Plant Disease Diagnosis

Prepared by

Cherry Relevante-Belagantol
Senior Researcher-Pathology
East-West Seed Philippines

Conrado H. Balatero
East-West Seed Group Research Manager-Pathology

Plant Pathology Department- Research and Development

 Learning Objectives

At the end of this training module, participants will be able to:

1. Apply/use the disease triangle in plant disease diagnosis.
2. Describe symptoms and signs of plant diseases caused
by biotic and abiotic agents.
3. Explain why laboratory-based tests are crucial in
diagnosis of plant diseases.
4. Identify diseases caused by fungi, bacteria and viruses
using appropriate diagnostic procedures.

Plant Pathology Department- Research and Development

 Basic Concepts

 Diagnosis is the first step in addressing the challenge of

plant diseases.
 Symptoms alone are usually not sufficient for accurate
diagnosis because the same symptoms can be caused by
different pathogens, or disease causing agents.
 Accurate diagnosis requires consideration of the basic
functions of the plant that are affected by the disease.

Plant Pathology Department- Research and Development

 The Disease Triangle

SUSCEPTIBLE
HOST
Disease

 Plant disease results when the host, pathogen, and an

environment favoring interaction occur together
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 Symptoms and Signs

 Symptoms are the expression of disease by a

plant as a response to the activities of the
pathogen.
 Symptoms may be localized or systemic.
 Signs are structures or products of a pathogen on
or in diseased plant.
 Biotic pathogens often produce signs, which are
evidence of their presence and can aid in
diagnosis.
 There are NO signs of abiotic factors/agents.

Plant Pathology Department- Research and Development

Common Symptoms of Plant Disease
Localized (e.g. leaf spots, canker)

Plant Pathology Department- Research and Development

Common Symptoms of Plant Disease
Systemic (e.g. wilt, blight, stunting, mosaic)

Plant Pathology Department- Research and Development

 Common Signs of Plant Pathogens

sclerotia

mycelia and
spores

bacterial ooze

Plant Pathology Department- Research and Development

 Field Diagnosis

 Diagnosis begins in the field or backyard.

 Some diseases may be diagnosed by the presence of
visible symptoms or signs.
 Field diagnosis requires good observation skills.
 Not all plant problems are attributable to pathogens (fungi,
bacteria, viruses)- consider the possibility of insect
infestation/environmental stresses.

Plant Pathology Department- Research and Development

 Basic Steps

• Determine identity of the host plant

1 • Recognize healthy plant appearance
Know what from an abnormal appearance
is normal/
abnormal

healthy

infected
healthy
healthy infected infected

Plant Pathology Department- Research and Development

 Basic Steps

• Identify plant part(s) affected

2 • Identify characteristic symptoms
Check for • Look for signs of biotic agents
symptoms • Correlate signs and symptoms with known
and signs diseases

Plant Pathology Department- Research and Development

 Basic Steps

3 • Check distribution /patterns of infection –

incidence and severity
Observe • Check for host specificity and incidence of the
patterns of same disease in neighboring fields
infections

Typical infection pattern of soil-borne pathogens

Photo: APS Press

Plant Pathology Department- Research and Development

 Basic Steps
Incidence
 Percentage of the crop affected

100% incidence of NCLB <10% incidence of ToLCV

 >50% (high); <30% (low); 31-50% (moderate)

Plant Pathology Department- Research and Development

 Basic Steps
Severity
 Measure of impact on crops
 >50% plant area affected (severe)
 <25% plant area affected (mild)
 26-50% plant area affected (moderate)

Mild

Moderate

Severe

Plant Pathology Department- Research and Development

 Basic Steps
• Review cultural management practices, cropping
4 history
• Take note of environmental conditions –
Get data on temperature, humidity, rainfall, etc.
cultural mgt./ • Take note of similar disease incidence in
environment neighboring fields

Plant Pathology Department- Research and Development

 Basic Steps
• If the biotic or abiotic cause cannot be identified
5 directly – collect samples for further laboratory
analysis
Collect and • Follow some do’s and don’ts in sample collection
send • Take pictures of the typical symptoms
samples • Send samples and pictures to nearest laboratory

Plant Pathology Department- Research and Development

 Clues For Abiotic Diseases

 Symptoms appear suddenly.

 Symptoms on individual plants are fairly uniform.
 Symptoms develop on several parts of individual plants or
on many plants.
 Different plant species in the area have similar symptoms.
 Symptoms may follow a chemical application patterns in
lines or rows.
 Foliar lesions have a distinct line between healthy and
affected tissue.
 Biotic signs are not present (unless the affected plant has
been invaded by secondary pathogens or saprophytes)

Plant Pathology Department- Research and Development

 BIOTIC OR ABIOTIC?

Plant Pathology Department- Research and Development

 BIOTIC OR ABIOTIC?

Plant Pathology Department- Research and Development

 BIOTIC OR ABIOTIC?

Plant Pathology Department- Research and Development

 BIOTIC OR ABIOTIC?

Plant Pathology Department- Research and Development

 BIOTIC OR ABIOTIC?

Plant Pathology Department- Research and Development

 PLANT DISEASE DIAGNOSIS GUIDE
LEAF FRUIT

Yellowing Mosaic Spots Spots Rot Deformed/

Mottled
Scabby/
uniform Viral Fungal
raised
Blossom-
end
veinal Bacterial Sunken
Bacterial Viral
Abiotic Stress/
Calcium
Fungal
STEM def.

ROOT
Collar Blight Spots
Rot Rot Galls Pale
Fungal/

Fungal Bacterial WHOLE PLANT Fungal Nematode Bacterial

Wilt
Wilting w/ yellowing Wilting w/o yellowing Stunted

Systemic Localized Bacterial Bacterial Root galls No galls

ooze ooze Viral
present absent Nematode
Fungal/
Nematode Fungal
Bacterial (Sclerotium)
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Abiotic
Diagnostic Procedure/Techniques for
Fungal Diseases
Isolation of biotic causes

Molecular
(PCR)
Microscopy

Plant Pathology Department- Research and Development

Diagnostic Procedure/Techniques for
Bacterial Diseases

Isolation of biotic causes

Molecular

Microscopy
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Diagnostic Procedure/Techniques for
Viral Diseases
Bioassay (using indicator hosts)

Molecular
(PCR)

Serology
(ELISA)

Plant Pathology Department- Research and Development

 Diagnosis of Viral Diseases
 ELISA(Enzyme-linked Immunosorbent
Assay) was developed in the late
1970’s
 It is a tool that is widely used by
pathologists of all kinds to detect and
study plant viruses and other plant
pathogens and the diseases they
cause
 It involves the reaction of antigens
(proteins foreign to the body) and
antibodies (specific molecules
produced by mammals in response to
the presence of the foreign protein)
 Some advantages of ELISA are as
follows: the tests are extremely
sensitive, large number of samples
can be tested concurrently, and results
are quantitative

Plant Pathology Department- Research and Development

 How does ELISA work?
Enzyme-linked
Antibody
(Step 3) Enzyme

Colored products
(detected if
(Step 2) Antigen is present)

Antigen binds (if

present in the sample)

Capture Antibody Substrate

(Step 4)
Microplate well (Step 1)

Plant Pathology Department- Research and Development

 ELISA Procedure

 Virus particles are

extracted from infected
leaf samples by grinding
in phosphate buffer

 A healthy leaf should be

included to serve as negative
control (healthy)

Plant Pathology Department- Research and Development

 ELISA Procedure
 The sap of each sample is carefully
loaded in individual wells of the
ELISA plate pre-coated with specific
antibody raised against the virus
 Washing step follows and the
enzyme-conjugated antibody is
added
 After an incubation period of 2-3hrs,
another washing step follows and the
substrate for the enzyme is added
 Reaction can be observed (10 min to
1 hr) as seen through the yellow
color that will appear in wells
containing the sap of leaf samples
where the virus was detected

Plant Pathology Department- Research and Development

 ELISA Procedure

 The reaction is measured

quantitatively using the ELISA
plate reader machine

 The results are shown in the

print-out. Positive result means
that the Abs reading/value of the
sample tested is 2X or higher
than the value of healthy control
e.g., PRSV-infected leaf = 1.0
healthy leaf= 0.10
The higher the value, the higher
the concentration of the virus in
the sample being tested

Plant Pathology Department- Research and Development

 DOT BLOT-ELISA

Plant Pathology Department- Research and Development

References:
 Gergerich, R.C., and V. V. Dolja. 2006. Introduction to Plant Viruses, the Invisible
Foe. The Plant Health Instructor. DOI: 10.1094/PHI-I-2006-0414-
01(http://www.apsnet.org/edcenter/intropp/PathogenGroups/Pages/PlantViruses.
aspx)
 Riley, M.B., M.R. Williamson, and O. Maloy. 2002. Plant disease diagnosis. The
Plant Health Instructor. DOI: 10.1094/PHI-I-2002-1021-
01(http://www.apsnet.org/edcenter/intropp/topics/Pages/PlantDiseaseDiagnosis.
aspx)
 Schumann, Gail L and Cleora D’Arcy. 2010. Essential Plant Pathology. 2nd ed.
2010. APS Press, USA.
 Windham Mark T. and Alan S. Windham. 2004. Plant Pathology Concepts and
Laboratory Exercises. CRC Press, USA.

Plant Pathology Department- Research and Development