Methods of use of seed-origin endophyte populations

• US 10,058,101 B2
• Filed: 02/17/2017
• Issued: 08/28/2018
• Est. Priority Date: 06/26/2013
• Status: Active Grant

• Current Assignee: Indigo Ag, Inc.
• Original Assignee: Indigo Agriculture, Inc.
• Inventors: von Maltzahn, Geoffrey, Flavell, Richard Bailey, Toledo, Gerardo V., Djonovic, Slavica, Marquez, Luis Miguel, Johnston, David Morris, Millet, Yves Alain, Lyford, Jeffrey, Naydich, Alexander, Sadowski, Craig
• Primary Examiner(s): IBRAHIM, MEDINA AHMED
• Application Number:
  US15/436,592
• Publication Number:
  US 20170164619A1
• Time in Patent Office: 557 Days
• Field of Search: None
• CPC Class Codes: A01C 1/00 Apparatus, or methods of use thereof, for testing or treating seed, roots, or the like, prior to sowing or planting
• CPC Class Codes: A01C 1/06 Coating or dressing seed
• CPC Class Codes: A01N 25/00 Biocides, pest repellants or attractants, or plant growth regulators, characterised by their forms, or by their non-active ingredients or by their methods of application , e.g. seed treatment or sequential application; Substances for reducing the noxious effect of the active ingredients to organisms other than pests
• 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
• CPC Class Codes: G01N 33/5097 involving plant cells immunoassays of plant cells G01N33/56961

Abstract

This application relates to methods and materials for providing a benefit to a seed or seedling of an agricultural plant (e.g., an agricultural grass plant), or the agricultural plant derived from the seed or seedling. For example, this application provides purified bacterial populations that include novel seed-origin bacterial endophytes, and synthetic combinations of seeds and/or seedlings (e.g., cereal seeds and/or seedlings) with heterologous seed-derived bacterial endophytes.

Claims

1. A method of treating seeds of a cereal agricultural plant, comprising mechanically or manually inoculating a plurality of cereal agricultural plant seeds with an agricultural formulation comprising an agriculturally acceptable carrier and a purified bacterial population that comprises a seed bacterial endophyte of the genus Enterobacter that is heterologous to the seed, and has a 16S nucleic acid sequence that is at least 99% identical to a 16S nucleic acid sequence selected from the group consisting of SEQ ID NO:

541, 595, 540, and 550, wherein the endophyte is heterologously disposed to the seed in an amount effective to colonize a plant germinated from the inoculated seed and to increase tolerance to drought of the inoculated seed as compared to a reference seed sowed under the same conditions, wherein the plant germinated from the inoculated seed has increased growth as compared to a plant germinated from the reference seed.

2. The method of claim 1, wherein the bacterial endophyte is obtainable from an interior seed compartment.

3. The method of claim 1, wherein the bacterial endophyte is obtainable from an exterior surface of a seed.

4. The method of claim 1, wherein the bacterial endophyte is obtainable from a different cultivar, variety or crop as compared to the seed.

5. The method of claim 1, wherein the bacterial endophyte colonizes the roots of a seedling germinated from the inoculated seed.

6. The method of claim 1, wherein the bacterial population includes two or more bacterial endophytes.

7. The method of claim 1, wherein the bacterial endophyte is obtained from rice seed, maize seed, wheat seed, or barley seed.

8. The method of claim 1, wherein the agricultural formulation further comprises a purified fungal population.

9. The method of claim 1, wherein the increased tolerance to drought occurs under conditions of biotic stress.

10. The method of claim 9, wherein the biotic stress is selected from the group consisting of nematode stress, insect herbivory stress, fungal pathogen stress, bacterial pathogen stress, and viral pathogen stress.

11. The method of claim 1, wherein the plant germinated from the inoculated seed has a larger amount of lateral roots and root-hairs as compared to the plant germinated from the reference seed.

12. The method of claim 1, wherein the plant germinated from the inoculated seed has increased shoot length as compared to the plant germinated from the reference seed.

13. The method of claim 1, wherein the plant germinated from the inoculated seed has increased seedling weight as compared to the plant germinated from the reference seed.

14. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is 100% identical to SEQ ID NO:

541.

15. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is 100% identical to SEQ ID NO:

595.

16. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is 100% identical to SEQ ID NO:

540.

17. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is 100% identical to SEQ ID NO:

550.

18. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is 100% identical to a 16S nucleic acid sequence of SEQ ID NO:

540, wherein the plant germinated from the synthetic combination has increased shoot length as compared to the plant germinated from the reference seed.

19. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is 100% identical to a 16S nucleic acid sequence of SEQ ID NO:

550, wherein the plant germinated from the synthetic combination has increased shoot length as compared to the plant germinated from the reference seed.

20. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is 100% identical to a 16S nucleic acid sequence of SEQ ID NO:

541, wherein the plant germinated from the synthetic combination has increased shoot length as compared to the plant germinated from the reference seed.

21. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is 100% identical to a 16S nucleic acid sequence of SEQ ID NO:

540, wherein the plant germinated from the synthetic combination has increased seedling weight as compared to the plant germinated from the reference seed.

22. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is 100% identical to a 16S nucleic acid sequence of SEQ ID NO:

550, wherein the plant germinated from the synthetic combination has increased seedling weight as compared to the plant germinated from the reference seed.

23. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is 100% identical to a 16S nucleic acid sequence of SEQ ID NO:

541, wherein the plant germinated from the synthetic combination has increased seedling weight as compared to the plant germinated from the reference seed.

24. The method of claim 1, wherein the effective amount is at least 10³ CFU/seed on the surface of the seed.

25. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is at least 99% identical to SEQ ID NO:

541.

26. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is at least 99% identical to SEQ ID NO:

595.

27. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is at least 99% identical to SEQ ID NO:

540.

28. The method of claim 1, wherein the seed bacterial endophyte of the genus Enterobacter has a 16S nucleic acid sequence that is at least 99% identical to SEQ ID NO:

550.

29. The synthetic combination of claim 1, wherein the cereal agricultural plant is selected from the group consisting of:

maize, rice, barley, or wheat.

30. The synthetic combination of claim 1, wherein the cereal agricultural plant is maize.

31. The synthetic combination of claim 1, wherein the cereal agricultural plant is rice.

32. The synthetic combination of claim 1, wherein the cereal agricultural plant is barley.

33. The synthetic combination of claim 1, wherein the cereal agricultural plant is wheat.