Field of Invention
This invention relates generally to the field of compositions and methods for
producing organic origin fertilizers in general and mycorrhizal origin fertilizers in
particular, to serve effectively as the biofertilizers of future that are cheap, have
cost effective production and above all are environment friendly in sharp contrast
to the fertilizers of chemical origin.
The invention focuses on not only generating a biofertilizer consortium but also
developing the process protocols for the same by means of new and innovative
highly efficient 'root organ culture' technique to generate a sort of cocktail
biofertilizer of mycorrhizal origin the benefit of which in the areas of crop
productivity and biocontrol is well known.
The invention more particularly describes the isolation and characterization,
including but not confined to, novel ectomycorrhizae-based bio-inoculums for use
in soil fertilization, biofertilizing activity and reclamation of industrially created
wastelands.
Background of Invention
The last few decades have witnessed in general a large amount of activity in the
general field of fertilizers and the biofertilizers in particular have attracted special
attention, primarily on account of them having been derived from natural sources
and having an essentially biological origin and their possible exploitation in the
numerous fields. It is here that the 'Mycorrhizal Assoiations' have been the
subject matter of experimental research to develop the various types of
biofertilizers and/or bio-inoculums for use and exploitation in diverse fields of
agriculture, forestry and ecological land reclamation exercises in various
programs. The prior art is flooded with a large number of patented inventions and
technical literature on the subject in question.
It is common knowledge that, agriculture is the science, art, and business of
cultivating the soil, producing crops, raising livestock; and farming. With respect
to cultivating the soil and producing crops, it is well known to add various
fertilizing and other compositions to the soil and/or plant foliage in order to
improve results. The agents that have been added to soil and/or plant tissues
include microbial agents, which impart some beneficial property to the soil and/or
plant to provide for desirable results.
The earliest reference of the term 'Mycorrhizae' finds mention in the 1885
German publication of Frank, according to which a mycorrhiza (Gk. UUKOC,
mykos, "fungus" and pi^a, riza, "roots",[ pi mycorrhizae, mycorrhizas) is a
symbiotic (generally mutualistic, but occasionally weakly pathogenic) association
between a fungus and the roots of a vascular plant. In a mycorrhizal association,
the fungus colonizes the host plant's roots, either intracellularly as in arbuscular
mycorrhizal fungi (AMF), or extracellularly as in ectomycorrhizal fungi. They are
an important component of soil life and soil chemistry.
With respect to the occurrence of the mycorrhizal associations, it is common
knowledge today, that, around 400 million years ago Rhynie Chert an early
Devonian period sedimentary deposit contained the earliest fossil assemblage
yielding plants preserved in sufficient detail to detect mycorrhizas - and they were
indeed observed in the stems of Aglaophyton major.
The studies have revealed that the 'mycorrhizas' are present in 92% of plant
families studied (80% of species) [Wang, B 2006], with arbuscular mycorrhizas
being the ancestral and predominant form and indeed the most prevalent
symbiotic association found in the plant kingdom. The structure of arbuscular
mycorrhizas has been highly conserved since their first appearance in the fossil
record, with both the development of ectomycorrhizas, and the loss of
mycorrhizas, evolving convergently on multiple occasions.
The significance of the mycorrhizal association can be judged from the fact that
today it is well established and backed by experimental evidence that the
mycorrhizal association has a key role to play in the following areas :
* Sugar-Water/Mineral exchange
* Absorption Mechanisms
* Disease and drought resistance
* Colonization of barren soil
* Resistance to toxicity
Thus in nutshell, an inference can be drawn to the effect that, the Ectomycorrhiza
has tremendous potential to benefit the plants by absorbing nitrogen,
phosphorus, potassium and calcium, converting complex organic molecules into
simpler available forms, protecting the roots from the pathogens, and producing
growth promoting substances (cytokinins). The Ectomycorrhizal association is
also capable of providing the trees with the ability to grow in disturbed soils such
as acidic and low fertility soils.
The US Patent No. 8,246,711 issued in favor of Marler discloses an invention
that describes the biotic fertilizers that build soil nutrients by accelerating the
growth of topsoil microorganisms. Biotic fertilizers are primarily aimed at
increasing populations of cyanobacteria, formally known as blue-green algae
and like organisms that have the ability to engage in photosynthesis reand to
engage in the-extraction of nitrogen from the atmosphere. Methods of biotic
fertilizer manufacture are presented that utilize animal waste product as well as
desirable compositions obtained thereby.
The US Patent No. 5,772,721 issued in favor of Kazemzadeh discloses an
invention that relates to a substantially odorless and sterilized fertilizer in pellet
form is produced by combining a dry organic waste material and a dry binder
material, then combining the resulting dry mixture with steam, water and/or
further organic waste in the form of sludge. The resulting material is provided to
an extruder for a more thorough, dispersive mixing, a pressure increase to at
least 100 psi, and heating above the glass transition temperature of the binder,
and more preferably to at least about 125.degree. C. to substantially sterilize the
material mass. The extruded material is forced through a die and released to an
ambient environment, rapidly reducing pressure of the material and thereby
lysing spores and microorganisms surviving conditions within the extruder. The
emerging material is segmented and dried, or segmented and tumbled to form
pellets, then dried. When dried to a moisture content of at most 7%, the pellets
further can be inoculated with an active agent within an adhesive, resulting in a
final moisture content of 10-14%.
The US Patent Application bearing No. 813070, published as US 2005/0132431
A 1 and belonging to Coughlan et al discloses an invention relating to the
methodology for synthesizing ectomycorrhiza in vitro. The invention in this patent
application further goes on to describe a method for colonizing a root-organ from
a plant with a mycorrhizal fungus that is representative of the naturally process of
colonization of plant roots by mycorrhizal fungi. The present invention also
relates to the use of this method for studying in vitro the colonization of plant
roots with a mycorrhizal fungus and to an in vitro model for studying the
colonization of plant roots with a mycorrhizal fungus.
The US Patent No. 6,133,196 issued in favor of Ocamb et al discloses an
invention wherein is described a novel method and composition is provided to
control Fusarium root rot and damping off on conifer seedlings. When certain
bacteria and an ectomycorrhizal fungus are placed in contact with the conifer
seed or seedling, the combination reduces or eliminates disease symptoms
caused by several Fusarium species.
The US Patent No. 6,808,917 issued in favor of Johnson describes an invention
which is an inoculum, a seed coated with the inoculum, a plant protected with the
inoculum, a method of producing the inoculum and a method of protecting a seed
or a plant with the inoculum. A preferred embodiment of the inoculum comprises
a combination of a fungus and a bacterium. Preferably, the fungus is a species of
Trichoderma and the bacterium is a species of Bacillus, preferably a sporeforming
strain of Bacillus. More preferably, the fungus is Trichoderma virens and
the bacterium is Bacillus amyloliquifaciens, although other combinations are also
preferred. Even more preferably, the fungus is Trichoderma virens GL-3 (ATCC
58678) and the bacterium is Bacillus amyloliquefaciens TJ1000 or 1BE (ATCC
BAA-390).
The US Patent No. 5,912,398 issued in favor of Goldstein et al describes an
invention wherein is disclosed a composition 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.
The US Patent No. 5,256,544 issued to Rogers et al. describes an industrial
scale continuous bioprocess for solubilizing rock phosphate ore by microbial
action. The method involves forming an aqueous mixture of phosphate
solubilizing microorganisms and phosphate ore particles of an appropriate size
and maintaining the mixture under conditions whereby the phosphate ore
particles are solubilized by a solubilizing agent produced and released by the
microorganisms. The mixture is then fractionated into an aqueous fraction
containing the soluble phosphate and a slurry fraction containing undissolved
solids. The soluble phosphate is removed from the aqueous fraction, and the
microorganisms present in the aqueous fraction are then recycled together with
the undissolved solids of the slurry fraction to continue the solubilization and
separation process.
The US Patent No. 5,021,077 issued in favor of Moore discloses an invention
that relates to a method of preparing high integrity natural nitrogenous granules
for agriculture by heating natural nitrogenous materials under alkaline conditions
until the materials develop adhesive properties, forming the materials into
granules by mechanical means, and heating the natural nitrogenous granules
until they harden; and the compositions formed by this method. The granules for
agriculture include natural fertilizers, secondary nutrients, micronutrients, and
natural animal feed protein supplements. The method also provides natural
based plant food and animal feed supplement granules containing natural or
synthetic additive substances which are useful in agriculture. The natural
materials used in the method include poultry waste, poultry feather meal hair
meal, seafood meal, blood meal, bone meal, soybean meal, food waste, and
grain by-products. The method provides natural nitrogenous granules which are
free of disagreeable odor by admixing reactive aldehyde compounds with the
natural nitrogenous materials prior to granule formation.
There remains an emerging need for an effective biofertilizer product and
developing effective biofertilization methods, which are more compatible with the
need for affordable and effective application without much of bias in favor of
usage of chemicals and/or chemically derived substances.
At the same time there also exists an urgently punctuated need for generating a
biofertilizer product by such means that there is a higher and a better propagule
production, an enhanced shelf life accompanied by an optimal degree of cost
effectiveness.
In view of the foregoing disadvantages inherent in the above-mentioned prior art
the general purpose of the present invention is:
to provide an improved combination of convenience and utility,
to include all the advantages of the prior art,
to attempt to overcome the major disadvantages/drawbacks of the prior art, and
to provide novel mycorrhizae-based biofertilizer consortium capable of serving as
effective biofertilizer and/or bio-origin based soil fertilization agents and growth
promoters as well as effective growth enhancers.
Summary of Invention
The present invention provides for a new and novel mycorrhizae based
biofertilizer inoculum and entails a method of production for same and also
focuses on developing the process protocols for the isolation and
characterization of the various formulations and ensuing compositions of the
mycorrhizal fungal inocula to generate the mycorrhizae based biofertilizer
consortium. The biofertilizer consortium is capable of serving as effective soil
fertilization agent, growth promoter as well as a growth enhancer
The methodology of the present invention has the potential to offer multifold
advantages of higher and better propagule production, an enhanced shelf life
and optimal degree of cost effectiveness and hence prospect for an easy
upscalibility in comparison to the conventional production methods. The inventors
are of the firm belief that an up scale version of this invention can go a long way
in contributing to at least a partial replacement of chemical fertilizers and thus the
present invention in addition to other novel, non-obvious and innovative features
also has the potential of being clean and environment friendly in terms of
application and usage.
The biofertilizer consortium in accordance with the technical and innovative
aspects contained in the present invention, is a multipurpose and multi-faceted
product-it is a growth enhancer, growth promoter, soil conditioner, bioremediator,
fruiting body producer with applications in food commercialization
and bio-control agent and has wide applications in agriculture, plantations
horticulture, forestry, and biofuels. It offers sustainable and environment-friendly
solutions to almost all cultivated plants and crops by enhancing nutrition and
yields up to 5%-25%, and curtailing chemical fertilizer inputs by 50%. It has also
shown immense potential in reclamation of stressed ecosystems like fly ash
dumps, sites loaded with alkali chlor sludge or distillery effluents, and other manmade
wastelands.
For a better understanding of the invention, its operating advantages and the
specific objects attained by its user, reference should be made to the
accompanying drawings and descriptive matter in which there are illustrated
embodiments of the invention.
Brief Description of Drawings
For a better understanding of the nature of the present invention, reference
should be made to the detailed description taken in conjunction with the
accompanying drawings in which:
Figure 1 is a diagrammatic depiction of the simplified flow chart for generating the
mycorrhizae based biofertilizer consortium.
Detailed Description of Invention
The exemplary embodiments described herein detail for illustrative purposes are
subject to numerous variations. It js understood that various omissions,
substitutions or equivalents are contemplated as circumstances may suggest or
render expedient, but is intended to cover the application or implementation
without departing from the spirit or scope of the invention.
Figure 1 is a diagrammatic depiction of the simplified flow chart for generating the
mycorrhizae based biofertilizer consortium.
The main inventive components of the invention that is the subject matter if the
present patent application, can be enumerated as below :
The invention consist of air dried
(i) Roots colonized with mycelia of ectomycorrhiza and AMF, vesicles of AM
(ii) Mycelia of AMF and ectomycorrhiza.
(iii) Spores of AMF.
The inventive concept in the present invention entails, the invention to be a ROC
based production of both AM and ectomycorrhiza that would in nutshell represent
a broad spectrum mycorrhizal product. It has the potential to provide a relatively
high propagule number of viable, pure and contamination free inoculum of both
AM and ectomycorrhiza. It also possesses tremendous cutting edge advantage
in terms of the easy upscalibility and the cost effectiveness. With this single
inoculation immense benefit can be provided to the plants and/or plant groups
that can colonize with AM and the ones that colonize with ectomycorrhiza. The
novel inventive concept contained in the present patent application is
strategically placed in terms of the advantageous impact that the invention can
have when addressing the agriculture and agro forestry systems or revegetation
programs, especially on higher altitudes. Depending on the requirement of the
soil and the plant this invention can be used either alone or in combination with
other Plant growth promoting microorganisms and chemical fertilizers.
In accordance with the preferred embodiment of working of the present invention,
the general chronological sequence for obtaining the novel mycorrhizae-based
biofertilizer consortium entails the following key steps:
(a). Isolation and selection of pre-grown ectomycorrhizae;
«
(b). Transformation;
(c). Addition of AM to the hairy roots;
(d). Addition of pre-grown ectomycorrhizae obtained from said step (a) to
colonized hairy roots obtained from said step (c);
(e). Incubation;
(f). Regular sub-culturing of plates containing said hairy roots said AM and «*M
ectomycorrhizae to obtain multiple copies; '
(g). Harvesting;
and,
(e). Airdrying.
The preferred embodiment of working of the present invention as shown above,
involves preparing the novel mycorrhizae-based biofertilizer consortium by the
following steps.
Plant tissue is transformed with Agrobacterium rhizogenes to induce hairy roots
formation. To these hairy roots plugs of pregrown ectomycorrhiza are added to
the petriplates of solidified nutritive media harboring hairy roots. This is followed
by incubation with AM, with BOD incubation being carried out at 26 degrees. This
is followed by regular subculturing, harvesting and air drying to obtain AM and
ectomycorrhiza.
In one of the preferred embodiments of working of the present invention, it is
contemplated that by effecting colonization of plant roots by the biofertilizer
consortium of the present invention, an enhanced plant growth rate can be
achieved on account of the inoculum composition's ability to stimulate better
uptake of nutrients like phosphorus and immobile trace elements, thereby
ensuring that an aim of making better nutrition available to plants is fully met with.
In another preferred embodiment of the present invention, it is contemplated that
by increasing mycorrhizal colonization on plant roots by the biofertilizer
consortium of the present invention, an increased tolerance against a wide range
of soil stresses such as the heavy metal toxicity, salinity, drought, and high soil
temperatures can be developed in the plants, thereby ensuring that in the
presence of the inoculum composition of the present invention, chances of plant
survival are greatly enhanced.
In yet another preferred embodiment of the present invention it is contemplated
to use the biofertilizer consortium of the present invention as an effective
biocontrol agent.
Although, a particular exemplary embodiment of the invention has been
disclosed in detail for illustrative purposes, it will be recognized to those skilled
the art that numerous variations or modifications of the disclosed invention,
including the rearrangement in the molecular configuration of the roc-mediated
ectomycorrhizae-based bio-inoculum and the consequently produced
mycorrhizae-based biofertilizer compositions and/or formulations as also its
method of use being amenable to modifications on account of an application in
diverse fields such as soil biofertilization, reclamation of industrially created
wastelands, serving as suitable biocontrol agent, effective enhancement of plant
productivity, facilitating enhanced nutrient uptake in plants, and exploitation in
field of biofuel crops as well as biopesticide formulation based biomarkers are
possible.
in
Accordingly, the invention is intended to embrace all such alterations,
modifications and variations as may fall within the spirit and scope of the present
invention.
Claims
I/We Claim :
1. A novel mycorrhizae-based biofertilizer consortium for use in diverse
applications, with said mycorrhizae-based biofertilizer consortium comprising;
(a). Air dried roots colonized with mycelia of ectomycorrhiza and AMF, vesicles of
AM;
(b). Air dried mycelia of AMF and ectomycorrhizae;
and,
(c). Air dried spores of AMF.
2. The novel mycorrhizae-based biofertilizer consortium of claim 1 wherein one
of the said application is as a biofetilizing agent
3. The novel mycorrhizae-based biofertilizer consortium of claim 1 wherein one
of the said application is as a plant growth enhancer.
4. The novel mycorrhizae-based biofertilizer consortium of claim 1 wherein one
of the said application is as a plant growth promoter.
5. The novel mycorrhizae-based biofertilizer consortium of claim 1 wherein one
of the said application is as a biocontrol agent.
6. The novel mycorrhizae-based biofertilizer consortium of claim 1 prepared from
roc-mediated method, wherein said composition is possessing better nutrient*
uptake facilitating attributes.
7. A roc-mediated method for obtaining novel mycorrhizae-based biofertilizer
consortium, said method comprising steps of:
(a). Isolation and selection of pre-grown ectomycorrhizae;
(b). Transformation;
(c). Addition of AM to the hairy roots;
(d). Addition of pre-grown ectomycorrhizae obtained from said step (a) to
colonized hairy roots obtained from said step (c);
(e). Incubation;
(f). Regular sub-culturing of plates containing said hairy roots said AM and said
ectomycorrhizae to obtain multiple copies;
(g). Harvesting;
and,
(e). Airdrying.
8. The method for obtaining novel mycorrhizae-based biofertilizer consortium in
accordance with claim 7, wherein said step (b) of transformation of plant tissue is
performed by Agrobactenum rhizogenes.
9. The method for obtaining novel mycorrhizae-based biofertilizer consortium in
accordance with claim 7, wherein said step (c) of incubation
BOD at 26 degrees.
step (c) of incubation is carried out in a
10. The method for obtaining novel mycorrhizae-based biofertilizer consortium in
accordance with claim 7, wherein said step (b) of transformation is responsible
for the production of hairy roots.