BIBLIOGRAPHY BASNGI, JAMIE BETH B. March...
BIBLIOGRAPHY
BASNGI, JAMIE BETH B. March 2006. Germplasm Evaluation for Tolerance
Against Bacterial Wilt (Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.). Benguet
State University, La Trinidad, Benguet.
Adviser: Jocelyn C. Perez, MSc.
ABSTRACT
Nine clones: 387410.7, Warishiro, 575003, 676070, 380579.3, 720045, 573275,
285378.27 and 720071 and variety Igorota were screened to identify potato germplasm
materials tolerant to bacterial wilt infection and resistant to late blight infection under
green house condition.
All entries were found to be tolerant to bacterial wilt as they were able to produce
marketable tubers despite the bacterial wilt infection recorded.
The clones that showed resistance to late blight infection were clones 720071,
285378.27 and 676070 respectively.
TABLE OF CONTENTS
Page
Title Page ……………………………………………………………………..
i
Abstract ………… …………………………………………………………..
i
Table of Contents …………………………………………………………….
ii
INTRODUCTION ……………………………………………………………
1
REVIEW OF LITERATURE
Causal Organism ……………………………………………………….
4
Epidemiology …………………………………………………………..
4
Host Range and Distribution …………………………………………...
5
Management ……………………………………………………………
6
Selection for Resistance to Bacterial Wilt ……………………………..
7
Potato Breeding for Resistance to Bacterial Wilt ……………………...
9
CIP Breeding Program …………………………………………………
11
MATERIALS AND METHODS
Evaluation using Potato Seed Tubers …………………………………..
12
Inoculation Preparation and Soil Inoculation ………………………….
13
Data Gathered …………………………………………………………..
13
Evaluation using Rooted Stem Cuttings ……………………………...
15
Preparation of Inoculum and Inoculation Method ……………………..
15
Data Gathered ………………………………………………………….
16
ii
RESULTS AND DISCUSSION
Percentage Bacterial Wilt Infection ……………………………………
17
Percentage Late Blight Infection ……………………………………….
20
Yield Parameters ……………………………………………………….
22
Number of Days from Planting to Bacterial Wilt Expression …………
25
SUMMARY, COCLUSION AND RECOMMENDATION
Summary ……………………………………………………………….
28
Conclusion ……………………………………………………………..
28
Recommendation ……………………………………………………....
29
LITERATURE CITED ……………………………………………………….
30
APPENDICES ………………………………………………………………..
34
iii
INTRODUCTION
Nature of the Study
Potato
(Solanum tuberosum Linn) is grown worldwide due to its nutritive and
money making value. It is one of the mankind’s most valuable food because it provides a
source of low cost energy to human diet and a source of minerals, proteins, vitamin A and
B2, carbohydrates and some elements like potassium and phosphorus (Kipps, 1979).
A single medium-sized potato contains about half the daily adult requirement of
vitamin C. Potato is very low in fat, with just 5 percent of the fat content of wheat, and
one-fourth the calories of bread. When it is boiled, it has more protein than maize, and
nearly twice the calcium (CIP, 1996).
In the Philippines, potato ranks third among the leading commercial vegetables in
terms of peso value and ranks first among the vegetable crops in Northern Luzon
(Buasen, 1978). In like manner, it ranks fourth among the major crops worldwide after
wheat, maize and rice.
However, different pest and diseases affect the production of potato. Bacterial
wilt caused by Ralstonia solanacearum (E.F. Smith) Yabuuchi et al. is one of the most
important, widespread and lethal bacterial diseases of plants (Ma, 1990). It limits the
production of potatoes, especially seed potatoes worldwide (CIP, 1992; Martin and
French, 1985; Schmiediche, 1984 and Rich, 1983).
Kelman (1953) considered the earliest known record of bacterial wilt to be on
tobacco in Indonesia in 1864, where entire fields were lost due to bacterial wilt. Bacterial
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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wilt was recorded on potatoes in the United States in 1890 (Kelman, 1953) and in
Australia in 1894 (Tyron, 1895).
The disease was reported in the Philippines by Reinking (1918) affecting tobacco,
pepper and tomato as cited by Valdez (1986). In 1922, Welles and Roldan reported that
it caused a serious disease on solanaceous crops in Southern Luzon. Series of study
conducted reveals that it became severe throughout the highland and lowland areas
producing potato (Perez et al., 1997).
The survival of the bacterium is influenced by temperature, humidity and other
physical and chemical soil factors. R. solanacearum may survive for many years in
certain soils and may disappear from one growing season to the next (Martin and French,
1996).
Bacterial wilt can be reduced only if various control components are combined.
An integrated disease management approach can lead to significant reduction or even
eradication of bacterial wilt (CIP, 1996).
Among the practices recommended, the use of resistant cultivars is considered to
play an important role. It is proven useful to control the potato strain, and is potentially
the most effective and ideal way to manage the disease (Jyothi et. al., 1993; French,
1996). It also delays the development of populations of insects resistant to the pesticide
currently being used (Ganga, 1992).
The study was conducted to screen potato clones and cultivars for tolerance to
bacterial wilt under green house condition, and evaluate their reaction to late blight
infection.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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The study was conducted at the bacterial wilt nursery, the Northern Philippines
Root Crops Research and Training Center (NPRCRTC) and the Department of Plant
Pathology laboratory, Benguet State University from September 2005 to February 2006.
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REVIEW OF LITERATURE
Causal Organism
The phytopathogen Ralstonia solanacearum has over 5000 genes, many of which
probably facilitate bacterial wilt disease development (Brown and Allen, 2001). It is a
gram-negative plant-pathogenic bacterium that causes bacterial wilt in a variety of plants
(Hayward, 1994).
It is a soil-borne pathogen that naturally infects roots. It exhibits a strong and
tissue-specific tropism within the host, specifically invading and highly multiplying in the
xylem vessels (Smith, 1896 and Yabuuchi et al., 1995). Once established in the xylem
vessels, the bacteria are able to enter the intercellular spaces of the parenchyma cells in
the cortex and pith in various areas of the plant. Here, R. solanacearum is able to
dissolve the cell walls and create slimy pockets of bacteria and cell debris. Production of
highly polymerized polysaccharides increases the viscosity of the xylem, which results in
plugging (Shew and Lucas, 1991).
High temperatures and high soil moisture generally favors Ralstonia
solanacearum, the exception being certain in Race 3 strains that are pathogenic on potato
and are able to grow well at lower temperatures (Stevenson et al., 2001). In certain soil,
R. solanacearum may survive for many years, in others, the bacterium may disappear
from one growing season to the next (Martin and French, 1996).
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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EPIDEMIOLOGY
Ralstonia solanacearum race 3 is a soil borne pathogen that persists in wet soils,
deep soil layers (>75 cm), and reservoir plants (Van der Wolf and Perombelon, 1997). Its
distribution in potato fields can be spotty, and is commonly found in areas that have poor
drainage (Stevenson et al., 2001). It is adapted to low temperatures, however its survival
in very cold temperatures is reduced. In a study conducted in potato fields (Dirk van
Elsas et al., 2000), R. solanacearum race 3 biovar 2 population densities declined at 15
and 20°C and was severely reduced at 4°C. Severe drought negatively impacted
population densities. Race 3 biovar 2 is most severe between 24-35°C (optimal
temperature of 27°C) and decreases in virulence when temperatures exceed 35 °C or fall
below 10 °C (Stansbury et al., 2001). In regions such as Australia, England, Kenya, and
Sweden the organism was not detected in previously diseased potato fields after two
years, suggesting that long-term survival in temperate regions is reduced (Van der Wolf
and Perombelon, 1997). In another study the bacterium persisted for 12 months in potato
fields (Dirk van Elsas et al., 2000). It is spread through infected potato tubers and can
move plant-to-plant through the soil (Stevenson et al., 2001).
Host Range and Distribution
Ralstonia solanacearum is a widely distributed pathogen found in tropical,
subtropical, and some temperate regions of the world (Fegan and Prior, 2004). The
species as a whole has a very broad host range and infects hundreds of species in many
plant families. The majority of hosts are dicots with the major exception being bananas
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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and plantains. Most economically important host plants are found in the Solanaceae or
nightshade family (Stevenson et al., 2001).
Specific host range and distribution of R. solanacearum depends on the race and
to some degree the biovar of the pathogen (Daughtrey, 2003). These host ranges and
distributions have been changing in recent years. Race 1 is endemic to the southern
United States. Race 3 Biovar 2 is a USDA “select agent” listed on the Agricultural
Bioterrorism Act of 2002 and is subject to regulatory actions including strict quarantines
since potato is extremely sensitive to this race (Ji et al., 2004).
Table below illustrates the relationship of race, biovar, host range, and geographic
distribution (Daughtrey, 2003).
Race
Host Range
Geographic distribution
Biovar
1
Wide
Asia, Australia
3, 4
Americas
1
2
Banana, other Musa spp.
Caribbean, Brazil, Worldwide
1
3
Potato, some other Solanaceae,
Worldwide except US and
2
Geranium; few other species
Canada
4
Ginger
Asia
3,4
5
Mulberry
Management
The great variability of Ralstonia solanacearum and the strong influence of
environmental conditions on resistance make bacterial wilt a disease difficult to manage
(French, 1996).
Among the practices recommended, the use of resistant cultivars is considered to
play an important role. It is proven useful to control the potato strain, and is potentially
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
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the most effective and ideal way to manage the disease (Jyothi et al., 1993; French,
1996). However, despite its importance, little progress has been made by either nationals
or international potato breeding program in achieving stable resistance in commercial
genotypes (Martin and French, 1996; Lopes and Quezado-Soarez, 1994).
Resistance is not general, but pathogen specific; a pathogen at one location may
over come the resistance effective at another location. More than one pathogen may occur
in a given field. Existence of different pathogen within races may reduce effectively
resistance at certain location, acceptance level of resistance should be determined for
definite use. When use for consumption, a certain percentage of infection may be
tolerated. But if seed production, it is preferable not to tolerate any bacterial wilt, because
few infested seed tubers can spread the disease over a wide area. Since the expression of
resistance is pathovar, and environmental specific, an essential step in development of
resistant varieties is local screening (Martin and French, 1996).
Selection for Resistance to Bacterial Wilt
The first thing to consider when planting potato must be the variety. One must
choose a variety that is adaptable to the locality in order to achieve maximum production.
Using the right variety ensures high yield and better quality. Series of varietal evaluation
must be conducted to determine the performance of a new or previously untried variety
(HARRDEC, 1996).
Clonal evaluation and selection is important in the breeding program. The
standard procedure involves the selection of healthy looking and high yielding plants in
the field. Tubers of each selected clones are then harvested and kept separately to be
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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planted in the next season. Plants are carefully inspected for any abnormalities, and if
found in the first generation (F1), their clones are rejected and removed right away from
the field. Hence, successful potato production begins with long-term labor and intensive
breeding (Beukema, 1985).
Kurupuaracchi (1995) conducted an on farm potato evaluation and found that not
all clones were as superior as those in the on station trail. Out of the 22 clones, only 2
clones exhibited comparable level of yield ability, adaptability and stability with their
popular local variety. Thus, as a role, new clones usually differ in cultural
characteristics; therefore, several series of evaluations must be made at different strategic
locations and seasons.
Barrozo (2001), among the twelve potato varieties screened against bacterial wilt
under green house condition revealed that all varieties were resistant as no above ground
symptoms were exhibited. However, belowground symptom showed that all cultivars
were infected.
The study conducted by Botangen (2003) at Bosleng, Paoay, Atok, Benguet shows
that all the seven clones evaluated differed in growth, pest and disease resistibility,
yield as well as processing qualities. It is important to evaluate at different locations
and seasons for further screening.
Simongo (1992) found that clone LBR 1-5 was the most prolific in apical cutting
and tuberlet production among the several clones evaluated. On the different on farm
location trials, the performance of the individual clone varies with clone LBR 1-9 being
the most promising with an average yield of 21.2t/ha and showed high resistance to late
blight, good eating and tuber qualities.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Potato Breeding for Resistance to Bacterial wilt
The cultivated potato, Solanum tuberosum subsp. tuberosum (2n = 4x = 48), is
one of the most important world food crops and demands continued genetic improvement
to meet the needs of a changing world. Potato breeding has been a cumbersome task due
to inherent biological factors: cytoplasmic nuclear sterilities, tetrasomic inheritance and
inbreeding depression. In addition, yield stasis exists within the potato germplasm of
North America (Douches, 1996).
The first systematic potato-breeding program for resistance to bacterial wilt
started in 1967 by Rowe and Sequeira (1972) at the University of Wisconsin. These
investigators began their program by intercrossing several resistant diploid clones of
Solanum phureja from the Central Colombian Collection, with clones of S. tuberosum
ssp. tuberosum. Field tests of S. phureja x spp. tuberosum hybrids revealed susceptibility
of this genetic material to Phytophthora infestans. In a further set of crosses, the
bacterial wilt-resistant hybrids were therefore crossed with Mexican late blight-resistant
clones, combining the two needed resistances. The Mexican germplasm used in this
crossing program consisted of ssp. tuberosum clones containing late blight resistance
genes derived from the wild hexaploid species S. demissum (Schmiediche, 1986).
In 1969, the University of Wisconsin sent 369 clones, representing 10 families, to
Peru where they were to be tested under natural conditions in fields heavily infected with
P. solanacearum. French and Herrera started a screening program at Huambos in the
Department of Cajamarca, Peru (Herrera, 1972). By 1974 the Wisconsin material had
changed hands three times and only seven of the original 369 clones had survived. Most
of the material had been lost due to causes other than bacterial wilt.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Apart from sending 369 clones of bacterial wilt resistant genetic material to Peru,
the University of Wisconsin sent similar sets of material to 20 countries around the
world. The success of the Wisconsin material is partly due to its high degree of
heterozygosity, which results from use of unreduced gametes in the tertaploid x diploid
crosses which were necessary components of this breeding program. The bacterial wilt
resistance of this material comes from the diploid species S. phureja. Expression of this
resistance at the tertaploid level becomes even more remarkable when the genomic
composition of the tertaploid BR clones is considered. Only one genome, disregarding
crossing over during meiosis, is S. phureja origin. Crossing BR clones with other
tertaploid clones that are not resistant to bacterial wilt will obviously dilute the S.
phureja-based resistance even more (Schmiediche, 1986).
The wild potato relative Solanum commersonii is reported to have good high-
temperature resistance to brown rot of potatoes. However, S. tuberosum and S.
commersonii have different ploidy and endosperm balance numbers and are therefore
sexually incompatible, so their cells were fused by somatic hybridization. The resulting
somatic hybrid plants were vigorous and potato-like in appearance, but their resistance
level was unknown. Six of the somatic hybrids were examined, the S. commersonii and S.
tuberosum parents, and cv. Atlantic with a virulent strain of R. solanacearum (race 3,
biovar 2) at 28 ºC. The result reveals that S. commersonii was significantly more
resistant to brown rot than the cultivated potatoes. Encouragingly, preliminary results
show no significant difference in disease resistance between the somatic hybrids and the
S. commersonii parent. The somatic hybrids are both self-compatible and fertile to S.
tuberosum back-crosses (Laferriere et al., 1997).
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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CIP Breeding Program
In 1977, data received from the National Potato Programs in CIP’s regions had
suggested that resistance based on S. phureja was effective against strains of P.
solanacearum that affect potatoes, as long as the resistant material was grown under
temperate climatic conditions. Such conditions can be found in tropical highland regions
or in the plains of northern India during the winter.
Until 1980 the only source of resistance to bacterial wilt had been S. phureja,
which had demonstrated its potential as well as its limitations. The narrow genetic base
of this resistance had become an object of concern for breeders, pathologists and
geneticists. At the end of 1980, a new clone (AVRDC 1287.19) from Taiwan developed
at the Asian Vegetable Research Development Centre (AVRDC) was found out to be
resistant to bacterial wilt and has heat adaptation. The source of resistance was, however,
not S. phureja but the two wild species S. raphanifolium and S. chacoense. The latter
was used in hesitation since its resistance, so effective under Taiwanese conditions, had
broken down under field conditions in Peru, where a strain of P. solanacearum different
from the Taiwan strain was present. However, the chance to introduce a new source of
resistance to bacterial wilt into the existing gene pool, and combining two sources of
resistance into one progeny, overrode doubts about the usefulness of this clone in CIP’s
breeding program (Schmiediche, 1986).
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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MATERIALS AND METHODS
A. Evaluation using potato seed tubers
Germplasm
Materials
Healthy
G2 tubers of ten different potato clones obtained from the
International Potato Center (CIP) and JICA were used in the study. The evaluation was
conducted under green house condition. Treatments were arranged following the
Complete Randomized Design (CRD) with 4 replications.
The clones and cultivars evaluated and their characteristics are as follows:
Clone Cultivar’s
Origin
Type of resistance
Name
LB Wart
BW
LBR-5 Igorota Philippines R
387410.7 LBr-9
CIP
R
Warishiro
Japan
575003 I-931
India
R S S
676070 Cruza
155
Mexico R
380579.3 BW-III
CIP
MR S
720045 Atzimba Mexico MR
S S
573275 ASN-69-1
Mexico R R S
285378.27
None
CIP
MR MR
720071 Monserrate Columbia
MR
R S
The bacterial wilt nursery was thoroughly prepared. The area was divided into 4
blocks, each block consisting of 10 plots measuring 0.47 x 2meters each. Seven hundred
grams of triple 14 and 12.5 kilograms of chicken dung were applied to each block before
planting. During hilling–up, 700 grams of triple 14 was applied in each block. Dithane
and Ridomil were used to protect the plant from fungal diseases.
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Inoculum Preparation and Soil Inoculation
Four kilograms of infected potato tubers were cut into small cubes and were
incorporated in the soil. Another 8 kilograms of infected tubers were cut into smaller
pieces, allowed to ooze in 10 gallons of tap water and applied as soil drench.
To enhance the rapid multiplication of the bacteria, the inoculated soil was
irrigated periodically and covered with plastic sheets to increase the soil temperature.
Data Gathered
1. Percentage Bacterial Wilt Infection. This was obtained using the formula:
%
infection
=
no.
of infected plant X 100
total no. of plants
2. Weight of marketable tubers (kg). These are the tubers free from disease
infection.
3. Weight of non-marketable tuber (kg). Marble size, rotten and diseased tubers
was considered as non-marketable.
4. Total weight of tubers infected with bacterial wilt (kg). This was determined
by weighing all the tubers infected with bacterial wilt.
5. Late blight infection. This was asses using the CIP rating scale:
Blight (%) CIP
Scale Description
0
1
No blight to be seen
0.1
1
Very few plants in larger plots with a
lesion, not more than two lesions per
10 m of row (30 plants)
1
2
Up to 10 small lesions
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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3
3
Up to 30 small lesion per plant or up
to one leaflet in each 20 attacked
10
4
Most plants are visibly attacked, and
one out of three leaflets infected, few
multiple infections per leaflet.
25
5
Nearly every leaflet with lesion.
Multiple infections per leaflet are
common. Field or plants in plot are
affected.
50
6
Every plant effected and half the leaf
area destroyed by blight. Plots looks
green, fleaked and brown; blight is
very obvious.
75
7
As previous, but three quarters of
each plant affected by blight. Lower
branches maybe overwhelmingly
killed off and only green leaves, if
any, are at the top of the plant.
Shape of plants maybe more spindly
due to extensive foliar loss. Plot
looked neither brown nor green.
91
8
Some leaves and most stems are
green. Plot looks brown with some
green patches.
97
9
Few green leaves, most with lesions,
remain. Many stems with lesions.
Plots looks brown.
100
9
All leaves and stems dead. No
visible
blight
left
to
evaluate.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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B. Evaluation using Stem cuttings
Two weeks old rooted stem cuttings from NPRCRTC were used in the study.
Treatments were arranged following the Complete Randomized Design (CRD) with 3
replications at three plants per replicate.
The stem cuttings were planted in plastic cups with sterilized soil and maintained
at the Department of Plant Pathology Green house.
The entries were as follows:
Clone Cultivar’s
Name
LBR-5 Igorota
575003 I-931
676070 Cruza
155
573275 ASN-69-1
285378.27 -
Preparation of Inoculum and Inoculation Method
First Trial
Inoculum of bacterial wilt was sourced from infected tubers. The tubers were
properly washed, cut and squeezed for the bacterial ooze to come out and was diluted in
10 ml distilled water. Using a sterilized wire loop, the bacterial suspension was streaked
in previously plated casamino-peptone glucose agar (CPGA) consisting of the following
in g/li of water:
10 g dextrose
1 g casamino acid
10 g peptone
18 g agar
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Surface growth of the bacterium was scraped and diluted in the desired volume of
distilled water. The soil was cultivated near the root system to create wounds before
drenching 30 ml of the bacterial suspension in each plant.
After 3 days, only 2 among the 45 stem cuttings wilted.
Second
Trial
In the second trial, the same germplasm materials were used but the inoculation
method was modified. Before planting, roots hairs of the stem cuttings were cut and were
dipped in the bacterial suspension for 3 minutes to allow entry of the bacteria in the plant
system.
In addition, infected potato tubers were cut into smaller pieces and allowed to
ooze out in the desired volume of distilled water. The soil was cultivated near the root
system to create wounds before drenching 60 ml of the bacterial suspension in each of the
plant 16 days after the first inoculation.
Bacterial population was determined using the spectrophotometer.
Data Gathered
1. Bacterial population. This was obtained by using the spectrophotometer.
2. Number of days from planting to bacterial wilt expression. Days were counted
from inoculation to symptom expression.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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RESULTS AND DISCUSSION
Evaluation Using Potato Seed Tubers
Percentage Bacterial Wilt Infection
Twenty three (23) days after planting (DAP), the variety Igorota and clone
387410.7 had the highest percentage of bacterial wilt infection with a mean of 27.50, and
at 44 DAP, 100% plant were dead due to bacterial wilt (Table 1). On the other hand,
clones 720071, 285378.27, 380579.3 and Warishiro showed a slow rate of bacterial wilt
infection from 23 to 58 DAP, (Plates 1 and 2).
Among the 10 entries, only the clone 720071 still had plant that is not infected
with bacterial wilt after 58 DAP.
The high incidence of bacterial wilt may have been influenced by the prevailing
weather conditions. Under greenhouse condition, the temperature was relatively higher
than the ambient temperature. The maximum and minimum temperatures during the last
week of September (BW rating started) were 26ºC and 17ºC respectively, compared to
the ambient condition with a maximum temperature of 24.5ºC and a minimum of 16.8ºC
(Table 2). The prevailing weather condition during the conduct of the study favors R.
solanacearum development as cited by Persley (1985) that the optimum temperature for
R. solanacearum development is from 25-35ºC. Moreover, Martin and French (1996)
cited that disease development of bacterial wilt is mainly influenced by temperature.
High temperature promoted bacterial wilt development.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Table 1. Mean percentage bacterial wilt infection of different clones/variety evaluated
CLONE/
DAYS
AFTER
PLANTING
VARIETY
23 30 37 44 51 58
Igorota 27.50
60.00a
80.00ab
100a 100a
100
387410.7
27.50
47.50ab
77.50a 100a 100a
100
Warishiro
2.50
15.00c
27.50bcd
57.50cde 90.00a
100
575003
15.00
30.00bc
52.50abc
90.00ab 100a
100
676070
10.00
30.00bc
50.00abc
77.50abc 97.50a 100
380579.3
5.00
12.50c
27.50cd
52.50cde 85.00ab 100
720045
7.50
22.50bc
37.50bcd
70.00bcd 95.00a 100
73275
7.50
22.50bc
47.50abc
75.00abc 92.50a
100
285378.27 7.50
7.50c
17.50cd
45.00d 72.50bc 100
720071
2.50
5.00c
12.50d
35.00e 60.00c 87.50
Means followed by a common letter are not significantly different at 5% level by DMRT
Plate 1. Clone 380579.3 showing wilt symptom at 30 DAP
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Plate 2. Infected tubers from clone 285378.27
Plate 3. Infected tubers from clone 676070
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Table 2. Mean weekly weather data
TEMPERATURE
AVERAGE
TOTAL
MONTH
ºC
RAINFALL
RELATIVE BRIGHT
Max Min
(mms and
HUMIDITY SUNSHINE
Tenths)
(%)
(mm)
______________________________________________________________________
September
Week 1
24.1 17.4
24.6
90
193.7
Week 2
25.1 17.7
7.2
89
169.7
Week 3
22.2 16.7
57.5
90
53.1
Week 4
24.5 16.8
2.3
87
293.1
October
Week 1
24.4 16
20
89
333.4
Week 2
24
15.6
10.4
86
252.8
Week 3
25.3 16
1.0
87
363.4
Week 4
24.9 16.2
3.0
84
287.1
November
Week 1
24
16.2
0.9
82
256.8
Week 2
25.6 15.4
4.2
82
321.1
Week 3
24.4 14.4
1.3
83
341.5
Week 4
25.6 15.3
0.0
77
322.2
February
Week 1
23.6 14.2
0.0
79
-
Week 2
25.4 14.3
2.3
80
-
Week 3
26
13.2
0.0
80
-
Week 4
26.9 12.9
0.0
74
-
Percentage Late Blight Infection
Among the 10 entries, clone 720071 showed a slow development rate of late
blight infection and was the only clone that reached 58 DAP (Table 3). Conversely,
clones 676070, 720045, 573275, 285378.27 and Warishiro, reached 51 DAP and had a
late blight rating ranging from 1 and 2, where no blight was seen; or very few plants in
larger plots with lesions and 2 with a maximum of 10 small lesions. On the other hand,
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
21
clone 387410.7 and variety Igorota only reached 37 DAP and had late blight ratings of 2.
The variety Igorota is rated as resistant to late blight. However the variety exhibited late
blight infection early in its growing period but is able to regenerate new leaves and
matures at 100-120 days.
Most of the entries are rated to be resistant to late blight but their resistance was
not quantified due to the early infection of bacterial wilt which caused their early
senescence.
The fluctuation of the prevailing weather condition played a great factor for the
manifestation of resistance or susceptibility of the clones/variety to late blight infection.
Thung (1974) explained that any environmental fluctuation may upset the pathogen and
shift host reaction to low and higher resistance or susceptibility.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
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Table 3. Mean percentage late blight infection of different clones/variety evaluated
CLONE/
DAYS
AFTER
PLANTING
VARIETY
30
37
44
51
58
________________________________________________________________________
Igorota
1.25bc
2.0
387410.7
2.00a
2.25
Warishiro
1.50b
1.75
3.0ab
2.0
575003
1.0c
1.25
2.0bcd
676070
1.0c
1.0
1.5d
1.5
380579.3
1.0c
1.25
2.75abc
2.75
720045
1.0c
1.50
2.50abcd 2.0
573575
1.0c
1.25
1.75cd
2.0
285378.27
1.0c
1.50
1.75cd
1.5
720071
1.0c
1.00
2.0bcd
1.75
1.5
Means followed by a common letter are not significantly different at 5% level by DMRT
Yield Parameters
Results reveal that clone 676070 had the highest marketable yield with a mean of
575g but also had the highest non marketable and bacterial wilt infected tubers (Table 4).
On the other hand, the least weight of marketable tubers was obtained in clone 720071
with a mean of 36.25g which also registered the lowest in non marketable tubers and
second lowest in bacterial wilt infected tubers with a mean of 21.25g.
The variety Igorota which is a late maturing variety registered one of the lowest
marketable tubers with a mean of 58.75g. Its low marketable yield is associated with its
early senescence due to bacterial wilt infection.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
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All the 10 entries evaluated were able to produce marketable tubers despite the
early bacterial wilt infection making them all tolerant to bacterial wilt.
Table 4. Yield parameters of different clones/variety evaluated
CLONE/ MARKET
ABLE
(g)
NON
BACTERIAL
WILT
VARIETY
MARKET
ABLE
(g)
INFECTED
(g)
Igorota
58.75
66.25
41.25
387410.7
48.75
55.00
3.75
Warishiro
72.50
52.50
8.75
575003
95.00
126.25
51.25
676070
575.00
250.00
388.75
380579.3
218.75
167.50
72.50
720045
82.50
113.75
25.00
573275
271.25
183.75
111.25
285378.27
161.25
193.75
193.75
720071
36.25
20.00
21.25
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
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Plate 4. Marketable, non-marketable, BW infected tubers of variety Igorota
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
25
Evaluation Using Stem Cuttings
Number of days from planting to bacterial wilt expression
Table 5 shows the number of days from planting to bacterial wilt expression.
Clone 285378.27 manifested bacterial wilt symptom at the shortest time having a mean of
24.210 days. The clone 575003 on the other hand exhibited the longest time of symptom
expression at a mean of 25.20 days.
The recorded temperature in the greenhouse during the month of February was
relatively higher than the ambient temperature. It was during the 3rd week of February
that wilting was observed in the sample plants. The maximum temperature in the green
house was 27ºC and minimum of 14ºC, compared to the ambient condition with a
maximum of 26ºC and a minimum of 13.2ºC (Table 2). Persley (1985) cited that the
optimum development of R. solanacearum is from 25-35ºC.
Table 5. Average number of days from transplanting to bacterial wilt symptom
expression
CLONES/
MEAN
NUMBER
OF
DAYS
CULTIVAR AFTER TRANSPLANTING
________________________________________________________________________
Igorota
24.887
575003 25.200
676070 24.663
573275 24.773
285378.27
24.210
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
26
Table 6 presents the bacterial population that was used during inoculation in the
first and second trial. Results revealed that there is a significant difference from the
bacterial population during the first and second trial.
During the termination of the study, not all the sample plants wilted. The
assymptomatic cuttings were tested using the water blank test which revealed that they
are infected with bacterial wilt (Plates 4 and 5).
The unsuccessful result obtained from the first trial may be due to the low
bacterial population that was inoculated to the sample cuttings. Nielson and Hyness
(1979), as cited by Nagpala (1986) reported that the possible variation of resistance or
susceptibility of crops were affected by the age of the plant at the time of inoculation. It
was reported that the best time to inoculate the plants should be about 4 weeks after
planting.
Table 6. Bacterial population of inoculum used during the trials
SAMPLE
AVERAGE BACTERIAL
POPULATION
TRIAL 1
TRIAL 2
DIFFERENCE
________________________________________________________________________
1
0.162
0.958
0.796
2
0.130
0.997
0.867
3
0.133
1.000
0.867
________________________________________________________________________
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
27
Plate 5. Wilted potato cuttings at 16 days after inoculation
Plate 6. Assymptomatic cuttings being tested for BW infection
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
28
SUMMARY, CONCLUSION AND RECOMMENDATION
Summary
Nine clones: 387410.7, Warishiro, 575003, 676070, 380579.3, 720045, 573275,
285378.27 and 720071 and the variety Igorota were evaluated to identify germplasm
materials that were tolerant to bacterial wilt infection under green house condition and to
determine their reaction to late blight infection.
Among the 10 entries evaluated, Igorota and clone 387410.7 showed the highest
percentage bacterial wilt infection at 23 DAP. On the other hand, clone 720071 exhibited
the slowest rate of bacterial wilt infection followed by clones 285378.27, 380579.3 and
Warishiro respectively.
Clones 720071, 285378.27 and 676070 showed resistance to late blight infection
with a rating of 1 or no blight or a maximum of 10 small lesions observed in the sample
plant.
The highest marketable yield was obtained from clone 676070 which ironically
registered the highest nonmarketable and bacterial wilt infected tubers.
Conclusion
Based on the results of the study, all the 10 entries evaluated were tolerant to
bacterial wilt as they were able to produce marketable tubers despite bacterial wilt
infection. Clones 720071, 285378.27 and 676070 confirmed their resistance to late blight
infection.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
29
Recommendation
Further evaluation must be conducted to assess the adaptability and yield of the
germplasm materials.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
30
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Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
34
APPENDICES
Appendix Table 1. Percentage bacterial wilt infection at 23 DAP
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
10 10
70
20
110
27.50
387410.7
50
20
20 20
110
27.50
Warishiro
10
0 0
0
10
2.50
575003
20 20
20
0 60
15
676070
10
0
10
20 40
10
380579.3
10
0
10
0 20
5
720045
0
20
10
0 30
7.5
573275
10
20
0
0 30
7.5
285378.27
10
10 0
20 30
7.5
720071
0
0 10
0 10
2.5
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
3.24 3.24
8.39 4.53 19.40
4.850
387410.7
7.11 4.53
4.53 4.53 20.70
5.175
Warishiro
3.24 0.71
0.71 0.71 5.37
1.343
575003
4.53 4.53
4.53 0.71 14.30
3.575
676070
3.24 0.71
3.24 4.53 11.75
2.930
380579.3
3.24 0.71
3.24 0.71 7.90
1.975
720045
0.71 4.53
3.24 0.71 9.19
2.210
573275
3.24 4.53
0.71 0.71 9.19
2.210
285378.27
3.24 0.71
0.71 4.53 9.19
2.210
720071
0.71 0.71
3.24 0.71 5.37
1.343
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
327.478 109.159
0.6702
Factor A
9
3112.413 345.824
2.1233ns 2.25 3.14
Error
27
4397.498 162.870
________________________________________________________________________
TOTAL
39
7,837.388
ns= not significant
Coefficient of Variation = 62.05%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Appendix Table 2. Percentage bacterial wilt infection at 30 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 60 40 100 40 240 60
387410.7
60 40 50 40 190
47.5
Warishiro
40 0 20 0 60 15
575003 40 40 40 0 120 30
676070 30 30 10 50 120 30
380579.3 20 10 20 0 50 12.5
720045 10 40 40 0 90 22.5
573275 30 40 0 20 90
22.5
285378.27 0
0
0
30 30 7.5
720071 0 0 20 0 20 5
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
7.78 6.36
10.02 6.36 30.52
7.630
387410.7
7.78 6.36
7.11 6.36 27.61
6.903
Warishiro
6.36 0.71
4.53 0.71 12.31
3.078
575003
6.36 6.36
6.36 0.71 19.79
4.948
676070
5.52 5.52
3.24 7.11 21.39
5.348
380579.3
4.53 3.24
4.53 0.71 13.01
3.253
720045
3.24 6.36
6.36 0.71 16.67
4.168
573275
5.52 6.36
0.71 4.53 17.12
4.280
285378.27
0.71 0.71
0.71 5.32 7.45
1.863
720071
0.71 0.71
4.53 0.71 6.66
1.665
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
907.490 302.497
0.9879
Factor A
9
11022.418 1224.713 3.997**
2.54 3.14
Error
27
8267.390 306.200
________________________________________________________________________
TOTAL
39
20,197.298
**= Highly Significant
Coefficient of Variation = 69.30%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
38
Appendix Table 3. Percentage bacterial wilt infection at 37 DAP
BLOCK
CLONE/
VARIETY
I II III IV
TOTAL
MEAN
Igorota 70 80 100 70 320 80
387410.7
80 60 90 80 310
77.5
Warishiro 50
0
40 20 110 27.5
575003 60 50 70 30 210
52.5
676070 40 60 40 60 200 50
380579.3 40 40 30 0 110 27.5
720045 20 70 60 0 150 37.5
573275 50 60 30 50 190
47.5
285378.27
20 20 0 30 70
17.5
720071 10 0 30 10 50 12.5
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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39
Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
8.39 8.97
10.2 8.39 35.77
8.943
387410.7
8.97 7.78
9.51 8.97 35.23
8.808
Warishiro
7.11 0.71
6.36 4.53 18.71
4.678
575003
7.78 7.11
4.13 5.52 24.54
6.135
676070
6.36 7.78
6.36 7.78 28.28
7.070
380579.3
3.36 6.36
5.52 0.71 18.95
4.738
720045
4.53 8.39
7.78 0.71 21.41
5.353
573275
7.11 7.78
5.52 7.11 27.52
6.880
285378.27
4.53 4.53
0.71 5.52 15.29
3.823
720071
3.24 0.71
5.52 3.24 12.71
3.178
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
2507.50 835.833
1.8547
Factor A
9
15162.50 1684.722 3.7384** 2.25 3.14
Error
27
1216.50 450.648
________________________________________________________________________
TOTAL
39
29,837.50
**= Highly Significant Coefficient of Variation = 37.26%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
40
Appendix Table 4. Percentage bacterial wilt infection at 44 DAP
BLOCK
CLONE/
VARIETY
I II III IV
TOTAL
MEAN
Igorota 100 100 100 100 400 100
387410.7
100 100 100 100 400 100
Warishiro
70 40 60 60 230
57.5
575003 100 100 90 70 360 90
676070 60 100 70 80 310 77.5
380579.3
70 50 50 40 210
52.5
720045 40 100 100 40 280 70
573275 70 100 50 80 300 75
285378.27
40 40 40 60 180 45
720071 40 20 50 30 140 35
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
41
Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
10.02 10.02
10.2 10.02 40.08 10.02
387410.7
10.02 10.02
10.02 10.02 40.08 10.02
Warishiro
8.39 6.36
7.78 7.78 30.31 7.578
575003
10.02 10.02
9.51 8.39 37.94 9.485
676070
7.78 10.02
8.39 8.97 35.16 8.790
380579.3
8.39 7.11
7.11 6.36 28.97 7.243
720045
6.36 10.02
10.02 6.36 32.76 8.190
573275
8.39 10.02
7.11 8.97 34.49 8.623
285378.27
6.36 6.36
6.36 7.78 26.86 6.715
720071
6.36 4.53
7.11 5.52 23.52 5.880
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
427.50 142.50
0.4998
Factor A
9
18372.50 2041.38
7.1604** 2.54 3.14
Error
27
7697.50 285.093
________________________________________________________________________
TOTAL
39
26,497.50
**=Highly Significant
Coefficient of Variation = 24.04%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
42
Appendix Table 5. Percentage bacterial wilt infection at 51 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 100 100 100 100 400 100
387410.7
100 100 100 100 400 100
Warishiro 90
70 100 100 360 90
575003 100 100 100 100 400 100
676070 100 100 90 100 390 97.5
380579.3 100 100 70
70 340 85
720045 100 100 100 80 380 95
573275 100 100 70 100 370 92.5
285378.27
70 80 60 80 290
72.5
720071 60 50 60 70 240 60
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
43
Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
10.02 10.02
10.02 10.02 40.08 10.02
387410.7
10.02 10.02
10.02 10.02 40.08 10.02
Warishiro
9.51 8.39
10.02 10.02 37.94 9.485
575003
10.02 10.02
10.02 10.02 40.08 10.02
676070
10.02 10.02
10.02 10.02 39.57 9.893
380579.3
10.02 10.02
9.51 8.39 36.82 9.205
720045
10.02 10.02
10.02 8.97 39.03 9.758
573275
10.02 10.02
8.39 10.02 38.45 9.613
285378.27
8.39 8.97
7.78 8.97 34.11 8.528
720071
7.78 7.11
7.78 8.39 31.06 7.765
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
267.50 89.167
0.8731
Factor A
9
6452.50 716.944
7.0199**
2.54 3.14
Error
27
2757.50 102.130
________________________________________________________________________
TOTAL
39
24,177.50
**=Highly Significant
Coefficient of Variation = 11.32%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
44
Appendix Table 6. Percentage bacterial wilt infection at 58 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 100 100 100 100 400 100
387410.7
100 100 100 100 400 100
Warishiro
100 100 100 100 400 100
575003 100 100 100 100 400 100
676070 100 100 100 100 400 100
380579.3
100 100 100 100 400 100
720045 100 100 100 100 400 100
573275 100 100 100 100 400 100
285378.27
100 100 100 100 400 100
720071 90 80 80 100 350 87.5
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
45
Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
10.02 10.02
10.02 10.02 40.08 10.02
387410.7
10.02 10.02
10.02 10.02 40.08 10.02
Warishiro
10.02 10.02
10.02 10.02 40.08 10.02
575003
10.02 10.02
10.02 10.02 40.08 10.02
676070
10.02 10.02
10.02 10.02 40.08 10.02
380579.3
10.02 10.02
10.02 10.02 40.08 10.02
720045
10.02 10.02
10.02 10.02 40.08 10.02
573275
10.02 10.02
10.02 10.02 40.08 10.02
285378.27
10.02 10.02
10.02 10.02 40.08 10.02
720071
9.51 8.97
8.97 10.02 37.47 9.368
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
27.50 9.167
Factor A
9
562.50 62.50
6.8182**
2.54 3.14
Error
27
247.50 9.167
________________________________________________________________________
TOTAL
39
837.50
**=Highly Significant
Coefficient of Variation = 3.07%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
46
Appendix Table 7. Late blight infection rating at 30 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 1 1 2 1 5
1.25
387410.7
2 2 2 2 8 2.0
Warishiro
1 2 1 2 6 1.5
575003 1 1 1 1 4 1
676070 1 1 1 1 4 1
380579.3
1 1 1 1 4 1
720045 1 1 1 1 4 1
573275 1 1 1 1 4 1
285378.27
1 1 1 1 4 1
720071 1 1 1 1 4 1
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
0.075 0.025
0.4030
Factor A
9
4.025
0.447
7.2090**
2.54 3.14
Error
27
1.675
0.062
________________________________________________________________________
TOTAL
39
5.775
**=Highly Significant
Coefficient of Variation = 21.20%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
47
Appendix Table 8. Late blight infection rating at 37 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 2 2 2 2 8 2
387410.7
2 2 3 2 9
2.25
Warishiro
2 2 1 2 7
1.75
575003 2 1 1 1 5
1.25
676070 1 1 1 1 4 1.0
380579.3
1 2 1 1 5
1.25
720045 2 1 1 2 6
1.50
573275 1 1 1 2 5
1.25
285378.27
1 2 1 2 6
1.50
720071 1 1 1 1 4 1.0
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
0.475 0.158
0.8104
Factor A
9
6.225
0.692
3.5403**
2.54 3.14
Error
27
5.275
0.195
________________________________________________________________________
TOTAL
39
11.975
**=Highly Significant
Coefficient of Variation = 29.97%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
48
Appendix Table 9. Late blight infection rating at 44 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 3 3 3 2 11
2.75
387410.7
4 3 4 3 14
3.5
Warishiro
4 2 3 3 12
3.0
575003 3 1 3 1 8 2.0
676070 1 2 2 1 6 1.5
380579.3
3 3 3 2 11
2.75
720045 3 2 3 2 10
2.5
573275 2 2 1 2 7
1.75
285378.27
1 3 1 2 7
1.75
720071 2 2 2 2 8 2.0
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
2.10 0.70
1.5882
Factor A
9
15.10
1.678
3.8067** 2.54 3.14
Error
27
11.90
0.441
________________________________________________________________________
TOTAL
39
29.10
**=Highly Significant
Coefficient of Variation = 28.25%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
49
Appendix Table 10. Late blight infection rating at 51 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 2 3 1 2 8 2
387410.7
4 3 4 3 14
3.5
Warishiro
2 2 2 2 8 2.0
575003 2 2 3 1 8 2.0
676070 1 2 2 1 6 1.5
380579.3
3 3 3 2 11
2.75
720045 2 2 2 2 8 2.0
573275 2 2 2 2 8 2.0
285378.27
1 2 1 2 6 1.5
720071 1 2 2 2 7
1.75
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
1.0 0.33
1.20
Factor A
9
13.10
1.456
5.24** 2.25 3.14
Error
27
7.500
0.278
________________________________________________________________________
TOTAL
39
21.60
**=Highly Significant
Coefficient of Variation = 25.10%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
50
Appendix Table 11. Late blight infection rating at 58 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 2 1 1 1 5
1.25
387410.7
3 2 2 2 9
2.25
Warishiro
2 1 1 1 5
1.25
575003 2 2 1 1 6
1.50
676070 1 2 1 1 5
1.25
380579.3
2 2 1 1 6
1.50
720045 2 1 1 2 6
1.50
573275 1 1 1 2 5
1.25
285378.27
1 2 2 2 6
1.50
720071 1 2 2 1 6
1.50
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
1.475 0.492 1.8247
Factor A
9
3.225 0.358
1.3299ns 2.54 3.14
Error
27
7.275 0.269
________________________________________________________________________
TOTAL
39
11.975
ns=Not Significant
Coefficient of Variation = 35.19%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
51
Appendix Table 12. Weight of marketable tubers (g)
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota 55 180 0 0 235
58.75
387410.7 160
0
0
35 195 48.75
Warishiro 245
0
0
45
290 72.5
575003 225 0 0 155 380 95
676070 600 665 625 410 2300 575
380579.3
250 400 190 35 875
218.75
720045 125 75 90 40 330
82.5
573275 230 260 240 355 1085
271.5
285378.27
0 245 200 200 645
161.25
720071 0 145 0 0 145
63.25
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
52
Transformed data
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota 7.45 13.44 0.71 0.71 22.31 5.578
387410.7
12.67 0.71 0.71 5.96 20.05 5.013
Warishiro
15.67 0.71 0.71 6.75 23.84 5.960
575003 15.02 0.71 0.71 12.47 28.91 7.228
676070 24.51 25.80 25.01 20.26 95.58 23.895
380579.3 15.83 20.01 13.80 5.96 55.6 13.900
720045 11.20 8.69 9.51 6.36 35.76 8.940
573275 15.18 16.14 15.51 18.85 65.68 16.420
285378.27
0.71 15.67 14.16 14.16 44.7 11.175
720071 0.71 12.06 0.71 0.71 14.19 3.548
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Replication
3
94.424 31.475 1.0297
Factor A
9
1447.218 160.802 5.2609**
2.54 3.14
Error
27
825.274 30.566
________________________________________________________________________
TOTAL
39
2366.915
**=Highly Significant
Coefficient of Variation = 54.39%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
53
Appendix Table 13. Weight of non- marketable tubers (g)
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota 60 105 5 95 265
66.25
387410.7
140 55 25 0 220 55
Warishiro
100 45 15 50 210
52.5
575003 100 165 110 130 505
126.25
676070 200 220 80 500
10000
250
380579.3
300 115 145 110 670 167.5
720045 150 75 35 195 455
113.75
573275 165 200 80 290 735
183.75
285378.27
370 130 120 155 775 193.5
720071 0 50 10 20 80 20
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
54
Transformed data
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota 7.78 10.27 2.35 9.77 30.17 7.542
387410.7
11.85 7.45 5.05 0.71 25.06 6.265
Warishiro
10.02 6.75 3.94 7.11 27.81 6.953
575003 10.02 12.86 10.52 11.42 44.81 11.205
676070 14.16 14.85 8.97 22.37 60.35 15.088
380579.3 17.33 10.75 12.06 10.51 50.65 12.663
720045 12.27 8.69 5.96 13.98 40.9 10225
573275 12.86 14.16 8.97 17.04 53.03 13.258
285378.27
19.25 11.42 10.98 12.47 54.12 13.530
720071 0.71 7.11 3.24 4.53 15.59 3.898
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Replication
3
116.104 38.701 3.8435
Factor A
9
496.030 55.114 5.4735**
2.54 3.14
Error
27
271.870 10.069
________________________________________________________________________
TOTAL
39
884.004
**=Highly Significant
Coefficient of Variation = 31.53%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
55
Appendix Table 14. Weight of Infected tubers (g)
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota
110 45 0 10 165
41.25
387410.7
15 0 0 0 15
3.75
Warishiro 0
0
0
35 35 8.75
575003 80 40 75 10 205
51.25
676070 150 350 760 295 1555
388.75
380579.3
50 180 15 45 290 72.5
720045 30 20 40 10 100 25
573275 115 5 255 70 445
11.25
285378.27
195 205 65 310 775
193.75
720071 35 50 0 0 85
21.25
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
56
Transformed data
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota 10.51 6.75 0.71 3.24 21.21 5.303
387410.7
3.94 0.71 0.71 0.71 6.07 1.518
Warishiro
0.71 0.71 0.71 5.96 8.09 2.023
575003 8.97 6.36 8.69 3.24 27.26 6.815
676070 12.27 18.72 27.58 17.19 75.76 18.94
380579.3 7.11 13.44 3.94 6.75 31.24 7.810
720045 5.52 4.53 6.36 3.24 19.65 4.913
573275 10.75 2.35 15.98 8.40 37.48 9.370
285378.27
13.98 14.34 8.09 17.62 54.03 13.508
720071 5.96 7.11 0.71 0.71 14.49 3.623
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Replication
3
8.204 2.735
0.1642
Factor A
9
1052.941 116.993 7.0271** 2.54 3.14
Error
27
449.579 16.649
________________________________________________________________________
TOTAL
39
15110.665
**=Highly Significant
Coefficient of Variation = 55.27%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
57
Appendix Table 15. Number of days from transplanting to bacterial wilt symptom
expression
CLONE/
REPLICATE
VARIETY
I II III
TOTAL
MEAN
1 23.33 26.00 26.00 74.66 24.887
2 24.66 26.00 26.00 75.60 25.200
3 23.66 24.66 26.00 73.99 24.663
4 23.30 25.33 26.00 74.32 24.773
5 23.33 23.66 23.3 72.63 24.210
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
2
9.401 4.701 3.9895
Factor A
4
7.562 0.391
0.3315ns 3.84 7.01
Error
18
9.426 1.178
________________________________________________________________________
TOTAL
14
20.390
ns= Not Significant
Coefficient of Variation = 4.39%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
BASNGI, JAMIE BETH B. March 2006. Germplasm Evaluation for Tolerance
Against Bacterial Wilt (Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.). Benguet
State University, La Trinidad, Benguet.
Adviser: Jocelyn C. Perez, MSc.
ABSTRACT
Nine clones: 387410.7, Warishiro, 575003, 676070, 380579.3, 720045, 573275,
285378.27 and 720071 and variety Igorota were screened to identify potato germplasm
materials tolerant to bacterial wilt infection and resistant to late blight infection under
green house condition.
All entries were found to be tolerant to bacterial wilt as they were able to produce
marketable tubers despite the bacterial wilt infection recorded.
The clones that showed resistance to late blight infection were clones 720071,
285378.27 and 676070 respectively.
TABLE OF CONTENTS
Page
Title Page ……………………………………………………………………..
i
Abstract ………… …………………………………………………………..
i
Table of Contents …………………………………………………………….
ii
INTRODUCTION ……………………………………………………………
1
REVIEW OF LITERATURE
Causal Organism ……………………………………………………….
4
Epidemiology …………………………………………………………..
4
Host Range and Distribution …………………………………………...
5
Management ……………………………………………………………
6
Selection for Resistance to Bacterial Wilt ……………………………..
7
Potato Breeding for Resistance to Bacterial Wilt ……………………...
9
CIP Breeding Program …………………………………………………
11
MATERIALS AND METHODS
Evaluation using Potato Seed Tubers …………………………………..
12
Inoculation Preparation and Soil Inoculation ………………………….
13
Data Gathered …………………………………………………………..
13
Evaluation using Rooted Stem Cuttings ……………………………...
15
Preparation of Inoculum and Inoculation Method ……………………..
15
Data Gathered ………………………………………………………….
16
ii
RESULTS AND DISCUSSION
Percentage Bacterial Wilt Infection ……………………………………
17
Percentage Late Blight Infection ……………………………………….
20
Yield Parameters ……………………………………………………….
22
Number of Days from Planting to Bacterial Wilt Expression …………
25
SUMMARY, COCLUSION AND RECOMMENDATION
Summary ……………………………………………………………….
28
Conclusion ……………………………………………………………..
28
Recommendation ……………………………………………………....
29
LITERATURE CITED ……………………………………………………….
30
APPENDICES ………………………………………………………………..
34
iii
INTRODUCTION
Nature of the Study
Potato
(Solanum tuberosum Linn) is grown worldwide due to its nutritive and
money making value. It is one of the mankind’s most valuable food because it provides a
source of low cost energy to human diet and a source of minerals, proteins, vitamin A and
B2, carbohydrates and some elements like potassium and phosphorus (Kipps, 1979).
A single medium-sized potato contains about half the daily adult requirement of
vitamin C. Potato is very low in fat, with just 5 percent of the fat content of wheat, and
one-fourth the calories of bread. When it is boiled, it has more protein than maize, and
nearly twice the calcium (CIP, 1996).
In the Philippines, potato ranks third among the leading commercial vegetables in
terms of peso value and ranks first among the vegetable crops in Northern Luzon
(Buasen, 1978). In like manner, it ranks fourth among the major crops worldwide after
wheat, maize and rice.
However, different pest and diseases affect the production of potato. Bacterial
wilt caused by Ralstonia solanacearum (E.F. Smith) Yabuuchi et al. is one of the most
important, widespread and lethal bacterial diseases of plants (Ma, 1990). It limits the
production of potatoes, especially seed potatoes worldwide (CIP, 1992; Martin and
French, 1985; Schmiediche, 1984 and Rich, 1983).
Kelman (1953) considered the earliest known record of bacterial wilt to be on
tobacco in Indonesia in 1864, where entire fields were lost due to bacterial wilt. Bacterial
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
2
wilt was recorded on potatoes in the United States in 1890 (Kelman, 1953) and in
Australia in 1894 (Tyron, 1895).
The disease was reported in the Philippines by Reinking (1918) affecting tobacco,
pepper and tomato as cited by Valdez (1986). In 1922, Welles and Roldan reported that
it caused a serious disease on solanaceous crops in Southern Luzon. Series of study
conducted reveals that it became severe throughout the highland and lowland areas
producing potato (Perez et al., 1997).
The survival of the bacterium is influenced by temperature, humidity and other
physical and chemical soil factors. R. solanacearum may survive for many years in
certain soils and may disappear from one growing season to the next (Martin and French,
1996).
Bacterial wilt can be reduced only if various control components are combined.
An integrated disease management approach can lead to significant reduction or even
eradication of bacterial wilt (CIP, 1996).
Among the practices recommended, the use of resistant cultivars is considered to
play an important role. It is proven useful to control the potato strain, and is potentially
the most effective and ideal way to manage the disease (Jyothi et. al., 1993; French,
1996). It also delays the development of populations of insects resistant to the pesticide
currently being used (Ganga, 1992).
The study was conducted to screen potato clones and cultivars for tolerance to
bacterial wilt under green house condition, and evaluate their reaction to late blight
infection.
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The study was conducted at the bacterial wilt nursery, the Northern Philippines
Root Crops Research and Training Center (NPRCRTC) and the Department of Plant
Pathology laboratory, Benguet State University from September 2005 to February 2006.
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REVIEW OF LITERATURE
Causal Organism
The phytopathogen Ralstonia solanacearum has over 5000 genes, many of which
probably facilitate bacterial wilt disease development (Brown and Allen, 2001). It is a
gram-negative plant-pathogenic bacterium that causes bacterial wilt in a variety of plants
(Hayward, 1994).
It is a soil-borne pathogen that naturally infects roots. It exhibits a strong and
tissue-specific tropism within the host, specifically invading and highly multiplying in the
xylem vessels (Smith, 1896 and Yabuuchi et al., 1995). Once established in the xylem
vessels, the bacteria are able to enter the intercellular spaces of the parenchyma cells in
the cortex and pith in various areas of the plant. Here, R. solanacearum is able to
dissolve the cell walls and create slimy pockets of bacteria and cell debris. Production of
highly polymerized polysaccharides increases the viscosity of the xylem, which results in
plugging (Shew and Lucas, 1991).
High temperatures and high soil moisture generally favors Ralstonia
solanacearum, the exception being certain in Race 3 strains that are pathogenic on potato
and are able to grow well at lower temperatures (Stevenson et al., 2001). In certain soil,
R. solanacearum may survive for many years, in others, the bacterium may disappear
from one growing season to the next (Martin and French, 1996).
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EPIDEMIOLOGY
Ralstonia solanacearum race 3 is a soil borne pathogen that persists in wet soils,
deep soil layers (>75 cm), and reservoir plants (Van der Wolf and Perombelon, 1997). Its
distribution in potato fields can be spotty, and is commonly found in areas that have poor
drainage (Stevenson et al., 2001). It is adapted to low temperatures, however its survival
in very cold temperatures is reduced. In a study conducted in potato fields (Dirk van
Elsas et al., 2000), R. solanacearum race 3 biovar 2 population densities declined at 15
and 20°C and was severely reduced at 4°C. Severe drought negatively impacted
population densities. Race 3 biovar 2 is most severe between 24-35°C (optimal
temperature of 27°C) and decreases in virulence when temperatures exceed 35 °C or fall
below 10 °C (Stansbury et al., 2001). In regions such as Australia, England, Kenya, and
Sweden the organism was not detected in previously diseased potato fields after two
years, suggesting that long-term survival in temperate regions is reduced (Van der Wolf
and Perombelon, 1997). In another study the bacterium persisted for 12 months in potato
fields (Dirk van Elsas et al., 2000). It is spread through infected potato tubers and can
move plant-to-plant through the soil (Stevenson et al., 2001).
Host Range and Distribution
Ralstonia solanacearum is a widely distributed pathogen found in tropical,
subtropical, and some temperate regions of the world (Fegan and Prior, 2004). The
species as a whole has a very broad host range and infects hundreds of species in many
plant families. The majority of hosts are dicots with the major exception being bananas
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and plantains. Most economically important host plants are found in the Solanaceae or
nightshade family (Stevenson et al., 2001).
Specific host range and distribution of R. solanacearum depends on the race and
to some degree the biovar of the pathogen (Daughtrey, 2003). These host ranges and
distributions have been changing in recent years. Race 1 is endemic to the southern
United States. Race 3 Biovar 2 is a USDA “select agent” listed on the Agricultural
Bioterrorism Act of 2002 and is subject to regulatory actions including strict quarantines
since potato is extremely sensitive to this race (Ji et al., 2004).
Table below illustrates the relationship of race, biovar, host range, and geographic
distribution (Daughtrey, 2003).
Race
Host Range
Geographic distribution
Biovar
1
Wide
Asia, Australia
3, 4
Americas
1
2
Banana, other Musa spp.
Caribbean, Brazil, Worldwide
1
3
Potato, some other Solanaceae,
Worldwide except US and
2
Geranium; few other species
Canada
4
Ginger
Asia
3,4
5
Mulberry
Management
The great variability of Ralstonia solanacearum and the strong influence of
environmental conditions on resistance make bacterial wilt a disease difficult to manage
(French, 1996).
Among the practices recommended, the use of resistant cultivars is considered to
play an important role. It is proven useful to control the potato strain, and is potentially
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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the most effective and ideal way to manage the disease (Jyothi et al., 1993; French,
1996). However, despite its importance, little progress has been made by either nationals
or international potato breeding program in achieving stable resistance in commercial
genotypes (Martin and French, 1996; Lopes and Quezado-Soarez, 1994).
Resistance is not general, but pathogen specific; a pathogen at one location may
over come the resistance effective at another location. More than one pathogen may occur
in a given field. Existence of different pathogen within races may reduce effectively
resistance at certain location, acceptance level of resistance should be determined for
definite use. When use for consumption, a certain percentage of infection may be
tolerated. But if seed production, it is preferable not to tolerate any bacterial wilt, because
few infested seed tubers can spread the disease over a wide area. Since the expression of
resistance is pathovar, and environmental specific, an essential step in development of
resistant varieties is local screening (Martin and French, 1996).
Selection for Resistance to Bacterial Wilt
The first thing to consider when planting potato must be the variety. One must
choose a variety that is adaptable to the locality in order to achieve maximum production.
Using the right variety ensures high yield and better quality. Series of varietal evaluation
must be conducted to determine the performance of a new or previously untried variety
(HARRDEC, 1996).
Clonal evaluation and selection is important in the breeding program. The
standard procedure involves the selection of healthy looking and high yielding plants in
the field. Tubers of each selected clones are then harvested and kept separately to be
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planted in the next season. Plants are carefully inspected for any abnormalities, and if
found in the first generation (F1), their clones are rejected and removed right away from
the field. Hence, successful potato production begins with long-term labor and intensive
breeding (Beukema, 1985).
Kurupuaracchi (1995) conducted an on farm potato evaluation and found that not
all clones were as superior as those in the on station trail. Out of the 22 clones, only 2
clones exhibited comparable level of yield ability, adaptability and stability with their
popular local variety. Thus, as a role, new clones usually differ in cultural
characteristics; therefore, several series of evaluations must be made at different strategic
locations and seasons.
Barrozo (2001), among the twelve potato varieties screened against bacterial wilt
under green house condition revealed that all varieties were resistant as no above ground
symptoms were exhibited. However, belowground symptom showed that all cultivars
were infected.
The study conducted by Botangen (2003) at Bosleng, Paoay, Atok, Benguet shows
that all the seven clones evaluated differed in growth, pest and disease resistibility,
yield as well as processing qualities. It is important to evaluate at different locations
and seasons for further screening.
Simongo (1992) found that clone LBR 1-5 was the most prolific in apical cutting
and tuberlet production among the several clones evaluated. On the different on farm
location trials, the performance of the individual clone varies with clone LBR 1-9 being
the most promising with an average yield of 21.2t/ha and showed high resistance to late
blight, good eating and tuber qualities.
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Potato Breeding for Resistance to Bacterial wilt
The cultivated potato, Solanum tuberosum subsp. tuberosum (2n = 4x = 48), is
one of the most important world food crops and demands continued genetic improvement
to meet the needs of a changing world. Potato breeding has been a cumbersome task due
to inherent biological factors: cytoplasmic nuclear sterilities, tetrasomic inheritance and
inbreeding depression. In addition, yield stasis exists within the potato germplasm of
North America (Douches, 1996).
The first systematic potato-breeding program for resistance to bacterial wilt
started in 1967 by Rowe and Sequeira (1972) at the University of Wisconsin. These
investigators began their program by intercrossing several resistant diploid clones of
Solanum phureja from the Central Colombian Collection, with clones of S. tuberosum
ssp. tuberosum. Field tests of S. phureja x spp. tuberosum hybrids revealed susceptibility
of this genetic material to Phytophthora infestans. In a further set of crosses, the
bacterial wilt-resistant hybrids were therefore crossed with Mexican late blight-resistant
clones, combining the two needed resistances. The Mexican germplasm used in this
crossing program consisted of ssp. tuberosum clones containing late blight resistance
genes derived from the wild hexaploid species S. demissum (Schmiediche, 1986).
In 1969, the University of Wisconsin sent 369 clones, representing 10 families, to
Peru where they were to be tested under natural conditions in fields heavily infected with
P. solanacearum. French and Herrera started a screening program at Huambos in the
Department of Cajamarca, Peru (Herrera, 1972). By 1974 the Wisconsin material had
changed hands three times and only seven of the original 369 clones had survived. Most
of the material had been lost due to causes other than bacterial wilt.
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Apart from sending 369 clones of bacterial wilt resistant genetic material to Peru,
the University of Wisconsin sent similar sets of material to 20 countries around the
world. The success of the Wisconsin material is partly due to its high degree of
heterozygosity, which results from use of unreduced gametes in the tertaploid x diploid
crosses which were necessary components of this breeding program. The bacterial wilt
resistance of this material comes from the diploid species S. phureja. Expression of this
resistance at the tertaploid level becomes even more remarkable when the genomic
composition of the tertaploid BR clones is considered. Only one genome, disregarding
crossing over during meiosis, is S. phureja origin. Crossing BR clones with other
tertaploid clones that are not resistant to bacterial wilt will obviously dilute the S.
phureja-based resistance even more (Schmiediche, 1986).
The wild potato relative Solanum commersonii is reported to have good high-
temperature resistance to brown rot of potatoes. However, S. tuberosum and S.
commersonii have different ploidy and endosperm balance numbers and are therefore
sexually incompatible, so their cells were fused by somatic hybridization. The resulting
somatic hybrid plants were vigorous and potato-like in appearance, but their resistance
level was unknown. Six of the somatic hybrids were examined, the S. commersonii and S.
tuberosum parents, and cv. Atlantic with a virulent strain of R. solanacearum (race 3,
biovar 2) at 28 ºC. The result reveals that S. commersonii was significantly more
resistant to brown rot than the cultivated potatoes. Encouragingly, preliminary results
show no significant difference in disease resistance between the somatic hybrids and the
S. commersonii parent. The somatic hybrids are both self-compatible and fertile to S.
tuberosum back-crosses (Laferriere et al., 1997).
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CIP Breeding Program
In 1977, data received from the National Potato Programs in CIP’s regions had
suggested that resistance based on S. phureja was effective against strains of P.
solanacearum that affect potatoes, as long as the resistant material was grown under
temperate climatic conditions. Such conditions can be found in tropical highland regions
or in the plains of northern India during the winter.
Until 1980 the only source of resistance to bacterial wilt had been S. phureja,
which had demonstrated its potential as well as its limitations. The narrow genetic base
of this resistance had become an object of concern for breeders, pathologists and
geneticists. At the end of 1980, a new clone (AVRDC 1287.19) from Taiwan developed
at the Asian Vegetable Research Development Centre (AVRDC) was found out to be
resistant to bacterial wilt and has heat adaptation. The source of resistance was, however,
not S. phureja but the two wild species S. raphanifolium and S. chacoense. The latter
was used in hesitation since its resistance, so effective under Taiwanese conditions, had
broken down under field conditions in Peru, where a strain of P. solanacearum different
from the Taiwan strain was present. However, the chance to introduce a new source of
resistance to bacterial wilt into the existing gene pool, and combining two sources of
resistance into one progeny, overrode doubts about the usefulness of this clone in CIP’s
breeding program (Schmiediche, 1986).
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MATERIALS AND METHODS
A. Evaluation using potato seed tubers
Germplasm
Materials
Healthy
G2 tubers of ten different potato clones obtained from the
International Potato Center (CIP) and JICA were used in the study. The evaluation was
conducted under green house condition. Treatments were arranged following the
Complete Randomized Design (CRD) with 4 replications.
The clones and cultivars evaluated and their characteristics are as follows:
Clone Cultivar’s
Origin
Type of resistance
Name
LB Wart
BW
LBR-5 Igorota Philippines R
387410.7 LBr-9
CIP
R
Warishiro
Japan
575003 I-931
India
R S S
676070 Cruza
155
Mexico R
380579.3 BW-III
CIP
MR S
720045 Atzimba Mexico MR
S S
573275 ASN-69-1
Mexico R R S
285378.27
None
CIP
MR MR
720071 Monserrate Columbia
MR
R S
The bacterial wilt nursery was thoroughly prepared. The area was divided into 4
blocks, each block consisting of 10 plots measuring 0.47 x 2meters each. Seven hundred
grams of triple 14 and 12.5 kilograms of chicken dung were applied to each block before
planting. During hilling–up, 700 grams of triple 14 was applied in each block. Dithane
and Ridomil were used to protect the plant from fungal diseases.
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Inoculum Preparation and Soil Inoculation
Four kilograms of infected potato tubers were cut into small cubes and were
incorporated in the soil. Another 8 kilograms of infected tubers were cut into smaller
pieces, allowed to ooze in 10 gallons of tap water and applied as soil drench.
To enhance the rapid multiplication of the bacteria, the inoculated soil was
irrigated periodically and covered with plastic sheets to increase the soil temperature.
Data Gathered
1. Percentage Bacterial Wilt Infection. This was obtained using the formula:
%
infection
=
no.
of infected plant X 100
total no. of plants
2. Weight of marketable tubers (kg). These are the tubers free from disease
infection.
3. Weight of non-marketable tuber (kg). Marble size, rotten and diseased tubers
was considered as non-marketable.
4. Total weight of tubers infected with bacterial wilt (kg). This was determined
by weighing all the tubers infected with bacterial wilt.
5. Late blight infection. This was asses using the CIP rating scale:
Blight (%) CIP
Scale Description
0
1
No blight to be seen
0.1
1
Very few plants in larger plots with a
lesion, not more than two lesions per
10 m of row (30 plants)
1
2
Up to 10 small lesions
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3
3
Up to 30 small lesion per plant or up
to one leaflet in each 20 attacked
10
4
Most plants are visibly attacked, and
one out of three leaflets infected, few
multiple infections per leaflet.
25
5
Nearly every leaflet with lesion.
Multiple infections per leaflet are
common. Field or plants in plot are
affected.
50
6
Every plant effected and half the leaf
area destroyed by blight. Plots looks
green, fleaked and brown; blight is
very obvious.
75
7
As previous, but three quarters of
each plant affected by blight. Lower
branches maybe overwhelmingly
killed off and only green leaves, if
any, are at the top of the plant.
Shape of plants maybe more spindly
due to extensive foliar loss. Plot
looked neither brown nor green.
91
8
Some leaves and most stems are
green. Plot looks brown with some
green patches.
97
9
Few green leaves, most with lesions,
remain. Many stems with lesions.
Plots looks brown.
100
9
All leaves and stems dead. No
visible
blight
left
to
evaluate.
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B. Evaluation using Stem cuttings
Two weeks old rooted stem cuttings from NPRCRTC were used in the study.
Treatments were arranged following the Complete Randomized Design (CRD) with 3
replications at three plants per replicate.
The stem cuttings were planted in plastic cups with sterilized soil and maintained
at the Department of Plant Pathology Green house.
The entries were as follows:
Clone Cultivar’s
Name
LBR-5 Igorota
575003 I-931
676070 Cruza
155
573275 ASN-69-1
285378.27 -
Preparation of Inoculum and Inoculation Method
First Trial
Inoculum of bacterial wilt was sourced from infected tubers. The tubers were
properly washed, cut and squeezed for the bacterial ooze to come out and was diluted in
10 ml distilled water. Using a sterilized wire loop, the bacterial suspension was streaked
in previously plated casamino-peptone glucose agar (CPGA) consisting of the following
in g/li of water:
10 g dextrose
1 g casamino acid
10 g peptone
18 g agar
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Surface growth of the bacterium was scraped and diluted in the desired volume of
distilled water. The soil was cultivated near the root system to create wounds before
drenching 30 ml of the bacterial suspension in each plant.
After 3 days, only 2 among the 45 stem cuttings wilted.
Second
Trial
In the second trial, the same germplasm materials were used but the inoculation
method was modified. Before planting, roots hairs of the stem cuttings were cut and were
dipped in the bacterial suspension for 3 minutes to allow entry of the bacteria in the plant
system.
In addition, infected potato tubers were cut into smaller pieces and allowed to
ooze out in the desired volume of distilled water. The soil was cultivated near the root
system to create wounds before drenching 60 ml of the bacterial suspension in each of the
plant 16 days after the first inoculation.
Bacterial population was determined using the spectrophotometer.
Data Gathered
1. Bacterial population. This was obtained by using the spectrophotometer.
2. Number of days from planting to bacterial wilt expression. Days were counted
from inoculation to symptom expression.
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RESULTS AND DISCUSSION
Evaluation Using Potato Seed Tubers
Percentage Bacterial Wilt Infection
Twenty three (23) days after planting (DAP), the variety Igorota and clone
387410.7 had the highest percentage of bacterial wilt infection with a mean of 27.50, and
at 44 DAP, 100% plant were dead due to bacterial wilt (Table 1). On the other hand,
clones 720071, 285378.27, 380579.3 and Warishiro showed a slow rate of bacterial wilt
infection from 23 to 58 DAP, (Plates 1 and 2).
Among the 10 entries, only the clone 720071 still had plant that is not infected
with bacterial wilt after 58 DAP.
The high incidence of bacterial wilt may have been influenced by the prevailing
weather conditions. Under greenhouse condition, the temperature was relatively higher
than the ambient temperature. The maximum and minimum temperatures during the last
week of September (BW rating started) were 26ºC and 17ºC respectively, compared to
the ambient condition with a maximum temperature of 24.5ºC and a minimum of 16.8ºC
(Table 2). The prevailing weather condition during the conduct of the study favors R.
solanacearum development as cited by Persley (1985) that the optimum temperature for
R. solanacearum development is from 25-35ºC. Moreover, Martin and French (1996)
cited that disease development of bacterial wilt is mainly influenced by temperature.
High temperature promoted bacterial wilt development.
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Table 1. Mean percentage bacterial wilt infection of different clones/variety evaluated
CLONE/
DAYS
AFTER
PLANTING
VARIETY
23 30 37 44 51 58
Igorota 27.50
60.00a
80.00ab
100a 100a
100
387410.7
27.50
47.50ab
77.50a 100a 100a
100
Warishiro
2.50
15.00c
27.50bcd
57.50cde 90.00a
100
575003
15.00
30.00bc
52.50abc
90.00ab 100a
100
676070
10.00
30.00bc
50.00abc
77.50abc 97.50a 100
380579.3
5.00
12.50c
27.50cd
52.50cde 85.00ab 100
720045
7.50
22.50bc
37.50bcd
70.00bcd 95.00a 100
73275
7.50
22.50bc
47.50abc
75.00abc 92.50a
100
285378.27 7.50
7.50c
17.50cd
45.00d 72.50bc 100
720071
2.50
5.00c
12.50d
35.00e 60.00c 87.50
Means followed by a common letter are not significantly different at 5% level by DMRT
Plate 1. Clone 380579.3 showing wilt symptom at 30 DAP
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Plate 2. Infected tubers from clone 285378.27
Plate 3. Infected tubers from clone 676070
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Table 2. Mean weekly weather data
TEMPERATURE
AVERAGE
TOTAL
MONTH
ºC
RAINFALL
RELATIVE BRIGHT
Max Min
(mms and
HUMIDITY SUNSHINE
Tenths)
(%)
(mm)
______________________________________________________________________
September
Week 1
24.1 17.4
24.6
90
193.7
Week 2
25.1 17.7
7.2
89
169.7
Week 3
22.2 16.7
57.5
90
53.1
Week 4
24.5 16.8
2.3
87
293.1
October
Week 1
24.4 16
20
89
333.4
Week 2
24
15.6
10.4
86
252.8
Week 3
25.3 16
1.0
87
363.4
Week 4
24.9 16.2
3.0
84
287.1
November
Week 1
24
16.2
0.9
82
256.8
Week 2
25.6 15.4
4.2
82
321.1
Week 3
24.4 14.4
1.3
83
341.5
Week 4
25.6 15.3
0.0
77
322.2
February
Week 1
23.6 14.2
0.0
79
-
Week 2
25.4 14.3
2.3
80
-
Week 3
26
13.2
0.0
80
-
Week 4
26.9 12.9
0.0
74
-
Percentage Late Blight Infection
Among the 10 entries, clone 720071 showed a slow development rate of late
blight infection and was the only clone that reached 58 DAP (Table 3). Conversely,
clones 676070, 720045, 573275, 285378.27 and Warishiro, reached 51 DAP and had a
late blight rating ranging from 1 and 2, where no blight was seen; or very few plants in
larger plots with lesions and 2 with a maximum of 10 small lesions. On the other hand,
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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clone 387410.7 and variety Igorota only reached 37 DAP and had late blight ratings of 2.
The variety Igorota is rated as resistant to late blight. However the variety exhibited late
blight infection early in its growing period but is able to regenerate new leaves and
matures at 100-120 days.
Most of the entries are rated to be resistant to late blight but their resistance was
not quantified due to the early infection of bacterial wilt which caused their early
senescence.
The fluctuation of the prevailing weather condition played a great factor for the
manifestation of resistance or susceptibility of the clones/variety to late blight infection.
Thung (1974) explained that any environmental fluctuation may upset the pathogen and
shift host reaction to low and higher resistance or susceptibility.
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Table 3. Mean percentage late blight infection of different clones/variety evaluated
CLONE/
DAYS
AFTER
PLANTING
VARIETY
30
37
44
51
58
________________________________________________________________________
Igorota
1.25bc
2.0
387410.7
2.00a
2.25
Warishiro
1.50b
1.75
3.0ab
2.0
575003
1.0c
1.25
2.0bcd
676070
1.0c
1.0
1.5d
1.5
380579.3
1.0c
1.25
2.75abc
2.75
720045
1.0c
1.50
2.50abcd 2.0
573575
1.0c
1.25
1.75cd
2.0
285378.27
1.0c
1.50
1.75cd
1.5
720071
1.0c
1.00
2.0bcd
1.75
1.5
Means followed by a common letter are not significantly different at 5% level by DMRT
Yield Parameters
Results reveal that clone 676070 had the highest marketable yield with a mean of
575g but also had the highest non marketable and bacterial wilt infected tubers (Table 4).
On the other hand, the least weight of marketable tubers was obtained in clone 720071
with a mean of 36.25g which also registered the lowest in non marketable tubers and
second lowest in bacterial wilt infected tubers with a mean of 21.25g.
The variety Igorota which is a late maturing variety registered one of the lowest
marketable tubers with a mean of 58.75g. Its low marketable yield is associated with its
early senescence due to bacterial wilt infection.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
23
All the 10 entries evaluated were able to produce marketable tubers despite the
early bacterial wilt infection making them all tolerant to bacterial wilt.
Table 4. Yield parameters of different clones/variety evaluated
CLONE/ MARKET
ABLE
(g)
NON
BACTERIAL
WILT
VARIETY
MARKET
ABLE
(g)
INFECTED
(g)
Igorota
58.75
66.25
41.25
387410.7
48.75
55.00
3.75
Warishiro
72.50
52.50
8.75
575003
95.00
126.25
51.25
676070
575.00
250.00
388.75
380579.3
218.75
167.50
72.50
720045
82.50
113.75
25.00
573275
271.25
183.75
111.25
285378.27
161.25
193.75
193.75
720071
36.25
20.00
21.25
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Plate 4. Marketable, non-marketable, BW infected tubers of variety Igorota
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
25
Evaluation Using Stem Cuttings
Number of days from planting to bacterial wilt expression
Table 5 shows the number of days from planting to bacterial wilt expression.
Clone 285378.27 manifested bacterial wilt symptom at the shortest time having a mean of
24.210 days. The clone 575003 on the other hand exhibited the longest time of symptom
expression at a mean of 25.20 days.
The recorded temperature in the greenhouse during the month of February was
relatively higher than the ambient temperature. It was during the 3rd week of February
that wilting was observed in the sample plants. The maximum temperature in the green
house was 27ºC and minimum of 14ºC, compared to the ambient condition with a
maximum of 26ºC and a minimum of 13.2ºC (Table 2). Persley (1985) cited that the
optimum development of R. solanacearum is from 25-35ºC.
Table 5. Average number of days from transplanting to bacterial wilt symptom
expression
CLONES/
MEAN
NUMBER
OF
DAYS
CULTIVAR AFTER TRANSPLANTING
________________________________________________________________________
Igorota
24.887
575003 25.200
676070 24.663
573275 24.773
285378.27
24.210
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Table 6 presents the bacterial population that was used during inoculation in the
first and second trial. Results revealed that there is a significant difference from the
bacterial population during the first and second trial.
During the termination of the study, not all the sample plants wilted. The
assymptomatic cuttings were tested using the water blank test which revealed that they
are infected with bacterial wilt (Plates 4 and 5).
The unsuccessful result obtained from the first trial may be due to the low
bacterial population that was inoculated to the sample cuttings. Nielson and Hyness
(1979), as cited by Nagpala (1986) reported that the possible variation of resistance or
susceptibility of crops were affected by the age of the plant at the time of inoculation. It
was reported that the best time to inoculate the plants should be about 4 weeks after
planting.
Table 6. Bacterial population of inoculum used during the trials
SAMPLE
AVERAGE BACTERIAL
POPULATION
TRIAL 1
TRIAL 2
DIFFERENCE
________________________________________________________________________
1
0.162
0.958
0.796
2
0.130
0.997
0.867
3
0.133
1.000
0.867
________________________________________________________________________
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
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Plate 5. Wilted potato cuttings at 16 days after inoculation
Plate 6. Assymptomatic cuttings being tested for BW infection
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
28
SUMMARY, CONCLUSION AND RECOMMENDATION
Summary
Nine clones: 387410.7, Warishiro, 575003, 676070, 380579.3, 720045, 573275,
285378.27 and 720071 and the variety Igorota were evaluated to identify germplasm
materials that were tolerant to bacterial wilt infection under green house condition and to
determine their reaction to late blight infection.
Among the 10 entries evaluated, Igorota and clone 387410.7 showed the highest
percentage bacterial wilt infection at 23 DAP. On the other hand, clone 720071 exhibited
the slowest rate of bacterial wilt infection followed by clones 285378.27, 380579.3 and
Warishiro respectively.
Clones 720071, 285378.27 and 676070 showed resistance to late blight infection
with a rating of 1 or no blight or a maximum of 10 small lesions observed in the sample
plant.
The highest marketable yield was obtained from clone 676070 which ironically
registered the highest nonmarketable and bacterial wilt infected tubers.
Conclusion
Based on the results of the study, all the 10 entries evaluated were tolerant to
bacterial wilt as they were able to produce marketable tubers despite bacterial wilt
infection. Clones 720071, 285378.27 and 676070 confirmed their resistance to late blight
infection.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
29
Recommendation
Further evaluation must be conducted to assess the adaptability and yield of the
germplasm materials.
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
30
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Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
34
APPENDICES
Appendix Table 1. Percentage bacterial wilt infection at 23 DAP
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
10 10
70
20
110
27.50
387410.7
50
20
20 20
110
27.50
Warishiro
10
0 0
0
10
2.50
575003
20 20
20
0 60
15
676070
10
0
10
20 40
10
380579.3
10
0
10
0 20
5
720045
0
20
10
0 30
7.5
573275
10
20
0
0 30
7.5
285378.27
10
10 0
20 30
7.5
720071
0
0 10
0 10
2.5
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Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
3.24 3.24
8.39 4.53 19.40
4.850
387410.7
7.11 4.53
4.53 4.53 20.70
5.175
Warishiro
3.24 0.71
0.71 0.71 5.37
1.343
575003
4.53 4.53
4.53 0.71 14.30
3.575
676070
3.24 0.71
3.24 4.53 11.75
2.930
380579.3
3.24 0.71
3.24 0.71 7.90
1.975
720045
0.71 4.53
3.24 0.71 9.19
2.210
573275
3.24 4.53
0.71 0.71 9.19
2.210
285378.27
3.24 0.71
0.71 4.53 9.19
2.210
720071
0.71 0.71
3.24 0.71 5.37
1.343
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
327.478 109.159
0.6702
Factor A
9
3112.413 345.824
2.1233ns 2.25 3.14
Error
27
4397.498 162.870
________________________________________________________________________
TOTAL
39
7,837.388
ns= not significant
Coefficient of Variation = 62.05%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Appendix Table 2. Percentage bacterial wilt infection at 30 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 60 40 100 40 240 60
387410.7
60 40 50 40 190
47.5
Warishiro
40 0 20 0 60 15
575003 40 40 40 0 120 30
676070 30 30 10 50 120 30
380579.3 20 10 20 0 50 12.5
720045 10 40 40 0 90 22.5
573275 30 40 0 20 90
22.5
285378.27 0
0
0
30 30 7.5
720071 0 0 20 0 20 5
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
7.78 6.36
10.02 6.36 30.52
7.630
387410.7
7.78 6.36
7.11 6.36 27.61
6.903
Warishiro
6.36 0.71
4.53 0.71 12.31
3.078
575003
6.36 6.36
6.36 0.71 19.79
4.948
676070
5.52 5.52
3.24 7.11 21.39
5.348
380579.3
4.53 3.24
4.53 0.71 13.01
3.253
720045
3.24 6.36
6.36 0.71 16.67
4.168
573275
5.52 6.36
0.71 4.53 17.12
4.280
285378.27
0.71 0.71
0.71 5.32 7.45
1.863
720071
0.71 0.71
4.53 0.71 6.66
1.665
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
907.490 302.497
0.9879
Factor A
9
11022.418 1224.713 3.997**
2.54 3.14
Error
27
8267.390 306.200
________________________________________________________________________
TOTAL
39
20,197.298
**= Highly Significant
Coefficient of Variation = 69.30%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Appendix Table 3. Percentage bacterial wilt infection at 37 DAP
BLOCK
CLONE/
VARIETY
I II III IV
TOTAL
MEAN
Igorota 70 80 100 70 320 80
387410.7
80 60 90 80 310
77.5
Warishiro 50
0
40 20 110 27.5
575003 60 50 70 30 210
52.5
676070 40 60 40 60 200 50
380579.3 40 40 30 0 110 27.5
720045 20 70 60 0 150 37.5
573275 50 60 30 50 190
47.5
285378.27
20 20 0 30 70
17.5
720071 10 0 30 10 50 12.5
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
8.39 8.97
10.2 8.39 35.77
8.943
387410.7
8.97 7.78
9.51 8.97 35.23
8.808
Warishiro
7.11 0.71
6.36 4.53 18.71
4.678
575003
7.78 7.11
4.13 5.52 24.54
6.135
676070
6.36 7.78
6.36 7.78 28.28
7.070
380579.3
3.36 6.36
5.52 0.71 18.95
4.738
720045
4.53 8.39
7.78 0.71 21.41
5.353
573275
7.11 7.78
5.52 7.11 27.52
6.880
285378.27
4.53 4.53
0.71 5.52 15.29
3.823
720071
3.24 0.71
5.52 3.24 12.71
3.178
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
2507.50 835.833
1.8547
Factor A
9
15162.50 1684.722 3.7384** 2.25 3.14
Error
27
1216.50 450.648
________________________________________________________________________
TOTAL
39
29,837.50
**= Highly Significant Coefficient of Variation = 37.26%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Appendix Table 4. Percentage bacterial wilt infection at 44 DAP
BLOCK
CLONE/
VARIETY
I II III IV
TOTAL
MEAN
Igorota 100 100 100 100 400 100
387410.7
100 100 100 100 400 100
Warishiro
70 40 60 60 230
57.5
575003 100 100 90 70 360 90
676070 60 100 70 80 310 77.5
380579.3
70 50 50 40 210
52.5
720045 40 100 100 40 280 70
573275 70 100 50 80 300 75
285378.27
40 40 40 60 180 45
720071 40 20 50 30 140 35
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
10.02 10.02
10.2 10.02 40.08 10.02
387410.7
10.02 10.02
10.02 10.02 40.08 10.02
Warishiro
8.39 6.36
7.78 7.78 30.31 7.578
575003
10.02 10.02
9.51 8.39 37.94 9.485
676070
7.78 10.02
8.39 8.97 35.16 8.790
380579.3
8.39 7.11
7.11 6.36 28.97 7.243
720045
6.36 10.02
10.02 6.36 32.76 8.190
573275
8.39 10.02
7.11 8.97 34.49 8.623
285378.27
6.36 6.36
6.36 7.78 26.86 6.715
720071
6.36 4.53
7.11 5.52 23.52 5.880
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
427.50 142.50
0.4998
Factor A
9
18372.50 2041.38
7.1604** 2.54 3.14
Error
27
7697.50 285.093
________________________________________________________________________
TOTAL
39
26,497.50
**=Highly Significant
Coefficient of Variation = 24.04%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
42
Appendix Table 5. Percentage bacterial wilt infection at 51 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 100 100 100 100 400 100
387410.7
100 100 100 100 400 100
Warishiro 90
70 100 100 360 90
575003 100 100 100 100 400 100
676070 100 100 90 100 390 97.5
380579.3 100 100 70
70 340 85
720045 100 100 100 80 380 95
573275 100 100 70 100 370 92.5
285378.27
70 80 60 80 290
72.5
720071 60 50 60 70 240 60
Germplasm Evaluation for Tolerance Against Bacterial Wilt
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Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
10.02 10.02
10.02 10.02 40.08 10.02
387410.7
10.02 10.02
10.02 10.02 40.08 10.02
Warishiro
9.51 8.39
10.02 10.02 37.94 9.485
575003
10.02 10.02
10.02 10.02 40.08 10.02
676070
10.02 10.02
10.02 10.02 39.57 9.893
380579.3
10.02 10.02
9.51 8.39 36.82 9.205
720045
10.02 10.02
10.02 8.97 39.03 9.758
573275
10.02 10.02
8.39 10.02 38.45 9.613
285378.27
8.39 8.97
7.78 8.97 34.11 8.528
720071
7.78 7.11
7.78 8.39 31.06 7.765
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
267.50 89.167
0.8731
Factor A
9
6452.50 716.944
7.0199**
2.54 3.14
Error
27
2757.50 102.130
________________________________________________________________________
TOTAL
39
24,177.50
**=Highly Significant
Coefficient of Variation = 11.32%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
44
Appendix Table 6. Percentage bacterial wilt infection at 58 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 100 100 100 100 400 100
387410.7
100 100 100 100 400 100
Warishiro
100 100 100 100 400 100
575003 100 100 100 100 400 100
676070 100 100 100 100 400 100
380579.3
100 100 100 100 400 100
720045 100 100 100 100 400 100
573275 100 100 100 100 400 100
285378.27
100 100 100 100 400 100
720071 90 80 80 100 350 87.5
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
45
Transformed data
CLONE/
BLOCK
VARIETY
I
II
III
IV TOTAL MEAN
Igorota
10.02 10.02
10.02 10.02 40.08 10.02
387410.7
10.02 10.02
10.02 10.02 40.08 10.02
Warishiro
10.02 10.02
10.02 10.02 40.08 10.02
575003
10.02 10.02
10.02 10.02 40.08 10.02
676070
10.02 10.02
10.02 10.02 40.08 10.02
380579.3
10.02 10.02
10.02 10.02 40.08 10.02
720045
10.02 10.02
10.02 10.02 40.08 10.02
573275
10.02 10.02
10.02 10.02 40.08 10.02
285378.27
10.02 10.02
10.02 10.02 40.08 10.02
720071
9.51 8.97
8.97 10.02 37.47 9.368
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
27.50 9.167
Factor A
9
562.50 62.50
6.8182**
2.54 3.14
Error
27
247.50 9.167
________________________________________________________________________
TOTAL
39
837.50
**=Highly Significant
Coefficient of Variation = 3.07%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
46
Appendix Table 7. Late blight infection rating at 30 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 1 1 2 1 5
1.25
387410.7
2 2 2 2 8 2.0
Warishiro
1 2 1 2 6 1.5
575003 1 1 1 1 4 1
676070 1 1 1 1 4 1
380579.3
1 1 1 1 4 1
720045 1 1 1 1 4 1
573275 1 1 1 1 4 1
285378.27
1 1 1 1 4 1
720071 1 1 1 1 4 1
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
0.075 0.025
0.4030
Factor A
9
4.025
0.447
7.2090**
2.54 3.14
Error
27
1.675
0.062
________________________________________________________________________
TOTAL
39
5.775
**=Highly Significant
Coefficient of Variation = 21.20%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
47
Appendix Table 8. Late blight infection rating at 37 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 2 2 2 2 8 2
387410.7
2 2 3 2 9
2.25
Warishiro
2 2 1 2 7
1.75
575003 2 1 1 1 5
1.25
676070 1 1 1 1 4 1.0
380579.3
1 2 1 1 5
1.25
720045 2 1 1 2 6
1.50
573275 1 1 1 2 5
1.25
285378.27
1 2 1 2 6
1.50
720071 1 1 1 1 4 1.0
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
0.475 0.158
0.8104
Factor A
9
6.225
0.692
3.5403**
2.54 3.14
Error
27
5.275
0.195
________________________________________________________________________
TOTAL
39
11.975
**=Highly Significant
Coefficient of Variation = 29.97%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
48
Appendix Table 9. Late blight infection rating at 44 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 3 3 3 2 11
2.75
387410.7
4 3 4 3 14
3.5
Warishiro
4 2 3 3 12
3.0
575003 3 1 3 1 8 2.0
676070 1 2 2 1 6 1.5
380579.3
3 3 3 2 11
2.75
720045 3 2 3 2 10
2.5
573275 2 2 1 2 7
1.75
285378.27
1 3 1 2 7
1.75
720071 2 2 2 2 8 2.0
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
2.10 0.70
1.5882
Factor A
9
15.10
1.678
3.8067** 2.54 3.14
Error
27
11.90
0.441
________________________________________________________________________
TOTAL
39
29.10
**=Highly Significant
Coefficient of Variation = 28.25%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
49
Appendix Table 10. Late blight infection rating at 51 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 2 3 1 2 8 2
387410.7
4 3 4 3 14
3.5
Warishiro
2 2 2 2 8 2.0
575003 2 2 3 1 8 2.0
676070 1 2 2 1 6 1.5
380579.3
3 3 3 2 11
2.75
720045 2 2 2 2 8 2.0
573275 2 2 2 2 8 2.0
285378.27
1 2 1 2 6 1.5
720071 1 2 2 2 7
1.75
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
1.0 0.33
1.20
Factor A
9
13.10
1.456
5.24** 2.25 3.14
Error
27
7.500
0.278
________________________________________________________________________
TOTAL
39
21.60
**=Highly Significant
Coefficient of Variation = 25.10%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
50
Appendix Table 11. Late blight infection rating at 58 DAP
CLONE/
BLOCK
VARIETY
I II III IV
TOTAL
MEAN
Igorota 2 1 1 1 5
1.25
387410.7
3 2 2 2 9
2.25
Warishiro
2 1 1 1 5
1.25
575003 2 2 1 1 6
1.50
676070 1 2 1 1 5
1.25
380579.3
2 2 1 1 6
1.50
720045 2 1 1 2 6
1.50
573275 1 1 1 2 5
1.25
285378.27
1 2 2 2 6
1.50
720071 1 2 2 1 6
1.50
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
3
1.475 0.492 1.8247
Factor A
9
3.225 0.358
1.3299ns 2.54 3.14
Error
27
7.275 0.269
________________________________________________________________________
TOTAL
39
11.975
ns=Not Significant
Coefficient of Variation = 35.19%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
51
Appendix Table 12. Weight of marketable tubers (g)
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota 55 180 0 0 235
58.75
387410.7 160
0
0
35 195 48.75
Warishiro 245
0
0
45
290 72.5
575003 225 0 0 155 380 95
676070 600 665 625 410 2300 575
380579.3
250 400 190 35 875
218.75
720045 125 75 90 40 330
82.5
573275 230 260 240 355 1085
271.5
285378.27
0 245 200 200 645
161.25
720071 0 145 0 0 145
63.25
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
52
Transformed data
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota 7.45 13.44 0.71 0.71 22.31 5.578
387410.7
12.67 0.71 0.71 5.96 20.05 5.013
Warishiro
15.67 0.71 0.71 6.75 23.84 5.960
575003 15.02 0.71 0.71 12.47 28.91 7.228
676070 24.51 25.80 25.01 20.26 95.58 23.895
380579.3 15.83 20.01 13.80 5.96 55.6 13.900
720045 11.20 8.69 9.51 6.36 35.76 8.940
573275 15.18 16.14 15.51 18.85 65.68 16.420
285378.27
0.71 15.67 14.16 14.16 44.7 11.175
720071 0.71 12.06 0.71 0.71 14.19 3.548
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Replication
3
94.424 31.475 1.0297
Factor A
9
1447.218 160.802 5.2609**
2.54 3.14
Error
27
825.274 30.566
________________________________________________________________________
TOTAL
39
2366.915
**=Highly Significant
Coefficient of Variation = 54.39%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
53
Appendix Table 13. Weight of non- marketable tubers (g)
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota 60 105 5 95 265
66.25
387410.7
140 55 25 0 220 55
Warishiro
100 45 15 50 210
52.5
575003 100 165 110 130 505
126.25
676070 200 220 80 500
10000
250
380579.3
300 115 145 110 670 167.5
720045 150 75 35 195 455
113.75
573275 165 200 80 290 735
183.75
285378.27
370 130 120 155 775 193.5
720071 0 50 10 20 80 20
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
54
Transformed data
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota 7.78 10.27 2.35 9.77 30.17 7.542
387410.7
11.85 7.45 5.05 0.71 25.06 6.265
Warishiro
10.02 6.75 3.94 7.11 27.81 6.953
575003 10.02 12.86 10.52 11.42 44.81 11.205
676070 14.16 14.85 8.97 22.37 60.35 15.088
380579.3 17.33 10.75 12.06 10.51 50.65 12.663
720045 12.27 8.69 5.96 13.98 40.9 10225
573275 12.86 14.16 8.97 17.04 53.03 13.258
285378.27
19.25 11.42 10.98 12.47 54.12 13.530
720071 0.71 7.11 3.24 4.53 15.59 3.898
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Replication
3
116.104 38.701 3.8435
Factor A
9
496.030 55.114 5.4735**
2.54 3.14
Error
27
271.870 10.069
________________________________________________________________________
TOTAL
39
884.004
**=Highly Significant
Coefficient of Variation = 31.53%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
55
Appendix Table 14. Weight of Infected tubers (g)
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota
110 45 0 10 165
41.25
387410.7
15 0 0 0 15
3.75
Warishiro 0
0
0
35 35 8.75
575003 80 40 75 10 205
51.25
676070 150 350 760 295 1555
388.75
380579.3
50 180 15 45 290 72.5
720045 30 20 40 10 100 25
573275 115 5 255 70 445
11.25
285378.27
195 205 65 310 775
193.75
720071 35 50 0 0 85
21.25
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
56
Transformed data
CLONE/
BLOCK
VARIETY
I II III
TOTAL
IV
MEAN
Igorota 10.51 6.75 0.71 3.24 21.21 5.303
387410.7
3.94 0.71 0.71 0.71 6.07 1.518
Warishiro
0.71 0.71 0.71 5.96 8.09 2.023
575003 8.97 6.36 8.69 3.24 27.26 6.815
676070 12.27 18.72 27.58 17.19 75.76 18.94
380579.3 7.11 13.44 3.94 6.75 31.24 7.810
720045 5.52 4.53 6.36 3.24 19.65 4.913
573275 10.75 2.35 15.98 8.40 37.48 9.370
285378.27
13.98 14.34 8.09 17.62 54.03 13.508
720071 5.96 7.11 0.71 0.71 14.49 3.623
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Replication
3
8.204 2.735
0.1642
Factor A
9
1052.941 116.993 7.0271** 2.54 3.14
Error
27
449.579 16.649
________________________________________________________________________
TOTAL
39
15110.665
**=Highly Significant
Coefficient of Variation = 55.27%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
57
Appendix Table 15. Number of days from transplanting to bacterial wilt symptom
expression
CLONE/
REPLICATE
VARIETY
I II III
TOTAL
MEAN
1 23.33 26.00 26.00 74.66 24.887
2 24.66 26.00 26.00 75.60 25.200
3 23.66 24.66 26.00 73.99 24.663
4 23.30 25.33 26.00 74.32 24.773
5 23.33 23.66 23.3 72.63 24.210
ANALYSIS OF VARIANCE
SOURCES OF DEGREES OF SUM OF MEAN OF COMPUTED TABULAR F
VARIATION FREEDOM SQUARES SQUARES F
0.05 0.01
Block
2
9.401 4.701 3.9895
Factor A
4
7.562 0.391
0.3315ns 3.84 7.01
Error
18
9.426 1.178
________________________________________________________________________
TOTAL
14
20.390
ns= Not Significant
Coefficient of Variation = 4.39%
Germplasm Evaluation for Tolerance Against Bacterial Wilt
(Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.)/ Jamie Beth Basngi. 2006
Document Outline
- Germplasm Evaluation for Tolerance
Against Bacterial Wilt (Ralstonia solanacearum (E.F. Smith) Yabuuchi et al.).
- BIBLIOGRAPHY
- ABSTRACT
- TABLE OF CONTENTS
- INTRODUCTION
- Nature of the Study
- REVIEW OF LITERATURE
- Causal Organism
- EPIDEMIOLOGY
- Host Range and Distribution
- Management
- Selection for Resistance to Bacterial Wilt
- Potato Breeding for Resistance to Bacterial wilt
- CIP Breeding Program
- MATERIALS AND METHODS
- Evaluation using potato seed tubers
- Inoculum Preparation and Soil Inoculation
- Data Gathered
- Evaluation using Stem cuttings
- Preparation of Inoculum and Inoculation Method
- Data Gathered
- RESULTS AND DISCUSSION
- Percentage Bacterial Wilt Infection
- Percentage Late Blight Infection
- Yield Parameters
- Number of days from planting to bacterial wilt expression
- SUMMARY, CONCLUSION AND RECOMMENDATION
- Summary
- Conclusion
- Recommendation
- LITERATURE CITED
- APPENDICES