Methods for enhanced root nodulation in legumes

Abstract

Disclosed herein are methods of increasing, enhancing, or accelerating root nodulation in a plant, accelerating growth of nitrogen fixing bacteria in nodules of a plant, increasing protein content in a plant, increasing yield of a plant, improving water retention of a plant, or reducing water use of a plant, the method comprising identifying a plant in need of root nodulation, and applying to the plant a composition comprising a protein component comprising yeast stress proteins resulting from subjecting a mixture obtained from the yeast fermentation to stress.

What is claimed is: 
     
         1 . A method of increasing, enhancing, or accelerating root nodulation in a legume plant, accelerating growth of nitrogen fixing bacteria in nodules of a legume plant, increasing protein content in a legume plant, increasing yield of a legume plant, improving water retention of a legume plant, or reducing water use of a legume plant, the method comprising:
 applying to the legume plant a composition comprising:
 a yeast preparation component comprising yeast stress proteins resulting from subjecting a mixture obtained from yeast fermentation to stress; 
 at least one non-ionic surfactant; 
 at least one anionic surfactant; and 
 water to make 100% volume; 
   
       wherein the yeast preparation component comprising yeast stress proteins is produced by a method comprising:
 growing yeast under aerobic fermentation conditions to obtain a fermentation mixture comprising fermented yeast cells, and proteins and peptides secreted therefrom; 
 subjecting the fermentation mixture to stress in order to stimulate the yeast cells to express heat shock proteins to obtain a stressed fermentation mixture; 
 separating the stressed fermentation mixture into a solid/precipitate and a supernatant; and 
 adjusting the pH of the supernatant to produce the yeast preparation component comprising yeast stress proteins. 
 
     
     
         2 . The method of  claim 1 , wherein the composition applied to the legume plant comprises:
 23% by volume of the yeast preparation.   
     
     
         3 . The method of  claim 1 , wherein the composition applied to the legume plant comprises:
 7.5% by volume non-ionic surfactant(s).   
     
     
         4 . The method of  claim 1 , wherein the composition applied to the legume plant comprises:
 2.5% by volume anionic surfactant(s).   
     
     
         5 . The method of  claim 1 , wherein the composition applied to the legume plant comprises:
 23% by volume of the yeast preparation;   7.5% by volume non-ionic surfactant(s); and   2.5% by volume anionic surfactant(s).   
     
     
         6 . The method of  claim 1 , wherein the at least one non-ionic surfactant comprises at least one of a dodecyl dimethylamine oxide, a coco diethanol-amide alcohol ethoxylate, a linear primary alcohol polyethoxylated, an alkylphenol ethoxylate, an alcohol ethoxylate, an EO/PO polyol block polymer, a polyethylene glycol ester, or a fatty acid alkanolamide. 
     
     
         7 . The method of  claim 1 , wherein the at least one anionic surfactant comprises at least one of sodium linear alkylbenzene sulphonate (LABS), sodium lauryl sulphate, sodium lauryl ether sulphate, a petroleum sulphonate, a linosulphonate, a naphthalene sulphonate, a branched alkylbenzene sulphonate, a linear alkylbenzene sulphonate, an alcohol sulphate, or PO and/or PO/EO sulfated alcohol. 
     
     
         8 . The method of  claim 1 , wherein the composition applied to the legume plant comprises:
 23% by volume of the yeast preparation;   7.5% by volume linear primary C12-C15 alcohol; and   2.5% by volume sodium lauryl ethyl sulfate.   
     
     
         9 . The method of  claim 1 , wherein the composition is applied to the soil near the plant. 
     
     
         10 . The method of  claim 1 , wherein the composition is applied through irrigation. 
     
     
         11 . The method of  claim 10 , wherein the irrigation is spray irrigation or drip irrigation. 
     
     
         12 . The method of  claim 1 , wherein the composition is applied with every watering cycle or on an intermittent basis. 
     
     
         13 . The method of  claim 1 , wherein the aerobic fermentation conditions comprise continuous aeration and agitation between 30 to 35° C. and at a pH of 4.0 to 6.0 for a period of at least 10 hours. 
     
     
         14 . The method of  claim 1 , wherein subjecting the fermentation mixture to stress comprises heating the fermentation mixture to 40-60° C. for at least 2 hours. 
     
     
         15 . The method of  claim 14 , wherein subjecting the fermentation mixture to stress further comprises cooling the fermentation mixture to 25° C. after heating it. 
     
     
         16 . The method of  claim 1 , further comprising adding sodium benzoate and propylene glycol to the supernatant, to produce the protein component comprising yeast stress proteins. 
     
     
         17 . The method of  claim 1 , wherein the stress further comprises physically or chemically disrupting the yeast after fermentation of the yeast. 
     
     
         18 . The method of  claim 1 , wherein the stress further comprises lysing the yeast after fermentation of the yeast. 
     
     
         19 . The method of  claim 1 , further comprising mixing the protein component with additional nutrients prior to the application to the plant. 
     
     
         20 . The method of  claim 1 , wherein the composition further comprises one or more of an anionic surfactant, a non-ionic surfactant, a cationic surfactant, and amphoteric surfactant. 
     
     
         21 . The method of  claim 1 , wherein the yeast is selected from the group consisting of  Saccharomyces cerevisiae, Kluyveromyces marxianus, Kluyveromyces lactis, Candida utilis  (Torula yeast),  Zygosaccharomyces, Pichia pastoris , and  Hansanula  polymorpha. 
     
     
         22 . The method of  claim 1 , where separating the stressed fermentation mixture into a solid/precipitate and a supernatant comprises centrifugation. 
     
     
         23 . The method of  claim 1 , wherein the legume plant is selected from the group consisting of alfalfa, clover, peas, beans, lentils, lupins, mesquite, carob, soy, peanuts, locust trees ( Gleditsia  or  Robinia ),  wisteria , and the Kentucky coffeetree ( Gymnocladus dioicus ). 
     
     
         24 . A soil mixture comprising soil and a composition comprising:
 a yeast preparation component comprising yeast stress proteins resulting from subjecting a mixture obtained from yeast fermentation to stress;   at least one non-ionic surfactant;   at least one anionic surfactant; and   water to make 100% volume;   
       wherein the yeast preparation component comprising yeast stress proteins is produced by a method comprising:
 growing yeast under aerobic fermentation conditions to obtain a fermentation mixture comprising fermented yeast cells, and proteins and peptides secreted therefrom; 
 subjecting the fermentation mixture to stress in order to stimulate the yeast cells to express heat shock proteins to obtain a stressed fermentation mixture; 
 separating the stressed fermentation mixture into a solid/precipitate and a supernatant; and 
 adjusting the pH of the supernatant to produce the yeast preparation component comprising yeast stress proteins. 
 
     
     
         25 . The method of  claim 24 , wherein the aerobic fermentation conditions comprise continuous aeration and agitation between 30 to 35° C. and at a pH of 4.0 to 6.0 for a period of at least 10 hours. 
     
     
         26 . The method of  claim 24 , wherein subjecting the fermentation mixture to stress comprises heating the fermentation mixture to 40-60° C. for at least 2 hours. 
     
     
         27 . The method of  claim 26 , wherein subjecting the fermentation mixture to stress further comprises cooling the fermentation mixture to 25° C. after heating it. 
     
     
         28 . The method of  claim 24 , further comprising adding sodium benzoate and propylene glycol to the supernatant, to produce the protein component comprising yeast stress proteins. 
     
     
         29 . The method of  claim 24 , wherein the stress further comprises physically or chemically disrupting the yeast after fermentation of the yeast. 
     
     
         30 . The method of  claim 24 , wherein the stress further comprises lysing the yeast after fermentation of the yeast. 
     
     
         31 . The method of  claim 24 , further comprising mixing the protein component with additional nutrients prior to the application to the plant. 
     
     
         32 . The method of  claim 24 , wherein the composition further comprises one or more of an anionic surfactant, a non-ionic surfactant, a cationic surfactant, and amphoteric surfactant. 
     
     
         33 . The method of  claim 24 , wherein the yeast is selected from the group consisting of  Saccharomyces cerevisiae, Kluyveromyces marxianus, Kluyveromyces lactis, Candida utilis  (Torula yeast),  Zygosaccharomyces, Pichia pastoris , and  Hansanula  polymorpha. 
     
     
         34 . The method of  claim 24 , where separating the stressed fermentation mixture into a solid/precipitate and a supernatant comprises centrifugation. 
     
     
         35 . The method of  claim 24 , wherein the composition applied to the legume plant comprises:
 23% by volume of the yeast preparation.   
     
     
         36 . The method of  claim 24 , wherein the composition applied to the legume plant comprises:
 7.5% by volume non-ionic surfactant(s).   
     
     
         37 . The method of  claim 24 , wherein the composition applied to the legume plant comprises:
 2.5% by volume anionic surfactant(s).   
     
     
         38 . The method of  claim 1 , wherein the composition applied to the legume plant comprises:
 23% by volume of the yeast preparation;   7.5% by volume non-ionic surfactant(s); and   2.5% by volume anionic surfactant(s).   
     
     
         39 . The method of  claim 24 , wherein the at least one non-ionic surfactant comprises at least one of a dodecyl dimethylamine oxide, a coco diethanol-amide alcohol ethoxylate, a linear primary alcohol polyethoxylated, an alkylphenol ethoxylate, an alcohol ethoxylate, an EO/PO polyol block polymer, a polyethylene glycol ester, or a fatty acid alkanolamide. 
     
     
         40 . The method of  claim 24 , wherein the at least one anionic surfactant comprises at least one of sodium linear alkylbenzene sulphonate (LABS), sodium lauryl sulphate, sodium lauryl ether sulphate, a petroleum sulphonate, a linosulphonate, a naphthalene sulphonate, a branched alkylbenzene sulphonate, a linear alkylbenzene sulphonate, an alcohol sulphate, or PO and/or PO/EO sulfated alcohol. 
     
     
         41 . The method of  claim 24 , wherein the composition applied to the legume plant comprises:
 23% by volume of the yeast preparation;   7.5% by volume linear primary C12-C15 alcohol; and   2.5% by volume sodium lauryl ethyl sulfate.   
     
     
         42 . A method of improving water retention of a legume plant, reducing water use of a legume plant, accelerating root nodulation in a legume plant, accelerating nitrogen fixation by a legume plant, accelerating growth of nitrogen fixing bacteria in nodules of a legume plant, increasing protein content in a legume plant, or increasing yield of a legume plant, the method comprising: growing the legume plant in the soil mixture of  claim 24 .