Mycorrhiza

Introduction, isolation, characterization,

screening, mass production, advantages and
application.

Presented by:
Sandeepkumar CH (UG14AGR1989)
 MYCORRHIZA

• The word mycorrhiza was coined by the German

scientist Albert Bernhard Frank in 1885.
• The word mycorrhiza is derived from the Greek words –
‘mukes’ meaning fungus and ‘rhiza’ meaning roots.
• Mycorrhiza (fungus-root) can be defined as a symbiotic
association between fungi and plant roots.
 Importance
• 95% of all the world's plant species form mycorrhizal
relationships with fungi and that in the majority of cases the plant
would not survive without them.
• Present in 95% of plants (83% Dicots, 79% Monocots and 100%
Gymnosperms).
• Brassicaceae, Cyperaceae, and Juncaceae- do not have
mycorrhizal associations (10-20%).
• The Orchidaceae are notorious as a family in which the absence
of the correct mycorrhizae is fatal even to germinating seeds
 Importance
• Mycorrhizae have existed for a very long time and can be
demonstrated in the fossilized roots of some of the earliest land
plants.
• Some scientists have suggested that plants were only able to move
on to land when they had developed mycorrhizal relationships
with fungi.
 Classification of mycorrhiza
Based on tropic level by A.B. Frank
• Ectotropic Mycorrhiza
• Endotropic Mycorrhiza
Based on morphological and anatomical feature
• Ectomycorrhiza
• Endomycorrhiza
• Ectendomycorrhiza
 Broadly classified into (Mark)
• Ectomycorrhiza (EcM)
• Endomycorrhiza (AM / VAM)
• Ectendomycorrhiza
• Monotropoid mycorrhiza
• Arbutoid mycorrhiza
• Orchid mycorrhiza
• Ericoid mycorrhiza
 Ectomycorrhiza or ectotrophic
mycorrhiza (EcM)
• Ectomycorrhizas, or EM, are typically formed between the roots
of around 10% of plant families, mostly woody plants including
the birch, dipterocarp, eucalyptus, oak, pine, deodar and rose
families , orchids, and fungi belonging to the Basidiomycota,
Ascomycota and Zygomycota.
• Commonly associated with trans temperate forest trees.
• Ectomycorrhizal fungi form a sheath or mantle around the root,
and hyphae emanate through the soil increasing the surface area.
• The fungus grows within the root cell wall but never penetrates the
cell interior.
• It grows between the cells of the cortex to form Hartig net.
• The Hartig net present outside the endodermis and meristematic
zones is the site for nutrient exchange.
• Colonization of root tips induces marked changes in the host
root morphology.
Fungus forming ectomycorrhizae
• Amanita muscaria
• Boletus variegatus
• Paxillus invalutus
• Rhizopogon vinicolor
• Entomoloma Amanita muscaria Entomoloma
• Sclerodendran
Ectomycorrhiza
Advantages of ectomycorrhiza
• Extensive multibranching hyphae increases the water holding
capacity of plants.
• Increase the tolerance to drought, high soil temperature, organic
and inorganic soil toxins, extremes of soil acidity to sulphur and
aluminium.
• Deter infection of feeder roots by some rot pathogens.
• Enhance the uptake of many nutrients. (P, Cu, Zn through Hartig
net)
• Disease control through barrier effect, competitive exclusion.
• Play a key role in afforestation.
 Endomycorrhiza or endotrophic
mycorrhiza
• Arbuscular mycorrhizae (often called AM) are the most common
and widespread of all mycorrhizae and are found in as many as
85%-90% of the world's plant species.
• Commonly associated with agricultural, horticulture crops in
addition to tropical trees.
• The external hyphal mantle or sheath is absent or scanty. The
fungal hyphae enters inside the root cortex and penetrates the
cortical cells.
• This is not a destructive parasitic association but endomycorrhiza
are present at certain times as a part of normal root development.
• AM fungi penetrate the cell walls of root cells.
• They grow between the cell wall and cell membrane forming
arbuscules.
• VAM fungi produce vesicles for lipid storage.
 Two main types of root colonization in
arbuscular mycorrhizae (AM).

A: Arum-type B: Paris-type

1: extraradical hyphae; 2: appressorium/hyphopodium; 3: arbusculum; 4:

vesiculum; 5: intercellular hyphae; 6: intracellular hyphae; 7: hyphal coils.
 Two main types of root colonization in
arbuscular mycorrhizae (AM).
• In the Arum-type the fungal hyphae grow intercellularly and
well-developed arbuscules are formed on branches entering the
neighboring cells.
• In the Paris-type the hyphae grow intracellularly, develop hyphal
coils in some cortical cells and smaller arbuscules develop on
these coils. Both the fungal and the plant partner influence the type
developed
 Fungi forming endomycorrhizae
• Endogone
• Glomus
• Sclerocystis
• Acaulospora
• Gigaspora Glomus Gigaspora
• Enterophophora
• Scutellispora
 Endomycorrhizae Ectomycorrhizae
Generally fungi produce its typical Fungi produce majority of its
structures, vescicles and arbuscules structure outside the root system.
inside the root system.
Commonly associated with Commonly associated with trans
agricultural, horticultural and temperate forest tree roots.
tropical trees.
Have a loose network of hyphae in Form a complete mantle or sheath
the soil and an extensive growth over the surface of the rot and
within the cortex cells of the plants. hyphae grows out into the soil.
Cannot be cultured on artificial Can be cultured on artificial media.
media.
Doesn’t cause morphological Cause morphological changes in
changes in roots. roots.
 Ectendomycorrhiza
• They share the features of both ecto- and endomycorrhiza.
• They have less developed hyphal mantle.
• The hyphae within the host penetrate its cells and grow within.
• These are found in both angiosperms and gymnosperms.
• Fungus associated are Ascomycetes.
• Hosts are Eucalyptus, Salix, Alnus etc.
 Monotropoid mycorrhiza
• The family Monotropaceae, which includes achlorophyllous
plants, develop the association.
• These plants entirely depend upon the fungus for
carbon and energy.
• Sheath, inter- and intracellular hyphae and peg-like haustoria
are present.

Monotropa sp.
 Arbutoid mycorrhiza

• These are found in the family Ericaceae.

• The fungi penetrate into the cortical cells forming extensive coils
of hyphae.
• The mycosymbionts are Basidiomycetes.
• Sheath, inter- and coiled intercellular hyphae are present.
 Orchid mycorrhiza
• At some point of time all Orchids are infected by orchidaceous
mycorrhiza – Basidiomycota.
• Orchids germinate only after infection by mycorrhiza. Ex:
Rhizoctonia sp.
• Within cells, hyphae form coils called pelotons which greatly
increase the interfacial surface area between orchid and fungus.
Cymbidum orchid Orchid pelotons stained Scaning
red in a light micrograph electronmicrograph of
of sectioned tissue. orchid pelotons
 Ericoid mycorrhiza
• This type occurs in the family Ericaceae.
• These plants have fine roots and the fungal members of
ascomycetes like Pezizella, Clavaria forms the association in the
outer region of cortical layer of roots.
• The ericoid fungal hyphae form a loose network over the hair root
surface the hyphae can also penetrate the epidermal cells, often at
several points in each cell and coiled hyphae fill the cell.
• Up to 80% of root volume can be fungal tissue and it is through
these coils that nutrient exchange is thought to occur
 Mycorrhiza Host range Types of relationship
Sheath, intercellular
Ectomycorrhiza Gymnosperms and Angiosperms
hyphae
Coiled intracellular hyphae, vesicle and
Endomycorrhiza (VAM) All groups of plant kingdom
arbuscules present

Ectendomycorrhiza Gymnosperms and Angiosperms Sheath optional, inter and intracellular hyphae

Sheath, inter and coiled intracellular hyphae

Monotropoid mycorrhiza Very restricted, Monotropaceae

Sheath, inter and coiled intracellular hyphae

Arbutoid mycorrhiza Very restricted, Ericales

Only coiled intracellular hyphae

Orchid mycorrhiza, Restricted, Orchidaceae

No sheath, no intercellular
Ericoid mycorrhiza Very restricted, Ericales hyphae, long,
coiled
 Isolation of Vesicular-Arbuscular
Mycorrhizal (VAM) spores from the soil

A. Sieving method
Requirements
• Soil sample
• 500 ml beaker
• Sieves of 710 µm, 250 µm, 75 µm and 45 µm.
• Bunsen burner
Procedure
• Take 200 ml water in 500 ml beaker.
• Heat the water to 40-50˚ C.
• Add 50 g of soil and mix well to form a suspension.
• Allow the heavier particles to settle down.
• Decant most of the suspension through a 710 µm sieve to remove
large organic matter and roots.
Procedure
• Add 200 ml of water to the suspension.
• Decant the suspension through 710 µm sieve.
• Decant this through 250 µm, 75 µm and 45 µm sieves
consequently.
• Collect the residue on the 45 µm sieve.
• Wash the residues well with water and collect the spore.
B. Floatation method
Requirements
• Soil sample
• Sucrose solutions (20, 40 and 60 %)
• Blender
• Fine sieve
• Centrifuge
• Centrifuge tube (50 ml)
Procedure
• Collect fresh soil samples from the field, mix them well and weigh
20 g soil.
• Transfer the soil into a blender.
• Blend it at high speed for 1-2 minutes so that the spores attached
to the soil particles or roots may become free.
• Filter the contents through a fine sieve and wash with strong
stream of water.
• Pour 10 ml of 20% sucrose into a centrifuge tube followed by the
same amount of 40% and 60% sucrose into the bottom of the tube.
 Procedure
• Take 10-15 ml of blended sieving and add onto the surface of 20%
sucrose layer.
• Centrifuge the contents for 3 minutes at 3000 rpm. Thereafter,
remove the debris which accumulate at the interfaces of 20-40%
and 40-60% of sucrose.
• Gently wash the spores present on fine sieve with a strong stream
of water so that sucrose should be removed.
• Collect the spores and observe under microscope.
Characterisation of Spores
 Mass Production : Problems and
prospects
• Being obligate symbionts AM fungi could be mass produced only
in the presence of living roots.
• Since AM fungal associations are universal and have been reported
in almost all terrestrial plants, these can be reproduced on a wide
range of host plants.
• There are several techniques reported for mass production of AM
inoculum.
 a. In vivo culture

• AM fungi are grown on roots of green house plants and chopped

mycorrhizal roots, often mixed with growth media containing
hyphae and spores, are used as source of inoculum.
• Soil could be replaced by inert substances such as vermiculite,
perlite, sand or a mixture of these for crude inoculum
production.
Mass production of VAM
Method of production

1. Tank for mass 2. Sprinkling of water in 3. Making of furrows to

multiplication of tank with vermiculite sow maize seeds
AM
Method of production

4. Sowing the 5. View of the maize 6. Vermiculite

seeds in furrows sown AM pit contained raised AM
infected maize plants
 b. In vitro/ axenic culture techniques

• i) Solution culture
• ii) Aeroponic culture
• iii) Root organ culture
i) Solution culture

• Involves growing infected roots in aqueous medium enriched with

mineral nutrients required for the growth of the roots under
controlled biotic and abiotic conditions.
ii) Aeroponic culture
Involves applying a fine mist of nutrient solutions to colonized roots
for AM fungal inoculum production.
iii) Root organ culture

• Use of a modified agar medium (MS rooting medium)/ liquid

medium for creation of increased amount of roots from callus
tissue and these roots are infected by AM spores or by surface
sterilized root bits obtained from mycorrhizal plant.
 Crop Mycorrhiza species
Barley, maize, wheat Glomus spp.
Bean Asaulospora morrowiae, Glomus, Gigaspora
Peanut Glomus fasciculatum, Sclerocystis dussi
Pea Glomus intraradices
Cotton Glomus sp., Sclerocystis sinuosa
Tomato, potato Gigaspora margarita, Glomus spp., Acaulospora sp.

Black pepper Entrophosphora colombiana, Scutellospora sp.

Cardamom Glomus fasciculautm
Citrus Glomus faciculatum, G. mosseae
Marigold Glomus mosseae
 Benefits of mycorrhiza
• Produce more vigorous and healthy plants.
• Increase plant establishment and survival at seedling or
transplanting.
• Enhance flowering and fruiting.
• Increase yields and crop quality.
• Improve drought tolerance, allowing watering reduction.
 Benefits of mycorrhiza
• Optimize fertilizers use, especially Phosphorus.
• Increase tolerance to soil salinity.
• Reduce disease occurrence.
• Contribute to maintain soil quality and nutrient cycling.
• Contribute to control soil erosion.
No mycorrhizal treatment Mycorrhizal treatment No mycorrhizal treatment Mycorrhizal treatment
 Application of VAM fungi

• Nursery application
100 g bulk inoculum is sufficient for one m2. The inoculum should
be applied a 2-3 cm below the soil at the time of sowing. The
seeds/cuttings should be sown/planted above the VAM inoculum to
cause infection.
 Application of VAM fungi

• For polythene bag raised crops

5 to 10 g bulk inoculum is sufficient for each packet. Mix 10 kg of
inoculum with 1000 kg of sand potting mixture and pack the potting
mixture in polythene bag before sowing.
 Application of VAM fungi
• For out-planting
20 g of VAM inoculum is required per seedling. Apply inoculum at the
time of planting.
• For existing trees
200 g VAM inoculum is required for inoculating one tree. Apply inoculum
near the root surface at the time of fertilizer application.
Few mycorrhizal products available
commercially
 REFERENCES
• Applied Microbiology – Moshrafuddin Ahmed & S K Basumatary
• Experiments in Microbiology, Plant Pathology and Biotechnology
– K R Aneja
• Practical Microbiology – R C Dubey and D K Maheshwari
• Bioinoculants – A Step Towards Sustainable Agriculture – R P Gupta,
Anu Kalia & Shammi Kapoor
• Biofertilizer Technology - S S Sandhu
 REFERENCES
Internet
• http://elte.prompt.hu/sites/default/files/tananyagok/StructureOfPlantsAn
dFungi/ch08s05.html
• http://www.davidmoore.org.uk/assets/mostly_mycology/diane_howarth
/monotropoid.html
• http://www.mycosym.com/EN/MycorrhizaEN.html
• https://www.google.co.in/search?q=ECTOMYCORRHIZA&dcr=0&so
urce=lnms&tbm=isch&sa=X&ved=0ahUKEwj_666P0ofZAhVMRY8K
HUOEDw0Q_AUICigB&biw=1366&bih=588#imgrc=b-
WD_IuzKXNP2M:
 Presented by

• Sandeep Kumar CH
• UG14AGR1989
• College of Agriculture, University of Agricultural Sciences, Raichur.
THANK YOU