MICROBIOLOGY AND PARASITOLOGY

LABORATORY NOTES/ ACTIVITIES

FINALS 3: HELMINTHS AND PROTOZOA

I. SPECIMEN COLLECTION AND TRANSPORT

A. Manner of Collection

1. Typical stool collection protocol consists of 3 specimens.

a. Outpatient: 1 specimen is collected every other day or a total of 3 specimens in 10 days.
Up to 6 specimens are collected in 14 days for Amebiasis.
b. Inpatient: Daily specimen collection is performs for 3 days.

2. Stool specimens for Ova and Parasite examination is best collected in clean, wide-mouthed containers
made of waxed cardboard or plastic with tight fitting lid to ensure retention of moisture and to prevent
accidental spillage.

3. Certain medications also affect specimen collection

a. Stool samples from patients whose therapy includes barium, bismuth, or mineral oil should
be collected prior to therapy or not until 5 to 7 days after the completion of therapy.
b. Patients who have taken antibiotics or antimalarial medications should be delayed for 2
weeks following therapy

4. Acceptable amount: Thumb sized/ walnut-sized.

2-5 grams or 5-6 tablespoons of watery stool.

5. Urine should not be allowed to contaminate the stool specimen.

6. Not be retrieved from toilet bowl water.

B. Important Considerations in Sample Collection

1. The specimen container should be labeled with
a. Patient’s name
b. Identification number
c. Physician’s name
d. Date and Time of collection
2. Requisition forms (paper/ computer-based) must accompany the specimen indicating the test(s)
requested.
3. The specimen should be placed into a zip lock plastic bag for transport to the laboratory. The paperwork
accompanying the specimen should be separated from the specimen container.
4. When handling all specimens, gloves and a protective coat should be worn at all times. Biohazard
hoods should also be used in laboratories when present

C. Stool Preservatives
1. Formalin:
2. Schaudinn’s solution: Considered to be the “Gold standard”
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LABORATORY NOTES/ ACTIVITIES
3. Polyvinyl Alcohol (PVA): Plastic resin that serves to adhere stool samples onto a slide
4. Merthiolate-iodine-formaldehyde (MIF): Enables separation of different strata of stool matter,
and sedimentation of stool particles that contain parasites.
5. Sodium Acetate Formalin: Viable alternative to the use of PVA and Schaundinn fixative
6. Modified Polyvinyl Alcohol: Other alternatives to mercury based PVA are the use of substitute
compounds containing copper sulfate or zinc sulfate.

D. Specimen Transport
 For best results, the stool should be brought to the lab right away. If this isn't possible, the stool
should be stored in preservative provided by the lab and then taken there as soon as possible.
 Raw stool samples should be transported cold and stored immediately at 2–4°C upon arrival. It is
best to test the stool sample as soon as they are received.
 Specimens collected using collection vials can be transported at room temperature and then stored
at 2–4°C for less than 3 days prior to processing.

MICROSCOPIC EXAMINATION
 To detect presence of parasites in stool specimen
 Involves direct wet preparation, concentrated technique, and permanently stained smear.
o Fresh specimen- all 3 procedures are performed
o Fixed specimen- only concentration and permanent stained smear is performed.
 Ocular Micrometer
a. Disk inserted into the eyepiece of the microscope.
b. Size is an important diagnostic feature.
c. Measured in microns (u/ um)

Direct Fecal Smear/ Direct Wet Preparation

 Slide made by mixing unfixed stool with saline (NSS) or iodine and subsequent examination of the
resultant mixture under the microscope.
 Results are only presumptive/ preliminary.

Kato-katz Technique
Kato Katz technique is used for qualitative and semi-quantitative diagnosis of intestinal helminthic
infestations; caused by Ascaris lumbricoides, Trichuris trichiura, hookworm, and especially Schistosoma
spp.
In the Kato-Katz technique, feces are pressed through a mesh screen to remove large particles. A
portion of the sieved sample is then transferred to the hole of a template on a slide. After filling the hole,
the template is removed and the remaining sample is covered with a piece of cellophane soaked in
glycerol. The glycerol clears the fecal material from around the eggs. The eggs are then counted and the
number is calculated per gram of feces.

Examination and results:

 Examine the whole area in a systematic zigzag pattern.
 Record the number and the type of each egg of each species on a recording form alongside the
sample number.
 Multiply the number of eggs by the appropriate number to give the number of eggs per gram (epg)-
standard measurement to assess the intensity of infection.
 MICROBIOLOGY AND PARASITOLOGY
LABORATORY NOTES/ ACTIVITIES

ANTIPROTOZOANS
CINCHONA ALKALOIDS Quinine Reserved for malarial strains resistant to other agents
AE: Cinchonism, Abortifacient
7-CHLORO-4- Chloroquine DOC for erythrocytic falciparum malaria
AMINOQUINOLINES Amiodaquine Highly supressive in P. vivax and P. falciparum
8-AMINOQUINOLINES Primaquine Only agent that can lead to radical cures of the P. vivax & ovale
9-AMINOACRIDINES Quinacrine For tx of giardiasis
Mefloquine For multi-drug resistant forms of Plasmodium falciparum

ANTHELMINTICS
NEMATODES
Piperazine Tx of pinworm roundworm
Pyrantel Pamoate Tx of pinworm, roundworm (ascaris)
Thiabendazole Antimitotic/ Antimicrotubule
Mebendazole Inhibits Microtubule synthesis
Albendazole
Ivermectin Tx of onchocerciasis (river blindness) – Oncocerca volvulus
Diethylcarbamazine Tx of Filiariasis
TREMATODES
Praziquantel Agent of choice for blood flukes (schistosomes)
Niridazole For schistosomiasis
Oxamniquine Tx of Schistosoma mansoni
Bithionol Agent of choice for liver fluke (Fasciola hepatica) and lung fluke (Paragonimus
westermani)
CESTODES
Niclosamide
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LABORATORY NOTES/ ACTIVITIES

NAME: DATE:

COURSE/YEAR/SECTION: SCORE:

FINALS 3: HELMINTHS AND PROTOZOA

Questions for research:

1. What are the advantages of using kato-katz technique?

2. What other methods by which we can determine the worm burden?

3. What parasites can be visualized using kato-katz?

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LABORATORY NOTES/ ACTIVITIES
4. What factors must be considered in evaluating stool sample?

5. As an allied medical student, what is the importance of studying about helminthes and protozoa
including the proper medication for various infections?