MALARIA

Made By- Dr. Pratibha Dhiwan

PG JR 2
Introduction
• Plasmodium falciparum is the major species associated with deadly infections
throughout the world.
• Only five species infect humans.
• Of the species that cause human disease, Plasmodium vivax and P. falciparum
cause 95% of infections.
• P. vivax may be responsible for 80% of the remaining infections, because this
species has the widest distribution in the tropics, subtropics, and temperate zones.
• P. falciparum is generally confined to the tropics, Plasmodium malariae is
sporadically distributed, and Plasmodium ovale is confined mainly to central
West Africa and some South Pacific islands.
• The fifth human malaria, Plasmodium knowlesi, a malaria parasite of longtailed
macaque monkeys, has been confirmed in human cases
• Reservoir of infection- Man, particularly children, in some parts of
Africa chimpanzees.
• Method of transmission- Inoculative method
- Transfusion malaria
- Congenital malaria
- Malaria in drug addicts
• Transmitting agent- Female anopheles.
• Infective form – sporozoites
• Portal of entry- skin
• Site of localisation- First in liver cells, then RBCs
Life Cycle Of Plasmodium
Metabolism of haemoglobin in Malaria
Plasmodium vivax (benign tertian malaria)
1 48-h cycle
2 Tends to infect young cells
3 Enlarged RBCs
4 Schüffner dots (true stippling) after 8–10 h
5 Delicate ring
6 Very amoeboid trophozoite
7 Mature schizont contains 12–24 merozoites
Morphology of malaria parasites
Plasmodium vivax:
1, Early trophozoite (ring form).

2, Late trophozoite with Schüffner dots (note enlarged red blood cell).

3, Late trophozoite with amoeboid cytoplasm (very typical of P. vivax).

4, Late trophozoite with amoeboid cytoplasm.

5,Mature schizont with merozoites and clumped pigment.

6, Microgametocyte with dispersed chromatin.

7, Macrogametocyte with compact chromatin.

Plasmodium vivax (note enlarged infected red blood cells
[RBCs]).
(1) Early trophozoite (ring form) (note one RBC contains
two rings—not that uncommon);

(2) older ring, note ameboid nature of rings;

(3) late trophozoite with Schüffner dots (note enlarged

RBC);

(4) developing schizont;

(5) mature schizont with 18 merozoites and clumped

pigment;

(6) microgametocyte with dispersed chromatin.

Plasmodium malariae (quartan malaria)
1 72-h cycle (long incubation period)
2 Tends to infect old cells
3 Normal size RBCs
4 No stippling
5 Thick ring, large nucleus
6 Trophozoite tends to form “bands” across the cell
7 Mature schizont contains 6–12 merozoites
Plasmodium malariae :
1, Early trophozoite (ring form).

2, Early trophozoite with thick cytoplasm.

3, Early trophozoite (band form).

4, Late trophozoite (band form) with heavy

pigment.

5, Mature schizont with merozoites arranged in

rosette.

6, Microgametocyte with dispersed chromatin.

7, Macrogametocyte with compact chromatin

Plasmodium malariae (note normal or smaller than
normal infected RBCs):-

(1)Early trophozoite (ring form);

(2)early trophozoite with thick cytoplasm;
(3)late trophozoite (band form);
(4)developing schizont;
(5)mature schizont with nine merozoites arranged in a
rosette;
(6) microgametocyte with compact chromatin
Plasmodium ovale
1 48-h cycle
2 Tends to infect young cells
3 Enlarged RBCs with fimbriated edges (oval)
4 Schüffner dots appear in the beginning (in RBCs with very young ring
forms, in contrast to P. vivax)
5 Smaller ring than P. vivax
6 Trophozoite less amoeboid than that of P. vivax
7 Mature schizont contains an average of 8 merozoites
Plasmodium Ovale
1, Early trophozoite (ring form) with Schüffner dots.

2, Early trophozoite (note enlarged red blood cell).

3, Late trophozoite in red blood cell with fimbriated edges.

4, Developing schizont with irregularly shaped red blood cell.

5, Mature schizont with merozoites (8) arranged irregularly.

6, Microgametocyte with dispersed chromatin.

7, Macrogametocyte with compact chromatin

Plasmodium ovale (note enlarged infected RBCs)
(1)Early trophozoite (ring form) with Schüffner dots (RBC
has fimbriated edges);

(2)early trophozoite (note enlarged RBC, Schüffner dots, and

RBC oval in shape);

(3)late trophozoite in RBC with fimbriated edges;

(4)developing schizont with irregular-shaped RBC;

(5)mature schizont with eight merozoites arranged

irregularly;

(6)microgametocyte with dispersed chromatin

Plasmodium falciparum (malignant tertian
malaria)
1 36–48–h cycle
2 Tends to infect any cell regardless of age, thus very heavy infection may result
3 All sizes of RBCs
4 No Schüffner dots (Maurer dots: may be larger, single dots, bluish)
5 Multiple rings/cell (only young rings, gametocytes, and occasional mature
schizonts are
seen in peripheral blood)
6 Delicate rings, may have two dots of chromatin/ring, appliqué or accolé forms
7 Crescent-shaped gametocyte
Plasmodium falciparum
1, Early trophozoite (accolé or appliqué form).

2, Early trophozoite (one ring is in headphone configuration/double chromatin

dots).

3, Early trophozoite with Maurer dots.

4, Late trophozoite with larger ring and Maurer dots.

5, Mature schizont with merozoites.

6, Microgametocyte with dispersed chromatin.

7, Macrogametocyte with compact chromatin

(1) Early trophozoites (the rings are in the headphone
configuration with double chromatin dots);

(2) early trophozoite (accolé or appliqué form);

(3) early trophozoites (note the multiple rings/cell);

(4) late trophozoite with larger ring (accolé or appliqué

form);

(5) crescent-shaped gametocyte;

(6) crescent-shaped gametocyte.

Pernicious Malaria
Series of phenomena occurring during the course of an infection of P. falciparum
which if not effectively treated threatens the life of the patient within 1 to 3 days

• Pathogenesis- The serious complications are the result of capillary blockage

consequent upon decreased effective circulating blood volume
• The blockade of the capillary blood vessels of the internal organs arises from
agglutination of parasitised erythrocytes

Clinical types-
1. Cerebral malaria- Manifested by hyperpyrexia, coma, paralysis
2. Algid malaria- cold and clammy skin with vascular collapse leading to
peripheral circulatory failure, along with either vomiting or watery diarrhoea
or passage of blood in faeces
3. Septicaemic malaria- high continued temperature, bilious remittent fever,
pneumonia & cardiac syncope.
Black water fever
It is a rare manifestation of falciparum malaria occurring in previously infected subjects and is
characterised by sudden intravascular haemolysis followed by fever and haemoglobinuria.

Etiology-
• It is most commonly associated with P. falciparum infection among non- immune
individuals who have resided in malarious countries for 6 moths to 1 year & have had
inadequate doses of quinine for both suppressive prophylaxis and treatment of repeated
clinical attacks.
• Other factors- cold, exposure to the sun, fatigue, trauma , pregnancy, parturition & X- ray
exposure.
Pathogenesis- Intravasular hemolysis metheamalbumineamia, hyperbilirubinemia,
haemoglobinuria
Main features of malarial infection
• Pigmentation of various organs
• Hyperplasia of reticuloendothelial system
• Parasitised erythrocytes filling the lumina of the capillaries of the
internal organs
• Vascular changes
• Degenerative changes in parenchymal cells
• Fibrosis
• Immunosuppression
Pathological Changes in Various Organs
Spleen
Macroscopic Appearance- moderately enlarged
• Color- slate- grey or black
• Capsule is thin
• Consistency- soft in acute & hard in chronic phase
• Cut surface- homogenous black area with scattered white trabeculae &
greyish white spots of Malphigian corpuscles along with hemorrhage
• Microscopic – Congestion of splenic sinusoids

• Enormous amounts of pigment- inside macrophages of red pulp

• Number of macrophages greatly increased

• White pulp free from pigment & parasites.

• Reticulin fibrils increased

Liver
Macroscopic appearance-

• Uniformly enlarged

• Color- chocolate red to slate grey or black

• Cut surface- dilated lobular vein, yellow area against a brownish

background
Microscopy-
• The central veins of the lobules & sinusoidal capillaries are dilated &
filled with parasitised RBCs.

• Kupffer cells- Increased in number & cytoplasm is filled with malarial

pigment & parasitised RBCs.

• Parenchymal cells of liver lying in central zone show fatty

degeneration, atrophy & centrilobular necrosis.

• The parasites are found in various stages of development

• Fibrosis in central or portal zone secendory to the process of

degeneration.
Bone Marrow-
• Unstained bone marrow films in chronic case appear slate grey or
black

• Malaria parasites detected in RBCs & neutrophils

• In acute P. falciparum infection- hypo, normo or mildly hypercellular,

erythropoiesis reduced

• In chronic P. falciparum infection- hypercellularity , erythroid

hyperplasia, dyserythropoieses, giant metamyelocytes & increased
eosinophils, lymphocytes, plasma cels & macrophages
• Bone marrow biopsy-
- Increased cellularity & increased macrophage activity often with
haemophagocytosis

- Sinusoids packed with parasitized red cells.

- In repeated attacks – bone marrow may be slate grey or black

- Hemozoin pigment present in macrophages, erythroid & granulocytic

precursors.

- Dyserythropoiesis & erythroid suppression

Kidney-
Divided in two groups:-

• In acute falciparum malaria- Renal anoxia syndrome (oliguria or

anuria & acute uraemia) aka renal tubular vascular syndrome or lower
nephron nephrosis

• Those found in relapsing quartan malaria, resembling nehrotic

syndrome- occur due antigen antibody compexes deposition in
glomerular basement membrane.
Pathology of cerebral malaria
• The entire capillary network of brain is distended and occluded by the parasitised
RBCs.
• More prominent in the grey mater & distribution is uniform in all location

Macroscopic appearance- Pial and cerebral vessels are markedly congested.

Cut surface- slate grey colour of the cortex & multiple hemorrhages in the
subcortical white matter and small infracts.
Microscopy-
-Dilatation & congestion of cerebral capillaries
-Perivascular haemorrhages, having the appearance of ring haemorrhages
-Scattered areas of softening due to degeneration of the nerve tissue, later invaded
by glial cells leading to Durck granuloma formation
Pathology of algid malaria
GIT- Mucous membrane pigmented to slate- grey colour
-Punctate haemorrhages seen
- Intestinal contents watery or dark brown with very little mucus
- Microscopically mucosal and submucosal capillaries are congested

Peripheral blood vessels- generalised vascular collapse or peripheral

circulatory failure

Adrenal Glands- necrosis of Zona fasciculata & hemorrhages with

congestion of Zona reticulata.
Pathology of septicaemic malaria
Heart- Microscopically reveals intensely congested coronary blood
vessels filled with parasitised RBCs along with fatty degeneration &
necrosis of heart muscle

Lung- Pneumonic symptoms, show small areas of hemorrhages with

patches of oedema & collapse.
-Alveolar capillaries congested & filled with parasitised erythtrocytes.
-Lung alveoli contain extravastated RBCs & pigmented monocytes.

Blood- High degree of parasitaemia & both schizogony &

gametogony occur in the peripheral circulation.
Clinical Characteristics of the Five Human Infections
Clinical Picture
• Anemia- hemolytic anemia , normocytic hypochromic with reduced
RBC count
• Leucocyte count- Increases during rising temperature, then returns to
normal after temperature comes down.
- Chronic cases – leukopenia
- Monocytes- increase ranging from 10- 20 percent
 Chemical changes in blood in malaria

• Plasma albumin is reduced. Albumin / globin ratio is reversed.

• Cholesterol level rises during rigor and falls during apyrexial period.

• Rise of blood sugar is probably correlated to adrenal function. Low blood

sugar in falciparum malaria.

• Rise of plasma potassium resulting from destruction of red blood cells.

• ESR is raised. It is related to plasma protein & change in the surface of
the red blood cells.
• A fall of pH and a loss of alkali reserve, due to increase of pyruvates
and lactates.
• Indirect bilirubin is increased.
• Biological changes in erythrocytes comprise increase in cellular
sodium with equivalent decrease in cellular potassium.
Laboratory diagnosis
• Examination of blood for parasite
- Time for taking blood
- Rules to be followed:-
Preparation of thin and thick film
Fluorescence Microscopy
• Red cells containing malaria parasites fluoresce when examined by
fluorescence microscopy after staining with acridine orange.
• The fluorescent dye conjugates with the gamma globulin of the serum
to be tested
• If such conjugated gamma globulin contains malarial antibody, it will
adhere to the relevant malarial parasite & will be recognised as
glistening particles under the fluorescent microscope.
• This has a sensitivity of about 90% in acute infections
• False-positive readings may occur with Howell–Jolly bodies and
reticulocytes.
Quantitative Buffy Coat Method
• The quantitative buffy coat (QBC) method (BD Diagnostic Systems) is
another procedure for detection of parasites by fluorescent
microscopy.
• The blood is centrifuged in capillary tubes that are coated with
acridine orange.
• It is fairly sensitive but requires expensive equipment and has the
disadvantage of false-positive results in the presence of Howell–Jolly
bodies and reticulocytes.
• When positive, identification of species requires examination of a
stained blood film, but it is useful as an initial screening test
 Antigen-Based Tests
• RDTs use small blood samples obtained by finger prick or by
venepunture and employ a ‘lateral diffusion’ system to generate
results.
• In general, a blood specimen to be tested (2–50 ml) is lysed in buffer
solution containing one or more malaria-specific ‘detection
antibodies’.
• The detection antibody is coupled to a visually observable label.
• Where specific antigen is present, a complex is formed between that
antigen and its cognate labelled antibody
• The labelled antigen–antibody complex generated is then bound by a
second ‘capture-antibody’ that recognizes the same antigen, and which
is immobilized as a line on the test strip.

• A positive result therefore generates a visible line of antigen–antibody

complex.

• A separate immobilized capture antibody recognizes the labelled

detection antibody alone; this control band will produce a line in the
absence of malaria antigen and confirms that the test has been
performed correctly and the result can be interpreted.
• The antigen targets detected by RDTs fall into two groups:-

• The first group of antigens are expressed in all malarial species.

Antigens from this group therefore confirm malarial infection is
present, but do not allow the parasite species to be determined.
Antigens from this group are Plasmodium aldolase (PMA) or parasite
lactate dehydrogenase (pLDH).

• The second group of antigens are specific for P. falciparum. Antigens

from this group are histidine-rich protein-2 of P. falciparum (pfHRP2)
or a P. falciparum-specific form of LDH (pfLDH).
• A ‘two line’ test uses an anti-pfHRP2 band together with the positive control band
to recognize P. falciparum only

• A ‘three line’ test uses a P. falciparum specific antibody band, together with PMA or
pLDH, together with the positive control antibody. The three line test can therefore
indicate the presence of P. falciparum, or if P. falciparum is not present the test will
indicate the presence of other malarial species.

• Since the second antibody is panmalarial, the three line test does not distinguish
between infection by P. vivax, P. ovale, or P. malariae, and will not detect mixed
infection.
Rapid diagnostic tests for malaria. An RDT of the ‘three line’ format, the image shows the output window.
(A) The results of a negative test in which the positive control line shows a positive band to indicate the technical
validity of the result.
(B) The positive lines in all three positions, i.e. a technically interpretable result (control line positive), with P.
falciparum-specific antigen detected (line 1), and pan-malarial antigen detected (line 2). This patient had a single-
species infection (P. falciparum), although the test does not allow a mixed infection to be excluded.
Nucleic Acid Detection
• Direct detection of the five species by using a specific DNA probe after PCR
amplification of target DNA sequences.
• Some laboratories are now using PCR for detection of malaria; the high
sensitivity, rapidity, and simplicity of some of the methods are becoming much
more relevant for diagnosis.
• Detection is possible for as few as 5 or 10 parasites per microliter of blood;
thus, PCR detects many more cases of low-level parasitemia than do thick blood
films.
• Although these tests are more sensitive than the traditional blood films, they are
primarily available as laboratory-developed tests and are not FDA approved.
• When a positive result is obtained using a nucleic acid–based test, it is still
recommended that a thin and thick blood smear be completed to quantitate the
level of parasitemia.
Automated Instruments

• Using automated flow cytometry hematology instruments, there are

potential limitations related to the diagnosis of blood parasite
infections.

• Failure to detect a light parasitemia is highly likely

Serological test

• It is also extremely difficult to differentiate a previous from an acute

malaria infection.

• Serology is primarily used to screen donor blood units and for

epidemiologic studies
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