Intestinal and

Extraintestinal Amebae
College of Medical Laboratory Science
Our Lady of Fatima University-Valenzuela

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 PROTOZOA
• Proto ( first), Zoa (animals)
• Single cell having a true nucleus, eukaryotic cells
• Unicellular organism that is composed of:
– Nucleus
– Cytoplasm
• Endoplasm: inner (often granulated), dense part
• Ectoplasm: outer (non-granulated), typically watery
– Structures for locomotion: pseudopodia, flagella, cilia,
undulating membrane
– Plasma membrane: controls secretions and excretions
– Cytostome: cell mouth
– Chromatoidal bodies: storage for glycogen protein

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 PROTOZOA
• Two stages commonly encountered:
– Trophozoite
• vegetative
• motile
• infective stage
• obtained from semi formed stool
– Cyst
• non-motile stage
• resistant- stage of certain protozoa
• transfer of infective stage
• has a thick wall
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 PROTOZOA
Encystation: stage forming a cyst or becoming enclosed to a
capsule, this event takes place in the rectum of the host as
feces are dehydrated or soon after the feces have been
excreted.
Excystation: escape from cyst or envelope, produces a
trophozoite from the cyst stage, and it takes place in the
large intestine of the host after the cyst has been ingested.
• Reproduction:
– Binary fission (asexual)
– Conjugation (Sexual)

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http://www.notesonzoology.com/protozoa/entamoeba-etymology-reproduction-and-life-cycle/1573
 Classification of Protozoan Parasites
• Phylum Sarcomastigophora

Subphylum Sarcodina Acathamoeba Entamoeba dispar

Endolimax nana Entamoeba gingivalis
Entamoeba coli Entamoeba histolytica
Iodamoeba butschlii Naegleria fowleri

Subphylum Mastigophora Chilomastix mesnili Dientamoeba fragilis

Giardia lamblia Trichomonas vaginalis
Trichomonas hominis Trichomonas tenax
Trichomonas vaginalis

Leishmania braziliensis Leishmania donovani

Leishmania tropica Trypanosoma cruzi
Trypanosoma brucei complex

• Phylum Ciliophora Balantidium coli

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 Classification of Protozoan Parasites (cont.)

• Phylum Apicomplexa Babesia spp

Cryptosporidium hominis
Cyclospora cayetanesis
Isospora belli
Plasmodium spp.
Toxoplasma gondii

• Phylum Microspora Enterocytozon bineusi

Encephalitozoon spp.
Vittaforma cornea
Pleistophora spp.
Brachiola vesicularum
Microsporidium spp.

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Phylum Sarcomastigophora
Subphylum Sarcodina

GENERAL RULES FOR AMEBAE

• All Entamoeba are commensal except for Entamoeba histolytica
• With pseudopodium (false feet) : finger-like structures for
movement
• Undergoes ENCYSTATION except for E. gingivalis and Dientamoeba
fragilis.
• Inhabits the large intestine except for E. gingivalis (gums)
• Amebiasis – presence of amoeba in any part of the body
(exclusively applied to E. histolytica)
• Asexually multiplies through Binary fission

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 Entamoeba histolytica
Morphologic Forms:
1. Trophozoite – divides through “binary fission”, capable of encystation
(overpopulation, pH change, food supply, availability of oxygen)
2. Precyst
3. Cyst – w/ protective thick cell wall (hyaline), capable of excystation
4. Metacyst

Infective stage : cyst (except for E. gingivalis)

MOT: ingestion of contaminated food and/or water with E. histolytica
cyst
TROPHOZOITE CYST
Vegetative & motile stage Non-motile, non-feeding stage
(feeding stage)
Found in watery, soft or semi Found in soft to formed stool
formed stool
Fragile Resistant to acidic pH 9
Life cycle of E. histolytica

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Morphologic comparison between E. histolytica and E. coli
A. Trophozoite
Point of E. histolytica E. coli
differentiation
Movement Unidirectional, progressive Sluggish, non-progressive
and non-directional
Shape of Fingerlike Blunted
pseudopodia
Manner or release of One at a time/ explosive Several at a time
pseudopodia
Nucleus uninucleated (central uninucleated (eccentric
karyosome) karyosome)
Inclusions RBC Bacteria, yeast, debris

Cytoplasm Clean looking Dirty looking

size bigger smaller

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Morphologic comparison between E. histolytica and E. coli

B. Cyst

Point of E. histolytica E. coli

differentiation
No. of nuclei Quadrinucleated More than 4

Chromatoidal bar Sausage, rod, cigar-shaped Broomstick, splinter-like

Nuclear membrane thin thick

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 TROPHOZOITE
Entamoeba histolytica Entamoeba coli

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 CYST
Entamoeba histolytica Entamoeba coli

www.cdc.gov

www.studyblue.com 14
 Pathogenesis
Symptoms

• Gradual onset of abdominal pain

• Diarrhea (with or without blood and mucus)
• In children, bloody diarrhea, fever and abdominal pain
• Abscess formation à Amoebic liver abscess

Pathology

Amoebic dysentery vs. Bacilliary Dysentery

Presence of Charcot-Leyden crystals found microscopically in the

stool in cases of Amoebic dysentery.

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 Pathogenesis
Amoebic dysentery Bacillary dysentery
Onset Gradual Acute
Signs/ Symproms No significant fever or Fever and usually vomiting
vomiting
Odor of feces Offensive, Fishy odor Odorless
Blood and mucus (+) Often watery and bloody
pH Acidic Alkaline
Pus cell/ PMN/ Neutrophils Few Numerous
Cellular exudates Scant Massive
Pyknotic residues Numerous Few
Charcot Leyden crystals Present Absent
Pathogenic amoeba Present Absent
Bacteria Few Numerous
Macrophages Absent Present

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Pathology
Common associated disease: Intestinal amebiasis, amebic colitis,
amebic dysentery, extraintestinal amebiasis
• Can cause ulceration “flask-shaped ulcer” in the intestines (cecum,
ascending colon and sigmoid)
Pathogenic determinants / Virulence factors:
1. Galactose-inhibitable adherence lectin (Gal Lectin): receptor
mediated adherence of amoeba to target cells
2. Amoeba spores
3. Cystein Proteinase: most important, tissue invading factor

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Extraintestinal amoebiasis

• Through the portal vein (liver), trophozoite reach other parts of

the body (liver, brain, lungs, kidneys)
a. amebic pneumonitis
b. venereal amebiasis (penile or vaginal amebiasis)

Asymptomatic carriers: cysts becomes unnoticed, ameba reproduce

but infected individual shows no clinical symptoms

Mode of transmission
Ingestion of contaminated food and/or water

Diagnostic Stage: identification of the cyst or trophozoite

Sample for ID: stool (examined within 30 minutes from collection)

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Laboratory Diagnosis:
1. Direct Fecal Smear
- saline solution : trophozoite motility
- saline + methylene blue : Entamoeba spp. stain blue
- saline + iodine : nucleus of E. histolytica can be
observed
2. Concentration Techniques
- Formalin Ether/ Ethyl Acetate Concentration Technique (FECT)
- Merthiolate Iodine Formalin Concentration (MIFC)

3. Culture : Robinson’s and Inoki medium

(more sensitive than stool microscopy but not routinely available)

4. Serologic Testing: ELISA, CIE, AGD, IHAT (can detect past infection),
IFAT

5. Molecular Testing : PCR

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Treatment
• To cure invasive disease at both intestinal and extraintestinal
site and to eliminate passage of cyst from intestinal lumen
Metronidazole : drug of choice for invasive amebiasis (Tinidazole
and secnidazole are also effective)
Diloxanide furoate: drug of choice for asymptomatic cyst passers
Percutaneous drainage of the liver abscess: Patients who do not
respond to metronidazole and need prompt relief of severe pain
Prevention and Control
• Proper hygiene
• Provision for sanitary disposal of human feces
• Improve access to clean and safe drinking water
• Good food preparation practices
• Avoid using “night soil”
• Food handler should be examined for cyst carriage
• Health education and promotion
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Non-Pathogenic species

1. Entamoeba
• Spherical nucleus with distinct nuclear membrane
lined with chromatin granules
• Small karyosome found near center of the nucleus
2. Endolimax
• Vesicular nucleus with a very large, irregularly
shapes karyosome anchored by achromatic fibrils
3. Iodamoeba
• Large, chromatin-rich karyosome surrounded by a
layer of achromatic globules and anchored to the
nuclear membrane by achromatic fibrils

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Non-Pathogenic species

Entamoeba Endolimax Iodamoeba

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Non-Pathogenic species

1. Entamoeba coli
• Harmless inhabitant of the colon
• Cysts: Size (12 – 25 microns)
• larger than E. histolytica
• Consists of 8 nuclei with very diffuse karyosomes
• May become hypernucleate with 16-32 nuclei
• May also contain needle-like chromatoidal bodies with irregular
fragmented/sharp/splintered ends
• Trophozoites: Size (18-28 microns)
• Smaller than E. histolytica
• Has one nucleus containing large, diffuse karyosomes
• Peripheral chromatin is usually dense and irregular
• Cytoplasm is usually rough and contain few to many ingested
debris

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Non-Pathogenic species

1. Entamoeba coli

Motility Sluggish, not progressive and non-directional

Pseudopodia Short and blunt; granular; slowly extruded

Inclusions Bacteria and other material; no RBCs ingested

Nucleus Rarely visible

Nuclear Thick; lined with coarse chromatin dots and
membrane bars
Karyosome Large; location is eccentric; surrounded by a
halo of non-staining material

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Non-Pathogenic species

2. Entamoeba dispar
• Morphologically similar to E. histolytica, but with different DNA and
RNA

3. Entamoeba hartmanni
• Similar to E. histolytica except much smaller and no RBC inclusions
• “small-race E. histolytica”

4. Entamoeba polecki
• Parasite of the pigs and monkeys (rarely infect humans)
• Entamoeba chattoni: found in apes and monkeys, identical to E.
polecki, identification via isoenzyme analysis

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Non-Pathogenic species
5. Entamoeba gingivalis
• Can be found in the mouth (gum and teeth surfaces)
• Abundant in cases of oral diseases
• No cyst stage, does not inhibit the intestines
• Transmission through kissing, droplet spray, sharing utensils

6. Entamoeba moshkovskii
• morphologically indistinguishable from those of the disease-
causing species E. histolytica and the non-pathogenic E. dispar,
but differs from them biochemically and genetically. Although
sporadic cases of human infection with E. moshkovskii have been
reported, the organism is considered primarily a free-living
amoeba.
• physiologically unique: osmotolerant, able to grow at room
temperature and able to survive at 0-41°C
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Non-Pathogenic species

7. Endolimax nana
• “Smallest amoeba”
• “Cross eyed cyst” – 4 eccentric nuclei

Motility Sluggish; random

Pseudopodia Blunt; hyaline

Cytoplasm Contain food vacuoles with ingested bacteria

Karyosome Large; central or eccentric in location; irregular

outline

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Non-Pathogenic species

8. Iodamoeba butschlii
• “iodine-cyst” because of its affinity to iodine
• Large glycogen vacuole/ body which stains deeply with iodine
• Uninucleated – resembling a “basket of flowers” shape

Motility Sluggishly progressive; with hyaline

pseudopodia
Inclusions Bacteria scattered throughout the cytoplasm;
RBCs are never ingested
Nucleus Not visible

Karyosome Large; centrally located; irregularly rounded;

surrounded by a layer of small granules

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 Endolimax nana cyst

Iodine stain Trichrome stain

Iodamoeba butschlii cyst 29

FREE LIVING PATHOGENIC AMOEBA

Acanthamoeba spp. (Acathamoeba castellani)

• Ubiquitous, free-living ameba
• With an active trophozoite stage with characteristic prominent
“thorn-like” appendages (acanthopodia) and resilient cyst stage
• Aquatic organism, can survive in contact lens cleaning solutions
• Most common ameba of freshwater and soil
• No flagellate state

MOT:
• aspiration or nasal inhalation: use of contaminated swimming
pools, deep well, etc.
• Direct invasion of the eye: contaminated saline

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FREE LIVING PATHOGENIC AMOEBA

Pathogenesis:
• Granulomatous Amoebic Encephalitis (GAE)
- destructive encephalopathy and associated meningeal irritation
- disease of the immunocompromised (AIDS)
- laboratory diagnosis: made post-mortem in most cases

• Amoebic keratitis (contact lens users)

- perforation of the cornea and results to subsequent loss of vision
- laboratory diagnosis: epithelial biopsy or corneal scrapings,
demonstration of cyst and trophozoites in stool ,PCR

Specimen: discharges, exudates, tissue secretions

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FREE LIVING PATHOGENIC AMOEBA
Naegleria fowleri
• Free-living protozoan with two vegitative forms: an ameba
(trophozoite form) and a flagellate (swimming form)
• Thermophilic organism that thrive best in hot springs and other
warm aquatic environment
• True pathogen

Pathogenesis:
• Fatal Primary amoebic encephalitis (PAM)
- patients initially complain of fever, headache, sore throat, nausea
and vomiting
- Hemorrhagic necrosis in post mortem examination of infected brain
- “Kernig’s sign”à diagnostic sign for meningitis where the patient is
unable to fully straighten his or her leg when the hip is flexed at 90
degrees because of hamstring stiffness
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FREE LIVING PATHOGENIC AMOEBA

MOT:
• Oral and intranasal routes while swimming in contaminated pools,
rivers and lakes

Diagnosis:
• PCR
• ELISA
• Bacteria-seeded agar culture (active trophozoites within 24 hrs)

Specimen: CSF

Prevention:
• Frequent cleaning
• Chlorination
• Salination
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Phylum Ciliophora
CILIATES
Balantidium coli
- Causative agent of “balantidiasis or balantidial dysentery”, similar to
amoebic dysentery
- Largest protozoan parasite
- Only parasitic ciliate
- Primarily associated with pigs
Morphology:
- Has trophozoite and cyst stage
Parts:
cytostome : entry of food
www.southampton.ac.uk
cytophage : excretes waste
2 disimilar nucleus (macro- and micronucleus)
2 contractile vacuoles
* encystation doesn’t result in increase in nuclei

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Pathogenic determinant:
- Hyaluronidase: causes the ulceration, secreted by trophozoite
- Ulceration is described as flask-shaped ulcer but with rounded
base and wider neck

MOT: ingestion of food/water contaminated with B. coli cyst

Specimen: stool
Diagnosis: microscopic demonstration of cyst and trophozoite in
direct fecal smear or concentration techniques, biopsy and bronchial
washing (pulmonary infection)
Infective stage: cyst
Treatment: Metronidazole
Prevention: same with other amoeba

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 REFERENCES
Belizario, V. and De Leon, W. (2015). Philippine Textbook
of Medical Parasitology. Third Edition. University of the
Philippines Manila. Ermita, Manila.

Mikhail A. Valdescona, RMT, MPH. PAR313 Lecture.

Our Lady of Fatima University. Valenzuela City.

Trinidad, J.A (2014). Lecture Notes for Parasitology Board

Review . Pioneer Educational review Center. Recto, Manila.

Domingo, E. (2012). Lecture Notes for Parasitology. Our

Lady of Fatima University. Valenzuela City.

http://www.cdc.gov/parasites/amebiasis/

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