Study of fungi:
Specimens are studied in one of the
two ways: those that cannot be grown under artificial conditions are studied
directly, and those that must or can be cultured or isolated are studied
indirectly.
1-Indirect study:
Many pathogens in plant material, soil and water must
first be isolated and obtained in pure culture before they can be studied.
These fungi are usually not visible to the unaided eye, although the disease
symptoms often are.
Isolation of fungi
Isolation from soil:
Dilution method:
In this technique1 gram(dry weight) of the SOIL to be studied is ground up (if necessary) and
dispersed in 9 ml of sterile water. One milliliter of this solution is
transferred to a second tube containing 9 ml of sterile water, resulting in a
0.01 dilution of the spore mass in the original material. The process is
repeated to yield dilutions of 0.001, 0.0001, and 0.00001 or even further if
necessary. A 1-ml portion from each dilution is pipetted to a separate Petri
dish, and cooled, melted agar medium poured over it. The plate should be moved
gently on the table in a figure-of-eight motion to effect proper dispersion.
Alternatively, the solution can be put on the surface of solidified medium and
spread evenly throughout.
After a few days incubation,
colonies will appear in varying densities, depending upon the amount of
dilution from the original material. The number of spores present in the
original sample can be calculated roughly by selecting the plates showing
40-100 colonies and writing down the colony count. With this information the
following calculation can be performed:
The accuracy of this
technique is low when only one plate is counted. There are numerous
contributing factors, including improper dispersion of spores during dilution,
failure to break up spore masses, or mutual inhibition of growth by certain
fungi. Greater accuracy is attained by doing several plates at the most
desirable dilution. This technique allow scientists to compare one soil with
another on a statistical basis.
Direct method:
It is a simple technique, requiring the placing of small
bits of the soil (0.005-0.015) on the surface of the agar or the pouring of
melted but cooled agar over the fragments .the particles are dispersed
throughout the medium by rotating the dish. After a few days incubation mould colonies appear on the surface, and
can be transferred into pure culture.