolerance, increased herbicide tolerance, increased herbivore resistance, improved nitrogen fixation, improved nitrogen utilization, improved nutrient use efficiency, improved root architecture, improved water use efficiency, increased biomass, increased root length, increased seedling root length, germination rate, increased seed weight, increased shoot length, increased seedling shoot length, increased shoot biomass, seedling mass, root surface area, increased yield, increased yield under water-limited conditions, increased kernel mass, improved kernel moisture content, increased metal tolerance, increased number of ears, increased number of kernels per ear, increased number of pods, nutrition enhancement, improved pathogen resistance, improved pest resistance, photosynthetic capability improvement, salinity tolerance, stay-green, vigor improvement, increased dry weight of mature seeds, increased fresh weight of mature seeds, increased number of mature seeds per plant, increased chlorophyll content, increased seed germination, increased number of pods per plant, increased length of pods per plant, reduced number of wilted leaves per plant, reduced number of severely wilted leaves per plant, increased number of non-wilted leaves per plant, increased plant height, earlier or increased flowering, increased protein content, increased fermentable carbohydrate content, reduced lignin content, male sterility, and increased antioxidant content.

3. The synthetic composition of claim 2, wherein said trait of agronomic importance is selected from the group consisting of:

germination rate, emergence rate, shoot biomass, root biomass, seedling root length, seedling shoot length, seedling mass, root surface area, and yield.

4. The synthetic composition of any of claims 1-3, wherein said synthetic composition further comprises an agronomic formulation that further comprises one or more of the following:

a stabilizer, or a preservative, or a carrier, or a surfactant, or an anticomplex agent, fungicide, nematicide, bactericide, insecticide, and herbicide, or any combination thereof.

5. The synthetic composition of any of claims 1-4, wherein said complex endophyte is present in an amount of at least about 10̂

2 CFU per plant element.

6. The synthetic composition of any of claims 1-5, wherein said complex endophyte comprises a host fungus from a class selected from the group consisting of:

Dothideomycetes, Sordariomycetes, Eurotiomycetes, Glomeromycetes, Basidiomycetes, Agaricomycetes, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

7. The synthetic composition of any of claims 1-5, wherein said complex endophyte comprises a bacterium from a class selected from the group consisting of:

Bacilli, Betaproteobacteria, Gammaproteobacteria, Actinobacteria, Nanohaloarchaea, Alphaproteobacteria, Bacilli, Cytophagia.

8. The synthetic composition of any of claims 1-5, wherein said complex endophyte comprises a host fungus from an order selected from the group consisting of:

Botryosphaeriales, Dothideales, Pleosporales, Coniochateles, Xylariales, Mucorales, Eurotiales, Diversisporales, Agaricales, Pezizales, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

9. The synthetic composition of any of claims 1-5, wherein said complex endophyte comprises a bacterium from an order selected from the group consisting of:

Bacillales, Burkholderiales, Enterobacteriales, Xanthomonadales, Actinomycetales, Rhizobiales, Bacillales, Cytophagales.

10. The synthetic composition of any of claims 1-5, wherein said complex endophyte comprises a host fungus from a family selected from the group consisting of:

Botryosphaeriaceae, Dothioraceae, Montagnulacea, Pleosporacea, Coniochaetaceae, Amphisphaeriaceae, Xylariacea, Rhizopodaceae, Trichocomaceae, Gigasporaceae, Hydnangiaceae, Tuberaceae, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

11. The synthetic composition of any of claims 1-5, wherein said complex endophyte comprises a bacterium from a family selected from the group consisting of:

Bacillaceae, Burkholderiaceae, Enterobacteriaceae, Xanthomonadaceae, Streptomycetaceae, Rhizobiaceae, Paenibacillaceae.

12. The synthetic composition of any of claims 1-5, wherein said complex endophyte comprises a host fungus from a genus selected from the group consisting of:

Boryosphaeria, Microdiplodia, Pestalotiopsis, Phyllosticta, Alternaria, Lecythophora, Daldinia, Rhizopus, Aspergillus, Gigaspora, Piriformospora, Laccaria, Tuber, Mucor, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

13. The synthetic composition of any of claims 1-5, wherein said complex endophyte comprises a bacterium from a genus selected from the group consisting of:

Dyella, Pantoea, Luteibacter, Ralstonia, Erwinia, Bacillus, Burkholderia, Streptomyces, Candidatus, Rhizobium, Paenibacillus.

14. The synthetic composition of any of claims 1-5, wherein said complex endophyte comprises a nucleic acid sequence at least 95% identical to a nucleic acid sequence selected from the group consisting of SEQ ID NO:

1 through SEQ ID NO;

333.

15. The synthetic composition of any of claims 1-14, wherein said complex endophyte is selected from those listed in Table 3 or Table 4.

16. The synthetic composition of any of claims 1-15, wherein said complex endophyte is associated with a plant element but is not directly contacting said plant element.

17. The synthetic composition of any of claims 1-16, wherein said plant element is selected from the group consisting of:

whole plant, seedling, meristematic tissue, ground tissue, vascular tissue, dermal tissue, seed, leaf, root, shoot, stem, flower, fruit, stolon, bulb, tuber, corm, keikis, and bud.

18. The synthetic composition of any of claims 1-17, wherein said plant element is from a plant selected from the group consisting of:

wheat, soybean, maize, cotton, canola, barley, sorghum, millet, rice, rapeseed, alfalfa, tomato, sugarbeet, sorghum, almond, walnut, apple, peanut, strawberry, lettuce, orange, potato, banana, sugarcane, potato, cassava, mango, guava, palm, onions, olives, peppers, tea, yams, cacao, sunflower, asparagus, carrot, coconut, lemon, lime, barley, watermelon, cabbage, cucumber, grape, and turfgrass.

19. A plurality of synthetic compositions of any of claims 1-18, confined within an object selected from the group consisting of:

bottle, jar, ampule, package, vessel, bag, box, bin, envelope, carton, container, silo, shipping container, truck bed, and case.

20. A plurality of synthetic compositions of any of claims 1-18, placed in a medium that promotes plant growth, said medium selected from the group consisting of:

soil, hydroponic apparatus, and artificial growth medium.

21. The plurality of synthetic compositions of claim 20, wherein said medium is soil, wherein said synthetic compositions are placed in the soil with substantially equal spacing between each seed.

22. A plurality of synthetic compositions of any of claims 1-19, wherein the synthetic compositions are shelf-stable.

23. A plant grown from the synthetic combination of any of claims 1-22, wherein said plant exhibits an improved phenotype of agronomic interest, selected from the group consisting of:

altered oil content, altered protein content, altered seed carbohydrate composition, altered seed oil composition, altered seed protein composition, increased chemical tolerance, increased cold tolerance, delayed senescence, increased disease resistance, increased drought tolerance, increased ear weight, growth improvement, health enhancement, increased heat tolerance, increased herbicide tolerance, increased herbivore resistance, improved nitrogen fixation, improved nitrogen utilization, improved nutrient use efficiency, improved root architecture, improved water use efficiency, increased biomass, increased root length, increased seedling root length, germination rate, increased seed weight, increased shoot length, increased seedling shoot length, increased shoot biomass, seedling mass, root surface area, increased yield, increased yield under water-limited conditions, increased kernel mass, improved kernel moisture content, increased metal tolerance, increased number of ears, increased number of kernels per ear, increased number of pods, nutrition enhancement, improved pathogen resistance, improved pest resistance, photosynthetic capability improvement, salinity tolerance, stay-green, vigor improvement, increased dry weight of mature seeds, increased fresh weight of mature seeds, increased number of mature seeds per plant, increased chlorophyll content, increased seed germination, increased number of pods per plant, increased length of pods per plant, reduced number of wilted leaves per plant, reduced number of severely wilted leaves per plant, increased number of non-wilted leaves per plant, increased plant height, earlier or increased flowering, increased protein content, increased fermentable carbohydrate content, reduced lignin content, male sterility, and increased antioxidant content.

24. The plant, plant element, or progeny of the plant of any of claims 1-23, wherein said plant or progeny of the plant comprises in at least one of its plant elements said complex endophyte, fungal host, or bacterial component.

25. A method of inoculating a plant with a fungal endophyte, comprising contacting a plant element of said plant with a formulation comprising a heterologous complex endophyte, wherein said complex endophyte comprises said fungal endophyte.

27. The method of claim 25 or 26, wherein the inoculation improves a trait of agronomic importance in the plant.

36. The method of any of claims 25-35, wherein the trait of agronomic importance is improved under normal watering conditions.

37. The method of any of claims 25-35, wherein the trait of agronomic importance is improved under conditions of water limitation.

38. The method of any of claims 25-37, wherein said plant reproductive element is a seed from a soybean plant, and wherein said complex endophyte comprises a fungus of the genus Dothideomyetes.

39. The method of any of claims 25-37, wherein said plant reproductive element is a seed from a wheat plant, and wherein said complex endophyte comprises a fungus of the genus Sodariomycetes.

40. The method of any of claims 25-39, wherein the complex endophyte is present in the formulation in an amount capable of modulating at least one of:

a trait of agronomic importance, the transcription of a gene, the expression of a protein, the level of a hormone, the level of a metabolite, and the population of endogenous microbes in plants grown from said seeds, as compared to isoline plants not associated with, or grown from plant elements associated with, said complex endophyte.

41. The method of any of claims 25-40, wherein said agronomic formulation further comprises one or more of the following:

a stabilizer, or a preservative, or a carrier, or a surfactant, or an anticomplex agent, fungicide, nematicide, bactericide, insecticide, and herbicide, or any combination thereof.

42. The method of any of claims 25-41, wherein said complex endophyte is present in an amount of at least about 10̂

2 CFU per plant element.

43. The method of any of claims 25-42, wherein said complex endophyte comprises a host fungus from a class selected from the group consisting of:

Dothideomycetes, Sordariomycetes, Eurotiomycetes, Glomeromycetes, Basidiomycetes, Agaricomycetes, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

44. The method of any of claims 25-42, wherein said complex endophyte comprises a bacterium from a class selected from the group consisting of:

Bacilli, Betaproteobacteria, Gammaproteobacteria, Actinobacteria, Nanohaloarchaea, Alphaproteobacteria, Bacilli, Cytophagia.

45. The method of any of claims 25-42, wherein said complex endophyte comprises a host Botryosphaeriales, Dothideales, Pleosporales, Coniochateles, Xylariales, Mucorales, Eurotiales, Diversisporales, Agaricales, Pezizales, selected from the group consisting of:

Botryosphaeriales, Dothideales, Pleosporales, Coniochateles, Xylariales, Mucorales, Eurotiales, Diversisporales, Agaricales, Pezizales, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

46. The method of any of claims 25-42, wherein said complex endophyte comprises a bacterium from an order selected from the group consisting of:

Bacillales, Burkholderiales, Enterobacteriales, Xanthomonadales, Actinomycetales, Rhizobiales, Bacillales, Cytophagales.

47. The method of any of claims 25-42, wherein said complex endophyte comprises a host fungus from a family selected from the group consisting of:

Botryosphaeriaceae, Dothioraceae, Montagnulacea, Pleosporacea, Coniochaetaceae, Amphisphaeriaceae, Xylariacea, Rhizopodaceae, Trichocomaceae, Gigasporaceae, Hydnangiaceae, Tuberaceae, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

48. The method of any of claims 25-42, wherein said complex endophyte comprises a bacterium from a family selected from the group consisting of:

Bacillaceae, Burkholderiaceae, Enterobacteriaceae, Xanthomonadaceae, Streptomycetaceae, Rhizobiaceae, Paenibacillaceae.

49. The method of any of claims 25-42, wherein said complex endophyte comprises a host fungus from a genus selected from the group consisting of:

Boryosphaeria, Microdiplodia, Pestalotiopsis, Phyllosticta, Alternaria, Lecythophora, Daldinia, Rhizopus, Aspergillus, Gigaspora, Piriformospora, Laccaria, Tuber, Mucor, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

50. The method of any of claims 25-42, wherein said complex endophyte comprises a bacterium from a genus selected from the group consisting of:

Dyella, Pantoea, Luteibacter, Ralstonia, Erwinia, Bacillus, Burkholderia, Streptomyces, Candidatus, Rhizobium, Paenibacillus.

51. The method of any of claims 25-42, wherein said complex endophyte comprises a nucleic acid sequence at least 95% identical to a nucleic acid sequence selected from the group consisting of SEQ ID NO:

1 through SEQ ID NO;

333.

52. The method of any of claims 25-51, wherein said complex endophyte is selected from those listed in Table 3 or Table 4.

53. The method of any of claims 25-52, wherein said complex endophyte is associated with a plant element but is not directly contacting said plant element.

54. The method of any of claims 25-53, wherein said plant element is selected from the group consisting of:

whole plant, seedling, meristematic tissue, ground tissue, vascular tissue, dermal tissue, seed, leaf, root, shoot, stem, flower, fruit, stolon, bulb, tuber, corm, keikis, and bud.

55. The method of any of claims 25-54, wherein said plant element is from a plant selected from the group consisting of:

wheat, soybean, maize, cotton, canola, barley, sorghum, millet, rice, rapeseed, alfalfa, tomato, sugarbeet, sorghum, almond, walnut, apple, peanut, strawberry, lettuce, orange, potato, banana, sugarcane, potato, cassava, mango, guava, palm, onions, olives, peppers, tea, yams, cacao, sunflower, asparagus, carrot, coconut, lemon, lime, barley, watermelon, cabbage, cucumber, grape, and turfgrass.

56. A plant element from the plant produced by the method of any of claims 25-55.

26. A method of inoculating a plant with a bacterial endophyte, comprising contacting a plant element of said plant with a formulation comprising a heterologous complex endophyte, wherein said complex endophyte comprises said bacterial endophyte.

28. A method of improving a trait of agronomic importance in a plant, comprising contacting a plant element with a formulation comprising a heterologous complex endophyte;

as compared to an isoline plant grown from a plant reproductive element not associated with said complex endophyte.

30. The method of claim 28 or 29, wherein the complex endophyte comprises a bacterium within a host fungus.

31. The method of claim 28 or 29, wherein the complex endophyte comprises a fungus within a host fungus.

29. A method of improving a trait of agronomic importance in a plant, comprising growing said plant from a plant reproductive element that has been contacted with a formulation comprising a heterologous complex endophyte;

as compared to an isoline plant grown from a plant reproductive element not associated with said complex endophyte.

32. A method of improving a trait of agronomic importance in a plant, comprising growing said plant from a plant reproductive element that has been contacted with a formulation comprising a heterologous complex endophyte, wherein the complex endophyte comprises a bacterium within a host fungus;

as compared to an isoline plant grown from a plant reproductive element not associated with said bacterium.

34. The method of any of claims 32-33, wherein the trait of agronomic importance is selected from the group consisting of:

altered oil content, altered protein content, altered seed carbohydrate composition, altered seed oil composition, altered seed protein composition, increased chemical tolerance, increased cold tolerance, delayed senescence, increased disease resistance, increased drought tolerance, increased ear weight, growth improvement, health enhancement, increased heat tolerance, increased herbicide tolerance, increased herbivore resistance, improved nitrogen fixation, improved nitrogen utilization, improved nutrient use efficiency, improved root architecture, improved water use efficiency, increased biomass, increased root length, increased seedling root length, germination rate, increased seed weight, increased shoot length, increased seedling shoot length, increased shoot biomass, seedling mass, root surface area, increased yield, increased yield under water-limited conditions, increased kernel mass, improved kernel moisture content, increased metal tolerance, increased number of ears, increased number of kernels per ear, increased number of pods, nutrition enhancement, improved pathogen resistance, improved pest resistance, photosynthetic capability improvement, salinity tolerance, stay-green, vigor improvement, increased dry weight of mature seeds, increased fresh weight of mature seeds, increased number of mature seeds per plant, increased chlorophyll content, increased seed germination, increased number of pods per plant, increased length of pods per plant, reduced number of wilted leaves per plant, reduced number of severely wilted leaves per plant, increased number of non-wilted leaves per plant, increased plant height, earlier or increased flowering, increased protein content, increased fermentable carbohydrate content, reduced lignin content, male sterility, and increased antioxidant content.

35. The method of claim 34, wherein said trait of agronomic importance is selected from the group consisting of:

germination rate, emergence rate, shoot biomass, seedling root length, seedling shoot length, seedling mass, root surface area, and yield.

33. A method of improving a trait of agronomic importance in a plant, comprising growing said plant from a plant reproductive element that has been contacted with a formulation comprising a heterologous complex endophyte, wherein the complex endophyte comprises a fungus within a host fungus;

as compared to an isoline plant grown from a plant reproductive element not associated with said fungus.

57. A method of improving a trait of agronomic importance in a plant, comprising isolating a bacterial endophyte from a complex endophyte, and growing said plant from a plant reproductive element onto which said bacterial endophyte is heterologously disposed;

as compared to an isoline plant grown from a plant reproductive element not associated with said bacterial endophyte.

58. The method of claim 57, wherein said trait of agronomic importance is selected from the group consisting of:

altered oil content, altered protein content, altered seed carbohydrate composition, altered seed oil composition, altered seed protein composition, increased chemical tolerance, increased cold tolerance, delayed senescence, increased disease resistance, increased drought tolerance, increased ear weight, growth improvement, health enhancement, increased heat tolerance, increased herbicide tolerance, increased herbivore resistance, improved nitrogen fixation, improved nitrogen utilization, improved nutrient use efficiency, improved root architecture, improved water use efficiency, increased biomass, increased root length, increased seedling root length, germination rate, increased seed weight, increased shoot length, increased seedling shoot length, increased shoot biomass, seedling mass, root surface area, increased yield, increased yield under water-limited conditions, increased kernel mass, improved kernel moisture content, increased metal tolerance, increased number of ears, increased number of kernels per ear, increased number of pods, nutrition enhancement, improved pathogen resistance, improved pest resistance, photosynthetic capability improvement, salinity tolerance, stay-green, vigor improvement, increased dry weight of mature seeds, increased fresh weight of mature seeds, increased number of mature seeds per plant, increased chlorophyll content, increased seed germination, increased number of pods per plant, increased length of pods per plant, reduced number of wilted leaves per plant, reduced number of severely wilted leaves per plant, increased number of non-wilted leaves per plant, increased plant height, earlier or increased flowering, increased protein content, increased fermentable carbohydrate content, reduced lignin content, male sterility, and increased antioxidant content.

59. The method of claim 58, wherein said trait of agronomic importance is selected from the group consisting of:

germination rate, emergence rate, shoot biomass, seedling root length, seedling shoot length, seedling mass, root surface area, and yield.

60. The method of any of claims 57-59, wherein the trait of agronomic importance is improved under normal watering conditions.

61. The method of any of claims 57-59, wherein the trait of agronomic importance is improved under conditions of water limitation.

62. The method of any of claims 57-61, wherein said complex endophyte comprises a host fungus from a class selected from the group consisting of:

Dothideomycetes, Sordariomycetes, Eurotiomycetes, Glomeromycetes, Basidiomycetes, Agaricomycetes, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

63. The method of any of claims 57-61, wherein said complex endophyte comprises a bacterium from a class selected from the group consisting of:

Bacilli, Betaproteobacteria, Gammaproteobacteria, Actinobacteria, Nanohaloarchaea, Alphaproteobacteria, Bacilli, Cytophagia.

64. The method of any of claims 57-61, wherein said complex endophyte comprises a host fungus from an order selected from the group consisting of:

Botryosphaeriales, Dothideales, Pleosporales, Coniochateles, Xylariales, Mucorales, Eurotiales, Diversisporales, Agaricales, Pezizales, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

65. The method of any of claims 57-61, wherein said complex endophyte comprises a bacterium from an order selected from the group consisting of:

Bacillales, Burkholderiales, Enterobacteriales, Xanthomonadales, Actinomycetales, Rhizobiales, Bacillales, Cytophagales.

66. The method of any of claims 57-61, wherein said complex endophyte comprises a host fungus from a family selected from the group consisting of:

Botryosphaeriaceae, Dothioraceae, Montagnulacea, Pleosporacea, Coniochaetaceae, Amphisphaeriaceae, Xylariacea, Rhizopodaceae, Trichocomaceae, Gigasporaceae, Hydnangiaceae, Tuberaceae, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

67. The method of any of claims 57-61, wherein said complex endophyte comprises a bacterium from a family selected from the group consisting of:

Bacillaceae, Burkholderiaceae, Enterobacteriaceae, Xanthomonadaceae, Streptomycetaceae, Rhizobiaceae, Paenibacillaceae.

68. The method of any of claims 57-61, wherein said complex endophyte comprises a host fungus from a genus selected from the group consisting of:

Boryosphaeria, Microdiplodia, Pestalotiopsis, Phyllosticta, Alternaria, Lecythophora, Daldinia, Rhizopus, Aspergillus, Gigaspora, Piriformospora, Laccaria, Tuber, Mucor, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

69. The method of any of claims 57-61, wherein said complex endophyte comprises a bacterium from a genus selected from the group consisting of:

Dyella, Pantoea, Luteibacter, Ralstonia, Erwinia, Bacillus, Burkholderia, Streptomyces, Candidatus, Rhizobium, Paenibacillus.

70. The method of any of claims 57-61, wherein the bacterium comprises a nucleic acid sequence at least 95% identical to a nucleic acid sequence selected from the group consisting of SEQ ID NO:

1 through SEQ ID NO;

333.

71. The method of any of claims 57-61, wherein said complex endophyte is selected from those listed in Table 3 or Table 4.

72. A plant element from the plant produced by the method of any of claims 57-70.

73. A method for preparing a synthetic composition, comprising associating the surface of a plurality of plant elements with a formulation comprising a purified microbial population that comprises a complex endophyte that is heterologous to the seed, wherein the complex endophyte is present in the formulation in an amount capable of modulating at least one of:

a trait of agronomic importance, the transcription of a gene, the expression of a protein, the level of a hormone, the level of a metabolite, and the population of endogenous microbes in plants grown from said seeds, as compared to isoline plants not associated with, or grown from plant elements associated with, said formulation.

74. The method of claim 73, wherein said trait of agronomic importance is selected from the group consisting of:

altered oil content, altered protein content, altered seed carbohydrate composition, altered seed oil composition, altered seed protein composition, increased chemical tolerance, increased cold tolerance, delayed senescence, increased disease resistance, increased drought tolerance, increased ear weight, growth improvement, health enhancement, increased heat tolerance, increased herbicide tolerance, increased herbivore resistance, improved nitrogen fixation, improved nitrogen utilization, improved nutrient use efficiency, improved root architecture, improved water use efficiency, increased biomass, increased root length, increased seedling root length, germination rate, increased seed weight, increased shoot length, increased seedling shoot length, increased shoot biomass, seedling mass, root surface area, increased yield, increased yield under water-limited conditions, increased kernel mass, improved kernel moisture content, increased metal tolerance, increased number of ears, increased number of kernels per ear, increased number of pods, nutrition enhancement, improved pathogen resistance, improved pest resistance, photosynthetic capability improvement, salinity tolerance, stay-green, vigor improvement, increased dry weight of mature seeds, increased fresh weight of mature seeds, increased number of mature seeds per plant, increased chlorophyll content, increased seed germination, increased number of pods per plant, increased length of pods per plant, reduced number of wilted leaves per plant, reduced number of severely wilted leaves per plant, increased number of non-wilted leaves per plant, increased plant height, earlier or increased flowering, increased protein content, increased fermentable carbohydrate content, reduced lignin content, male sterility, and increased antioxidant content.

75. The method of claim 74, wherein said trait of agronomic importance is selected from the group consisting of:

germination rate, emergence rate, shoot biomass, seedling root length, seedling shoot length, seedling mass, root surface area, and yield.

76. The method of any of claims 73-75, wherein the trait of agronomic importance is improved under normal watering conditions.

77. The method of any of claims 73-75, wherein the trait of agronomic importance is improved under conditions of water limitation.

78. The method of any of claims 73-77, wherein said synthetic composition further comprises an agronomic formulation that further comprises one or more of the following:

a stabilizer, or a preservative, or a carrier, or a surfactant, or an anticomplex agent, fungicide, nematicide, bactericide, insecticide, and herbicide, or any combination thereof.

79. The method of any of claims 73-78, wherein said complex endophyte is present in an amount of at least about 10̂

2 CFU per plant element.

80. The method of any of claims 73-79, wherein said complex endophyte comprises a host fungus from a class selected from the group consisting of:

Dothideomycetes, Sordariomycetes, Eurotiomycetes, Glomeromycetes, Basidiomycetes, Agaricomycetes, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

81. The method of any of claims 73-79, wherein said complex endophyte comprises a bacterium from a class selected from the group consisting of:

Bacilli, Betaproteobacteria, Gammaproteobacteria, Actinobacteria, Nanohaloarchaea, Alphaproteobacteria, Bacilli, Cytophagia.

82. The method of any of claims 73-79, wherein said complex endophyte comprises a host fungus from an order selected from the group consisting of:

Botryosphaeriales, Dothideales, Pleosporales, Coniochateles, Xylariales, Mucorales, Eurotiales, Diversisporales, Agaricales, Pezizales, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

83. The method of any of claims 73-79, wherein said complex endophyte comprises a bacterium from an order selected from the group consisting of:

Bacillales, Burkholderiales, Enterobacteriales, Xanthomonadales, Actinomycetales, Rhizobiales, Bacillales, Cytophagales.

84. The method of any of claims 73-79, wherein said complex endophyte comprises a host fungus from a family selected from the group consisting of:

Botryosphaeriaceae, Dothioraceae, Montagnulacea, Pleosporacea, Coniochaetaceae, Amphisphaeriaceae, Xylariacea, Rhizopodaceae, Trichocomaceae, Gigasporaceae, Hydnangiaceae, Tuberaceae, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

85. The method of any of claims 73-79, wherein said complex endophyte comprises a bacterium from a family selected from the group consisting of:

Bacillaceae, Burkholderiaceae, Enterobacteriaceae, Xanthomonadaceae, Streptomycetaceae, Rhizobiaceae, Paenibacillaceae.

86. The method of any of claims 73-79, wherein said complex endophyte comprises a host fungus from a genus selected from the group consisting of:

Boryosphaeria, Microdiplodia, Pestalotiopsis, Phyllosticta, Alternaria, Lecythophora, Daldinia, Rhizopus, Aspergillus, Gigaspora, Piriformospora, Laccaria, Tuber, Mucor, or any of the corresponding anamorph or teleomorph taxonomy of the preceding.

87. The method of any of claims 73-79, wherein said complex endophyte comprises a bacterium from a genus selected from the group consisting of:

Dyella, Pantoea, Luteibacter, Ralstonia, Erwinia, Bacillus, Burkholderia, Streptomyces, Candidatus, Rhizobium, Paenibacillus.

88. The method of any of claims 73-79, wherein said complex endophyte comprises a nucleic acid sequence at least 95% identical to a nucleic acid sequence selected from the group consisting of SEQ ID NO:

1 through SEQ ID NO;

333.

89. The method of any of claims 73-88, wherein said complex endophyte is selected from those listed in Table 3 or Table 4.

90. The method of any of claims 73-89, wherein said complex endophyte is associated with a plant element but is not directly contacting said plant element.

91. The method of any of claims 73-90, wherein said plant element is selected from the group consisting of:

whole plant, seedling, meristematic tissue, ground tissue, vascular tissue, dermal tissue, seed, leaf, root, shoot, stem, flower, fruit, stolon, bulb, tuber, corm, keikis, and bud.

92. The method of any of claims 73-91, wherein said plant element is from a plant selected from the group consisting of:

wheat, soybean, maize, cotton, canola, barley, sorghum, millet, rice, rapeseed, alfalfa, tomato, sugarbeet, sorghum, almond, walnut, apple, peanut, strawberry, lettuce, orange, potato, banana, sugarcane, potato, cassava, mango, guava, palm, onions, olives, peppers, tea, yams, cacao, sunflower, asparagus, carrot, coconut, lemon, lime, barley, watermelon, cabbage, cucumber, grape, and turfgrass.

93. A plant element from the plant produced by the method of any of claims 73-92.

94. A method of improving the efficacy of a bacterial endophyte in an application, comprising utilizing a complex endophyte, wherein said complex endophyte comprises said bacterial endophyte.

96. The method of claim 94 or claim 95, wherein said application is selected from the group consisting of:

agriculture, plant improvement, water quality improvement, snow or ice production, bioremediation, industrial compound production, pharmaceutical compound production, and production of bioengineered substances.

97. The method of claim 94 or claim 95, wherein said application is a production method of a composition belonging to a class of compound selected from the group consisting of:

acids, alcohols, amino acids, amylases, antibiotics, biogases, bioplastics, citric acid, enzymes, esters, fatty acids, flavoring agents, glutamic acid, human or animal hormones, human growth hormone, ice, insulin, lactic acid, lipases, lipids, minerals, nitrogen, oils, nucleic acids, pectinases, preservatives, proteins, snow, sugars, vaccines, viruses, vitamins, and waxes.

95. A method of improving the efficacy of a fungal endophyte in an application, comprising utilizing a complex endophyte, wherein said complex endophyte comprises said fungal endophyte.

98. A method of improving the performance of a bacterial endophyte in an application, comprising identifying a complex endophyte comprising a bacterium comprising a nucleic acid sequence with at least 95% identity to that of the bacterial endophyte, and substituting the complex endophyte for the bacterial endophyte in said application.

99. The method of claim 98, wherein said bacterial endophyte is further associated with a plant element.

100. The method of claim 98, wherein said bacterial endophyte is Gram-negative.

101. The method of claim 98, wherein said fungal endophyte has improved sporulation capability.

102. The method of claim 98, wherein said characteristic is selected from the group consisting of:

efficacy, survivability, shelf-stability, tolerance to an antibiotic, tolerance to reduced environmental moisture.