PROTOZOA (Sarcodina) Sexual:

 SYNGAMY - Sexual cells will unite to form a zygote

PROTOZOOLOGY containing many daughter cells inside.
 Intestinal Amoeba (Pathogenic)  CONJUGATION - two cells attached to one
another; exchange nuclear materials they
 Commensal Amoeba
separate.
 Free-living Pathogenic Amoeba
 Flagellates
Protozoa
 Ciliates
 has only 2 stages:
 Other Intestinal Protozoa
1. Trophozoite – vegetative/ motile stage
 Malarial Parasite
2. Cyst – infective stage
 Blood and Tissue Flagellates – non-motile stage
 Coccidian – non-feeding stage

Protozoa Trophozoite Cyst

 Unicellular (Encystation)
 Motile
 Varies shape, size, locomotion/locomotor Cysts Trophozoite
apparatus method group (Excystation)

Protoplasm Subphylum Sarcodina

- Most important part of protozoa  Is a subphylum of the Phylum
Sarcomastigophora, of unicellular life forms
CYTOPLASM that move by cytoplasmic flow.
1. Ectoplasm  Some species use cytoplasmic extensions
a.) Locomotorapparatus called pseudopodia for locomotion or feeding.
o Pseudopodia or “false feet” – Amoeba o Entamoeba histolytica
o Flagella – Flagellates o Non-pathogenic amoeba
o Cilia - Ciliates o Free Living amoeba
o Undulating membrane – Trichomonas spp.
b.) Structure for Procurement of food
Mouth  CYTOSOME
c.) Structure excretion of metabolic waste
Anus  CYTOPYGE
2. Endoplasm - vital function/processes
 Ribosomes
 Mitochondria
 Nucleus
1. Macronucleus – vegetative function
(response to food)
2. Micronuclues – reproductive function
 Lysosomes
 Vacuole – regulates the osmotic pressure

Reproduction:
Asexual - most of the protozoa reproduce by
 BINARY FISSION - parasite divides transversely.
 MULTIPLE SCHIZOGONY - nucleus divides into
multiple daughter cells
 ENDODYOGENY - single internal body results into
two daughter cells
 INTESTINAL AMOEBIASIS
Entamoeba histolytica
 Most dangerous; pathogenic Asymptomatic/ Chronic Stage
 Produce histolytic enzyme  Hyaluronidase  Presence of cysts in stool
(causes lysis of tissue)  Seen in well-formed stool  cysts stage
HABITAT: Colon, walls of rectum and cecum  Produced AMEBOMA – lesion in intestine
MOT: Ingestion of contaminated food or water
(fecal-oral route), Sexually transmitted Symptomatic/ Acute Amoebic Dysentery
FINAL HOST: Man  Passing out trophozoite
 Bloody mucoid stool with fishy foul odor
4 STAGES:
A. Trophozoite (12-30 um) Acute Stage/ Amoebic Diarrhea
 Movement : unidirectional  1 direction  Trophozoite passer
 Karyosome : centrally located  Watery diarrhea
 Cytoplasm : clean looking  Suffer from tenesmus(painful defecation)
 Ingest red blood cell (Hematophagus
Trophozoite) PATHOGENESIS of Amoebiasis
 Forms lesion (Napkin ring lesion)
1. NON-INVASIVE
B. Pre-cystic stage - Amoeba colony on intestinal mucosa
 Start to form nucleus - Asymptomatic cyst passer
 Throw anything that is not needed 2. INVASIVE
 To survive condition outside the ulcer - Necrosis of mucosa  Ulcers, dysentery
C. CYST STAGE (12-15 um) - Ulcer enlargement  Severe dysentery, colitis,
 Contains 1-4 nucleus (mature cyst) peritonitis
 TETRANUCLEATED CYSTS - Metastasis  Extraintestinal amoebiasis
 Infective stage – 4 nucleus (quadrinucleated
stage)
 Possess a chromatoidal bar (cigar shape/
sausage)  Used as food reserve
 Thick wall
 Resist in harsh condition
D. METACYSTIC TROPHOZOITE
 The tetranucleated cyst will divide to form
trophozoite
MANIFESTATIONS: DIAGNOSIS (Extraintestinal Amoebiasis):
 Ulcer enlargement  severe dysentery
 Perforation of intestinal wall  peritonitis Extraintestinal(hepatic)
 Local abscesses - Symptoms
 2 bacterial infections - History of dysentery
 Occasional ameboma (= amebic granuloma) - Enlarged liver
- Serology (PCR)
 Ameboma= inflammatory thickening of intestinal - Imaging (CT, MRI, ultrasound)
wall around the abscess (can be confused with - Abscess aspiration
tumor) - Reddish brown liquid
- Trophozoites at abscess wall
EXTRAINTESTINAL AMOEBIASIS
 Metastasis via blood stream Culture medium
 Primarily liver (portal vein)  Boeck’s & Drbohlavs Medium
o other sites less frequent  Balamuth’s Medium
 Ameba - free stools common  Chick Embryo Culture
 Diamond’s Culture medium
Amebic Liver Abscess  Schaffer Ryden Fyre Medium
 Chocolate-colored ‘pus’  TYI-S-33 Medium – commonly used for
o necrotic material E.histolytica
o usually bacteria free
 Lesions expand STAINS
 Further metastasis Trophozoite:
 Quensel’s Methylene Blue
Pulmonary Amoebiasis  MIF solution
 Rarely primary Cyst:
 Rupture of liver abscess through diaphragm  Lugols iodine
 2 bacterial infections common  Di Antoni Solution
 Fever, cough, dyspnea, pain
TREATMENT
DIAGNOSIS (Intestinal Amoebiasis):  Asymptomatic
STOOL - Iodoquinolor Paromomycin
Acute: watery, mucoid, bloody, mushy (w/ trophozoite)  Symptomatic
Chronic: Formed stool (w/ cysts) - Metronidazole or Tinidazole
 Drain liver abscess
Techniques:
Acute: DFS (Direct Fecal Smear)
PREVENTION and CONTROL
Chronic:
 Avoid fecal-oral transmission not normally
 Brine Floatation Technique
associated with travelers diarrhea
 ZnSO4 Floatation Technique
 MIFCT (Merthiolate Iodine Formaldehyde
Concentration Technique)
 FECT (Formalin Ether/Ethyl Acetate
Concentration Technique)
 DFS (Direct Fecal Smear)

SEROLOGICAL TEST:
 IFAT (Indirect Fluorescent Antibody Testing)
 IHAT (Indirect Hemagglutination Test)
 EIA (ELISA) – Enzyme Immunoassay (Enzyme
Linked Immunosorbent Assay)
 PCR (Polymerase Chain Reaction)
 Non - Pathogenic Amoebas

 Entamoeba dispar
 Entamoeba hartmanii
 Entamoeba coli
 Entamoeba gingivalis
 Entamoeba polecki  The cysts measure usually 11 to 15 μm(range 9
 Entamoeba moshkovskii to 18 μm) and their shape varies from spherical
 Endolimax nana to oval.
 Iodamoeba butschlii  The trophozoites measure usually 15 to 20
 Blastocystis hominis μm(range 10 to 25 μm).

COMMENSAL AMOEBA
Entamoeba coli
Entamoeba dispar
 Cyst
 Same morphologic features as Entamoeba
- Mature, infective cyst in stool specimen,
histolytica but has genetic and biochemical
- Large, spherical or ovoidal shaped cyst, 15-
differences
25 micrometer in
 Attributed as non-pathogenic Entamoeba diameter
histolytica strain - Usually contains 8 nuclei,
 E. dispar is negative to PCR(Gal/GalNac lectin) but 16 nucleated cyst can
while E. histolytica is positive to PCR be found sometimes
(Gal/GalNac lectin) - Each nuclei has eccentric
karyosome
Entamoeba hartmanni - Peripheral nuclear chromatin asymmetrically
 Cysts distributed, giving an uneven thickness look
- 6-8 um to nuclear membrane
- 4 nuclei (mature) - Sometimes, chromatoid body with splintered
- blunt chromatoid bodies ends(look-like a witch broomstick) can be
- CB persist in mature cysts seen in cytoplasm
 Trophozoites
 Trophozoites (20-25 um)
- 8-10 um
- Broad blunt pseudopodia
 Nuclear structure
- Movement : Sluggish
- Peripheral chromatin
- Karyosome : Eccentric
- Small karyosome
- Cytoplasm : Dirty looking
 Small race of E. histolytica
 Small in size  E. coli is bigger than E. histolytica

Entamoeba moshkovskii Endolimax nana

 Identical morphology as E. histolytica  Cysts  “cross eyed cysts”
 Free-living (sewerage) - 6-8 mm
 Temperature: Difference w/ E. histolytica - 4 nuclei
o E. moshkovskii  0 - 41C  Trophozoites
o E. histolytica  20 – 40 C - 5-12 um

Entamoeba polecki
 Rarely found in humans
 Commensal of pigs & monkeys
 Mature cyst has one nucleus
Iodamoeba bütschlii Blastocystosis
 Cysts  Gastrointestinal pathology is controversial
- 10-12 mm  Majority not associated with symptoms
- 1 nucleus  Some produce a wide variety of intestinal
- Glycogen vacuole disorders: abdominal cramps, irritable bowel
 Trophozoites syndrome, bloating, flatulence, mild to moderate
- 12-15 mm diarrhea without fecal WBC or blood, nausea,
 Nuclear structure vomiting, low grade fever and malaise
- No peripheral chromatin  Among immunosuppressed individuals
 Nucleus: Achromatic granule
Diagnosis:
 Fecal examination
Blastocystis hominis o Hematoxylin or trichrome staining is used
 Inhabitant of the lower intestinal tract of to differentiate the various stages
humans and other animals Treatment:
 Formerly classified as yeast under the genus Metronidazole, Iodoquinol, Trimetophrim-
Schizosaccharomyces sulfamethoxazole
 Proposed life cycle but need validation
 Reproduction: Binary Fission
 Known to occur in six morphological forms : Entamoeba gingivalis
(1) vacuolated, (2) amoeba-like, (3) granular,  Found in oral cavity
(4) multiple fission, (5) cyst and (6)  No cyst stage
avacuolarform  Trophozoites nearly identical to E. histolytica
 Main type causing diarrhea – Vacuolated form  Periodontal disease  pyorrhoea –
 spherical, 5-10 μm inflammation of gums
 Have a large, central vacuole that pushes the
cytoplasm and nuclei to the periphery of the
cell
 Can retrieve people with HIV

Amoeba-like forms
- occasionally observed in stool samples
- Exhibit active extension and retraction of
pseudopodia
- Intermediate stage between the vacuolar and
pre-cystic form
- Ingest bacteria to enhance encystment
Granular forms
- Mainly observed in old cultures
- 10-60 μm
- Granular content develop into daughter cells of
the ameba-form when the cell ruptures
Multiple fission
- Arise from vacuolated forms
- Believed to produce many vacuolated forms
 Free-living Pathogenic Amoeba Laboratory Diagnosis
 In Naegleria infections, the diagnosis can be
Naegleria fowleri made by microscopic examination of
 Ubiquitous genus found in fresh water lakes and cerebrospinal fluid (CSF).
ponds  A wet mount may detect motile trophozoites,
 PAM (Primary Amoebic Meningoencephalitis) and a Giemsa-stained smear will show
first recognized by Fowler (1965) trophozoites with typical morphology.
o initially thought to be Acanthamoeba
 Naegleria fowleri is only species associated with PREVENTION
PAM  Avoid swimming on stagnant water
 Free-living amoebo-flagellate, can exist as an  Chorination cannot kill N. fowleri
amoeba (trophozoite and cyst form) and as a
flagellate (swimming form)
 Trophozoite is with characteristic Acanthamoeba spp.
lobosemonopseudopodium and a very  Ubiqitousamoeba of the soil and water
prominent nucleus with a centrally-located  Human cases first reported in the early 70's
karyosome  Majority of patients are chronically ill,
 Mode of Transmission: entry into the olfactory immunocompromised, or debilitated with other
neuroepithelium, nasal cavity and olfactory bulbs diseases
 Infective stage: trophozoites and cyst  A small free-living amoeba characterized by an
active trophozoite stage and a dormant cyst stage
 Trophozoites exhibit a characteristically single and
large nucleus and small, spiny filaments for
locomotion (acanthopodia)

Clinical Features
 Acute primary amebic
meningoencephalitis(PAM) is caused by
Naegleria fowleri.
 1-14 days incubation period
 symptoms usually within a few days after  Cysts are spherical, 15-20 μm in diameter, having a
swimming in warm still waters thick double wall. The outer wall may be spherical
 rapid clinical course, death in 4-5 days after or wrinkled, the inner wall appear stellate or
onset of symptoms polyhedral. (Acanthamoebatrophozoitesand a cyst,
 4 known survivors treated with Amphotericin B (trichromestain).
 It presents with severe headache and other
meningeal signs, fever, vomiting, and progresses
rapidly (<10 days) and frequently to coma and
death.

SYMPTOMS:
 Frontal headache
 Fever w/ congested nasal passages  Mode of Transmission: Entry through the eyes,
 Irritation of the meninges nasal passages to the lower respiratory tract and
 Kernig’s sign through ulcerated or broken skin
 Nausea, vomiting, disorietation  Infective stage: cyst and trophozoite
Acanthamoebaspp.: free living amoebae of the Other Intestinal Protozoan:
Acanthamoeba genus cause two clinical syndrome:
1) Granulomatous amoebic encephalitis (GAE) Dientamoeba fragilis trophozoite
2) Amoebic keratitis - A disseminated form of GAE  Dientamoeba fragilisis a flagellate that must be
is described in individuals with the Acquired morphologically differentiated from the amebas,
Immunodeficiency Syndrome (AIDS) especially Endolimax nana and Entamoeba
hartmanni.
Acanthamoeba Meningoencephalitis  Trophozoites usually measure 9 to 12 μm (range 5
 Portal of entry through respiratory tract (inhalation to 15 μm).
of cysts) or wounds in skin  Most trophozoites are typically binucleate, some
 Presumed hematogenous dissemination to the have only one nucleus.
CNS  Trophozoitein stool specimes, Trichrome stained
 Infection associated with immunosuppression  No cystic stage
 Onset is insidious with headache, personality  Karyosome centrally placed in the nucleus and
changes, slight fever characteristically made of several granules
 Progresses to coma and death in weeks to months
 Cranial nerve palsies, Hemiparesis, ataxia, seizure

Amoebic Keratitis
 Predisposing factors
- ocular trauma
- contact lens (contaminated cleaning solutions)
 Symptoms
- ocular pain
- corneal lesions
 Treatment
- difficult, limited success
 Diagnosis
- Tissue biopsy (histological examination):
trophozoites, cyst
o Brain, corneal scrapings

TREATMENT
Supportive:
1. Rifampicin & AmphothericinB
2. 5-fluorocytosine
3. Itraconazole
4. Ketoconazole
5. Pentamidine
6. Neomycin
7. Miconazole