PROTOZOAN INFECTIONS

M L S 0 4 2 L EC T UR E
CLI N I CA L PA R A SI TOLGY
P R E PA RED BY: DI A DE M CR U Z, R M T, M S C
Intestinal amebae
Entamoeba histolytica
➢ Most invasive

➢ Cause colitis and liver abscess

➢ The trophozoites are highly motile and possess pseudopodia

Microscopic examination (stool specimens)

▪ Progressive and directional movement of trophozoites

Trophozoites

▪ Hyaline pseudopodium : clear, glasslike ectoplasm, or outer layer is extruded, and the granular endoplasm flows into it
(Ingested red blood cells)

▪ Ability to colonize and invade the large bowel, while cysts are never found within invaded tissues.

▪ Multiply by binary fission.

Cysts

▪ Spherical, and the size may vary from 10 to 20 μm.

▪ Highly refractile hyaline cyst wall, one to four nuclei, and rod-shaped (or cigar-shaped) chromatoidal bars.
Entamoeba histolytica
➢ Consists of two stages: an infective cyst and an invasive
trophozoite form.

➢ The quadrinucleate cyst is resistant to gastric acidity and

desiccation, and can survive in a moist environment for several
weeks.

➢ Ingestion from fecally-contaminated material

Pathogenesis and Clinical Manifestations

➢ Mechanisms for virulence are: production of enzymes or other

cytotoxic substances, contact-dependent cell killing, and
cytophagocytosis.

➢ Trophozoites adhere to the colonic mucosa through a galactose

inhibitable adherence lectin (Gal lectin) -> kill mucosal cells by
activation of their caspase-3, leading to their apoptotic death
engulfment.
Entamoeba histolytica
Diagnosis

➢ Microscopic detection: trophozoites and cysts in stool specimens

➢ Ideally, a minimum of three stool specimens collected on different days should be examined.

➢ The detection of E. histolytica trophozoites with ingested red blood cells is diagnostic of amebiasis.

➢ Charcot-Leyden crystals can also be seen in the stool.

Treatment and Prognosis

➢ To cure invasive disease at both intestinal and extraintestinal sites

➢ To eliminate the passage of cysts from the intestinal lumen.

➢ Metronidazole is the drug of choice

Epidemiology

➢ Humans are the major reservoirs of infection.

➢ MOT: Ingestion of food and drink contaminated with cysts from human feces, and direct fecal-oral contact

➢ Amebic infection is prevalent in the Indian subcontinent, Africa, East Asia, and South and Central America.
Commensal amebae
Entamoeba hartmanni
➢ Smaller trophozoite than E. histolytica (3-12 um)

➢ Mature cysts measure 4 to 10 μm, and have rod-shaped

chromatoid material with rounded or squared ends.

➢ Does not ingest red blood cells

Entamoeba coli
Trophozoites

➢ Measure 15 to 50 μm in diameter.

➢ More vacuolated or granular endoplasm with bacteria and debris, but no red blood cells

➢ A narrower, less-differentiated ectoplasm

➢ Broader and blunter pseudopodia used more for feeding than locomotion

➢ More sluggish, undirected movements

➢ Thicker, irregular peripheral chromatin with a large, eccentric karyosome in the nucleus
Entamoeba coli
Cyst

➢ its larger size (10 to 35 μm in diameter)

➢ More nuclei (eight versus four in E. histolytica)

➢ More granular cytoplasm

➢ Splinter-like chromatoidal bodies.

➢ Iodine staining reveals dark-staining, perinuclear masses, which are glycogen.

Entamoeba polecki
➢ Parasite found in the intestines of pigs and monkeys. Rarely, it can infect
humans

➢ Motility of trophozoites is sluggish. A small karyosome is centrally located in the

nucleus.

➢ Cyst is consistently uninucleated, and chromatoidal bars are frequently angular

or pointed.

➢ Prominent nuclear membrane and karyosome

Entamoeba gingivalis
➢ Can be found in the mouth. lives on the surface of gum and teeth, in
gum pockets, and sometimes in the tonsillar crypts (Oral disease).
➢ Transmission is most probably direct: through kissing, droplet spray, or
by sharing utensils
Trophozoite
➢ Measures 10 to 20 μm.
➢ It moves quickly, and has numerous blunt pseudopodia.
➢ Food vacuoles that contain cellular debris (mostly leukocytes, which is
characteristic of this species) and bacteria are numerous.
No cyst stage
Trophozoites

➢Small, with a diameter of 5 to 12 μm, and exhibit sluggish movement.

➢Blunt, hyaline pseudopodia, and the nucleus has a large, irregular karyosome.

➢Food vacuoles found in the cytoplasm may contain bacteria.

Cysts

➢ Quadrinucleated when mature.

Iodamoeba butschlii
Trophozoite (averages 9 to 14 μm in diameter )

➢ Large, vesicular nucleus with a large, central karyosome, surrounded by achromatic granules.

➢ No peripheral chromatin granules on the nuclear membrane.

Cyst

➢ 9 to 10 μm in diameter (ranging from 6-16 μm),

➢ Uninucleated, and has a large glycogen body

Diagnosis
❑ Stool examination

❑ Formalin ether/ethyl acetate concentration technique (FECT) and iodine stain are useful to
differentiate the species.

❑ For E. gingivalis, a swab between the gums and teeth is examined for trophozoites.

❑ Cysts are recovered from formed stools, while trophozoites are recovered from watery or semi-
formed stools.

❑ Trophozoites are best demonstrated by direct fecal smear.

❑ In recovering cysts, the use of concentration techniques like FECT and zinc sulfate flotation is
useful
Free-living Amebae
Acanthamoeba spp.
➢ Ubiquitous, free-living ameba that is the etiologic agent of Acanthamoeba keratitis (AK) and
granulomatous amebic encephalitis (GAE).

➢ Active trophozoite stage with characteristic prominent “thorn-like” appendages

(acanthopodia)

➢ Highly resilient cyst stage into which it transforms when environmental conditions are not
favorable.

➢ It is an aquatic organism that is found in a myriad of natural and artificial environments, and
can survive even in contact lens cleaning solutions.

Trophozoites

➢ Exhibit a characteristic single large nucleus with a centrally-located, densely staining nucleolus

➢ Large endosome

➢ Finely granulated cytoplasm

➢ Large contractile vacuole

➢ Small, spiny filaments for locomotion known as acanthapodia

Acanthamoeba spp.
➢ No flagellated stage exists as part of the life
cycle.
➢ The trophozoites replicate by mitosis (nuclear
membrane does not remain intact).
➢ IS: trophozoites
➢ Entry can occur through the eye, the nasal
passages to the lower respiratory tract, or
ulcerated or broken skin
Diagnosis
➢AK = epithelial biopsy or corneal scrapings for
recoverable ameba with characteristic staining
patterns on histologic analysis.
➢GAE = usually made post mortem in most cases.
Naegleria fowleri
➢ Two forms of trophozoites: ameboid and ameboflagellate

Ameboid (10 to 35 µm; rounded = 10 to 15 µm in diameter)

➢ Cytoplasm is granular and contains many vacuoles.

➢ Single nucleusis large and has a large, dense karyosome

➢ Lacks peripheral chromatin

➢ Thermophilic organisms which thrive best in hot springs and

other warm aquatic environments.

Pathogenesis and Clinical Manifestations

➢ Rapidly destructive and fatal meningoencephalitis termed

primary amebic meningoencephalitis (PAM).

➢ Survive in elevated temperatures and reproduces rapidly in

temperatures above 30°C.
Naegleria fowleri
Diagnosis

➢ Compatible history

➢ PCR and immunostaining

➢ Serology utilizing ELISA

Treatment

➢ Amphotericin B in combination with clotrimazole

Prevention and Control

➢ Easily killed by chlorination of water at 1 ppm or higher.

Ciliates and Flagellates
Balantidium coli
Trophozoite (30 to 150 μm long and 25 to 120 μm wide)

➢ Locomotion = cilia arranged in a longitudinal pattern extending from the oral to the
caudal region

➢ Cytostome = an oral apparatus, which it acquires food

➢ Cytopyge = which it excretes waste.

➢ Two dissimilar nuclei = macronucleus (bean-shaped) and micronucleus (round)

➢ Two contractile vacuoles that act as osmoregulatory organelles.

➢ Extrusive organelles called mucocysts which are located beneath the cell membrane.

Cysts (40 to 60 μm in diameter)

➢ Spherical to slightly ovoid in shape and measure

➢ Covered with thick cell walls (double-walled).

➢ Unlike amebae, encystation does not result in an increase in number of nuclei.

Balantidium coli
➢ MOT: ingestion of food and/or water contaminated with cysts.

➢ Incubation period = 4 to 5 days.

➢ Ingested cysts excyst in the small intestines and become

trophozoites.

➢ Cause pathologic changes in the colonic wall and mucosa.

➢ Reproduction occurs asexually (asymmetric binary fission) or

sexual reproduction (conjugation)

➢ IS: Cysts

Pathogenesis and Clinical Manifestation

➢ Attacking the intestinal epithelium and creating a characteristic

ulcer with a rounded base and wide neck

Diagnosis

➢ Direct examination or concentration (sedimentation or flotation)

techniques
Giardia duodenalis
A flagellate that lives in the duodenum, jejunum, and upper ileum.

Asexual life cycle (trophozoites and quadrinucleated infective cyst stages)

Trophozoites (9 to 12 µm long by 5 to l5 µm wide)

➢ Pyriform or teardrop shaped, pointed posteriorly, with a pair of ovoidal nuclei, one on
each side of the midline.

➢ The dorsal side is convex, while the ventral side is concave with a large adhesive disc
used for attachment.

➢ Bilaterally symmetrical, with a distinct medial line called the axostyle.

➢ Erratic tumbling motion by four pairs of flagella arising from superficial organelles in
the ventral side of the body

➢ Longitudinal binary fission and are found in diarrheic stools.

➢ Covered with variant-specific surface proteins (VSPs)

Giardia duodenalis
Cysts (8 to 12 µm long by 7 to 10 µm wide.)

Ovoid; young cyst (two nuclei), mature cysts (four nuclei)

flagella retracted into axonemes, the median or parabasal body, and

deeply stained curved fibrils surrounded by a tough hyaline cyst wall
secreted from condensed cytoplasm

MOT: Transferred to the mouth via contaminated hands, food, or

water.

IS: mature cysts

Pathogenesis and Clinical Manifestations

➢ 1 to 4 weeks (average of 9 days) for the disease to manifest

➢ Acute cases (abdominal pain, cramping, associated with diarrhea).

There is also excessive flatus with an odor of “rotten eggs” due to
hydrogen sulfide.

➢ Chronic infection (steatorrhea)

Giardia duodenalis
Diagnosis

➢ Diagnosis is made by demonstration of trophozoites and/or cysts in stool specimens.

➢ Trophozoites (direct fecal smears) = having a floating leaf-like motility.

➢ Cysts in stools = concentration techniques

➢ Duodeno-jejunal aspiration

➢ Enterotest®

➢ Antigen detection tests and immunofluorescent tests

Treatment

➢ Metronidazole

➢ Alternative: tinidazole, furazolidone, Albendazole

Trichomonas vaginalis
Only in the trophozoite stage.
➢ Pyriform shape, measuring 7 to 23 µm
➢ Four free anterior flagella that appear to arise from a
simple stalk, and a fifth flagellum embedded in the
undulating membrane.
➢ Median axostyle and a single nucleus.
➢ Found in the urogenital tract/ vagina/ renal pelvis
/urethra / prostate/ epididymis
➢ Multiply by binary fission
➢ MOT: sexual intercourse.
Trichomonas vaginalis
Pathogenesis and Clinical Manifestations
➢ Inflammation of the vaginal mucosa
➢ liquid vaginal secretions (greenish or yellow in color)
➢ Include secondary bacterial infection of the urogenital tract
➢ Punctate hemorrhages of the cervix, the so-called strawberry
cervix
➢ In males (asymptomatic) = urethritis, prostatitis
Diagnosis
➢ Saline preparation of vaginal fluid
➢ The accepted gold standard is culture which takes 2 to 5 days
Treatment
➢ Metronidazole or tinidazole
Non-Pathogenic Flagellates
Trichomonas hominis
➢ Occurs only as a trophozoite

➢ Pyriform shape and measures 7 to 13 µm.

➢ Five anterior flagella and a posterior flagellum projecting from an undulating membrane.

➢ Cytostome and the nucleus are situated at the anterior end.

➢ Axostyle extends from anterior to posterior along the mid-axis.

➢ MOT: fecal contamination of food and drinks.

➢ Habitat: cecal area of the large intestine of human and other primates.

➢ Non-invasive

➢ Pass out with diarrheic stools.

Trichomonas tenax
➢ Pyriform flagellate (only in the trophozoite stage): 5 to 12 µm,

➢ Four free equal flagella and a fifth one on the margin of an undulating
membrane which does not reach the posterior end of the body, and
lacks a free posterior extension.

➢Single nucleus and a cytostome (Binary fission)

➢ MOT: Droplet spray from the mouth, kissing, or common use of

contaminated dishes and drinking glasses.

➢ Habitat: tartar around the teeth, in cavities of carious teeth, and in

necrotic mucosal cells in the gingival margins.

➢ Resistant temperature change; survive for several hours in drinking

water.

➢ Diagnosis = swabbing the tartar between the teeth, the gingival

margin, or tonsillar crypts.

➢ Pulmonary trichomoniasis (w/ the presence of bacteria)

➢ Treatment: metronidazole
Chilomastix mesnili
➢ Habitat: cecal region of the large intestine.

➢ It has well-defined trophic and cystic stages.

➢ Trophozoite: asymmetrically pear-shaped (spiral groove extending through the

middle half of the body) - 6 to 10 µm.

➢ Boring and spiral forward movement

➢ Cyst is pear- or lemon-shaped, broadly rounded at one end and somewhat

bluntly conical at the other end which has a knob-like protruberance

➢ MOT: ingestion of cysts in food and drinks.

➢ Diagnosed by microscopic examination of feces and demonstration of

trophozoites or cysts.

➢ Preventive and control measures: sanitation and personal hygiene

END