Lab diagnosis of Trematodes, Blood Flagellates,Plasmodium

and Protozoans

Prepared by: Faiz Ullah 2nd batch MLT

T. b. rhodesiense and T. b. gambiense

T. b. rhodesiense and T. b. gambiense trypanosomes● Trypanosomes measure 13–42 micro meter in length and may show a variety of forms (pleomorphic).* * In the early stages of acute African trypanosomiasis,

long slender trypanosomes (often seen dividing) can be found. In the later stages, intermediate and trypanosomes, some having no free flagellum,may be seen.

● Single flagellum arises from the kinetoplast. It extends forwards along the outer margin of undulating

membrane and usually beyond it as a free anterior flagellum.

T. b. rhodesiense and T. b. gambiense

● Small dot-like kinetoplast stains darkly.● Nucleus stains dark-mauve and is usually centrally placed but posterior nuclear forms may also be seen.● Cytoplasm stains palely and contains granules.

Note: Microscopically, the trypanosomes of T. b. rhodesienseand T. b. gambienselook the same.

T. b. rhodesiense and T. b. gambiense

Morphology of a trypanosome of T.b.gambiense orT.b.rhodesiense

Left: T.b.gambiense/T.b.rhodesiensein thick blood film. Right: T.b.gambiense/T.b.rhodesiense in thin blood film.

T. cruzi trypanosomes

● Usually C-shaped, measuring 12–30 micro meter in length with a narrow membrane and freeflagellum.

● Has a large, round to oval, dark-red staining kinetoplast at the posterior end.

● Nucleus is centrally placed and stains redmauve. Note: Occasionally slender forms of T. cruzican be seen

which have an elongated nucleus, subterminal kinetoplast, and shorter free flagellum.

T. cruzi trypanosomes

Right: T.cruziin Giemsa stained thin blood film.Left: T.rangeli.

Amastigotes of Leishmania species

Structurally, the amastigotes of Leishmania species that cause VL, CL and MCL are similar. There are variations in size between species.

● Small, round to oval bodies measuring 2–4 micro meter

● Can be seen in groups inside blood monocytes (less commonly in neutrophils), in macrophages in aspirates or skin smears, or lying free between cells.

● The nucleus and rod-shaped kinetoplast in each amastigote stain dark reddish-mauve.

● The cytoplasm stains palely and is often difficult to see when the amastigotes are in groups.

Amastigotes of Leishmania species

Leishmaniaamastigotes in monocyte in a Giemsastained blood film

Giemsa stained amastigotes of L.donovani. Right:As seen in bone marrow. Left: As seen in splenic aspirate.

Plasmodium Species• The appearance of each of the four species of plasmodia that

infect humans is sufficiently different to allow their differentiation in stained smears

• The parasitized erythrocyte in P. vivax and P. ovale infections is pale, enlarged, and contains numerous Schüffner’s dots

• All asexual stages (trophozoite, schizont, merozoite) may be seen simultaneously

• Cells infected by P. ovale are elongated and frequently irregular or fimbriated in appearance

• In P. malariae infections, the RBCs are not enlarged and contain no granules. The trophozoites often present as “band” forms, and the merozoites are arranged in rosettes around a clump of central pigment.

• In P. falciparum infections, the rings are very small and may contain two chromatin dots rather than one

Plasmodium Species• There is often more than one parasite per cell, and

parasites are frequently seen lying against the margin of the cell

• Intracytoplasmic granules known as Maurer’s dots may be present but are often cleft shaped and fewer in number than Schüffner’s dots

• Schizonts and merozoites are not present in the peripheral blood. Gametocytes are large and banana shaped

Plasmodium Species

Trophozoites - Plasmodium falciparum: Early trophozoites have the characteristic signet ring shape. Also, unique to P. falciparum is the presence of multiple trophozoites in one cell.

Trophozoites - Plasmodium vivax: Red blood cells infected by P. vivax are often larger than uninfected red blood cells. They approximately 1.5 times the size of a normal cell

Trophozoites - Plasmodium malariae: Characteristic trophozoites of P. malariae showing the ring shape and the tendency of infected cells to be of normal or smaller size (arrows).

Gametocyte - Plasmodium falciparum: The gametocytes of P. falciparum have a crescent or banana shape.

Gametocyte - Plasmodium malariae: The gametocytes of P. malariae havea round shape about the size of red blood cells. They have a fine granular appearance.

Gametocyte - Plasmodium ovale: A round gametocyte that is larger than normal red blood cells. It has a granular appearance as well as Schuffner's dots

Schizont - Plasmodium malariae: A schizont containing merozoites (6 to 12) giving a coarse granular appearance

Schizont - Plasmodium vivax: A schizont showing the large number of merozoites typical of this species (16-24). Also note the larger size compared to a normal red blood cell

A red blood cell showing the Schuffner's dots characteristic of cells infected by Plasmodium vivax and Plasmodium ovale

Egg of S. mansoni

● It is pale yellow-brown, large, and oval, measuring about 150*60 micro meter.● Has a characteristic side (lateral) spine.Note:Sometimes the spine may appear terminal likethat of an S. haematobiumegg but if the egg is rolledover by pressing gently on the cover glass the spinewill be seen to be lateral.● Contains a fully developed miracidium.

Egg of S. japonicum

● It is colourless or pale yellow-brown, large and round to oval, measuring about 90*65 micro meter.

● A very small hook-like spine (rudimentary spine) can sometimes be seen projecting from the egg wall but often it is hidden by faecal debris and red cells.

● Contains a fully developed miracidium.

Egg of S. mansoni & S. japonicum

Egg of S.mansoni with lateral spine. Egg of S.japonicum. Rudimentary spine is not visible.

Egg of S. haematobium

● It is pale yellow-brown, large and oval in shape, measuring about 145*55 micro meter.● Has a characteristic small spine at one end

(terminal spine).● Contains a fully developed miracidium. Note: Sometimes the miracidia hatch from the eggs and

can be seen ‘swimming’ in the urine ● A living egg (in fresh unpreserved urine) shows what is

called flame cell movement, i.e. flickering of the excretory flame cells

Egg of S. haematobium

S.haematobiumegg and redcells in urine

Positionof flame cell

Egg of F. hepatica

● It is yellow-brown, large and oval. F. hepatica eggs measure 130–145*70–90micro meter. F. gigantica eggs are larger, measuring 156–197*90–104 micro meter.

● Has an indistinct operculum (lid).● Contains an unsegmented ovum surrounded by many yolk cells.

Note: Morphologically Fasciolaeggs resemble those of Fasciolopsis buski and Echinostoma species.

Egg of F. hepatica

Eggs of F.hepatica.

Egg of F. buski

● It is yellow-brown, large and oval, measuring 130–154*78–98 micro meter.

● Has a small operculum (lid) which is usually difficult to see.● Contains an unsegmented ovum surrounded by yolk

cells.

Note: Morphologically the eggs of F. buskiresemble those of F. hepatica, F. gigantia(similar but smaller), and Echinostomaspecies.

Egg of F. buski

Egg of F.buski.

Trophozoite of E. histolytica

● Average size is about 25*20 micro meter

● Shows active amoeboid movement (directional) in fresh warm specimen.

● In dysenteric specimens, the amoebae contain ingested red cells. This is diagnostic of E. histolytica.● Single nucleus is present which has a central

karyosome (not always visible).

Trophozoite of E. histolytica

Morphology of E.histolyticatrophozoite

Cysts of E. histolytica/E. dispar

● Round, measuring 10–15 micro meter

● Contain, 1, 2 or 4 nuclei with a central karyosome (special staining techniques are required to show details of nuclear structure).

● Chromatoid bodies (aggregations of ribosomes) can be seen particularly in immature cysts. They do not stain with iodine but can be stained with Burrow’s stain or Sargeaunt’s stain

Cysts of E. histolytica/E. dispar

Morphology of immature and matureE.hisolytica/E.disparcysts.

Trophozoite of G. lamblia

● Small pear-shaped flagellate with a rapid tumbling and spinning motility, often likened to a falling leaf.

● Measures 12–15*5–9 micro meter ● Has a large concave sucking disc on the ventral surface

(attaches trophozoite to the intestinal mucosa).● It has four pairs of flagella, two axonemes,and two nuclei

which stain well (Giemsa or Field’s technique).● A single or two curved median bodies are present (function unknown).

T

Cyst of G. lamblia

● Small and oval measuring 8–12 * about 6 micro meter

● Internal structures include four nuclei grouped at one end (sometimes difficult to see), axonemes, median bodies, and remains of flagella. These structures can be seen in unstained preparations but are more clearly defined in an iodine preparation.

Trophozoite & Cyst of G. lamblia

Morphology of G.lamblia. Left: cyst. Right: trophozoite

Cyst of B. coli

● Large, round, thick walled, measuring 50–60 micro meter in diameter.

● Cilia (inside wall) can sometimes be seen in younger cysts.

● Macronucleus stains well with iodine.

Trophozoite of B. coli

● Large, easily seen oval shaped ciliate with a rapid revolving motility, measuring 50–200*40-70 micro meter

● With careful focusing, the beating cilia can be seen particularly in the region of the funnel-shaped cytostome.

● Often contains ingested red cells.● A large macronucleus may be seen. A very small

micronucleus lies close to the macronucleus but this can only be seen instained preparations.

● One of two contractile vacuoles may be visible.

Trophozoite & Cyst of B. coli

Morphology of B.colicyst & B.coli cyst in saline preparation

B.coli trophozoite in saline preparation.

Trichomonas vaginalis

Trichomonas vaginalis