Environmental Engineering Reference
In-Depth Information
bottle containing sterilized water and brought to
the laboratory safely for observation.
after one transfer to the medium, additional
transfers usually ensure pure cultures.
2. Small bits of boiled hempseed cotyledons are
added to an actively growing gross culture and
the fungus is allowed to develop zoosporangia
on this fresh bait. A micropipette is placed
opposite a discharge pore and the zoospores
are collected as they emerge from the zoospo-
rangium. The zoospore suspension, thus col-
lected, is then placed within the glass ring
partially submerged in the glucose-glutamate
agar (Table 12.2 ), and the plate is incubated at
18-22 °C. When the hyphae appear outside
the glass ring, they are removed along with a
small block of agar and transferred to a Petri
dish containing enough sterilized, deionized
water to cover the block. The fungus, develop-
ing in this culture, is bacteria-free.
From Soil Samples
About 10 g weight of collected soil is placed in a
fresh bottle containing 25 ml distilled deionized
water and is shaken vigorously for few minutes
and then poured into the Petri dish to a depth of
about 1 cm. After the particulate matter has set-
tled down within 1-2 h, 4-5 boiled hempseed
cotyledons are fl oated on the water and are incu-
bated at 20-22 °C. Following the appearance of
hyphal growth on the hempseed baits within
48-72 h, they are transferred to sterilized
Corning Petri dishes containing sterilized, deion-
ized water and incubated at 18-20 °C and are
identifi ed on the formation of reproductive struc-
tures there.
The identifi cation and characterization of
these members must be based on pure cultures.
For the purpose two different techniques have
been used:
1. A young, actively growing colony is removed
from the gross culture, washed thoroughly
several times and blotted between sterilized,
dry pieces of fi lter paper to remove excess of
water and bacteria. A tuft of hyphae is cut
from this culture and is, then, transferred to an
isolation medium (Table 12.2 ) and incubated
at 18-20 °C. With the growth on the semi-
solid medium, bacteria-free hyphal tips are
removed aseptically along with a small block
of agar and transferred to the fresh media. If
the growth is not free from contamination
Endophytic Hyper-parasitism
It is beyond the scope of this article to deal in
extenso with the organisms parasitizing the mem-
bers of oömycetes. However, certain important
contributions in this fi eld have been given. The
fi rst substantial publication is that of Fischer
( 1882 ) describing the biology of parasitism on
water moulds. Another important contribution
explaining host-parasite relation is of Held ( 1972 ,
1973 , 1974 ). Members of oömycetes have been
found to be parasitized by several unicellular
forms like Phlyctochytrium (Sparrow and
Dogma, 1973 ; Johnson, 1975 , 1976 ; Milanez,
1967 ; Johnson and Howard 1972 ; Karling, 1976),
Rhizophydium (Beneke and Rogers, 1970;
Table 12.2 Composition of glucose-glutamate isolation and culture medium
EDTA
200 mg
Zinc chloride
40.0 mg
Pot. hypophosphate
87 mg
Ferric chloride
1.3 mg
Pot. dihydr. phos.
68 mg
DL-Methionine
50.0 mg
Mag. chloride
160 mg
Sod. monoglutamate
500.0 mg
Calcium chloride
66 mg
D -glucose
3.0 g
Manganese chloride
75 mg
Water
1.0 l
Modifi ed from Scott et al. ( 1963b )
Agar (Difco purifi ed): 15.0 g
pH adjusted to 6.5 with pot. hydroxide
 
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