ANALISIS PENGARUH INOVASI PRODUK DAN PER
Mycopathologia (2005) 159: 153–157
Springer 2005
Enhancing biological control of basal stem rot disease (Ganoderma
boninense) in oil palm plantations
A. Susanto, P.S. Sudharto & R.Y. Purba
Indonesian Oil Palm Research Institute, Jl. Bigjen Katamso No. 51, Medan 20158, Indonesia
Abstract
Basal Stem Rot (BSR) disease caused by Ganoderma boninense is the most destructive disease in oil palm,
especially in Indonesia and Malaysia. The available control measures for BSR disease such as cultural
practices and mechanical and chemical treatment have not proved satisfactory due to the fact that
Ganoderma has various resting stages such as melanised mycelium, basidiospores and pseudosclerotia.
Alternative control measures to overcome the Ganoderma problem are focused on the use of biological
control agents and planting resistant material. Present studies conducted at Indonesian Oil Palm Research
Institute (IOPRI) are focused on enhancing the use of biological control agents for Ganoderma. These
activities include screening biological agents from the oil palm rhizosphere in order to evaluate their
effectiveness as biological agents in glasshouse and field trials, testing their antagonistic activities in large
scale experiments and eradicating potential disease inoculum with biological agents. Several promising
biological agents have been isolated, mainly Trichoderma harzianum, T. viride, Gliocladium viride,
Pseudomonas fluorescens, and Bacillus sp. A glasshouse and field trial for Ganoderma control indicated that
treatment with T. harzianum and G. viride was superior to Bacillus sp. A large scale trial showed that the
disease incidence was lower in a field treated with biological agents than in untreated fields. In a short term
programme, research activities at IOPRI are currently focusing on selecting fungi that can completely
degrade plant material in order to eradicate inoculum. Digging holes around the palm bole and adding
empty fruit bunches have been investigated as ways to stimulate biological agents.
Key words: biological control, Ganoderma boninense, oil palm, Trichoderma spp.
Introduction
During the last two decades there has been a rapid
expansion in the areas planted with oil palm.
Major oil palm developments in Indonesia include
not only Sumatra, but also Borneo, Celebes, Papua, and Banten. This expansion has involved
both the use forest land and the conversion of
existing plant plantations. One of the major constraints to oil palm cultivation is the presence of
disease. Of the diseases that occur in oil palm
plantations, Basal Stem Rot caused by Ganoderma
boninense is the most destructive [1, 2].
In several oil palm plantations in Indonesia,
BSR has caused large losses where 50% or more of
the productive plants have died [3], and currently
BSR is the major disease in oil palm plantations
[4]. Previously, G. boninense was only found on
older plants, but recently it has been found in
younger plants where symptoms appear earlier
and are more severe, leading to greater replanting.
Historically control of BSR has involved cultural techniques, and mechanical and chemical
control. The failure to control the disease is due to
the characteristics of G. boninense. The fungus is
soil borne and so fungicides may be ineffective due
to degradation in the soil before they can reach
their target. In addition, G. boninense also has
many forms of resting stages including resistant
mycelium, basidiospores, chlamydospores, and
pseudosclerotia. In order to combat these characteristics the best approach to the control of BSR
154
may be biological control and the utilisation of
resistant oil palms. Breeding for resistance is a
long-term activity, whereas biological control may
be developed over a shorter time scale.
Materials and methods
Collection of candidate fungal and bacterial
biocontrol agents
Isolation of biocontrol agent candidates was
undertaken from oil palm rhizospheres in North
Sumatra, West Sumatra, Lampung, and Banten.
The choice of plantation area was based on the
stage of development of the plants, the plantation
age, the plant condition and the history of the
plantation. Plant development stages that were
considered were immature plant, productive
plant, and old plant. The age of the plantation included first, second, third, and fourth generation
plantings.
The
history
of
the
plantation considered its previous usage, and
whether it had been used for forest, tea, cocoa,
rubber, and coffee. All of the observations were
conducted on healthy and infected plants where
possible. Isolation of candidate fungus biocontrol
agents was by dilution plating on Martin
Agar + chloramphenicol medium as described by
Johnson and Curl [5]. The fungi isolated were
counted and the population density was calculated
after 1 week. The fungi obtained were purified and
identified. A similar method was also applied to
the isolation of bacterial biocontrol agents, but
with serial dilutions to 10)7 plated on nutrient agar
(Table 1).
Table 1. Population and inhibition capacity of candidate biocontrol agents
No.
Isolate
Population/g soil
1
2
3
4
5
6
7
8
10
11
14
15
16
20
21
22
24
9
17
19
25
12
13
18
23
26
27
28
T. harzianum-1
T. harzianum-9
T. harzianum-11
T. harzianum-21
T. harzianum-26
T. harzianum-29
T. harzianum-34
T. harzianum-39
T. harzianum-45
T. harzianum-58
T. harzianum-88
T. harzianum-91
T. harzianum-95
T. harzianum-107
T. harzianum-112
T. harzianum-119
T. harzianum-131
G. viride-44
G. viride-98
G. viride-105
G. viride-136
T. viride-70
T. viride- 82
T. viride-102
T. viride-123
T. viride-138
Bacillus sp.-10
Pseudomonas
fluorescens-1
P. fluorescens-58
P. fluorescens-63
1.3
5.4
1.7
5.3
1.3
2.2
3.5
2.3
4.0
1.3
1.3
1.3
1.5
4.1
2.7
4.0
4.0
6.7
1.3
1.3
4.0
8.4
1.3
2.4
4.0
6.7
5.6
1.8
29
30
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
104
104
105
104
104
106
105
105
104
104
104
104
105
105
104
104
104
104
104
104
104
105
104
105
104
104
108
108
4.6 · 108
4.1 · 108
Inhibition (%)
Plantation area
85.6
92.0
83.8
91.1
87.0
84.0
87.7
93.0
96.0
92.6
94.6
96.7
93.0
96.1
92.7
94.5
97.8
92.0
87.6
85.9
84.0
84.0
90.0
89.1
90.0
92.1
54.2
49.6
Banten Sehat TBM
Banten Sehat TM
Banten Sakit TM
Lampung Sehat T III
Lampung Sakit T III
Sumbar Sehat TM I
Sumbar Sakit TM I
Sumut Sehat TBM IV
Sumut Sakit TBM IV
Sumut Hutan Sehat T
Sumut Sehat II TBM
Sumut Sakit II TBM
Sumut Sehat II TM
Sumut Sakit II T
Sumut Sehat III TBM
Sumut III TBM
Sumut Sakit III TM
Sumut Sakit TBM IV
Sumut Sakit II TM
Sumut Sehat II T
Sumut Sehat III T
Sumut Teh Sehat TBM
Sumut Karet Sehat T
Sumut Sehat II T
Sumut Sehat III TM
Sumut Sakit III T
Lampung Sakit TBM IV
Banten Sehat TBM I
jk
defg
k
efg
hijk
k
hij
bcdef
abc
cdef
abcd
a
bcdef
ab
bcdef
abcde
a
defg
hij
ijk
k
k
fgh
ghi
fgh
defg
l
m
42.2 n
50.4 m
Sumut Sakit II T
Sumut Sakit III TBM
Entries followed by the same letter are not significantly different by Duncan’s Multiple Range Test at the 5% level.
155
Table 2. Percentage of oil palm seedling death at 12 months after inoculation
Cross
Disease incidence (%)
Plant height (cm)
Frond numbers
Vigour
BO909 T · BO 936 T
BO 906 T · BO 796 P
M1314 D · BO 932 T
BO2581 D · BO2581 D
BO 5462 D · BO 358 P
BO 5585 D · BO 5510 D
0.00
2.78
1.11
6.11
2.78
1.67
88.52 c
103.60 a
97.18 b
60.65 d
100.55 ab
85.42 c
9.08 d
11.05 c
12.23 a
11.13 c
11.38 bc
11.63 b
2.98
2.88
2.92
2.58
2.80
2.87
b
b
b
a
b
b
a
a
a
b
a
a
Entries followed by the same letter are not significantly different by Duncan’s Multiple Range Test at the 5% level.
Inhibition effectiveness of candidate biocontrol
agents
A dual culture method was used to evaluate the
effectiveness of the candidate biocontrol agents.
After 4 days incubation this was calculated as a
percentage inhibition from the formula:
KP ¼
ab
100%
a
where KP = inhibition effectiveness, a = the
radius of the G. boninense colony without the
biocontrol agent, b = the radius of the G. boninense colony with the biocontrol agent (Table 2).
Efficacy of biocontrol agents in glasshouse trials
Plant materials used were BO909 T · BO 936 T,
BO 906 T · BO 796 P, M1314 D · BO 932 T,
BO2581 D · BO2581 D, BO 5462 D · BO 358 P,
and BO 5585 D · BO 5510 D. Sawdust + PDA
was used for the production of the G. boninense
inocula. Inocula were divided in to smaller aliquots of 163 cm3 in plastic measuring cylinders.
Seedlings were inoculated when they were transplanted from the pre-nursery to the main nursery.
One inoculated root of each oil palm seedling was
put into a plastic bag and tied, and this was then
covered with soil in the main nursery plastic bag.
The experimental design was a completely random
block design factorial of 6 · 5 · 2 · 2 with three
replications for each combination. The first factor
was type of cross used for the oil palm material
(where V1 = T · T, V2 = T · P, V3 = D · T, V4
= D · D selfing, V5 = D · P, V6 = D · D). The
second factor was the biocontrol agent used (T0 =
without biocontrol agents, T1 = T. harzianum, T2
= G. viride, T3 = Bacillus sp., T4 = T. harzianum
+ G. viride + Bacillus sp.). The third factor was
the G. boninense inoculation (G0 = without
inoculation and G1 = inoculation with G. boninense), and the fourth factor was the addition of
chitin (K0 = without chitin and K1 = plus
chitin). The biocontrol agents were added as 10 g
formulation for fungal biocontrol agents and
10 ml formulation for bacterial biocontrol agents.
For the T4 treatment, 5 g T. harzianum, 5 g G. viride, and 5 ml Bacillus sp. were used respectively. Chitin was added at 1 g per polybag. The
conidial density of T. harzianum and G. viride
preparations were 4 · 106 conidium/ml, whereas
Bacillus sp. was used at 7 · 108 cfu/ml. The variables recorded were disease incidence of basal stem
rot, plant height and number of leaves, plant vigour, and development of the biocontrol agent
population. Observations were made monthly for
1 year.
Results and discussion
Collection of candidate biocontrol agents
One hundred and forty fungi were isolated from
various oil palm rhizospheres and these included
18 isolates of T. harzianum, 5 isolates of T. viride, 4
isolates of Gliocladium viride, 28 isolates of
A. flavus, 4 isolates of A. niger, 13 isolates of A.
fumigatus, 14 isolates of Penicillium citrinum, 4
isolates of Rhizopus, 6 isolates of P. chrysogenum,
12 isolates of P. commune, and 32 isolates of
P. funiculosum. The majority of the isolates were
considered as candidate biocontrol agents. The
population densities of each species were similar
between plantations and were about 104–105 cfu/g
soils. Seventy two isolates of bacteria were obtained and these included isolates of Pseudomonas
fluorescens and Bacillus sp.
156
Inhibition activity of candidate biocontrol agents
Twenty six fungi and four bacteria showed potential biocontrol activities in dual culture analysis. Fungi were categorised as biocontrol agents if
they showed an inhibition capacity >80%, and
bacteria were considered biocontrol agents if they
showed an inhibition capacity of >40%. The isolate that gave the highest inhibition capacity was
T. harzianum (97.8%). This was isolated from
a third generation oil palm planting in
North Sumatra. The isolate was similar to another
T. harzianum isolate from a BSR infected
immature fourth generation planting in North
Sumatra, to an isolate from healthy immature
second generation planting in North Sumatra, to
an isolate from infected immature second generation planting in North Sumatra, and to an isolate
from infected productive first generation planting
in West Sumatra. T. harzianum was the most
common species among the candidate biocontrol
agents.
Only four bacteria showed potential biocontrol
properties, all of the bacteria gave lower inhibition
values than the fungi. The highest inhibition
capacity was shown by an isolate of Bacillus sp.
from Lampung (54.2%). The remaining bacterial
biocontrol agents all had inhibition capacities of
between 40 and 50%.
The presence of Trichoderma and Gliocladium
was influenced by soil porosity, soil pH, and other
chemical properties of the soil. The abundance of
Trichoderma in soil depends on its ability to degrade organic matter and its resistance to other
microorganisms. The differences in the abundancies of the biocontrol agents may also be influenced by the climatic conditions at each site.
Table 3. The effect of biocontrol agents on Basal Stem Rot
incidence and oil palm vigour
Treatment
Disease incidence (%) Plant vigour
Control
T. harzianum
G. viride
Bacillus sp.
T. harzianum +
G. viride +
Bacillus sp.
18.06 a
0.00 c
0.00 c
9.72 b
8.33 bc
2.67
3.00
3.00
2.71
2.82
b
a
a
b
b
Entries followed by the same letter are not significantly different
by Duncan’s Multiple Range Test at the 5% level.
The D · D cross may have the highest disease
incidence, due to selfing leading to an accumulation of susceptible traits, as selfing apparently
did not result in an accumulation of resistance
traits.
The addition of candidate biocontrol agents
(T. harzianum and G. viride) was found to significantly reduce Basal Stem Rot incidence (Table 3).
Disease was only seen in the negative controls and
with the Bacillus sp. and mixed inoculum. The
ability of Bacillus sp. to inhibit G. boninense appeared to be lower than either of the fungal biocontrol agents, and every test involving Bacillus sp.
gave higher disease incidences. In the mixed inocula it may be that the Bacillus sp. competed with
the T. harzianum and G. viride isolates.
The oil palms treated with T. harzianum and
G. viride were significantly taller than the negative
controls and the palms treated with Bacillus sp.
and the mixed treatments. Adding of T. harzianum
and G. viride also appeared to give significantly
higher plant vigour than the control, Bacillus sp.
and mixed species treatments.
Efficiency of biocontrol agents in glasshouse trial
The disease incidence of Basal Stem Rot was not
consistent for all oil palm crosses. The cross with
the highest disease incidence was the selfed D · D
at 6.11%, compared to the other crosses where the
disease incidence ranged from 1.11 to 2.78%.
The T · T had not shown any disease symptoms
by the end of the 1 year period. This does not
indicate that T · T was resistant to G. boninense,
as this cross is known to develop BSR in the field,
and one possibility is that the 1 year period was
not enough for the pathogen to infect T · T.
Conclusions
This study has isolated 30 candidate biocontrol
agents consisting of 17 isolates of T. harzianum, 4
isolates of G. viride, 5 isolates of T. viride, one
isolate of Bacillus sp., and 3 isolates of Pseudomonas fluorescents. After 1 year of inoculation,
T. harzianum and G. viride appeared to prevent
Basal Stem Root in glasshouse trials, whereas,
Bacillus sp. had very little capacity to prevent
infection by G. boninense.
157
References
1. Semangun H. Penyakit–penyakit tanaman perkebunan di
Indonesia. Yogyakarta: Gadjah Mada University Press,
1990: 808.
2. Treu R. Macro fungi in oil palm plantations of South East
Asia. J Gen Mycol 1998; 12: 10–14.
3. Turner PD. Oil Palm Diseases and Disorders. Oxford:
Oxford University Press, 1981: 280.
4. Darmono TW. Ganoderma in oil palm in Indonesia: current
status and prospective use of antibodies for the detection of
infection. In: Harman GE, Kubicek CP, eds. Trichoderma
and Gliocladium Volume 1: Enzymes, biological control and
commercial applications. UK: Taylor & Francis Ltd., 1998:
393.
5. Johnson LF, Curl EA. Methods for Research on the
Ecology of Soil Borne Plant Pathogens. Minnesota: Burgess
Publishing Company, 1972: 247.
Address for correspondence: A. Susanto, Indonesian Oil Palm
Research Institute, Jl. Bigjen Katamso No. 51, Medan 20158,
Indonesia
Phone: +61-7862466/77/50; Fax: +61-7862488;
E-mail: iopri@idola.net.id
Springer 2005
Enhancing biological control of basal stem rot disease (Ganoderma
boninense) in oil palm plantations
A. Susanto, P.S. Sudharto & R.Y. Purba
Indonesian Oil Palm Research Institute, Jl. Bigjen Katamso No. 51, Medan 20158, Indonesia
Abstract
Basal Stem Rot (BSR) disease caused by Ganoderma boninense is the most destructive disease in oil palm,
especially in Indonesia and Malaysia. The available control measures for BSR disease such as cultural
practices and mechanical and chemical treatment have not proved satisfactory due to the fact that
Ganoderma has various resting stages such as melanised mycelium, basidiospores and pseudosclerotia.
Alternative control measures to overcome the Ganoderma problem are focused on the use of biological
control agents and planting resistant material. Present studies conducted at Indonesian Oil Palm Research
Institute (IOPRI) are focused on enhancing the use of biological control agents for Ganoderma. These
activities include screening biological agents from the oil palm rhizosphere in order to evaluate their
effectiveness as biological agents in glasshouse and field trials, testing their antagonistic activities in large
scale experiments and eradicating potential disease inoculum with biological agents. Several promising
biological agents have been isolated, mainly Trichoderma harzianum, T. viride, Gliocladium viride,
Pseudomonas fluorescens, and Bacillus sp. A glasshouse and field trial for Ganoderma control indicated that
treatment with T. harzianum and G. viride was superior to Bacillus sp. A large scale trial showed that the
disease incidence was lower in a field treated with biological agents than in untreated fields. In a short term
programme, research activities at IOPRI are currently focusing on selecting fungi that can completely
degrade plant material in order to eradicate inoculum. Digging holes around the palm bole and adding
empty fruit bunches have been investigated as ways to stimulate biological agents.
Key words: biological control, Ganoderma boninense, oil palm, Trichoderma spp.
Introduction
During the last two decades there has been a rapid
expansion in the areas planted with oil palm.
Major oil palm developments in Indonesia include
not only Sumatra, but also Borneo, Celebes, Papua, and Banten. This expansion has involved
both the use forest land and the conversion of
existing plant plantations. One of the major constraints to oil palm cultivation is the presence of
disease. Of the diseases that occur in oil palm
plantations, Basal Stem Rot caused by Ganoderma
boninense is the most destructive [1, 2].
In several oil palm plantations in Indonesia,
BSR has caused large losses where 50% or more of
the productive plants have died [3], and currently
BSR is the major disease in oil palm plantations
[4]. Previously, G. boninense was only found on
older plants, but recently it has been found in
younger plants where symptoms appear earlier
and are more severe, leading to greater replanting.
Historically control of BSR has involved cultural techniques, and mechanical and chemical
control. The failure to control the disease is due to
the characteristics of G. boninense. The fungus is
soil borne and so fungicides may be ineffective due
to degradation in the soil before they can reach
their target. In addition, G. boninense also has
many forms of resting stages including resistant
mycelium, basidiospores, chlamydospores, and
pseudosclerotia. In order to combat these characteristics the best approach to the control of BSR
154
may be biological control and the utilisation of
resistant oil palms. Breeding for resistance is a
long-term activity, whereas biological control may
be developed over a shorter time scale.
Materials and methods
Collection of candidate fungal and bacterial
biocontrol agents
Isolation of biocontrol agent candidates was
undertaken from oil palm rhizospheres in North
Sumatra, West Sumatra, Lampung, and Banten.
The choice of plantation area was based on the
stage of development of the plants, the plantation
age, the plant condition and the history of the
plantation. Plant development stages that were
considered were immature plant, productive
plant, and old plant. The age of the plantation included first, second, third, and fourth generation
plantings.
The
history
of
the
plantation considered its previous usage, and
whether it had been used for forest, tea, cocoa,
rubber, and coffee. All of the observations were
conducted on healthy and infected plants where
possible. Isolation of candidate fungus biocontrol
agents was by dilution plating on Martin
Agar + chloramphenicol medium as described by
Johnson and Curl [5]. The fungi isolated were
counted and the population density was calculated
after 1 week. The fungi obtained were purified and
identified. A similar method was also applied to
the isolation of bacterial biocontrol agents, but
with serial dilutions to 10)7 plated on nutrient agar
(Table 1).
Table 1. Population and inhibition capacity of candidate biocontrol agents
No.
Isolate
Population/g soil
1
2
3
4
5
6
7
8
10
11
14
15
16
20
21
22
24
9
17
19
25
12
13
18
23
26
27
28
T. harzianum-1
T. harzianum-9
T. harzianum-11
T. harzianum-21
T. harzianum-26
T. harzianum-29
T. harzianum-34
T. harzianum-39
T. harzianum-45
T. harzianum-58
T. harzianum-88
T. harzianum-91
T. harzianum-95
T. harzianum-107
T. harzianum-112
T. harzianum-119
T. harzianum-131
G. viride-44
G. viride-98
G. viride-105
G. viride-136
T. viride-70
T. viride- 82
T. viride-102
T. viride-123
T. viride-138
Bacillus sp.-10
Pseudomonas
fluorescens-1
P. fluorescens-58
P. fluorescens-63
1.3
5.4
1.7
5.3
1.3
2.2
3.5
2.3
4.0
1.3
1.3
1.3
1.5
4.1
2.7
4.0
4.0
6.7
1.3
1.3
4.0
8.4
1.3
2.4
4.0
6.7
5.6
1.8
29
30
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
·
104
104
105
104
104
106
105
105
104
104
104
104
105
105
104
104
104
104
104
104
104
105
104
105
104
104
108
108
4.6 · 108
4.1 · 108
Inhibition (%)
Plantation area
85.6
92.0
83.8
91.1
87.0
84.0
87.7
93.0
96.0
92.6
94.6
96.7
93.0
96.1
92.7
94.5
97.8
92.0
87.6
85.9
84.0
84.0
90.0
89.1
90.0
92.1
54.2
49.6
Banten Sehat TBM
Banten Sehat TM
Banten Sakit TM
Lampung Sehat T III
Lampung Sakit T III
Sumbar Sehat TM I
Sumbar Sakit TM I
Sumut Sehat TBM IV
Sumut Sakit TBM IV
Sumut Hutan Sehat T
Sumut Sehat II TBM
Sumut Sakit II TBM
Sumut Sehat II TM
Sumut Sakit II T
Sumut Sehat III TBM
Sumut III TBM
Sumut Sakit III TM
Sumut Sakit TBM IV
Sumut Sakit II TM
Sumut Sehat II T
Sumut Sehat III T
Sumut Teh Sehat TBM
Sumut Karet Sehat T
Sumut Sehat II T
Sumut Sehat III TM
Sumut Sakit III T
Lampung Sakit TBM IV
Banten Sehat TBM I
jk
defg
k
efg
hijk
k
hij
bcdef
abc
cdef
abcd
a
bcdef
ab
bcdef
abcde
a
defg
hij
ijk
k
k
fgh
ghi
fgh
defg
l
m
42.2 n
50.4 m
Sumut Sakit II T
Sumut Sakit III TBM
Entries followed by the same letter are not significantly different by Duncan’s Multiple Range Test at the 5% level.
155
Table 2. Percentage of oil palm seedling death at 12 months after inoculation
Cross
Disease incidence (%)
Plant height (cm)
Frond numbers
Vigour
BO909 T · BO 936 T
BO 906 T · BO 796 P
M1314 D · BO 932 T
BO2581 D · BO2581 D
BO 5462 D · BO 358 P
BO 5585 D · BO 5510 D
0.00
2.78
1.11
6.11
2.78
1.67
88.52 c
103.60 a
97.18 b
60.65 d
100.55 ab
85.42 c
9.08 d
11.05 c
12.23 a
11.13 c
11.38 bc
11.63 b
2.98
2.88
2.92
2.58
2.80
2.87
b
b
b
a
b
b
a
a
a
b
a
a
Entries followed by the same letter are not significantly different by Duncan’s Multiple Range Test at the 5% level.
Inhibition effectiveness of candidate biocontrol
agents
A dual culture method was used to evaluate the
effectiveness of the candidate biocontrol agents.
After 4 days incubation this was calculated as a
percentage inhibition from the formula:
KP ¼
ab
100%
a
where KP = inhibition effectiveness, a = the
radius of the G. boninense colony without the
biocontrol agent, b = the radius of the G. boninense colony with the biocontrol agent (Table 2).
Efficacy of biocontrol agents in glasshouse trials
Plant materials used were BO909 T · BO 936 T,
BO 906 T · BO 796 P, M1314 D · BO 932 T,
BO2581 D · BO2581 D, BO 5462 D · BO 358 P,
and BO 5585 D · BO 5510 D. Sawdust + PDA
was used for the production of the G. boninense
inocula. Inocula were divided in to smaller aliquots of 163 cm3 in plastic measuring cylinders.
Seedlings were inoculated when they were transplanted from the pre-nursery to the main nursery.
One inoculated root of each oil palm seedling was
put into a plastic bag and tied, and this was then
covered with soil in the main nursery plastic bag.
The experimental design was a completely random
block design factorial of 6 · 5 · 2 · 2 with three
replications for each combination. The first factor
was type of cross used for the oil palm material
(where V1 = T · T, V2 = T · P, V3 = D · T, V4
= D · D selfing, V5 = D · P, V6 = D · D). The
second factor was the biocontrol agent used (T0 =
without biocontrol agents, T1 = T. harzianum, T2
= G. viride, T3 = Bacillus sp., T4 = T. harzianum
+ G. viride + Bacillus sp.). The third factor was
the G. boninense inoculation (G0 = without
inoculation and G1 = inoculation with G. boninense), and the fourth factor was the addition of
chitin (K0 = without chitin and K1 = plus
chitin). The biocontrol agents were added as 10 g
formulation for fungal biocontrol agents and
10 ml formulation for bacterial biocontrol agents.
For the T4 treatment, 5 g T. harzianum, 5 g G. viride, and 5 ml Bacillus sp. were used respectively. Chitin was added at 1 g per polybag. The
conidial density of T. harzianum and G. viride
preparations were 4 · 106 conidium/ml, whereas
Bacillus sp. was used at 7 · 108 cfu/ml. The variables recorded were disease incidence of basal stem
rot, plant height and number of leaves, plant vigour, and development of the biocontrol agent
population. Observations were made monthly for
1 year.
Results and discussion
Collection of candidate biocontrol agents
One hundred and forty fungi were isolated from
various oil palm rhizospheres and these included
18 isolates of T. harzianum, 5 isolates of T. viride, 4
isolates of Gliocladium viride, 28 isolates of
A. flavus, 4 isolates of A. niger, 13 isolates of A.
fumigatus, 14 isolates of Penicillium citrinum, 4
isolates of Rhizopus, 6 isolates of P. chrysogenum,
12 isolates of P. commune, and 32 isolates of
P. funiculosum. The majority of the isolates were
considered as candidate biocontrol agents. The
population densities of each species were similar
between plantations and were about 104–105 cfu/g
soils. Seventy two isolates of bacteria were obtained and these included isolates of Pseudomonas
fluorescens and Bacillus sp.
156
Inhibition activity of candidate biocontrol agents
Twenty six fungi and four bacteria showed potential biocontrol activities in dual culture analysis. Fungi were categorised as biocontrol agents if
they showed an inhibition capacity >80%, and
bacteria were considered biocontrol agents if they
showed an inhibition capacity of >40%. The isolate that gave the highest inhibition capacity was
T. harzianum (97.8%). This was isolated from
a third generation oil palm planting in
North Sumatra. The isolate was similar to another
T. harzianum isolate from a BSR infected
immature fourth generation planting in North
Sumatra, to an isolate from healthy immature
second generation planting in North Sumatra, to
an isolate from infected immature second generation planting in North Sumatra, and to an isolate
from infected productive first generation planting
in West Sumatra. T. harzianum was the most
common species among the candidate biocontrol
agents.
Only four bacteria showed potential biocontrol
properties, all of the bacteria gave lower inhibition
values than the fungi. The highest inhibition
capacity was shown by an isolate of Bacillus sp.
from Lampung (54.2%). The remaining bacterial
biocontrol agents all had inhibition capacities of
between 40 and 50%.
The presence of Trichoderma and Gliocladium
was influenced by soil porosity, soil pH, and other
chemical properties of the soil. The abundance of
Trichoderma in soil depends on its ability to degrade organic matter and its resistance to other
microorganisms. The differences in the abundancies of the biocontrol agents may also be influenced by the climatic conditions at each site.
Table 3. The effect of biocontrol agents on Basal Stem Rot
incidence and oil palm vigour
Treatment
Disease incidence (%) Plant vigour
Control
T. harzianum
G. viride
Bacillus sp.
T. harzianum +
G. viride +
Bacillus sp.
18.06 a
0.00 c
0.00 c
9.72 b
8.33 bc
2.67
3.00
3.00
2.71
2.82
b
a
a
b
b
Entries followed by the same letter are not significantly different
by Duncan’s Multiple Range Test at the 5% level.
The D · D cross may have the highest disease
incidence, due to selfing leading to an accumulation of susceptible traits, as selfing apparently
did not result in an accumulation of resistance
traits.
The addition of candidate biocontrol agents
(T. harzianum and G. viride) was found to significantly reduce Basal Stem Rot incidence (Table 3).
Disease was only seen in the negative controls and
with the Bacillus sp. and mixed inoculum. The
ability of Bacillus sp. to inhibit G. boninense appeared to be lower than either of the fungal biocontrol agents, and every test involving Bacillus sp.
gave higher disease incidences. In the mixed inocula it may be that the Bacillus sp. competed with
the T. harzianum and G. viride isolates.
The oil palms treated with T. harzianum and
G. viride were significantly taller than the negative
controls and the palms treated with Bacillus sp.
and the mixed treatments. Adding of T. harzianum
and G. viride also appeared to give significantly
higher plant vigour than the control, Bacillus sp.
and mixed species treatments.
Efficiency of biocontrol agents in glasshouse trial
The disease incidence of Basal Stem Rot was not
consistent for all oil palm crosses. The cross with
the highest disease incidence was the selfed D · D
at 6.11%, compared to the other crosses where the
disease incidence ranged from 1.11 to 2.78%.
The T · T had not shown any disease symptoms
by the end of the 1 year period. This does not
indicate that T · T was resistant to G. boninense,
as this cross is known to develop BSR in the field,
and one possibility is that the 1 year period was
not enough for the pathogen to infect T · T.
Conclusions
This study has isolated 30 candidate biocontrol
agents consisting of 17 isolates of T. harzianum, 4
isolates of G. viride, 5 isolates of T. viride, one
isolate of Bacillus sp., and 3 isolates of Pseudomonas fluorescents. After 1 year of inoculation,
T. harzianum and G. viride appeared to prevent
Basal Stem Root in glasshouse trials, whereas,
Bacillus sp. had very little capacity to prevent
infection by G. boninense.
157
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Address for correspondence: A. Susanto, Indonesian Oil Palm
Research Institute, Jl. Bigjen Katamso No. 51, Medan 20158,
Indonesia
Phone: +61-7862466/77/50; Fax: +61-7862488;
E-mail: iopri@idola.net.id