Microbial Fermentation Methods and Compositions
The present invention provides methods for the cultivation of the Methylobacterium genus of bacteria. In particular the method provides methods for the efficient and inexpensive cultivation of these bacteria. Additionally, the invention provides methods for the utilization of these bacterial cultures to improve plant agriculture.
1 .- 115 . (canceled)
116 . A method for obtaining a Methylobacterium preparation comprising:
(i) preparing a culture medium comprising a mono-culture or co-culture of Methylobacterium, wherein said medium comprises a liquid phase and a solid phase, wherein the liquid and solid phase are provided as sterile components, wherein the solid phase comprises a plurality of particles which are suspended in the liquid phase, and wherein the solid phase does not comprise viable photosynthetic microorganisms;
(ii) growing said Methylobacterium from an initial titer in said culture medium, wherein said culture medium is essentially free of contaminating microorganisms, wherein said particles provide for a decrease in doubling time of said Methylobacterium grown in said culture medium relative to the doubling time of the Methylobacterium grown in liquid media alone, and wherein the Methylobacterium titer in said culture medium is increased at least ten-fold relative to the initial titer of the Methylobacterium in the culture medium;
(iii) producing a fermentation broth with a Methylobacterium titer of at least 5×10 8 colony-forming units per milliliter; and
(iv) harvesting the Methylobacterium from said fermentation broth, thereby obtaining the Methylobacterium preparation.
117 . The method of claim 116 , wherein said method comprises the further step of drying the harvested Methylobacterium.
118 . The method of claim 117 , wherein drying is effected by spray drying, lyophilization, desiccation, heating, and combinations thereof.
119 . The method of claim 116 , wherein the titer of said Methylobacterium strain is increased at least 100-fold relative to the initial titer of the Methylobacterium in the culture medium.
120 . The method of claim 116 , wherein the particles are about 2 microns to about 1000 microns in average length or average diameter.
121 . The method of claim 116 , wherein said solid phase comprises about 0.02% to about 20% of said media by mass.
122 . The method of claim 116 , wherein said solid phase comprises about 0.02% to about 0.5% of said media by mass.
123 . The method of claim 116 , wherein at least 99.9% of the microorganisms present in the fermentation broth are Methylobacterium.
124 . The method of claim 116 , wherein said solid phase provides for adherent growth of said Methylobacterium and/or wherein said solid phase does not serve as a carbon source for said Methylobacterium.
125 . The method of claim 116 , wherein said solid phase comprises a solid substance selected from the group consisting of a polysaccharide, a diatomaceous earth, a salt crystal, and combinations thereof.
126 . The method of claim 125 , wherein said solid phase comprises salt crystals formed during preparation of the culture medium.
127 . The method of claim 125 , wherein said polysaccharide is selected from the group of a cellulosic polysaccharide, a chitinous polysaccharide, and a galactan polysaccharide.
128 . The method of claim 116 , wherein said Methylobacterium is selected from the group consisting of M. aminovorans, M. chloromethanicum, M. dichloromethanicum, M. extorquens, M. fujisawaense, M. mesophilicum, M. organophilum, M. radiotolerans, M. rhodesianum, M. rhodinum, M. thiocyanatum, M. nodulans, M. cerastii, M. gossipiicola, Methylobacterium sp. strain LMG6378, M. phyllosphaerae, M. oryzae, M. platani, M. populi, and M. zatmanii.