Protozoan Maricelle Manlutac

PROTOZOANS
Maricelle D. Manlutac, RMT, MPH, MLS (ASCPi)

Faculty
Protozoans
 Unicellular organisms
 No cell wall
 Contains at least one nuclei
 2 regions of cytoplasm
 Some contain vacuoles
 With special organs for locomotion
Protista
Sarcomastigophora Ciliophora Apicomplexa

-Cilia -Apical complex
• Balantidium coli • Plasmodium

Mastigophora Sarcodina
• Babesia
-Flagella -Pseudopodium
• Toxoplasma
• Isospora
• Giardia • Entamoeba • Cyclospora
• Chilomastix • Endolimax • Cryptosporidium
• Iodamoeba
• Trichomonas
• Acanthamoeba
• Trypanosoma • Naegleria
• Leishmania
Microspora
• Dientamoeba
• Enterocytozoon
• Encephalitozoon
Trophozoite Cyst
 Motile
 Non-motile
 Feeding stage
 Non-feeding stage
 Vegetative stage
 Found in formed stool
 Found in diarrheal and liquid
 Infective stage
stool
 Amoeba excreted as
trophozoite cannot mature to
cyst
 Use saline not iodine

Amoeba: Rule
 All amoeba are non-pathogenic except E. histolytica
 All amoeba encyst except E. gingivalis
 Cyst is the IS of all amoeba except E. gingivalis
 All amoeba are found in the GIT except E. gingivalis
 **E. gingivalis - their food vacuoles contain cellular debris mostly

leukocytes (WBC) not RBc, which is a distinguishing characteristic of
this parasite
Amoeba
Trophozoite Entamoeba coli Entamoeba histolytica
Movement Sluggish / Non-Progressive/ Progressive

Non- directional Unidirectional
Shape of Blunt Finger-like
Pseudopodia
Release of Several at a time One at a time in an
Pseudopodia explosive manner
Nucleus Mononucleated Mononucleated
Karyosome Eccentric Center
Inclusion body Bacteria, debris RBC

“Dirty looking” “Clear looking”
Amoeba
Cyst Entamoeba coli Entamoeba histolytica
Number of nuclei Greater than 4 Quadrinucleated

(6-8) (1-4)
Chromatoidal bars/ Broomstick in appearance Sausage like
bodies Cigar shape
Rod-shape
Entamoeba coli
VS
Entamoeba
histolytica
Entamoeba coli
Entamoeba coli
Entamoeba histolytica
 Pathology: Amebiasis
Abdominal pain and diarrhea with or without blood and
mucus
Ulceration (narrow neck and broad base)
Fulminant colitis with severe bloody diarrhea in children
Extra-intestinal form: Amebic liver abscess
Lab Test:
1. Stool Exam (DFS)
2. Concentration techniques
3. Stool culture: Robinson’s and Inoki medium
4. PCR/ ELISA
5. Serological test
Amoeba
Endolimax nana
Trophozoite Cyst
 Sluggish motility  Quadrinucleated
 Blunt pseudopodium  “Cross-eyed appearance”
 Inclusion body: debri, bacteria  Comma shaped chromatoidal

bars
 Karyosome: large, irregular
Endolimax nana
Amoeba
Iodamoeba butschlii
Trophozoite Cyst
 Large vesicular nucleus

 Uninucleated
 Achromatic granules
 Large glycogen body
 High affinity with iodine

 “Iodine cyst”
Iodamoeba butschlii
Flagellates
Giardia lamblia
Trophozoite Cyst
 “Old man’s face with eye glasses”  Ovoid

 Bilaterally symmetrical
 Quadrinucleated (rare 2 but
 Pear shaped never 3)
 4 pairs of flagella  Axostyle is still present

 Falling leaf motility
 IS
 Two nuclei- powerful ventral sucking
disc
 Axostyle/ Axial Rod/ Axoneme
 Parabasal body
 Acts as energizing body
Flagellates
Giardia lamblia
Pathology: Travelers' diarrhea
Malabsorption- Steatorrhea ( fats in stool: 5g in 24 hrs)
Excessive flatus – “Rotten eggs”
Lab Test:
1. Stool Exam (DFS)
2. Doudenal Fluid – Entero Test: capsule with nylon thread-trophozoite
3. Serological – cyst wall protein 1 (CWP1) antigen

Giardia lamblia
Giardia lamblia
Flagellates
Chilomastix mesnili
Trophozoite Cyst
 3 anterior flagella
 Mononucleated with distinct
 1 cytosomal flagellum protuberance
 Sheperd’s crook
 “American lemon”
 Asymmetrical
 “Nipple like cyst”
 Spiral groove
 Cork-screw like motility
Chilomastix mesnili
Flagellates
Trichomonas vaginalis
Trophozoite Cyst
 Pear shape
 NO CYST FORM
 4 anterior flagella
 Prominent axostyle – caudal

appearance
 Short undulating membrane
 Rapid jerky tumbling motility
 Siderophil granules
Flagellates
Pathology: Vulvovaginitis
Non specific prostatitis in males
Inflamed Cervix: “Strawberry like”
Lab Test:
1. Microscopic- urine, seminal fluid, prostatic fluid, penile/ vaginal discharge,
and cervical smear
2. Culture Method- Diamond Modified Medium, Feinberg and Whittington
Medium
3. InPouch TV Test
InPouch TV Test
 NO CYST FORM
Trichomonas species
Characteristics T. vaginalis T. hominis T. tenax
Habitat Genitalia Intestine Oral cavity

Size Largest Medium Smallest
Nucleus Ovoidal Ovoidal Rounded
Undulating Less than ½ of the As long as the 2/3 of the costa
membrane costa costa
Inclusion bodies Siderophil granules None None
Sx for dx Urine, vaginal Stool Oral scrappings

swab/discharge/
prostatic secretion
Ciliates
Balantidium coli
Trophozoite Cyst
 Ovoidal/ Bean-shaped
 2 kinds of nuclei (kidney and dot  Ovoid/ rouded

like)
 Cilia is covered with cystic wall
 Covered with cilia
 “Thrown ball like motility”  2 nuclei- macro and micro
 “Rolling motility”
Parts:
 Cytostome
 Macronucleus
 Micronucleus
 Cytopyge
Ciliates
Balantidium coli
Pathology: Ciliary dysentery
Balantidiasis
Ulcer- Flask shaped (rounded base and wide neck)
Tenesmus and diarrhea
Lab Test: Stool Exam

Balantidium coli
Apicomplexa
 No apparent means of locomotion

 Apical complex
 located at the anterior portion of the parasite
 Invasion and penetration of target cells
Asexual
 Complex life cycle
Sexual
 Includes: Plasmodium, Babesia, Toxoplasma, Isospora, Cyclospora,

Cryptosporidium
Plasmodium
 Causes Malaria
 Principal mosquito vector: Female Anopheles flavirostris
 Obligate intracellular parasite
Life cycle includes:
1. Asexual reproduction (SCHIZOGONY) in man
2. Sexual reproduction (SPOROGONY) in arthropod vector

Plasmodium
1. Plasmodium falciparum
 Malignant tertian malaria
2. Plasmodium vivax
 Benign tertian malaria
3. Plasmodium malariae
 Quartan malaria
4. Plasmodium ovale
 Ovale tertian malaria
** Plasmodium knowlesi
 Fifth human malaria parasite
Malaria
A. IS to IH: Sporozoites
IH: Man
Asexual: Schizogony : Schizont:

Merozoite
B.
DH: Mosquito IS to DH: Gametocytes
Sexual: Sporogony : Oocyst: Sporozoites

Asexual Cycle
Vector bites man, injecting sporozoites in
the bloodstream
After a number of erythrocytic

cycles, some merozoites Cryptozoites develop in liver cells to
develop to gametocytes(macro merozoites, completing Exoerythrocytic
and micro) stage
Schizont contains merozoites, Merozoites enter the circulating

released on RBC rupture, each free RBC to become ring
to invade other new cells trophozoites then mature to
schizont
Sexual Cycle
Macrogametocyte and
microgametocyte ingested by
vector in blood meal
Infectious sporozoites
released from oocyst
Gametes released in gut and unite to
migrate to salivary glands of
form zygote (motile ookinete)
mosquito waiting to deliver
to man’s bloodstream
Encystment in gut wall

produce oocyst
Pathogenesis
1. Paroxysm : Chills and fever
a. Every 36 hours: P. falciparum
b. Every 48 hours: P. vivax and P. ovale
c. Every 72 hours: P. malariae
2. Anemia
3. P. falciparum infection- most likely fatal
a. Cerebral malaria- RBC, organisms and pigment block brain vessels
b. Blackwater fever- sudden massive intravascular hemolysis with

resultant hemoglobinuria
Resistance to Malaria
1. Duffy negative phenotype: Fy (a-b-)
 Marker of African Black race
 In P. vivax and P. knowlesi
2. Patient with Sickle Cell Anemia
3. G6PD deficiency
Laboratory Examination
1. Examination of blood film – gold standard
a) Thick blood film- screening purposes; quantify
**Dehemoglobinized Giemsa
b) Thin blood film- identification of species/ stage

**Fix with Methanol Giemsa
2. QBC (Quantitative Buffy Coat) Technique

Fluorescent dye: Acridine orange
3. Rapid Diagnostic Test

a) Optimal assay- parasitic LDH (pLDH)
b) Para Sight F
c) Malaquick - HRP2Ag: Histidine Rich Protein 2 Antigen
4. Serological Test
Blood film
Rapid Diagnostic Tests
 Immunochromatographic methods
Antigens being detected:
a. Histidine rich protein II (HRP II) – P. falciparum
b. Plasmodium lactate dehydrogenase (pLDH)- if P. falciparum or

not, but not among the non- P. falciparum
c. Plasmodium aldolase- all Plasmodium species

RDT
Points of P. falciparum P. vivax P. malariae P. ovale
Differentiation
Age of RBC All types Young RBC Old RBC Young RBC
Growing Usually absent Amoeboid Band form/ Fimbriated,

Trophozoite (larger RBC) Compact ring round, compact
No. of 8-32 12-24 6-12 4-8

merozoites in **in rosette
schizonts **Fruit pie
Stipplings Maurer’s dots Schuffner’s Ziemann’s dots James dots
**Stephen Christopher dots **Schuffner’s
**Ring forms Multiple Single Single Single

infected
Applique or Present Absent Absent Absent
Accole
Gametocytes Macro: Banana Large, round, Large, round, Spherical

shape/ crescent oval oval
Micro: Sausage **ovoid
shape
Plamodium vivax (Trophozoite)
 Growing trophozoite
 Schuffner’s dots- reddish

granules
 Enlarge RBC
Plasmodium vivax
(Schizonts)
 Average number: 16
merozoites
Plasmodium falciparum
(Trophozoite)
 Ring form: Early trophozoite stage
 Normal RBC size

Plasmodium falciparum
(Gametocyte)
 Infective stage to FH

Protozoan Maricelle Manlutac

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PROTOZOANS

Maricelle D. Manlutac, RMT, MPH, MLS (ASCPi)

Sarcomastigophora Ciliophora Apicomplexa

• Balantidium coli • Plasmodium

 Use saline not iodine

 All amoeba are non-pathogenic except E. histolytica

 All amoeba encyst except E. gingivalis

 Cyst is the IS of all amoeba except E. gingivalis

 All amoeba are found in the GIT except E. gingivalis

 **E. gingivalis - their food vacuoles contain cellular debris mostly

Movement Sluggish / Non-Progressive/ Progressive

Karyosome Eccentric Center

Inclusion body Bacteria, debris RBC

Cyst Entamoeba coli Entamoeba histolytica

Number of nuclei Greater than 4 Quadrinucleated

 Sluggish motility  Quadrinucleated

 Blunt pseudopodium  “Cross-eyed appearance”

 Inclusion body: debri, bacteria  Comma shaped chromatoidal

 Large vesicular nucleus

 High affinity with iodine

 “Old man’s face with eye glasses”  Ovoid

 4 pairs of flagella  Axostyle is still present

 Axostyle/ Axial Rod/ Axoneme

Pathology: Travelers' diarrhea

Malabsorption- Steatorrhea ( fats in stool: 5g in 24 hrs)

Excessive flatus – “Rotten eggs”

1. Stool Exam (DFS)

2. Doudenal Fluid – Entero Test: capsule with nylon thread-trophozoite

3. Serological – cyst wall protein 1 (CWP1) antigen

 Prominent axostyle – caudal

 Short undulating membrane

 Rapid jerky tumbling motility

Characteristics T. vaginalis T. hominis T. tenax

Habitat Genitalia Intestine Oral cavity

Inclusion bodies Siderophil granules None None

Sx for dx Urine, vaginal Stool Oral scrappings

 2 kinds of nuclei (kidney and dot  Ovoid/ rouded

Pathology: Ciliary dysentery

Ulcer- Flask shaped (rounded base and wide neck)

Tenesmus and diarrhea

Lab Test: Stool Exam

 No apparent means of locomotion

 Includes: Plasmodium, Babesia, Toxoplasma, Isospora, Cyclospora,

 Principal mosquito vector: Female Anopheles flavirostris

 Obligate intracellular parasite

Life cycle includes:

1. Asexual reproduction (SCHIZOGONY) in man

2. Sexual reproduction (SPOROGONY) in arthropod vector

 Benign tertian malaria

Asexual: Schizogony : Schizont:

Sexual: Sporogony : Oocyst: Sporozoites

After a number of erythrocytic

Schizont contains merozoites, Merozoites enter the circulating

Encystment in gut wall

a. Every 36 hours: P. falciparum

b. Every 48 hours: P. vivax and P. ovale

c. Every 72 hours: P. malariae

3. P. falciparum infection- most likely fatal

a. Cerebral malaria- RBC, organisms and pigment block brain vessels