Diagnosing Plant Problems: Plant Diseases National Plant Diagnostic Network First Detector Training...

National Plant Diagnostic NetworkNational Plant Diagnostic Network

Diagnosing Plant Problems: Plant

DiseasesNational Plant Diagnostic Network

First Detector Training

Williamson, Riley, and Maloy. December 2006.Williamson, Riley, and Maloy. December 2006.Update, August 2008.Update, August 2008.NPDN Publication No. 0013NPDN Publication No. 0013

MODULE 4 – Diagnosing plant problems: Plant Diseases

Adapted from “Plant Disease Diagnosis” by M. B. Riley, M. R. Williamson and O. Maloy.

In: The Plant Health Instructor. DOI:10.1094/PHI-I-2002-1021-01

Used by permission of the American Phytopathological Society


Plant Problem Diagnosis

Diagnosis –process used for the identification of problems.

Tools:– Plant sciences, especially plant

pathology and entomology

– Arts of investigation and detective work


Plant Problem Diagnosis: Preliminary Considerations

Know Normal Appearance of Plant Proper plant identification is critical.

Scientific vs. common names Variety or cultivar

Recognize healthy plant appearance. What’s normal for specific plant? Compare with healthy plants of same genus,

species and cultivar.


Plant Problem Diagnosis Steps

1. Consider the Possible Agents: Abiotic – non-living agent

Damage from chemicals, weather, mechanical

Nutritional problems

Biotic – living agent Pathogens - parasitic microorganisms that

cause diseases Pests – insects or mammals feeding on or

damaging plants



2. Consult literature resources for possible diseases and disorders.

Indexes listing hosts and their pathogens

Websites providing information Books with background info and

host/pathogen lists Compendia for information on specific

crops



3. Investigate symptom progression. Biotic disease – symptoms progress

and nearby plants become infected. Abiotic disease – generally a lack of

symptom progression. Does not spread. Exception – nutritional symptoms

progress slowly.



4. Ask questions When was the problem noticed? Was the damage sudden or gradual? How old are affected plants? Percentage of plants affected?

What is the degree of injury?



5. Observe patterns. Determine prevalence of problem

Large area/All plants – generally abiotic

Scattered, localized – generally biotic Check for distribution of symptoms

Uniform – generally abiotic Random – generally biotic


Observation of Patterns

Scattered symptom distribution

Random pattern of damage

Loblolly pine killed by southern pine beetle

Photo: R. Billings, Texas Forest Service, Photo: R. Billings, Texas Forest Service, http://www.forestryimages.orghttp://www.forestryimages.org


Observation of Patterns

Symptoms distributed in a large area. Damage pattern is uniform.

Cotton field with chemical damage

Used by permission of M. Williamson



6. Review cultural practices. Proper planting technique Fertilizer and pesticide application Irrigation frequency

7. Review environmental conditions. Temperature extremes Drought or excess rain Soil type and conditions

8. Check host specificity – many hosts may mean an abiotic problem



9. Check for symptoms and signs. Symptoms - plant reactions or

alterations of a plant’s appearance due to a disease or disorder

Signs - actual pathogen, parts or by-products seen on a diseased host plant


Symptoms

Identify and classify symptoms Underdevelopment Overdevelopment Necrosis or death Alteration of normal appearance Wilting

Identify plant parts affected May need to identify source of symptoms

within plant system


SymptomsSymptoms

Underdevelopment Underdevelopment Stunting of plants, Stunting of plants,

leaves leaves Shortened Shortened

internodesinternodes Inadequate Inadequate

chlorophyll chlorophyll productionproduction

Caused by many Caused by many types of pathogenstypes of pathogens

Photo: J. Dunez, www.forestryimages.org

Chrysanthemum stunt viroid infection of chrysanthemum showing stunting and earlier blooming of affected plants; healthy plants in middle.


Symptoms - Overdevelopment

Galls form on all plant parts, caused by many pathogens.

Oak leaf blister, caused by Taphrina caerulescens.

Gall on stem of eastern redcedar caused by Agrobacterium tumefaciens.

Overgrowth of leaf tissue causes thickening and distortion.

Photo: Edward L. Barnard, Florida Department of Agriculture and Consumer Services, www.ipmimages.org

Photo: Joseph O'Brien, USDA Forest Service, www.ipmimages.org


Symptoms

Over-development - more biotic causes Root knot nematodes - root galls Callus formation around cankers Tissue proliferation -some downy mildews

and phytoplasmas Some insects and mites also cause galls.


Symptoms

Over-development Abiotic causes Oedema Fasciation can be

physiological or due to biotic causes.

Some tree burlsOedema of camellia, a physiological disorder

Used by permission of M. Willliamson


Symptoms and Signs

Tissue Necrosis Fungal leaf spots Usually round, not vein-

limited Elongated on narrow

leaves or stems May have alternating

zones of light and dark tissue

Sporulation or mycelia may be evident

Target spot of tobacco, caused by Thanatephorus cucumeris..

Photo: Paul Bertrand, University of Georgia, www.ipmimages.org


Symptoms and Signs

Tissue Necrosis

Bacterial leaf spots Often dark and water-

soaked Often vein-limited,

giving angular shape Bacterial “flow”

observed under microscope

Bacterial leaf spot of sweet pepper caused by Xanthomonas campestris pv. vesicatoria

Photo: Volcani Center Archives, Agricultural Research Organization, www.ipmimages.org


Symptoms and Signs

Tissue Necrosis

Fruit rots Firm or soft and

watery Colors vary Fungal sporulation

may be present Fungal and bacterial

causesBrown rot of peach, caused by Monilinia fruticola

Photo: Clemson University - USDA Cooperative Extension Slide Series, , www.ipmimages.org


Symptoms

Tissue Necrosis

Cankers = localized necrotic lesions Sunken or swollen or

both Mainly caused by

fungi and bacteria Mechanical injury

can causeSugar maple canker caused by Nectria spp.

Photo: Joseph O'Brien, USDA Forest Service, www.ipmimages.org


SymptomsSymptoms

Tissue NecrosisTissue Necrosis

Blight = rapid death Blight = rapid death or dieback.or dieback. Also from coalescing Also from coalescing

leaf spots, e.g. early leaf spots, e.g. early blight of tomato blight of tomato

Mainly fungal and Mainly fungal and bacterial causesbacterial causes

Fire blight caused by Fire blight caused by Erwinia amylovoraErwinia amylovora

Photo: Robert L. Anderson, USDA Forest Service, www.ipmimages.org


SymptomsSymptoms


Dieback - many causesDieback - many causes Girdling cankers. Follow Girdling cankers. Follow

dieback to detectdieback to detect Dieback pathogens w/out Dieback pathogens w/out

cankerscankers Root problemsRoot problems Mechanical, chemical or Mechanical, chemical or

cold damagecold damage

Pitch canker of Virginia pine, caused Pitch canker of Virginia pine, caused by by Fusarium subglutinansFusarium subglutinans

Photo: Edward L. Barnard, Florida Department of Agriculture and Consumer Services, www.ipmimages.org


SymptomsSymptoms

Tissue NecrosisTissue NecrosisRoot rotsRoot rots

Root lesionsRoot lesions Darkening and Darkening and

softening of rootssoftening of roots Sloughing off of outer Sloughing off of outer

tissuestissues Yellowing of foliage Yellowing of foliage

and stunting of plantsand stunting of plants Fungal and bacterial Fungal and bacterial

causescauses

Root rot of boxwood, caused by Root rot of boxwood, caused by Pythium Pythium sp.sp.



SymptomsSymptoms


Damping offDamping off Rapid death of Rapid death of

seedlings and cuttingsseedlings and cuttings Stem infected at soil Stem infected at soil

line, seedling topplesline, seedling topples Spreads rapidly under Spreads rapidly under

crowded conditionscrowded conditions Mainly fungal agentsMainly fungal agents

Damping off of tobacco, caused by Damping off of tobacco, caused by Rhizoctonia Rhizoctonia spp. and spp. and Pythium Pythium spp. spp.

Photo: Clemson University - USDA Cooperative Extension Slide Series, www.ipmimages.org


SymptomsSymptoms

WiltingWilting

Biotic factors Biotic factors Root, crown or stem rotsRoot, crown or stem rots Vascular wiltsVascular wilts Root crown or stem Root crown or stem

damage from insects or damage from insects or animalsanimals

Mainly fungal and Mainly fungal and bacterial causesbacterial causes

Used by permission of M.WilliamsonUsed by permission of M.Williamson

Stem wilt of Stem wilt of ExacumExacum from INSV from INSV infectioninfection


SymptomsSymptoms

WiltingWiltingAbiotic factorsAbiotic factors

Damage from Damage from weather extremesweather extremes

Dry or flooded soilDry or flooded soil Mechanical damage Mechanical damage

to roots, crown or to roots, crown or stemstemCold damage on azalea Cold damage on azalea

stem. Shoots wilt above stem. Shoots wilt above damage.damage.

Used by permission of M. WilliamsonUsed by permission of M. Williamson


SymptomsSymptoms

Alteration of normal Alteration of normal appearanceappearance

MosaicMosaic Irregular patches of Irregular patches of

discolored tissue.discolored tissue. Often with distortionOften with distortion Viruses mainlyViruses mainly

Mosaic symptoms on cucumber due to Mosaic symptoms on cucumber due to a viral infectiona viral infection

Photo: David B. Langston, University of Georgia, www.ipmimages.org


SymptomsSymptoms


Ringspot Ringspot On leaves and fruits.On leaves and fruits. Distinct ring shaped Distinct ring shaped

lesions, often in lesions, often in concentric zonesconcentric zones

VirusesVirusesTomato spotted wilt virus (TSWV) on geranium leaf

Photo: Dr. Backhaus, Biologische Bundesanstalt für Land- und Forstwirtschaft, www.ipmimages.org


SymptomsSymptoms


Abiotic causesAbiotic causes Chemical damageChemical damage Nutritional deficienciesNutritional deficiencies

Cotton damage by the herbicide 2,4-D



SymptomsSymptoms


YellowingYellowing Often nutritional, Often nutritional,

cultural or cultural or environmentalenvironmental

Root malfunction – Root malfunction – biotic or abioticbiotic or abiotic

Nematode infestationNematode infestation ““Yellows” phytoplasma Yellows” phytoplasma

diseasesdiseasesBurford hollies yellowing due Burford hollies yellowing due to root malfunction. Soil is to root malfunction. Soil is poorly drained.poorly drained.



SymptomsSymptoms

Be aware of symptom variability.Be aware of symptom variability.

• May have more than one problem.May have more than one problem.• More than one pathogen may be involved.More than one pathogen may be involved.• Pathogens have varying levels of virulence.Pathogens have varying levels of virulence.• Environmental conditions can affect symptom Environmental conditions can affect symptom

expression.expression.• Host genetics and physiology can effect symptom Host genetics and physiology can effect symptom

expression.expression.


Symptom variability Symptom variability due to multiple pathogens due to multiple pathogens

Peach seedling on left Peach seedling on left infected with both infected with both Prune dwarf virusPrune dwarf virus (PDV) and (PDV) and Prunus Prunus necrotic ringspot virusnecrotic ringspot virus (PNRSV)(PNRSV)

Seedling in middle Seedling in middle infected with PDV infected with PDV alonealone

Seedling on right Seedling on right infected with PNRSV infected with PNRSV alonealone

Peach seedlings infected by single or multiple viruses show symptom variability.

Used by permission of S. Scott


SignsSigns

Fungal signsFungal signs Evidence of Evidence of

pathogen on tissue.pathogen on tissue. Spores, mycelia or Spores, mycelia or

fruiting bodies.fruiting bodies. Use hand lens or Use hand lens or

knife for field knife for field detection. detection.

White mycelia of White mycelia of ArmillariaArmillaria sp. on sp. on tree trunk affected by Armillaria tree trunk affected by Armillaria root and stem rot.root and stem rot.

Photo: USDA Forest Service Archives, USDA Forest Service, www.ipmimages.org


SignsSigns

Fungal signsFungal signsFruiting bodiesFruiting bodies Shape of fruiting Shape of fruiting

body aids in fungal body aids in fungal identification.identification.

Tissue location may Tissue location may help differentiate help differentiate between species.between species.

White rust of chrysanthemum, fruiting White rust of chrysanthemum, fruiting bodies of bodies of Puccinia horianaPuccinia horiana

Photo: Central Science Laboratory, Harpenden Archives, British Crown, www.ipmimages.org


SignsSigns

Fungal signsFungal signs• Powdery mildew Powdery mildew

fungi form mycelia fungi form mycelia and spores on tissue and spores on tissue surface.surface.

• Powdery material Powdery material rubs off.rubs off.

• Leaves often Leaves often distorted, discolored distorted, discolored

• Dark, round fruiting Dark, round fruiting bodies form in fall.bodies form in fall.

Oidium Oidium sp. causing powdery sp. causing powdery mildew on mildew on Viburnum Viburnum suspensumsuspensum..



Symptoms and SignsSymptoms and Signs

Downy mildews often Downy mildews often produce angular leaf produce angular leaf spots.spots.

Downy mildews Downy mildews develop sporulation develop sporulation on leaf undersides on leaf undersides only.only.

Downy mildew of veronica, caused by Downy mildew of veronica, caused by Peronospora sordidaPeronospora sordida


Downy mildew of Buddleia Downy mildew of Buddleia caused by caused by Peronospora harotiiPeronospora harotii..




SignsSigns

Bacterial signsBacterial signs ““Streaming” from freshly Streaming” from freshly

cut stem in water.cut stem in water. ““Stringing” from cut Stringing” from cut

stems pushed together, stems pushed together, then pulled apart.then pulled apart.

Bacteria “flow” observed Bacteria “flow” observed from tissue mount on from tissue mount on compound microscope. compound microscope.

Bacterial ooze can be Bacterial ooze can be observed on-site with observed on-site with some diseases.some diseases.

Bacterial streaming from tomato infected by Ralstonia solanacearum.



Plant Problem Diagnosis StepsPlant Problem Diagnosis Steps

12. Laboratory Examination and Testing12. Laboratory Examination and Testing

Dissecting microscopeDissecting microscope Compound microscopeCompound microscope Electron microscopeElectron microscope Moist chamber incubationMoist chamber incubation CulturingCulturing Additional tests for biotic agentsAdditional tests for biotic agents Tests for abiotic agentsTests for abiotic agents


Laboratory Examination Laboratory Examination Examination using dissecting Examination using dissecting

microscopemicroscope• Observe sporulation Observe sporulation

that’s invisible to that’s invisible to naked eye.naked eye.

• Closer view of Closer view of structures seen by structures seen by naked eyenaked eye

• Nematologist can ID Nematologist can ID nematodes to nematodes to genus.genus.



Laboratory ExaminationLaboratory ExaminationExamination using microscope Examination using microscope

of higher magnificationof higher magnification ID fungi to genus or to ID fungi to genus or to

species.species. View bacterial flow; View bacterial flow;

observe morphology at observe morphology at highest magnification.highest magnification.

View virus inclusion View virus inclusion bodies.bodies.

Using electron Using electron microscope, view microscope, view viruses, phytoplasmasviruses, phytoplasmas


Rust spores, Rust spores, Puccinia Puccinia sp. sp.


Laboratory testsLaboratory tests

Moist IncubationMoist Incubation Goal – to induce Goal – to induce

sporulationsporulation Important for Important for

obligate fungal obligate fungal pathogenspathogens

Avoid overly moist Avoid overly moist conditions.conditions.

Can surface sterilizeCan surface sterilize


Moist chamberMoist chamber



Fungal isolation Fungal isolation Surface sterilize.Surface sterilize. Use margin of Use margin of

diseased area.diseased area. Nutritive agar media, Nutritive agar media,

can be selectivecan be selective Diagnostician can Diagnostician can

often ID to genus often ID to genus from appearance of from appearance of cultureculture


Fungal culture.Fungal culture.


Isolate identificationIsolate identification

Fungal identification – Other methodsFungal identification – Other methods Examination and comparison with Examination and comparison with

drawings in literaturedrawings in literature Utilizing keys in literatureUtilizing keys in literature Literature sources for host diseases Literature sources for host diseases

and disordersand disorders Serological, molecular, other testsSerological, molecular, other tests



Bacterial isolationsBacterial isolations Grind tissue in Grind tissue in

sterile water.sterile water. Streak suspension Streak suspension

on bacterial medium.on bacterial medium. Can use selective Can use selective

mediamedia Transfer single, Transfer single,

isolated colonyisolated colony

Cultures of Ralstonia solanacearum

Used by permission of K. OngUsed by permission of K. Ong


Isolate identificationIsolate identification

Bacterial identificationBacterial identification Biochemical testsBiochemical tests Carbohydrate utilization (BIOLOG)Carbohydrate utilization (BIOLOG) Fatty acid methyl ester analysis Fatty acid methyl ester analysis

(FAME)(FAME) Molecular methodsMolecular methods Polymerase chain reaction (PCR)Polymerase chain reaction (PCR) Serological tests, e.g. ELISASerological tests, e.g. ELISA


Identification of other pathogensIdentification of other pathogens

Non-culturable pathogensNon-culturable pathogens

Viruses, viroidsViruses, viroids PhytoplasmasPhytoplasmas Fastidious bacteriaFastidious bacteria


Identification of non-culturable Identification of non-culturable pathogenspathogens

MethodsMethods Serological tests, e.g. ELISA Serological tests, e.g. ELISA Electron microscopyElectron microscopy Staining for virus-induced inclusion Staining for virus-induced inclusion

bodiesbodies Molecular tests, e.g. PCRMolecular tests, e.g. PCR Enzyme activity testingEnzyme activity testing Host range testingHost range testing


Identification of other pathogensIdentification of other pathogensELISA testsELISA tests

Multi-well test for Multi-well test for multiple samplesmultiple samples

Individual ELISA kitIndividual ELISA kit

UsedUsed by permission of M. Williamsonby permission of M. Williamson


Identification of NematodesIdentification of Nematodes

Extraction from Soil and Plant MaterialExtraction from Soil and Plant Material Soil extraction using centrifugal flotationSoil extraction using centrifugal flotation Baerman funnel techniques for soil and Baerman funnel techniques for soil and

tissuetissue Mist chamber extraction for nematodes in Mist chamber extraction for nematodes in

root or stem tissueroot or stem tissue

Direct examination of nematodes in Direct examination of nematodes in tissuetissue..



Diagnostic tests for abiotic causesDiagnostic tests for abiotic causesSoil, waterSoil, water

pH testingpH testingNutrient analysisNutrient analysisSoluble salts analysis Soluble salts analysis Analysis for chemicalsAnalysis for chemicals

Plant tissue testsPlant tissue testsNutrient analysisNutrient analysisAnalysis for chemicalsAnalysis for chemicals


Plant Disease Diagnosis StepsPlant Disease Diagnosis Steps

13) Investigate pathogenicity.13) Investigate pathogenicity.

Is identified organism a known pathogenIs identified organism a known pathogen

- of host?- of host?

- of hosts in same genus?- of hosts in same genus?

Consult literature to determineConsult literature to determineIf not a host, Kochs Postulates may be next stepIf not a host, Kochs Postulates may be next step A lengthy process, rarely used in routine A lengthy process, rarely used in routine

diagnosesdiagnoses Problem arises when organism cannot be culturedProblem arises when organism cannot be cultured


Koch’s PostulatesKoch’s Postulates

Robert Koch – German physician in late Robert Koch – German physician in late 1800s1800s

Proved that a bacterium caused anthrax Proved that a bacterium caused anthrax in cattlein cattle

Developed steps proving the bacterium Developed steps proving the bacterium was causalwas causal

Same steps used to prove pathogenicity Same steps used to prove pathogenicity for plant disease organismsfor plant disease organisms


Koch’s PostulatesKoch’s Postulates

1.1. Note constant association of organism Note constant association of organism with diseased plants and consistent, with diseased plants and consistent, observable symptoms.observable symptoms.

2.2. Isolate and characterize organism in Isolate and characterize organism in pure culture.pure culture.

3.3. Inoculate healthy plant with organism, Inoculate healthy plant with organism, and observe the same disease and and observe the same disease and symptoms.symptoms.

4.4. Re-isolate same organism.Re-isolate same organism.


Plant Disease Diagnosis StepsPlant Disease Diagnosis Steps

14. Final Diagnosis14. Final Diagnosis

All information compiled and analyzedAll information compiled and analyzedOrganism identified, found to be pathogen of Organism identified, found to be pathogen of hosthost

ORORLab tests, other information reveal an abiotic Lab tests, other information reveal an abiotic causecause

15. Develop control recommendations and 15. Develop control recommendations and present to grower. present to grower.


• Meg Williamson, Clemson Meg Williamson, Clemson UniversityUniversity

• Melissa Riley, Clemson UniversityMelissa Riley, Clemson University

• Otis Maloy, Washington State Otis Maloy, Washington State UniversityUniversity

AuthorsAuthors


• This publication can be used for non-profit, This publication can be used for non-profit, educational use only purposes. educational use only purposes. Photographers retain copyright to Photographers retain copyright to photographs or other images contained in photographs or other images contained in this publication as cited. This material was this publication as cited. This material was developed as a core training module for developed as a core training module for NPDN First Detector Training. The authors NPDN First Detector Training. The authors and website should be properly cited. and website should be properly cited. Images or photographs should also be Images or photographs should also be properly cited and credited to the original properly cited and credited to the original source.source.

• Publication Number: 0013Publication Number: 0013• Publication Date: December 2006Publication Date: December 2006

Diagnosing Plant Problems: Plant Diseases National Plant Diagnostic Network First Detector Training...

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