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Plasmodium: Diagnosis

Laboratory Diagnosis

Microscopy – detecting & identifying malarial parasites in peripheral blood films.
Concentrating parasites in venous blood by centrifugation when they cannot be found in blood films
Using a rapid malaria Ag or enzyme detection test
Other tests – Hb, PCV, Blood glucose, total WBC & platelet count.

Examination of blood Film

Collection of blood
Best prepared directly from capillary blood
In EDTA bulb (used within 30 mins)

Time of collection

As soon as possible if malaria is suspected
Before administering antimalarials
During pyrexia lphase

Types of blood Film

Two types:

1.Thick films :

30 to 40 times more sensitive than thin films
More suitable for detection of malarial parasite when they are few in number
Blood is not fixed, RBCs are lysed during staining (only parasitic forms will be seen)

2. Thin films :

To confirm the Plasmodium species
Assists in the identification of mixed infections
Blood is fixed, parasites are seen within the RBCs
Also helps in assessing the response to treatment.

Fixation and Staining

Fixation – thin films are fixed with absolute alcohol for 1 to 2 mins.
Staining – films are stained with Romanowsky stain: Giemsa, field’s, Wright’s
Giemsa – 10% solution for 10 mins

Identifying Blood Film

Look for the different morphological forms of the parasite in blood smear:

Trophozoites / ring forms
Schizont(Not seen in Plasmodium falciparum)
Gametocytes

Trophozoites / ring forms

Character	P. vivax	P. falciparum
Size	2.5µ (1/3rd of RBC)	1.25 to 1.5 µ
Cytoplasm	Thick opposite to nucleus	Uniform thickness
Nucleus	One/ ring	Can have >1
Number of rings	One ring/ RBC	>1/ RBC
Location in RBCs	Always inside RBCs	Inside as well as on the surface (accole’ forms)
Type of RBC infected	Preferentially young RBCs & reticulocytes	All types

Schizont

Character	P. vivax	P. falciparum
Size of RBC	Increases to twice its size	Does not change
No of merozoites	16	8 to 32
Arrangement of merozoites	Symmetric in form of rosette	Asymmetrical
Presence in peripheral blood	Present	Absent

Gametocytes

Character	P. vivax	P. falciparum
Shape – Male Female	Spherical Spherical	Cresentic Sausage shaped
Nucleus – M F	Central, diffuse Peripheral,small	Central, diffuse Central,compact
Infected RBC	Enlarged	Deformed, with its membrane, stretched.

Important Points

Schüffner’s dots

Exclusively found in Plasmodium ovale and Plasmodium vivax
Morphologic alterations in infected host erythrocytes that are visible as multiple brick-red dots

Maurer’s dot:

Fine granular precipitates or irregular cytoplasmic particles present in red blood cells infected with the trophozoites of Plasmodium falciparum

Counting the % age of parasitised RBCs

On thin blood films
When falciparum malaria parasitemia is high

Method of counting:

Select an area where no of RBCs is roughly 250.
Count the no of parasitised RBCs in 4 such fields i.e. approximately 1000 RBCs.
Divide by 10 to obtain the percentage.
*WHO – if it is >5%, then the parasitemia is heavy & prognosis is poor.

Rapid Diagnostic Test

Developed to diagnose falciparum malaria rapidly & without a microscope.
Can also detect vivax malaria
Immunochromatographic test
Three tests are available commercially
Detects HRP2 Ag (Histidine-rich protein) or specific pLDH (parasite lactate dehydrogenase) or Pan malarial plasmodium aldolase
Both HRP2 & pLDH are produced by the parasites during their growth & differentiation in RBCs.

Exam Question

Laboratory Diagnosis

Microscopy – detecting & identifying malarial parasites in peripheral blood films.
Concentrating parasites in venous blood by centrifugation when they cannot be found in blood films
Using a rapid malaria Ag or enzyme detection test
Other tests – Hb, PCV, Blood glucose, total WBC & platelet count.

Thin films :

To confirm the Plasmodium species

Identifying Blood Film

Look for the different morphological forms of parasite in blood smear:

Trophozoites / ring forms
Schizont(Not seen in Plasmodium falciparum)
Gametocytes

Schüffner’s dots

Exclusively found in Plasmodium ovale and Plasmodium vivax
Morphologic alterations in infected host erythrocytes that are visible as multiple brick-red dots

Maurer’s dot:

Fine granular precipitates or irregular cytoplasmic particles present in red blood cells infected with the trophozoites of Plasmodium falciparum

Immunochromatographic test

Three tests are available commercially
Detects HRP2 Ag (Histidine-rich protein) or specific pLDH (parasite lactate dehydrogenase) or Pan malarial plasmodium aldolase
Both HRP2 & pLDH are produced by the parasites during their growth & differentiation in RBCs.

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Plasmodium: Diagnosis

Plasmodium: Diagnosis

Schüffner’s dots

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Plasmodium: Diagnosis

Schüffner’s dots

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