2. INTRODUCTION
Most of the parasitic infections cannot be conclusively diagnosed.
On the basis of clinical features and physical examination, laboratory
diagnosis depends upon:
- Microscopy
- Culture
- Serological test
- Skin test
- Molecular method
- Animal inoculation
- Xenodiagnosis
- Imaging
- Hematology
3. SPECIMENS
An appropriate clinical
specimen should be collected
for definitive diagnosis of
parasitic infections.
• Stool
• Blood
• Urine
• Sputum
• Cerebrospinal fluid (CSF)
• Tissue and aspirates
• Genital specimens
This Photo by Unknown Author is licensed under CC BY-NC
4. DEFINITION OF STOOL
• Human feces is called as Stool
• From latin word Faex meanin residue
• It is the waste residue of indigestive materials of an animals
digestive tract expelled through the anus during defecation
• Scatology or Caprology is the study of feces
5. COMPOSITION OF STOOL
• ¾ Water,
• ¼ Solid Undigested and Unabsorbed food Intestinal secretions,
• Mucous Bile pigments
• Salts Bacteria
• Inorganic material Epithelial cells, Leukocytes
6. STOOL EXAMINATION
HOW TO COLLECT A STOOL SAMPLE?
• Because of the fragile nature of many intestinal parasites and the need to maintain their
morphology for accurate identification, reliable microscopic diagnosis can’t be made unless
the stool is collected properly.
• 10 g (approx.) of fresh and uncontaminated faeces is taken in a clean plastic container.
• Container should be free from antiseptics and disinfectants.
• Label all samples clearly with Patient’s name, reference no, date and time of collection.
• All samples should be accompanied by a requisition form from the physician giving relevant
clinical details and recent travel history.
• Samples and forms of patients with a confirmed or suspected diagnosis of certain infectious
diseases like AIDS and Hepatitis should be clearly labeled as “Risk of infection” or “Biohazard”
• Most viable parasites are susceptible to desiccation or temp variation and hence care should
be taken.
• When there is a time lapse (>4 days) between collection and observation, a preservative can
be added to retain the morphology as near to the original as possible.
• Any whole worms or segments passed should be kept in a separate container.
7. COLOUR OF STOOL
• Human fecal matter is normally yellowish brown in colour which
results from a combination of bile and bilirubin.
• VARIATIONS
• Bright Red/Maroon Tan/Clay Blood streak White Yellow Pale
greasy Green Black Blue
9. MICROSCOPY IN
PARASITOLOGY
• The microscope is the parasitologist’s main
tool.
• Should be binocular & Most suitable
objectives are- 10x, 40x & 100x.
• The microscope must be covered and
immersion oil should be removed from the
lens- with xylene or ether when not in use.
• Eyepiece micrometer: measuring the size of
suspected parasites in faeces is helpful for
identification.
10. M AT E R I A L S
• Microscope slides
• Cover slips
• Sodium chloride solution
• Lugol’s Iodine Solution
• Wooden applicator
• Fresh stool
• Gloves
11. VARIOUS STAINING METHODS
• Giemsa (Malaria) Stain
• Trichrome Stain
• Methylene Blue Phosphate Pre-Stain
• Modified Acid-Fast Staining
• Chromotrope Staining
• Calcofluor White Staining
12. WET MOUNT
• Simplest and easiest technique for examination of faeces, and
this method should be performed in all laboratories in the
peripheral level.
• Can be prepared directly from faecal material or concentrated
specimens.
• Basic types are:
• Saline mount
• Iodine mount
• Buffered Methylene Blue mount
13. PREPARING A FAECAL
SPECIMEN
• Place a drop of saline in the
center of left half of the
slide.
• If the presence of amoebic
trophozoites is suspected
warm saline (37c) should be
used.
• Place a drop of iodine in the
center of right half of the
slide.
14. PREPARING A WET
FILM
• With an applicator stick
(match or toothpick), pick
up a small portion of the
specimen (size of a match
head) and mix the drop of
saline.
• Eosin 1% aqueous solution,
can be used for staining
wet films.
15. SALINE WET MOUNT
• Used for initial microscopic examination of
stools.
• Employed primarily to demonstrate worms’s
eggs, larvae, cysts & trophozoites.
• Can also reveal the presence of RBC’s &
WBC’s.
• Particularly useful for detecting live motile
trophozoites of E.histolytica, Balantidium coli
and Giardia lamblia.
• Rhabditiform larvae Of Strongyloides
stercoralis are detected in freshly passed
stool.
16. IODINE WET MOUNT
• Used mainly to stain glycogen and the
nuclei of cysts, if present.
• Cysts can be specifically identified in
this mount.
• Protozoan cyst stained with iodine
show yellow-gold cytoplasm, brown
glycogen material and pale refractile
nuclei.
17. BUFFERED METHYLENE BLUE MOUNT
• Should be prepared each time amoebic trophozoites are seen in a
saline wet mount or even when its presence is suspected.
• Stains only trophozoites of amoeba
• It does not stain amoebic cyst or trophozoites and cyst of
falgellates.
• Nucleus and the inclusions such as RBC or yeast cells stain dark
blue.
• Cytoplasm stains light blue.
18. EXAMINATION
• Put the slide with the mount on the
microscope stage and focus under 10x.
• Regulate the light in the microscope field
with the substage diaphragm.
• Examine the entire cover slip area with 10x,
focus the objective on top left corner & move
the slide systematically backwards &
forwards, up & down.
• When organisms are seen switch to the high-
dry objective and increase the light by
opening the substage diaphragm to observe
the detailed morphology.
• This is a systematic examination.
• If mounts aren’t examined this way, any
parasites present will usually be found.
19. NO R M A L VA L U E S
• Undigested food materials – None to small amount
• Starch – None
• Eggs, Cysts, Parasitic fragments – None
• Yeasts – None
• Leukocytes – None
