Department of Agriculture-Cordillera Administrative Region

Republic of the Philippines
Department of Agriculture –CARFU
Cordillera Highland Agricultural Resources Management
Project
Sto. Tomas Road, Dairy Farm, Baguio City
Tele/Fax Nos. 444-79-94/444-83-29
PO Box 1158
E-Mail: charm@mazcom com
SURVEY,
IDENTIFICATION
AND MAPPING OF
SOIL BORNE
PLANT
PATHOGENS IN
THE FARMING
AREAS OF
MOUNTAIN
PROVINCE
JOAN DIMAS-BACBAC
CHARLES A. PICPICAN

RATIONALE:
In the Commodity Analysis and Planning (CAP) workshop conducted in
Sagada, Sabangan, Bontoc and Tadian, the farmers identified a “ soil borne”
disease that was affecting their bell pepper crops. From the workshop, it was
furthermore learned that the “condition” was spreading and that it was starting
to cause economic damage. The farmers therefore recommended that a soil
survey be done to determine which areas have the “disease” and which areas
are free from it.
Soil-borne pathogens cause heavy yield losses in the world’s major staple
and vegetable crops, their severity often being exacerbated by edaphic and
climatic stresses imposed on plants grown increasingly under more marginal
conditions in response to mounting land pressure.
Diagnosis of any condition entails a thorough study of the infected plants
to determine whether the condition is indeed due to pathogenic factors or due to
environmental/soil conditions. Microscopic and cultural studies have to be done
to describe the associated organism(s) and to determine its (their) identity. Only
when the identity of a pathogen is known can a management scheme be
determined. Furthermore, part of any successful disease management program
is the knowledge of the incidence of the pathogen in affected areas and the
severity of the disease in these areas.
OBJECTIVES
This study aimed to survey the major vegetable planting areas of Mt.
province to gather information about an identified soil-borne condition affecting
vegetable crops in the area. Specifically, it aimed to:
1) Isolate and identify the cause of the condition;
2) Describe the morphological and physiological characteristics of the
pathogen;
3) Sample soils from vegetable producing areas of Mt. Province and
measure soil populations of the pathogen;
4) Determine the incidence and severity of the disease in Mt. Province;
and
5) Map the incidence and severity of the disease in the area

METHODOLOGY
The study was done in three major phases: (1) soil sampling and
collection of disease specimens; (2) disease diagnosis and laboratory work; and
(3) terminal report writing.
A. Soil Sampling and collection of Specimens
Soil samples were collected from areas pre-identified on consultation with
CHARMP Extension Support Services (ESS) personnel. Geographic Information
System (GIS) personnel were also involved in site visits so they could gather
coordinates for mapping.
At the site, an ocular assessment was made of the condition of the
standing crop. Presence of diseases and disease severity were determined. The
cropping history of the site was also determined from interviews with the farmer.
Three sites per barangay were sampled with three random samples taken
per site. Site samples were then combined in the laboratory and a composite
sample taken for examination. Samples were stored in proper bags in the lab
while waiting processing.
Samples of diseased plants were also taken at the sites. Specimens were
placed in polyethylene bags and labeled with date and place of collection.
Samples were then brought to the laboratory for incubation and processing.
B. Laboratory Work and Disease Diagnosis
Determining Population of Microbes. Ten grams of each composite
soil sample was diluted in 100 ml of sterile distilled water. A standard dilution
series was performed up to 10 -8 . Only the last three dilutions were plated out
following standard procedures. Plates were incubated at 30 0 C for 24 hours.
Colony counts were then performed and the microbial population was then
counted and population per microbe type was expressed as cfu/g soil.
Isolation of Soil Microbes. From the composite soil, sample, ten
grams was diluted in 100 ml of sterile distilled water. One ml of the soil
suspension was plated out on the appropriate differential media. Inoculated
plates were then incubated at 30 0 C for 224-48 hours. Growth of microorganisms
was observed and described. Re-isolation of different microorganisms was done
on previously prepared and plated standard media for bacteria and fungi. These
plates were then incubated at the conditions previously stated.

Cultural and Morphological Characterization of Isolates. After a
24-48 hour incubation period, cultural and morphological characteristics of the
isolated microbes were determined through ocular examination of pure culture
and microscopic examination of semi-permanent slide mounts of the microbes.
Morphology of the bacterial isolates was determined through the Gram stain
procedure. Observations were noted.
Biochemical Characterization of Bacteria. Standard procedures for
biochemical characterization of bacterial isolates were done to aid identification.
Identification of Isolated Microbes. Identification of known and
unfamiliar or newly-isolated microbes was done using recorded symptomatology
and morphological, cultural and biochemical characteristics. For unfamiliar or
newly isolated microbes, pathogenicity was confirmed through pathogenicity
tests.
Documentation. All results and observations were duly noted and photo
documentation was made of all laboratory results.
C. Mapping of Soilborne Diseases
Members of the CHARMP GIS unit accompanied the researchers to the
sampling sites. Locations and coordinates were duly noted. Data on the incidence
and severity of the soilborne diseases was summarized then shared with the GIS
unit personnel. They plotted the data and produced the maps.
RESULTS AND DISCUSSIONS
a. Location and Description of Sampling Sites
Fifteen sites were sampled representing five municipalities of Mt. Province
(Table 1). From each municipality, three sites were selected and three samples
were taken from each site.

Table 1. Municipalities covered and the sampling sites from each municipality
MUNICIPALITY
SAMPLING SITE
Sabangan Napua *
Camatagan *
Tambiangan
Bauko
Tadian
Sagada
Bontoc
*
not CHARMP –covered area
Tapapan
Leseb )
Maba-ay
Lubon
Bantey
Poblacion
Ambasing
Patay
Madongo
Bayyo
Talubin
Gonogon *
Some non-CHARMP areas were sampled on the recommendation of the
local AT’s . On the other hand, some CHARMP sites were not sampled due to
absence of a standing crop. This is because the probable presence of a soilborne
pathogen is usually presumed from symptoms of above-ground plant parts.
The smallest area sampled was 50 m 2 in four out of the five
municipalities. In Bontoc, the smallest site sampled was 80 m 2 . The largest
sampling site varied over the areas and ranged from 300 to 500 m 2. . The soil pH
on the other hand, was more homogenous. Lowest pH in all sites was 3.5 while
the highest ranged from 5.0 to 6.5. These readings classify the soil in the area in
the acidic to slightly acidic range. The acidic lower readings are not favorable for
vegetable crop production and are usually implicated in increasing the severity of
such soilborne diseases as the clubroot of crucifers.

Table 2. Estimated area and soil pH of sampling sites
SITE ESTIMATED AREA ( M 2 ) SOIL Ph_________
Smallest Largest Low High
Bauko 50 500 3.5 6.0
Bontoc 50 500 3.5 6.0
Sabangan 50 500 3.5 6.0
Sagada 50 400 3.5 6.5
Tadian 80 400 3.5 5.0
B. Cropping History of Sampling Sites
Knowledge of the cropping history of an area provides insights into
incidence and severity of many plant diseases and eventually aids in the
development of disease management measures. Based on interviews with the
farmers and observations, the cropping history of the sites was generally
homogenous (Table 3). Monocropping was common either in the form of
planting the same crop in succession or planting a crop from the same family.
For example, sweet pepper was followed by sweet pepper or by another
Solanaceous crop like potato or tomato.

Table 3. Crop succession in the sampling sites
SAMPLING SITE
CURRENT CROP______________
Same AS Previous Same Family as Different Crop
Crop Previous Crop Family
Bauko (n=12) 3 3 6
Bontoc (n=8) 4 0 4
Sagada (n=11) 3 2 6
Sabangan(n=10) 3 3 4
Tadian (In=8) 2 1 5
TOTAL ( n=49) 15 9 25
Planting the same crop in succession or a crop from the same family
constitutes monocropping. Among 49 current cropping, half were planted to the
same crop or belonged to the same family as the previous crop. This finding is
especially important because monocropping provides the pathogen with a
perpetually available food source. Thus, the soilborne pathogen can multiply
undeterred by food resource limitations.
The increase in pathogen population increases the inoculum reservoir in
the soil and eventually results to increased disease severity. Furthermore, the
large inoculum reservoir enhances the changes for dissemination of the
pathogen to neighboring fields.
C. Identity of suspected Soilborne Plant Disease of Sweet Pepper
Based on symptomatology ( Figure 1a and Figure 1b) and the cultural and
morphological characteristics (figure 3) of the isolated microorganisms, the
disease of sweet pepper that was the major complaint of the Mt. Province
farmers is the bacterial blight of pepper. This disease is also known as the brown
rot of Solanaceous crops and is caused by Ralstonia solanacearum (E.F. Smith)
Yabuuchi et. Al – the same bacterium that causes bacterial wilt in potato,
tomato, eggplant, tobacco, banana and other Musa species.

Figure 1a.
Sweet pepper plant showing the
most common symptoms of
bacterial wilt
Among these plants, disease incidence is particularly common in tobacco,
tomato, potato, pepper, eggplant, peanut and banana. Buddenhagen and
Kelman (1964) defined the strains attacking potato at lower temperature as race
3, those attacking banana and Heliconia as race 2, and others as race 1. Five
biovars are differentiated by the use of carbohydrates. High correlations have
been recognized among biovars, pathotypes, phage types and bacteriocinogenic
types.
Symptoms of Bacterial Wilt. Acute wilting is particularly notable in
susceptible young plants. Once wilting is established, the plants quickly wither
and die without recovering. Different symptoms may develop depending on the
plant species, cultivar, growth stage, and environmental conditions such as
temperature. They included aboveground symptoms such as yellowing, dwarfing,
stunting and development of adventitious roots and underground symptoms such
as decay of tubers of potato and ginger. In some plants, flowers and fruits may
fall prematurely
Discoloration of the vascular system from pale yellow to dark is a common
feature of diseased plants. The surface of a traverse section is moist, and
droplets of milky bacterial ooze exude on the invaded tissue ( Figure 2b).
Infected tuber of potato and ginger externally appear healthy when disease has
not advanced, but section reveals the discoloration of the vascular region and
bacterial exudation from invaded tissue.
Occurrence of Bacterial Ooze. To determine the presence of bacteria
in the vascular bundles of the plant, a piece of the stem about 2 inches long is

cut and cleaned of surface dirt. One end of the sample is submerged in clean tap
water or distilled water. The sample is held in place by piercing the other end
with an unraveled paper clip that is hooked to the side of the water-containing
vessel. After about 5 minutes, a mucous-like substance oozes out of the
submerged stem rot. This confirms that the wilt is caused by a bacterium
( Figure 2b) .
Figure 2b.
Bacterial ooze from a cut end of the stem of an
infected pepper plant .
Morphological Characteristics of R. solanacearum. Ralstonia
solanacearum is a Gram-negative rod (Figure 4) measuring about 0.5-0.7 x 0.5-
2.0 um. It is none-spore forming, noncapsulate and nitrate-reducing. Also, it is a
catalase-positive, ammonia-forming, strictly aerobic and monotrichous bacterium.
Confirmation of the presence of Ralstonia solanacearum bears significance
to farmers growing vegetables in Mountain Province. The bacterium is one of the
most versatile and most widespread plant pathogens. It is present in many forms
called races ( Buddenhagen, 1954) and isolates from the Cordillera ( Lando,
2002) possess a high degree of genetic variability. This genetic variability further
translates to its versatility in exploiting various niches and hosts. The pathogen
furthermore is easily spread through irrigation water, contaminated tools and
other farm implements, through seed or other planting materials and in some
instances, by insects.

Figure 3. Typical colonies of Ralstonia solanacearum
on differential media (Triphenyl tetrazolium chloride
agar)
Figure 4. Gram-stained slide of Ralstonia
solanacearum (1000x)
Occurrence of R. solanacearum in Mt. Province. The occurrence of
R. solanacearum in Mt. Province can be seen in Figure 5. Without studies to
trace the genetic characteristics of the isolates, the source of the pathogen can
only be surmised at this point. The pathogen may be native or may have been
carried latent in potato seed tubers. Earlier studies done in other countries
showed that the pathogen could be isolated from newly cultivated areas. In this
study, newly cultivated areas were free of the pathogen at the time of sampling
based on above-ground symptomatology. This may therefore point to the
contaminated planting material as the most probably source of the pathogen in
the area.
The knowledge of the distribution of Ralstonia in the province has
epidemiological significance. At a glance, the viewer can see where the pathogen
exists and will know to avoid obtaining planting material from these sites. Also,
the viewer will be able to avoid planting susceptible crops in the areas where the
bacterium has been found to occur. The knowledge will prevent further spread of
the pathogen and also prevent further economic loss.

D. OTHER SOILBORNE MICROORGANISMS ISOLATED
Twenty-nine soilborne microorganisms occurred in soils taken from the
sampling sites. Of these microorganisms, only 23 were identified. Eighteen
isolates were pathogenic and among them, six were potentially destructive
(Table 4). Among the other microorganisms, five isolates were potential
biocontrol agents.
Figure 5.
Map showing the distribution of
Ralstonia solanacearum in Mt.
Province sampling sites.
Table 4. Identified soilborne microorganisms isolated from soils in the sampling areas
POTENTIALLY OTHER LESS DESTRUCTIVE POTENTIAL
DESTRUCTIVE PATHOGENS BIOCONTROL AGENTS
Fusarium Deightoniella Nodulosporium Trichoderma
Plasmodiophora Aspergillus Clasdosphorium Penicillium
Rhizoctonia Phymatotrichopsis Moniales Bacillus
Sclerotium Culvularia Amerosperium Pseudomonas
Verticillum Aureobasidum Trichothecium Paecelomyces
Xanthomonas Rhizopus Hansfordia

Soilborne Plant Pathogen. The presence of the potentially destructive
pathogens presents a problem for vegetable farmers in Mt. Province (Figure 6).
As shown in Table 5, the diseases taken individually are capable of causing huge
economic losses. When present together (Figure 7), options for crop rotation are
generally reduced and disease management become more difficult.
For example, the presence of R. solanacearum and Plasmodiophora
brassicae together eliminates all solanaceous plants ( potato, tomato, eggplant,
pepper) and all crucifers ( cabbage, Chinese cabbage, pechay, mustard, broccoli,
cauliflower) from the choices that can be used in crop rotation. The Solanaceae
sand the Brassicae represent most of the major vegetable cash crop. The
inability to plant them means a loss of potential income for the farmers in the
area.
Table 5. Soilborne pathogens and their potential for destruction
PATHOGEN DISEASE(S) CAUSED ECONOMICALLY AVERAGE DAMAGE
IMPORTANT HOST ESTIMATES (%)
Fusarium Root & stem rot Wide host range 30 %
Plasmodiophora Clubroot all Brassicas 57 %
Rhizoctonia root & stem rots Most crops 35 %
Sclerotium Root & stem rots Most crops 25 %
Verticillium Vascullar wilts Wide host range 15 %
Xanthomonas Black rot Brassicas 55 %
*From ocular assessment of standing crop.
Potential Biocontrol Agents. One important finding is the isolation of
potential biological control ( biocontrol) agents (Table 4, Figure 8). Biological
control is defined as “ any condition under which or practice whereby survival or
activity of a pathogen is reduced through the agency of any living organism
( except man) with the result that there is a reduction in the incidence of the
disease caused by that pathogen” ( Garrett, 1970). Baker and cook (1974) have
given a broader definition that “ biological control is the reduction of inoculum
density or disease-producing activities of a pathogen of a parasite in its active or
dormant state, by one or more organisms, accomplished naturally or through

manipulation of the environment, host, or antagonist, or by mass introduction of
one or more antagonists”.
Figure 6. Some of the potentially destructive plant pathogens isolated
from the samples
Figure . Map showing occurrence of plant pathogens in the sampling
sites

Table 6 shows the potential for each of the potential biocontrol agents
that were isolated from the study area and that could be exploited in disease
management strategies against major pathogens occurring in the area.
Table 6. Biological control potential of some soilborne microorganisms isolated
from the sampling sites
_________________________________________________________________
POTENTIAL BIO CONTROL AGENT SOME GENERA OF PATHOGENS
REPORTEDLY CONTROLLED*
Trichoderma
Fusarium, Rhizoctonia , Pythium,Phytopthora,
Verticillum, Sclerotium, Botrytis
Peniciullum
Bacillus
Pseudomonas
Paecelomyces
penicillum, Botrytis
Rhizoctonia , Monilia, Fusarium, Streptomyces
Monilia, Penicillum, Botrytis, Fusarium
Meloidogyne, Heterodera, etc, (Root knot and
Cyst nematode)
Biological control agents work in several ways to control pathogenic
microorganisms. Biocontrol microorganisms may produce one or more
metabolites ( most often antibiotics), which either inhibit or kill the pathogens.
They may compete with the pathogens for food or they may parasitize the
pathogens. They may also prevent the spores of pathogenic fungi from
germinating. Biocontrol microbes have also been known to stimulate the
resistance response of the host.
If proven to be effective, these could form an integral part of integrated
management measures that can be taken against the identified plant diseases in
the area. One major advantage of using these biocontrol agents is that they are
most often highly specific to their hosts and thus have minor impact on the
native microbial biodiversity in the soil. Furthermore, unlike conventional
pesticides, they do not adversely affect the applicator. Aside from their ability to
suppress or kill pathogens, some biocontrol agents have the capacity to produce
growth factors and thus promote plant growth.

Unidentified Microorganisms Isolated. Figure 9 shows the
photomicrographs of some of the microorganisms that we were not able to
identify. This does not mean, however, that these unidentified microbes are
unimportant. On the contrary, the importance of these microorganisms must be
emphasized.
They could be potential pathogens or biocontrol agents. As such, it is of
importance that their identity be known and confirmed. Once their identity is
known, adequate steps can be taken to prevent possible disease outbreaks in
the case of the pathogens and/or determine their biocontrol potential. They
could play other important ecological roles such as mycorrhizal fungi. However,
their roles and ultimate importance can be determined only when their identity is
known.
Figure 9. Photomicrographs of unknown microorganisms
isolated from sampling sites

VI. SUMMARY , CONCLUSIONS AND RECOMMENDATIONS
SUMMARY
A study was conducted to identify a reported soilborne disease attacking
sweet pepper plants CHARM Project-covered areas in Mt. Province and to
determine the occurrence of this disease. The sampling sites had a range of soil
pH from 3.0 to 6.5. The smallest area was 50 m 2 . These sampling sites were
planted to various vegetables and some to rice.
Ralstonia solanacearum, which is the causal organism of bacterial wilt of
pepper, was isolated from twelve out of 47 sites sampled. Of these 12 sites,
severity in seven sites was higher than 20 %. Seven other soilborne pathogens
of potential economic importance were isolated. These pathogens included
Fusarium, Rhizoctonia , Sclerotium, Verticillum, and Xanthomonas. The presence
of Plasdiophor brassicae, the causal organism of clubroot of crucifers was also
noted.
Three minor plant pathogens and 13 other microorganisms were isolated
from the sites. Five of these microorganisms belonged to genera with noted
biological control capabilities while the rest remain unidentified.
CONCLUSIONS
Based on the data gathered from the study and the observation made on
site, the following are concluded:
1. Some soils in the vegetable-producing areas of Mt. province are
infested with Ralstonia solanaceraum.
2. Based on the incidence, bacterial wilt is still limited in the sampling
sites.
3. Severity of the disease can be directly related to cropping pattern. In
areas with a 90 % severity observed, the previous crop was usually a
solanaceous crop.
4. Other major pathogens exist whose impact may increase with
continuing vegetable culture in the area.
5. Exploitation of native biocontrol agents can be used as integral part of
disease management. However, the biocontrol potential of these
identified agents must be verified first.

RECOMMENDATIONS
The following are therefore recommended:
1. Appropriate disease management measures should be employed areas
where bacterial wilt and major soilborne pathogens were found. These
measures include:
a) Use of clean seed or planting material.
Buy potato seeds only from accredited suppliers to be assured
that this is clean
b) Correct crop rotation
Crops in rotation should not belong to same family ( eg.
Crucifers, solanaceous crops, etc)
c) Use of cultural management practices which can inhibit or kill
the pathogens
c.1) Irrigation – flooding versus nematodes, fungi and some
bacteria
c.2) Avoid water logging
d.) Apply lime on the soil depending on the result of the soil
sample
e) Enhance native antagonistic microorganisms
e.1. Incorporation of organic matter eg: compost and animal
manures
e.2. Keep soil aerated
e.3. Observe care in use of fungicides and fertilizers.
2) Further study should be done to :
a) Include all of the vegetable producing sites in CHARMP-covered
site in the Province. It would also be better if the entire
province were surveyed.
b) Determine the incidence and severity of major diseases on
crops throughout the year.
c) Identify alternative hosts of the pathogens identified
d) Plot soil characteristics in the area ( pH, OM, NPK)
e) Evaluate efficiency of isolated potential biocontrol agents