WO2011099021A1

WO2011099021A1 - Product that makes phosphorous available to plants

Info

Publication number: WO2011099021A1
Application number: PCT/IN2010/000179
Authority: WO
Inventors: Babubhai C. Patel; Chetan S. Patel
Original assignee: Individual
Current assignee: Individual
Priority date: 2010-02-09
Filing date: 2010-03-24
Publication date: 2011-08-18
Anticipated expiration: 2012-08-09

Abstract

This invention is based upon composition and method of preparing an improved phosphate solubilizing bacterial based product from Bacillus megaterium and Pseudomonas putida that makes phosphorous available to plant, which are unavailable due to higher soil PH. It also increases efficiency of chemical fertilizer.

Description

PRODUCT THAT MAKES PHOSPHOROUS AVAILABLE TO PLANTS

PREAMBLE OF INVENTION- This invention is particular described the nature of the invention and the manner in which it is to be performed.

FIELD OF INVENTION- The present invention relates generally to a composition and method of making a bacterial based product that makes phosphorous available to plant which are unavailable due to higher soil PH.

PRIOR ART-

In the existing system as given in United States Patent Application No. 6638730 Granted Patent wherein the invention relates to a composition useful for qualitative screening of phosphate solubilizing microorganism, and also relates to a synergistic composition for qualitative screening of phosphate solubilizing microorganisms, based upon visual observation.

In the existing system as given in United States Patent Application No. 5147441 - Granted Patent wherein the a biological product for actively producing nitrogen and phosphate enrichment of soil comprising a nitrogen-fixing effective amount of Azospirillum sp. M32, an amount of Bacillus polymixa M7 sufficient to produce polymyxin, an anti-microbial peptide active against Gram negative bacteria which converts phosphorous materials fixed in the soil to which it is applied to a phosphate in plant-available forms, and a nutrient for the bacteria comprising a cellulosic material containing cellulose as an integral component selected from the group consisting of straw, xylan, hemicellulose and lignin, as well as other organic carbon or nitrogen materials.

In the existing system as given in United States Patent Application No. 5912398 - Patent Application where in a composition is disclosed for providing phosphate fertilizer to the root zone of plants. The composition comprises a microorganism capable of producing and secreting a solubilization agent, a carbon source for providing raw material for the microorganism to convert into the solubilization agent, and rock phosphate ore for providing a source of insoluble phosphate that is solubilized by the solubilization agent and released as soluble phosphate. The composition is provided in a physical form, such as a granule, that retains the microorganism, carbon source, and rock phosphate ore, but permits water and soluble phosphate to diffuse into the soil. A method of using the composition for providing phosphate fertilizer to plants is also disclosed.

In the existing system as given in European Patent Application No. EP1049377 - Patent Application where in a mycorrhiza have growth-retardant activity for Poa annua and may be used to control the growth of P. annua in high quality sport or amenity turf consisting mainly of Agrostis or Festuca species. Strains of VA mycorrhiza selected from genera Glomus, Acaulospora, Entrophosphora, Gigaspora, Scutellospora and Scierocytis may be used. The VA mycorrhiza is applied to the soil as a formulated product in which the VA mycorrhizae constitute the principal component having biological activity on the growth of turf grass. Dose rates of inoculum having an MPN of from about 200 to 1000 or more propagules per gram are effective and may be formulated with a clay or other solid carrier. The amount of formulated product applied to the turf may be from about 5 g/m2 to about 1 kg/m2. The infective prop are selected from fungal spores, mycelium, hyphae, and fragments of mycorrhiza infected roots. These may be applied to sport or amenity turf, or other grassy area in need of P. annua control, or an area in which high quality turf-grass is to be laid or sown.

In the existing system as given in United States Patent Application No. 4589226 - Patent application wherein a package and method of its production, providing a means for transferring soluble phosphorus from a source supply to plant root systems without loss of phosphorus to surrounding soil structure by use of microorganisms. A source of soluble phosphorus is supported along with a plant seed and dormant, plant-beneficial microorganisms within the cell matrix of a cellulosic support device which may be formed into a configuration such as a capsule or ribbon. The cellulosic support device may also contain nutrients for encouraging plant growth upon germination of the plant seed. The fungal inoculum is obtained by propagation of the microorganism in symbiotic association with the root systems of host plants either hydroponically, or in soil, depending upon the microorganism species used. Spores, cryoprotected hyphae and the seed of a vector plant are combined at low temperature with a cellulosic material which is used to form the packages. The formed packages are rapidly freeze-dried, and stored. The carrier system is inoculated into soil and, under proper environmental conditions, the plant seed germinates and the microorganisms become active and develop a symbiotic relationship with the growing plant. Growth of the fungus in association with the roots of the vector plant distributes the microorganisms within the soil and serves to introduce them to other plants, allowing the microorganisms to recover phosphorus and other constituents from the soil and to deliver them to their host plant by a process of translocation. The growing plant provides a vehicle for inoculating other nearby plant roots with the microorganisms.

In the existing system as given in United States Patent Application No. 20090308121 - Patent Application wherein the invention relates to eco-friendly compositions and methods for providing plant growth enhancing formulations comprising mixtures of microbial isolates. In particular, numerous bacterial and fungal strains were isolated from a variety of soil types, from rhizospheres and from root nodules of leguminous plants, in designed combinations, for providing plant growth and plant productivity enhancing formulations. These specifically designed polymicrobial formulations would further provide protection against plant pathogens lowering the need for nitrogen containing fertilizers, solubilize minerals, protect plants against pathogens, and make available to the plant valuable nutrients, such as phosphate, thus reducing and eliminating the need for using chemical fertilizers and chemical pesticides.

In the existing system as given in United States Patent Application No. 5586411 -Patent Application wherein said methods and compositions for increasing the availability of soluble phosphate and fixed nitrogen for legume rRhizobium symbioses involving co- inoculating legume seeds with a phosphate-solubilizing soil fungus, Penicillium bilaii, and Rhizobium spp. prior to planting. The two types of microorganisms do not appear to compete for nutrients in the rhizosphere of legumes and the P. bilaii provides a source of available phosphorus for use by the plant without adversely affecting the nitrogen fixation ability of the Rhizobium spp.; and indeed such ability is enhanced. The invention is used to increase the efficiencies of nodulation, nitrogen fixation and legume crop production.

In the existing system as given in Indian Patent Application No. 00183/KOLNP/2004 - Patent Application wherein invention relates to product(s) containing alive microorganism^) suitable for soil treatment, microorganisms multiplying under different climatic and natural circumstances, as well as the procedure for the production of the products, furthermore procedure for the treatment of the soil and plants with the products. More detailed, the invention relates to a procedure for preparing the products from any of the micro-organisms specified below, or from the mixture thereof. Furthermore, the invention relates to a procedure for the creation of the cultures of the micro-organisms to be used. Subject of the invention are the micro-organisms themselves, as well. More detailed, the invention relates to a procedure for the treatment of the soil and the plants with a product containing at least one of the micro-organisms Azospirillum brasilense ssp. SW51 (NCAIM /P/ B 001293), Azotobacter vinelandii spp. M657 (NCAIM IV I B 001291 ), Pseudomonas fluorescens var. SW1 1 (NCAIM /P/ B 001296), Bacillus polymyxa var. SW17 (NCAIM /P/ B 001295/, Bacillus megaterium var. M326 (NCAIM /P/ B 001301/ -, furthermore the products multiplying and existing in the environment of the plant in question, containing the listed micro-organisms and their production.

In the existing system as given in Indian Patent Application No. 85/CAL/2000- Patent Application wherein invention relates to invention relates to a process for preparing a novel biofertiliser strains of nitrogen fixer and phosphate solubiiiser to promote soil health in particular for growth of tea plants comprising of nitrogen fixer and phosphate solubiiiser in equal proportion wherein the nitrogen fixer is prepared by the steps of a) transferring the stock culture Azotobacter Chroococcum (BAH) into mother culture modified Ashley's medium, b) allowing the mother culture medium to grow for 3-7 days, c) transferring the grown mother culture to main culture medium and d) allowing the main culture medium to grow ge for another 3-7 days to obtain Azotobacter choococcum (BAH) strains and in which the phosphate solubiiiser is prepared from as a mixture of Bacillus subtilis (BB 15), Pseudomonas Putida (BP19) Aspergillus awamori (FA16) and Aspergitlus flavus (FA18) in a ratio of 5:2:2: 1 in Pikovskaya's medium wherein Bacillus Subtilis (BB15) and Pseudomonas putida (BP 19) are prepared as in a manner of preparing Azotobacter Choococcum and Aspergillus awamori (FA 16) and Aspergillus flavus (FA 18) (molasses or yeast medium) are prepared by transferring the stock culture to mother culture medium, allowing the mother culture to grow for 15 days, transferring the grown mother culture to mairi culture medium and allowing it to grow again for another 15 days to obtain Aspergillus awamori (FA 16) and Aspergillus flavus (FA 18) strains.

OBJECT OF THE INVENTION Traditionally, the known phosphate solubilizing bacteria product has different method of its preparation. The present invention is based upon phosphate solubilizing bacteria from Bacillus megaterium , Pseudomonas putida that makes phosphorous available to plant, which are un available due to higher soil PH. It also increases efficiency of chemical fertilizer.

STATEMENT OF INVENTION- The inventor has invented a composition and method of making a bacterial based product that makes phosphorous available to plant, which are unavailable due to higher soil PH. It also increases efficiency of chemical fertilizer.

DETAILED DESCRIPTION OF INVENTION-

The ability of a few soil microorganisms to convert insoluble forms of phosphorus to an accessible form is an important trait in plant growth-promoting bacteria for increasing plant yields. The use of phosphate solubilizing bacteria as inoculants increases the P uptake by plants

The phosphate solubilizing bacteria have the capability to change the insoluble form of phosphorus into soluble one. Accordingly, in one aspect, this invention provides the composition and method of preparing phosphate solubilizing bacteria from Bacillus megaterium and Pseudomonas putida that makes phosphorous available to plant, which are un available due to higher soil PH. It also increases efficiency of chemical fertilizer.

Material and Method used in preparing phosphate solubilizing bacteria is as under Step-1 Nucleus culture (Maintain culture)

Pure culture of bacteria (Bacillus megatherium , Pseudomonas putida) is inoculated aseptically on plate having 20 ml PAC-09 Agar media; such two to three plates are generally inoculated. Such inoculated plate is maintained in BOD incubator at 27 ± 1 °C for 6 to 7 days with 12/12 hour lighting cycle. (Composition for one liter of PAC-09 agar media is 10 gm Sucrose, 0.1 gm MgS0₄7H₂0, 0.2 gm NaCl, 0.05 gm FeS0₄, 5 gm Ca₃(P0₄)₂, 0.5 gm (NH^SC^, 0.2 gm KC1, 0.5 gm Yeast extract, 0.05 gm MnS0₄, 20 gm Agar agar and 1000 ml distilled water).

Step-2 Starter culture (In 100 ml flask)

Culture grown on plate is inoculated aseptically in a flask (250 ml capacity) having 100 ml PAC-09 broth media with the help of inoculating loop, and is allowed to grow at 27 ± 1 °C for at least 3 to 4 days. For superior growth, flask is put on shaker with agitation of media at 150 RPM. (Composition for one liter of PAC-09 broth media is 10 gm Sucrose, 0.1 gm MgS0₄7H₂0, 0.2 gm NaCl, 0.05 gm FeS0₄, 5 gm Ca₃(P0₄)₂, 0.5 gm (NH₄)₂S0₄, 0.2 gm KC1, 0.5 gm Yeast extract, 0.05 gm MnS0₄ and 1000 ml distilled water). Step-3 Growth culture (In 1000 ml flask)

After sufficient growth in 250 ml flask as specified in step-2, 50 ml culture from this grown culture is inoculated aseptically in 500 ml PAC-09 broth media in a 1000 ml capacity flask for further growth at 27 ± 1 °C for at least 3 to 4 days on shaker with 150 RPM, such two flasks is prepared. (Composition for one liter of PAC-09 broth media is 10 gm Sucrose, 0.1 gm MgS0₄7H₂0, 0.2 gm NaCl, 0.05 gm FeS0₄, 5 gm Ca₃(P0₄)₂, 0.5 gm (NH₄)₂S0₄, 0.2 gm KC1, 0.5 gm Yeast extract, 0.05 gm MnS0₄ and 1000 ml distilled water).

Step-4 Seed Fermentor (In small fermentor of 10 lit capacities)

One (1) lit grown culture from step-3 is inoculated aseptically in small fermenter having 10 litter PAC-09 broth media, and allow for further growth at 27 ± 1 °C for at least 3 to 4 days with agitation by motor at 100 RPM. (Composition for one liter of PAC-09 broth media is 10 gm Sucrose, 0.1 gm MgS0₄7H₂0, 0.2 gm NaCl, 0.05 gm FeS0₄, 5 gm Ca₃(P0₄)₂, 0.5 gm (Ntt^SC^, 0.2 gm KC1, 0.5 gm Yeast extract, 0.05 gm MnS0₄ and 1000 ml distilled water).

Step-5 Production Fermentor (In Big fermentor of 200 lit capacities)

Ten (10) lit culture grown in step-4 is inoculated aseptically in Big fermenter having 200 litter PAC-09 media (Composition for one liter of PAC-09 broth media is 10 gm Sucrose, 0.1 gm MgS0₄7H₂0, 0.2 gm NaCl, 0.05 gm FeS0₄, 5 gm Ca₃(P0₄)₂, 0.5 gm (NH₄)₂S0₄, 0.2 gm KC1, 0.5 gm Yeast extract, 0.05 gm MnS0₄ and 1000 ml distilled water), and allow for further growth at 27 ± 1 °C for at least 3 to 4 days with agitation by motor at 100 RPM. Step-6 Final out put from big fermentor mixed with starch @ 4 to 5 % which act as a stabilizer and melto dextrin @ 1 to 2 % which act as a drying agent or with 5 % skim milk powder and agitate continuously by using agitator.

Step-7 Mix culture prepared as per step-6, is then Spray dried at definite 160 °C in let and 70 to 72 °C out let temp in spray dryer unit.

Step-7 Collecting dry bacterial powder

Material dried in spray dryer unit is then collected in clean stainless steel vessels and stored at 5 °C up to further use.

Step-8 Mix with suitable carrier

At the time of formulation, material previously dried is mixed with dextrose @ 10 % (10 gm bacterial powder with 90 gm dextrose powder).

Step-9 Packaging

Material prepared as above is then packed in 100 gm, 250 gm and 500 gm packing by using pouch packing machine.

The final material prepared has to be used in following form: The final material prepared has to be used in following form:

Dose in one Acre: In case of Seed treatment add 100 gm per seed required per hector For soil application add 100 gm to 250 gm per hector

Claims

CLAIMS I claim,

1. A composition and method of preparing an improved phosphate solubilizing bacterial based product from Bacillus megatherium and Pseudomonas putida that makes phosphorous available to plant, which are un available due to higher soil PH, the method comprising steps of nucleus culture, starter culture, growth culture, seed fermentor, production fermentor, mixing of final output, drying, collecting, mixing with dextrose and packing the final output.

2. A composition and method of preparing improved phosphate solubilizing bacterial product according to claim 1 , by inoculating aseptically pure culture of Bacillus megatherium and Pseudomonas putida on two or three plate having 20 ml PAC- 09 Agar media and maintain in BOD incubator at 27 ± 1 °C for 6 to 7 days with 12/12 hr lighting cycle, wherein, composition for one liter of PAC-09 agar media contains 10 gm Sucrose, 0.1 gm MgSo_{4 H2}0, 0.2 gm NaCl, 0.05 gm FeSo₄, 5 gm Ca3(Po₄)2, 0.5 gm (N_H4₎2So , 0.2 gm KC1, 0.5 gm Yeast extract, 0.05 gm MnS₀4, 20 gm Agar agar and 1000 ml distilled water.

3. A composition and method of preparing improved phosphate solubilizing bacterial product according to claim 1 , wherein culture grown on plate as per method cited in claim 2 is inoculated aseptically in a flask of 250 ml capacity having 100 ml PAC-09 broth media with the help of inoculating loop, and allowing to grow it at 27 ± 1 °C for at least 3 to 4 days, where as for superior growth, flask is put on shaker with agitation of media at 150 RPM, wherein, composition for one liter of PAC-09 broth media contains 10 gm Sucrose, 0.1 gm MgS0₄7H₂0, 0.2 gm NaCl,

0.05 gm FeS0₄, 5 gm Ca₃(P0₄)₂, 0.5 gm

0.2 gm KC1, 0.5 gm Yeast extract, 0.05 gm MnS0₄ and 1000 ml distilled water).

4. A composition and method of preparing improved phosphate solubilizing bacterial product according to claim 1 , wherein after sufficient growth in 250 ml flask as per method cited in claim 3, 50 ml culture from this grown culture is inoculated aseptically in 500 ml PAC-09 broth media in a 1000 ml capacity flask for further growth at 27 ± 1 °C for at least 3 to 4 days on shaker with 150 RPM, such two flasks is prepared, wherein composition for one liter of PAC-09 broth media contains 10 gm Sucrose, 0.1 gm MgS0₄7H₂0, 0.2 gm NaCl, 0.05 gm FeS0₄, 5 gm Ca₃(P0₄)₂, 0.5 gm (NH^SC^, 0.2 gm KC1, 0.5 gm Yeast extract, 0.05 gm

. MnS0₄ and 1000 ml distilled water).

5. A composition and method of preparing improved phosphate solubilizing bacterial product according to claim 1, One (1) lit grown culture as per method cited in claim 4, is inoculated aseptically in small fermenter having 10 litter PAC-09 broth media, and allowing it for further growth at 27 ± 1 °C for at least 3 to 4 days with agitation by motor at 100 RPM, wherein composition for one liter of PAC-09 broth media contains 10 gm Sucrose, 0.1 gm MgS0₄7H₂0, 0.2 gm NaCl, 0.05 gm FeS0₄, 5 gm Ca₃(P0₄)₂, 0.5 gm (NH₄)₂S0₄, 0.2 gm KG, 0.5 gm Yeast extract, 0.05 gm MnS0₄ and 1000 ml distilled water).

6. A composition and method of preparing improved phosphate solubilizing bacterial product according to claim 1, ten (10) lit culture grown as per method cited in claim 5 is inoculated aseptically in Big fermenter having 200 litter PAC-09 media, wherein composition for one liter of PAC-09 broth media contains 10 gm Sucrose, 0.1 gm MgS0₄7H₂0, 0.2 gm NaCl, 0.05 gm FeS0₄, 5 gm Ca₃(P0₄)₂, 0.5 gm (NH₄)₂S0₄, 0.2 gm KC1, 0.5 gm Yeast extract, 0.05 gm MnSQ₄ and 1000 ml distilled water), and allow for further growth at 27 ± 1 C for at least 3 to 4 days with agitation by motor at 100 PM.

7. A composition and method of preparing improved phosphate solubilizing bacterial product according to claim 1 , wherein, final out put from big fermentor, as per method cited in claim 6, is mixed with starch @ 4 to 5 % which act as a stabilizer and melto dextrin @ 1 to 2 % which act as a drying agent or with 5 % skim milk powder and agitate continuously by using agitator.

8. A composition and method of preparing improved phosphate solubilizing bacterial product according to claim 1 , wherein, mix culture prepared as per method cited in claim 7, is then Spray dried at definite 160 °C in let and 70 to 72 °C out let temp in spray dryer unit.

9. A composition and method of preparing improved phosphate solubilizing bacterial product according to claim 1, wherein, material dried in spray dryer unit as per method cited in claim 8 is then collected in clean stainless steel vessels and stored at 5 °C up to further use.

10. A composition and method of preparing improved phosphate solubilizing bacterial product according to claim 1 , wherein, at the time of formulation material previously dried as per method cited in claim 9 is mixed with dextrose @ 10 %, i.e. 10 gm bacterial powder with 90 gm dextrose powder and then packed as final phosphate solubilizing bacterial product.