COMMENSAL AMEBAE

Significance of commensal amebae in stool:

1. They may be mistaken for the pathogenic E. histolytica
2. Indication of fecal contamination of food or water

Parasite biology:
Entamoeba

 Spherical nucleus with distinct nuclear membrane

lined with chromatin granules and small karyosome
found near the center of the nucleus.
 Trophozoites usually have only 1 nucleus

Endolimax

 Has vesicular nucleus with relatively large and

irregularly-shaped karyosome anchored to the nucleus
by achromatic fibrils

Iodamoeba

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  Characterized by a large chromatin-rich karyosome
and surrounded by a layer of achromatic globules and
anchored to the nuclear membrane by achromatic
fibrils

Stages of development:
1. Trophozoite
2. Precyst
3. Cyst
4. Metacystic trophozoite
Note:
Exception of E. gingivalis

NON PATHOGENIC AMEBAE

Entamoeba dispar
 Morphologically similar to E. histolytica, but their DNA and ribosomal RNA are different.
 Its isoenzyme pattern is different from that of E. histolytica
 In reference diagnosis laboratories, molecular analysis by PCR-based assays is the method of choice for
discriminating between the pathogenic species (E. histolytica) and the nonpathogenic species (E.
dispar).

Agarose gel (2%) analysis of a PCR diagnostic test for differentiation between E. histolytica and E. dispar

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 Lanes 1 - 4: Amplification with the Psp3/Psp51 PCR primer pair specific for
E. histolytica.  Diagnostic band size - 876 bp.
Lanes 6 - 9: Amplification with the NPsp3/NPsp51 PCR primer pair specific
for E. dispar.  Diagnostic band size - 876 bp.
Lanes 1 and 6: E. histolytica 200:NIH, zymodeme II (positive control for E.
histolytica).
Lanes 2 and 7: E. dispar 351:IMMiT, zymodeme I (positive control for E.
dispar).
Lanes 3 and 8: Specimen from a patient with a liver abscess (positive with
E. histolytica primers and negative with E. dispar primers).  E. histolytica
333:IMMiT, zymodeme XIV.
Lanes 4 and 9: Specimen from an asymptomatic patient (positive with E.
dispar primers and negative with E. histolytica primers).  E. dispar
389:IMMiT, zymodeme I.
Lane 5: Molecular base pair standard, 100-bp ladder (from 600 to 1,000
bp).

Entamoeba hartmanni
 morphologic characteristics similar to E. histolytica (except trophozoites have a max diameter of 12 μm
and cysts have a max diameter of 10 μm, and cysts often have a single nucleus).
 Historically, E. hartmanni has been called the small race of E. histolytica.
 Smaller than E. histolytica and does not ingest RBC
 More sluggish in movement
 Mature cyst measure from 5-10um, quadrinucleated and have a coarse cytoplasm

 Immature cysts usually have chromatoidal bars, which may be short with tapered ends, or thin and
bar-like.
Note: Differentiation requires careful measurement of a representative sample of organisms with a
properly calibrated ocular micrometer.

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Entamoeba coli
 Cosmopolitan in distribution; harmless inhabitant of the colon.
 Troph can be differentiated from E. histolytica by the ffg:
a. more vacuolated/granular endoplasm with bacteria and debris but no RBC
b. narrower, less differentiated ectoplasm
c. broader and blunter pseudopodia
d. more sluggish undirectional movements
e. thicker, irregular peripheral chromatin with a large eccentric karyosome in the nucleus.

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Entamoeba coli

Trophozoite Cyst
 More vacuolated or granular endoplasm with  Large size
bacteria and debris but no rbc  Greater # of nuclei
 Narrow, less differentiated cytoplasm  More granular cytoplasm
 Broader, blunter pseudopodia  Splinter-like chromatoidal bodies
 More sluggish, undirected movements
 Thicker irregular peripheral chromatin with large
eccentric karyosome in the nucleus

Entamoeba polecki
 Parasite of pigs and monkeys
 Rarely infect humans
 Cyst is consistently uninucleated
 In stained fecal smear – nuclear membrane and karyosome are very prominent

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Entamoeba gingivalis

 Resides in the mouth ( gum and teeth surface,

gum pockets)
 Troph is 10-20 µm
 No cyst stage
 Moves quickly - numerous pseudopodia
 Lives on the gum and teeth surface, gum pockets
& tonsillar crypts.
 Abundant in cases of oral disease
 Transmission:
a. kissing
b. droplet spray
c. sharing utensils

Note:
E. gingivalis – a swab between gums and teeth is examined for trophozoites.

Endolimax nana
 smallest ameba to infect humans (6-15um)
Trophozoites:
- sluggish movement, and the characteristic Endolimax nucleus-large irregular karyosome.
Cysts:
- contain 4 nuclei (although smaller numbers may be seen).
-Cysts are easily differentiated from those of other amebae but may be confused with Blastocystis
hominis.

Iodamoeba butschlii
Trophozoite:
 9-14um long (ranging from 6-20um)
 Identified by its large vesicular nucleus with large endosome surrounded by achromatic granules
 No peripheral chromatin granules on the nuclear membrane
Cyst:
 Uninucleated; has a large glycogen body which stains deeply with iodine
 Diagnosis
 Examination of feces or tissues: ( 3 negative SE using a sensitive conc tech)
a. Formed stool is examined with saline or iodine for cyst
b. Liquid or semi-formed stools is examined for trophozoite

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  Treatment
No treatment is necessary because these amebae do not cause disease.
 Prevention and Control
Infection may be prevented through proper disposal of human waste and good personal hygiene.

FREE LIVING PATHOGENIC AMEBAE

Acanthamoeba
Small free-living ameba characterized by:
-active trophozoites
-dormant cyst
Ubiquitous organism.
Trophozoite:
Feed on Gram(-) bacteria, blue-green algae or yeasts.
Reproduce by binary fission.
Exhibits small, spiny filaments for locomotion known as acanthapodia.
Moves sluggishly with polydirectional movement.
Can be cultivated in PYGC(proteose-peptone, yeast extract, glucose and cysteine) medium containing
antibiotics.

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 Cyst Trophozoite

Pathogenesis and Clinical Manifestations

(GAE) granulomatous amebic encephalitis
- occurs in chronically ill and debilitated individuals, those with impaired immune defense mechanisms,
and some who are under immunosuppressive therapy.

Diagnosis
Diagnosis of Acanthamoeba encephalitis is made only after death in the majority of cases.
Specific diagnosis depends on demonstrating the trophozoites or cysts in tissues using histopathologic stains
and microscopy.

Treatment
-5-fluorocytosine
-Ketoconazole

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 -Itraconazole
-Pentamidine
-Amphotericin B

Prevention and Control

Boiling water is the best possible way of killing the trophozoites and cysts.
Cleaning should be done with masks if there is threat of infection.

Naegleria
 Free-living amebo-flagellate

exist as an:
a. ameba(troph form)
b. flagellate (swimming form).

Life Cycle:
trophozoite stage that can transform reversibly into non-reproductive flagellate stage or a resistant
cyst.

Non pathogenic species:

N.gruberi
N.philippinensis

Mode of transmission:
a. Oral and intranasal routes while swimming in contaminated pools, lakes, and rivers.
b. In arid regions through inhalation of dust-borne cyst

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 N.fowleri – pathogenic specie; able to survive at 46°C.
Agent of the following diseases:
-Gastritis
-Diarrhea
-Primary Amebic Meningoencephalitis (PAM)

Diagnosis
-Presence of trophozoites in the brain and CSF (PAM)
-Culture method
-PCR and ELISA
Treatment
-Amphotericin B
-Clotrimazole
-Azithromycin

Prevention and Control

a. No known means of prevention and control of Naegleria infection.
b. Avoid diving into and swimming in warm and stagnant freshwater pools, water discharge pools,
unchlorinated poorly maintained swimming pools, or mud-lined lakes and ponds.

OTHER INTESTINAL PROTOZOANS

Blastocystis hominis

 Inhabitant of lower intestinal tract

 Formerly classified as yeast (genus Schizosaccharomyces)
 Lacks cell wall but has mitochondrion with protozoan morphology.

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Reproduction:

a. asexual – binary fission

b. sporulation – under strict anaerobic conditions.

Optimal growth - 37◦C in the presence of bacteria.

5 Morphological forms of Blastocystis hominis

Forms Characteristics
Vacuolated -most predominant forms in fecal specimens
-spherical in shape (5-10um in dm)
-a large central vacuole pushes the cytoplasm and nuclei to the periphery.
-main type of Blastocystis causing diarrhea.
Ameba-like -occasionally observed in stool samples.
-exhibit active extension and retraction of pseudopodia.
-appears to be an intermediate stage between the vacuolar form and pre-cystic
form.
Granular -observed from old cultures.
-dm varies from 10-60um.
-granular contents develop into daughter cells of the ameba-form when the cell
ruptures.

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 Multiple fission -forms arise from vacuolated forms.
-it is believed that these multiple fission forms produce many vacuolated forms.
Cyst & avacuolar -size is about 3-55um & has a very prominent & thick osmophilic electron dense
wall.
-assumed that the thick-walled cyst might be responsible for external
transmission while those cysts with thin-walls might be the cause of reinfection
within a host’s intestinal tract.

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