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(2l) Application Number: KElP/ 2009/ 000945

(51) Int.CI.7: A 01 N/63, C 05F 11/08, C 09 K/17, C 12N 1/20
   
(22) Filing Date: 15/07/2009

(73) Owner:PHYTOMEDIA INTERNATIONAL LIMITED of  P.O BOX 79509-00200, NAIROBI., Kenya

(72) Inventor: CHARLES A. NJOROGE of P.O. BOX 79509-00200, NAIROBI, Kenya

(74) Agent/address for correspondence:                               

(54) Title: COMPOSITIONS FOR INOCULATING PLANTS.

(57) Abstract: The present invention relates to methods for producing a composition for inoculating leguminous crops in general and in particular to a method of isolating novel Rhizobium strains which are biologically active and which have high nitrogen fixation ability and to methods of creating inoculating compositions containing the said Rhizobium strains made by a process comprising the following steps; Preparing a suitable medium for growing a selected strain of Rhizobium; Mixing the selected strain of Rhizobium to the medium developed above and allowed to multiply at the temperature of between 27 and 28 degrees centigrade for 46 hours; Cooling the above mixture generated above to a temperature between 8 and 12 degrees centigrade; Adding a solution of the fertilizer medium to the cooled mixture.

NQVE!,: COMPOSITIONS FOR INOCULATING PLANTS

FIELD OF INYJtNTION


The present invention relates to methods for producing a composition for inoculating leguminous crops in general and in particular to a method of isolating novel Rhizobium strains which are biologically active and which have high nitrogen fixation ability and to methods of creating inoculating compositions containing the said Rhizobium strains.

BACK GROYND OF THE INVENTION


The present invention refers to a method of isolating biologically active Rhizobium strains that are then mixed with a suitable medium or carrier to produce the said inoculating composition.

The symbiotic relationship between legumes and Rhizobium is known phenomenon and commonly seen as nodules at the roots of legumes. The legumes provide a niche and the Rhizobium in return fixes nitrogen directly from atmospheric nitrogen converting it to useful nitrogenous compounds.

There are a number of strains of Rhizobium bacteria and different strains are adapted to different symbiotic relationships and perform differently to deferent kinds of legumes and as taught in this invention are also adapted differently to various climatic and environmental conditions.

An example is that Rhizobium japonicum adopts and converts more nitrogen in roots of Soya beans while Rhizobium leguminosarum is more effective in peas. Rhizobium trifolii is well known to be more effective in nodulating clover crop and Rhizobium phaseoli is excellent in nodulating green pea.
 
It is therefore common practice that in commercial agricultural practices farmers will use the strains that are more effective and give the best results. This means that the inoculants that are best suited for the legume seed will be that which is selected from the strain that will produce the highest number of nodules that have the best yield in forms of nitrogen fixation.


It is therefore prudent that in the development of Rhizobium inoculants, the developer should ensure that it is the most effective strain is contained in the composition that is to be used as inoculants for a specific kind of legumes.

In the current invention it has been demonstrated that even a high nitrogen fixing Rhizobium strain may receive competition from strain that already exist in an agricultural area and therefore the choice of a strain has to be done in consideration of geographical location and other environmental stress factors.

The aforementioned is true since the living bacteria in the soil is continuously being subjected to environmental pressure and naturally tend to mutate and are therefore selected through the pressure of the ecological niche. This means that many of the soils in agricultural areas contain indigenous strains which have evolved and are adapted for survival in competitive co-existence with other strains in that particular environment.

This means that a farmer may get an inoculating composition which contains the highest nitrogen fixing capability but fail to nodulate in the farm due to competition from other more adopted strains which are indigenous and more adapted to that environment and have been previously selected by environmental pressures for competition in that precise environment or certain climatic area.

SUMMARY OF INVENTION


The object of the present invention is a method for preparing an inoculating composition

containing selected strains of Rhizobium bacteria. Another aspect of the present invention 2
 

is the is the process of isolating of novel strains of Rhizobium bacteria useful as a crop inoculants for legume species which exhibit high rate of adaptation and are able to enhance nitrogen fixation activity than other indigenous strains in a geographical area.

It is another object of the present invention to provide a composition for inoculating plants that include strains selected by this method which are more predictable to enhance the growth of legume crop plants than those that are currently present in the market.

Another object of the invention is identifying the naturally predominant Rhizobum strain by comparative analysis of cultures.

It is another objective of the invention composition containing seeds coated with a fertilizer composition.

It is another objective of the invention to provide a composition for inoculating plants that in which the choice of strain of Rhizobium is selected taking into consideration many other parameters and not just how active it is but should be able to withstand competition from indigenous strains of Rhizobium in a selected geographical area.

Another object of the invention is exposing the culture of naturally predominant strain to mutagenic agent and selecting among the multigenised bacteria for mutants having high nitrogen fixation ability in association with the legumes. The present invention is further directed towards the novel strains created through the use of this process.

It is another object of the present invention to provide a method of producing a composition for inoculation of plants that contains novel Rhizobium strains which are able to compete with previous selected strains by environmental pressures for competition in that precise environment or certain climatic areas.

It is an important feature of the present invention to provide a composition for inoculating plants that are able to select for strain competitiveness and prevalence in particular agricultural environmental conditions without the necessity for simulating the
competitive conditions on a bacterial strain in artificially controlled environment, a situation which has never been able to be achieved to date.

Another object of the invention is to provide a method of preparing a composition for inoculating plants that in a agricultural useful efficient and practical and is carried by a medium that is an organic fertilizer and the composition has been able to exhibit excellent results in terms of nodulating plants and achieving the maximum nitrogen fixation rate.

The results of above invention is an inoculating composition with a fertilizer incorporated as one or more components of the culture medium containing the inoculants useful in industrial and agricultural activities.

Another object of the invention is a composition for inoculating plants prepared in accordance with the method described above.

Other objects, advantages features of the present invention will become more evident after the following detailed description, the example and embodiments not being limiting and given only to illustrate the invention but does not limit the scope of the current invention.


DETAILED DESCRIPTION OF INVENTION


The process of isolating novel Rhizobium strains according to the present invention proceeds through a series of steps described below.

First it is necessary to isolate a number of rhizobium cultures from a selected geographical area which are then analyzed to identify naturally predominant strains of rhizobium. The identified naturally predominant strains are then subjected to mutigenised colonies which have been selected for high nitrogen fixation.


After the high nitrogen fixation mutants strains are selected, the strains are returned to the field test to determine if they can remain competitive and maintain sufficiently high increase yield than the indigenous Rhizobium cultures in a given geographically location. The process begins with the identification of the competitive strains. Unlike the prior endeavors in creating rhizobium inoculating compositions, no effort has been taken to select and design to achieve competitiveness in that actual field conditions and that specific geographical and climatically conditions.

The first step in the selection of the strain directed towards identifying those indigenous naturally predominant strains that already exist in selected geographical growing areas for the specific crop that is selected. For example, if peas are chosen, a site will be selected where peas are grown and identify strains of indigenous rhizobium that are predominant in that area. The indigenous strains referred herein are those that are naturally present and the strains which have evolved and been able to adapt that environment especially soil types and climatic conditions.

The process of determining and isolating the mutant strains proceeds first by large scale collection of nodules from legume growing area that has been selected. The legume plant should be selected from a field that has not been inoculated with any Rhizobium inoculants. The plants are then removed from the field and the nodules are harvested from the plants that have been grown from this field.

The nodules are then surface sterilized and then crashed so that the Rhizobium colonies that live in the nodules can be extracted. Cells from the crashed nodules are placed in a suitable growth media like nutrients agar medium andre-cultivated in a laboratory. After growing the strains the researcher can then proceed through the strain identification process

The identification of these strains is done using state of the art technology and currently most bacteria are identified and classified based on the result of a series of tests. The most important steps that are characteristics of this current invention include examination
 
1. Compositions for inoculating plants made by a process comprising the following steps; preparing a suitable medium for growing a selected strain of Rhizobium;

Mixing the selected strain of Rhizobium to the medium developed above and allowed to multiply at the temperature of between 27 and 28 degrees centigrade for 46 hours; Cooling the above mixture generated above to a temperature between 8 and 12 degrees centigrade; Adding a solution of the fertilizer medium to the cooled mixture.

2.    Compositions for inoculating plants according to claim I that contains novel Rhizobium strains which are able to compete with previous selected strains selected by environmental pressures in that precise environment and climatic conditions.

3.    Compositions for inoculating plants according to claim 1 comprising a Rhizobium strain selected according to claim 2 carried in a soil fertilizing agent.

4.    Compositions for inoculating plants according to claim 1 where the seeds coated with a

mixture of the selected Rhizobium strain and a soil fertilizing agent.


5. Compositions for inoculating plants according to claim 1 that is useful in industrial and agricultural activities.

ABSTRACT
The present invention relates to methods for producing a composition for inoculating leguminous crops in general and in particular to~method of isolating novel Rhizobium strains which are biologically active and which have high nitrogen fixation ability and to methods of creating inoculating compositions containing the said Rhizobium strains. made by a process comprising the following steps; Preparing a suitable medium for growing a selected strain of Rhizobium; Mixing the selected strain ofRhizobium to the medium developed above and allowed to multiply at the temperature of between 27 and 28 degrees centigrade for 46 hours; Cooling the above mixture generated above to a temperature between 8 and 12 degrees centigrade; Adding a solution of the fertilizer medium to the cooled mixture.

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