Introduction
INTRODUCTION
Sal (Shorea robusta Gaertn. f.) is an angiospermic dicot perennial tree species
of the family dipterocarpaceae. This tree is a native to southern Asia ranging south of
the Himalaya from Myanmar in the east to India and Bangladesh to Nepal. In India it
extends from Assam, Bengal, Orissa, and Jharkhand west to the Shivalik hills in
Haryana and east of Yamuna. The range also extends through the Western Ghats and
to the eastern Vindhya and Satpura ranges of central India. It is often a dominant tree
where it occurs. Sal is a moderate to slow growing, and can attain heights of 30 to 35
m and a trunk diameter of up to 2-2.5 m. The leaves are 10–25 cm long and 5–15 cm
broad. In wetter areas, it is evergreen; in drier areas, it is deciduous, shedding most of
the leaves in between February to April, leafing out again in April -May. The sal
flowers are whitish in color, appear in early summer. These come out in axially
racemose panicles covered with white pubescence. They fruit during summer and the
seeds ripen from May-June. These tend to germinate even while on the tree and
accordingly begin to fall soon after germination (Champion and Seth, 1968) (Figs.1ab, Fig.2c).
Sal is one of the most important sources of hardwood timber in India. Sal
density has significantly reduced from Sal Dense 65.61 % in 1976 to Sal Dense
11.12% in the year 1999 followed by Sal Open 11.18 % and Sal Medium 18.24 %.
The overall change has been estimated to be 42.11% of the total forested area
(Chauhan et. al., 2003). This is an indication why protection of sal forest is necessary.
A mycorrhiza in general is a symbiotic relation between a fungus and the
roots of a vascular plant. In a mycorrhizal association, the fungus colonizes roots of
host plants, either intracellularly as in arbuscular mycorrhizal fungi or extracellularly
as in ectomycorrhizal fungi (Harley and Smith, 1983). Mycorrhizae form a
mutualistic relationship with the roots of most plant species. This mutualistic
association provides the fungus with relatively constant and direct access to
carbohydrates, such as glucose and sucrose supplied by the plant. The carbohydrates
are translocated from their source to root tissue and on to fungal partners. In return,
the plant gains the benefits from the mycobiont in terms of water and mineral
nutrients thus improving the plant's mineral absorption capabilities. Plant roots alone
1
Introduction
may be incapable of taking up phosphate ions that are demineralized for example, in
soils with a basic pH. The mycelium of the mycorrhizal fungus can, however, access
these phosphorus sources, and make them available to the plants they colonize.
(Bowen et. al., 1974, Kumar Dinesh et. al., 1968). The mechanisms of increased
absorption are both physical and chemical. Mycorrhizal mycelia are much smaller in
diameter and longer in length than the smallest root, and thus can explore a greater
volume of soil, providing a larger surface area for absorption. Also, the cell
membrane chemistry of fungi is different from that of plants. Mycorrhizae are
especially beneficial for the plant partner in nutrient-poor soils (Harley, 1959).
Mycorrhizal plants are often more resistant to diseases, such as those caused by
microbial soil-borne pathogens, and are also more resistant to the effects of drought.
Plants grown in sterile soils and growth media often perform poorly without the
addition of spores or hyphae of mycorrhizal fungi to colonise the plant roots and aid
in the uptake of soil mineral nutrients (Hatch, 1937). The absence of mycorrhizal
fungi can also slow plant growth in early succession or on degraded landscapes. Fungi
have been found to have a protective role for plants rooted in soils with high metal
concentrations, such as acidic and contaminated soils. Pine trees inoculated with
Pisolithus tinctorius planted in several contaminated sites displayed high tolerance to
the prevailing contaminant, survivorship and growth (Albert, et. al. 1981). One study
discovered that existence of zinc-tolerant strains of Suillus bovinus conferred
resistance to Pinus sylvestris. This was due to binding of the metal to the mycelium of
the fungus, without affecting the exchange of beneficial substances (Behrmann et. al.
1992). Mycorrhizae are present in 92% of plant families studied (80% of species),
with arbuscular mycorrhizae being the ancestral and predominant form, and indeed
the most prevalent symbiotic association found in the plant kingdom (Harley, 1959).
The structure of arbuscular mycorrhizae has been highly conserved since their first
appearance in the fossil record, with both the development of ectomycorrhizae, and
the loss of mycorrhizae, evolving convergently on multiple occasions (Bakshi, 1957,
1966, 1971). Sal establishes a mycorrhizal relationship with some basidiomyceteous
and gasteromyceteous fungal forms which helps the phycobiont to survive in soil
having nutrient at minimum and in return mycobionts gets shelter and prepared food
from the phycobionts (Bakshi, 1974)
2
Introduction
1.1 Mycorrhiza
Mycorrhizal symbiosis refers to the association of the fungi with plant roots.
This relationship is predominantly mutualistic that is, with both partners benefiting
from the association. There are seven types of mycorrhizal association, but common
to all types is the net movement of carbon, generally (but not always) from the plant
host to the fungus partner. In return, a fungus may confer increased nutrient supply,
defense against pathogenic attack, and drought resistance to its partner plant.
According to the A.B. Frank 1885 “Mycorrhiza is a dynamic condition of
mutual exploitation in which both partners gets benefit from each other and a
balance between invasive and defensive force is maintained”.
A mycorrhiza is a mutualistic symbiosis between a fungus and the roots of a
plant. This interaction result in recognizable fungal structure on or within roots.
Mycorrhizal fungi gain carbohydrates (simple sugar) from plant roots, and enhance
plant uptake of inorganic nutrients, particularly P and N. Mycorrhizae can provide
plant with protection against drought, high soil, temperature, acidity, heavy metal,
some pathogens, nematodes, insect and other soil organisms (Bakshi, 1974).
More than 90% of all plant families studied (80% of species) in both
agricultural and natural environments form mycorrhizal association and they can be
essential for plant nutrition. Mycorrhiza is found in a wide range of habitats, high
latitudes and altitude and aquatic ecosystems. There is little exception to the rule that
mycorrhizae are found in all plant species that are economically important to man.
Mycorrhizal association is widespread amongst plant families and appears to
have evolved and spread with the early land plants (effectively enabling early plants
with small ineffective roots to invade and become established in the terrestrial
environment).
1.2 Types of mycorrhizae
Mycorrhizae are classified into three categories
1.2.1
Ectotrophic Mycorrhiza
1.2.2
Endotrophic Mycorrhiza
3
Introduction
1.2.3
Ectendotrophic Mycorrhiza
This classification is based on the location of the fungal hyphae in relation to the root
tissues of the plant ecto means outside the root, endo means inside.
1.2.1 Ectotrophic Mycorrhiza
Ectotrophic Mycorrhiza (ECM) is a form of Mycorrhizal association in
which a well developed mycelium forms a mantle on the outside of the root. In ECM
the fungus forms a Mantle or sheath around the rootlets and also enters the root
forming an intercellular set of hyphae called the Hartig net (quoted in Hatch 1936).
The hyphae of the hartig nets do not form haustoria. In ECM the fungal component is
usually a basidiomycete or sometimes an ascomycete. ECM occurs on certain groups
of temperate shrubs and trees such as beeches, oaks, willows, poplars, cottonwoods,
and pines. The associations are most common in vegetation experiencing seasonal
growth, where they are thought to extend the growing period. In addition,
ectomycorrhizae are common on trees growing in the cold, dry conditions close to the
Arctic Circle and high on the slopes of mountains where they make the trees better
able to survive in harsh conditions. In an ectomycorrhizal association, the fungus
forms a thick mat, called a mantle, on the outside of the young roots, and it also grows
in between epidermal cells and into the cortex of the root interior. Within the root, the
fungus never penetrates any of the cells but instead remains confined to the
intercellular spaces where it forms a network called a Hartig net. The fungal
filaments, called hyphae, also extend outward from the root where they increase the
volume of soil available to be "mined" for nutrients. They also increase the surface
area for the absorption of water and mineral salts, particularly phosphates but also
NH4, K, Cu2+, Zn2+, and NO3-. Once the root is colonized by the fungus, the
production of root hairs slows or even ceases as the absorptive role of the root hairs is
taken over by the hyphae of the ectomycorrhizal fungus (Bakshi, 1974; Hatch 1936,
1937).
Ectomycorrhizas can link together groups of trees, the submerged mycelium
acting as what has been described as a ‘wood-wide-web’. Ectomycorrhizal fungi
depend on the plant host for carbon sources, most being uncompetitive as saprotrophs.
With few exception (Tricholoma fumosum), the fungi are unable to utilize cellulose
4
Introduction
and lignin; but the fungi are able to utilize nutrients, particularly phosphate and
ammonium ions, which the root cannot access. Host plants grow poorly when they
lack ectomycorrhizae. This ectomycorrhizal group is reasonably homogenous, but a
subgroup ectendomycorrhizas, has been appended (Hatch, 1937).
The ectomycorrhizal fungi (ECM) do not show a high degree of host
specificity, either in natural or artificial condition. It is common to find mycorrhizae
belonging to several different fungi on the root system of a single tree. Norway
spruce (Picea albies) can form ectomycorrhizal symbiosis with over 100 different
fungal species and the fly agaric (Amanita muscaria); can infect the roots of trees as
varied as birch, eucalyptus, spruce, douglas fir and tropical pines (Trappe, 1962).
1.2.2. Endotrophic Mycorrhizae
Endomycorrhizae are characterized by intracellular infection within the root.
Endotrophic mycorrhizae are caused by the invasion of absorbing roots by specific
phycomycetous fungi. The hyphae are present on the root surface only one individual
threads and penetrate directly into the root hairs and other cells of epidermis hyphae
grow on cortex cells within the cells the hyphae may appear as coils, swellings or
minute branches there is no penetration of meristem or stele.
Far more common are the endomycorrhizae, which have a zygomycete as the
fungal component and which actually penetrate the cell walls of the root cortex.
Although the hyphae do not enter the cytoplasm of the cortical cells, in most cases
they cause the plasma membrane to bulge inward, forming highly branched structures
called arbuscules and terminal swellings called vesicles. Thus, this type of
endomycorrhizae is referred to as arbuscular mycorrhizae (AM) or vesiculararbuscular mycorrhizae (VAM).
Arbuscular mycorrhiza is the most common type of mycorrhizal association
and was probably the first to evolve; the fungi are members of the lower fungi. AM
fungi are distantly related to other Zygomycetes and are now placed in phylum
Glomeromycota. AM fungi are obligate biotrophs, and they are associated with roots
of about 80% of plant species (that is equivalent to about two-thirds of all lands
plants, or around 90% of all vascular plants), including many crop plants (Mukerji et.
al., 2002).
5
Introduction
The arbuscules are in intimate contact with the cortical cells and provide an
increased surface area over which carbohydrates can pass from the plant to the fungus
and mineral elements from the fungus to the plant. The vesicles are thought to
function as storage compartments for the fungus. As with the ectomycorrhizae, the
fungal hyphae extend from the root into the soil and increase the surface area for
absorption, but there is no mantle or Hartig net, and root hairs are often present. AM
are found on almost all herbaceous angiosperms, some gymnosperms, and many ferns
and mosses. Endomycorrhizae are particularly important in the tropics where the soils
are typically poor in phosphates. Studies have indicated that roots associated with
mycorrhizal fungi can take up phosphate four times faster than roots without such
fungi. Mycorrhizal fungi are particularly effective in utilizing highly insoluble rock
phosphorus, Ca3 (PO4)2, that cannot be used by plants. The fungal hyphae make
phosphates available to the plant by converting them to a soluble form.
Ericoid endomycorrhizas are mycorrhizas of Erica (heather), Calluna (ling)
and Vaccinium (bilberry) are plants that endure moorlands and similar challenging
environments. Fungi are members of the Ascomycota (an example is Hymenoscyphus
ericea). The plant’s rootlets are covered with a sparse network of hyphea; the
fungus digests polypeptides saprotrophically and passes absorbed nitrogen to the plant
host; in extremely harsh condition the mycorrhiza may even provide the host with
carbon
sources (by metabolizing polysaccharides and proteins for their carbon
content).
Two specialized subgroups may be separated out of the ericoid
endomycorrhizal groups Arbutoid endomycorrhiza and Monotropoid endomycorrhiza
(the mycorrhizal association formed by the achlorophyllus plants of the
Montropaceae).
Orchidaceous endomycorrhizas are similar to ericoid mycorrhizas but their
carbon nutrition even is more dedicated to supporting the host plant as the young
orchid seedling is non-photosynthetic and depends on the fungus partner utilizing
complex carbon sources in the soil, and making carbohydrates available to the young
orchid. All orchids are achlorophyllus in the early seedling stages but usually
chlorophyllus as adults, so in this case the seedling stage orchid can be interpreted as
6
Introduction
parasitizing the fungus. A characteristic fungus example is the basidiomycete genus
Rhizoctonia.
1.2.3. Ectendotrophic Mycorrhizae
The Ectendotrophic mycorrhizae share characters of both, form the hyphal
mantle and inside the root form intercellular as well as intracellular hyphae.
Ectendomycorrhizas caused by E-strain fungi are often found in the early seedling
stages of species of Pinus and other conifers. The sheath is very thin, almost absent in
some cases. A well developed Hartig net is found just behind the meristematic region.
Further back, the cells are penetrated by the fungus and almost filled by coiled hyphae
surrounded by host plasmalemma. In the sapling or adult stage of tree growth they are
usually
supported
by
ectomycorrhizas
but,
according
to
Harley
(1959),
ectendomycorrhizas have been found in adult trees. Similar mycorrhizas have been
reported from some angiosperms (Harley, 1959). A few experiments (Harley and
Smith, 1983) have shown improved growth, presumably with enhanced mineral
nutrition, of the plant host.
1.3 Ectomycorrhizal Fungi
Ectomycorrhizal fungi are, economically, one of the most important groups
of fungi. These are the fungi that form a symbiotic relationship with a plant forming a
sheath around the root tip of the plant. The fungus then form a Hartig net which
means that there is an inward growth of hyphae (fungal cell growth from) which
penetrate the plant root structure (Harley, 1959).
The mycorrhizae are more or less annual and that their formation depends on
the availability of carbohydrates from the trees. The carbohydrate production is
variable depending on the levels of photosynthesis and tree growth (Richards and
Wilson, 1963b).
In
general,
the
fungi
involved
in
ectomycorrhizae
come
under
Basidiomycetes from the families, Amanitaceae, Boletaceae Continariaceae,
Russulaceae, Tricholomataceae, Rhizopogonaceae, and Sclerodermataceae. They are
included in genera Amanita, Boletus, Cantharellus, Cortinarius, Rhizopogon, Russula,
7
Introduction
Scleroderma and Cenococcum many of these fungi show a wide host spectrum
(Trappe, 1962; Kumar and Lakhanpal,1984; Lakhanpal, 1991).
1.4 Other Associations
Two other types of mycorrhizae are found in the heather and orchid families.
In heather (family - Ericaceae), the fungus secretes enzymes into the soil that convert
materials, particularly nitrogen-containing compounds, into nitrate or ammonical
forms that can be taken up more readily. In orchids (family - Orchidaceae), the seeds
contain a mycorrhizal fungus essential for seed germination. Within the seed, the
hyphae absorb stored carbohydrates and transfer them to the plant embryo.
Some plants, such as those of the mustard (Brassicaceae) family and the
sedge, (Cyperaceae) family lack mycorrhizae. In addition, most plants growing in
flooded soils (or under hydroponics) do not form mycorrhizae nor do plants grown
where conditions are extremely dry or saline. Also, plants growing in very fertile i.e.,
nutrient-rich soils have less-developed mycorrhizae compared to plants growing in
nutrient-poor soils.
1.5 Mycorrhizosphere effects
The term “Mycorrhizosphere” was suggested by Rambelli (1973) to
describe the soil surrounding influenced by mycorrhizas. Evidence for the
existence and possible significance of mycorrhizosphere effects has been
reviewed by a number of authors (Fogel, 1988; Linderman, 1988).
A key features of the Mycorrhizosphere is the presence of mycorrhizal
hyphae that surround the root and extend out from it in the form of dense
extramatrical
hyphea.
These
hyphae
may
extend
the
limits
of
the
mycorrhizosphere considerably and can enter rhizosphere of uninfected roots.
Interaction with other soil microorganisms may be direct or indirect, through
the effect on the host plant.
Mycorrhizal infection may cause changes in the quality and quantity of
the
root
exudates
and
secretions,
enhance
the
nutrient
and
elemental
composition, alter the hormone balance of host roots, and increase respiratory
8
Introduction
losses of CO2 from the root surface. Since mycorrhizal hyphae often
dramatically influence the distribution and absorptive surface area of root
systems they present a considerable surface area across which direct interaction
may take place with the microbial flora. A “mycosphere” may thus develop
around mycorrhizal hyphae in which enhanced microbial population of altered
species composition may occur.
The extramatrical hyphae themselves may exude substances that cause
soil and organic matter to aggregate (Sutton and Sheppard, 1976) providing
micro sites for growth of bacteria, fungi, actinomycetes and algae. In cases in
which the root is more or less completely surrounded by fungal material, such
as the sheathed lateral roots of ectomycorrhizal plants, most or all of the
substances entering the soil may do so through fungal hyphae. Interaction may
be stimulatory, inhibitory or neutral (Bowen and Thoedorou, 1979). Stimulatory
effects of microorganisms on ectomycorrhizal colonization and development
have been reported by MacAfee and Fortin (1988). The in vitro growth of
different ectomycorrhizal isolates can also be both stimulated or inhibited by
Actinomycetes isolated from the mycorrhizosphere of Pinus resinosa (Richter et
al., 1989). It seems that distinct microbial communities may have evolved in
response to the presence of specific mycorrhizal associations, and the isolations
and selection of the appropriate “helper” organisms for use as co-inoculants,
may offer scope of additive or synergistic growth stimulation.
While exudation of specific compounds from the root or mycelium
may
stimulate
microorganisms
beneficial
to
the
symbiosis,
extracellular
metabolites may, also have an antibiotic effect on certain phytopathogenic
microorganisms (Kope
and
Fortin,
1989).
The
continuous
supply
of
carbohydrates that mycorrhizal fungi receive from their hosts is probably
important in terms of providing the energy for synthesis of the wide range of
compounds that are undoubtedly involved in these microbial interaction. It is
often assumed that mycorrhizal fungi are at a competitive advantage with
respect to the saprophytic flora because of this direct supply of plant
assimilates.
9
Introduction
Interaction with saprophic population could thus influence decomposition
processes. Abuzinadah and Read (1986, a, b) suggested that the increased rates of
Pine litter decomposition following exclusion of ectomycorrhizal root could be
caused by the removal of successful competition for limited organic nitrogen
by ectomycorrhizal fungi with proteolytic enzymes. Death, decomposition and
leakage of organic compounds from decomposing mycorrhizal hyphae represents
a potential input of carbon into the soil ecosystem. The proteolytic activity of
certain ectomycorrhizal species is that organic compounds released from dying
hyphae could be used directly by living mycorrhizal hyphae and the carbon
recycled internally within the ectomycorrhizal association, restricting losses to
the soil ecosystem through immobilization.
Nutrient cycling may take place on different spatial scales, and that cycling at
very small scales-nutrient micro-loops may contribute to over all cycling at larger
scales. Many “short circuits” in traditional pathways of nutrient cycling are emerging
and the significance of these needs to be more closely investigated (Abuzinadah and
Read, 1986, a, b).
1.6 Spread of mycorrhizal colonization
In most countries, where pines and other ectotrophic trees like eucalyptus is
not indigenous, mycorrhizal fungi have probably first arrived in the roots of potted
plants. According to historians, early settlers often brought trees from their home
country and planted them around their new homes (Stephens and Kidd, 1953; Pryor,
1956). Thus, probably large numbers of living mycorrhizal seedlings of pine, oak, and
other European trees have been planted in South Africa, Australia, New Zealand and
Latin America as long as two or three hundred years ago. In those days there were no
plant quarantine regulations restricting the import of living plants.
Probably mycorrhizal fungi have also arrived in many countries through
botanical gardens. According to old records, the first specimens of many exotic trees
were brought to the gardens as potted plants.
Mycorrhizal infection may also have arrived as spores attached to imported
seed. In tropical and subtropical countries it is customary to extract pine seed in direct
sunshine right in the midst of the woods, where the seed can easily become
10
Introduction
contaminated with both fungal spores. Formerly there were no quarantine regulations
to require disinfection of imported seed. So far no definite proof is available that a
mycorrhizal fungus arrived anywhere as spores attached to imported seed. Many
observations, however, strongly support such an assumption (Singh and Kumar, 1966;
Perry et al, 1982; Pigott, 1982,).
It is often impossible to determine with certainty the origin of mycorrhizal
infection in nurseries which have not been intentionally inoculated. In countries where
inoculated nurseries and plantations already exist, infection may be transported to new
ones through spore flight or, for instance soil adhering to tools, car tires or shoes of
foresters moving from one nursery to another.
1.7 Ecological Importance of Mycorrhizae
The importance of mycorrhizae in ecosystems became particularly apparent in
the 1960s when plants grown in green houses were transplanted into areas such as
slag heaps, landfills, and strip-mined areas in order to reclaim the land. With few
exceptions, such plants did not survive in those infertile areas. The reason of this
failure lies in the fact that greenhouse soil is often sterilized to prevent the growth of
pathogens, and the sterilization process kills the mycorrhizal fungi and other growth
promoting microorganisms as well. Today, such reclamation attempts are much more
successful because mycorrhizal fungi are inoculated with the plants when they are
transplanted into the reclaimed areas. Similarly, attempts to grow certain species of
European pines in the United States were unsuccessful until mycorrhizal fungi from
their native soils were added at the time of transplanting (Mikola, 1970, 1973, Marx,
et al, .1985. Mohan, 1991, 2004).
Mycorrhizae are thought to have played an important role in the colonization
of the land by plants some four hundred million years ago (Harley, 1959). Studies of
fossil plants have shown that endomycorrhizae were prevalent at that time, and such
associations may have been crucial in helping plants make the transition from the
nutrient-rich sea to the nutrient-poor land (Mohan, 2004; Bakshi, 1974; Mitchell, et
al.1937).
11
Introduction
Nutritional Study of Mycorrhizal Fungi:
Mycorrhizal roots lack root hairs the fungal sheath extending to the soil
absorb nutrients (Ruehle, 1983). The Hartig net acts as a liaison tissue between the
fungal sheath and host cells. The ectomycorrhizal habit increases the surface area of
the root system and hence affords better intake of nutrients such as nitrogen,
phosphorus and potassium from the surrounding soil, using intact plant as well as
excised roots of pine (Pinus radiata) many workers have demonstrated the transfer of
isotopically labeled phosphorus, nitrogen calcium and sodium from the soil into the
roots through the fungal mycelium. The labeled isotopes were detected in all parts of
the plants including leaves (Bowen, et al., 1974).
The pattern of movement of ions has been investigated in ectomyorrhizal
fungus of Pinus sylvestris. The mycorrhizal association of this large rooted species
can easily be distinguished into an outer fungal sheath and an inner host core. The
host core consists of the Hartig net of the fungus between of the host. Experiment was
done by Harley and his Associates in England with excised mycorrhizal roots. Results
have shown that 80-90% of the absorbed phosphate remains accumulated in the
fungal sheath (Bowen, et al., 1974).
Melin and Nisson (1950) reported that the phosphate accumulation in the
fungal sheath may vary in intact as well as decapitated plants depending on the rates
of transpiration. Since decapitated plants accumulated more of phosphates in the
sheath than intact ones. It is likely that the sheath acts as a phosphate reservoir and
releases the nutrients during certain deficient conditions while under normal
conditions a steady uptake of phosphate to the plants is maintained by the fungal
hyphae.
1.8 Role of Ectomycorrhizal fungi in biocontrol
Ectomycorrhizal fungi are known to enhance the uptake of water and plant
nutrients, increases resistance to root pathogens and promote plant growth (Maccomb
and Griffith, 1946; Linderman, 1988). Initial
evidence
for
the
role
of
ectomycorrhizal fungi in disease suppression was provided by a number of
field observation that showed seedling or trees of both angiosperms and
12
Introduction
gymnosperms to be more resistant to root pathogens than their non-mycorrhizal
counterparts (Kope and Fortin, 1989).
As with other biological control methods of soil-borne diseases , the
major problem associated with the use of ectomycorrhizal fungi as biocontrol
agents is their poor survival and slow growth rate. To date the protective influence
of ectomycorrhizal fungi has been found to be extremely variable, therefore,
unreliable for large-scale biocontrol. It is thought that a clearer understanding of
ectomycorrhizal fungal physiology will help optimize the use of these organism
as biocontrol agents (Marx, 1969a,b,c).
Keeping the above in view the present study has been undertaken with the
following objectives.
1.9 Objectives
1.
Survey, collection and identification of ectomycorrhizal fungi and other
growth promoting microbes in laboratory.
2.
To isolate and culture ectotrophic mycorrhizae and other growth promoting
microbes in laboratory.
3.
To study the relationship between mycorrhizae and growth of sal seedling.
4.
To develop a consortium of inoculum including mycorrhizal fungi and other
growth promoting microbes for sal.
13