Microscopic Diagnosis of Malaria

The direct microscopic visualization of the parasite on the thick and/or thin blood smears has been the "gold standard" for malaria diagnosis.

Thick Blood smear: (Click here to view a video clip)
Thick smears consist of a thick layer of dehemoglobinized (lysed) red blood cells (RBCs). Thus, thick smears allow a more efficient detection of parasites (increased sensitivity). However, they do not permit an optimal review of parasite morphology.
Allow the smear to dry thoroughly. Insufficiently dried smears (and/or smears that are too thick) can detach from the slides during staining. You can accelerate the drying by using a fan or hair dryer. Do not fix thick smears with methanol or heat. If there will be a delay in staining smears, dip the thick smear briefly in water to hemolyse the RBCs.

Thin Blood Smear:(Click here to view a video clip)
Thin smears consist of blood spread in a layer such that the thickness decreases progressively toward monolayer. Fix the smear by dipping them in absolute methanol.

Microscopic examination:
First screen the thick/thin smear at a low magnification (10× or 20× objective lens), to detect large parasites(microfilaria) then examine the smear using oil immersion objective. NCCLS recommend examination of at least 300 oil immersion fields for the determination of "No Parasite Seen".

Diagnostic Points for Plasmodium falciparum

1. Red Cells are not enlarged.
2. Rings appear fine and delicate and there may be several in one cell.
3. Some rings may have two chromatin dots.
4. Presence of marginal or applique forms.
5. It is unusual to see developing forms in peripheral blood films.
6. Gametocytes have a characteristic crescent shape appearance. However, they do not usually appear in the blood for the first four weeks of infection.
7. Maurer's dots may be present.

Diagnostic Points for P. vivax

1. Red cells containing parasites are usually enlarged.
2. Schuffner's dots are frequently present in the red cells as shown above.
3. The mature ring forms tend to be large and coarse.
4. Developing forms are frequently present.

Diagnostic Points for P. malariae

1. Ring forms may have a squarish appearance.
2. Band forms are a characteristic of this species.
3. Mature schizonts may have a typical daisy head appearance with up to ten merozoites.
4. Red cells are not enlarged.
5. Chromatin dot may be on the inner surface of the ring.

Diagnostic Points for P. ovale

1. Red cells enlarged.
2. Comet forms common (top right).
3. Rings large and coarse.
4. Schuffner's dots, when present, may be prominent.
5. Mature schizonts similar to those of P. malariae but larger and more coarse.

Quantifying parasites:
% parasitemia = (parasitized RBCs/total RBCs) × 100


Click here for some helpful hints and case presentations.
Click here for self assessment on microscopic diagnosis of Malaria.

For More Information follow the links:

• Preparation of blood smears
• Staining for malaria parasites
• Laboratory diagnosis of Plasmodium falciparum
• Laboratory diagnosis of Plasmodium vivax
• Laboratory diagnosis of Plasmodium ovale
• Laboratory diagnosis of Plasmodium malariae
• Comparison of the Plasmodium Species Which Cause Human Malaria
• Plasmodium spp. Blood Stage Parasites, Thick Blood Smears
• Plasmodium spp. Blood Stage Parasites, Thin Blood Smears

Courtesy: CDC & Royal Perth Hospital