Medical Parasitology Lab.

Concentration techniques

 The microscopic examination of feces is required for the

recognition and identification of intestinal parasites:
1. Direct Microscopy:
Advantages
Useful for the observation of motile protozoan
trophozoites.
Disadvantages
May not detect ova, cysts and larvae which are present
in scant numbers.

Raed Z. Ahmed, Medical Parasitology Lab.,2012

2. Concentration techniques :
Advantages
Maximizes the numbers of organisms detected which may
be too scanty to be seen by direct microscopy alone.
Worm eggs, larvae, and protozoan cysts may be recovered.
Disadvantages
Destroys trophozoite stages. Most concentration methods
destroy trophozoites stages.

The purpose of concentrating feces is to increase possibility to

finding ova, cyst, or larvae in samples that not be able to seen
by direct microscopy.
Raed Z. Ahmed, Medical Parasitology Lab.,2012

Concentration Methods
1. Sedimentation method
Modified Formal- Ether sedimentation technique
Acid- Ether sedimentation technique
2. Flotation method
Saturated Salt Solution technique
Sheathers Sugar Centrifugal Flotation technique
Zinc Sulphate Centrifugal Flotation technique

Raed Z. Ahmed, Medical Parasitology Lab.,2012

Sedimentation Methods

MODIFIED FORMAL- ETHER

SEDIMENTATION
Raed Z. Ahmed, Medical Parasitology Lab.,2012

Materials and Method

Libra
Applicator stick
Glass centrifugal tubes
Beaker
Wire sieve
Vortex or whirlimixer
Centrifuge.
Reagent:
Reagent I: 10% formalin solution in distilled water.
Reagent II: diethyl ether or ethyl acetate .
Raed Z. Ahmed, Medical Parasitology Lab.,2012

Procedures
1. Emulsify 1 gm. of feces in 7 ml of 10% formalin in a
centrifuge tube.
2. Strain the suspension through a brass wire sieve, and
collect in beaker.
3. Pour the filtrate into a 15 ml boiling tube and add 3 ml
of ether, then mix well 15 sec on vortex or whirlimixer
or 1 min by hand.
4. Transfer the ether- formalin suspension back into the
washed centrifuge tube, and centrifuge at 3,000 rpm for
1 min.
Raed Z. Ahmed, Medical Parasitology Lab.,2012

Procedures

(cont.)

5. Loosen the fatty layer and debris at the top of the tube
with an applicator stick and invert the tube quickly to
discard the supernatant.
6. On righting the tube, a few drops only should remain
with the sediment, mix the sediment well and transfer
one drops onto a glass slide and cover it with coverslip.
7. Scan the whole coverslip using 10x objective, turning
into 40x for confirmation of identification of parasites.

Modified Formal- Ether

Sedimentation
Formalin- Ether or Formalin- Ethyl acetate method is the recommended
concentration procedures.
Most types of worm eggs (round worms, tapeworms, schistosomes, and
other fluke eggs), larvae, and protozoan cysts may be recovered by this
method.
Advantages:
1. Speed: one sample can be processed in 5 minutes.
2. Broad spectrum: it will recover most ova, cyst and larvae.
3. The morphology of most parasites is retained for easy identification.
Disadvantages:
1. Requires several pieces of apparatus which does not make it an easy.
2. The preparation contains some debris.
3. Ether is flammable. Formalin is an irritant.
4. Hymenolepis nana and Fasciola spp. do not concentrate well.
Raed Z. Ahmed, Medical Parasitology Lab.,2012

Sedimentation Methods

ACID- ETHER SEDIMENTATION

TECHNIQUE
Raed Z. Ahmed, Medical Parasitology Lab.,2012

Materials and Method

Libra
Applicator stick
Glass centrifugal tubes
Beaker
Wire sieve
Vortex or whirlimixer
Centrifuge.
Reagent:
Reagent I: 15% Hydrochloric acid.
Conc. HCl 40 ml + 60 ml Distilled water.

Reagent II: diethyl ether or ethyl acetate.

Raed Z. Ahmed, Medical Parasitology Lab.,2012

Procedures
1. Mix thoroughly 1 gm. feces with 3 ml of 15% of
hydrochloric acid and then mix well.
2. Add and additional 5-6 ml of 15% HCl and mix.
3. Strain the suspension through a wire sieve into beaker.
4. Place suspension in a glass centrifuge tube and make up
to the 10 ml with distilled water.
5. Add 4 ml of ether, stopper the tube and shake
vigorously 20 -30 sec using vortex.

Raed Z. Ahmed, Medical Parasitology Lab.,2012

Procedures

(cont.)

6. Centrifuge 2-3 min at 1500 rpm, the suspension now

will be layered.
7. Loosen plug of debris with applicator stick and
immediately pour off liquid.
8. Transfer one drops onto a glass slide and cover it with
coverslip.
9. Scan the whole coverslip using 10x objective, turning
into 40x for confirmation of identification of parasites.

Raed Z. Ahmed, Medical Parasitology Lab.,2012

Intestinal Protozoa

Giardia lamblia
Raed Z. Ahmed, Medical Parasitology Lab.,2012

Giardia lamblia
It is the most common flagellate of the intestinal tract that
cause giardiasis, Traveler's diarrhea.
There is two diagnostic stages for Giardia lamblia :
1. Cyst (infective stage).
2. Trophozoite (motile form, motility by flagella).
Diagnosis:
. Stool examination to see cyst stage, or trophozoite
stage if the sample is fresh.

Raed Z. Ahmed, Medical Parasitology Lab.,2012

Life cycle

Raed Z. Ahmed, Medical Parasitology Lab.,2012

Giardia lamblia cyst

Raed Z. Ahmed, Medical Parasitology Lab.,2012

Giardia lamblia Trophozoite

Raed Z. Ahmed, Medical Parasitology Lab.,2012