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Title: DIAGNOSTIC PARASITOLOGY

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Title: DIAGNOSTIC PARASITOLOGY

1
DIAGNOSTIC PARASITOLOGY

THURSDAY 4-6 PM
SPRING 2007

2
WHY PERFORM THIS TYPE OF TESTING

1. Travel
2. Control Issues
3. Epidemiologic Considerations
4. Compromised Patients
5. Approach to Therapy

3
WHO SHOULD PERFORM TESTING

1. Laboratory Personnel
a. CLIA 88-High- complexity procedures
b. Few procedures are automated
c. Organism identification relies on
morphologic characteristics
2. Nonlaboratory Personnel
a. CLIA 88-Waived tests
b. Wet mount examinations
c. No over-the-counter testing methods

4
Where should testing be performed

1. Inpatient Setting
a. Hospital or offsite location
b. Stat procedures
Blood smears for malaria
CSF for free-living amebae
2. Outpatient or Referral Setting
a. Routine testing
b. Batch tested

5
Where should testing be performed

3. Decentralized Physician Office Labs
Over-the-Counter (Home Care)
a. Usually not considered appropriate
for diagnostic parasitology
tesing

6
What Factors Should Precipitate Testing

1. Travel History
2. Immune Status of the Patient
3. Clinical Symptoms
4. Documented Previous Infection
5. Contact with Infected Individuals
6. Screening outbreak situations
7. Occupational Screening food handlers
8. Therapeutic Failure retested after therapy

7
What Testing Should Be Performed

1. Screening Test
a. Wide range in both sensitivity and
specificity
b. OP Exam
c. Monoclonal antibody based test
d. Tests on on request

8
What Testing Should Be Performed

2. Routine Methods
a. May be screening methods OP
b. Blood smears for malaria
c. Pinworm tapes
d. Occult blood tests
e. Specimens from other body sites
(urine sputum duodenal aspirates)

9
What Testing Should Be Performed

3. Special Testing
a. Parasite culture
b. Serologic Testing for parasitic
disease
4. Other (Nonmicrobiological) Testing
a. Routine urinalysis
b. Hematology CBC
c. Chemistry profiles

10
PARASITE CLASSIFICATION

SIX MAJOR DIVISIONS
1. PROTOZOA (INTESTINAL)
A. AMEBAE
1. SINGLE CELLED
2. PSEUDOPODS (MOTILITY)
3. TROPHOZOITE STAGE
4. CYST STAGE
5. FECAL-ORAL TRANSMISSION
6. MOUTH-MOUTH (ENTAMOEBA
GINGIVALIS)
7. ENTAMOEBA HISTOLYTICA
SIGNIFICANT ORGANISM

11
PARASITE CLASSIFICATION

PROTOZOA
2. FLAGELLATES
A. FLAGELLA
B. FECAL-ORAL TRANSMISSION
C. TROPHOZOITE CYST
STAGES
D. DIENTAMOEBA FRAGILIS
TRICHOMONAS (TROPHOZOITE)
E. GIARDIA LAMBLIA MOST COMMON
PATHOGEN

12
PARASITE CLASSIFICATION

PROTOZOA
3. CILIATES
A. SINGLE-CELLED
B. MOVE BY CILIA
C. FECAL-ORAL TRANSMISSION
D. BALANTIDIUM COLI ONLY
HUMAN PATHOGEN
E. TROPHOZOITE CYST STAGES

13
PARASITE CLASSIFICATION

PROTOZOA
4. COCCIDIA
A. INGESTION OF MEAT
B. FECAL-ORAL TRANSMISSION
VIA CONTAMINATED FOOD
AND/OR WATER
C. INFECTIVE STAGE OOCYST
D. CRYPTOSPORIDIUM CYCLOSPORA
ISOPORA SARCOCYSTIS

14
PARASITE CLASSIFICATION

PROTOZOA
5. MICROSPORIDIA
A. RANGE 1 TO 2.5 UM
B. INFECTIVE FORM SPORE
C. MODIFIED TRICHROME
STAINS
D. INFECTIONS THRU INGESTION
INHALATION OR DIRECT
INOCULATION FROM ENVIRONMENT

15
PARASITE CLASSIFICATION

PROTOZOA (OTHER BODY SITES)
1. AMEBAE
A. PATHOGENIC FREE LIVING
ORGANISMS
ASSOCIATED WITH WARM
FRESH-
WATER ENVIRONMENTS.
B. FOUND IN CNS EYES AND OTHER
BODY
SITES.
C. NAEGLERIA ACANTHAMOEBA
BALAMUTHIA

16
PARASITE CLASSIFICATION

PROTOZOA (OTHER BODY SITES)
2. FLAGELLATES
A. TRICHOMONAS VAGINALIS
IN THE GENITOURINARY
SYSTEM ACQUIRED BY
SEXUAL TRANSMISSION
B. TRICHOMONAS TENAX
FOUND IN MOUTH
NONPATHOGENIC

17
PARASITE CLASSIFICATION

3. PROTOZOA (OTHER BODY SITES)
1. COCCIDIA
A. IMPORTANT IN THE
COMPROMISED PATIENT
B. DISSEMINATE FROM THE
INTESTINAL TRACT TO
OTHER BODY SITES
C. CRYPTOSPORIDIUM

18
PARASITE CLASSIFICATION

PROTOZOA (OTHER BODY SITES)
4. MICROSPORIDIA
A. MICROSPORIDIA (1 TO 2.5 UM)
B. INTESTINE TO OTHER BODY
SITES
C. MODIFIED TRICHOME STAINS
TO DETECT

19
PARASITE CLASSIFICATION

PROTOZOA (BLOOD AND TISSUE)
1. SPOROZA
A. ALL ARE ARTHROPOD BORNE
B. PLASMODIUM (MOSQUITOES)
C. BABESIA (TICK BORNE)
D. EXAM BOTH THICK AND THIN BLOOD
FILMS

20
PARASITE CLASSIFICATION

FLAGELLATES
1. LEISHMANIAE
A. RECOVERY AND ID RELATED
TO BODY SITE.
B. AMASTIGOTES LIMITED TO THE
SITE OF THE LESION
C. CUTANEOUS MUCOCUTANEOUS
VISCERAL
D. SAND FLY

21
PARASITE CLASSIFICATION

FLAGELLATES
2. TRYPANOSOMES
A. ID TO THE SPECIES LEVEL
BASED ON GEOGRAPHIC
EXPOSURE HISTORY AND
CLINICAL SYMPTOMS
B. TRYPOMASTIGOTE FORM
C. AFRICAN SLEEPING SICKNESS

22
PARASITE CLASSIFICATION

NEMATODES (INTESTINAL)
1. ROUNDWORMS
A. ELONGATE-CYLINDRICAL
B. ACQUIRED BY INGESTION
OF EGGS OR PENETRATION
OF THE SKIN BY LARVAL FORMS
FROM THE SOIL

23
PARASITE CLASSIFICATION

NEMATODES (TISSUE)
1. ROUNDWORMS
A. RARELY SEEN IN THE U.S.
B. TRICHINELLA SPIRALIS
C. ACQUIRED BY INGESTION OF
OF INFECTIVE RAW OR
POORLY COOK MEAT (PORK
BEAR WALRUS)

24
PARASITE CLASSIFICATION

NEMATODES (BLOOD AND TISSUE)
1. FILARIAL WORMS
A. ARTHROPOD BORN
B. ID BY LARVAL WORMS
(MICROFILARIAE) IN BLOOD
OTHER BODY FLUIDS OR SKIN

25
PARASITE CLASSIFICATION

CESTODES (INTESTINAL)
1. TAPEWORMS
A. INGESTION OF LARVAL FORMS
IN POORLY COOKED OR RAW
MEATS OR FRESHWATER
FISH INFECTED BEETLES
B. TAPEWORM CONSISTS OF A
CHAIN OF EGG-PRODUCING UNITS
CALLED PROGLOTTIDS

26
PARASITE CLASSIFICATION

CESTODES (TISSUE)
1. TAPEWORMS
A. TISSUE INFECTION
B. TAENIA SOLIUM
C. ECHINOCOCCUS
GRANULOSUS

27
PARASITE CLASSIFICATION

TREMATODES (INTESTINAL)
1. FLATWORMS
A. FLAT WITH ORAL AND VENTRAL
SUCKERS
B. REQUIRE A FRESHWATER
SNAIL TO SERVE AS AN
INTERMEDIATE HOST
C. INFECTIONS ARE FOODBORNE
(FRESHWATER FISH MOLLUCKS
FRESHWATER PLANTS)

28
PARASITE CLASSIFICATION

TREMATODES (LIVER AND LUNG)
1. FLATWORMS
A. REQUIRE A FRESHWATER SNAIL
TO SERVE AS INTERMEDIATE
HOST
B. INFECTIONS ARE FOODBORNE
(FRESHWATER FISH CRAYFISH
CRABS OR PLANTS)
C. CLONORCHIS PARAGONIMUS EXAMPLES

29
PARASITE CLASSIFICATION

TREMATODES (BLOOD)
1. FLATWORMS
A. SEXES ARE SEPARATE
B. INFECTION IS ACQUIRED BY
SKIN PENETRATION BY THE
CERCARIAL FORMS THAT ARE
RELEASED FROM FRESHWATER
SNAILS.
C. WORMS RESIDE IN THE BLOOD VESSELS
OVER THE SMALL LARGE INTESTINE
OR
THE BLADDER.
D. SCHISTOSOMES

30
PARASITE CLASSIFICATION

PENTASTOMIDS
1. TONGUE WORMS

31
PARASITE CLASSIFICATION

ACANTHOCEPHALA
1. THORNY-HEAD WORMS

32
COLLECTION OPTIONS
33
1. SAFETY

A. HANDLE ALL FRESH SPECIMENS
CAREFULLY
B. USE UNIVERSAL PRECAUTIONS
GLOVES PROPER CONTAINERS
SAFETY CABINETS DISCARD
POLICIES

34
2. FRESH STOOL SPECIMENS

A. INTERFERING SUBSTANCES
SHOULD BE AVOIDED WHEN STOOL
SPECIMENS ARE COLLECTED
a. Collect before barium is used
b. Mineral oil bismuth antibotics
antimalarial agents.
c. Collection delayed 5 to 10 days

35
COLLECTION METHOD

1. CLEAN CONTAINERS OR FIXATIVE
VIALS.
2. CONTAMINATION WITH URINE OR
WATER SHOULD BE AVOIDED.
3. LABELED WITH PATIENT NAME TIME
AND DATE OF COLLECTION

36
NUMBER OF SPECIMENS

1. NORMAL EXAMINATION FOR STOOL
PARASITES BEFORE THERAPY
INCLUDE 3 SPECIMENS.
2. THREE SPECIMENS POOLED
CONTROVERSIAL
3. POSTTHERAPY SPECIMENS NOT
COLLECTED UNLESS PATIENT BECOMES
SYMPTOMATIC AGAIN.

37
COLLECTION TIMES

1. THREE SPECIMENS COLLECTED ON
SEPARATE DAYS.
2. WITHIN NO MORE THAN 10 DAYS.
3. EXCEPTION PATIENT WITH SEVERE
WATERY DIARRHEA ORGANISMS
MIGHT BE MISSED BECAUSE OF
TREMEDOUS DILUTION FACTOR RELATED
TO FLUID LOSS.

38
SPECIMEN TYPE STABILITY AND PRESERVATION

1. FRESH SPECIMENS MANDATORY
FOR RECOVERY OF MOTILE
TROPHOZOITES.
2. LIQUID STOOL SHOULD BE
EXAMINED OR PRESERVED WITHIN
30 MINUTES OF PASSAGE (TROPS)
3. SOFT STOOL EXAM OR PRESERVED
WITHIN 1 HOUR OF PASSAGE (TROPS
CYSTS) DIENTAMOEBA FRAGILIS TROPHOZOITES
4. FORMED STOOL SHOULD BE EXAMINED OR PRESERVED
WITHIN 24 HOUR OF PASSAGE.

39
PRESERVATION OF STOOL SPECIMENS

1. DELAY IN RECEIVING STOOL
SPECIMENS PLACE IN FIXATIVES
2. FORMALIN SODIUM ACETATE-
ACETIC ACID-FORMALIN (SAF)
SCHAUDINNS FLUID AND
POLYVINYL ALCOHOL (PVA)
3. KEEP IN MIND A PERMANENT STAINED
SMEAR IS MANDATORY.
4. MAY WANT TO PERFORM SCREENING METHODS
FLUORESCENT-ANTIBODY (FA) OR ENZYME
IMMUNOASSAY
(EIA). MAKE SURE THE FIXATIVE IS COMPATIBLE
WITH THE
KIT YOU ARE USING.
5. DISPOSAL REGULATIONS FOR COMPOUNDS CONTAINING
MERCURY ARE
BECOMING MORE STRICT

40
FOMALIN FIXATIVE

FORMALIN
1. ALL-PURPOSE FIXATIVE FOR
HELMINTH EGGS AND LARVAE
AND PROTOZOAN CYSTS.
2. 5 AND 10 CONCENTRATIONS
A. 5 FOR PRESERVATION OF
PROTOZOAN CYSTS
B. 10 FOR PRESERVATION OF
HELMINTH EGGS AND LARVAE
3.

41
FORMALIN

PROS
GOOD OVERALL FIXATIVE FOR STOOL CONCENTRATE
EASY TO PREPARE LONG SHELF LIFE
CONCENTRATED SEDIMENT CAN BE USED WITH THE NEW
IMMUNOASSAY METHODS

CONS
DOES NOT PRESERVE TROPHOZOITES WELL
DOES NOT ADEQUATELY PRESERVE ORGANISM MORPHOLOGY
FOR A GOOD
PERMANENT STAINED SMEAR

42
SODIUM ACETATE-ACETIC ACID-FORMALIN (SAF)

PROS
CAN BE USED FOR CONCENTRATION AND PERMANENT
STAINED SMEARS
CONTAINS NO MERCURY COMPOUNDS
EASY TO PREPARE LONG SHELF LIFE
CONCENTRATED SEDIMENT CAN BE USED WITH THE NEW
IMMUNOASSAY METHODS

CONS
POOR ADHESIVE PROPERTIES ALBUMIN-COATEDS
RECOMMENDED
PROTOZOAN MORPHOLOGY BETTER IF IRON HEMATOXYLIN
STAINS USED FOR PERMANENT STAINED SMEARS
(TRICHROME NOT AS GOOD.
MAY BE MORE DIFFICULT TO USE THAN PVA FIXATIVE
HOWEVER THIS IS REALLY NOT A LIMITING FACTOR.

43
SCHAUDINNS FLUID

PRO
FIXATIVE FOR SMEARS PREPARED FROM FRESH FECAL
SPECIMENS OR SAMPLES FROM THE INTESTINAL MUCOSAL
SURFACES
PROVIDES EXCELLENT PRESERVATION OF PROTOZOAN
TROPHOZOITES AND CYSTS

CONS
NOT GENERALLY RECOMMENDED FOR USE IN
CONCENTRATION PROCEDURES
CONTAINS MERCURIC CHLORIDE WHICH IS A DISPOSAL
PROBLEM
POOR ADHESIVE QUALITIES WITH LIQUID OR MUCOID
SPECIMENS

44
POLYVINYL ALCOHOL (PVA)

1. PVA IS A PLASTIC RESIN THAT IS
INCORPRORATED INTO
SCHAUDINNS FIXATIVE.
2. PVA POWDER SERVES AS AN
ADHESIVE FOR THE STOOL
MATERIAL WHEN THE STOOL-PVA
MIXTURE IS SPREAD ONTO THE GLASS
.
3. FIXATION IS STILL ACCOMPLISHED BY
SCHAUDINNS FLUID ITSELF.

45
POLYVINYL ALCOHOL (PVA)

PRO
CAN PREPARE PERMANENT STAINED SMEARS AND PERFORM
CONCENTRATION TECHNIQUES (LESS COMMON)
PROVIDES EXCELLENT PRESERVATION FO PROTOZOAN
TROPHOZOITES AND CYSTS
LONG SHELF LIFE IN TIGHTLY SEALED CONTAINERS AT
ROOM TEMPERATURE.
ALLOWS SPECIMENS TO BE SHIPPED TO LABORATORY FOR
SUBSEQUENT EXAMINATION.

CONS
TRICHURIS EGGS AND GIARDIA CYSTS ARE NOT
CONCENTRATED AS EASILY. STRONGYLOIDES LARVAL
MORPHOLOGY IS POOR.
ISOSPORA OOCYSTS MAY NOT BE VISIBLE IN
PVA-PRESERVED MATERIAL.
CONTAINS MERCURY COMPOUNDS (SCHAUDINNS FLUID)
MAY TURN WHITE AND GELATINOUS WHEN IT BEGINS TO
DEHYDRATE OR WHEN REFRIGERATED.
DIFFICULT TO PREPARE IN THE LAB.
SPECIMENS CONTAINING PVA CANNOT BE USED WITH THE
IMMUNOASSAY DETECTION KITS.

46
MODIFIED PVA

PRO
CAN BE USED FOR PERMANENT STAINED SMEARS AND
CONCENTRATION TECHNIQUES.
MANY WORKERS PERFER THE ZINC SUBSTITUTES OVER
THOSE PREPARED WITH COPPER SULFATE.
ZINC-BASED FIXATIVES APPEAR TO BE SOME OF THE
BETTER ALTERNTIVE.
DOES NOT CONTAIN MERCURY COMPOUNDS.

CONS
OVERALL PROTOZOAN MORPHOLOGY OF TROPS AND CYSTS
IS POOR WHEN PRESERVED IN THE COPPER
SULFATE-BASED FIXATIVE.
STAINING CHARACTERISTICS OF PROTOZOA NOT
CONSISTENT.
SMALL PROTOZOAN CYSTS (SUCH AS ENDOLIMAX NANA)
IDENTIFICATION MAY BE DIFFICULT.

47
SINGLE-VIAL COLLECTION SYSTEMS

PRO
CAN PREPARE PERMANENT STAINED SMEARS AND PERFORM
CONCENTRATION TECHNIQUES.
CAN PERFORM IMMUNOASSAY PROCEDURES.
DO NOT CONTAIN MERCURY COMPOUNDS.
UNLESS ORGANISM NUMBERS ARE SMALL ACCEPTABLE
ORGANISM RECOVERY AND ID IS POSSIBLE.

CONS
OVERALL PROTOZOAN MORPHOLOGY OF TROPHOZOITES AND
CYSTS IS NOT AS GOOD AS THAT WITH MERCURIC
CHLORIDE-BASED FIXATIVES.
STAINING CHARACTERISTICS OF PROTOZOA NOT
CONSISTENT.

48
PROCEDURE NOTES