Endophytes and related methods

Abstract

The present invention relates to a method for identifying and/or characterising an endophyte strain, said method including providing a plurality of samples of endophytes, subjecting said endophytes to genetic analysis, subjecting said endophytes to metabolic analysis and selecting endophytes having a desired genetic and metabolic profile. The present invention also relates to novel endophytes having a desired toxin profile wherein the endophyte produces significantly less toxic alkaloids compared with a control endophyte such as standard toxic (ST) endophyte; and/or significantly more alkaloids conferring beneficial properties compared with a control endophyte such as ST endophyte. The present invention also relates to endophyte variants having a desired genetic and metabolic profile, wherein said endophyte variants possess genetic and/or metabolic characteristics that result in a beneficial phenotype in a plant harbouring or otherwise associated with the endophyte variant. Preferably said endophyte variants are generated by polyploidisation or induced chromosome doubling.

1 . A method for identifying and/or characterising an endophyte strain, said method including:
 providing a plurality of samples of endophytes;   subjecting said endophytes to genetic analysis;   subjecting said endophytes to metabolic analysis; and   selecting endophytes having a desired genetic and metabolic profile.   
     
     
         2 . A method according to  claim 1 , further including assessing geographic origin of the endophytes and selecting endophytes having a desired genetic and metabolic profile and a desired geographic origin. 
     
     
         3 . A method according to  claim 1  wherein the plurality of samples of endophytes are provided by a method including:
 providing a plurality of plant samples; and 
 isolating endophytes from said plant samples. 
 
     
     
         4 . A substantially purified or isolated endophyte having a desired toxin profile, wherein the endophyte produces significantly less toxic alkaloids compared with a control endophyte; and/or significantly more alkaloids conferring beneficial properties compared with a control endophyte. 
     
     
         5 . An endophyte according to  claim 4 , wherein said beneficial properties include improved tolerance to water and/or nutrient stress and/or improved resistance to pests and/or diseases in the plant with which the endophyte is associated. 
     
     
         6 . An endophyte according to  claim 5 , wherein said alkaloids conferring beneficial properties are selected from the group consisting of peramine, N-formylloline, N-acetylloline and norloline. 
     
     
         7 . An endophyte according to  claim 4 , wherein said toxic alkaloids include ergovaline. 
     
     
         8 . An endophyte according to  claim 4 , wherein said endophyte is isolated from a fescue species. 
     
     
         9 . An endophyte according to  claim 8 , wherein said fescue species is tall fescue. 
     
     
         10 . An endophyte according to  claim 4 , wherein said endophyte is from the genus  Neotyphodium.    
     
     
         11 . An endophyte according to  claim 4 , wherein said toxic alkaloids are present in an amount less than approximately 1 μg/g dry weight. 
     
     
         12 . An endophyte according to  claim 4 , wherein said alkaloids conferring beneficial properties are present in an amount of between approximately 5 and 100 μg/g dry weight. 
     
     
         13 . A substantially purified or isolated endophyte selected from the group consisting of E1, NEA10, NEA11, NEA12, NEA13, NEA14, NEA16, NEA17, NEA18, NEA19, NEA20, NEA21 and NEA23. 
     
     
         14 . A plant inoculated with an endophyte according to  claim 13 , wherein the inoculated plant is otherwise free of endophytes. 
     
     
         15 . A plant, plant seed or other plant part derived from a plant according to  claim 14  and stably infected with an endophyte according to  claim 13 . 
     
     
         16 . A method of producing a modified plant comprising stably infecting a host plant with an endophyte according to  claim 13 . 
     
     
         17 . A method of analysing metabolites in a plurality of endophytes, said method including:
 providing:   a plurality of endophytes; and   a plurality of isogenic plants;   inoculating each isogenic plant with an endophyte;   culturing the endophyte-infected plants; and   analysing the metabolites produced by the endophyte-infected plants.   
     
     
         18 . A method of quantifying endophyte content of a plant, said method including measuring copies of a target sequence by quantitative PCR. 
     
     
         19 . A substantially purified or isolated nucleic acid encoding a polypeptide or transcription factor involved in sexual reproduction or vegetative hyphal fusion in an endophyte. 
     
     
         20 . A genetic construct including a nucleic acid according to  claim 10 . 
     
     
         21 . An endophyte variant having a desired genetic and metabolic profile, wherein said endophyte variant possesses genetic and/or metabolic characteristics that result in a beneficial phenotype in a plant harbouring, or otherwise associated with, the endophyte variant and wherein said beneficial phenotype is selected from the group consisting of improved tolerance to water and/or nutrient stress, improved resistance to pests and/or diseases, enhanced biotic stress tolerance, enhanced drought tolerance, enhanced water use efficiency, reduced toxicity and enhanced vigour; in the plant with which the endophyte is associated, relative to a control endophyte such as standard toxic (ST) endophyte or to a no endophyte control plant. 
     
     
         22 . An endophyte variant according to  claim 21 , wherein said endophyte variant is generated by polyploidisation or induced chromosome doubling. 
     
     
         23 . An endophyte variant according to  claim 22 , wherein said endophyte variant is generated by treating an endophyte with colchicine or a similar compound that induces polyploidisation or induced chromosome doubling. 
     
     
         24 . An endophyte variant according to  claim 21 , wherein said endophyte variant is generated by subjecting an endophyte to X-ray mutagenesis or exposing an endophyte to ionising radiation. 
     
     
         25 . An endophyte variant according to  claim 21 , wherein said endophyte variant is generated from an endophyte which is isolated from a  Lolium  species. 
     
     
         26 . An endophyte variant according to  claim 25 , wherein said  Lolium  species is  Lolium perenne.    
     
     
         27 . An endophyte variant according to  claim 21 , wherein said endophyte variant is generated from an endophyte of the genus  Neotyphodium.    
     
     
         28 . An endophyte variant according to  claim 27 , wherein said endophyte variant is generated from an endophyte of a species selected from the group consisting of  N. uncinatum, N. coenophialum  and  N. lolii.    
     
     
         29 . An endophyte variant selected from the group consisting of NEA12dh5, NEA12dh6, NEA12dh13, NEA12dh14, and NEA12dh17. 
     
     
         30 . A plant inoculated with an endophyte according to  claim 29 , wherein the inoculated plant is otherwise free of endophytes. 
     
     
         31 . A plant, plant seed or other plant part derived from a plant according to  claim 30  and stably infected with an endophyte according to  claim 29 . 
     
     
         32 . A method of producing a modified plant comprising stably infecting a host plant with an endophyte according to  claim 29 .