Methods for improving plant responses to pests and pathogens

Abstract

A method for increasing plant responses to pests and / or pathogens by increasing the production of one or more plant defense compounds derived from anthranilates in plants. Also provided is a method for identifying and selecting microbial strains that can be used in plant treatments to increase plant responses to attacks by pests and / or pathogens.

Description

[Technical Field]

[0001] Cross-reference of priority application This patent application claims the benefits of U.S. Provisional Patent Application No. 63 / 561,055 filed on 4 March 2024, U.S. Provisional Patent Application No. 63 / 606,485 filed on 5 December 2023, and U.S. Provisional Patent Application No. 63 / 504,299 filed on 25 May 2023.

[0002] Sequence List This application includes a sequence listing submitted electronically in XML file format, which is incorporated herein by reference in its entirety. The XML file, created on 16 May 2024, is named P14472WO00.xml and has a size of 251,036 bytes. [Background technology]

[0003] Plants have developed a variety of defense mechanisms to protect themselves from attacks by various organisms, including fungi, bacteria, viruses, nematodes, and insects. These defense mechanisms include structural barriers, the production of chemicals toxic to invaders, the production of chemicals that attract natural enemies of target pests or pathogens, and hypersensitivity responses characterized by rapid cell death at the point of infection. Despite these natural protective mechanisms, crop losses due to damage from plant pathogens and pests occur every year, imposing a significant cost on the global economy. New methods are needed to mitigate the impact of pest and pathogen attacks on plants.

[0004] Summary of the Invention This specification provides methods for improving a plant's response to attack by pathogens or pests by increasing the levels of one or more plant defense compounds produced in the plant. Such methods include treating a plant, a part of a plant, or a seed with a microbial strain that is not pathogenic to the plant, wherein the treatment increases the expression in the plant of one or more gene transcripts involved in the production of plant defense compounds; and growing the plant in the presence of a pathogen or pest, thereby increasing the levels of one or more plant defense compounds in the plant compared to a control plant, improving the plant's response to the pathogen or pest compared to a control plant, wherein the control plant is not genetically modified or treated with the microbial strain. In some embodiments, the microbial strain used in the methods provided herein expresses one or more genes in the siderophore production pathway and / or one or more genes in the polyketide production pathway. In some embodiments, such microbial strains enhance the induced systemic resistance (ISR) plant defense response in the treated plant or treated seed, plant grown from a part of a plant, or plant grown in treated soil. In some embodiments, the plant defense response provides the production of metabolites that repel or otherwise reduce the effects of plant pests or pathogens on the plant. In some embodiments, the plant pest is an insect. In some embodiments, the protein in the polyketide production pathway that induces the plant response to the pathogen or pest is located on a plasmid. In some embodiments, the protein in the polyketide production pathway is encoded by the gene or a variant thereof on SEQ ID NO: 87. In some embodiments, the protein in the polyketide production pathway that induces the plant response to the pathogen or pest is encoded by the gene or a variant thereof on SEQ ID NO: 86. In some embodiments, the protein in the polyketide production pathway that induces the plant response to the pathogen or pest is encoded by a polynucleotide having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity with SEQ ID NOs: 36-50.In some embodiments, the protein in the polyketide production pathway that induces a plant response to a pathogen or pest comprises a polypeptide sequence having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to any one of SEQ ID NOs. 21-35. In some embodiments, the protein in the microorganism that induces a plant response to a pathogen or pest is a homolog or ortholog of any one of SEQ ID NOs. 21-35. In some embodiments, the gene in the polyketide production pathway is the bfmBAB_2 gene. In some embodiments, bfmBAB_2 comprises a polynucleotide sequence having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NOs. 36. In some embodiments, the bfmBAB_2 gene encodes a protein having the sequence of SEQ ID NOs. 21. In some embodiments, the bfmBAB_2 gene encodes a protein having at least 70% identity with SEQ ID NO: 36 and / or at least 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity with SEQ ID NO: 21. In some embodiments, the bacterial strain is Methylobacterium or Methylorubrum. In some embodiments, the bacterial strain is NLS0042 (NRRL B-50932). In some embodiments, the bacterial strain is not NLS0042 (NRRL B-50932). In some embodiments, the plant defense compound reduces insect larval foraging on plant roots. In some embodiments, insect larvae are repelled from the plant roots. In some embodiments, the plant defense compound reduces insect larval foraging on plant roots compared to a control plant. In some embodiments, the plant is maize and the insect pest is maize root worm. In some embodiments, root regeneration after exposure to pathogens or pests is increased compared to control plants.

[0005] In some embodiments provided herein, a method for improving a plant's response to attack by a pathogen or pest increases the level of one or more plant defense compounds derived from anthranilates produced in the plant. Such a method includes the steps of: modifying a plant genome to increase the expression of one or more gene transcripts involved in the production of one or more plant defense compounds derived from anthranilates in the plant; and / or treating a plant, a part of a plant, or a seed with a microbial strain that is not pathogenic to the plant, wherein the treatment increases the expression of one or more gene transcripts involved in the production of plant defense compounds derived from anthranilates in the plant; and cultivating the plant in the presence of a pathogen or pest, thereby increasing the level of one or more plant defense compounds in the plant compared to a control plant, improving the plant's response to the pathogen or pest compared to a control plant, wherein the control plant is not genetically modified or treated with the microbial strain. In some embodiments, the expression of one or more gene transcripts related to the production of anthranilates and / or the conversion of anthranilates to plant defense compounds is increased compared to a control plant. In some embodiments, transcription of genes encoding anthranilate synthase protein components is increased compared to control plants. In some embodiments, transcription of anthranilate synthase alpha or beta subunit component genes is increased compared to control plants. In some embodiments, transcription of genes encoding anthranilate N-benzoyltransferase is increased compared to control plants. In some embodiments, the plant defense compound is an anthranilate ester. In some embodiments, the anthranilate ester is selected from the group consisting of methyl anthranilate, dimethyl anthranilate, ethyl anthranilate, phenylethyl anthranilate, and menthyl anthranilate. In some embodiments, the plant defense compound is an anthranilate-derived phytoalexin. In some embodiments, the microbial strain that enhances the plant's response to pathogens or pests is a bacterial strain.In some embodiments, the bacterial strain is Methylobacterium or Methylorubrum. In some embodiments, the bacterial strain is NLS0042 (NRRL B-50932). In some embodiments, the bacterial strain is not NLS0042 (NRRL B-50932). In some embodiments, the plant defense compound reduces insect larval foraging on plant roots. In some embodiments, the plant defense compound reduces insect larval foraging on plant roots compared to a control plant. In some embodiments, the plant is maize and the insect pest is maize root worm. In some embodiments, root regeneration after exposure to the pathogen or pest is increased compared to a control plant. In some embodiments, the expression of anthranilate synthase protein component having the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 4, or a protein having the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 4, is increased compared to a control plant. In some embodiments, the expression of anthranilate synthase proteins or protein components containing amino acid sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NOs. 9, 10, 12, 14, 15, 17, 18, or 20 is increased compared to control plants. In some embodiments, the expression of the gene encoding anthranilate N-benzoyltransferase of SEQ ID NOs. 6 or 7 is increased compared to control plants. In some embodiments, the expression of the gene encoding anthranilate N-benzoyltransferase having at least 70%, 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NOs. 6 or 7 is increased compared to control plants. In some embodiments of the methods provided herein, the plant defense compound is not an indole derivative.

[0006] In some embodiments of the methods provided herein, the insect pest is a thrips, and the treated plant is corn, soybean, cotton, peanut, potato, tomato, or strawberry. In some embodiments, the insect pest is an aphid, and the plant is soybean, cotton, wheat, potato, tomato, strawberry, or pepper plant. In some embodiments, the insect pest is the fall armyworm, and the plant is soybean, cotton, wheat, rice, or strawberry plant. In some embodiments, the insect pest is a leafhopper, and the plant is rice, potato, tomato, or bayberry plant. In some embodiments, the insect pest is a lepidoptera, and the plant is soybean, tomato, pepper, or bayberry. In some embodiments, the insect pest is a nematode, and the plant is soybean, cotton, potato, or tomato plant. In some embodiments, the insect pest is a horsehair worm and the plant is a corn, soybean, or potato plant. In some embodiments, the insect pest is a mirid bug and the plant is a cotton, tomato, or strawberry plant. In some embodiments, the insect pest is a cutworm and the plant is a corn or strawberry plant. In some embodiments, the insect pest is a flea beetle and the plant is a Brassica plant. In some embodiments, the insect pest is a gall midge and the plant is a soybean plant. In some embodiments, the insect pest is a stink bug and the plant is a tomato plant. In some embodiments, the insect pest is a Colorado potato beetle and the plant is a potato plant. In some embodiments, the insect pest is a rice water weevil and the plant is a rice plant. In some embodiments, the insect pest is a wheat sawfly, a grain beetle, or a wheat mite and the plant is a wheat plant. In some embodiments, the insect pest is an alfalfa hopper, a tobacco budworm larva, or a ground bug, and the plant is a peanut plant. In some embodiments, the insect pest is a ground worm, and the plant is a soybean plant. In some embodiments, the insect pest is a maize root worm, and the plant is a maize plant.

[0007] A method is provided for reducing or repelling pathogens or pests by increasing the levels of one or more plant defense compounds, such microbial strains protect plant roots from attack by pathogens or pests. In some embodiments, the proteins in the polyketide production pathway that induce a plant response to pathogens or pests reside on a recruitable plasmid, and optionally, the recruitable plasmid is heterologous to the bacterial strain containing the recruitable plasmid. In some embodiments, the proteins in the polyketide production pathway are encoded by the gene on SEQ ID NO: 87 or a variant thereof. Variants of SEQ ID NO: 87 include (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NO: 87, and / or (ii) a DNA sequence encoding the protein encoded by SEQ ID NO: 87. In some embodiments, the proteins in the polyketide production pathway that induce a plant response to pathogens or pests are encoded by the gene on SEQ ID NO: 86 or a variant thereof. A variant of SEQ ID NO: 86 comprises (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NO: 86, and / or (ii) a DNA sequence encoding the protein encoded by SEQ ID NO: 86. In some embodiments, the protein in the polyketide production pathway that induces a plant response to a pathogen or pest is encoded by polynucleotide SEQ ID NOs: 36-50. In some embodiments, the protein in the polyketide production pathway that induces a plant response to a pathogen or pest comprises a polypeptide having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to any one of SEQ ID NOs: 21-35. In some embodiments, the protein in a microorganism that induces a plant response to a pathogen or pest is a homolog or ortholog of any one of SEQ ID NOs: 21-35. In some embodiments, the gene in the polyketide production pathway is the bfmBAB_2 gene. In some embodiments, bfmBAB_2 has the sequence of sequence number 36.In some embodiments, the bfmBAB_2 gene encodes a protein having the sequence of SEQ ID NO: 21. In some embodiments, the bfmBAB_2 gene encodes a protein having at least 70% identity to SEQ ID NO: 36 and / or at least 80% identity to SEQ ID NO: 21. In some embodiments, the bacterial strain is Methylobacterium or Methylorubrum. In some embodiments, the bacterial strain is NLS0042 (NRRL B-50932). In some embodiments, the bacterial strain is not NLS0042 (NRRL B-50932). In some embodiments, the plant defense compound reduces insect larval foraging on plant roots. In some embodiments, insect larvae are repelled from the plant roots. In some embodiments, the plant defense compound reduces insect larval foraging on plant roots compared to a control plant. In some embodiments, the plant is maize and the insect pest is maize root worm. In some embodiments, root regeneration after exposure to the pathogen or pest is increased compared to a control plant. In some embodiments, the plant defense compound reduces insect larval foraging on plant roots. In some embodiments, the plant defense compound reduces insect larval foraging on plant roots compared to a control plant. In some embodiments, the plant is corn and the insect pest is maize rhizomeworm. In some embodiments, the bacterial strain is Methylobacterium or Methylorubrum. In some embodiments, the bacterial strain is NLS0042 (NRRL B-50932). In some embodiments, the bacterial strain is not NLS0042 (NRRL B-50932). In some embodiments, the plant defense compound reduces insect larval foraging on plant roots. In some embodiments, the plant defense compound reduces insect larval foraging on plant roots compared to a control plant. In some embodiments, the plant is corn and the insect pest is maize rhizomeworm.

[0008] A method for reducing or repelling pathogens or pests by treating soil, plants, parts of plants, or seeds with a microbial strain to produce metabolites derived from such microbial strain, wherein such metabolites enhance the defense mechanisms of the plants, parts of plants, or seeds, and the defense mechanisms protect the plant roots from attack by pathogens or pests. In some embodiments, a method for reducing or repelling pathogen or pest damage includes treating soil, plants, parts of plants, or seeds with a microbial strain such that the microbial strain expresses metabolites, and cultivating plants in the presence of pathogens or pests such that the treated plants, parts of plants, or seeds are more repelling or have reduced pathogen or pest damage compared to a control plant, while the control plant is not genetically modified or treated with such microbial strain. A method is also provided for reducing or repelling pathogens or pests by treating soil, plants, parts of plants, or seeds with a microbial strain to produce one or more metabolites and / or peptides derived from such microbial strain, wherein the one or more metabolites and / or peptides enhance the defense mechanisms of the plant, part of a plant, or seed, and these defense mechanisms protect the plant roots from attack by pathogens or pests. In some embodiments, a method for reducing or repelling pathogen or pest damage includes treating soil, plants, parts of plants, or seeds with a microbial strain in which the microbial strain expresses one or more genes or gene pathways involved in metabolite biosynthesis, and one or more metabolites enhance the plant's response to pathogens and / or pests; and cultivating plants in the presence of pathogens or pests so that the treated plants, parts of plants, or seeds repel or reduce pathogen or pest damage compared to control plants, while control plants are either not treated with the microbial strain or are treated with it. In some embodiments, the microbial strain produces siderophores or polyketides. In some embodiments, the polyketides produced by the microbial strain are antimicrobial compounds.In some embodiments, the protein in the polyketide production pathway that induces a plant response to a pathogen or pest is located on a recruitable plasmid. In some embodiments, the protein in the polyketide production pathway is encoded by the gene on SEQ ID NO: 87 or a variant thereof. A variant of SEQ ID NO: 87 includes (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NO: 87, and / or (ii) a DNA sequence encoding the protein encoded by SEQ ID NO: 87. In some embodiments, the protein in the polyketide production pathway that induces a plant response to a pathogen or pest is encoded by the gene on SEQ ID NO: 86 or a variant thereof. A variant of SEQ ID NO: 86 includes (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NO: 86, and / or (ii) a DNA sequence encoding the protein encoded by SEQ ID NO: 86. In some embodiments, the proteins in the polyketide production pathway that induce a plant response to a pathogen or pest are encoded by SEQ ID NOs: 36-50. In some embodiments, the proteins in the polyketide production pathway that induce a plant response to a pathogen or pest have one of the sequences from SEQ ID NOs: 21-35. In some embodiments, the proteins in microorganisms that induce a plant response to a pathogen or pest are homologs or orthologues of one of the sequences from SEQ ID NOs: 21-35. In some embodiments, the gene in the polyketide production pathway is the bfmBAB_2 gene. In some embodiments, bfmBAB_2 has the sequence from SEQ ID NO: 36. In some embodiments, the bfmBAB_2 gene encodes a protein having the sequence from SEQ ID NO: 21. In some embodiments, the bfmBAB_2 gene has at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity with SEQ ID NO: 36, and / or at least 80%, 85%, 90%, 95%, 98%, 99%, or 100% identity with SEQ ID NO: 21.In some embodiments, the plant is a cereal crop such as maize, rice, wheat, rye, oats, barley, and millet, or the plant is grass, soybeans, strawberries, or cotton, and the insect pest is a cutworm. In some embodiments, the cutworm is the fall armyworm. In some embodiments, the cutworm is a Spodoptera species, including, but not limited to, S. frugiperda and S. exiqua. In some embodiments, the plant is a Solanaceae plant such as tomato, tobacco, eggplant, pepper, and potato, and the insect pest is a hawk moth. In some embodiments, the hawk moth is a Manduca species, including M. quinquemaculata and M. sexta. In some embodiments, the plant is soybeans, and the insect pest is a soybean inchworm. In some embodiments, the plant is tobacco or cotton, and the pest is a thrips. In some embodiments, the thrips is the citrus thrips. In some embodiments, the bacterial strain is Methylobacterium or Methylorubrum. In some embodiments, the bacterial strain is NLS0042 (NRRL B-50932). In some embodiments, the bacterial strain is not NLS0042 (NRRL B-50932). In some embodiments, the plant defense compound reduces insect larval foraging on plant roots. In some embodiments, the plant defense compound reduces insect larval foraging on plant roots compared to a control plant. In some embodiments, the plant is corn and the insect pest is maize root worm.

[0009] A method is provided herein for identifying a microbial strain that enhances a plant's response to a pathogen or pest, wherein the microbial strain is not pathogenic to the plant. Such a method includes: (i) treating a plant, a part of a plant, or a seed of a plant with at least a first microbial strain that is not pathogenic to the plant to obtain treated seeds and / or a treated plant; (ii) growing the treated plant in the presence of the pathogen or pest, or growing a plant from a part of a treated plant or a treated seed; (iii) taking one or more tissue samples from the plant and from an untreated control plant, wherein the tissue samples are taken during a growth stage while the pest or pathogen is attacking the plant tissue; and (iv) assaying the samples to identify an increased production of one or more plant defense compounds derived from anthranilates in the treated plant compared to a control plant, thereby identifying a microbial strain that enhances a plant's response to the pathogen or pest. In some embodiments, the control plant is an untreated plant. In some embodiments, the control plant is treated with a different microorganism. In some embodiments, such a method further includes the step of selecting a sample for analysis from the treated plant that shows reduced damage from the pathogen or pest compared to the control plant. In some embodiments, the sample is assayed to determine the level of one or more gene transcripts related to the production of anthranilates and / or the conversion of anthranilates to plant defense compounds. In some embodiments, the sample is assayed to determine the level of one or more plant defense compounds derived from anthranilates. In some embodiments, the pathogen or pest is a fungus, bacterium, nematode, insect, or virus. In some embodiments, a portion of the treated plant is selected from the group consisting of leaves, stems, shoots, flowers, fruits, buds, roots, tubers, rhizomes, runners, bulbs, and corms. In some embodiments, the collected tissue sample is selected from the group consisting of leaf, stem, shoot, flower, fruit, bud, root, tuber, rhizome, runner, bulb, and corm tissue samples.In some embodiments, plant tissue samples are analyzed to determine the level of one or more gene transcripts encoding anthranilate synthase protein components or the level of anthranilate synthase protein components (e.g., by an enzyme assay or immunoassay). In some embodiments, the anthranilate synthase protein components are alpha or beta subunits. In some embodiments, plant tissue samples are analyzed to determine the level of gene transcripts encoding anthranilate N-benzoyltransferase or the level of anthranilate N-benzoyltransferase (e.g., by an enzyme assay or immunoassay). In some embodiments, the plant defense compound is an anthranilate ester. In some embodiments, the anthranilate ester is selected from the group consisting of methyl anthranilate, dimethyl anthranilate, ethyl anthranilate, phenylethyl anthranilate, and menthyl anthranilate. In some embodiments, the plant defense compound is an anthranilate-derived phytoalexin.

[0010] A further method that can be used to identify microbial strains that enhance a plant's response to a pathogen or pest, wherein the microbial strain is not pathogenic to the plant, comprises screening a sample containing one or more microbial strains for the presence of one or more genes encoding one or more proteins for the production of compounds that enhance a plant's response to a pathogen or pest. In some embodiments, the microbial strain is identified by the presence of one or more genes in the siderophore biosynthesis pathway. In some embodiments, the microbial strain is identified by the presence of one or more genes in the polyketide biosynthesis pathway. In some embodiments, the polyketide is an antimicrobial compound. In some embodiments, the proteins in the polyketide production pathway that induce a plant's response to a pathogen or pest are located on a recruitable plasmid. In some embodiments, the proteins in the polyketide production pathway are encoded by the gene on SEQ ID NO: 87, or by variants of those genes containing DNA sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to the protein-coding sequence of SEQ ID NO: 87. In some embodiments, proteins in the polyketide production pathway that induces a plant response to a pathogen or pest are encoded by the gene on SEQ ID NO: 86, or by variants of that gene containing DNA sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to the protein-coding sequence of SEQ ID NO: 86. In some embodiments, proteins in the polyketide production pathway that induces a plant response to a pathogen or pest are encoded by polynucleotides containing DNA sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to one or more of SEQ ID NOs: 36-50. In some embodiments, the proteins in the polyketide production pathway that induces a plant response to a pathogen or pest include an amino acid sequence having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity with any one of SEQ ID NOs. 21-35.In some embodiments, the protein in the microorganism that induces a plant response to a pathogen or pest is a homolog or ortholog of any one of SEQ ID NOs. 21-35. In some embodiments, the gene in the polyketide production pathway is the bfmBAB_2 gene. In some embodiments, bfmBAB_2 contains a polynucleotide sequence having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NO. 36. In some embodiments, the bfmBAB_2 gene encodes a protein having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NO. 21. In some embodiments, the bfmBAB_2 gene encodes a protein having at least 70%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NO: 36, and / or at least 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NO: 21. In some embodiments of any of the above methods and compositions, the microbial strain is a bacterial strain or a fungal strain. In some embodiments, the bacterial strain is a Methylobacterium or Methylorubrum strain. In some embodiments, the plant is selected from the group consisting of corn, wheat, rye, rice, alfalfa, barley, oats, rye, sorghum, millet, soybeans, Brassica, Hemp, tobacco, potatoes, peanuts, carrots, cotton, coffee, coconuts, sugar beets, oats, barley, tomatoes, pumpkins, cucumbers, gourds, lettuce, peppers, peas, onions, green beans, bay beans, sunflowers, safflower, sweet potatoes, cassava, coffee, coconuts, conifers, turfgrass, leafy vegetables, microgreens, herbs, fruit plants such as strawberries, and fruit trees including but not limited to apple trees and nut trees, as well as ornamental plants. In some embodiments, the plant is a maize plant. In some embodiments, the pest is an insect. In some embodiments, the tissue sample is a root sample. In some embodiments, the pest is a maize rhizome worm.

[0011] Methods and compositions that can be used to treat plants with microorganisms identified using the methods described herein in order to enhance the plant's response to one or more pathogens or pests are also provided herein. In some embodiments, the microbial compositions provided herein include a microbial strain that enhances the plant's defense response to pathogens and / or pests, and additional components for enhancing the long-term storage of the microbial strain, facilitating the treatment of plants and / or parts of plants, and / or functioning as plant growth regulators. In some embodiments, the microbial strains provided herein are in a solid composition as an essentially dry product having, for example, about 5% or less moisture. In some embodiments, the solid composition is in powder or granular form. In some embodiments, the composition provided herein is a liquid culture containing water, oil, and / or polymers. In some embodiments, the microorganisms provided herein are provided as a fluid liquid. In some embodiments, the microorganisms are stabilized in a suspension-like concentrate in which the continuous phase is miscible with water but is not water. For example, the continuous phase in such a suspension-like concentrate can be a polymer such as a polyether. In some embodiments, the microbial composition is provided as an oil dispersion. In some embodiments, the microorganism is encapsulated in a protective carrier. In some embodiments, the composition provided herein comprises Methylobacterium strain NLS0042 (NRRL B-50932) or a derivative thereof as a dry powder. In some embodiments, the composition comprising Methylobacterium strain NLS0042 (NRRL B-50932) or a derivative thereof comprises additional components to facilitate long-term storage as a dry composition. In some embodiments, the long-term stability of Methylobacterium strain NLS0042 (NRRL B-50932) or a derivative thereof in the composition comprising the dry powder and / or on treated seeds is enhanced compared to an undried composition or composition lacking additional components that enhance the long-term storage of the microbial strain and / or facilitate the processing of the plant and / or parts of the plant.In some embodiments, a composition comprising Methylobacterium strain NLS0042 (NRRL B-50932) or a derivative thereof comprises one or more oligosaccharides or polysaccharides. In some embodiments, a composition comprising Methylobacterium strain NLS0042 (NRRL B-50932) or a derivative thereof comprises one or more polysaccharides selected from gums including but not limited to dextrin, maltodextrin, disaccharides, starch, chitosan, alginates, and gums including, but not limited to, karaya gum, jaguar gum, xanthan gum, glucomannan, tragacanth gum, konjac gum, polysaccharide gum, mucus, gum arabic, and other natural gums. In some embodiments, a dry composition comprising Methylobacterium strain NLS0042 (NRRL B-50932) or a derivative thereof, with enhanced stability compared to other compositions, further comprises maltodextrin, trehalose, and / or glucomannan. [Brief explanation of the drawing]

[0012] [Figure 1] This figure shows the results of a feeding selection assay when larvae choose between NLS0042-treated and untreated corn roots. A vast majority of larvae chose the untreated roots over the NLS0042-treated roots. The circles indicate the percentage of larvae making a given selection in each of the 12 replicates of this experiment. [Figure 2] A figure showing the results of a feeding selection assay when selecting between two NLS0042-treated maize roots. In this case, most larvae do not select and instead remain in the central petri dish from which they started. The circles indicate the percentage of larvae that make a given selection in each of the 12 replicates of this experiment. [Figure 3] A figure showing the results of a feeding selection assay when selecting between two untreated corn roots. The circles indicate the percentage of larvae making a given selection in each of the 12 replicates of this experiment. [Modes for carrying out the invention]

[0013] definition As used herein, the term "and / or" is construed to mean that each of two or more designated features or components is specifically disclosed, with or without the other. Accordingly, as used herein in phrases such as "A and / or B," the term "and / or" is intended to include "A and B," "A or B," "A" (alone), and "B" (alone). Similarly, as used in phrases such as "A, B, and / or C," the term "and / or" is intended to include each of the following embodiments: A, B, and C; A, B, or C; A or C; A or B; B or C; A and C; A and B; B and C; A (alone), B (alone), and C (alone).

[0014] As used herein, the terms “include,” “includes,” and “including” are to be interpreted as encompassing at least the features they refer to, or including the items they refer to, but not excluding any additional, unspecified features or items.

[0015] As used herein, the term “biological preparation” refers to a component of a composition for treating a plant or part of a plant that consists of or is derived from a microorganism. Biological preparations include biological control agents, other beneficial microorganisms, microbial extracts, natural products, plant growth activators, or plant defense agents. Non-limiting examples of biological control agents include bacteria, fungi, beneficial nematodes, and viruses. In certain compositions, a biological preparation may include a single culture or co-culture of Methylobacterium, or a combination of separately cultured Methylobacterium strains or isolates.

[0016] As used herein, the term “Methylobacterium” refers to the genera and species of the family methylobacteriaceae, including bacterial species of the genus Methylobacterium and the proposed genus Methylorubrum (Green and Ardley (2018)). Methylobacterium includes pink-pigmented facultative methylotropic bacteria (PPFM), as well as non-pink-pigmented Methylobacterium nodulans, and colorless mutants of Methylobacterium isolates. For example, “Methylobacterium” refers to, but is not limited to, the bacterial species listed below, and any new Methylobacterium species that have not yet been reported or described and can be characterized as Methylobacterium or Methylorubrum based on phylogenetic analysis. Methylobacterium adhaesivum, Methylobacterium oryzae, Methylobacterium aerolatum, Methylobacterium oxalidis, Methylobacterium aquaticum, Methylobacterium persicinum, Methylobacterium brachiatum, Methylobacterium phyllosphaerae, Methylobacterium brachythecii, Methylobacterium phyllostachyos, Methylobacterium bullatum, Methylobacterium platani, Methylobacterium cerastii, Methylobacterium pseudosasicola, Methylobacterium currus, Methylobacterium radiotolerans, Methylobacterium dankookense, Methylobacterium soli, Methylobacterium frigidaeris, Methylobacterium specialis, Methylobacterium fujisawaense, Methylobacteriumtardum、Methylobacterium gnaphalii、Methylobacterium tarhaniae、Methylobacterium goesingense、Methylobacterium thuringiense、Methylobacterium gossipiicola、Methylobacterium trifolii、Methylobacterium gregans、Methylobacterium variabile、Methylobacterium haplocladii、Methylobacterium aminovorans(Methylorubrum aminovorans)、Methylobacterium hispanicum、Methylobacterium extorquens(Methylorubrum extorquens)、Methylobacterium indicum、Methylobacterium podarium(Methylorubrum podarium)、Methylobacterium iners、Methylobacterium populi(Methylorubrum populi)、Methylobacterium isbiliense、Methylobacterium pseudosasae(Methylorubrum pseudosasae)、Methylobacterium jeotgali、Methylobacterium rhodesianum(Methylorubrum rhodesianum)、Methylobacterium komagatae、Methylobacterium rhodinum(Methylorubrum rhodinum)、Methylobacterium longum、Methylobacterium salsuginis(Methylorubrum salsuginis)、Methylobacterium marchantiae、Methylobacterium suomiense(Methylorubrum suomiense、Methylobacterium mesophilicum、Methylobacteriumthiocyanatum (Methylorubrum thiocyanatum), Methylobacterium nodulans, Methylobacterium zatmanii (Methylorubrum zatmanii), or Methylobacterium organophilum.

[0017] As used herein, “mineral nutrients” (sometimes simply “nutrients”) are micronutrients or macronutrients that are necessary or useful for plants or parts of plants, including, but not limited to, nitrogen (N), potassium (K), calcium (Ca), magnesium (Mg), phosphorus (P), and sulfur (S), as well as the micronutrients chlorine (Cl), iron (Fe), boron (B), manganese (Mn), zinc (Z), cobalt (Co), copper (Cu), molybdenum (Mo), and nickel (Ni).

[0018] As used herein, “vitamin” refers to an organic compound that is necessary in small amounts for normal growth and metabolism. Vitamins are important for the growth of humans and / or animals, and some vitamins have been reported to be beneficial to plants. Vitamins include, but are not limited to, vitamin A (all-trans retinol and all-trans retinyl esters, as well as all-trans beta-carotene and other provitamin A carotenoids), vitamin B1 (thiamine), vitamin B2 (riboflavin), vitamin B3 (niacin), vitamin B5 (pantothenic acid), vitamin B6 (pyridoxine), vitamin B7 (biotin), vitamin B9 (folic acid or folate), vitamin B12 (cobalamin), vitamin C (ascorbic acid), vitamin D (calciferol), vitamin E (tocopherol and tocotrienol), and vitamin K (quinone).

[0019] As used herein, the term “strain” should include all isolates of such strain.

[0020] As used herein, the term “recruitable plasmid” refers to a plasmid that can be transferred from a donor strain to a recipient strain. A recruitable plasmid as defined herein contains the cis-acting DNA element (i.e., oriT) necessary for conjugation and has a functional origin of replication in Methylobacterium. Other elements necessary for conjugation may also be encoded on a recruitable plasmid. A conjugate or autotransmissible plasmid is considered a recruitable plasmid for use in the manner defined herein, containing the cis-acting DNA necessary for conjugation, encoding all the genes necessary for DNA transfer into a recipient cell / line or isolate. As used herein, “variant” refers, when used in relation to a Methylobacterium isolate, to any isolate having chromosomal genomic DNA with at least 99%, 99.9%, 99.8%, 99.7%, 99.6%, or 99.5% sequence identity with the chromosomal genomic DNA of a reference Methylobacterium isolate, such as the deposited Methylobacterium isolate provided herein. Isolate variants can be obtained from a variety of sources, including soil, plants, or plant material, as well as water, particularly water related to plants and / or agriculture. Variants include derivatives obtained from deposited isolates. Methylobacterium isolates or strains can be sequenced (as taught, e.g., by Sanger et al. (1977), Bentley et al. (2008), or Caporaso et al. (2012)), and genome-scale comparisons of sequences can be performed using sequence analysis tools such as BLAST (taught by Altschul et al. (1990)) or clustalw (www.ebi.ac.uk / Tools / msa / clustalw2 / ) (Konstantinidis et al. (2005)). Variants can be identified, for example, by the presence of the 16S sequence of a reference strain, and variants also exhibit plant productivity-enhancing traits of the reference strain.

[0021] As used herein, "derivative", when used in connection with a Methylobacterium isolate, refers to any Methylobacterium obtained from the deposited Methylobacterium isolate provided herein. Derivatives of a Methylobacterium isolate include, but are not limited to, derivatives obtained by selection, derivatives selected by mutagenesis and selection, and genetically transformed Methylobacterium obtained from a Methylobacterium isolate. A "derivative" can be identified, for example, based on genetic identity to the strain or isolate from which it was obtained, and generally exhibits chromosomal genomic DNA having at least 99%, 99.9%, 99.8%, 99.7%, 99.6%, or 99.5% sequence identity to the chromosomal genomic DNA of the strain or isolate from which it is derived.

[0022] As used herein, "sequence identity" or "percent identity" refers to a measure of genomic similarity at the nucleotide level between the coding regions of two genomes when used to assess whether a particular Methylobacterium strain is a variant or derivative of a Methylobacterium strain provided herein. Sequence identity between the coding regions of bacterial genomes can be calculated, for example, by determining the average nucleotide identity (ANI) score using FastANI (Jain et al. "High throughput ANI analysis of 90K prokaryotic genomes reveals clear species boundaries", Nat Communications 9, 5114 (2018)), and Han et al. ("ANI tools web: a web tool for fast genome comparison within multiple bacterial strains"; Database, 2016, 1-5).

[0023] As used herein, "leafy vegetable plants" refers to vegetable crops having edible leaves, including but not limited to spinach, kale, lettuce (including romaine, butterhead, iceberg, and looseleaf lettuce), collard greens, cabbage, beet greens, cress, Swiss chard, arugula, escarole, endive, bok choy, and the leaves of turnips. Leafy vegetable plants also, as used herein, refer to plants cultivated for the harvest of microgreens and / or herbs, including but not limited to lettuce, cauliflower, broccoli, cabbage, cress, arugula, garlic, onion, leek, amaranth, mustard spinach, beans, spinach, melons, cucumbers, pumpkins, basil, celery, coriander, radishes, radicchio, chicory, dill, rosemary, French tarragon, basil, shiso, carrots, fennel, beans, peas, chickpeas, and lentils. Leafy vegetable plants also refer to mixtures of various leafy vegetable plants, such as mesclun or other mixed salad vegetables or mixed microgreens. As used herein, "leafy vegetable plants" also encompasses other Brassicaceae or Brassicaceae vegetables not specifically named herein.

[0024] As used herein, "fruit" or "fruiting plants" can be fleshy fruiting plants, including but not limited to melons (including watermelon and cantaloupe), berries (including strawberries, blueberries, blackberries, and raspberries), grapes, kiwis, mangoes, papayas, pineapples, bananas, peppers, tomatoes, pumpkins, and cucumber plants. As used herein, "fruit" or "fruiting plants" can also refer to fruit-bearing trees or fruiting fruit trees, including but not limited to apples, peaches, pears, lemons, limes, oranges (and other citrus fruits), cherries, plums, apricots, nectarines, elderberries, pomegranates, persimmons, papayas, figs, avocados, and guavas.

[0025] As used herein, “ornamental” plants refer to plants cultivated primarily for display purposes and not for functional purposes, including but not limited to perennial plants and woody shrubs, ornamental grasses, potted flowers, cut flowers, and bulbs, including but not limited to tulips, hyacinths, daffodils, petunias, and carnations, as well as potted flowers produced from vegetatively propagated cuttings, including but not limited to poinsettias and chrysanthemums.

[0026] As used herein, “genetic element” refers to an element within a DNA or RNA molecule that contains a series of adjacent nucleotides, each at least 20 nucleotides long and up to 50, 100, 1,000, or 10,000 or more nucleic acid lengths. A genetic element may include, for example, a different group of adjacent nucleic acids, such as the genome of a plant-associated microorganism, which contains introns and exons. Genetic elements may reside on chromosomes or on extrachromosomal elements such as plasmids. In eukaryotic plant-associated microorganisms, genetic elements may reside in the nucleus or in mitochondria. In some embodiments, a genetic element is a functional genetic element (e.g., a gene) that codes for a peptide or protein.

[0027] As used herein, the terms “homologous” or “ortholog” refer to related genetic elements or proteins encoded by those genetic elements, determined based on the degree of sequence identity. These terms describe the relationship between a genetic element or encoded protein found in one isolate, species, or strain and a corresponding or equivalent genetic element or protein found in another isolate, species, or strain. As used herein, a particular genetic element in a first isolate, species, or strain is considered equivalent to a genetic element present in a second isolate, species, or strain if the protein encoded by that genetic element in the first isolate, species, or strain has at least 50 percent identity. Percentage identity can be determined using a number of software programs available in the art, including BLASTP, ClustalW, ALLALIGN, DNASTAR, SIM, SEQALN, NEEDLE, SSEARCH, etc.

[0028] As used herein, the term “metabolites” refers to substances produced during the metabolism of a microbial strain, including but not limited to siderophores, non-ribosomal peptides, polyketides, or combinations thereof.

[0029] If a term is provided in the singular form, other embodiments described by the plural form of that term are also provided.

[0030] To the extent that any of the definitions provided herein conflicts with any definition provided in any patent or non-patent reference incorporated herein by reference, any patent or non-patent reference cited herein, or any patent or non-patent reference found elsewhere, the definitions provided herein shall be used herein.

[0031] Further explanation The methods and compositions provided herein may be used to improve the plant response to attacks by pests and / or pathogens by increasing the levels of one or more plant defense compounds in the plant. In some embodiments of the methods provided herein, a plant, part of a plant, or seed reduces or repels pathogens or pests after treatment of the soil, plant, part of a plant, or seed with a microbial strain that produces metabolites, and such metabolites induce a defense response in the plant, protecting the plant from attacks by pathogens or pests. In some embodiments, the plant response is improved by increasing the levels of one or more plant defense compounds derived from anthranilic acid in the plant. Anthranilates, the conjugate base of anthranilic acid, are synthesized in plants from colismate by the action of the enzyme anthranilate synthase (EC 4.1.3.27), a heterotetrameric enzyme consisting of two α and two β subunits. This reaction is a branching point from the aromatic amino acid pathway to tryptophan biosynthesis, and the tryptophan binding site involved in feedback inhibition of tryptophan synthesis is located in the α subunit. Overexpression of the Trp-insensitive anthranilate synthase (AS) α subunit has been reported to increase tryptophan levels in transgenic plants and is hypothesized to increase the content of indole-related compounds involved in insect resistance in biosynthetic pathways branching off from the tryptophan pathway. Anthranilates are also involved in the production of other plant secondary metabolites, including anthranilate esters and anthramides, in various plants. Methyl anthranilate (MA) is naturally present in various plants and is used as a repellent against birds and insects. MA occurs in very small amounts on maize roots and has been reported to induce a strong repellent response by neonatal rhizome insect larvae in laboratory bioassays (Bernklau et al. (2016) J. Economic Entomology 109:1683-1690).

[0032] In some embodiments, the anthranilate-derived plant defense compound is not derived from indole. In some embodiments, the anthranilate-derived plant defense compound is an anthranilate ester. In some embodiments, the anthranilate ester is selected from the group consisting of methyl anthranilate, dimethyl anthranilate, ethyl anthranilate, phenylethyl anthranilate, and menthyl anthranilate. In some embodiments, the anthranilate-derived plant defense compound is a phytoalexin. In some embodiments, the phytoalexin is an amide biosynthesized after conversion from anthranilate to N-benzoylanthranilate catalyzed by anthranilate N-benzoyltransferase (EC2.3.1.44).

[0033] In some embodiments of the methods provided herein, a plant genome is modified to provide increased levels of plant defense compounds derived from anthranilates. In some embodiments, the plant genome is modified to enhance the expression of one or more gene transcripts involved in the production of such plant defense compounds. In other embodiments, the plant genome is modified to reduce the expression of genes contributing to tryptophan synthesis and to provide an accumulation of anthranilates for use in enzymatic reactions resulting in the production of anthranilate esters and anthramides. In some embodiments, the plant is genetically modified to express heterologous transcripts of genes involved in the production of plant defense compounds. Heterologous transcripts may be derived, for example, from different plant sources, microbial sources, or may be produced synthetically. Various plant regulators are used in such ways, including tissue-specific promoters that target increased expression in specific one or more tissues that are under attack by plant pathogens or pests. In some embodiments, root, leaf, green tissue, fruit, tuber, seed, or vascular tissue-specific promoters are used. Constitutive promoters are also used in the methods to provide expression in a wide range of plants, plant parts, and plant tissues. In some embodiments, the expression of natural plant gene transcripts is increased.

[0034] In some embodiments, gene modification is achieved by gene editing techniques that enable specific recombination of endogenous genes in plant genomes, including those involving CRISPR / CAS (such as those disclosed in U.S. Patent Applications Publications 2015 / 0344912, 2016 / 0138008, 2018 / 0179547, 2020 / 0172886, and 2022 / 0282244, which are incorporated herein by reference in their entirety), meganucleases, zinc finger nucleases (ZFNs), and transcription activator-like effector nucleases (TALENs). In certain embodiments, gene editing reagents can be used to increase the expression of one or more plant genes involved in the production of anthranilate-derived plant defense compounds by incorporating DNA containing a transcriptional enhancer element (e.g., U.S. Patent Application Publication US2016 / 0168584, which is entirely incorporated herein) into the promoters of those genes, either by homology-directed repair (HDR) or non-homologous end joining (NHEJ). In certain embodiments, the promoter or the promoter containing the enhancer insertion is operably ligated to an anthranilate synthase protein component, which optionally contains the amino acid sequences of SEQ ID NOs. 2, SEQ ID NOs. 4, or a sequence having at least 90%, 95%, 98%, or 99% sequence identity thereto. In certain embodiments, a promoter including a promoter or enhancer insertion is operably ligated to a gene encoding an anthranilate N-benzoyltransferase, which optionally contains the polypeptide sequence of SEQ ID NO: 6 and / or SEQ ID NO: 7, or a sequence having at least 90%, 95%, 98%, or 99% sequence identity thereto, and is increased compared to a control plant. In certain embodiments, a promoter including a promoter or enhancer insertion is operably ligated to a maize gene provided in Table 6 and optionally located in maize or other monocotyledonous plants.

[0035] In other embodiments provided herein, levels of anthranilate-derived plant defense compounds are increased as a result of treating the plant, including seeds, or a portion thereof, with one or more microbial strains that are not pathogenic to the plant, and the treatment results in increased expression in the plant of one or more gene transcripts involved in the production of anthranilate-derived plant defense compounds. In some embodiments, the microbial strains are not pathogenic to the treated plant, even though they may be pathogenic to other plants not treated by the methods described herein. In some embodiments, the microbial strains are beneficial to the treated plant.

[0036] In some embodiments of the methods provided herein, plants or parts of plants or seeds reduce or repel pathogens or pests after the soil, plants, parts of plants, or seeds are treated with a microbial strain that produces metabolites, such metabolites that enhance the defense mechanisms of the plants, parts of plants, or seeds, which protect the plants or plant roots from attack by pathogens or pests. In some embodiments, the plant defense mechanisms include increased production of anthranilates and / or anthranilate-derived compounds. In some embodiments, the production of anthranilates and / or anthranilate-derived compounds in the roots is enhanced. In some embodiments, the plant defense response protects the plant roots from attack. In some embodiments, the plant response is improved by treating the plants, parts of plants, seeds, or soil with a microorganism that expresses genes involved in inducing an ISR response, such as genes involved in the biosynthesis of siderophores and / or polyketides. In some embodiments, the proteins in the polyketide production pathway that induce the plant response to pathogens or pests are located on a recruitable plasmid. In some embodiments, the protein in the polyketide production pathway is encoded by a gene on DNA having the sequence of SEQ ID NO: 87, or a variant thereof comprising (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NO: 87, and / or (ii) a DNA sequence encoding the protein encoded by SEQ ID NO: 87. In some embodiments, the protein in the polyketide production pathway that induces a plant response to a pathogen or pest is encoded by a gene on DNA having the sequence of SEQ ID NO: 86, or a variant thereof comprising (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NO: 86, and / or (ii) a DNA sequence encoding the protein encoded by SEQ ID NO: 86.In some embodiments, the proteins in the polyketide production pathway that induces a plant response to a pathogen or pest are encoded by polynucleotides comprising DNA sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NOs. 36-50. In some embodiments, the proteins in the polyketide production pathway that induces a plant response to a pathogen or pest include polypeptide sequence plant responses to pathogens or pests, encoded by polynucleotides comprising DNA sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to any one of SEQ ID NOs. 21-35. In some embodiments, the proteins in microorganisms that induce a plant response to a pathogen or pest are homologs or orthologues of any one of SEQ ID NOs. 21-35. In some embodiments, the gene in the polyketide production pathway is the bfmBAB_2 gene. In some embodiments, the bfmBAB_2 gene contains the polynucleotide sequence of SEQ ID NO: 36. In some embodiments, the bfmBAB_2 gene encodes a protein having the sequence of SEQ ID NO: 21. In some embodiments, the bfmBAB_2 gene encodes a protein containing a polynucleotide having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NO: 36, and / or an amino acid sequence having at least 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NO: 21. In some embodiments, the bacterial strain is Methylobacterium or Methylorubrum. In some embodiments, the bacterial strain is NLS0042 (NRRL B-50932). In some embodiments, the bacterial strain is not NLS0042 (NRRL B-50932).In some embodiments, a method for reducing or repelling pathogen or pest feeding damage includes treating soil, plants, parts of plants, or seeds with a microbial strain, wherein the microbial strain expresses metabolites; and cultivating plants in the presence of pathogens or pests, thereby the treated plants, parts of plants, or seeds being more repelling of pathogens or pests or having reduced pathogen or pest feeding damage compared to control plants, while the control plants are not genetically modified or treated with the microbial strain. In some embodiments, plant defense compounds reduce insect larval foraging on plant roots. In some embodiments, plant defense compounds reduce insect larval foraging on plant roots compared to controls.

[0037] In some embodiments, the plant is treated with a bacterial strain to increase the expression in the plant of one or more gene transcripts involved in the production of anthranilate-derived plant defense compounds, or to reduce or repel plant pathogens or pests. In some embodiments, the plant defense compound is derived from anthranilate.

[0038] Provided are microbial strains containing heterologous DNA that can confer resistance, act as repellents, and confer tolerance, reduced damage, foraging, reduced infection, and / or reduced invasion of plants by pests or pathogens, as well as methods for producing such microbial strains. In some embodiments, the DNA is transferred from a microbial strain that can confer resistance, act as a repellent, and confer tolerance, reduced damage, foraging, reduced infection, and / or reduced invasion of plants by pests or pathogens, to a separate microbial strain lacking that DNA. In certain embodiments, the heterologous DNA transferred to a distinctly different microbial strain encodes a protein having any one sequence of SEQ ID NOs. 21-35, or a protein sequence having at least 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to any one of SEQ ID NOs. 21-35. In some embodiments, the DNA is transferred from a microbial strain that induces a plant response to insects and / or pathogens onto a recruitable plasmid. In some embodiments, the recruitable plasmid is transferred from one Methylobacterium or Methylorubrum strain to a second Methylobacterium or Methylorubrum strain. In some embodiments, the recruitable plasmid includes SEQ ID NO: 87, or a variant thereof, comprising (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NO: 87, and / or (ii) a DNA sequence encoding the protein encoded by SEQ ID NO: 87. In some embodiments, the recruitable plasmid includes SEQ ID NO: 86, or a variant thereof, comprising (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NO: 86, and / or (ii) a DNA sequence encoding the protein encoded by SEQ ID NO: 86.In further embodiments, recruitable plasmids, which are transferred to microorganisms to confer the ability to induce a plant defense response, encode one or more proteins comprising amino acid sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NOs. 21–35. In some embodiments, the genes encoding proteins that enhance the plant's response to insects and / or pathogens are heterologous to the microbial host. In some embodiments, the genes encoding one or more of SEQ ID NOs. 21–35, or their derivatives, variants, homologs, or orthologues, are genetically engineered to prepare recombinant constructs that provide expression of one or more proteins comprising amino acid sequences having at least SEQ ID NOs. In some embodiments, such recombinant constructs include regulatory sequences for providing expression of the one or more proteins in a target microbial host. In some embodiments, a recombinant DNA construct for the expression of one or more proteins containing amino acid sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity with SEQ ID NOs. 21–35 is stably integrated into the genome of a target microbial host. In some embodiments, a construct for the expression of one or more proteins containing amino acid sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity with SEQ ID NOs. 21–35 is introduced and maintained on a plasmid or other extrachromosomal element of a target microbial host. In some embodiments, one or more genes encoding proteins containing amino acid sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity with SEQ ID NOs. 21–35 are located in an operon and expressed from there. In other embodiments, one or more genes encoding proteins containing amino acid sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity with SEQ ID NOs. 21-35 are present in each recombinant expression construct.Also provided are recombinant DNA constructs comprising heterologous promoters operably linked to one or more genes encoding proteins containing amino acid sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NOs. 21-35, as well as microbial cells comprising the recombinant DNA constructs. In some embodiments, a microbial strain engineered to express one or more proteins containing amino acid sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NOs. 21-35 is a bacterial strain. In certain embodiments, the aforementioned or otherwise provided DNA molecules are introduced into microbial strains other than Methylobacterium species #4 (NLS0042, NRRL B-50932). In certain embodiments, the aforementioned or otherwise provided DNA molecules are introduced into microbial strains of Table 1 selected from the group consisting of Methylobacterium species #1-3, #5-53, and #54. In certain embodiments, the aforementioned or otherwise provided DNA molecules are introduced into Methylorubrum species, including Methylorubrum species #63 in Table 1. In some embodiments, a microbial strain is a fungal strain that has been engineered to express one or more proteins containing amino acid sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NOs. 21-35.

[0039] Bacterial strains used in this method include Actinomycetes, Agrobacterium, Arthrobacter, Alcaligenes, Aureobacterium, Azobacter, Azorhizobium, Azospirillum, Azotobacter, Beijerinckia, Bacillus, Brevibacillus, Burkholde. ria, Chromobacterium, Clostridium, Clavibacter, Comomonas, Corynebacterium, Curtobacterium, Enterobacter, Flavobacterium, Gluconacetobacter, Gluconobacter, Herbaspirillum, Hydrogenophage, Kleb This includes, but is not limited to, bacterial strains of the genera Siella, Luteibacter, Lysinibacillus, Mesorhizobium, Methylobacterium, Methylorubrum, Microbacterium, Ochrobactrum, Paenibacillus, Pantoea, Pasteuria, Phingobacterium, Photorhabdus, Phyllobacterium, Pseudomonas, Rhizobium, Rhodococcus, Bradyrhizobium, Serratia, Sinorhizobium, Sphingomonas, Streptomyces, Stenotrophomonas, Variovorax, Xanthomonas, and Xenorhadbus. In some embodiments, the bacteria are selected from the group consisting of Bacillus amyloliquefaciens, Bacillus cereus, Bacillus firmus, Bacillus lichenformis, Bacillus pumilus, Bacillus sphaericus, Bacillus subtilis, Bacillus thuringiensis, Chromobacterium suttsuga, Pasteuria penetrans, Pasteuria usage, and Pseudomonas fluorescens.

[0040] In some embodiments, plants are treated with beneficial fungi, including but not limited to the genera Acremonium, Alternaria, Ampelomyces, Aspergillus, Aureobasidium, Beauveria, Botryosphaeria, Cladosporium, Cochliobolus, Colletotrichum, Coniothyrium, Embellisia, Epicoccum, Fusarium, Gigaspora, Gliocladium, Glomus, Laccaria, Metarhisium, Muscodor, Nigrospora, Paecilonyces, Paraglomus, Penicillium, Phoma, Pisolithus, Podospora, Rhizopogon, Scleroderma, Trichoderma, Typhula, Ulocladium, and Verticillium. In certain embodiments, the fungi are Beauveria bassiana, Coniothyrium minitans, Gliocladium vixens, Muscodor albus, Paecilomyces lilacinus, and Trichoderma polysporum.

[0041] In some embodiments, the plants are treated with a Methylobacterium or Methylorubrum bacterial strain. In some embodiments, the Methylobacterium or Methylorubrum strain is a deposited strain disclosed in Table 1. [Table 1-1] [Table 1-2]

[0042] In certain embodiments, microbial strains provided and used in methods that can confer resistance, repellency, tolerance, reduced damage, reduced infection, reduced foraging, and / or reduced invasion to, or by, the pests or pathogens disclosed herein are obtained by transferring DNA from a microbial strain that can confer resistance, act as a repellent, and confer tolerance, reduced damage, foraging, reduced infection, and / or reduced invasion by increasing the production of one or more plant defense compounds derived from anthranilates in plants, to a microbial strain that does not confer such resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion, and then recovering or selecting a new microbial strain conferred with pest or pathogen resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion by the transferred DNA. In other embodiments, DNA transferred from a microbial strain capable of conferring resistance to pests or pathogens, acting as a repellent, and conferring tolerance, reduced damage, foraging, reduced infection, and / or reduced invasion encodes a protein having any one sequence of SEQ ID NOs. 21-35, or a protein having a sequence with at least 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to any one of SEQ ID NOs. 21-35. In some embodiments, the DNA is transferred from a microbial strain that induces a plant response to insects and / or pathogens onto a recruitable plasmid. In some embodiments, the recruitable plasmid is transferred from one Methylobacterium or Methylorubrum strain to a second Methylobacterium or Methylorubrum strain. In some embodiments, the recruitable plasmid includes (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to (ii) a DNA sequence encoding the protein encoded by (ii) 87.In some embodiments, the recruitable plasmid includes SEQ ID NO: 86, or a variant thereof, comprising (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NO: 86, and / or (ii) a DNA sequence encoding the protein encoded by SEQ ID NO: 86. In further embodiments, the recruitable plasmid, which is transferred to a microorganism to confer the ability to induce a plant defense response, encodes one or more proteins comprising polypeptide sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NOs: 21–35. In some embodiments, the recruitable plasmid, which is transferred to a microorganism to confer the ability to induce a plant defense response, comprises one or more sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NOs: 36–50. In some embodiments, a recruitable plasmid, which is transferred to a microorganism to confer the ability to induce a plant defense response, encodes one or more proteins containing polypeptide sequences having at least 70%, 80%, 85%, 90%, 95%, 98%, 99%, or 100% sequence identity to SEQ ID NOs. 51–85, or one or more proteins having sequences of at least 70%, 80%, 85%, 90%, 95%, 98%, or 100% sequence identity to SEQ ID NOs. 51–85, facilitates the transfer of the mobile plasmid.

[0043] In certain embodiments, the DNA that can confer resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion to or by a pest or pathogen by increasing the production of one or more plant defense compounds by plants, derived from anthranilates, is DNA from NLS0042 or its derivatives (e.g., a DNA donor strain), and / or the microbial strain that does not confer such resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion (e.g., a DNA recipient strain) is a strain other than NLS0042 provided in Table 1. Methods for transferring DNA from a donor strain (e.g., NLS0042) to a recipient strain (e.g., another strain in Table 1) include, but are not limited to, the DNA transfer method disclosed in U.S. Patent Application Publication No. US2021 / 0171961, which is incorporated herein by reference in its entirety.

[0044] In some embodiments of the methods provided herein, the gene transcript involved in the production of one or more plant defense compounds from anthranilates encodes an AS alpha or beta component. In some embodiments, the gene transcript encodes an AS beta component. In some embodiments, the AS beta gene transcript is a maize plant transcript. In some embodiments, the expression of the AS beta subunit protein having the sequence of SEQ ID NO: 2 or its homolog or orthologue is increased. Homologs and orthologues of SEQ ID NO: 2 include Arabidopsis thaliana proteins AT1G24909, AT1G25155, AT1G24807, AT1G25083, ASB2, and ASB1, as well as rice proteins OASB1 (Os04g0463500) and OASB2 (Os03g0718000). In some embodiments, the gene transcript encodes an AS alpha component. In some embodiments, the AS alpha gene transcript is a maize plant transcript. In some embodiments, the expression of the AS alpha subunit protein having the sequence of SEQ ID NO: 4 or its homolog or orthologue is increased. Homologs and orthologues of Sequence ID No. 4 include the Arabidopsis thaliana proteins AT3G55870, ASA1, and ASA2, as well as the rice proteins OASA2 (Os03g0264400) and OASA1 (Os03g0826500). Additional plant A alpha and beta subunit genes can be identified, for example, from plant genome sequences. In some embodiments, genes encoding AS alpha or beta protein components are transcribed to produce multiple transcripts and translated proteins. Examples of additional maize AS genes for use in the methods described herein are provided in Example 4.

[0045] In other embodiments of the methods disclosed herein, the gene transcript involved in the production of one or more plant defense compounds from anthranyrates encodes an anthranyrate N-benzoyltransferase protein that catalyzes the production of N-benzoylanthranylate from benzoyl-CoA and anthranyrates, a reaction involved in the production of anthramidophyte alexins. The enzyme EC2.3.1.144 may be referred to as anthranyrate N-hydroxycinnamoyl / benzoyltransferase due to its ability to use other thioesters of coenzyme A as donors in its reaction with anthranyrates, including cinnamoyl-CoA, 4-coumaroyl-CoA, and salicyloyl-CoA. Compounds resulting from such reactions with other donor substrates may also be involved in the production of plant defense compounds. In one embodiment of the methods described herein, the gene transcript involved in the production of one or more plant defense compounds encodes a maize anthranyrate N-benzoyltransferase protein represented by SEQ ID NO: 6 and / or SEQ ID NO: 7. Other plant anthranilate N-benzoyltransferase proteins, including DcHCBT2_Z84386 from Dianthus caryophyllus, are also of interest in the methods described herein.

[0046] In other embodiments of the methods disclosed herein, a gene transcript involved in the production of a protein that enhances a plant's response to a pathogen or pest encodes a protein having at least 80% sequence identity with SEQ ID NO: 21. In some embodiments, the gene transcript encodes a protein containing an amino acid sequence having at least 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99%, or 100% sequence identity with SEQ ID NO: 21.

[0047] The methods disclosed herein are applicable to improving plant responses to a variety of plant pests and pathogens, including bacterial and fungal pathogens, viruses, nematodes, and insects. Such pathogens may attack one or more parts of a plant, including but not limited to leaves, stems, shoots, flowers, fruits, buds, roots, tubers, rhizomes, runners, bulbs, and corms. An improved plant response to a pathogen or pest results in reduced damage or other adverse effects of the pathogen or pest. Adverse effects of a pathogen or pest attack on a plant include, but are not limited to, damage or necrosis of any kind of plant tissue, reduction of any kind of plant yield, any reduction in the value of crop plant products, and / or production of undesirable metabolites or growth products of the pathogen or pest, including but not limited to fungal metabolites or mycotoxins, or fungal growth by-products.

[0048] In certain embodiments, the methods provided herein are for Alternaria, Ascochyta, Aspergillus, Bipolaris, Botrytis, Bremia, Cercospora, Cochliobolus, Colletotrichum, Diplodia, Erysiphe, Exserohilum, Fusarium, Gaeumanomyces, Macrophonina, Magnaporthe, Nectria, Peronospora, Phakopsora, and Phialophora. It improves the plant response to fungal pathogens selected from the group consisting of species Phoma, Phymatotrichum, Phytophthora, Plasmopara, Puccinia, Podosphaera, Pyrenophora, Pyricularia, Pythium, Rhizoctonia, Sclerotium, Sclerotinia, Septoria, Stagonospora, Thielaviopsis, Uncinula, Ustilago, Venturia, and Verticillium.

[0049] In some embodiments, the methods provided herein improve the plant response to bacterial pathogens selected from the group consisting of Pseudomonas, Ralstonia, Agrobacterium, Xanthomonas, Erwinia, Xylella, Dickeya, Pectobacterium, Clavibacter, and Candidatus species.

[0050] In some embodiments, when plants are attacked by insects, an improved response can result in a reduction of damage directly caused by insects and / or damage caused by plant pathogens transmitted by insects. Viruses that affect crops include topoviruses and geminiviruses. Some common viruses that can have a serious impact on plants include tomato yellow necrosis virus, beet curly top virus, tomato yellow leaf curl virus, cucumber mosaic virus, potato virus y, potato virus x, cauliflower mosaic virus, African cassava mosaic virus, plum pox virus, brom mosaic virus, potato virus, tobacco mosaic virus, tomato yellow necrosis virus, tomato yellow leaf curl virus, cucumber mosaic virus, cauliflower mosaic virus, African cassava virus, plum pox virus, and brom mosaic virus.

[0051] Biting insects cause damage such as leaf blemishes or spots, leaf curling, and stunted or deformed fruits, in addition to the effects caused by vector pathogens. Biting insects include leafhoppers, thrips, and aphids, and attack the vascular tissue of plants, such as those present in roots, stems, leaves, and other plant organs.

[0052] In some embodiments of the methods and compositions described herein, an improved plant response to insects having chewing mouthparts (chewing insects) can be obtained. Damage caused by chewing insects can take various forms. In some cases, leaves or flowers may be completely consumed by some insect, or the plant or part of the plant may appear torn and have chewed edges or centers. In some cases, only the upper or lower surface may be consumed, which may be observed as a brown, burnt appearance or openings between leaf veins. Chewing damage to the inside of a plant may be referred to as burrowing damage or burrowing damage. Chewing and biting pests may bite and chew the leaves, stems, buds, flowers, and / or roots of plants. Damage from such pests can include leaf drop from widespread feeding, tunnel formation from insects such as leaf miners burrowing into plant leaves, ring bark stripping from beetles feeding on living trees, and root damage from insect feeding that can lead to lodging. Common chewing insect pests include snails, slugs, caterpillars, burrowing insects, armyworms, grubs, and beetles. Non-limiting examples of crops and target pests in which the methods, microbial strains, and compositions provided herein are used include: peppers—aphids and lepidopterans; tomatoes—stink bugs, aphids, whiteflies, beet leafhoppers (BCTV vector), tobacco hawk moths; peanuts—potato leafhoppers, bean beetles, lepidopteran insects; rapeseed—flea beetles; soybeans—fall armyworms, soybean inchworms, nematodes; cotton—citrus thrips, fall armyworms, nematodes, aphids; rice—fall armyworms, rice water weevils; and maize—maize root worms, armyworms, beetles, maize leaf and root aphids, ground worms, mites, armyworm larvae, and horsehair worms.

[0053] In some embodiments of the methods described herein, the insect pest is the maize rhizome worm (CRW), a member of the broad genus Diabrotica. In some embodiments, the methods described herein enhance the response of maize plants to attacks by the Western maize rhizome worm (WCR), leading to reduced lodging and / or increased yield compared to control plants. In some embodiments of the methods and compositions described herein, the improved response of maize plants to CRW feeding is obtained by treatment of maize plants, part, or seeds with beneficial microorganisms. In some embodiments, maize plants are treated with beneficial bacteria. In some embodiments, maize plants are treated with Methylobacterium or Methylorubrum species. In some embodiments, maize plants are treated with deposited Methylobacterium or Methylorubrum species listed in Table 1. In some embodiments, maize plants are treated with NLS0042 (NRRL B-50932). In some embodiments, maize plants are treated with Methylobacterium or Methylorubrum species other than NLS0042 (NRRL B-50932). In some embodiments, an improved response of maize plants to CRW herbivory is obtained by modifying the maize plant genome to increase the expression of one or more gene transcripts involved in the production of one or more plant defense compounds. In some embodiments, the maize plant genome is modified to increase the expression of native maize gene transcripts. In some embodiments, the maize plant genome is modified to increase the expression of heterologous gene transcripts, for example, from a microbial source or from a non-maize plant. In some embodiments, the plant is modified to increase the expression of anthranilate synthase subunit transcripts. In some embodiments, the expression of alpha and / or beta subunit transcripts is increased. In some embodiments, the expression of anthranilate N-benzoyltransferase is increased.In some embodiments, the expression of anthranilate synthase beta subunit transcripts and anthranilate N-benzoyltransferase transcripts is increased. In some embodiments, the expression of one or more gene transcripts encoding proteins having the sequence of SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, or SEQ ID NO: 7 is increased.

[0054] The plants that can be processed and / or genetically modified in the manner provided herein include a wide range of plants, including but not limited to crops, leafy vegetables, fruits or fruit trees, ornamental plants, turfgrasses, and trees grown for commercial production. Such plant species include, but are not limited to, maize, soybeans, Brassicaceae or Brassica genus vegetables (e.g., B. napus, B. rapa, B. juncea), alfalfa, rice, rye, wheat, barley, oats, sorghum, millet (e.g., pearl millet (Pennisetum glaucum), foxtail millet (Panicum miliaceum), proso millet (Setaria italica), and finger millet (Eleusine coracana)), sunflowers, safflower, carrots, peppers, tomatoes, pumpkins, cucumbers, melons, and other gourds, beans, peas, chickpeas, lentils, tobacco, potatoes, peanuts, cotton, berries, grapes, kiwis, mangoes, papayas, pineapples, bananas, species of hemp (including, but not limited to, Cannabis sativa and industrial hemp varieties), sweet potatoes (Ipomoea This includes batatus, cassava, coffee, coconut, ornamental plants (including but not limited to azaleas, hydrangeas, hibiscus, roses, tulips, daffodils, petunias, carnations, poinsettias, and chrysanthemums), conifers (including but not limited to pines such as loblolly pine, slash pine, Ponderosa pine, Contorta pine, and Monterey pine, fir such as Douglas fir, American hemlock, Sitka spruce, redwood, European fir and balsam fir, and cedars such as Western red cedar and Alaska yellow cedar), and turfgrasses (including but not limited to annual bluegrass, annual ryegrass, Canadian bluegrass, bog grass, bentgrass, wheatgrass, Kentucky bluegrass, timothy grass, ryegrass, redtop, Bermuda grass, St. Augustine grass, and Zoysia grass).

[0055] Identification and selection of microorganisms that enhance plant responses to pests or pathogens. This specification also provides a method for identifying microorganisms that enhance a plant's response to one or more pathogens or pests, wherein the microorganism is not the pathogen of the plant, and the plant response is enhanced by the increased production in the plant of one or more plant defense compounds derived from anthranilates. In some embodiments, a method for identifying microorganisms that enhance a plant's response to one or more pathogens or pests includes the steps of: treating a plant, a part of a plant, or a seed of a plant with at least a first microbial strain that is not the pathogen of the plant to obtain treated seeds and / or a treated plant; growing the treated plant in the presence of the pathogen or pest, or growing a plant from a part of a treated plant or a treated seed; taking one or more tissue samples from the plant and from an untreated control plant, wherein the tissue samples are taken during a growth stage while the pest or pathogen is attacking the plant tissue; and assaying the samples to identify a microorganism that provides increased production of one or more plant defense compounds derived from anthranilates. In some embodiments, such a method includes the additional step of selecting a sample for analysis of the level of one or more plant defense compounds from a treated plant, which shows reduced damage from the pathogen or pest compared to an untreated control plant, or reduced damage from the plant pathogen or pest compared to another plant treated with the microorganism.

[0056] In some embodiments, a method for selecting a microbial strain that enhances a plant's response to a pathogen or pest includes the steps of: (i) assaying one or more tissue samples from a plant treated with at least a first microbial strain that is not a plant pathogen, or from a plant grown from part of a plant or seeds treated with the first microbial strain, for increased levels of one or more plant defense compounds derived from anthranilates compared to one or more control tissue samples from an untreated control plant, wherein the tissue samples are taken from the treated and untreated plants while the pest or pathogen is attacking the plant tissue or after it has attacked; and (ii) selecting a microbial strain that is not a plant pathogen and provides increased levels of one or more plant defense compounds, thereby selecting a microbial strain that enhances a plant's response to a pathogen or pest.

[0057] In some embodiments, plant tissue extracts or plant parts, such as germinated seedlings, are treated and assayed in vitro, for example, in a culture dish or test tube. In some embodiments, a method for selecting a microbial strain that enhances a plant's response to a pathogen or pest includes: (i) assaying one or more plant tissue samples treated with at least a first microbial strain that is not a pathogen of the plant, compared to one or more untreated control plant tissue samples, wherein the tissue samples were exposed to the pest or pathogen during or after treatment with the first microbial strain; and (ii) selecting a microbial strain that provides an increased level of one or more plant defense compounds, thereby selecting a microbial strain that enhances a plant's response to the pathogen or pest.

[0058] Additional methods may also be used to identify microorganisms that enhance a plant's response to one or more pathogens or pests, where the microorganism is not the pathogen of the plant, and the plant response is enhanced by gene expression in the microorganism. In certain embodiments, such methods may include subjecting a sample to nucleic acid analysis and determining that the sample contains nucleic acids expressing one or more proteins involved in the production of proteins that enhance a plant's response to pathogens or pests. In some embodiments, microorganisms are identified as having one or more genes encoding proteins involved in the biosynthesis of polyketides. In some embodiments, the genes encoding one or more polyketide synthesis proteins are encoded by sequences present in SEQ ID NO: 86, SEQ ID NO: 87, or variants thereof. Variants of SEQ ID NO: 87 include (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NO: 87, and / or (ii) a DNA sequence encoding the protein encoded by SEQ ID NO: 87. Variants of SEQ ID NO: 86 include (i) a DNA sequence having at least 80%, 85%, 90%, 95%, 98%, or 99% sequence identity to SEQ ID NO: 86, and / or (ii) a DNA sequence encoding the protein encoded by SEQ ID NO: 86. In some embodiments, the gene encoding the polyketide synthesis protein is a homolog or ortholog of the polyketide synthesis protein expressing the sequence present in SEQ ID NO: 86 and / or SEQ ID NO: 87. In some embodiments, the polyketide biosynthesis protein coding sequence identified in microorganisms encodes a protein having at least 80% identity to the protein having the sequences of SEQ ID NOs: 21–35. In some embodiments, such a polyketide biosynthesis protein coding sequence has at least 70% identity to the polyketide biosynthesis protein coding sequences of SEQ ID NOs: 36–50.In some embodiments, the polyketide biosynthesis protein has the sequence of SEQ ID NO: 21 or has approximately 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99%, or 100% sequence identity to SEQ ID NO: 21. Nucleic acid analysis for identifying such sequences includes, but is not limited to, sequencing-based techniques, using BLAST to compare sequences, nucleic acid hybridization, polymerase chain reaction (PCR), mass spectrometry, nanopore-based detection, branched DNA analysis, and combinations thereof. In some embodiments, nucleic acid analysis is used to identify 10 per gram of sample. 3 , 10 4 , 10 5 , 10 6 Microbial strains present at or above concentrations can be detected. Target samples for identifying microbial strains that enhance a plant's response to pathogens or pests include various water sources, such as soil samples, plants, plant parts, residual plant material, and water from paddy fields or crop irrigation.

[0059] Various methods can be used to treat plants with microorganisms and identify or select one or more microorganisms that enhance the plant's response to a target pathogen or pest. Such methods may include, but are not limited to, spraying, coating, partially coating, dipping, and / or absorbing plants, parts of plants, or seeds with one or more microorganisms or compositions containing such microorganisms. Compositions containing microorganisms for application to plants may be, but are not limited to, aqueous or non-aqueous liquids, dry compositions, or emulsions. In certain embodiments, plant seeds or cuttings may be dipped and / or absorbed in a composition containing a microbial strain. In certain embodiments, seed absorption and / or dipping may be carried out with gentle agitation. Seed treatment may be achieved using both continuous and / or batch seeding machines. In certain embodiments, coated seeds may be prepared by slurring seeds with a coating composition containing a microbial strain. Alternatively, microbial strains may be applied to the soil or other growing medium in which the plants are cultivated. Soil treatment or application may include, but is not limited to, in-furrow application (e.g., before, during, and / or after seeding), soil drenching, and distribution of granular or other dry formulations into the soil (e.g., before, during, and / or after seeding or plant growth). Treatment of plants grown in hydroponic systems may include pre-germination seed treatment, foliar application to germinated plants or parts thereof, and application in liquid solutions used in the hydroponic system.

[0060] Plants, plant parts, or seeds for use in screening methods to identify microorganisms that enhance plant responses to target pathogens or pests may be any plant that is attacked and damaged by a target pest or pathogen, or plant parts or seeds from such a plant. Plants may include, but are not limited to, crop plants, leafy vegetable plants, fruiting plants or trees, ornamental plants, turfgrasses, or trees cultivated for commercial production, such as conifers or nut trees. Such plant species include, but are not limited to, maize, soybeans, Brassicaceae or Brassica genus vegetables (e.g., B. napus, B. rapa, B. juncea), alfalfa, rice, rye, wheat, barley, oats, sorghum, millet (e.g., pearl millet (Pennisetum glaucum), foxtail millet (Panicum miliaceum), proso millet (Setaria italica), and finger millet (Eleusine coracana)), sunflowers, safflower, carrots, peppers, tomatoes, pumpkins, cucumbers, melons, and other gourds, beans, peas, chickpeas, lentils, tobacco, potatoes, peanuts, cotton, berries, grapes, kiwis, mangoes, papayas, pineapples, bananas, species of hemp (including, but not limited to, Cannabis sativa and industrial hemp varieties), sweet potatoes (Ipomoea This includes batatus, cassava, coffee, coconut, ornamental plants (including but not limited to azaleas, hydrangeas, hibiscus, roses, tulips, daffodils, petunias, carnations, poinsettias, and chrysanthemums), conifers (including but not limited to pines such as loblolly pine, slash pine, Ponderosa pine, Contorta pine, and Monterey pine, fir such as Douglas fir, American hemlock, Sitka spruce, redwood, European fir and balsam fir, and cedars such as Western red cedar and Alaska yellow cedar), and turfgrasses (including but not limited to annual bluegrass, annual ryegrass, Canadian bluegrass, bog grass, bentgrass, wheatgrass, Kentucky bluegrass, timothy grass, ryegrass, redtop, Bermuda grass, St. Augustine grass, and Zoysia grass).

[0061] In some embodiments, the microorganisms used to treat plants, parts of plants, or plant seeds in an assay for identifying microorganisms that enhance the plant response to a target pathogen or pest are beneficial microorganisms that provide additional benefits to the treated plants. In some embodiments, the beneficial microorganisms are bacterial strains. Bacterial strains that can be used in the assays described herein for identifying microorganisms that enhance the plant response to a target pathogen or pest include Actinomycetes, Agrobacterium, Arthrobacter, Alcaligenes, Aureobacterium, Azobacter, Azorhizobium, Azospirillum, Azotobacter, Beijerinckia, Bacillus, Brevibacillus, Burkholderia, Chromobacterium, Clostridium, Clavibacter, Commonas, Corynebacterium, Curtobacterium, Enterobacter, Flavobacterium, Gluconacetobacter, Gluconobacter, and Herbaspirill This includes, but is not limited to, non-pathogenic bacterial strains of the genera *Lysinibacillus*, *Hydrogenophage*, *Klebsiella*, *Luteibacter*, *Lysinibacillus*, *Mesorhizobium*, *Methylobacterium*, *Methylorubrum*, *Microbacterium*, *Ochrobactrum*, *Paenibacillus*, *Pantoea*, *Pasteuria*, *Phingobacterium*, *Photorhabdus*, *Phyllobacterium*, *Pseudomonas*, *Rhizobium*, *Rhodococcus*, *Bradyrhizobium*, *Serratia*, *Sinorhizobium*, *Sphingomonas*, *Streptomyces*, *Stenotrophomonas*, *Variovorax*, *Xanthomonas*, and *Xenorhadbus*. In some embodiments, the beneficial microorganism is a fungal strain.In some embodiments, the fungal strains tested in assays to identify enhanced plant responses to attacks by pathogens or pests are non-pathogenic strains of the genera Acremonium, Alternaria, Ampelomyces, Aspergillus, Aureobasidium, Beauveria, Botryosphaeria, Cladosporium, Cochliobolus, Colletotrichum, Coniothyrium, Embellisia, Epicoccum, Fusarium, Gigaspora, Gliocladium, Glomus, Laccaria, Metarhisium, Muscodor, Nigrospora, Paecilonyces, Paraglomus, Penicillium, Phoma, Pisolithus, Podospora, Rhizopogon, Scleroderma, Trichoderma, Typhula, Ulocladium, or Verticillium.

[0062] In some embodiments, microorganisms that enhance the plant response to pathogens or pests enhance the induced systemic resistance (ISR) response in plants. Some plant-beneficial microorganisms, such as plant growth-promoting rhizosphere bacteria (PGPRs), are capable of inducing broad-spectrum induced systemic resistance (ISR). Some PGPR strains induce responses similar to pathogen-induced acquired systemic resistance (SAR). In some embodiments, one or more genes involved in inducing the ISR response are jasmonic acid-dependent defense genes. In some embodiments, genes involved in inducing the ISR response are identified in microorganisms and encode siderophores. In some embodiments, one or more gene clusters contributing to ISR generation in treated plants are involved in the biosynthesis of metabolites. In some embodiments, gene clusters contributing to ISR generation are polyketide synthase gene clusters, siderophore gene clusters, non-ribosomal peptide synthases, or a combination thereof.

[0063] In some embodiments, a method for identifying or selecting a microbial strain that enhances a plant's response to a pathogen or pest described herein includes the step of cultivating a plant treated in the presence of the plant's pathogen or pest. In other embodiments, a plant or part of a plant is exposed to the pest or pathogen in vitro, for example, in a culture medium. The plant can be exposed to the pest or pathogen in any plant growth medium, including, but not limited to, soil, liquids such as hydroponic media, and nutrient culture media such as those used for plant tissue culture or micropropagation. Thus, in some embodiments, the treated part of a plant may be part of a plant for use in micropropagation, such as stem tips, anthers, petals, pollen, and other plant tissues, or it may be undifferentiated plant tissue such as germinating seedlings or callus.

[0064] The target pathogens or pests for use in the assay methods described herein include bacterial and fungal pathogens, as well as insect pests. Fungal pathogens include strains of Alternaria, Ascochyta, Aspergillus, Bipolaris, Botrytis, Bremia, Cercospora, Cochliobolus, Colletotrichum, Diplodia, Erysiphe, Exserohilum, Fusarium, Gaeumanomyces, Macrophomina, Magnaporthe, Nectria, Peronospora, Phakopsora, Phialophora, Phoma, Phymatotrichum, Phytophthora, Plasmopara, Puccinia, Podosphaera, Pyrenophora, Pyricularis, Pythium, Rhizoctonia, Sclerotium, Sclerotinia, Septoria, Stagonospora, Thielaviopsis, Uncinula, Ustilago, Venturia, and Verticillium. The target bacterial pathogens for use in the screening methods provided herein include strains of Pseudomonas, Ralstonia, Agrobacterium, Xanthomonas, Erwinia, Xylella, Dickeya, Pectobacterium, Clavibacter, and Candidatus. The target insects for use in the described screening methods may be any insect that causes damage to plants, including, but are not limited to, leafhoppers, thrips, aphids, snails, slugs, caterpillars, burrowing insects, burrowing insects, armyworms, grubs, and beetles. Most insect pests have four distinctly different developmental stages: egg, larva, pupa, and adult. Spiny and chewing insects have three developmental stages: egg, nymph, and adult. In some embodiments of the screening methods disclosed herein, the insect pests are applied to plants to enable screening for an enhanced response of the plants to the pests. Pests can be applied at any stage of growth where pathogens attack and damage plants.In some embodiments, the target pest is naturally present in the plant growth medium, for example, when the plant is cultivated in field soil. In some embodiments of the screening methods described herein, the target insect is applied to a plant, a part of a plant, or a plant culture medium at the larval stage. In some embodiments of the screening methods described herein, the target insect pest is a maize rhizome worm (CRW), a member of the broad genus Diabrotica.

[0065] A method for identifying microbial strains that enhance a plant's response to a pathogen or pest, as described herein, further comprises the step of taking one or more tissue samples from a treated plant. In some embodiments, the tissue samples are taken at a growth stage of the plant, plant part, or plant tissue where the target pest or pathogen is attacking the plant, plant part, or plant tissue. The tissues to be sampled are selected based on the target pest and may include leaves, stems, shoots, flowers, fruits, buds, roots, tubers, rhizomes, runners, bulbs, corms, sprouting seedlings, and callus tissue.

[0066] The methods for identifying microbial strains that enhance a plant's response to a pathogen or pest, as described herein, further include the step of assaying one or more isolated tissue samples to identify increased production of anthranilate-derived plant defense compounds, and / or increased production of transcripts encoding proteins associated with anthranilate production and / or conversion of anthranilate to plant defense compounds. In some embodiments, samples from treated plants grown in the presence of a target pathogen or pest are assayed to identify increased levels of one or more gene transcripts associated with anthranilate production and / or conversion of anthranilate to plant defense compounds, compared to levels of such transcripts in control samples from untreated plants or from plants treated with microorganisms that do not enhance a plant's response to a target pathogen or pest. Assays of gene transcript levels include RNA extraction and quantitative RNA-seq analysis, microarray analysis, high-throughput sequencing, etc. In some embodiments, samples are assayed using metabolomics approaches that capture volatile and non-volatile metabolites to identify plants having increased levels of one or more plant defense compounds. In some embodiments, microorganisms that enhance a plant's response to a target pathogen or pest are identified by the presence of increased levels of gene transcripts encoding anthranilate synthase alpha or beta subunit protein components, compared to levels of such transcripts from untreated plants or from plants treated with microorganisms that do not enhance the plant's response to the pathogen or pest. In some embodiments, the increased level of transcripts encoding the beta subunit component indicates a microorganism that enhances the plant's response to the target pathogen or pest.In some embodiments, microorganisms that enhance a plant's response to a target pathogen or pest are identified by the presence of increased levels of gene transcripts encoding enzymes involved in the synthesis of one or more anthranilate-derived plant defense compounds in a sample from treated plants, compared to such transcripts in untreated plants or from plants treated with microorganisms that do not enhance the plant's response to the pathogen or pest. In some embodiments, the plant defense compound is an anthranilate-derived phytoalexin. In some embodiments, increased levels of transcripts encoding anthranilate N-benzoyltransferase indicate microorganisms that improve the plant's response to attack by pathogens or pests. In some embodiments, a sample is assayed to determine increased expression of one or more gene transcripts encoding proteins having sequences of SEQ ID NO: 2, SEQ ID NO: 4, SEQ ID NO: 6, and / or SEQ ID NO: 7.

[0067] The target pathogens or pests for use in the assay methods described herein include bacterial and fungal pathogens, as well as insect pests. Fungal pathogens include strains of Alternaria, Ascochyta, Aspergillus, Bipolaris, Botrytis, Bremia, Cercospora, Cochliobolus, Colletotrichum, Diplodia, Erysiphe, Exserohilum, Fusarium, Gaeumanomyces, Macrophomina, Magnaporthe, Nectria, Peronospora, Phakopsora, Phialophora, Phoma, Phymatotrichum, Phytophthora, Plasmopara, Puccinia, Podosphaera, Pyrenophora, Pyricularis, Pythium, Rhizoctonia, Sclerotium, Sclerotinia, Septoria, Stagonospora, Thielaviopsis, Uncinula, Ustilago, Venturia, and Verticillium. The target bacterial pathogens for use in the screening methods provided herein include strains of Pseudomonas, Ralstonia, Agrobacterium, Xanthomonas, Erwinia, Xylella, Dickeya, Pectobacterium, Clavibacter, and Candidatus. The target insects for use in the screening methods described herein may be any insect that causes damage to plants, including, but are not limited to, leafhoppers, thrips, aphids, snails, slugs, caterpillars, burrowing insects, burrowing insects, armyworms, grubs, and beetles. In some embodiments of the screening methods described herein, the plant is maize, and the target insect pest is maize rhizome worm (CRW), a member of the broad genus Diabrotica.

[0068] Composition for treating plants In certain embodiments of any of the methods identified herein, a composition for treating plants to improve their response to attacks by pests and / or pathogens comprises a microorganism that enhances the plant response by increasing the production of one or more plant defense compounds derived from anthranilates and at least one additional component. In other embodiments, the composition comprises a microorganism that enhances the plant response by increasing the production of one or more plant defense compounds and at least one additional component. In some embodiments, the additional component may be an additional active ingredient, e.g., a pesticide or a second biological formulation. In certain embodiments, the pesticide may be an insecticide, fungicide, herbicide, nematicide, or other biocide. Non-limiting examples of insecticides and nematicides include carbamates, diamides, macrocyclic lactones, neonicotinoids, organophosphates, phenylpyrazoles, pyrethrins, spinosine, synthetic pyrethroids, tetronic acid, and tetramic acid. In certain embodiments, insecticides and nematicides include abamectin, aldicarb, aldoxycarb, bifenthrin, carbofuran, chlorantraniliporle, clothianidin, cyfluthrin, cyhalothrin, cypermethrin, deltamethrin, dinotefuran, emamectin, ethiprole, phenamifos, fipronil, flubendiamide, fostiazate, imidacloprid, ivermectin, lambdacyhalothrin, milbemectin, nitenpyram, oxamyl, permethrin, thioxazafen, spinetram, spinosad, spirodiclofen, spirotetramat, tefluthrin, thiacloprid, thiamethoxam, and thiodicarb. Useful fungicides, though not limited to specific examples, include aromatic hydrocarbons, benzimidazoles, benzthiadiazoles, carboxamides, carboxylic acid amides, morpholines, phenylamides, phosphonates, quinone external inhibitors (e.g., strobilurin), thiazolidinedions, thiophanates, thiophenecarboxamides, and triazoles.Specific examples of fungicides include acibenzoral-S-methyl, azoxystrobin, venalaxyl, bixafen, boscalid, carbendazim, cyproconazole, dimethomorph, epoxyconazole, fluopyram, fluoxastrobin, fluthianil, flutolanil, fluxapyroxad, fosetyl-Al, ipconazole, isopyrazam, kresoxim-methyl, mephenoxam, metalaxyl, metconazole, mycrobutanil, orysastrobin, penflufen, penthiopyrad, picoxystrobin, propiconazole, prothioconazole, pyraclostrobin, sedaxane, silthiofam, tebuconazole, tifluzamide, thiophanate, tolclofos-methyl, trifloxystrobin, and triticonazole. Non-exclusive examples of other biocides include, for example, isothiazolinone, 1,2-benzothiazolinone-3-one (BIT), 5-chloro-2-methyl-4-isothiazolinone-3-one (CIT), 2-methyl-4-isothiazolinone-3-one (MIT), octylisothiazolinone (OIT), dichlorooctylisothiazolinone (DCOIT), and butylbenzisothiazolinone (BBIT), 2-bromo-2-nitropropane-1,3-diol (Bronopol), 5-bromo-5-nitro-1,3-dioxane (Bronidox), tris(hydroxymethyl)nitromethane, 2,2-dibromo-3-nitrilopropionamide (DBNPA), and alkyldimethylbenzylammonium chloride. Non-exclusive examples of herbicides include ACCase inhibitors, acetanilide, AHAS inhibitors, carotenoid biosynthesis inhibitors, EPSPS inhibitors, glutamine synthase inhibitors, PPO inhibitors, PS II inhibitors, and synthetic auxins. Specific examples of herbicides include acetochlor, cretodym, dicamba, flumioxazine, fomesafen, glyphosate, glufosinate, mesotrione, quizalopop, saflufenacil, sulcotrione, and 2,4-D.

[0069] In some embodiments, the compositions or methods disclosed herein may include additional active ingredients selected from the group consisting of clothianidin, ipconazole, imidacloprid, metalaxyl, mefenoxam, thioxazafen, azoxystrobin, thiomethoxam, fluopyram, prothioconazole, pyraclostrobin, and sedaxane.

[0070] In some embodiments, the second biological preparation may be an additional beneficial microorganism, a microbial extract, a plant extract, a yeast extract, plant chitosan, a natural product, a plant growth activator, or a plant defense agent. Non-limiting examples of the second biological preparation may include bacteria, fungi, beneficial nematodes, and viruses. In certain embodiments, the second biological preparation is a Methylobacterium or Methylorubrum strain, including but not limited to the strains listed in Table 1. In some embodiments, the plant is treated with metanotropic bacteria. The metanotrophic bacterial strains useful in the compositions and methods described herein include bacterial species from genera selected from the group consisting of Methyloacidimicrobium, Methyloacidiplilum, Methylobacter, Methylocaldum, Methylocapsa, Methylocella, Methylococcus, Methylocystis, Methyloferula, Methylogaea, Methyloglobus, Methylohalobius, Methylomagnum, Methylomarinum, Methylomicrobium, Methylomonas, Methyloparacoccus, Methyloperedens, Methyloprofundus, Methylosarcina, Methylosinus, Methylosoma, Methylosphaera, Methylothermus, and Methylovulum. In some embodiments, the metanotrophic strain is a species of Methylobacter, Methylocystis, Methylomicrobium, Methylomonas, Methylosarcina, or Methylosinus. In some embodiments, the metanotroph is a strain of Methylomicrobium lacus, Methylosarcina fibrata, Methylosinus trichosporium, Methylosinus sporium, Methylocystis rosea, Methylocystis parvus, or Methylocystis hirsuta.In some embodiments, Methylomicrobium lacus is the deposited strain NRRL B-68261. In some embodiments, Methylocystis hirsuta is the deposited strain NRRL B-68262. In some embodiments, the Methylocystis species is a deposited strain selected from NRRL B-68282, NRRL B-68283, NRRL B-68284, NRRL B-68285, NRRL B-68286, NRRL B-68319, NRRL B-68321, NRRL B-68323, and NRRL B-68347. In some embodiments, the Methylosarcina species is the deposited strain NRRL B-68281. In some embodiments, the Methylosinus genus is a deposited strain selected from NRRL B-68320, NRRL B-68322, and NRRL B-68348.

[0071] In certain embodiments, the second biological formulation is Actinomycetes, Agrobacterium, Arthrobacter, Alcaligenes, Aureobacterium, Azobacter, Azorhizobium, Azospirillum, Azotobacter, Beijerinckia, Bacillus, Brevibacillus, Burkholderia, Chromobacterium, Clostridium, Clavibacter, Commonas, Corynebacterium, Curtobacterium, Enterobacter, Flavobacterium, Gluconacetobacter, Gluconobacter, Herbaspiri llum, Hydrogenophage, Klebsiella, Luteibacter, Lysinibacillus, Mesorhizobium, Microbacterium, Ochrobactrum, Paenibacillus, Pantoea, Pasteuria, Phingobacterium, Photorhabdus, Phylloba The bacteria may be of the genera cterium, Pseudomonas, Rhizobium, Rhodococcus, Bradyrhizobium, Serratia, Sinorhizobium, Sphingomonas, Streptomyces, Stenotrophomonas, Variovorax, Xanthomonas, and Xenorhadbus. In certain embodiments, the bacteria are selected from the group consisting of Bacillus amyloliquefaciens, Bacillus cereus, Bacillus firmus, Bacillus lichenformis, Bacillus pumilus, Bacillus sphaericus, Bacillus subtilis, Bacillus thuringiensis, Chromobacterium suttsuga, Pasteuria penetrans, Pasteuria usage, and Pseudomona fluorescens.

[0072] In certain embodiments, the second biological preparation may be a fungus belonging to the genera Acremonium, Alternaria, Ampelomyces, Aspergillus, Aureobasidium, Beauveria, Botryosphaeria, Cladosporium, Cochliobolus, Colletotrichum, Coniothyrium, Embellisia, Epicoccum, Fusarium, Gigaspora, Gliocladium, Glomus, Laccaria, Metarhisium, Muscodor, Nigrospora, Paecilonyces, Paragalus, Penicillium, Phoma, Pisolithus, Podospora, Rhizopogon, Scleroderma, Trichoderma, Typhula, Ulocladium, and Verticilium. In certain embodiments, the fungus is Beauveria bassiana, Coniothyrium minitans, Gliocladium vixens, Muscodor albus, Paecilomyces lilacinus, or Trichoderma polysporum. In certain embodiments, the composition comprises several additional biological components comprising a population including any combination of the above-mentioned bacteria or fungal genera or species.

[0073] In further embodiments, the second biological formulation may include, but is not limited to, various Bacillus, Pseudomonas, Coniothyrium, Pantoea, Streptomyces, and Trichoderma species. The microbial biopesticide may be a bacterium, fungus, virus, or protist. Particularly useful biopesticide microorganisms include various Bacillus subtilis, Bacillus thuringiensis, Bacillus pumilis, Pseudomonas syringae, Trichoderma harzianum, Trichoderma virens, and Streptomyces lydicus strains. Other microorganisms added may be genetically modified isolates or wild-type isolates available as pure cultures. In certain embodiments, it is expected that the second biological formulation may be provided in the composition in the form of spores. In further embodiments, the second biological formulation may be a plant growth activator or plant defense agent comprising, but not limited to, a biostimulant including, seaweed extract or humic acid salt, harpin, Reynoutria sachalinensis, jasmonic acids, lipochitooligosaccharides, and isoflavones.

[0074] In some embodiments for plant treatment, microbial inoculation is performed as a dry powder or particles into a non-aqueous continuous phase containing a non-aqueous solvent (e.g., a non-aqueous miscible solvent). In some embodiments, the microbial inoculation is uniformly dispersed in the non-aqueous continuous phase. Such compositions include additional components to enhance the mixing of the microbial inoculant with the aqueous composition containing the pesticide and / or to enhance the stability of the microbial inoculation in such aqueous composition. For non-limiting examples of components useful in such compositions, see, for example, U.S. Patent Publication US2023 / 0337681, which is incorporated herein by reference in its entirety. Dry microbial powders for use in such compositions can be produced using a variety of methods, including but not limited to spray drying, freeze-drying, air drying, fluidized bed drying, electrospray drying, or other drying methods. For non-limiting examples of methods for preparing dry microbial compositions, see, for example, U.S. Patent Publication US2022 / 0312772, which is incorporated herein by reference in its entirety.

[0075] In certain embodiments, compositions used to treat plants, plant parts, or plant seeds contain microbial strains that enhance the plant's defense response against pathogens and / or pests, as well as agriculturally acceptable excipients or agriculturally adjuvants. Agriculturally acceptable excipients include, but are not limited to, wood flour, clay, activated carbon, diatomaceous earth, fine-particle inorganic solids, and calcium carbonate. Clays and inorganic solids that can be used include, but are not limited to, calcium bentonite, kaolin, pottery clay, talc, perlite, mica, vermiculite, silica, quartz powder, montmorillonite, and mixtures thereof. Agriculturally acceptable excipients also include talc, graphite, polyethylene wax-based powders (such as Fluency Agent), protein powders, e.g., soy protein powder, or combinations of protein powder and lipids, e.g., lecithin or vegetable oil. In some embodiments, compositions include additional components to facilitate or enhance long-term storage and / or stability as a dry composition. In some embodiments, the long-term stability of the dried powder and / or treated seeds is enhanced compared to other compositions (e.g., compositions lacking components that facilitate long-term storage and / or stability, including stability on a portion of the treated plant or seeds). Additional components that can facilitate or enhance long-term storage and / or stability may include, but are not limited to, one or more oligosaccharides or polysaccharides. In some embodiments, the polysaccharides are selected from dextrins, maltodextrins, disaccharides, starches, chitosan, alginates, and gums including, but not limited to, karaya gum, jaguar gum, xanthan gum, glucomannan, tragacanth gum, konjac gum, polysaccharide gums, mucilage, gum arabic, and other natural gums. In some embodiments, any combination of the above or other agriculturally acceptable excipients and / or agriculturally acceptable adjuvants is used.Agriculturally acceptable adjuvants that promote adhesion to seeds include polyvinyl acetate, polyvinyl acetate copolymer, hydrolyzed polyvinyl acetate, polyvinylpyrrolidone-vinyl acetate copolymer, polyvinyl alcohol copolymer, polyether, polyvinyl methyl ether, polyvinyl methyl ether-maleic anhydride copolymer, wax, latex polymer, cellulose including ethylcellulose and methylcellulose, hydroxymethylcellulose, hydroxypropylcellulose, hydroxymethylpropylcellulose, polyvinylpyrrolidone, alginates, dextrin, maltodextrin, polysaccharides, fats, oils, and This includes, but is not limited to, proteins, karaya gum, jaguar gum, tragacanth gum, polysaccharide gums, mucus, gum arabic, shellac, vinylidene chloride polymers and copolymers, soy-based protein polymers and copolymers, lignosulfonates, acrylic copolymers, starch, polyvinyl acrylate, zein, gelatin, carboxymethylcellulose, chitosan, polyethylene oxide, acrylimide polymers and copolymers, polyhydroxyethyl acrylate, methylacrylimide monomer, alginates, ethylcellulose, polychloroprene, and syrups, or mixtures thereof. Other useful agriculturally acceptable adjuvants that can promote coating include, but are not limited to, polymers and copolymers of vinyl acetate, polyvinylpyrrolidone-vinyl acetate copolymers, and water-soluble waxes. Furthermore, agriculturally acceptable adjuvants also include a variety of lubricants such as talc, graphite, polyethylene wax-based powders (e.g., Fluency Agent), protein powders, e.g., soy protein powder, or combinations of protein powders and lipids, e.g., lecithin or vegetable oils (which can provide smooth flow and separation (unification) of seeds). Various surfactants, dispersants, anticoagulants, foaming control agents, and dyes disclosed herein and in U.S. Patent No. 8,181,388 can be adapted for use with compositions for treatment with microbial strains that enhance the plant response to pathogens and / or pests.In some embodiments, the dry composition comprises a Methylobacterium strain, as well as other components selected from the group consisting of maltodextrin, trehalose, glucomannan, soy protein, soy-based protein polymers and copolymers, talc, and graphite.

[0076] Methods of application for treating plants with microorganisms and additional components include spraying, coating, partially coating, dipping, drenching, and / or absorbing seeds, plants, or parts of plants with the composition. In certain embodiments, seeds and / or seedlings are exposed to the composition in the soil or other plant growing medium in which the plants or plants resulting from the seeds are cultivated. Examples of methods of application in which microbial strains are provided in the soil include furrow application, soil drenching, etc. In certain embodiments, the effective amount of one or more microbial strains provided in the treatment of seeds or parts of plants that provide an enhanced plant response to pathogens and / or pests is at least about 10 per seed or part of the treated plant. 3 , 10 4 , 10 5 , or 10 6 It is CFU.

[0077] Additional Embodiments In addition, the following non-limiting embodiments are included in this disclosure.

[0078] Embodiment 1. A method for improving a plant's response to a pathogen or pest, wherein the method is (i) modifying the plant genome to increase the expression of one or more gene transcripts involved in the production of one or more plant defense compounds derived from anthranilates in the plant, and / or treating the plant, part of the plant, or seeds with a microbial strain that is not pathogenic to the plant, wherein the treatment increases the expression in the plant of one or more gene transcripts involved in the production of plant defense compounds derived from anthranilates, and (ii) A method comprising growing the plant in the presence of the pathogen or pest, thereby increasing the level of one or more plant defense compounds in the plant compared to a control plant, and improving the plant's response to the pathogen or pest compared to a control plant, wherein the control plant is not modified or treated as in (i), and increasing the level of plant defense compounds produced in the plant by growing it.

[0079] 2. The method according to Embodiment 1, wherein the expression of one or more gene transcripts or polypeptides related to the production of anthranilates and / or the conversion of anthranilates to plant defense compounds is increased.

[0080] 3. The method according to Embodiment 2, wherein the gene transcript encodes an anthranylate synthase protein component, or the polypeptide is an anthranylate synthase protein component.

[0081] 4. The method according to Embodiment 3, wherein the anthranilate synthase protein component is an alpha or beta subunit.

[0082] 5. The method according to Embodiment 2, wherein the gene transcript encodes an anthranilate N-benzoyltransferase, or the polypeptide is an anthranilate N-benzoyltransferase.

[0083] 6. The method according to Embodiment 1, wherein the plant defense compound is an anthranilate ester.

[0084] 7. The method according to Embodiment 6, wherein the anthranilate ester is selected from the group consisting of methyl anthranilate, dimethyl anthranilate, ethyl anthranilate, phenylethyl anthranilate, and menthyl anthranilate.

[0085] 8. The method according to Embodiment 1, wherein the plant defense compound is an anthranilate-derived phytoalexin.

[0086] 9. The method according to any one of Embodiments 1 to 8, wherein the microbial strain is a bacterial strain.

[0087] 10. The method according to any one of Embodiments 1 to 9, wherein the microbial strain contains one or more genes encoding proteins involved in polyketide biosynthesis.

[0088] 11. The method according to Embodiment 10, wherein the one or more genes are located on a plasmid containing (i) one or more proteins having sequences having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to one or more of SEQ ID NOs: 21-35, (ii) one or more proteins having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to SEQ ID NOs: 21, (iii) one or more sequences from SEQ ID NOs: 36-50, and / or (iv) SEQ ID NOs: 86, or variants thereof having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% identity to SEQ ID NOs: 86.

[0089] 12. The method according to any one of Embodiments 9 to 11, wherein the bacterial strain is a Methylobacterium or Methylorubrum strain.

[0090] 13. The method according to any one of Embodiments 9 to 12, wherein the bacterial strain is a bacterial strain other than NLS0042 (NRRL B-50932) containing heterologous DNA from NLS0042 (NRRL B-50932) that confers a phenotype of increased expression in the plant of one or more gene transcripts involved in the production of plant defense compounds to a bacterial strain other than NLS0042, and optionally, the heterologous DNA encodes one of more proteins having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity with one or more of SEQ ID NOs: 21 to 35.

[0091] 14. The method according to Embodiment 13, wherein the bacterial strain other than NLS0042 is one of the bacterial strains shown in Table 1.

[0092] 15. The method according to any one of Embodiments 1 to 14, wherein the plant defense compound reduces foraging by insect larvae on plant roots compared to the control plant.

[0093] 16. The method according to Embodiment 3 or 4, wherein the anthranilate synthase protein component has the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO: 4.

[0094] 17. The method according to Embodiment 5, wherein the expression of the gene encoding an anthranilate N-benzoyltransferase containing the polypeptide sequence of SEQ ID NO: 6 or SEQ ID NO: 7 is increased compared to the control plant.

[0095] 18. The method according to any one of Embodiments 1 to 17, wherein the plant defense compound is not an indole derivative.

[0096] 19. The method according to any one of Embodiments 1 to 18, wherein the plant is a crop plant shown in Table 10, and / or the pathogen or pest is shown in Table 10 and / or 11, or the plant is a maize plant.

[0097] 20. A method for identifying microbial strains that enhance the plant's response to pathogens or pests, (i) to obtain treated seeds and / or treated plants by treating a plant, a part of a plant, or a seed of a plant with at least a first microbial strain that is not a pathogen of the plant, (ii) Growing the treated plant in the presence of the pathogen or pest, or growing a plant from a part of the treated plant or from treated seeds, (iii) Taking one or more tissue samples from the plant and from an untreated control plant, wherein the tissue samples are taken during or after the growth stage while the pest or pathogen is attacking the plant and the untreated control plant. (iv) A method comprising assaying the sample and identifying an increased production of one or more plant defense compounds derived from anthranilates in the treated plant compared to an untreated control plant, thereby identifying a microbial strain that enhances the plant's response to the pathogen or pest.

[0098] 21. A method for selecting a microbial strain that enhances a plant's response to a pathogen or pest, (i) Assaying one or more tissue samples from a plant treated with at least a first microbial strain that is not a plant pathogen, or from a plant grown from part or seeds of a plant treated with the first microbial strain, for increased levels of one or more plant defense compounds derived from anthranilates compared to one or more control tissue samples from an untreated control plant, wherein the tissue samples are collected from the treated and untreated plants while the pest or pathogen is attacking the plant tissue or after it has attacked. (ii) A method comprising selecting a microbial strain that provides the increased levels of the one or more plant defense compounds, rather than the plant pathogen, thereby enhancing the plant's response to the pathogen or pest.

[0099] 22. The method according to Embodiment 20 or 21, further comprising the step of selecting a sample for analysis from the treated plant that shows reduced damage from the pathogen or pest compared to the untreated control plant.

[0100] 23. The method according to Embodiment 20 or 21, wherein the sample is assayed to determine the level of one or more gene transcripts or polypeptides related to the production of anthranilates and / or the conversion of anthranilates to plant defense compounds.

[0101] 24. The method according to Embodiment 20, wherein the sample is assayed to determine the level of one or more plant defense compounds derived from anthranilates.

[0102] 25. The method according to Embodiment 20 or 21, wherein the pathogen or pest is a fungus, bacterium, nematode, insect, or virus.

[0103] 26. The method according to Embodiment 20, wherein a portion of the treated plant is selected from the group consisting of leaves, stems, shoots, flowers, fruits, buds, roots, tubers, rhizomes, runners, bulbs, and corms.

[0104] 27. The method according to Embodiment 20, wherein the collected tissue sample is selected from the group consisting of leaf, stem, shoot, flower, fruit, bud, root, tuber, rhizome, runner, bulb, and corm tissue sample.

[0105] 28. The method according to any one of embodiments 20 to 27, wherein the levels of one or more gene transcripts encoding anthranilate synthase protein components are determined in the treated and untreated tissues.

[0106] 29. The method according to Embodiment 28, wherein the anthranilate synthase protein component is an alpha or beta subunit.

[0107] 30. The method according to Embodiment 23, wherein the levels of the gene transcript encoding anthranilate N-benzoyltransferase are determined in the treated and untreated tissues.

[0108] 31. The method according to Embodiment 21 or 24, wherein the plant defense compound is an anthranilate ester.

[0109] 32. The method according to Embodiment 31, wherein the anthranilate ester is selected from the group consisting of methyl anthranilate, dimethyl anthranilate, ethyl anthranilate, phenylethyl anthranilate, and menthyl anthranilate.

[0110] 33. The method according to Embodiment 21 or 24, wherein the plant defense compound is an anthranilate-derived phytoalexin.

[0111] 34. The method according to any one of Embodiments 20 to 33, wherein the microbial strain is a bacterial strain or a fungal strain.

[0112] 35. The method according to Embodiment 34, wherein the bacterial strain is a Methylobacterium or Methylorubrum strain.

[0113] 36. The method according to any one of Embodiments 20 to 35, wherein the plant is selected from the group consisting of corn, wheat, rye, rice, alfalfa, barley, oats, rye, sorghum, millet, soybeans, Brassica, Hemp, tobacco, potatoes, peanuts, carrots, cotton, coffee, coconut, sugar beet, oats, barley, tomatoes, pumpkins, cucumbers, gourds, lettuce, peppers, peas, onions, green beans, sunflowers, safflower, sweet potatoes, cassava, coffee, coconut, conifers, turfgrass, leafy vegetables, microgreens, herbs, fruit trees and other fruit plants, and ornamental plants.

[0114] 37. The method according to embodiment 36, wherein the plant is corn.

[0115] 38. The method according to embodiment 36 or 37, wherein the tissue sample is a root sample.

[0116] 39. The method according to any one of embodiments 20 to 38, wherein the pest is a corn root worm.

[0117] 40. The first microbial strain is obtained by transferring DNA from a second microbial strain, which can confer resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion to or from the pest or pathogen by increasing the production of one or more plant defense compounds derived from anthranilates in treated plants, to a third microbial strain, which does not confer resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion to or from the pest or pathogen in plants treated with the third microbial strain, optionally the second microbial strain being NLS0042, and / or optionally the third microbial strain being a microbial strain other than NLS0042 provided in Table 1, optional The method according to any one of Embodiments 20 to 39, wherein, by selection, the imported DNA encodes one or more proteins having sequences with at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to one or more of SEQ ID NOs: 21 to 35, or (ii) is present on a plasmid containing SEQ ID NOs: 86, or a variant thereof having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to SEQ ID NOs: 86, and / or (iii) contains one or more sequences from SEQ ID NOs: 37 to 50, or a variant thereof having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% identity to one or more of SEQ ID NOs: 37 to 50.

[0118] 41. A method for monitoring pest or pathogen resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion in plants, which can confer said pest or pathogen resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion, and which involves at least one tissue sample from a plant treated with a first microbial strain that is not a pathogen of the plant, or from a plant grown from a portion or seeds of a plant treated with the first microbial strain, (i) Increased levels of one or more plant defense compounds derived from anthranilates, and / or (ii) Assaying for increased expression of one or more gene transcripts or polypeptides related to the production of anthranilates and / or the conversion of anthranilates to plant defense compounds, A method wherein the increased levels and / or increased expression are compared to one or more control tissue samples from an untreated control plant, the tissue samples being collected from the treated plant and the untreated control plant while the pest or pathogen is attacking the plant tissue or after the attack, thereby indicating that the increased levels of the one or more plant defense compounds, gene transcripts, and / or polypeptides in the tissue samples from the treated plant, compared to the control plant, indicate increased pest or pathogen resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion in the treated plant.

[0119] 42. The method according to Embodiment 41, wherein the treated plant does not exhibit increased levels of the one or more plant defense compounds, gene transcripts, and / or polypeptides, and the method further comprises retreatment with the first microbial strain and / or treatment with another biological control agent, insecticide, fungicide, or pesticide.

[0120] 43. A method for selecting a microbial strain that enhances a plant's response to a pathogen or pest, comprising: (i) assaying one or more tissue samples from a plant treated with at least a first microbial strain that is not a plant pathogen, or from a plant grown from part or seeds of a plant treated with the first microbial strain, for increased levels of one or more plant defense compounds derived from anthranilates compared to one or more control tissue samples from an untreated control plant, wherein the tissue samples are taken from the treated and untreated plants while the pest or pathogen is attacking the plant tissue or after it has attacked; and (ii) selecting a microbial strain that is not a plant pathogen and provides the increased levels of the one or more plant defense compounds, thereby selecting a microbial strain that enhances a plant's response to the pathogen or pest.

[0121] 44. A method for selecting a microbial strain that enhances a plant's response to a pathogen or pest, comprising: (i) assaying one or more plant tissue samples treated with at least a first microbial strain that is not a pathogen of the plant for increased levels of one or more plant defense compounds derived from anthranilates compared to one or more untreated control plant tissue samples, wherein the tissue samples were exposed to the pest or pathogen during or after treatment with the first microbial strain; and (ii) selecting a microbial strain that provides the increased levels of the one or more plant defense compounds, thereby selecting a microbial strain that enhances a plant's response to the pathogen or pest.

[0122] 45. A method for improving a plant's response to a pathogen or pest, wherein the method is (i) modifying the plant genome to increase the expression of one or more gene transcripts involved in the production of one or more plant defense compounds in the plant, and / or treating the plant, part of the plant, or seeds with a microbial strain that is not pathogenic to the plant, wherein the treatment increases the expression of one or more gene transcripts involved in the production of plant defense compounds in the plant, and (ii) A method comprising growing the plant in the presence of the pathogen or pest, thereby increasing the level of one or more plant defense compounds in the plant compared to a control plant, and improving the plant's response to the pathogen or pest compared to a control plant, wherein the control plant is not modified or treated as in (i), and increasing the level of plant defense compounds produced in the plant by growing it.

[0123] 46. ​​The method according to Embodiment 45, wherein the plant defense compound is derived from an anthranilate.

[0124] 47. A method for eliminating pathogens or pests, wherein the method is (i) treating soil, plants, parts of plants, or seeds with a non-pathogenic microbial strain, wherein the treatment increases the expression in the plant of one or more gene transcripts involved in the production of plant defense compounds. (ii) A method for increasing the level of plant defense compounds produced in a plant, comprising cultivating the plant in the presence of the pathogen or pest such that the level of one or more plant defense compounds in the plant increases compared to a control plant, the plant repels the pathogen or pest better than the control plant, and the control plant is not modified or treated as in (i).

[0125] 48. The method according to Embodiment 47, wherein the plant defense compound is derived from an anthranilate.

[0126] 49. The method according to Embodiment 47, wherein the microbial strain produces metabolites, and such metabolites increase the production of such plant defense compounds compared to the control plant.

[0127] 50. The method according to Embodiment 49, wherein the metabolite is selected from the group consisting of siderophores, non-ribosomal peptides, polyketides, or combinations thereof.

[0128] 51. The method according to Embodiment 50, wherein the microbial strain comprises one or more gene clusters encoding proteins involved in the biosynthesis of the metabolites.

[0129] 52. The method according to any one of Embodiments 47 to 51, wherein the microbial strain comprises one or more genes located on a plasmid containing (i) one or more proteins having a sequence having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to one or more of SEQ ID NOs: 21 to 35, (ii) one or more proteins having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to SEQ ID NOs: 21, (iii) one or more sequences from SEQ ID NOs: 36 to 50, and / or (iv) SEQ ID NOs: 86, or a variant having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% identity to SEQ ID NOs: 86, and optionally the plant is a crop plant shown in Table 10, and / or the pathogen or pest is shown in Table 10 and / or 11.

[0130] 53. A method for selecting a microbial strain that confers pest or pathogen resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion to a plant, comprising identifying in the microbial strain (i) one or more polynucleotides encoding a protein in the polyketide biosynthesis pathway, and / or (ii) one or more proteins in the polyketide biosynthesis pathway.

[0131] 54. The method according to Embodiment 53, wherein the polynucleotide is identified by detecting at least one polynucleotide present on a plasmid containing (i) one or more proteins having a sequence having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to one or more of the sequence numbers 21-35, (ii) one or more proteins having at least 80% sequence identity to sequence number 21, (iii) one or more sequences having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to one or more of the sequence numbers 36-50, and / or (iv) sequence number 86, or a sequence having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to sequence number 86.

[0132] 55. The method according to Embodiment 53 or 54, wherein the polynucleotide is identified by nucleic acid amplification, hybridization, and / or sequencing techniques.

[0133] 56. The method according to Embodiment 53 or 54, wherein the protein is identified by detecting (i) one or more proteins having a sequence having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity with one or more of SEQ ID NOs: 21 to 35, or (ii) one or more proteins having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity with SEQ ID NO: 21.

[0134] 57. The method according to Embodiment 56, wherein the protein is identified by immunoaffinity and / or mass spectrometry techniques.

[0135] 58. The method according to any one of Embodiments 53 to 57, further comprising the step(s) of isolating and / or culturing the identified microbial stain containing the polynucleotide and / or protein.

[0136] 59. The method according to any one of Embodiments 53 to 58, wherein the microbial strain comprises a gene that (i) encodes one or more proteins having a sequence having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to one or more of SEQ ID NOs: 21 to 35, (ii) encodes one or more proteins having at least 80% sequence identity to SEQ ID NOs: 21, or (iii) comprises one or more sequences from SEQ ID NOs: 36 to 50, or variants thereof having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to SEQ ID NOs: 36 to 50.

[0137] 60. The microbial strain (i) encodes one or more proteins having a sequence that has at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity with one or more of sequence codes 21-35; (ii) encodes one or more proteins having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity with sequence code 21; (iii) encodes one or more of sequence codes 36-50, or sequence The method according to any one of Embodiments 53 to 58, wherein a DNA gene is modified on a plasmid containing one or more sequences having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to any one of numbers 36 to 50, and / or (iv) Sequence ID No. 86, or a sequence having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to Sequence ID No. 86.

[0138] 61. A method for improving a plant's response to a pathogen or pest, wherein the method is (i) a plant, part of a plant, or seed encoding one or more proteins having a sequence that has at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to NLS0042, or (i) one or more sequences from sequence numbers 21-35; (ii) one or more proteins having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% sequence identity to sequence number 21; (iii) one or more sequences from sequence numbers 37-50. , or treatment with a DNA-containing microbial stain present on a plasmid containing a variant having at least 80%, 85%, 90%, 95%, 97%, 98%, or 99% identity to one or more of SEQ ID NOs. 37-50, and / or (iv) SEQ ID NOs. 86, 85%, 90%, 95%, 97%, 98%, or 99% identity to SEQ ID NOs. 86, (ii) A method comprising cultivating the plant or a plant grown from the seeds in the presence of the pathogen or pest such that the plant's response to the pathogen or pest is improved compared to a control plant, and the plant is not a maize plant.

[0139] 62. The method according to Embodiment 61, wherein the plant is selected from the group consisting of chili peppers, tomatoes, bayberries, Brassica plants, soybeans, cotton, and rice.

[0140] 63. The method according to Embodiment 61 or 62, wherein the level of one or more plant defense compounds in the plant is increased compared to a control plant.

[0141] 64. The method according to any one of embodiments 61 to 63, wherein the pathogen or pest is an insect pest selected from the group consisting of aphids, lepidoptera, stink bugs, whiteflies, sugar beetle, tobacco hawk moth, potato leafhopper, bean ladybug, flea beetle, fall armyworm, soybean inchworm, citrus thrips, and rice water weevil.

[0142] 65. The method according to any one of Embodiments 61 to 64, wherein the improved response of the treated plant or a plant grown from the treated part or seeds includes improved pest or pathogen resistance, tolerance, reduced damage, reduced infection, and / or reduced intrusion compared to an untreated control plant.

[0143] 66. The method according to any one of embodiments 61 to 65, wherein the plant is a crop plant shown in Table 10, and / or the pathogen or pest is shown in Table 10 and / or 11. [Examples]

[0144] Example 1: Analysis of upregulated maize genes in field trials Field trials were established to include a trapping crop to attract maize rhizome worms (CRW) or to continuously crop maize for several years to increase CRW pressure. Plants were either treated with Methylobacterium strain NLS0042 or left untreated (UNT) in the all-factor design. The maize varieties used in the trials were P1197AM and P1197AMXT, as described in the Pioneer 2022 Corn Hybrid-Herbicide Management Guide. ·Optimum® AcreMax® insect protection system with AM-YGCB, HX1, LL, and RR2. Includes a single-bag integrated refuge solution for above-ground insects. In EPA-designated cotton-growing counties, 20% of separate corn borer refuges must be planted with Optimum AcreMax products. ·AMXT (Optimum® AcreMax® XTreme) - Includes a single-bag integrated refuge solution for above- and below-ground insects. Major components include the Agrisure® RW trait, Bt trait, and Herculex® XTRA gene. In EPA-designated cotton-growing counties, 20% of separate corn borer refuges must be planted with Optimum AcreMax XTreme products. ·Methylobacterium in the form of spray-dried powder was applied to the seeds in the field at planting at a rate of at least 1×10 6 CFU per seed.

[0145] Roots were collected when CRW larval damage was at its peak. Root samples from each of the four treatments (NLS0042 on P1197AM maize, UNT on P1197AM maize, NLS0042 on P1197AMXT maize, and UNT on P1197AMXT maize) were excavated from the soil, as much mud as possible was removed, and the roots were immediately frozen on dry ice. The samples were returned to the laboratory, and the roots were pulverized under liquid nitrogen. RNA was extracted from the roots using the Qiagen RNeasy mini kit. The RNA was treated with DNase using DNase Max, and the quality was checked by Bioanalyzer. Quantseq (3' RNAseq) was performed using Lexogen (Greenland, NH). Using the Bluebee platform and the B73 maize genome as reference, samples exhibiting the desired CRW phenotype (NLS0042-treated plants with a lower node damage score (NIS) than UTC plants with the same genetic background, and UTC plants with a higher root damage score than NLS0042-treated plants with the same genetic background) were analyzed for differential expression (DE) genes. The NIS, also known as the Iowa root damage score, ranges from 0 to 3, where 0 is no damage and 3 is three nodes damaged within 2 inches of the stem (Oleson et al., 2005). For each of the four treatments, there were three samples for each condition. [Table 2]

[0146] The differentially expressed gene codes were compared with maize gene annotations v3-v5 obtained from MaizeGDB. Differentially expressed genes with significant p-values ​​after multiple comparison correction were manually searched for in addition to the annotation information. [Table 3]

[0147] Two of the nine genes upregulated beyond UNT by NLS0042 in AMXT maize were found to be associated with anthranilates in corn. The transcripts of anthranilate N-benzoyltransferase protein 2 and anthranilate synthase homolog 1 (AS beta subunit) were significantly increased in Methylobacterium-treated AMXT maize plants compared to untreated maize plants. Smaller increases in gene expression were also observed for transcripts encoding other anthranilate synthase protein components.

[0148] Example 2: Greenhouse assay results Maize plants were treated with NLS0042, formulated as a freeze-dried powder and applied as a seed treatment before sowing, as shown in Table 4, and compared to untreated seeds (UTC). An additional set of non-PPFM-treated seeds received jasmonic acid applied to seedlings as a foliar spray (1 mM concentration, sprayed until runoff) before infection with the larvae of the western maize rhizomatous insect Diabrotica virgifera, which served as a positive control for ISR. Two types of destructive harvests occurred in parallel sets of pots at 3, 6, and 9 days after larval invasion: 1) roots and soil were transferred to a Berlese funnel for live larval extraction, then counted, weighed, and individually measured for length; and 2) roots were removed and treated for RNA extraction and gene expression analysis. Parallel sets of seedlings were potted in 2-gallon pots and grown until near maturity, and root damage was assessed at the VT growth stage using the Iowa Root Damage Scale (NIS). [Table 4] [Table 5]

[0149] Larval survival, biomass, and length did not appear to be adversely affected by either seed treatment with Methylobacterium NLS0042 or seedling drenching with jasmonic acid. Both NLS0042 as seed treatment and jasmonic acid as foliar drenching showed a tendency toward reduced root damage compared to the untreated control, as measured via the Iowa node damage score (NIS), where 0 represents no larval root damage pressure and 3 represents extremely high root damage pressure (Table 5). Gene expression results comparing NLS0042-treated samples (NLS0042_UTC) and jasmonic acid-treated samples (UTC_JA) with untreated controls showed that, compared to untreated control plants, both NLS0042 seed-treated plants and jasmonic acid-irrigated plants showed high signals at 9DAI for jasmonic acid-regulating genes, namely ribosome inactivating protein 2 (rip2), terpene synthase (tps23), and maize protease inhibitor (mpi).

[0150] Example 3: Method for selecting microorganisms that enhance the maize plant response to CRW. Assays for identifying and / or selecting plant-beneficial microorganisms that enhance the response of maize plants to maize rhizome worm larvae are performed as follows: Maize seeds are treated with non-maize pathogenic microorganisms, e.g., strains of Methylobacterium or Methylorubrum, sown in growth medium, and grown to seedling stage V2–V4. The plants are removed from the growth medium and gently washed to remove the growth medium from the roots. The root system is placed in a petri dish lined with moist cellulose filter paper and allowed to invade with early-stage maize rhizome worm larvae. The larvae are allowed to feed on the maize roots and are then removed. Roots and / or root exudates as well as headspace are evaluated by genetic and / or metabolomics assays to determine the levels of anthranilate-derived plant defense compounds and / or gene transcripts associated with the production of anthranilate-derived plant defense compounds. Microorganisms that increase the production of anthranilate-derived plant defense compounds and / or increase the levels of gene transcripts associated with the production of anthranilate-derived plant defense compounds, compared to levels in untreated control plants, are selected as microorganisms that enhance the maize plant's response to maize rhizome insects.

[0151] Further steps for selecting samples for analysis include evaluating the insect feeding behavior of larvae before analyzing root samples. After feeding, larvae are gently removed from the roots using a paintbrush and positioned away from the roots. Larval behavior is tracked, and the time it takes to reach the root surface is recorded. Larval behavior using control roots from untreated plants is compared to that using roots from treated plants. The presence of chemical feeding inhibitors such as methyl anthranilate in the root exudate either prevents larvae from returning to the corn roots or causes them to take significantly longer to return to the treated plant roots compared to the roots of untreated control plants.

[0152] A selection assay for analyzing the insect feeding behavior of larvae is performed using roots treated with NLS0042 as described above, along with control roots, using the following treatment. 1) Untreated 10-day-old plants free from WCR damage 2) 10-day-old plants treated with NLS0042 that show no WCR damage. 3) Untreated 10-day-old plants with 3 days of WCR larval feeding pressure. 4) 10-day-old NLS0042-treated plants subjected to 3 days of WCR larval feeding pressure.

[0153] When the choice was between NLS0042-treated roots and untreated roots, the larvae overwhelmingly chose the untreated roots (Figure 1). This basic condition was tested in four of the six possible combinations of treatments and was only slightly important if the roots had been subjected to WCR larval feeding pressure prior to the selection test.

[0154] If the selection was between two NLS0042-treated roots, most larvae would not make a selection and instead remain in the central petri dish where they started (Figure 2). This means that these larvae are unable to feed.

[0155] When the choice lies between two untreated roots, the larvae slightly prefer the untreated root that had been subjected to WCR larval feeding pressure prior to the selection test and choose one of the untreated roots (Figure 3).

[0156] These results indicate that NLS0042-treated plants release volatile chemicals (or multiple chemicals) that suppress WCR larval feeding damage. These results are consistent with the WCR larval response to methyl anthranilate.

[0157] Example 4. Analysis of metabolite production by Methylobacterium The contribution of metabolites produced by Methylobacterium NLS0042 to reducing maize root damage caused by maize rhizome insects is investigated. Two biosynthetic gene clusters involved in metabolite production were identified in the NLS0042 genome. From each cluster, two genes, asbA, predicted to be involved in siderophore production, and bfmBAB, predicted to be involved in the biosynthesis of antimicrobial compounds, were selected for the generation of knockout mutants. Allelic exchange vectors for each gene target were assembled and transported into NLS0042 via conjugation and integration at the target site. Counter-selection and screening were performed to identify knockout mutants. PCR evidence supporting the generation of knockout mutants for both gene targets was obtained. Sequencing of regions expected to contain deletions confirmed the generation of the desired knockout mutants. The asbA and bfmBAB mutant strains are tested in the herbivore selection assay as described above to determine whether the mutations block the ability of Methylobacterium to repel CRW larval herbivory.

[0158] Example 5. Selective assay and identification of volatile substances produced by treated maize plants. An additional selection assay is performed using treated and untreated maize roots as described above. The treatments include 1) untreated seeds, 2) seeds treated with NLS0042, 3) seeds treated with NLS0042mut1, and 4) seeds treated with NLS0042mut2. Ten newborn larvae are placed in the center, and the larval positions are recorded in the dark after 5 hours. The following sets of treatments are evaluated in at least 10 replicates. Process 1 vs. Process 2 Process 1 vs. Process 3 Process 1 vs. Process 4 Process 2 vs. Process 3 Process 2 vs. Process 4 Process 3 vs. Process 4

[0159] Larval selection between treatments was analyzed, and the volatile compounds involved in selection were identified as follows:

[0160] Untreated non-Bt maize seeds are rinsed and incubated overnight in distilled water at 25°C to accelerate germination. Seeds are dried, weighed, and treated under a biosafety cabinet at a rate of 1E6cfu / seed according to protocol. Maize seeds are planted and allowed to germinate and grow for at least 10 days without larval feeding to develop priming activity characteristic of ISR and defensive volatile chemicals(s). After 10 days of growing, a vacuum is applied to extract volatile substances onto a solid substrate. The substrate is used to identify known plant volatiles using GC-MS with methyl anthranilate as a control compound.

[0161] The two treatments in this experiment are an untreated control and the original NLS0042 powder. The comparison is made between the volatile substances produced in the untreated plants 10 days after cultivation and those produced by the NLS0042-treated plants.

[0162] Example 6. Maize gene, sequence, and homolog [Table 6-1] [Table 6-2]

[0163] Example 7. Evaluation of insect feeding behavior of larvae before analysis of root samples. A larval feeding selection assay was performed essentially as described in Example 5. The behavior of larvae using control roots from untreated plants was compared with that of roots from treated plants, and the results are provided in Table 7. The presence of methyl anthranilate in the root exudate prevented larvae from returning to the maize roots or caused them to take a significantly longer time to return to the treated plant roots compared to the roots of the untreated control plants. [Table 7]

[0164] Example 8. Selective assay and identification of volatile substances produced by treated maize plants. Additional selection assays were performed using treated and untreated maize roots as described above. Treatments included 1) untreated seeds, 2) seeds treated with NLS0042, and 3) seeds treated with NLS0042mut2 (bfmBAB mutant). The NLS0042mut2 strain was obtained as described in Example 4. Ten neonatal larvae were placed in the center, and the larval positions were recorded after incubation in the dark. The results are shown in Table 8 below. The bfmBAB mutant significantly reduced the repellent effect of NLS0042, demonstrating that this gene contributes to the induction of the repellent effect of NLS0042 in maize plants. Other genes identified in the polyketide synthase pathway are similarly evaluated to identify similar effects on induced insect defense responses in maize plants. [Table 8]

[0165] Example 9. Microbial screening for polyketide biosynthesis sequences. The method for identifying microorganisms based on the presence of a sequence is as follows: The whole or partial genome sequence of a microorganism, or a metagenomic sequence from an environmental sample, is obtained. For isolated microorganisms or combinations of cultured microorganisms, the sequence of interest is identified using BLAST (Basic Local Alignment Search Tool, Altschul et al. 1990) or other nucleic acid analysis software. The sequence may be either a nucleotide or a protein sequence. Sequences can also be searched using annotations for genes, gene clusters, and / or protein domains. Alternatively, the microorganism of interest can be identified using standard PCR or qPCR primers specific to the sequence of interest. Environmental samples containing multiple characterized or uncharacterized microorganisms are also screened in this manner, and microorganisms with positive signals are purified from aliquots of the original sample. Colony PCR and / or aliquot dilution methods are used to identify positive microorganisms. The sequences used in the analysis are provided in Table 9. [Table 9]

[0166] In this way, sequences present on (i) sequences of sequence number 87, sequence number 86, and / or their variants, or (ii) sequences of sequence numbers 21-35, or sequences encoding one or more proteins having homologs or orthologues thereof, are identified. Microorganisms identified in this way are screened to identify microorganisms that enhance plant defense responses against pathogens and / or insect pests.

[0167] Example 10. Field test analysis of the effect of Methylobacterium strain NLS0042 on the invasion of special crops. Field tests will be conducted on peppers to determine the ability of NLS0042 to enhance plant defenses against aphids and caterpillars.

[0168] Field trials will be conducted on tomatoes to determine the ability of NLS0042 to enhance plant defenses against aphids and stink bugs.

[0169] Field trials will be conducted on bayberries to determine the ability of NLS0042 to enhance plant defenses against potato leafhoppers, bean beetles, and lepidopterans.

[0170] Field trials will be conducted on rapeseed species to determine the ability of NLS0042 to enhance plant defenses against the flea beetle.

[0171] Plants, plant parts, and / or seeds are treated with NLS0042 as a foliar spray, seed treatment, drenching (e.g., soil drenching), furrow treatment, or a combination thereof. The treated plants are grown in the presence of naturally occurring and / or artificially supplemented invasions of target insect pests. The plants are evaluated for yield and insect damage and compared to control plants to identify the enhanced plant response to pests resulting from treatment with NLS0042.

[0172] Example 11. Greenhouse tomato test to assess the effectiveness of NLS0042 against whiteflies. Tomato plants, parts of them, and / or seeds are treated with NLS0042 as a foliar spray, seed treatment, drenching, or a combination thereof. Whiteflies are inoculated into the treated plants and untreated control plants. Whitefly counts are performed weekly after inoculation and compared to counts in control plants not treated with NLS0042 to identify enhanced plant responses to whiteflies.

[0173] Example 12. Tomato test to assess the effect of NLS0042 on leafhoppers. Treated and untreated tomato plants are exposed in a greenhouse to leafhoppers carrying the Beet Curly Top Virus (BCTV) pathogen. NLS0042 is applied as a foliar spray, seed treatment, drenching, or a combination thereof. Treated plants are evaluated by digital PCR to determine the viral load. Treated and untreated controls are transplanted to field trials to determine the effect on plant vitality (hyperspectral imaging), yield, and fruit quality. The presence of insects is also determined using insect nets and / or sticky cards.

[0174] Example 13. Greenhouse experiment to evaluate the effect of NLS0042 on caterpillar pests in Solanaceae, soybean, cotton, and rice crops. Tomato, eggplant, pepper, soybean, cotton, and rice plants are treated with NLS0042 as a foliar spray, seed treatment, drenching, or a combination thereof. Pre-weighed tobacco hawk moth caterpillars are allowed to feed on treated and untreated control tomato plants at various phenological stages over a period of time. Pre-weighed soybean inchworm and fall armyworm caterpillars are allowed to feed on treated and untreated control soybean plants at various phenological stages over a period of time. Pre-weighed fall armyworm caterpillars are allowed to feed on treated soybean, cotton, and rice plants, as well as untreated control soybean, cotton, and rice plants, at various phenological stages over a period of time. During the treatment period of all experiments, data on caterpillar growth, mortality, volume, and developmental milestones were collected and analyzed to identify the enhanced plant response to insect pests resulting from treatment with NLS0042.

[0175] In the second experiment, artificial diets fortified with 10% leaf material from treated and untreated control plants are prepared. Sphinx moth, fall armyworm, and soybean inchworm caterpillars are allowed to feed on, develop, and complete their life cycles using these diets. Data on life history characteristics are collected to further evaluate the effect of NLS0042 treatment on plants against insect pests.

[0176] A selection assay is performed between treated and control plants to evaluate the effect of NLS0042 treatment on herbivore selection and to assess potential antibiotic effects.

[0177] Example 14. Greenhouse experiment to evaluate the effect of NLS0042 on thrips and aphid pests in cotton. Population assays are performed using a known number of citrus thrips or aphids. The thrips or aphids are allowed to feed on and develop in NLS0042-treated plants and untreated control plants, and their population growth is monitored over a long period. Electrophysiological experiments are also performed using a technique called electroosmography, which provides information on how NLS0042-treated plants and untreated control plants change in their antibiotic and repellent properties.

[0178] Selection assays are also performed between treated and control plants to evaluate the effect of NLS0042 treatment on herbivore selection and to assess potential antibiotic effects.

[0179] Example 15. Evaluation of the effects of microorganisms containing genes that induce plant defense responses against insects and / or pathogenic pests. Microbial strains containing one or more genes for the expression of one or more polyketide synthesis proteins having any one sequence from sequence numbers 21-35, or its homolog or orthologue, are identified by genome screening and / or selection as described herein. Alternatively, such strains are produced by the transfer of genes encoding the polyketide synthesis proteins identified herein by plasmid transfer and / or genetic transformation with recombinant constructs. The microbial strains are used to treat target plants in greenhouses, grow boxes, and / or field assays, and the plants are evaluated for enhanced plant defense responses against target pests. Treatments include foliar application, absorption or drenching, and seed treatment. Table 10 below shows the plants and pests to be evaluated. [Table 10-1] [Table 10-2] [Table 10-3] [Table 11]

Claims

1. A method for improving a plant's response to a pathogen or pest, wherein the method is (i) treating a plant, part of a plant, or seed with a non-pathogenic microbial strain such that the treatment increases the expression in the plant of one or more gene transcripts involved in the production of anthranilate-derived plant defense compounds, and / or modifying the plant genome to increase the expression in the plant of one or more gene transcripts involved in the production of anthranilate-derived plant defense compounds, and (ii) A method comprising growing the plant in the presence of the pathogen or pest, thereby increasing the level of one or more plant defense compounds in the plant compared to a control plant, and improving the plant's response to the pathogen or pest compared to a control plant, wherein the control plant is not modified or treated as in (i), and increasing the level of plant defense compounds produced in the plant by growing it.

2. The method according to claim 1, wherein the expression of one or more gene transcripts or polypeptides related to the production of anthranilates and / or the conversion of anthranilates to plant defense compounds is increased.

3. The method according to claim 2, wherein the gene transcript encodes an anthranylate synthase protein component, or the polypeptide is an anthranylate synthase protein component.

4. The method according to claim 3, wherein the anthranilate synthase protein component is an alpha or beta subunit.

5. The method according to claim 2, wherein the gene transcript encodes an anthranilate N-benzoyltransferase, or the polypeptide is an anthranilate N-benzoyltransferase.

6. The method according to claim 1, wherein the plant defense compound is an anthranilate ester.

7. The method according to claim 6, wherein the anthranilate ester is selected from the group consisting of methyl anthranilate, dimethyl anthranilate, ethyl anthranilate, phenylethyl anthranilate, and menthyl anthranilate.

8. The method according to claim 1, wherein the plant defense compound is a phytoalexin derived from an anthranilate.

9. The method according to claim 1, wherein the microbial strain is a bacterial strain.

10. The method according to claim 1, wherein the microbial strain comprises one or more genes encoding proteins involved in polyketide biosynthesis.

11. The method according to claim 10, wherein the one or more genes are located on a plasmid comprising (i) one or more proteins having sequences having at least 80% sequence identity with one or more of sequence numbers 21 to 35, (ii) one or more proteins having at least 80% sequence identity with sequence number 21, (iii) one or more sequences from sequence numbers 36 to 50, and / or (iv) sequence number 86 or a variant thereof having at least 80% identity with sequence number 86.

12. The method according to claim 11, wherein the bacterial strain is a Methylobacterium or Methylorubrum strain.

13. The method according to claim 11, wherein the bacterial strain is a bacterial strain other than NLS0042 (NRRL B-50932) that contains heterologous DNA from NLS0042 (NRRL B-50932) which confers a phenotype of increased expression in the plant of one or more gene transcripts involved in the production of plant defense compounds to a bacterial strain other than NLS0042, and optionally the heterologous DNA encodes one of more proteins having at least 80% sequence identity with one or more of SEQ ID NOs: 21 to 35.

14. The method according to claim 13, wherein the bacterial strain other than NLS0042 is one of the bacterial strains shown in Table 1.

15. The method according to claim 1, wherein the plant defense compound reduces foraging by insect larvae on plant roots compared to the control plant.

16. The method according to claim 3, wherein the anthranilate synthase protein component has the amino acid sequence of SEQ ID NO: 2 or SEQ ID NO:

4.

17. The method according to claim 5, wherein the expression of the gene encoding an anthranilate N-benzoyltransferase containing the polypeptide sequence of SEQ ID NO: 6 or SEQ ID NO: 7 is increased compared to the control plant.

18. The method according to claim 1, wherein the plant defense compound is not an indole derivative.

19. The method according to any one of claims 1 to 18, wherein the plant is a maize plant.

20. A method for identifying microbial strains that enhance a plant's response to pathogens or pests, (i) to obtain treated seeds and / or treated plants by treating a plant, a part of a plant, or a seed of a plant with at least a first microbial strain that is not a pathogen of the plant, (ii) Growing the treated plant in the presence of the pathogen or pest, or growing a plant from a part of the treated plant or from treated seeds, (iii) Taking one or more tissue samples from the plant and from an untreated control plant, wherein the tissue samples are taken during or after the growth stage while the pest or pathogen is attacking the plant and the untreated control plant. (iv) A method comprising assaying the sample and comparing it with an untreated control plant to identify an increased production of one or more plant defense compounds derived from anthranilates in the treated plant, thereby identifying a microbial strain that enhances the plant's response to the pathogen or pest.

21. A method for selecting a microbial strain that enhances the plant's response to a pathogen or pest, (i) Assaying one or more tissue samples from a plant treated with at least a first microbial strain that is not a plant pathogen, or from a plant grown from a portion or seeds of a plant treated with the first microbial strain, for increased levels of one or more plant defense compounds derived from anthranilates compared to one or more control tissue samples from an untreated control plant, wherein the tissue samples are collected from the treated and untreated plants while the pest or pathogen is attacking the plant tissue or after it has attacked. (ii) A method comprising selecting a microbial strain that provides the increased levels of the one or more plant defense compounds, rather than the plant pathogen, thereby enhancing the plant's response to the pathogen or pest.

22. The method according to claim 20 or 21, further comprising the step of selecting a sample for analysis from the treated plant that shows reduced damage from the pathogen or pest compared to the untreated control plant.

23. The method according to claim 20 or 21, wherein the sample is assayed to determine the level of one or more gene transcripts or polypeptides related to the production of anthranilates and / or the conversion of anthranilates to plant defense compounds.

24. The method according to claim 20, wherein the sample is assayed to determine the level of one or more plant defense compounds derived from anthranilates.

25. The method according to claim 20 or 21, wherein the pathogen or pest is a fungus, bacterium, nematode, insect, or virus.

26. The method according to claim 20, wherein a portion of the treated plant is selected from the group consisting of leaves, stems, shoots, flowers, fruits, buds, roots, tubers, rhizomes, runners, bulbs, and corms.

27. The method according to claim 20, wherein the collected tissue sample is selected from the group consisting of leaf, stem, shoot, flower, fruit, bud, root, tuber, rhizome, runner, bulb, and corm tissue sample.

28. The method according to claim 23, wherein the levels of one or more gene transcripts encoding anthranilate synthase protein components are determined in the treated tissue and the untreated tissue.

29. The method according to claim 28, wherein the anthranilate synthase protein component is an alpha or beta subunit.

30. The method according to claim 23, wherein the level of the gene transcript encoding anthranilate N-benzoyltransferase is determined in the treated tissue and the untreated tissue.

31. The method according to claim 21 or 24, wherein the plant defense compound is an anthranilate ester.

32. The method according to claim 31, wherein the anthranilate ester is selected from the group consisting of methyl anthranilate, dimethyl anthranilate, ethyl anthranilate, phenylethyl anthranilate, and menthyl anthranilate.

33. The method according to claim 21 or 24, wherein the plant defense compound is an anthranilate-derived phytoalexin.

34. The method according to claim 20 or 21, wherein the microbial strain is a bacterial strain or a fungal strain.

35. The method according to claim 34, wherein the bacterial strain is a Methylobacterium or Methylorubrum strain.

36. The method according to claim 20 or 21, wherein the plant is selected from the group consisting of corn, wheat, rye, rice, alfalfa, barley, oats, rye, sorghum, millet, soybeans, Brassica, Hemp, tobacco, potatoes, peanuts, carrots, cotton, coffee, coconut, sugar beets, oats, barley, tomatoes, pumpkins, cucumbers, gourds, lettuce, peppers, peas, onions, green beans, sunflowers, safflower, sweet potatoes, cassava, coffee, coconuts, conifers, turfgrass, leafy vegetables, microgreens, herbs, fruit trees and other fruit plants, and ornamental plants.

37. The method according to claim 36, wherein the plant is corn.

38. The method according to claim 37, wherein the tissue sample is a root sample.

39. The method according to claim 20 or 21, wherein the pest is a corn root worm.

40. The first microbial strain is obtained by transferring DNA from a second microbial strain, which can confer resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion to or from the pest or pathogen by increasing the production of one or more plant defense compounds derived from anthranilates in treated plants, to a third microbial strain that does not confer resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion to or from the pest or pathogen in plants treated with the third microbial strain, and optionally the second microbial strain is NLS0042 and / or optional The method according to claim 20 or 21, wherein, optionally, the third microbial strain is a microbial strain other than NLS0042 provided in Table 1, and optionally, the imported DNA is present on a plasmid containing (i) a sequence having at least 80% sequence identity to one or more of SEQ ID NOs: 21 to 35, or (ii) SEQ ID NOs: 86, or a variant thereof having at least 80% sequence identity to SEQ ID NOs: 86, and / or (iii) a sequence having at least 80% identity to one or more of SEQ ID NOs: 37 to 50, or a variant thereof having at least 80% identity to one or more of SEQ ID NOs: 37 to 50.

41. A method for monitoring pest or pathogen resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion in plants, which can confer said pest or pathogen resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion, and which involves at least one tissue sample from a plant treated with a first microbial strain that is not a pathogen of the plant, or from a plant grown from a part or seed of a plant treated with the first microbial strain, (i) Increased levels of one or more plant defense compounds derived from anthranilates, and / or (ii) Assaying for increased expression of one or more gene transcripts or polypeptides related to the production of anthranilates and / or the conversion of anthranilates to plant defense compounds, A method wherein the increased levels and / or increased expression are compared to one or more control tissue samples from an untreated control plant, the tissue samples being collected from the treated plant and the untreated control plant while the pest or pathogen is attacking the plant tissue or after the attack, thereby indicating that the increased levels of the one or more plant defense compounds, gene transcripts, and / or polypeptides in the tissue samples from the treated plant, compared to the control plant, indicate increased pest or pathogen resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion in the treated plant.

42. The method according to claim 41, wherein the treated plant does not exhibit increased levels of the one or more plant defense compounds, gene transcripts, and / or polypeptides, and the method further comprises retreatment with the first microbial strain and / or treatment with another biological control agent, insecticide, fungicide, or pesticide.

43. A method for selecting a microbial strain that enhances a plant's response to a pathogen or pest, comprising: (i) assaying one or more tissue samples from a plant treated with at least a first microbial strain that is not a plant pathogen, or from a plant grown from part or seeds of a plant treated with the first microbial strain, for increased levels of one or more plant defense compounds derived from anthranilates compared to one or more control tissue samples from an untreated control plant, wherein the tissue samples are taken from the treated and untreated plants while the pest or pathogen is attacking the plant tissue or after it has attacked; and (ii) selecting a microbial strain that is not a plant pathogen and provides the increased levels of the one or more plant defense compounds, thereby selecting a microbial strain that enhances the plant's response to the pathogen or pest.

44. A method for selecting a microbial strain that enhances a plant's response to a pathogen or pest, comprising: (i) assaying one or more plant tissue samples treated with at least a first microbial strain that is not a pathogen of the plant for increased levels of one or more plant defense compounds derived from anthranilates compared to one or more untreated control plant tissue samples, wherein the tissue samples were exposed to the pest or pathogen during or after treatment with the first microbial strain; and (ii) selecting a microbial strain that provides the increased levels of the one or more plant defense compounds, thereby selecting a microbial strain that enhances the plant's response to the pathogen or pest.

45. A method for improving a plant's response to a pathogen or pest, wherein the method is (i) modifying the plant genome to increase the expression of one or more gene transcripts involved in the production of one or more plant defense compounds in the plant, and / or treating the plant, part of the plant, or seeds with a microbial strain that is not pathogenic to the plant, wherein the treatment increases the expression of one or more gene transcripts involved in the production of plant defense compounds in the plant, and (ii) A method comprising growing the plant in the presence of the pathogen or pest, thereby increasing the level of one or more plant defense compounds in the plant compared to a control plant, and improving the plant's response to the pathogen or pest compared to a control plant, wherein the control plant is not modified or treated as in (i), and increasing the level of plant defense compounds produced in the plant by growing it.

46. The method according to claim 45, wherein the plant defense compound is derived from an anthranilate.

47. A method for eliminating pathogens or pests, wherein the method is (i) treating soil, plants, parts of plants, or seeds with a non-pathogenic microbial strain, wherein the treatment increases the expression of one or more gene transcripts involved in the production of plant defense compounds in the plant. (ii) A method for increasing the level of plant defense compounds produced in a plant, comprising cultivating the plant in the presence of the pathogen or pest such that the level of one or more plant defense compounds in the plant increases compared to a control plant, the plant repels the pathogen or pest better than the control plant, and the control plant is not modified or treated as in (i).

48. The method according to claim 47, wherein the plant defense compound is derived from an anthranilate.

49. The method according to claim 47, wherein the microbial strain produces metabolites, and such metabolites increase the production of such plant defense compounds compared to the control plant.

50. The method according to claim 49, wherein the metabolite is selected from the group consisting of siderophores, non-ribosomal peptides, polyketides, or combinations thereof.

51. The method according to claim 50, wherein the microbial strain comprises one or more gene clusters encoding proteins involved in the biosynthesis of the metabolites.

52. The method according to claim 47, wherein the microbial strain comprises (i) one or more proteins having a sequence having at least 80% sequence identity with one or more of sequence numbers 21 to 35, and (ii) one or more genes encoding one or more proteins having at least 80% sequence identity with sequence number 21.

53. A method for selecting a microbial strain that confers pest or pathogen resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion to a plant, comprising identifying in the microbial strain (i) one or more polynucleotides encoding a protein in the polyketide biosynthesis pathway, and / or (ii) one or more proteins in the polyketide biosynthesis pathway.

54. The method according to claim 53, wherein the polynucleotide is identified by detecting at least one polynucleotide present on a plasmid containing (i) a sequence having at least 80% sequence identity to one or more sequences from SEQ ID NOs: 21 to 35, (ii) a sequence having at least 80% sequence identity to SEQ ID NOs: 21, (iii) one or more sequences having at least 80% sequence identity to one or more sequences from SEQ ID NOs: 36 to 50, or any one of SEQ ID NOs: 36 to 50, and / or (iv) SEQ ID NOs: 86, or a sequence having at least 80% sequence identity to SEQ ID NOs:

86.

55. The method according to claim 54, wherein the polynucleotide is identified by nucleic acid amplification, hybridization, and / or sequencing techniques.

56. The method according to claim 53, wherein the protein is identified by (i) detecting one or more proteins having a sequence having at least 80% sequence identity with one or more of sequence numbers 21 to 35, or (ii) detecting one or more proteins having at least 80% sequence identity with sequence number 21.

57. The method according to claim 56, wherein the protein is identified by immunoaffinity and / or mass spectrometry techniques.

58. The method according to claim 53, further comprising the step(s) of isolating and / or culturing the identified microbial stain containing the polynucleotide and / or protein.

59. The method according to any one of claims 53 to 58, wherein the microbial strain comprises a gene that (i) encodes one or more proteins having a sequence having at least 80% sequence identity with one or more of sequence codes 21 to 35, (ii) encodes one or more proteins having at least 80% sequence identity with sequence code 21, or (iii) one or more sequences from sequence codes 36 to 50, or variants thereof having at least 80% sequence identity with sequence codes 36 to 50.

60. The method according to any one of claims 53 to 58, wherein the microbial strain is genetically modified with DNA, wherein it is located on a plasmid containing (i) one or more proteins having at least 80% sequence identity to one or more of sequence IDs 21 to 35, (ii) one or more proteins having at least 80% sequence identity to sequence ID 21, (iii) one or more sequences having at least 80% sequence identity to one or more of sequence IDs 36 to 50, and / or (iv) sequence ID 86, or a sequence having at least 80% sequence identity to sequence ID 86.

61. A method for improving a plant's response to a pathogen or pest, wherein the method is (i) Treating a plant, part of a plant, or seed with a microbial stain containing DNA present on a plasmid containing NLS0042, or (i) one or more proteins having sequences having at least 80% sequence identity to one or more of SEQ ID NOs: 21-35, (ii) one or more proteins having at least 80% sequence identity to SEQ ID NOs: 21, (iii) one or more sequences from SEQ ID NOs: 37-50, or variants thereof having at least 80% identity to one or more of SEQ ID NOs: 37-50, and / or (iv) SEQ ID NOs: 86, or variants thereof having at least 80% identity to SEQ ID NOs: 86, (ii) A method comprising cultivating the plant or a plant grown from the seeds in the presence of the pathogen or pest such that the plant's response to the pathogen or pest is improved compared to a control plant, and the plant is not a maize plant.

62. The method according to claim 61, wherein the plant is selected from the group consisting of chili peppers, tomatoes, bayberries, Brassica plants, soybeans, cotton, and rice.

63. The method according to claim 61, wherein the level of one or more plant defense compounds in the plant is increased compared to a control plant.

64. The method according to claim 61, wherein the pathogen or pest is an insect pest selected from the group consisting of aphids, lepidoptera, stink bugs, whiteflies, sugar beet leafhoppers, tobacco hawk moths, potato leafhoppers, bean ladybugs, flea beetles, fall armyworms, soybean inchworms, citrus thrips, and rice water weevils.

65. The method according to any one of claims 61 to 64, wherein the improved response of the treated plant or a plant grown from the treated part or seeds includes improved pest or pathogen resistance, tolerance, reduced damage, reduced infection, and / or reduced invasion compared to an untreated control plant.

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