Germinability of sclerotia of Macrophomina phaseolina


Ayanru, D.K.G.; Green, R.J.J.

Canadian Journal of Botany 56(9): 1107-1112

1978


The germination of sclerotia of M. phaseolina [snap bean pathogen] from 3-20 day old PDA [potato dextrose agar] cultures incubated at 32.degree. C and 24.degree. C varied from 98 to 25% and 75 to 30%, respectively. Germinability of sclerotia from cultures of all ages tested was rapidly lost during storage in desiccators for 24 h at 6.degree. C or in screw-top vials at room temperature for 30 days. Germination of sclerotia stored in screw-top vials at 6.degree. C for 30 days declined less rapidly- There was a linear relationship between the size of the sclerotium and the number of germ tubes emerging from it. The role of sclerotium size in survival and what factors may be responsible for loss of germinability with increasing age of source culture are discussed.

Germinability
of
sclerotia
of
Macrophomina
phaseolina
1,2
D.
K.
G.
AYAN
RU
3
AND
R.
J.
GREEN,
JR.
Deparimeni
of
Bolany
and
Plata
Pathology,
Purdne
Universily,
Wes!
Loleyene,
IN,
U.S.A.
47907
Received
December
22,
1976
AYANRU,
D.
K.
G.,
and
R.
J.
GREEN,
JR.
1978.
Germinability
of
sclerotia
of
Macrophomina
phaseolina.
Can.
J.
Bot.
56:
1107-1112.
The
germination
of
sclerotia
of
Macrophomina
phaseolina
from
3-
to
20-day-old
PDA
cultures
incubated
at
32°C
and
24°C
varied
from
98
to
25%
and
75
to
30%,
respectively.
Germinability
of
sclerotia
from
cultures
of
all
ages
tested
was
rapidly
lost
during
storage
in
desiccators
for
24
h
at
6°C
or
in
screw-top
vials
at
room
temperature
for
30
days.
Germination
of
sclerotia
stored
in
screw-top
vials
at
6°C
for
30
days
declined
less
rapidly.
There
was
a
linear
relationship
between
the
size
of
the
sclerotium
and
the
number
of
germ
tubes
emerging
from
it.
The
role
of
sclerotium
size
in
survival
and
what
factors
may
be
responsible
for
loss
of
germinability
with
increasing
age
of
source
culture
are
discussed.
AYANRU,
D.
K.
G.,
et
R.
J.
GREEN,
JR.
1978.
Germinability
of
sclerotia
of
Macrophomina
phaseolina.
Can.
J.
Bot.
56:
1107-1112.
Les
auteurs
ont
etudie
la
germination
des
sclerotes
provenant
de
Macrophomina
phaseolina
cultive
sur
extrait
de
pomme
de
terre
&lose
et
add
itionne
de
dextrose.
La
germination
des
sclerotes
provenant
de
cultures
agees
de
3
a
20
jours
et
incubees
a
32°C
et
24°C
variait
respective-
ment
de
98
a
25%
et
de
75
a
30%.
Le
pouvoir
de
germination
des
sclerotes
provenant
des
cultures
de
tousles
ages
testes
se
perdait
rapidement
au
cours
de
l'entreposage
a
6°C
pendant
24
h
dans
des
dessicateurs,
ou
encore
a
la
temperature
de
la
piece
pendant
30
jours
dans
des
fioles
a
bouchon
filete;
dans
ce
dernier
cas,
le
pouvoir
de
germination
diminuait
morns
rapidement.
On
a
constate
('existence
d'une
relation
lineaire
entre
la
dimension
du
sclerote
et
le
nombre
de
tubes
germinatifs
qui
en
emergent.
Les
auteurs
discutent
le
role
de
la
dimension
du
sclerote
dans
la
survie,
ainsi
que
les
facteurs
possiblement
responsabl
es
de
la
baisse
du
pouvoir
de
germination
avec
('augmenta-
tion
de
l'age
de
la
culture
d'oU
le
sclerote
provient.
[Traduit
par
le
journal]
1107
Introduction
A
study
of
the
ontogeny
and
structure
of
sclerotia
of
Macrophomina
phaseolina
(Maubl.)
Ashby
(Wyllie
and
Brown
1970)
revealed
that
all
cells
of
sclerotia
grown
on
cellophane
discs
over
potato
dextrose
agar
(PDA)
for
14
days
appear
to
have
the
ability
to
germinate.
Actual
germination
capability
of
the
sclerotia
was
not
tested.
A
knowledge
of
the
effect
of
culture
age
on
germination
and
that
of
the
conditions
under
which
sclerotia
could
be
stored
for
extended
periods
of
time
without
rapid
loss
of
viability
could
be
of
practical
importance.
It
could
make
possible
the
use
of
a
single
source
of
in-
oculum
for
extended
studies
needing
such
unifor-
mity.
Sclerotia
of
M.
phaseolina
in
cucurbit
roots
may
survive
10
months
dry
storage
in
the
labora-
tory
(Chaffer
and
Akhtar
1968),
but
the
conditions
for
the
maturation
and
longevity
of
individual
sclerotia
are
unknown.
Sclerotia
of
M.
phaseolina
are
heterogeneous,
'Journal
series
paper
5749,
Purdue
University,
Agricultural
Experiment
Station,
West
Lafayette.
IN,
U.S.A.
-Research
supported,
in
part,
by
funds
from
the
Mint
Re-
search
Grant.
3
Present
address:
Department
of
Biological
Sciences,
Uni-
versity
of
Benin,
Benin
City,
Nigeria.
varying
in
shape
and
size
from
25
x
22.4
p.m
to
152
x
32
p.m
or
more
(Taubenhaus
1913).
Haigh
(1928)
found
at
least
three
classes,
as
indicated
by
the
constant
differences
in
size
of
the
sclerotia
that
occur
over
several
generations.
The
role
of
sclerotium
size
in
germination
and
its
influence
on
the
survival
in
soil
or
in
dry
storage
have
not
been
reported.
In
this
study,
we
report
on
the
germinabil-
ity
of
the
sclerotia
of
M.
phaseolina
as
affected
by
its
size,
specified
conditions
of
dry
storage,
and
the
age
of
the
culture
from
which
they
are
har-
vested.
Materials
and
Methods
An
isolate
of
M.
phaseolina
from
snap
bean
preserved
on
PDA
slants
at
4-6°C
was
used
in
the
study.
Sclerotia
of
the
fungus
were
produced
at
32
±
1°C
(type
H)
and
24
±
1°C
(type
L)
on
cellophane
discs
(cellulose
xanthate)
spread
over
PDA
in
petri
dishes
150
mm
diameter
x
20
mm
deep.
Cultures
were
harvested
after
3,
6,
10,
and
20
days
of
incubation,
respectively,
by
scraping
sclerotia
from
discs
with
a
sterile
spatula.
Harvested
sclerotia
were
air
dried
for
3
h
at
a
room
temperature
of
24°C.
To
detach
individual
sclerotia
from
agglutinated
masses,
the
dried
material
was
ground
in
a
mortar
with
a
pestle.
Sclerotia
were
segregated
into
different
size
categories
by
sieving
the
ground
material
through
nos.
100-, 120-,
140-,
160-,
180-,
200-,
and
325-mesh
sieves
(U.S.
standard
series)
having
openings
of
149,
125,
105,
91,
81,
74,
and
44
p.m,
respectively.
Sclerotia
retained
on
no.
100
sieve
were
discarded
after
weighing
them,
and
the
sclerotia
from
the
other
sieves
were
collected.
The
number
of
1108
CAN.
J.
BOT.
VOL.
56.
1978
TABLE
1.
The
effect
of
age
of
culture
incubated
at
32°C
on
the
size
and
weight
composition
of
sclerotia
following
3
h
air
drying
and
grinding
Sclerotia
sizes
Sclerotia
counts,
no.
sclerotia/0.01
g
%
composition
by
weight
Sieve
size
or
no.°
Sieve
openings,
gm
3
6
6
10
20
30
100
149
29
18
10
7
5
120
125
8
400+316
17
20
15
12
9
140
105
10
300+417
14
18
17
15
12
160
97
15
700+573
25
29
42
46
54
180
81
21
100+679
6
4
5
4
3
200
74
30
550+888
6
6
6
7
5
325
44
44
600+2999
3
5
5
9
12
°U.S.
standard
series.
bAge
of
cultures
in
days.
sclerotia/0.1
g
and
the
weight
of
sclerotia
in
each
size
category
were
determined.
For
storage
purposes,
0.05
g
of
sclerotia
of
the
different
classes
were
placed
in
24-ml
glass
screw-top
vials
and
stored
either
at
6°C
or
at
room
temperature.
The
germinability
(total
and
rate
of
germination)
of
sclerotia
in
each
vial
was
determined
after
5,
10,
20,
and
30
days
of
storage
at
the
two
temperatures.
For
germination
tests,
sclerotia
were
washed
thoroughly
in
six
changes
of
sterile
phosphate
buffer
(pH
6.5)
by
a
repeated
process
of
centrifugation
(1475
g)
for
3
min
and
decantation.
After
washing,
sclerotia
were
plated
by
spraying
them
on
cel-
lophane
discs
spread
over
PDA
in
petri
plates
using
Devilbiss
atomizers.
Some
300
±
35
sclerotia
were
sprayed
on
each
plate
following
appropriate
dilutions
with
sterile
deionized
water.
Incubation
was
at
32
±
1°C
for
15
h,
the
predetermined
time
required
for
90%
germination
of
viable
sclerotia.
To
fix,
pre-
serve,
and
stain
the
germ
tubes
of
sclerotia.
the
plates
were
sprayed
with
lactophenol
aniline
blue
and
stored
at
4°C
until
counts
olgermination
were
made.
Plates
were
divided
into
eight
sectors,
and
alternate
sectors
were
screened
under
the
Stereo
Zoom
7
microscope
(Bausch
&
Lomb)
at
a
140
x
magnification.
Of
200
sclerotia
examined
on
each
plate.
the
percentage
of
germinating
sclerotia
with
one
to
three
germ
tubes
(X)
and
those
with
four
or
more
(4+)
germ
tubes
(Y)
were
determined.
Germ
tubes
shorter
than
the
diameter
of
a
sclerotium
were
not
counted.
All
experiments
were
done
in
duplicate
and
repeated
three
times.
To
determine
whether
observed
poor
germination
of
10-
and
20-day-old
sclerotia
was
the
result
of
an
endogenous
or
en-
vironmentally
induced
dormancy,
the
following
was
done.
Sclerotia
were
leached
continuously
for
3
days
at
4°C.
using
the
millipore
filter
apparatus
method
(Emmatty
and
Green
1969).
As
alternatives
to
leaching,
sets
of
sclerotia
were
soaked
in
5
x
10
-3
M
concentration
of
either
ethanol.
methanol,
chloroform
(5
x
10
-4
Al),
or
deionized
water
for
3
days
at
4°C.
At
6-h
intervals.
the
germinability
of
treated
sclerotia
was
determined
by
plating
them
on
cellophane
discs
over
PDA
after
thorough
washing.
The
rates
of
germination
and
the
total
germination
of
the
10-
and
20-day-old
sclerotia
were
further
tested
by
spraying
washed
sclerotia
on
cellophane
discs
spread
on
washed
and
dried
Whatman
no.
1
filter
paper
soaked
in
solutions
of
the
following
sugars
and
amino
acids
at
5
x
10
A4
concentrations:
0-fructose,
D-glucose,
maltose.
D-ribose.
and
sucrose;
DL-
phe
nylalanine
and
DL-serine.
Results
For
all
cultures
of
different
ages
(Table
1),
the
greatest
weight
of
harvested
sclerotia
was
retained
on
the
no.
160
sieves
(97-p,m
pore
size).
Sclerotia
of
this
size
(between
97
and
105
pLm
diameter)
contri-
buted
approximately
25, 29,
42,
46,
and
54%
of
the
yield
from
cultures
incubated
for
3,
6,
10,
20,
and
30
days,
respectively,
whether
at
32°C
or
24°C.
On
the
average,
the
number,
weight,
and
size
variations
of
sclerotia
were
from
8400/0.01
g
for
sclerotia
having
the
size
range
of
120-140
p.m
to
44
600/0.01
g
for
sclerotia
with
the
size
range
of
40-70µm
(Table
1).
For
the
purpose
of
statistical
analysis,
all
germi-
nation
data
were
arcsin
transformed.
Germination
of
sclerotia
of
all
sizes
was
affected
by
the
ages
of
the
culture
from
which
they
were
harvested
(Fig.
1).
The
differences
in
germination
were
also
highly
significant
(1%
level).
Sclerotia
from
3-,
6-,
10-,
and
20-day-old
cultures
(type
H)
had
98,
55,
33,
and
25%
germination,
respectively.
At
these
corres-
ponding
culture
ages,
95,
38,
12,
and
10%
of
the
germinating
sclerotia
had
4+
germ
tubes.
Sclerotia
from
3-,
6-,
10-,
and
20-day-old
type
L
cultures
showed
75, 78,
38.
and
30%
germination,
respec-
tively,
with
65,
61,
15,
and
10%
of
those
germinat-
ing
having
4+
germ
tubes.
Incubation
temperatures
of
cultures
from
which
sclerotia
were
obtained
significantly
affected
both
the
rate
of
germination
and
the
total
germination
of
the
sclerotia.
Following
4-,
6-,
8-,
10-,
12-,
and
15-h
incubation,
sclerotia
from
3-day-old
type
H
cul-
tures
showed
5,
9,
37,
76,
89,
and
98%
germination,
respectively.
After
15-h
incubation,
90%+
of
the
germinating
sclerotia
had
4+
germ
tubes.
Com-
parative
rates
of
germination
of
sclerotia
from
3-day-old
type
L
cultures
at
the
corresponding
six
incubation
periods
were
0,
6,
23,
45,
59,
and
75%;
only
65%
of
those
germinating
after
15
h
incubation
had
4+
germ
tubes.
However,
the
reverse
was
the
case
for
the
germination
of
sclerotia
from
6-day-old
type
H
cultures
whose
total
germination
after
15
h
AYANRU
AND
GREEN
1109
100
-
;Sclerotic
with
4+
germ
tubes
U
Sclelotia
with
I—
3
germ
tubes
80
Type
H
Type
L
6
0-
k.
40-
J
k.
20
Io
20
20
3
ACE
Of
CUL
TURE
(DAYS)
FIG.
I.
The
percentage
germination
of
sclerotia
(overall
means
of
all
classes
tested)
from
cultures
of
different
ages
grown
at
32°C
(type
H)
and
24°C
(type
L)
following
a
3-11
air
drying,
grinding,
and
incubation
for
15
h
at
32°C.
incubation
was
only
55%,
with
38%
of
them
having
4+
germ
tubes,
while
sclerotia
from
6-day-old
type
L
culture
had
a
comparatively
higher
total
germina-
tion
of
75%,
with
61%
of
these
having
4+
germ
tubes.
No
significant
differences
(5%
level)
were
found
between
the
total
germination
and
the
germi-
nation
rates
of
sclerotia
from
both
type
H
and
L
cultures
10
days
old
and
older.
The
size
of
the
sclerotium
significantly
(5%
level)
affected
its
germination
(Table
2).
For
all
viable
sclerotia
(those
from
cultures
6
days
old
or
younger),
but
with
the
exception
of
sclerotia
from
6-day-old
type
L
cultures,
there
was
a
fairly
consis-
tent
decrease
in
germination
as
the
size
of
the
sclerotium
decreases.
The
overall
picture
of
sclero-
tial
germination
is
one
of
rapid
decline
with
age
of
source
culture.
However,
when
the
germination
data
of
the
different
sizes
of
sclerotia
from
cultures
of
different
ages
are
presented
separately,
a
new
picture
emerges
(Fig.
2).
The
germination
of
sclerotia
of
the
nos.
180,
200,
and
325
sieve
size
groups
from
type
L
cultures
peaked
at
the
age
of
6
days
compared
with
all
other
groups
of
sclerotia
with
highest
total
germination
at
3
days.
The
size
of
the
sclerotium
also
affected
the
ulti-
mate
number
of
germ
tubes
that
emerged
from
it
and
the
rate
of
their
radial
growth.
A
significant
(5%
level)
positive
correlation
(r
=
0.87)
was
found
between
the
size
of
the
sclerotium
and
the
total
number
of
germ
tubes
emerging.
The
germination
of
both
type
H
and
type
L
sclerotia
of
all
sizes
(from
both
type
H
and
type
L
cultures)
stored
at
room
temperature
declined
rapidly
(Fig.
3).
Overall
germination
of
all
sizes
of
sclerotia
from
3-day-old
type
H
cultures
stored
for
0,
5,
10,
20,
and
30
days
at
room
temperature
were
98,
83,
46, 41,
and
20%,
respectively.
Comparative
germination
of
sclerotia
of
this
group
stored
at
6°C
for
the
corresponding
time
period
were
98,
93,
84,
83,
and
78%,
respectively.
Similar
germination
rates
were
obtained
from
3-day-old
type
L
sclerotia
stored
at
identical
temperatures.
Germination
of
.1
20
0
0
0
—A
3
60
1110
CAN.
J.
BOT.
VOL.
56,
1978
TABLE
2.
Percentage
germination
of
sclerotia
of
different
sizes
harvested
from
cultures
of
different
ages
following
a
3-h
air
drying,
grinding,
and
incubation
for
15
h
at
32°C
Age
of
cultures,
days
3
6
10
20
Sclerotium
size
(gm)b
X`
Y
X
Y
X
Y
X
Y
Type
1-1°
125-147
2
95
21
54
16
22
16
24
105-125
2
96
20
50
19
14
13
15
97-105
2
92
26
33
18
12
17
10
81-97
1
94
25
32
18
9
11
7
74-81
4
86
29
23
18
8
15
7
44-74
9
70
27
22
21
2
17
3
Type
L
125-147
4
84
16
53
23
17
19
17
105-125
9
76
15
59
20
12
20
15
97-105
6
76
17
61
20
11
21
6
81-97
8
48
13
68
22
10
20
9
74-97
8
34
19
64
21
14
19
7
44-74
9
25
29
32
25
4
15
5
°Types
H
and
L
are
cultures
incubated
at
32
and
24°C,
respectively.
°The
respective
sizes
of
sclerotia
were
those
retained
on
nos.
120,
140,
160,
180,
200,
and
325
sieves
(U.S.
standard
series).
`Germination
classes:
X
=
sclerotia
with
one
to
three
germ
tubes;
Y
=
those
with
four
or
more
germ
tubes.
0
0
TYPE
H
A
A
A
TYPE
L
4
60
a
20
0
5
0
6
A
20
0
a
//
3
6
10
20
3
AGE
OF
CULTURE
FIG.
2.
The
comparative
effect
of
age
of
culture
on
the
germi-
nation
(%)
of
sclerotia
of
different
sizes
(1,
2,
3,
4,
5.
and
6
are
sclerotia
with
diameters
of
125, 105,
97,
81,
74,
and
44
respectively)
derived
from
cultures
incubated
at
32
±
1°C
(type
H)
and
24
±
1°C
(type
L).
160
3
do
_
0
Type
L
1
0
Type
H
0
-
0
r.#
0-0
A
—A
#
A
4#
0
N
0-0
#
0
Fro
a
2
Type
H
4
Type
L
-‹
0
10
0
10
STORAGE
LENGTH
(DAYS)
FIG.
3.
The
comparative
germination
of
sclerotia
of
different
sizes
(140.
160,
and
180
are
sclerotia
with
size
ranges
of
105-125,
97-105,
and
81-97
p.m,
respectively)
harvested
from
cultures
incubated
at
32°C
(type
H)
and
24°C
(type
L)
following
their
storage
in
screw-top
vials
for
varying
time
periods
at
6°C
(1
and
3)
and
24°C
(2
and
4).
sclerotia
from
6-,
10-,
and
20-day-old
type
H
cul-
tures
stored
at
room
temperature
for
30
days
dropped
from
75,
32,
and
22%
to
7,
5,
and
3%,
respectively.
Similar
declines
in
germination
rates
were
obtained
for
the
6-,
10-,
and
20-day-old
type
L
sclerotia
stored
under
the
same
conditions
of
temp-
erature.
However,
although
there
was
only
a
slight
decrease
in
germination
for
all
classes
of
sclerotia
stored
at
6°C,
the
decline
in
germination
was
very
rapid
when
sclerotia
were
stored
in
desiccators
containing
calcium
chloride
at
6°C.
Within
24
h,
the
rate
of
germination
of
sclerotia
from
3-day-old
type
H
cultures
stored
in
desiccators
at
6°C
dropped
from
98
to
12%.
Similar
rapid
decline
in
germina-
tion
rates
were
observed
for
sclerotia
from
types
H
and
L
cultures
of
all
ages
following
storage
in
desic-
cators
at
6°C.
The
germinability
of
sclerotia
from
the
10-
and
20-day-old
types
H
and
L
cultures
was
further
tested
by
incubating
them
at
23
and
32°C
for
24
h.
There
was
no
significant
improvement
in
the
ger-
mination
rates
of
sclerotia
incubated
for
24
h
com-
pared
with
those
of
sclerotia
incubated
for
15
h.
Assessment
of
percentage
germination
of
sclerotia
incubated
for
more
than
24
h
was
generally
imprac-
I00
0
6
r
A
'zt
60
0
A
ri
6
10
20
(DAYS)
AYANRU
AND
GREEN
1111
tical
and
inaccurate
because
of
fast
growing
and
spreading
germ
tubes.
There
was
also
no
significant
increase
in
the
germination
of
the
poorly
germinat-
ing
sclerotia
from
10-
or
20-day-old
cultures
sprayed
on
cellophane
discs
over
washed
filter
paper
soaked
in
individual
sugars,
amino
acids,
and
(or)
their
mixtures.
The
germination
rate
of
sclerotia
from
3-day-old
type
H
cultures
that
served
as
control
was
80%+
on
discs
over
filter
paper
moistened
with
deionized
water
or
with
the
organic
supplements.
Continuous
leaching
of
the
poorly
germinating
sclerotia
from
the
10-day-old
and
older
cultures
with
deionized
water
or
soaking
them
in
water,
ethanol,
methanol,
or
chloroform
for
up
to
3
days
at
4°C
did
not
significantly
improve
either
the
rate
of
germination
or
total
germination.
Discussion
The
sclerotium
of
M.
phaseolina
appears
to
ma-
ture
rapidly
within
3
days,
particularly
when
pro-
duced
in
cultures
incubated
at
30°C+.
Maturity
of
sclerotia
from
cultures
incubated
at
lower
tempera-
ture
of
23-25°C,
as
determined
by
germinability
and
resistance
to
air
drying,
appears
complete
within
6-7
days.
Such
sclerotial
maturation
is
much
more
rapid
than
the
14
days
previously
reported
(Wyllie
and
Brown
1970).
Also,
there
appears
to
be
no
obligatory
resting
period
for
the
sclerotium
since
germination
of
sclerotia
from
3-
or
6-day-old
cul-
tures
occurs
readily.
Observed
poor
germination
of
sclerotia
from
10-
and
20-day-old
cultures
is,
perhaps,
the
result
of
environmental
dormancy
since
ultrastructural
examination
of
14-day-old
sclerotia
(Wyllie
and
Brown
1970)
revealed
that
all
cells
of
the
sclerotium
are
capable
of
germination.
However,
an
exogenous
supply
of
nutrients
did
not
enhance
germination.
The
positive
correlation
between
increasing
age
of
cultures
and
increasing
proportions
of
sclerotia
retained
on
no.
160
sieves
(Table
1)
is
probably
due
to
this
being
the
most
typical
size
of
sclerotium
for
this
fungus
under
the
conditions
provided.
Aggre-
gates
of
sclerotia
from
older
cultures
were
more
readily
broken
apart
into
discrete
units
than
those
from
young
cultures.
This
did
not,
however,
affect
germination
since
the
germination
of
only
individu-
ally
separated
sclerotia,
rather
than
of
aggregates
with
two
or
more
sclerotid,
was
tested.
The
significantly
higher
germination
rates
of
sclerotia
from
3-day-old
type
H
cultures
over
those
of
type
L
may
be
due
to
the
different
stages
of
maturity
of
the
sclerotia.
By
the
3rd
day
of
incuba-
tion,
sclerotia
obtained
from
cultures
incubated
at
the
higher
temperature
were
apparently
mature,
resisted
the
3-h
air
drying
prior
to
mechanical
grind-
ing,
and
germinated
rather
readily.
Most
of
the
sclerotia
from
the
type
L
cultures
were
still
imma-
ture
after
3
days,
and
their
germinability
was
re-
duced
by
air
drying.
Sclerotia
from
3-
or
6-day-old
(type
H
and
L)
cultures
germinated
readily
when
tested
without
air
drying.
On
the
contrary,
sclerotia
from
10-
or
20-day-old
cultures
had
low
germina-
tion
rates
consistently
when
tested
with
or
without
air
drying.
The
effect
of
stage
of
maturity
on
germi-
nation
has
been
reported
(Cagley
1936)
at
least
for
nondiasporic
spores
of
other
fungi.
Differential
ef-
fects
of
incubation
temperature
on
rates
of
accumu-
lation
of
metabolic
products
in
sclerotia
is
a
proba-
ble
explanation
for
observed
differences
in
germi-
nation.
Initiation
of
sclerotia
depends
on
the
ac-
cumulation
of
metabolic
substances
(Wheller
and
Waller
1965).
Since
the
high
temperature
of
35°C
is
the
most
favourable
for
mycelial
growth
in
Sclerothan
sp.
(Norton
1953)
and
some
other
fungi,
it
appears
that
in
this
fungus
temperature
most
favourable
for
mycelial
growth
also
induces
the
production
of
early
maturing
most
viable
sclerotia.
The
loss
of
viability
(inability
to
germinate)
of
sclerotia
with
increasing
age
of
source
culture
is
due
partly
to
the
effect
of
desiccation.
The
rapid
decline
in
viability
of
sclerotia
stored
in
desiccators
at
6°C
and
in
vials
at
room
temperature
indicates
that
the
sclerotium
of
this
fungus
survives
poorly
under
dry
conditions.
In
this
respect,
the
sclerotia
of
M.
phaseolina
are
comparable
with
those
of
Phymatotrieum
oninirortn,
which
do
not
survive
drying
for
over
l'/,
h
in
the
air
of
the
laboratory
(King
et
al.
1931).
The
suggestion
has
been
made
(Hoffmaster
et
al.
1943)
that
the
"sclerotium
of
this
fungus
is
an
immature
pycnidium.
Also,
since
the
sclerotium
lacks
a
rind
and
its
cells
have
cyto-
plasmic
continuity
via
septal
pores
(Wyllie
and
Brown
1970),
sensitivity
to
desiccation
is
perhaps
not
unexpected.
Such
ultrastructural
features
suggest
that
a
lack
of
permeability
to
water,
gases,
and
nutrients
is
unlikely
to
be
the
cause
of
the
sclerotium's
decreased
viability
with
age
of
source
culture.
Endogenous
metabolic
inhibitors,
if
present
in
older
sclerotia,
were
not
detected
or
were
not
re-
moved
from
sclerotia
by
leaching
with
water
or
soaking
in
ethanol,
methanol,
chloroform,
or
deionized
water.
Also,
since
nutrient
amendments
did
not
significantly
increase
germination,
the
slow
rates
of
germination
and
low
total
germination
of
sclerotia
from
10-
and
20-day-old
cultures
may
be
attributable
to
normal
physiological
degeneration
resulting
from
senescence
as
has
been
reported
at
1112
CAN.
J.
BOT.
VOL.
56,
1978
least
for
nondiasporic
cultures
of
other
fungi
(Coch-
rane
1958).
Perhaps
the
accumulation
of
certain
amino
acids,
organic
acids,
and
lipids
in
sclerotia
from
older
cultures
poisons
the
cells
of
the
sclerotium.
Total
lipids
and
organic
acids,
while
remaining
constant
in
some
fungi,
have
been
shown
to
increase
with
age
in
cells
of
M.
phaseolina
(Gottlieb
and
Van
Etten
1966).
Lack
of
appropriate
enzyme
systems
for
catalizing
such
accumulated
metabolites
in
sclerotia
from
older
cultures
may
also
be
a
factor.
Ayanru
and
Green
(1974)
attri-
buted
inhibition
of
sclerotia]
germination
by
DL-
methionine
to
toxic
effects
and
poor
utilization
of
amendments,
such
as
lactose
and
L-cystine,
to
lack
of
appropriate
adaptive
or
constitutive
enzymes.
Since
larger
sclerotia
had
consistently
higher
germination
rates,
produced
more
numerous
faster
growing
germ
tubes,
and
presumably,
contain
greater
amounts
of
endogenous
nutrients
for
ger-
mination
than
smaller
sclerotia,
their
inoculum
po-
tential
will
also
obviously
be
greater.
Such
features
will
enhance
the
survival
in
soil
of
larger
sclerotia
and
sustain
their
viability
during
repeated
flushes
of
new
growth.
AYANRU,
D.
K.
G..
and
R.
J.
GREEN,
JR.
1974.
Alteration
of
germination
patterns
of
sclerotia
of
Macrophomina
phaseolina
on
soil
surfaces.
Phytopathology,
64:
595-601.
CAGLEY,
D.
M.
1936.
Spores
and
spore
germination
in
wild
cultivated
mushrooms
(Psalliom
spp.).
Trans.
Br.
Mycol.
Soc.
20:
225-241.
CHAFFER,
A.,
and
P.
AKHTAR.
1968.
Survival
of
Macro-
phomina
phaseoli
(Maubl.)
Ashby
on
cucurbit
roots
(Luffa
actaangulata).
Mycopathol.
Mycol.
Appl.
35:
245-248.
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V.
W.
1958.
Physiology
of
fungi.
John
Wiley
&
Sons.
Inc.,
New
York.
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-
r
-
rv,
D.
A..
and
R.
J.
GREEN,
JR.
1969.
Fungistasis
and
the
behaviour
of
yerlicillimn
albo-atram
in
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Phytopathol-
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59:
1590-1595.
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D.,
and
J.
L.
VAN
ETTEN.
1966.
Changes
in
fungi
with
age.
I.
Chemical
composition
of
Rhizoctorikt
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and
Sclerothon
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J.
Bacteriol.
91:
161-168.
HAIGH,
J.
C.
1928.
Macrophomina
phaseoli
(Maubl.)
Ashby.
The
pycnidial
stage
of
Rhizoctunia
batatico/a
(Taub.).
Trop.
Agric.
(Ceylon),
70:
77-79.
HOFFMASTER,
D.
E.,
J.
H.
MCLAUGHLIN,
W.
W.
RAY,
and
K.
S.
CHESTER.
1943.
The
problem
of
dry
rot
caused
by
Macrophomina
phaseoli.
Phytopathology,
33:
1113-1114.
KING,
C.
J.,
H.
F.
Loomis,
and
C.
HOPE.
1931.
Studies
on
sclerotia
and
mycelial
strands
of
the
cotton
root
rot
fungus.
J.
Agric.
Res.
43:
827-840.
NORFON,
D.
C.
1953.
Linear
growth
of
Sclerotion
bataticola
through
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Phytopathology,
43:
633-636.
TAUBENHAUS,
J.
J.
1913.
The
black
rots
of
the
sweet
potato.
Phytopathology,
3:
159-166.
WHELLER,
B.
E.
J..
and
J.
M.
WALLER.
1965.
The
production
of
sclerotia
by
Sclerothon
colysil.
IL
The
relationship
between
mycelia'
growth
and
initiation
of
sclerotia.
Trans.
Br.
Mycol.
Soc.
48:
303-314.
WYLLIE,
T.
J.,
and
M.
F.
BROWN.
1970.
Ultrastructural
forma-
tion
of
sclerotia
of
Macrophomina
phaseoli.
Phytopathology,
60:
524-528.