BIBLIOGRAPHY
BAWAYAN, JENNIE T. OCTOBER, 2008. Pathogenicity Test of Fusarium
oxysporum f. sp. fragariae. Benguet State University, La Trinidad, Benguet.
Adviser: Asuncion L. Nagpala, PhD.
ABSTRACT
Pathogenicity of Fusarium oxysporum isolated from strawberry obtained from
Balili, Pomology, and Swamp was inoculated to chrysanthemum, tomato and the source
plant to evaluate the ability of the isolates to cause root rot and discoloration of crowns
on all the test plants.
At spore concentrations of 6.46x106/ml (Balili isolate), 8.53x106/ml (Pomology
isolate) and 1.60x106/ml (Swamp isolate), root rotting symptom and crown discoloration
was observed in strawberry plants dipped in the fungal suspension for one hour before
planting after two to four weeks. Chrysanthemum and tomato dipped at the same time
and the same amount of spore suspension did not develop root rotting or crown
discoloration. Both plants did not also show any form of above ground symptoms after
two to four weeks.
Based on this result, Fusarium isolated from strawberry is Fusarium oxysporum f
sp. Fragariae which infected strawberry only.

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TABLE OF CONTENTS

Page
Bibliography. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
i
Abstract. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
i
Table of Contents. . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
ii
INTRODUCTION . . . . . . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
1
REVIEW OF LITERATURE
The Crop. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
4
The Pathogen. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
4
General Characteristics of Fusarium oxysporum f. sp. fragariae . . . . . . . .
5
Disease Caused by Fusarium oxysporum f. sp. fragariae . . . . . . . . . . . . . .
6
Symptoms Caused by Fusarium oxysporum f. sp. fragariae . . . . . . . . . . .
7
Management of Fusarium oxysporum f. sp. fragariae. . . . . . . . . . . . . . . . .
8
MATERIALS AND METHODS
Isolation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
9
Microscopic Observation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
9
Standardization of Inoculum . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
9
Pot Bioassay . . . . . . . . . . . . . .. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
10
Assessment of Symptoms on Inoculated Plants
11
Data Gathered. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
11


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RESULTS AND DISCUSSION
Above Ground Symptoms . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
12
Below Ground Symptoms. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
14
No. of Dead Strawberry Plants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
15
Chrysanthemum Inoculated Plants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
16
Tomato Inoculated Plants . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
18
Fusarium oxysporum Isolates. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 19
SUMMARY, CONCLUSION AND RECOMMENDATION
Summary. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
21
Conclusion. . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
21
Recommendation . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
22
LITERATURE CITED . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
23
APPENDICES . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .
25

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1
INTRODUCTION

Strawberry, Fragaria x ananassa Duch originated in Europe, around 1750. It is a
hybrid between the pistillate South American F.chiloensis Duch. and North American F.
virginiana Duch. For many centuries strawberry is still a favorite fruit of the temperate
world. The fruit was valued for its delicious flavor and fragrance, and for health restoring
qualities. Today in the United States strawberry is being grown in home gardens and in
every state of the Union. The crop is also produced commercially in different climatic
zones of the country like Atlantic Coast, South Atlantic and Gulf, Florida, East Central
and Northeast, Great Plains and Rocky Mountains, North Pacific Coast or Pacific
Northwest and California (Wilhelm and Nelson, 1996).

In Benguet, there is about 35 percent of the farming populations engaged in
strawberry industry. According to the Municipal Agriculturist office (MAO) of the
municipality of La Trinidad, the average production per hectare on November 2005 to
April 2006 was 18.5 metric tons, but declined 14 metric tons on 2006 to 2007. This
decline in production was attributed due to improper growing practices of farmers, poor
quality of planting materials, and most of all, the presence of pest and diseases.

Strawberries are high-valued export crop that are grown in the temperate areas of
Western Australia. Soil born fungal pathogens such as root and crown rots are important
diseases of commercial strawberry crops and causes considerable yield reduction
worldwide. Survey done in Swan Coastal Plain North Perth on root and crown rot during
2005 and 2006, showed high incidence of the disease. Crown and roots of individual
plants including soil surrounding roots were tested for the presence of soil pathogens.
Research showed that Fusarium oxysporum f. sp. fragariae was consistently isolated
Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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from crowns and roots and Phytophthora cactorum was detected from roots, crowns and
soil. Other pathogens associated with the crown and roots of strawberry are Pythium spp.,
Phoma spp., Rhizoctonia spp., Colletotrichum spp., and Mycrophomina spp. (Golzar, et
al., 2007).

Morocko,( 2006) also stated that strawberry cultivation in many regions of the
world is constrained by serious disease that can affect the root system, crown, and the
basal part of the petioles causing death of the host and considerable reduction of the
yield. Among these are root rot and crown rot caused by Fusarium spp., Verticillium spp,
Phytophthora spp., and disorders caused by Colletotrichum spp.

In Japan the pathogen Fusarium shares in the black root rot disease complex on
strawberries together with Pythium spp., Rhizoctonia spp., and the lesion nematode
Pratylenchus penetrans. Its is a very serious disease of cultivated strawberries and the
damage caused by this disease has spread to open -culture farming in cooler regions such
as Akita Prefecture (Takashaki et al, . 2006). Pecnold (2001) also stated that black root
rot is the most common of all root disease in Indiana, United States. Since temperature in
Benguet is closely similar to weather conditions of this countries it’s no wonder that in
(2001), Ngitew first reported the occurrence of Fusarium wilt in the growing areas of
Benguet and recently, Reyes (2008) characterized the species infecting strawberry as
Fusarium oxysporum based on cultural and morphological traits but pathogenicity of the
isolate was not done.
Differentiating
between
Fusarium species responsible for the strawberry vascular
wilt and rotting of crown and roots through patogenicity is vital in the development and
implementation of effective control strategies. It is there for necessary to conduct
Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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pathogenicity test of the Fusarium associated on the crown and roots of strawberry for
the formulation proper management of the disease. Correct diagnosis will help strawberry
growers to improve their strawberry production thereby increasing their yield.
Objectives of the Study
1. Test the pathogenicity of Fusarium oxysporum isolated from strawberry,
2. Test the pathogenicity of Fusarium oxysporum on tomato, and chrysanthemum,
and strawberry

Time and Place of the Study
The experiment was conducted at Benguet State University, Department of Plant
Pathology Laboratory and Greenhouse from August to September 2008.







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REVIEW OF LITERATURE

The Crop
Strawberry is a perennial herb of the genus Fragaria of the rose family.
Strawberries are first grown in temperate regions throughout the world, but were first
cultivated in the United States and have become widely distributed crop in farms and
gardens. The white flower, have five calyx, five petals, many stamens, and numerous
seeds distributed on the berries (Redmund, 2007).

According to Hermano (1999), strawberry is a subtropical plant grown for about a
century as traditional crop in the high lands. Because of this crop, Baguio- Benguet has
been noted and popularly known as "Strawberry Region" of the country. Strawberry
production started in early years of the present century. This crop was introduced and was
found to be adaptable to the region. The introduction and evaluation were done probably
by Americans who established an Agricultural School in 1916 at La Trinidad, Benguet as
cited by (Reyes, 2008).

Strawberries thrives best in well drained, clay loam, loamy soils at a soil pH
ranging from 5.5-6.5 and temperature ranging from 140C to 230C ( Ngitew, 2003).

The Pathogen

Reyes (2008) cited that Fusarium is a filamentous fungus widely distributed on
plants and in the soil. It is found in normal micro flora of commodities, such as rice,
beans, soybean, and other crops. While most species are common at tropical and
subtropical areas, some inhabit the soil in cold climates. The fungus can stay in the soil
for the whole season or plant debris as a dormant mycelium or black-speck-sized bodies
Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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(microsclerotia).This microsclerotia can remain viable in the soil for many years. Under
favorable environmental conditions, they germinate and produce threadlike fungal
structures (hyphae).The hyphae can penetrate the root hairs or through wounds in the
rootlets, once inside the root the fungus invades and destroys the water-conducting tissue.
Destruction of water-conduction tissue results in reduced water uptake by the plant; thus
plants wilt and wither. As fungal colonies grow older, they produce microsclerotia in
infected host tissue and the disease cycle is completed (Anonymous, 2008).
According to Ellis (2006), the genus Fusarium currently contains over 20 species
of Fusarium solani, F. oxysporum, and F. chlamydosporum as the most common. Some
are plant pathogens causing root and stem rot, vascular wilt or fruit rot. Other species
causes storage rot and are important mycotoxin producers. Several species, notably F.
oxysporum, F. soloni, F. moniliforme, are recognized being pathogenic to man and
animals causing mycotic kertitis, onychomycosis and hyalohyphomycosis.

General Characteristics of Fusarium sp.
Colonies of Fusarium are fast growing, pale or brightly colored (depending on the
species) and may not have a cottony aerial mycelium. The color of the thallus varies from
whitish to yellow, brownish, pink, reddish or lilac shades. Species of Fusarium typically
produce both macro-and micro conidia from slender phialides. Microconidia are hyaline,
two- to several- celled, fusiform-to sickle shaped, mostly with an elongated apical cell
pedicellate basal cell. Microconidia are 1- to 2-celled, hyaline, pyriform to avoid, straight
or curved. Chlamydoconidia may be present or absent (Ellis, 2006).

Reyes (2008) characterizes Fusarium oxysporum as fast growing (4.1 cm colony
diameter in 1 week at 280 and 300 c) and produced white and pale violet pigment on PDA.
Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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Macroconidia has 3 septations and falcate to almost straight in shape. Abundant
microconidia that are oval, ellipsoidal and kidney-shaped and were formed in false heads
from short phailides. Chlamydospores were formed after three weeks of incubation in
Carnation leaf agar (CLA). While Fusarium solani is a slow growing (2.9 cm at 280 and
2.05 at 300 colony diameter after three weeks) and produce cream to pale brown and
white pigment in PDA. Its macroconidia has three septation, and sausage-shaped. It
produced very few microconidia that are oval and ellipsoidal in shape and were formed in
false–heads from long phailides. Chlamydospores were formed after two weeks of
incubation on PDA. The result of the cultural and morphological characterization of
Fusarium oxyporum done by Reyes (2008) is inconformity with the description of
Fusarium in the published laboratory manual for Fusrium by Burgess et al., (1994).
On the other hand, Nagarajan et al., (2004), added that Fusarium oxysporum f. sp.
fragariae is a fungal pathogen causing strawberry wilt disease. Research done on the
genetic variation of the 22 F. oxysporum f. sp. fragariae isolates showed a high level of
genetic variations.

Diseases Caused by Fusarium spp.
According to Agrios (1997), Fusarium causes vascular wilts primarily on annual
vegetables and flowers, herbaceous perennial ornamentals, plantation crops, weeds and of
mimosa tree (silk tree). Most of the vascular wilt- causing Fusaria belongs to the
Fusarium oxysporum. Different host plants are attacked by special forms or races of the
fungus.

Black root rot is caused by complex interactions of environmental factors, soil
fungi, nematodes such as Pratylenchus penetrans, fertilizer burn, soil compaction,
Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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herbicide damage, drought, and excess salt, water or improper soil pH. Several fungi are
implicated in the disease, including Rhizoctonia spp., Pythium spp., and Fusarium spp.
(Los and Schroeder, 2007).
A research in Oregon in 1930's and 1940's by Pscheidt (2007), implicated
Rhizoctonia spp., Fusarium spp., and Ramularia spp. with root rot of strawberry. He also
added that winter injury to roots encourages infection by Fusarium spp.

Symptoms Caused by Fusarium
oxysporum


Agrios (2005) described the symptoms caused by Fusarium oxysporum as
follows:
Fusarium oxysporum causes vascular wilt and rotting of crown and roots. The
leaves of infected plants or of parts of infected plants lose turgidity, become flaccid and
lighter green to greenish yellow, droop and finally wilt, turn yellow then brown and die.
Wilted leaves maybe flat and curled. Young tender shoots also wilt and die. In cross
sections of infected stems and twigs, discolored, brown areas appear as a complete or
interrupted ring consisting of discolored vascular tissue. In xylem vessels of infected
stems and roots, mycelium and spores of the causal fungus maybe present. Some of the
vessels maybe clogged with mycelia, spores, or polysaccharide produced by the fungus.
Clogging is increased further by gels and gums formed by the accumulation and
oxidation of breakdown of products of plant cells attacked by fungal enzyme. The
oxidation and translocation of such breakdown of product seem to be responsible for the
brown discoloration of affected vascular tissue.


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Management of Fusarium Disease

Black root rot is favored by wet soils and soils low in organic matter. As a result,
proper site selection and preparation are both important management tools for this disease
complex. Soil drainage should be good, low-lying areas that have a tendency to be poorly
drained should not be planted with strawberries.
Crop rotation by planting the area into cover crops for at least one growing
season, even if strawberries were not grown previously, to build organic matter in the
soil. Annual ryegrass, sudan grass and sorghum sudan can be grown after strawberry.

In the absence of a site with good soil drainage, the strawberry should be planted
in raised beds. The raised beds will allow excess soil water to drain from the strawberry
root system, creating an environment less favorable to disease causing fungi, and lessen
soil compaction that will occur near the root system. When planting, use only white
rooted plants which have been purchased from a reputable nursery.
Cultural practices which favor good plant growth and development must be done.
Soil and or tissue analysis should be performed each year to determine optimum fertilizer
application, and minimize soil compaction. Over or under irrigation of strawberry field
should be avoided (Pritts and Handley, 1991).

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MATERIALS AND METHODS

A. Isolation of Fusarium
Following the standard procedure in isolating Fusarium wilt, roots with infection
were cut into 1-2 mm sections, sterilized in10 % sodium hypochlorite (Clorox) for 1
minute, blot dried on sterile tissue paper and plated in water agar. The plates were
incubated for two to three days to allow the pathogen to grow. The pathogen was sub-
cultured unto carnation leaf agar (CLA) after one week.

B. Microscopic Observation
The characterization done by Reyes (2008) on Fusarium oxysporum from
strawberry which was based on the laboratory manual for Fusarium (Burgess et al., 1994)
was used as basis during the microscopic observation. These are: presence of sickle-
shaped macroconidia and oval shaped microconidia formed in false heads on short
monophialides; chlamydospores formed after 2-3 weeks; presence of septa on the macro
conidia formed on hyphae and branched conidiophore and the formation of the
conidogenoues cells.

C. Standardization of Inoculum
Pure-culture isolates obtained from strawberry plants grown from Balili,
Pomology, and Swamp was added with 10 ml sterile distilled water. Sterilized wire loop
was used to scrape the fungal growth to detach the spore and mycelia on the surface of
carnation leaf agar (CLA). An amount of 0 .1 ml. of Fusarium inocula was deposited on
the ridge of the haemacytometer after which, spores were counted. Standardization was
done thrice. The average count of the three trials from the five squares was multiplied
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with 20,000 to obtain the total spore count/ 0. 1ml.

D. Pot Bioassay

To confirm if the Fusarium isolated and identified by Reyes in 2007-2008 from
strawberry which is Fusarium oxysporum f. sp fragariae, the different isolates from
strawberry grown in Balili, Pomology, and Swamp, was inoculated in four week old
seedlings of Fragaria x ananassa cv. Sweet Charlie, Chrysanthemum (Dendrathema x
grandiflorum) and Tomato (Lycopersicun esculentum) to test their pathogenicity.
Chrysanthemum, Tomato, and Strawberry runners were dipped separately for one hour in
500 ml volume suspension of Fusarium oxysporum at a spore concentrations / ml of 6.46
x106(Balili), 8.53 x106 (Pomology), and 1.60 x106 (Swamp). The test plants after dipping
were planted on a black polyethylene bags (5”x 6” diameter) containing 800g of
sterilized soil. Untreated plants were dipped in tap water. The experiment was laid out
following the Complete Randomized Design (CRD) with three (3) replicates and with a
five (5) sample plants per replicate. The treatments are as follows:

Fusarium Isolate
T0- Un-inoculated (Control)
T1 –Fusarium oxysporum isolated from strawberry obtained from Balili area
T2 – Fusarium oxysporum isolated from strawberry obtained from Pomology area
T3 – Fusarium oxysporum isolated from strawberry obtained from Swamp area



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E.
Assessment of Symptoms on Inoculated Plants
Assessment of symptoms was done two weeks after planting. Presence of above
ground symptoms such as wilting, yellowing, stunting and death of seedlings were
carefully noted. Infected plants was counted and recorded. Below ground symptoms like
crown discoloration and root rotting were observed after two (2) and five (5) weeks.

Data Gathered
1. Above ground symptoms- yellowing, wilting stunting and seedling death was
observed two weeks after inoculation until the termination of the study.
2. Below ground symptoms- black root rot and crown discoloration was observed
and recorded at termination of the experiment.
3. Number of infected plants- number of wilted and dead plants were counted and
recorded after two weeks and thereafter.
4. Re-isolation of Fusarium from inoculated strawberry plants.



Experimental Set-up


Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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RESULTS AND DISCUSSION

Above Ground of Symptoms

Wilting, yellowing and stunting and death of strawberry plants inoculated with the
different isolates of Fusarium appeared two weeks after planting. The symptoms
progressed on the third until the fourth week of the experiment. Golzar, et al., (2007)
described that the infected strawberry plants wilt and die either slowly or rapidly soon
after the first symptoms are noted due to destruction of the roots. Affected plants die
more rapidly in very dry than in moist soils. Infected young plants often wither and die
without producing any fruits.
Morocko (2006) also stated that the common symptoms of root rot on
strawberries shows patchy appearance of stunted plants in the field, and discoloration of
leaves that turns yellow. Similarly, Pritts and Handley (1991) described strawberry
infected plants to have general lack of vigor with poor runner growth and small berries.
Plants may collapse when water demand is high during spring growth, during or after
fruiting, or during drought stress. Figure 1 shows wilted, yellowing and stunted plants.







A
B
C

gFigure 1. Strawberry plants showing wilting, yellowing and stunting: (a) inoculated with
6.46 x106 /ml spores (Balili isolate), (b) inoculated with 1.60x106spores / ml

(Swamp isolate), (c) inoculated with 8.53 x 106 spores/ ml (Pomology isolate)

Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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Except for strawberry that showed above ground symptoms of infection after
inoculation, pathogenicity test showed that Fusarium isolated from strawberry is not
pathogenic to the other test plants, (Table 1) Chrysanthemum and tomato dipped in
different spore concentrations of Fusarium (6.46 x106, Balili isolate), (8.53x106,
Pomology isolate) and (1.60x6 Swamp isolate) were not infected by the fungus Fusarium
oxysporum (Figure 2).

Table 1. Above ground symptoms observed two weeks after inoculation until the
fifth week


SYMPTOM INOCULATED
PLANTS
Balili Isolate
Chrysanthemum
Tomato
Strawberry
• Wilting
- - +
• Yellowing of
- - +
leaves
• Stunting
- - +
Pomology Isolate



• Wilting
- - +
• Yellowing of
- - +
leaves
• Stunting
- - +
Swamp



• Wilting
- +
• Yellowing of
- - +
leaves
• Stunting
- - +

This result corroborates the findings of Freeman and Rodriguez (1993) in their
wilt – screening for resistance against Fusarium. Fusarium oxysporum f. sp. melonis
which is specific to muskmelon (Cucumis melo L.) is unable to cause disease in
watermelon (Citrullus lanatus (Matsuma and Nakai).



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A
B
Figure 2. Chrysanthemum (a) and tomato (b) inoculated with the same amount of spore
concentration of Fusarium oxysporum without above ground symptoms
after two to four weeks.


Below Ground Symptoms
Strawberry plants that showed wilting, yellowing and stunting also exhibited
rotting on roots that is colored black. The crown of infected plants also showed brown to
black discoloration. According to Pritts and Hardley (1991), infected roots are rotted at
their tips or appear mottled with black lesions among the white roots and blackening of
the entire root system. In the early stage of black root rot, the root core is white (Figure
3), if the plants are severely affected, both the core and the tissue will be black.


A
B







Figure 3. Sample of infected roots with white core, pointed with arrow (a) and (b)
healthy roots
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H
I





A
B
C
Figure 4. Healthy and infected strawberry roots and crown(a) Healthy strawberry roots
(un-inoculated) (b) healthy and infected crowns inoculated with 8.53 x 106
spores/ ml (Pomolog isolate) (c) with root rot, inoculated with 1.60 x106
spores /ml (Swamp isolate)

Number of Dead Strawberry Plants
Table 2 shows the number of dead strawberry plants, three weeks after planting
and until the termination of the experiment. Out of fifteen sample plants per treatment,
four from those inoculated with Balili isolate, six inoculated with Pomology isolate, and
five from those inoculated with Swamp isolate, wilted and died. Number of dead plants
increased after 28 days with three additional from Balili isolate, four from Pomology
isolate and one from Swamp Isolate. The highest total infected plants were recorded from
the treatment inoculated with the Pomology isolate, (Figure 5). This also showed that the
higher the spore concentration used, the more number of plants died.





Figure 5. Sample of dead strawberry seedling
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Table 2. Wilted plants observed and recorded 21 and 28 days after inoculation

TREATMENTS
21 DAYS
28 DAYS
TOTAL
Control/ un-inoculated
-
-
-
Balili Isolate
4
3


7
Pomology Isolate
6
4
10
Swamp Isolate
5
1
6

Chrysanthemum Inoculated Plants
No above and below ground sym

ptoms on chrysanthemum were noted after three
and four weeks after panting and until the termination of the experiment. (Table 3 and
Figure 6).

Table 3. Chrysanthemum plants inoculated with the different isolates of Fusarium
Oxysporum

TREATMENTS
21 DAYS
28 DAYS
TOTAL
Control(un-inoculated) -
-
-
Balili Isolate
-
-
-
Pomology Isolate
-
-
-
Swamp Isolate
-
-
-





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A
B










C
D



Figure 6. Inoculated roots of chrysanthemum with out symptoms (a) Un- inoculated
(b) healthy roots inoculated with 6.46 x106, spores/ ml (Balili isolate)

(c) healthy roots inoculated with 8.53 x 106 spores/ ml (Pomology isolate
(d) healthy roots inoculated with 1.60 x106 spores /ml (Swamp isolate)



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Tomato Inoculated Plants
Similarly, tomato plants inoculated with the different isolates of Fusarium did not
show any symptom of infection on leaves, stems and roots (Table 4). This corroborates
the findings of Golzar, et al., (2007), whereby Fusarium isolated from strawberry and
inoculated to tomato and cucumber plants did not cause infection, (Figure 6).

Table 4. Tomato plants inoculated the different isolates of Fusarium oxysporum

TREATMENNTS 3rd WEEK
4th WEEK
TOTAL
Control/ untreated
-
-
-
Balili Isolate

-
-
-
Pomology Isolate
-

-
-
Swamp Isolate
-
-
-












A
B





Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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C
D



Figure 7. Inoculated roots of tomato with out symptoms(a) Un- inoculated (b) healthy
roots inoculated with 6.46 x106, spores/ ml (Balili isolate) (b) healthy roots
inoculated with 8.53 x 106 spores/ ml(Pomology isolate) (c) healthy roots
inoculated with 1.60 x106 spores /ml(Swamp isolate)

Fusarium oxysporum Isolates
Re-isolation of infected root and crown of strawberry inoculated with Fusarium
oxysporum was done. Figure 8 shows the original culture and re- isolated Fusarium
oxysporum from Balili, Pomology, and Swamp.










A
B
Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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A
C





C
F
D










E
F

Figure 8. (a) Macroconidia in hypha from the original Balili isolate (b) Conidia in a
developing branched conidiophore re- isolated from the inoculated
strawberry, (c) Branched conidiophore with macroconidia from the original
Pomology isolate (d) Branched conidiophore with macroconidia re-isolated
from the inoculated strawberry, (e) Macroconidia in hyphae from the original
Swamp isolate (f) Macrocnidia in hyphae re- isolated from the inoculated
strawberry (400x).

Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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SUMMARY, CONCLUSION AND RECOMMENDATION

Summary

Pathogenicity test of Fusarium oxysporum isolated from Strawberry obtained
from Balili, Pomology and Swamp was inoculated on chrysanthemum, tomato and
strawberry to determine if root rot and crown rot or discoloration symptoms will develop
in all the test plants.

Results of the study revealed that the three isolates of Fusarium from Balili,
Pomology, and Swamp were pathogenic to strawberry only. No wilting, yellowing,
stunting, and root rotting symptom developed both for chrysanthemum and tomato
inoculated with spore concentrations of Fusarium at (6.46 x 106 /ml (Balili isolate),
8.53x106/ ml (Pomology isolate) and 1.60x 106/ml (Swamp isolate). Fusarium above
ground symptoms produced by strawberry inoculated plants was stunting, and wilting.
Symptoms that developed in roots were in the form of root rot and brown to black
discoloration of the crown.

Conclusion

From the results, the Fusarium obtained from strawberry caused infection only on
the inoculated strawberry seedlings. Inoculated chrysanthemum and tomatoes were not
infected. They did not show any symptoms of wilting, yellowing, stunting, and root rot
on chrysanthemum and tomatoes, which indicates that Fusarium infecting strawberry is
Fusarium oxysporum f. sp. Fragariae having infected strawberry only.


Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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Recommendation
Based on the above findings, it is recommended that appropriate management of
Fusarium oxysporum f. sp. fragariae be done in the areas where strawberries are grown
to improve production.



















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LITERATURE CITED

AGRIOS, G. N. 1997. Plant Pathology.4th Edition. University of Florida, Academic
Press Inc., San Diego, California. Pp.34. 523-525 &538-540

ANONYMOUS, 2007. Fusarium species. Retrieved 12 September from
http://search.yahoo.com/searchei=utf-8&fr=fUSARIUM

ELLIS, D. 2006. Mycology Online, Fusarium sp. The University of Adelaide Australia.
Retrievedfromhttp://www.mycology.adelaide.edu.au/Fungal_Descriptions//Fusari
um/

FREEMAN, S. AND RODRIGUEZ, R.J. A Rapid Technique for Assessing
Pathogonicity of Fusarium Oxysporum f. sp. Niveum and F. o. melonis on
cucurbits. Vol. 77.No.12.

GOLZAR, A., PHILLIPS, D., AND MACK, S. 2007. Occurrence of Strawberry root and
crown rot in Western Australia. Department of Agriculture and Food Western
Australia, Locked Bag 4, Bentley Delivery Centre, WA 6983, Australia.

HERMANO, F. G.1999. Strawberry Production Management and Technology. Benguet
State University La Trinidad, Benguet.

MOROCKO, I. 2006. Characterization of the Strawberry Pathogen and Biocontrol
Possibilities. Swedish University of Agricultural Sciences.Pp.7, 8, 9.

NAGARADIAN, G., HYEON NAM1, M., YOUNG SONG, J., JOON YOO2, S., GI
KIM, H. 2004. Genetic variation in Fusarium oxysporum f. sp. fragariae
Population based on RAPD and DNA RLFP Analysis. The Plant Pathology
Journal, Department of Agriculture Biology, Chugnam National University,
Daejeon 305-764, and Korea.

NGITEW, A. 2003. Survey and Identification of Fungal Diseases of Strawberry. BS
Thesis. Benguet State University, La Trinidad Benguet. Pp. 13.

LOS, L. M. and M. L. SCHROEDER. 2007. Black Root Rot of Strawberry. University of
Connecticut. Integrated Pest Management. Retrieved 25 September from
http://www.hort.uconn.edu/ipm/homegrnd/htms/25strrot.htm

MUNICIPAL AGRICULTURIST OFFICE. 2007. La Trinidad, Benguet.

PECKNOLD, P. C. 2001. Strawberry Root Disease. Purdue University Cooperative
Extension Service West Lefayette, IN 47907. Retrieved
fromhttp://www.ces.purdue.edu/extmedia/BP/BP-46.html

Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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PRITTS, M. and HANDLEY, D. 1991.Bramble Production Guide. Northeast
Agricultural Engineering Service Pub. No. NRAES-35. Pp. 189.

REYES, M. 2008.Characterization of Fusarium spp. associated with the crown and roots
of Strawberry (Fragaria x ananassa Duch) in La Trinidad Benguet.Pp.6-13.

PSCHEIDT, J. W. Ed. 2007. Strawberry Root Rot Complex. OSU Extension Office.
Retrieved from http://plant-disease.ippc.orst.edu/disease.

TAKAHASHI H., Y. YOSHIDA, and H. KANDA, 2007. WB-B33, A New Fusarium
Wilt-Resistant Strawberry Line. ISHS Acta Horticulture 760:IHC2006 - II
International Symposium on Plant Genetic Resources of Horticultural Crops.
Retrieved 25 September from http://www.actahort.org/books/760/760_57.htm

WILHELM, S., NELSON, R. 2006. Pest Management Systems for Strawberry Diseases
Vol.111.Pp.395.















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APPENDICES

Appendix Table 1. Above ground symptoms observed two weeks after inoculation until
the fifth week.

SYMPTOM INOCULATED
PLANTS
Balili Isolate
Chrysanthemum
Tomato
Strawberry
• Wilting
- - +
• Yellowing of
- - +
leaves
• Stunting
- - +
Pomology Isolate



• Wilting
- - +
• Yellowing of
- - +
leaves
• Stunting
- - +
Swamp



• Wilting
- +
• Yellowing of
- - +
leaves
• Stunting
- - +


Appendix Table 2. Wilted/dead plants observed and recorded 21 and 28 days after
inoculation

TREATMENTS
21 DAYS
28 DAYS
TOTAL
Control/ un-inoculated
-
-
-
Balili Isolate
4
3


7
Pomology Isolate
6
4
10
Swamp Isolate
5
1
6








Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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Appendix Table 3. Chrysanthemum plants inoculated with the different isolates of
Fusarium oxysporum

Treatments
21 DAYS
28 DAYS
TOTAL
Control/un-inoculated
-
-
-
Balili Isolate
-
-
-
Pomology Isolate
-
-
-
Swamp Isolate
-
-
-


Appendix Table 4. Tomato plants inoculated the different isolates of Fusarium
oxysporum

TREATMENTS 3rd WEEK
4th WEEK
TOTAL
Control/ untreated
-
-
-
Balili Isolate
-
-
-
Pomology Isolate
-
-
-
Swamp Isolate
-
-
-

Pathogenicity Test of Fusarium oxysporum f. sp. fragariae / Jennie T. Bawayan. 2008

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Document Outline

• Pathogenicity Test of Fusarium oxysporum f. sp. fragariae.
  • BIBLIOGRAPHY
  • ABSTRACT
  • TABLE OF CONTENTS
  • INTRODUCTION
    • Objectives of the Study
    • Time and Place of the Study
  • REVIEW OF LITERATURE
    • The Crop
    • The Pathogen
    • General Characteristics of Fusarium sp.
    • Diseases Caused by Fusarium spp
    • Symptoms Caused by Fusariumoxysporum
    • Management of Fusarium Disease
  • MATERIALS AND METHODS
    • Isolation of Fusarium
    • Microscopic Observation
    • Standardization of Inoculum
    • Pot Bioassay
    • Assessment of Symptoms on Inoculated Plants
    • Data Gathered
  • RESULTS AND DISCUSSION
    • Above Ground of Symptoms
    • Below Ground Symptoms
    • Number of Dead Strawberry Plants
    • Chrysanthemum Inoculated Plants
    • Tomato Inoculated Plants
  • SUMMARY, CONCLUSION AND RECOMMENDATION
    • Summary
    • Conclusion
    • Recommendation
  • LITERATURE CITED
  • APPENDICES

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