CHAPTER-VIII Bioassay of antagonists on disease...
Transcript of CHAPTER-VIII Bioassay of antagonists on disease...
148
CHAPTER-VIII
Bioassay of antagonists on disease suppression and plant growth under
green house condition.
8.1 Background
Since the pathogens that cause root disease of tea are soil borne, it becomes essential
to study the effect of soil microbes for their antagonistic/ hyperparasitic behaviour against the
test pathogens U. zonata and F. lamaoensis, under study .The diseases transmitted from the
infected plant to the healthy plant through the infected root.
Various soil microbes which act as effective biocontrol agents against soil borne
pathogens are well-known for their beneficial effect as plant growth promoter (Barka et al .,
2000 ; Chakraborty et al.,2005). In the present study, an attempt was made to study the effect
of selected antagonistic microbial strains isolated from tea soil and was found effective in in-
vitro antagonistic test. Bioassay of highly antagonistic microbial strains screened in-vitro is
very essential to determine their potentiality under natural condition, therefore a nursery trial
was conducted for assessing the effect of different treatments on growth parameters and plant
health. Treatments were applied in the root zone as there are evidence that actinomycetes are
quantitatively and qualitatively important in the rhizosphere (Barakate et al., 2002; Crawford
et al.,1993; Doumbou et al., 2001; Miller et al., 1989, 1990), where they may influence plant
growth and protect plant roots from invasion by root pathogenic fungi (Lechevalier,1988).
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8.2 Materials and Methods
8.2.1 Evaluation of screened antagonist strains under nursery conditions
The nursery trial was conducted in a glasshouse at Tocklai Experimental Station,
Jorhat (Latitude 26043
/ 46.38
// N; Longitude 94
013
/ 43.44
//E), with tea seedlings for a period of
18 months (fig 8.1). The isolates used in this trial were selected on the basis of their efficacy
to inhibit the growth of two test pathogens under in- vitro conditions (antagonistic test
conducted in chapter VII)
8.2.2 Soil used
The soil used in the nursery trial for filling up the polythene sleeves (34cmx19cm)
was collected from the tea plantation area under Tocklai Experimental Station. Black
polythene sleeves (34cm×19cm) were filled with 6 kg of uniformly broken soil. The soil was
of sandy loam texture with moisture content % 17.5 and PH
of 4.5-5.3. The microbial load of
the soil sample was estimated before filling up of the sleeves (Table 8.1) to have an idea of
the biological status of the soil.
8.2.3 Planting Material
Cuttings of tea plants of clone TV1 cultivars of Assam variety, (Camellia sinensis
var. assamica) were procured from nursery of Borbheta Experimental plot (Jorhat) having
latitude 26.71o, longitude 94.20
o and elevation 322ft. The cuttings were transported to glass
house and set in the campus of Tocklai Experimental Station where they were raised in
nursery bed for the study (fig 8.2). After the formation of callus, the tea plantlets were
transferred to the polythene sleeves and maintained well through regular monitoring.
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8.2.3 .1 Description of selected clone
TV 1 Type : Assam China hybrid
Frame : Compact and dense
Shoot size (2L+B) : Medium
Weight
Fresh (gm) : 0.59
Dry (gm) : 0.12
Leaf type (size) : medium
Rooting : very good
Leaf shape : broadly lanceolate
Pubescence : medium
Drought resistance : good
Susceptible to disease: High to moderate depending on pathogen
Involved.
8.2.4 Experimental Treatments
Nursery trial was conducted to assess biocontrol potential of selected PSB and
actinomycetes isolates against two primary root disease causing pathogens of tea by dual
inoculation of the pathogens (U.zonata or F.lamaoensis) and the antagonistic isolates of PSB
(MM/PH/BST and MM/PH/KMP) and actinomycetes (MM/PH/AC-02 and MM/PH/AC-09)
following Ganesan and Gnanamanickam, (1987). The experiment was conducted with
thirteen treatments consisting 4 isolates and two reference strains of Trichoderma viride, one
absolute control (no pathogen and no biocontrol agent) and another two control with only
pathogens (ordinary control).The experiment was performed in the plastic sleeves with tea
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plantlets under greenhouse condition with 13 treatments and 5 replications following the
Completely Randomized Block Design (CRBD).The layout of the nursery trial plot is shown
in fig 8.3. The treatments (antagonistic isolates) used for nursery evaluation of the selected
PSB and actinomycetes isolates in controlling the root disease causing pathogens as well as in
enhancing growth of tea plants are given below:
Sl. No. Treatments
1. MM1- MM/PH/AC-02+ Ustulina zonata (U.zonata)
2. MM2- MM/PH/AC-09+ U.zonata
3. MM3- MM/PH/KMP + U.zonata
4. MM4- MM/PH/BST + U.zonata
5. MM5- MM/PH/AC-02+ Fomes lamaoensis (F. lamaoensis )
6. MM6- MM/PH/AC-09+ F. lamaoensis
7. MM7- MM/PH/KMP+ F. lamaoensis
8. MM8- MM/PH/BST+ F. lamaoensis
9. MM9 –Trichoderma viride + U.zonata
10. MM10- T. viride + F. lamaoensis
11. MM11- Control (only U.zonata )
12. MM12- Control (only F. lamaoensis )
13. MM13- Absolute control (No pathogen and no antagonists)
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The plants were kept under optimum shade condition throughout the experimental
period. The plants were watered regularly and plant protection measures were taken as and
when necessary to protect the plants against insect pest damage.
8.2.4.1 Preparation of inocula for treatment
Tea bushes infected with charcoal stump rot were collected from root disease infested
area of Kakojan Tea Estate (28, Deberapara div.). For brown root rot disease, infected tea
bushes were taken from Tyroon Tea Estate, (Kheremia section No.10). Infected tea roots
were initially washed properly with tap water to remove all the soil particles, and again rinsed
with distilled water. The roots were made into small rectangular blocks (4cm x 2cm x1cm),
and incubated at 25 ± 2o C in conical flasks under moist condition. After 15-20 days the
infested wooden blocks were found to be covered by thick subfelty mycelia with colour
ranging from brownish white to brown in case of F. lamaoensis (Brown root rot) and thick
charcoal like encrustation and black lining in all the surfaces of the wooden blocks in case of
U. zonata (Charcoal stump rot).
8.2.4.2 Pure culture of applicable strains
Pure cultures of four selected strains (MM/PH/BST,MM/PH/KMP, MM/PH/AC-02
and MM/PH/AC-09) were selected on the basis of in-vitro antagonistic screening, and
prepared in broth of Pikovskaya‘s medium (MM/PH/BST, MM/PH/KMP) and Kenknights
medium (MM/PH/AC-02, MM/PH/AC-09) under laboratory conditions. The cultures were
incubated at their optimum temperatures till the appearance of full growth of the strains.
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8.2.4.3 Treatment application
The incubated tea root bits were soaked in the pure culture broth of the test strains and
kept overnight before application of treatments. One year tea plants raised in the polythene
sleeves for nursery trial were inoculated artificially in the root zone with pure inocula
cultured on fresh infested tea root bits under laboratory condition. The freshly infested tea
root bits with and without implication of pure culture of antagonistic strains that are to be
applied as treatments were introduced in the soil by making four holes around the plants so
that the bits touches the root of the target tea plants. The treatment application was conducted
following the method of Barthakur (1997) with some modification. Thirty target plants were
taken for each treatment and equal number of plants were not subjected to any treatment were
considered as control.
8.2.6 Plant growth measurements
The experiments were maintained for eighteen months. Stem girth, root length, fresh
weight and dry weight of biomass per plant were recorded by non-destructive methods on the
day of inoculation and at quarterly intervals (after every three months) after application of the
treatment.
8.2.5.1 Root length
On expiry of the harvesting schedule, plants were carefully uprooted and washed
gently the root system to get them free of soil. The root length was recorded by measuring
from base of the stem to tip of the longest root and expressed in centimetres.
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8.2.5.2 Stem girth
Stem girth was determined by measuring the diameter of the stems at collar region
using a vernier calliper applying the formula 2πr and expressed in centimetres.
8.2.5.3 Total Biomass content
Both the fresh and dry weight of the uprooted plants was recorded. Uprooted plants
were first air dried and then oven dried at 600C to a constant weight for recording the dry
biomass and were expressed in g per plant.
8.2.6 Periodic observation of plant health and disease incidence
Visual observation of the plant health was recorded after every harvesting by
recording the number of healthy plants, plants with stunted growth, plants started wilting,
plants with acute wilting and dead. On the basis of plant health, disease occurrence and
incidence of mortality, a score chart was prepared for recording the periodic observation.
Score Chart.
Score 1: Healthy
Score 2: Stunted growth
Score 3: Initial wilting
Score 4: Progressive wilting
Score 5: Dead
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8.2.6.1 Determination of disease incidence
The tea plants exhibiting stunted growth, initial wilting, progressive wilting and
death were examined thoroughly by uprooting the plants. The uprooted tea plants after each
harvesting were washed properly with tap water and the disease incidence was determined
from external appearance, according to characteristic symptoms mentioned in I.T.A.
Memorandum No.8 (Tunstall, 1940)
To determine the incidence of fungi on roots, the modified technique of the method of
Short et al., (1980) was used where root from each uprooted plants after each harvesting were
cut into one cm long root pieces, surface sterilized with 1% Ca(OCl)2 for 3 min and
transferred onto potato dextrose agar plates containing streptomycin (0.2gm/litre). After
incubation for 5 to 7 days at 280C, incidence of root infecting fungi and percent disease
reduction (DIR) were recorded using the formula as given by Shameem, (2006).
Total number of plants infected by the pathogen
Disease incidence % = x 100
Total number of plants
.
No. of plants infected in control – No. of plants infected in treatment
% DIR= ———————————————————————————— x 100
No. of plants infected in control
8.2.7 Experimental design and statistical analysis
Nursery experiment was arranged in a complete randomized block design with four
replicates for each treatment as well as control. Statistical analysis of the obtained data was
subjected to standard analysis of variance procedure. The values of LSD were calculated at
5% level according to the method described by Gomez and Gomez (1984).
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8.3 Results
8.3.1 Effect of PSB and actinomycetes isolates on growth parameters of tea plants
After the 1st harvest, growth parameters like stem girth, root length, fresh weight and
dry weight of tea plants was found to be increased as a result of treatments in MM5
(MM/PH/AC-02 and F.lamaoensis) and in MM9 (T.viride and U.zonata) (Table 8.2 and Fig.
8.a). Except MM3 (MM/PH/KMP and U.zonata), MM8 (MM/PH/BST and F.lamaoensis)
and MM11 (only U.zonata), all other treatments showed better stem girth.
More root length was observed in the plants treated with MM5 (MM/PH/AC-02 and
F.lamaoensis), MM7 (MM/PH/KMP and F.lamaoensis), MM9 T.viride and U.zonata ) and
MM10 (T.viride and F.lamoensis ) than untreated plants.
Fresh weight of the plants under all the treatments were found higher except in
treatment MM3 (MM/PH/KMP and U.zonata) and MM11 (only U.zonata) which was at par
with the untreated control. Dry weight of the plants with treatment MM1, MM5, MM7 and
MM9 were found significantly higher than untreated control. All the findings were found to
be statistically significant.
The data recorded after 2nd
harvest (Table 8.3 and Fig. 8.b) revealed that the plants
treated with MM9 (T.viride and U.zonata) showed maximum growth in stem girth, root
length and fresh weight over untreated plants. But, in case of dry weight, plants treated with
MM8 (MM/PH/BST and F.lamaoensis) showed the highest value. All the plants treated with
combination of antagonist microbes and pathogen showed better growth in stem girth, root
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length, fresh weight and dry weight of the plants than the untreated plants, while the plants
grown with the pathogen only did not showed any significant difference with the untreated
plants.
The data recorded after 3rd
harvesting is presented in Table 8.4. and Fig. 8.c Plants
treated with MM4 (MM/PH/BST and U.zonata) showed maximum stem girth followed by
treatment MM8 (MM/PH/BST and F.lamaoensis). In case of root length, fresh and dry
weight biomass, MM8 recorded the highest. All the treatments of antagonist isolates in
combination with the test pathogen (treatments MM1, MM2, MM3, MM4, MM5, MM6,
MM7, MM8) were found to increase all the growth parameters significantly,(fig 8.4). But the
plants treated with only pathogens (treatments MM11 and MM12) did not show any
significant variation with the untreated plants (treatment MM13).
After the 4th harvest, the growth parameters like stem girth, root length and fresh
weights were found to be increased, with treatments MM2, MM4, MM6 and MM8 than the
untreated plants. Among all the treatments, MM2 (MM/PH/AC-09 and U. zonata) was found
to be the best followed by treatment MM8 (MM/PH/BST and F. lamaoensis). Plants treated
with only pathogens (treatments MM11 and MM12) showed significantly lesser growth in all
the growth parameters, however the results were at par with the untreated plants in case of
dry weight(fig 8.5&fig 8.6)
The records of the 5th harvest (Table 8.6 and Fig. 8.e) clearly states the maximum
effect of treatment MM2 (MM/PH/AC-09 and U.zonata), MM4 (MM/PH/BST and
U.zonata), MM6 (MM/PH/AC-09 and F.lamaoensis) and MM8 (MM/PH/BST and F.
lamoensis) on all the growth parameters. All the treatments, except MM11 (only U. zonata)
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and MM12 (only F. lamaoensis) showed significant (P<0.05) increase in growth parameters
over untreated plants.
According to the data recorded after 6th harvest (Table 8.7 and Fig. 8.f) treatment
MM2 (MM/PH/AC-09 and U.zonata) was found to be highly effective in increasing stem
girth, root length and dry weight in comparison to all other treatments, effect of treatment
MM11 (only U. zonata) and MM12 (only F. lamaoensis) was found to be the lowest among
all the other treatments.
8.3.2 Periodic observation of plant health and disease incidence
Observation on the plant health and development of morphological disease symptoms
was taken after every three months from the date of treatment applications. Some of the
plants were found healthy throughout the observation, while some showed disease incidence
according to the type of treatment.
Records of the periodic observation on plant health and disease occurrence of treated
tea plants under greenhouse trial were presented in Table 8.8.The data reveal that all the
antagonist isolates (MM1, MM2, MM3, MM4, MM5, MM6, MM7, MM8) effectively check
disease occurrence. Plants treated with antagonist exhibited good health at the time of 1st
harvesting irrespective of treatments. Initial wilting in one plant and stunted growth in two
plants was noted at the time of 2nd
harvest, in plants treated with U. zonata (Treatment
MM11) while rest of the plants were healthy. At the time of 3rd
harvesting all other plants
except the treatment MM11 showed healthy growth. In the treatment MM11, progressive
wilting was noticed in one plant along with initial wilting in two and stunted growth in other
two. Stunted growth (1plant) and initial wilting (2 plants) were also observed in plants treated
with only F. lamaoensis (Treatment MM12). At the time of 4th harvesting, four plants from
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MM11 and three plants from MM12 treated plants showed progressive wilting. Initial wilting
was recorded in plants with treatment MM1 (3 plants), MM2 (1plant), MM3 (2 plants), MM5
(1 plant) and MM10 (1 plant). At the time of 5th
and 6th
harvesting all the plants taken for
observation in MM11 and MM12, were found dead with withered leaves (fig 8.7). Uprooting
of dead plants recorded in MM11 treated plants showed symptoms of charcoal stump rot
infection (fig 8.8) and that in MM12 treated plants showed initiation of brown root rot
symptoms.
8.3.2.1 Determination of disease incidence
Appearance of small white, turning black cushions were evidenced in root of
progressive wilting and dead plants under treatment MM11 (U.zonata) (fig 8.9). Infected root
when peeled of carefully, white delicate fans of mycelium were observed. When the root was
cut off longitudinally black lines were detected when they were observed minutely which
shows their resemblance with the characteristics of charcoal stump rot. After uprooting of
progressive wilting and dead plants of MM12 (F.lamaoensis), the roots were found to be
rough with soil even after washing, cream to brown mycelium were also found encrusted to it
which were considered to be infected by brown root rot ( fig 8.10).
Overall percentage of disease incidence was recorded to be optimum (50%) in
plants treated with MM11 (only U.zonata) followed by 43% in plants treated with MM12
(only F.lamaoensis) (Table 8.7 and Fig. 8.f). Disease incidence was found to be less (6.67%)
in the plants treated with treatment MM2 (MM/PH/AC-09+ U.zonata) and MM6
(MM/PH/AC-09+ F. lamaoensis) in each of the cases.
The percentage of disease reduction by the inoculated antagonist PSB and
actinomycetes varied from 61.54 to 86.67% (Table 8.7 and fig 8.g). Isolate MM/PH/AC-09
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showed maximum control (86.67%) of U. zonata, followed by the same isolate against F.
lamaoensis, with 84.62 % reduction in disease over control (treated with only pathogens).
8.4 Discussion
Periodic observation on plant health and disease incidence of the plants under
greenhouse trial indicated that, all the four antagonists were effective in checking the disease
occurrence as compared to the plants treated with only pathogen (control). Among the
thirteen treatments including untreated plants (Absolute control), the progress of disease
occurrence was less in MM/PH/AC-09 and MM/PH/BST treated plants with both the test
pathogens. The severity of diseases substantially increased in plants treated with only
pathogens. From the investigation it was found that percent of disease incidence was 50 and
43 in plants treated with only U.zonata and F.lamaoensis respectively. 100 percent disease
incidence was not recorded in any of the treatment because due to time unavailability our
nursery trial was confined to 18 months, which was not enough for both the pathogens to
attack the tea plants completely. According to the previous record it was known that charcoal
stump rot (U.zonata) requires a period of six months to four years to attack plant completely
and Brown root rot (F.lamaoensis ) attacks tea plants completely above three year but young
tea plants are affected within few months.
In the present study, all the antagonistic isolates utilized as treatment were able to
improve the plant growth leading to increase in stem girth, root length, Fresh and dry weight
biomass over the pathogen treated as well as plants without any treatments, under nursery
condition which correlates with the work of Phukan et al. (2011),where they reported several
indigenous species of Bacillus, Pseudomonas, Azotobacter, Azospirillum isolated from the tea
soil showing plant growth promoting as well as antagonistic effect against Black rot and
blister blight disease of tea in both nursery and commercial trial and also with the work of
Mishra (2006). Previous work of many workers explains that growth promotion occurs as a
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result of direct stimulation of plant growth, induction of host plant systemic resistance or
suppression of plant pathogens (Dwivedi and Johri, 2003; Lugtenberg et al., 2002; Fakhouri
et al., 2001; Van Loon et al., 1998). Isolates MM/PH/AC-09 and MM/PH/BST was found to
be highly effective in increasing the growth parameters of the plants significantly (P<0.05) as
well as in decreasing disease occurrence which justify the fact that antagonism is one of the
important trait of Plant growth promoting microbes, therefore Such microorganism have been
applied to a wide range of plants for the purpose of plant growth enhancement and disease
control (Barka et al., 2000 and Chakraborty et al., 2005). The increment in growth
parameters in response to microbial antagonists inoculation endorsed the fact that the test
strains were having one or more growth promoting mechanism including mobilization and
efficient uptake of nutrients ( Biswas et al.,2000), solubilization of insoluble phosphates
(Alikhani et al., 2006), and inhibition of fungal growth (Nautiyal, 1997).
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Table 8.1: Microbial load of the soil used for nursery trial.
Type of microbes Population
density
Bacteria
39.6 x10-5
cfu/g
Fungi 12.0 x10-4
cfu/g
Actinomycetes 2.3 x10-4
cfu/g
Azotobacter 1.3 x10-5
cfu/g
Azospirillum ND
Phosphate solubilizer 1.0 x10-5
cfu/g
ND- not detected, cfu- colony forming units.
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Table 8.2: Effect of different treatments on stem girth, root length and
biomass of the plant (1st harvesting, Dec 09-Mar 10 )
Treatments Stem girth (cm) Root length
(cm)
Fresh weight of
plant
(gm)
Dry weight of
plant
(gm)
MM1 1.48ab 48.60ab 117.80abcd 59.22a
MM2 1.35cd 41.40e 115.44de 55.74bc
MM3 1.27de 38.00f 113.34e 53.24c
MM4 1.40bc 44.28cde 118.40abc 56.50b
MM5 1.50a 49.52a 120.56a 61.41a
MM6 1.46ab 41.68de 115.94cde 55.16bc
MM7 1.39bc 46.00abc 119.26ab 60.80a
MM8 1.25e 44.88bcde 115.14de 56.48b
MM9 1.55a 49.66a 120.40a 61.82a
MM10 1.31cde 46.24abc 115.80cde 54.04bc
MM11 1.25e 44.16cde 113.34e 55.50bc
MM12 1.33cde 46.32abc 116.82bcd 56.50b
MM13 1.30de 45.28bcd 113.58e 54.18bc
S.Ed () 0.04 1.66 1.24 1.23
CD(0.05) 0.08 3.37 2.51 2.49
The figures followed by same letter(s) do not differ significantly
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Table 8.3: Effect of different treatments on stem girth, root length and
biomass of the plant (2nd
harvesting, Mar10-june-10)
Treatments Stem girth (cm) Root length (cm) Fresh weight of
plant
(gm)
Dry weight of
plant
(gm)
MM1 1.64bc 54.20c 140.60cd 67.50bc
MM2 1.55fg 47.46f 132.60f 60.40e
MM3 1.58ef 46.40f 126.68gf 58.10f
MM4 1.62cd 52.60cd 135.80e 61.60de
MM5 1.66b 57.48b 142.52c 69.48ab
MM6 1.64bc 50.50de 131.42f 56.66fg
MM7 1.60de 52.40cd 139.60d 63.42d
MM8 1.67b 58.46b 145.38b 71.60a
MM9 1.75a 61.66a 148.50a 66.42c
MM10 1.54gh 53.50cd 127.50g 61.26de
MM11 1.51h 47.74ef 125.42gh 55.22gh
MM12 1.52gh 46.48f 124.60h 53.48h
MM13 1.54gh 47.26f 126.50gh 57.30fg
S.Ed () 0.02 1.39 1.24 1.05
CD(0.05) 0.03 2.81 2.50 2.13
The figures followed by same letter(s) do not differ significantly
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Table 8.4: Effect of different treatments on stem girth, root length and biomass
of the plant (3rd
harvesting June10-sep-10)
Treatments Stem girth (cm) Root length (cm) Fresh weight of
plant
(gm)
Dry weight of
plant
(gm)
MM1 1.65f 51.50de 145.10c 70.18bc
MM2 1.69cd 59.20a 150.10b 75.32a
MM3 1.71bc 51.60de 142.50d 65.06d
MM4 1.75a 56.40b 148.71b 72.20b
MM5 1.68de 54.20c 139.20e 62.30e
MM6 1.69cd 59.20a 155.60a 75.60a
MM7 1.65f 53.50cd 135.51f 61.60e
MM8 1.72b 58.60a 157.48a 76.56a
MM9 1.71bcd 51.30de 141.71d 68.20c
MM10 1.66ef 51.00e 141.56d 62.40e
MM11 1.56h 48.16f 129.40g 58.30f
MM12 1.62g 49.28ef 130.20g 60.20ef
MM13 1.66ef 50.20f 133.31f 60.26ef
S.Ed () 0.01 1.03 1.15 1.04
CD(0.05) 0.02 2.08 2.33 2.11
The figures followed by same letter(s) do not differ significantly
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Table 8.5: Effect of different treatments on stem girth, root length and biomass
of the plant (4th
harvesting, Dec10-Mar-11)
Treatments Stem girth (cm) Root length (cm) Fresh weight of
plant
(gm)
Dry weight of
plant
(gm)
MM1 1.75d 53.08cd 150.82de 72.60de
MM2 1.85a 58.50a 160.06a 78.42a
MM3 1.75d 53.30cd 149.20e 72.28de
MM4 1.82b 58.50a 156.48bc 76.60abc
MM5 1.75d 55.50bc 148.78ef 72.52de
MM6 1.81b 57.50ab 158.50ab 77.64ab
MM7 1.74d 52.50d 152.12d 74.84cd
MM8 1.78c 56.70ab 155.22c 75.28bc
MM9 1.74d 52.10d 152.40d 70.10ef
MM10 1.75d 53.64cd 148.78ef 72.20de
MM11 1.65g 47.60e 145.80g 68.10f
MM12 1.68f 48.50e 146.24fg 59.26f
MM13 1.71e 52.54d 148.78ef 68.88f
S.Ed () 0.01 1.16 1.25 1.23
CD(0.05) 0.03 2.35 2.53 2.48
The figures followed by same letter(s) do not differ significantly
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Table 8.6 : Effect of different treatments on stem girth, root length and biomass
of the plant (5th
harvesting, Sep10-Dec-10)
The figures followed by same letter(s) do not differ significant
Treatments Stem girth (cm) Root length (cm) Fresh weight of
plant
(gm)
Dry weight of
plant
(gm)
MM1 1.87bc 59.20cd 188.40bc 81.34c
MM2 2.12a 64.26a 195.48a 88.56b
MM3 1.84cd 57.16de 182.68d 87.22
MM4 2.11a 62.38ab 196.44a 82.60
MM5 1.90bc 60.10bc 183.52d 76.28
MM6 1.93b 61.32bc 195.48a 82.28
MM7 1.78d 57.16de 184.58cd 88.56
MM8 1.91bc 60.10bc 190.12b 77.22
MM9 1.88bc 59.24cd 185.50cd 82.60
MM10 1.88bc 57.34de 183.52d 88.12
MM11 1.53e 41.30f 172.16e 68.46
MM12 1.43f 42.38f 171.56e 65.42
MM13 1.87bc 56.36e 181.62d 75.91
S.Ed () 0.04 1.08 2.04 1.26
CD(0.05) 0.07 2.19 4.13 2.56
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Table 8.7: Effect of different treatments on stem girth, root length and biomass
of the plant (6st harvesting, Mar11-june-11)
The figures followed by same letter(s) do not differ significantly
Treatments Stem girth (cm) Root length (cm) Fresh weight of
plant
(gm)
Dry weight of
plant
(gm)
MM1 2.02bc 59.24c 235.48fg 100.36ef
MM2 2.20a 64.84a 242.46b 110.20a
MM3 1.96c 58.66c 238.36de 100.96e
MM4 2.14ab 64.60a 244.82a 111.62d
MM5 2.00bc 61.36bc 236.62ef 99.40ef
MM6 2.11ab 62.72ab 242.46b 107.38b
MM7 2.00bc 58.66c 234.52g 100.40ef
MM8 2.08abc 61.46bc 241.72b 107.12bc
MM9 2.06bc 59.24c 239.60cd 104.56cd
MM10 2.08abc 58.68c 240.50bc 102.32de
MM11 1.69d 42.56d 161.72i 97.70f
MM12 1.75d 43.52d 172.56h 100.96e
MM13 1.96c 58.68c 234.52g 100.64ef
S.Ed () 0.06 1.26 0.91 1.32
CD(0.05) 0.12 2.55 1.85 2.66
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Table 8.8 Periodic observation on plant health and disease incidence
Treatment
Status of plant health and disease incidence after treatments
3 months 6 months 9 months 12 months 15 months 18 months
MM1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 3 1 3 3 1 1 4 3 4 2 4 3 4 5
MM2 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 2 1 1 3 1 1 1 1 3 3 5 1 2 3 4
MM3 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 3 3 1 1 2 4 5 1 4 1 4 1
MM4 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 2 1 1 4 1 1 5 3 4
MM5 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 3 1 1 1 1 3 1 4 5 2 4 1
MM6 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 3 1 1 3 1 1 1 1 1 4 4 3
MM7 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 2 1 1 4 1 4 5 4 3 3 4
MM8 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 4 1 1 1 4 4
MM9 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 3 2 4 1 4 1 2 4 5 5
MM10 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 3 1 1 1 1 2 3 4 5 5 4 4 4
MM11 1 1 1 1 1 1 1 2 2 3 2 2 3 3 4 1 4 4 4 4 5 5 4 4 4 4 5 5 5 5
MM12 1 1 1 1 1 1 1 1 1 1 1 1 3 3 2 1 4 4 4 2 5 4 4 4 5 5 5 5 5 4
MM13 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 1 4 2 1 3 3
170
Table 8.9 Details of the plant health and disease incidence at the completion
of study.
Treatments Total numbers
of plants
Numbers of
diseased plants
%Disease reduction
over control
MM1 30 5 66.67
MM2 30 2 86.67
MM3 30 4 73.33
MM4 30 3 80.00
MM5 30 4 69.23
MM6 30 2 84.62
MM7 30 5 61.54
MM8 30 3 76.92
MM9 30 5 66.67
MM10 30 6 53.85
MM11(control) 30 15 00.00
MM12(control) 30 13 00.00
MM13(Absolute control)
30 1 00.00
N.B: Control denotes the plants treated with only pathogen and Absolute
control denotes the plants without any treatments
171
Table 8.10 Effect of antagonists on charcoal stump rot
Treatments % disease incidence
MM/PH/AC-02 + U.zonata 16.67
MM/PH/AC-09 + U.zonata 6.67
MM/PH/KMP + U.zonata 13.33
MM/PH/BST + U.zonata 10.00
T.viride (reference ) +U.zonata 16.67
U.zonata (control) 50.00
S.Ed (±) 0.8755
CD(0.05) 2.145
Table 8.11 Effect of antagonists on Brown root rot
Treatments % disease incidence
MM/PH/AC-02 + F.lamaoensis 13.33
MM/PH/AC-09 + F.lamaoensis 6.67
MM/PH/KMP + F.lamaoensis 16.67
MM/PH/BST + F.lamaoensis 10.00
T.viride (reference ) + F.lamaoensis 20.00
F.lamaoensis (control) 43.33
S.Ed (±) 0.8944
CD(0.05) 2.191
172
Fig 8.a. Effect of different treatments on stem girth, root length and
biomass of the plant (1st harvesting, Dec 09-Mar 10)
Fig. 8.b. Effect of different treatments on stem girth, root length and biomass of
the plant (2nd
harvesting, Mar10-june-10)
0
0.2
0.4
0.6
0.8
1
1.2
1.4
1.6
1.8
0
20
40
60
80
100
120
140
MM1 MM2 MM3 MM4 MM5 MM6 MM7 MM8 MM9 MM10MM11MM12MM13
Ste
m g
irth
(cm
)
Ro
ot le
ng
th (
cm
), F
resh
weig
ht
of
pla
nt
(gm
), D
ry w
eig
ht
of
pla
nt
(gm
)
Treatment
RL (cm) Fwt
Dwt Stem girth (cm)
1.35
1.4
1.45
1.5
1.55
1.6
1.65
1.7
1.75
1.8
0
20
40
60
80
100
120
140
160
MM1 MM2 MM3 MM4 MM5 MM6 MM7 MM8 MM9 MM10MM11MM12MM13
Ste
m g
irth
(cm
)
Ro
ot le
ng
th (
cm
), F
resh
weig
ht
of
pla
nt
(gm
), D
ry w
eig
ht
of
pla
nt
(gm
)
Treatment
RL (cm) Fwt
Dwt Stem girth (cm)
173
Fig. 8.c. Effect of different treatments on stem girth, root length and biomass of the
plant (3rd
harvesting June10-sep-10)
Fig. 8.d. Effect of different treatments on stem girth, root length and biomass of the
plant (4th
harvesting, Dec10-Mar-11)
1.45
1.5
1.55
1.6
1.65
1.7
1.75
1.8
0
20
40
60
80
100
120
140
160
180
MM1 MM2 MM3 MM4 MM5 MM6 MM7 MM8 MM9 MM10MM11MM12MM13
Ste
m g
irth
(cm
)
Ro
ot le
ng
th (
cm
), F
resh
weig
ht
of
pla
nt
(gm
), D
ry w
eig
ht
of
pla
nt
(gm
)
Treatment
RL (cm) Fwt
Dwt Stem girth (cm)
1.55
1.6
1.65
1.7
1.75
1.8
1.85
1.9
0
20
40
60
80
100
120
140
160
180
MM1 MM2 MM3 MM4 MM5 MM6 MM7 MM8 MM9 MM10MM11MM12MM13
Ste
m g
irth
(cm
)
Ro
ot le
ng
th (
cm
), F
resh
weig
ht
of
pla
nt
(gm
), D
ry w
eig
ht
of
pla
nt
(gm
)
Treatment
RL (cm) Fwt
Dwt Stem girth (cm)
174
Fig. 8.e. Effect of different treatments on stem girth, root length and biomass of the
plant (5th
harvesting, Sep10-Dec-10)
Fig. 8.f. Effect of different treatments on stem girth, root length and biomass of the
plant (6th
harvesting, Mar11-june-11)
0
0.5
1
1.5
2
2.5
0
50
100
150
200
250
MM1 MM2 MM3 MM4 MM5 MM6 MM7 MM8 MM9 MM10MM11MM12MM13
Ste
m g
irth
(cm
)
Ro
ot le
ng
th (
cm
), F
resh
weig
ht
of
pla
nt (g
m),
Dry
weig
ht o
f p
lan
t (g
m)
Treatment
RL (cm) Fwt
Dwt Stem girth (cm)
0
0.5
1
1.5
2
2.5
0
50
100
150
200
250
300
MM1 MM2 MM3 MM4 MM5 MM6 MM7 MM8 MM9 MM10MM11MM12MM13
Ste
m g
irth
(cm
)
Ro
ot le
ng
th (
cm
), F
resh
weig
ht
of
pla
nt
(gm
), D
ry w
eig
ht
of
pla
nt
(gm
)
Treatment
RL (cm) Fwt
Dwt Stem girth (cm)
175
Fig 8.g: Graphical representation of percent disease incidence and disease
reduction over control.
0
10
20
30
40
50
60
70
80
90
100
Treatments %Disease incidence %Disease reduction over control
Treatments
176
M 10 M 10 M 9 M95 4 5 4 5 4 5 4
M 4 M 3 M 2 M15 3 5 3 5 3 5 3
M 4 M 3 M 2 M15 5 5 5 5 5 5 5
M 11 M 11 M 11 M115 2 5 2 5 2 5 2
M 8 M 7 M 6 M55 5 5 5 5 5 5 5
M 8 M 7 M 6 M55 3 5 3 5 3 5 3
M 13 M 13 M 13 M135 2 5 2 5 2 5 2
M 12 M 12 M 12 M125 2 5 2 5 2 5 2
ENTRANCE
E
S
W
N
Fig 8.3 Layout of nursery trial plots
MM1- MM/PH/AC-02+ U.zonata
MM2- MM/PH/AC-09+ U.zonata
MM3- MM/PH/KMP + U.zonata
MM4- MM/PH/BST + U.zonata
MM5- MM/PH/AC-02+ F . lamoensis
MM6- MM/PH/AC-09+ F . lamoensis
MM7- MM/PH/KMP+ F . lamoensis
MM8- MM/PH/BST+ F . lamoensis
MM9 –T..viride + U.zonata
MM10- T..viride + F . Lamoensis
MM11- U.zonata
MM12- F . Lamoensis
MM13- Without Treatment.
177
fig 8.1: Site of Nursery trial
fig 8.2:
Cutting of TV1
cultivars raised in
nursery bed
178
fig 8.4: Observation of treated plants at the time of third harvesting
A- Treated with only pathogens; B- Treated with pathogen and antagonist.
fig 8.5: Observation of treated plants at the time of fourth harvesting
A-Treated with (MM/PH/AC-09+U.zonata ;
B-Treated with (MM/PH/BST + F.lamaoensis)
A
B
A
B
C
D
179
fig 8.6(a): Treated with pathogens +antagonists
fig 8.6(b): Treated with only pathogens
fig 8.6(c): untreated plants
180
Fig8.7: Observation of plants treated with only pathogens
at the time of sixth harvesting.
fig 8.8: Observation of root in uprooted plants after sixth harvesting
A- Healthy plant B- Diseased plant
A B
B
181
Fig 8.9: Root infected with U.zonata after 12 months
Of treatment application
Fig 8.10: Root infected with F.lamaoensis after 18 months
o f treatment application