The Protozoa
Introduction:
The Phylum Protozoa is classified into four
subdivisions according to the methods of
locomotion.
 The amoebae (Sarcodina) move by means of
pseudopodia.
 The flagellates (Mastigophora) typically move by
long, whiplike flagellae.
 The ciliates (Ciliata) are propelled by rows of cilia
that beat with a synchronized wavelike motion.
 The sporozoans (Sporozoa) lack specialized
organelles of motility.
The Protozoa
General:
 There are about 45,000 protozoan species; around
8000 are parasitic, and around 25 species are
important to humans.
 Diagnosis - must learn to differentiate between the
harmless and the medically important. This is most
often based upon the morphology of respective
organisms.
 Transmission - mostly person-to-person, via fecaloral route; fecally contaminated food or water; other
means include sexual transmission, insect bites or
insect feces.
The Protozoa
General:


Trophozoite - the motile vegetative stage;
multiplies via binary fission; colonizes
host.
Cyst - the inactive, non-motile, infective
stage; survives the environment due to
the presence of a cyst wall. Cysts do not
multiply, however, some organisms divide
within the cyst wall.
The Protozoa
Diagnostic Features:
 Nuclear structure - important in species differentiation.
 Size - helpful in identifying organisms; must have
calibrated objectives on the microscope in order to
measure accurately.
 Cytoplasmic inclusions - chromatoid bars (coalesced
RNA); red blood cells; food vacuoles containing
bacteria, yeast, etc.
 Appearance of cytoplasm - smooth & clean or
vacuolated.
 Type of motility - directional or non-directional; sluggish
or fast.
The Protozoa
Nuclear Structure:
Chromatin - nuclear DNA. Present as “peripheral”
chromatin and the karyosome.
 Karyosome - a small mass of chromatin within the
nuclear space. Also called “endosome” or
“centrosome.”
 Peripheral Chromatin - chromatin adhering to the
nuclear membrane.
 Nuclear membrane - membrane surrounding all
nuclear material.
Chromatoid body or “bar” - coalesced RNA within
the cytoplasm of the cyst stage.

The Protozoa
Intestinal Protozoa - The Amoebae
General Life cycle  The definitive host ingests the infective cyst stage from
fecal contamination in environment.
 The cyst passes into the small intestine & excystation
occurs with transformation to the trophozoite stage.
 Trophozoites colonize the host, multiplying asexually
via binary fission. They can remain near the lumen (nonpathogens) or invade the wall of the intestine & multiply
(pathogens).
 Cysts and trophozoites are passed in the feces of the
infected host.
The Protozoa
Intestinal Protozoa - The Amoebae
Entamoeba histolytica

Epidemiology - Occurs worldwide; the
highest incidence and prevalence is in
areas with poor sanitation.

Pathology and Clinical Manifestations - the
most pathogenic of all; causes amoebic
dysentery; can become extra-intestinal; can
be fatal. Hepatic abscess is the most
common and dangerous complication.

Chronic infections may last for years; often
confused with colitis, cancer.

Distribution - worldwide, mostly in tropics
and sub-tropics.
The Protozoa
Intestinal Protozoa - The Amoebae
Entamoeba histolytica

Morphology & Laboratory Identification trophozoites range 12 to 30 microns in
diameter; nucleus has an even distribution of
peripheral chromatin and a small, compact,
centrally located karyosome; cytoplasm is
smooth and granular; inclusions, if present,
are red blood cells; cysts range 10 to 20
microns in diameter and contains four nuclei
when mature. Cigar-shaped chromatoid bars
may be present in some cysts.
The Protozoa
Intestinal Protozoa - The Amoebae
Entamoeba hartmanni



Epidemiology - similar to E. histolytica
Formerly called the “small race” of
Entamoeba histolytica.
Technologists must be able to
differentiate this organism from E.
histolytica because E. hartmanni is
non-pathogenic.
The Protozoa
Intestinal Protozoa - The Amoebae
Entamoeba hartmanni
 Morphology & Laboratory Identification This organism is morphologically
similar to E. histolytica. The difference
lies in the sizes of the respective
organisms. Trophozoites will measure
less than 12 microns, while cysts will
measure less than 10 microns.
The Protozoa
Intestinal Protozoa - The Amoebae
Entamoeba coli


Significance - this is a harmless commensal;
must be differentiated from pathogens.
Morphology - trophozoites range from 10 to 35
microns in diameter; cysts range from 10 to 30
microns in diameter and contain 8 to 16 nuclei
when mature; the nucleus exhibits an
eccentric karyosome with irregular, coarse
chromatin. The cytoplasm is heavily
vacuolated, containing yeast, bacteria, and
debris.
The Protozoa
Intestinal Protozoa - The Amoebae
Entamoeba gingivalis
 Infective site - the mouth; the organism
thrives in diseased gums, but is not
considered a causal agent. It is
destroyed in stomach if swallowed.
 Transmission - contact with fomites
(drinking glasses, eating utensils, etc.);
kissing.
 Morphology - resembles E. histolytica,
but has no cyst stage. It is the only
species which ingests leucocytes.
The Protozoa
Intestinal Protozoa - The Amoebae
Endolimax nana
 Occurrence - occurs in about 14% of
the US population; 21% worldwide.
 Pathogenicity - none.
 Morphology - trophozoites range from 5
to 10 microns in diameter. The nucleus
contains a large, blot-like karyosome;
there is little or no peripheral chromatin.
Cysts are usually sub-oval, measuring 4
to 6 by 6 to 10 microns.
The Protozoa
Intestinal Protozoa - The Amoebae
Iodamoeba butschlii
 Pathogenicity - none.
 Morphology - the cyst is often
called the “iodine cyst” due to the
presence of a large glycogen
vacuole which stains dark brown
with iodine.
The Protozoa
Naegleria fowleri trophozoite in CSF
Tissue Dwelling Amoebae
Naegleria fowleri

Classification - an ameboflagellate; a freeliving organism alternating between amoeboid
and flagellated forms; only the amoeboid form
is found in tissues.

Life cycle - the amoeba gains entry via the
nasal mucosa, usually during a swimming
event; it moves along the olfactory nerve,
gaining access to the brain via the cribriform
plate. Cases are invariably fatal. Infections do
not spread from person-to-person.
The Protozoa
Tissue Dwelling Amoebae
Naegleria fowleri
 Symptoms - Dramatic and rapidly
progressive. Headache, fever, nausea
& vomiting occur within 1 to 2 days.
Meningoencephalitis, irrational
behavior, coma & death usually occur
within 9 days of exposure.
 Diagnosis - Usually made at autopsy.
CSF contain motile amoebae. Early
diagnosis is critical. Amoebae in CSF
specimens can be cultured on nonnutrient agar containing bacteria.
The Protozoa
Tissue Dwelling Amoebae
Acanthamoeba spp.

Life cycle - also a free-living amoeba. The
amoeba reaches the brain hematogenously
after entering a wound or lesion on the skin.
More commonly, the organism is associated
with getting into eyes via contaminated or
homemade cleaning solutions.

Symptoms - slow onset (10 or more days).
Presents as chronic, granulomatous lesions in
brain. In eye lesions, the infection resembles a
herpes virus infection.

Acanthamoeba keratitis - associated with users
of extended-wear contact lenses.
The Protozoa
Superclass Mastigophora - the flagellates. Inhabit the mouth,
bloodstream, gastrointestinal, or urogenital tracts.
Morphological Characteristics

Flagellum(ae) - organelles of locomotion; an extension of
ectoplasm; moves with a whip-like motion.

Axostyle - a supporting mechanism, a rod-shaped structure; not all
flagellates have these.

Undulating membrane - a protoplasmic membrane with a flagellar
rim extending out like a fin along the outer edge of the body of
some flagellates.

Costa - a thin, firm rod-like structure running along the base of the
undulating membrane.

Cytosome - a rudimentary mouth; also referred to as a gullet.
The Protozoa
Superclass Mastigophora - the flagellates.
Identification of a flagellate is based upon:
 Size.
 Shape.
 Motility.
 Number and morphology of nuclei.
 Number and location of flagellae.
 Location in the body of the host.
The Protozoa
Intestinal flagellates
Giardia lamblia

Most common protozoan parasite in the U.S.A.

Life cycle - man ingests cysts from fecally
contaminated environment; the organism
excysts in the upper intestine; trophozoites
multiply and attach to the intestinal mucosa;
often enter the gall bladder. Trophozoites and
cysts are passed in the feces.

Diagnosis - identification of cysts or
trophozoites in stool specimens or duodenal
contents.
The Protozoa
Intestinal flagellates
Giardia lamblia
Morphology - very distinctive. Dorsalventrally flattened, and Bi-laterally
symmetrical.
 Cyst - 9 x 12 micrometers and contain 2
to 4 nuclei; parabasal bodies are present.
 Trophozoite - Four pairs of flagella - one
pair located anterior, two pair located
ventrally, and one pair located
posteriorly. An axostyle and parabasal
bodies are present.
The Protozoa
Intestinal flagellates
Giardia lamblia

Epidemiology - prevalence 1 to 30%, common
in children’s day care centers; can be
transmitted in water. Sexual transmission has
been well documented.

Pathology and Clinical Manifestations symptoms can be severe; diarrhea, foulsmelling, greasy, mucus-laden stools,
flatulence, nausea, cramps. Most infections
are asymptomatic; chronic cases experience
weight loss, malabsorption of fat, protein,
folic acid, and fat-soluble vitamins.
The Protozoa
Intestinal flagellates
Dientamoeba fragilis
 General - Formerly classified as an
amoeba; electron microscopy and
immunological studies have suggested
a flagellate nature.
 Laboratory diagnosis - detection of binucleated trophozoites; fragmented
karyosomes consisting of 4 to 8
granules of chromatin.
The Protozoa
Intestinal flagellates
Dientamoeba fragilis
Diagnostic stage - the trophozoite
in feces. There is no cyst stage.
 Morphology - 1 or 2 nuclei, with
little or no peripheral chromatin;
karyosome is divided into 4 to 8
distinct granules.
 May rarely ingest rbc’s.
The Protozoa
Intestinal flagellates
Dientamoeba fragilis
 Pathology - infection is usually
asymptomatic; can be associated
with diarrhea, anorexia, abdominal
pain.
 Association with pinworm - the
organism may be transmitted from
host to host within the egg of
Enterobius vermicularis.
 Distribution - worldwide, there is a 1%
to 20% prevalence.
The Protozoa
Intestinal flagellates
Chilomastix mesnili
 A non-pathogen - must be
differentiated from Giardia.
 Found in cecum and colon.
 Transmission - by ingestion of
mature cysts.
The Protozoa
Intestinal flagellates
Chilomastix mesnili
Morphology  Trophozoite - 4 flagella (3
anterior, 1 associated with the
cytostome; one nucleus,
always located anteriorly.
 Cyst - lemon shape; 1 nucleus;
cytostome may be seen.
The Protozoa
The Trichomonads
Characteristics  Undulating membrane - protoplasmic
membrane with flagellar rim extending
out like a fin along outer edge of body.
 Flagella - several in a tuft, provides
locomotion.
 Axostyle - functions for support.
 Costa - firm rod-like structure running
along base of the undulating
membrane.
 Cytostome - rudimentary mouth.
The Protozoa
The Trichomonads
Trichomonas hominis
 Commensal - must differentiate from
pathogens.
 Transmission - direct person-to-person
fecal transmission; no cyst stage.
 Morphology - “arc-shaped” exhibits a
wobbly, jerky, motility.
 Must differentiate from T. vaginalis - in
instances where feces is contaminated
with urine.
The Protozoa
The Trichomonads
Trichomonas vaginalis
 Life cycle - trophozoite lives in the
vagina, urethra, epididymis, and
prostate; multiplies via longitudinal
fission; no cyst stage.
 Mode of infection - sexual intercourse
or fomites.
 Diagnosis - identification of
trophozoites in body fluids (wet
mounts of discharges) or on PAP
smears.
The Protozoa
The Trichomonads
Trichomonas vaginalis
 Pathology - Females: vaginal discharge;
burning, Itching, or chafing. Frequency
of urination or dysuria. Males: frequently
asymptomatic. If the prostate is
involved, the patient may develop
discharge, dysuria, and enlargement of
prostate with tenderness.
 Morphology - has an axostyle and short
undulating membrane that extends less
than half the body length; 4 flagellae.
The Protozoa
Class Ciliophora - The Ciliates
Balantidium coli
 Epidemiology - Rarely found in USA.
This is the only ciliate parasite of
humans.
 Largest parasitic protozoan trophozoite is 30-120 x 25-125 microns;
the cyst averages 50 - 70 microns in
diameter.
 Life cycle - The cyst is ingested via
fecal contamination in environment;
cysts excyst in the small intestine;
trophozoites migrate to large intestine.
The Protozoa
Class Ciliophora - The Ciliates
Balantidium coli
 Pathology & Symptoms - Many
infections are asymptomatic, organism
feeding on bacteria at surface of
mucosa. Severe infections - with the aid
of hyaluronidase, the organism burrows
into submucosa, producing ulcers.
Symptoms - dysentery, abdominal pain,
nausea & vomiting, fever, headache.
 Diagnosis - Diagnosed by observing
cysts & trophozoites in fecal samples.
The Protozoa
Class Ciliophora - The Ciliates
Balantidium coli
 Morphology - Large, oval shape;
two nuclei, 1 large kidney shaped
(macronucleus) & 1 small
micronucleus (micronucleus not
often seen); body surface covered
by longitudinal rows of cilia;
cytostome present.
 Primary animal reservoir - pigs,
monkeys.
The Protozoa
Intestinal Coccidia
General  Organisms infecting humans include Isospora,
Sarcocystis, Cryptosporidium, Cyclospora &
Toxoplasma.
 Some have 2-host life cycle.
 Sexual & asexual reproduction - Schizogony - asexual
binary fission; Sporogony - sexual reproduction.
 Diagnostic stages are often difficult to locate. Acid fast
stains are used to visualize. Oocysts do not stain with
iodine or permanent stains like trichrome.
The Protozoa
Intestinal Coccidia
Sarcocystis suihominis
Sarcocystis bovihominis
Sarcocystis lindemanni



Pathology - Sarcocystis bovihominis & S.
suihominis are intestinal infections. Sarcocystis
lindemanni is a muscle infection.
Humans are definitive hosts for S. suihominis &
bovihominis; intermediate hosts are pig (sui-) and
cow (bovi-); humans are intermediate hosts for S.
lindemanni; the definitive host is unknown.
Infective stage - sarcocysts in meat (intestinal);
ingestion of oocysts from animal feces (muscle).
The Protozoa
Intestinal Coccidia
Isospora belli
 Definitive host - humans.
 Schizogony - takes place upon initial
infection when sporozoites invade the
intestinal epithelium and multiply.
Sporogony - progeny of asexual
reproduction initiate development into
gametes (male & female); fertilized
gametes develop into oocysts which are
passed as the infectious stage in the
feces.
The Protozoa
Intestinal Coccidia
Isospora belli
 Diagnostic/infective stage - mature
oocyst - contains two sporocysts, each
of which contain four sporozoites (do
not stain with iodine).
 Intermediate hosts - none.
 Pathology - ranges from asymptomatic
to acute, severe dysentery.
The Protozoa
Intestinal Coccidia
Cryptosporidium parvum




Site of infection - primarily an intestinal
infection, can become systemic in AIDS
patients.
Originally considered an animal parasite
(rodents, cattle and sheep). Organisms from
some (fish, fowl and reptiles) do not infect
humans.
Infections in immunocompromised people can
be serious, even fatal.
Identification - oocysts are 2 - 5 microns in
diameter; do not stain with iodine; and are
acid-fast.
The Protozoa
Intestinal Coccidia
Cryptosporidium parvum


Transmission - the endemic cycle is maintained
via person-to-person, fecal-oral route
transmission; a relatively common finding in
“day-care” diarrhea; can be sexually
transmitted; big potential for being waterborne
due to significant resistance to disinfectants.
Pathology - most infections cause severe
diarrhea. In the immunosuppressed patient, the
condition is protracted and life threatening.
There is no drug effective against this parasite.
The Protozoa
Intestinal Coccidia
Cyclospora cayetanensis


General - The first outbreak in the USA
occurred in medical residents in 1980; a large
multi-state outbreak occurred the summer of
1996. During this outbreak, Texas had more
than 100 cases over a period of about six
weeks, while no more than one case had been
reported during any previous year.
Transmission - fecally contaminated food or
water. The 1996 outbreak was associated with
contaminated raspberries.
The Protozoa
Intestinal Coccidia
Cyclospora cayetanensis



Animal reservoirs - not known. Other
Cyclospora species are known to infect
a variety of animals, but C. cayetanensis
is the name designated for the only one
known to infect humans.
Pathology - diarrhea similar to that
experienced with Cryptosporidium
infections.
Identification - Oocysts are 10 microns
in diameter, and are variably acid-fast.
The Protozoa
Microsporidium spp.
 General - at this time, infections are
thought to be limited to AIDS patients.
 Transmission - person-to-person, via
the fecal-oral route; can be sexually
transmitted.
 Identification - a very small organism,
spores average 1.0 by 1.7 microns.
They stain a reddish-pink color with
Chromotrope stain. Some stained
spores exhibit a dark staining “belt”
across the middle of the organism.