FLAGELLATES

MODULE 3: PROTOZOOLOGY
 LEARNING OBJECTIVES

➢ Describe the morphology of the parasites under protozoan flagellates.

➢ Discuss the life cycle of each parasites.
➢ Employ appropriate laboratory test for the diagnosis and identification of a parasite.
➢ Differentiate one specie from the other in terms of morphology, distribution, and pathophysiology.
➢ Formulate plans for the prevention and control of parasitic infections.
➢ Explain the importance of studying protozoan flagellates in terms of each acquisition, diagnosis, prevention,
and control.
 LEARNING OUTCOMES

➢Recall the morphology, life cycle, pathogenesis,

epidemiology of different flagellated protozoans.
➢Distinguish the diagnostic features of each flagellated
protozoans.
WHAT IS A PARASITE? ARE FLAGELLATES PARASITES?
HOW ARE THEY CLASSIFIED AND WHAT ARE THE FLAGELLATES OF
CLINICAL IMPORTANCE?

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 PARASITE

A parasite is basically an organism that derives all their benefits from another organism while
harming that organism in the process.

Parasites can be divided into 2 major groups:

the Protozoa

the Helminths.
➢All Protozoa fall under Kingdom Protista.
➢The major organisms causing disease in man belong to
Phylum Sarcomastigophora and Apicomplexa.

➢Under Phylum Sarcomastigophora are 2 subphyla:

1. Subphylum Mastigophora
2. Subphylum Sarcodina REVIEW
➢Subphylum Mastigophora
- Whose organelles of locomotion are whip-like
structures arising from the ectoplasm called flagella.
FLAGELLATE S
CLASSIF IC AT I ON
OF PROTOZOAN
 1. Parasitic (Pathogenic Flagellates):

➢ Intestinal Flagellates
CLASSIFICATION Giardia lamblia
BASED ON ITS
PATHOGENICITY: Dientamoeba fragilis
➢ Genital flagellate
Trichomonas vaginalis
 ➢ Blood and Tissue flagellates
Trypanosoma cruzi
Trypanosoma brucei gambiense
Trypanosoma brucei rhodesiense
CLASSIFIC ATION Leishmania spp.
BASED ON ITS
PATHOGENICITY
II. Non-Pathogenic Flagellates:
* Trichomonas hominis
* Trichomonas tenax
* Chilomastix mesnili
GIARDIA LAMBLIA
 INTESTINAL FLAGELLATES

• GIARDIA LAMBLIA

- also known as Giardia intestinalis or Giardia duodenalis.

- worldwide distribution; more prevalent in warm climates and in children.
- cause epidemic and endemic diarrhea.
- Disease: Giardiasis
- Habitat: duodenum, jejunum, and upper ileum of humans.
- 2 stages: trophozoites and (quadrinucleated) infective cyst stages.
LATERAL VIEW OF GIARDIA
LAMBLIA
LIFE CYCLE
OF GIARDIA
LAMBLIA
 Trophozoite attach to mucosal cells of duodenum and
jejunum through ventral suckers.
* The spectrum of disease produced by Giardia is called
Giardiasis.
* Giardiasis lead to chronic diarrhea and malabsorption of fat
and carbohydrates.
• Diarrhea symptoms are due to the following mechanisms:
PATHOGENESIS - coating of intestinal mucosa by large number of
trophozoites.
- damage to epithelial brush border of the intestinal
mucosa
- alteration in gut activity
- increase secretions of fluid into lumen.
CLINICAL MANIFESTATIONS
 • IP: 1- 4 weeks; average of 9 days
• 50% of infected patients - asymptomatic.
• Acute cases:
- abdominal pain, described as cramping, associated with
diarrhea;
excessive flatus with an odor of “rotten eggs” due to
hydrogen sulfide.
CLINICAL
• Diarrhea in 89% of cases; malaise and flatulence.
MANIFESTATIONS
Spontaneous recovery occurs within 6 weeks in mild to
moderate cases.
• Chronic infection:
- steatorrhea; weight loss, profound malaise, and low-
grade fever.
 Diagnostic Techniques are:

I. Direct fecal smears –

- Demonstration of G. lamblia trophozoites and/or cysts
in stool specimens; trophozoite as having a floating
leaf-like motility.
- Concentration technique is recommended to
DIAGNOSIS detect cysts in stools.
II. If the parasite is not found in the feces, duodeno
jejunal aspiration may be done.
- Examination of the duodenal contents for
trophozoites gives a higher percentage of positive
findings compared to examination of feces.
 III. Enterotest (String test) -
demonstrate Giardia trophozoites.
IV. Antigen detection tests and
immunofluorescent tests are already
available as commercial kits.
A. Immunochromatographic
assays detect the presence of Giardia
antigen in stool.
= Cyst wall protein 1 (CWP1) is
DIAGNOSIS one of the antigens used for these
diagnostic test
B. Direct fluorescent antibody
assays
- the gold standard in diagnosis
as such assays have the highest
combination of sensitivity and
specificity.
 TREATMENT

• Metronidazole
- 250 mg three times a day for 5 to 7 days
- Pediatric dose: 15 mg/kg/day in three divided doses).
- usually well tolerated in adults and has a cure rate of 90%.
• Alternative Drugs:
a. Tinidazole - single dose of 2 g for adults; 50 mg/kg in children)
b. Furazolidone - 100 mg four times daily for 10 days for adults;
Pediatric dose: 6 mg/kg/day in four divided doses for 7 to 10 days).
c. Albendazole - 400 mg/day for 5 days in adults and 10 mg/kg/day for
5 days in children.
- is equally effective as metronidazole at the above doses; not available
in the Philippines.
• Nitazoxanide - has likewise been used effectively in drug-resistant cases.

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• Prompt treatment of asymptomatic - reduce cyst passage and
possible transmission;
• high risk groups:
- food handlers, institutionalized patients, children attending
day-care, and day-care workers.
 EPIDEMIOLOGY

• Giardia has a worldwide distribution.

• In the Philippines, the prevalence: 1.6 to 22.0%; groups in areas with poor
sanitation and hygiene practices - higher prevalence of giardiasis.
• higher in male adults than females, also in other countries.
• Direct oral-anal sexual contact : men who have sex with men
• Outbreaks of giardiasis : reported outside the Philippines. In water-borne
(recreational water or drinking water).
• Foodborne outbreaks.
• The low infective dose, prolonged communicability, and relative resistance to
chlorine - transmission of Giardia through drinking and recreational water, food,
and person-to-person contact.
 • Methods of prevention and control
include:
- proper or sanitary disposal of human
PREVENTION excreta,
AND CONTROL
- avoid contaminated water,
- avoid risky sexual behaviors/ practices
- wash hands with soap and water after
bathroom use and before eating
DIENTAMOEBA
FRAGILIS

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• originally described as amoeba but is actually a flagellate with only the trophozoite stage
known.
• Name Dientamoeba fragilis is derived from the binucleate nature of trophozoite
(Dientamoeba) and the fragmented appearance of its nuclear chromatin (fragilis).
• No cyst stage; this protozoan is closely related to and resembles Trichomonas.
• Habitat: the mucosal crypts of the appendix, cecum and the upper colon.
• The exact life cycle is unknown
• Direct human to human transmission: via the fecal-oral route or via transmission of
helminth eggs particularly that of Enterobius vermicularis serve as a vector.
• stools from macaques, gorillas, and swine were found to carry D. fragilis, thus animal
reservoirs may also be potential sources of human infections
 MORPHOLOGY

• relatively small, varying from 3 - 22 microns in diameter.

• has only a trophozoite stage
• in a permanently stained preparation, one, two or rarely three nuclei can be seen, two
being the most common.
• nuclear chromatin is usually fragmented into three to five granules but these have not
been visualized by Giemsa Stain
• normally no peripheral chromatin on the nuclear membrane.
• cytoplasm is usually vacuolated; may contain ingested debris (mostly bacteria; rarely
RBC’s ) as well as some large uniform granules.
• cytoplasm can also appear uniform and clean with a few inclusions.
PATHOGENESIS
• Organisms infect mucosal crypts of the large intestine that are located close to the mucosal
epithelium, from the cecum to the rectum; however, the cecum and proximal colon are usually
affected. This parasite is not known to be invasive and does not cause cellular damage. It may
invoke an eosinophilic inflammatory response in the colonic mucosa; thus, symptoms are
related to the superficial colonic mucosal irritation. Similar to some other parasites
(eg, Cyclospora cayetanensis, Giardia lamblia, Cryptosporidium parvum), the parasite D
fragilis has been demonstrated to cause disease in humans regardless of their immune status.
CLINICAL MANIFESTATIONS
• Dientamoeba fragilis does not invade tissues
• usually asymptomatic but if symptomatic. the onset of infection: usually accompanied by
loss of appetite, colicky abdominal pain, and intermittent diarrhea with excess mucus,
abdominal tenderness, a bloating sensation, and flatulence.
• 11% of patients has anal pruritus. This is due to co-infection with Enterobius.
• Chronic infection of this organism: mimic the symptoms of diarrhea-predominant
irritable bowel syndrome (IBS),
• experts have suggested ruling out infection with this organism first before diagnosing a
patient as having IBS.
LIFE CYCLE OF DIENTAMOEBA
FRAGILIS
 DIAGNOSIS

• by observation of binucleate trophozoites in multiple fixed and stained fresh

stool samples.
• Fresh stool samples are necessary
• Purged stool specimens provide more suitable material for examination than the average
formed stool.
• Even when formed, D. fragilis may be misdiagnosed as other amoebae. This organism is not
detected by stool concentration methods.
• Prompt fixation of the fresh specimen with polyvinyl alcohol fixative or Schaudinn’s
fixative has been found to be helpful.
 TREATMENT

• Iodoquinol at 650 mg three times daily for 20 days. The pediatric dose is 40
mg/kg/day in three doses, also for 20 days.
• Tetracycline and metronidazole have also been found to be effective.
• Paromomycin, in cases of refractory cases.
 EPIDEMIOLOGY

• The organism has a world-wide distribution with varying infection rates ranging from 0.4
to as high as 42%.
• Unlike majority of parasitic infections, D. fragilis is more prevalent in well-developed
countries as opposed to disadvantaged and resource poor nations.The parasite is also
endemic in crowded communities (i.e institutions), populations with unsatisfactory
sanitation conditions, and individuals who travel to underprivileged countries.[
 PREVENTION AND CONTROL

• Specific recommendations for prevention and control cannot be made until there
is more specific information concerning the method of transmission. However,
prevention rests on limiting oral transmission.
• Personal hygiene and sanitation measures recommended like:
1. Proper hand washing after using the toilet
2. Protect food from flies and cockroaches.
3. Protected sexual practices.
GENITAL FLAGELLATES

TRICHOMONAS VAGINALIS

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• TRICHOMONAS VAGINALIS:
- a uninucleated flagellated protozoa that causes a non-viral sexually
transmitted disease known as Trichomoniasis.
- correlates strongly with the number of sexual partners
most prevalent non-viral sexually transmitted infection.
• Habitat: vagina and prostate; found only in humans; no animal reservoir.
- found in the urogenital tract.
In women: in the vagina but may ascend as far as the renal pelvis.
In men: can be isolated from the urethra, prostate, and less frequently in
the epididymis.
 MORPHOLOGY
SHAPE: Pear-shaped or ovoid

SIZE: About 10 x 7um

AXOSTYLE: Single, situated on midline and runs posteriorly

to the margin of the body, from which it often protrudes.
NUCLEUS: Single, elongated & central

FLAGELLA: 4 anterior flagellae and an undulating membrane TROPHOZOITE

that extends about 2/3rds of its length.
CYTOSTOME: a depression at the anterior end through
which it ingests only food particle.
It exists only as a trophozoite; no cyst form.
 PATHOGENESIS
• Transmission: sexual contact.
• The risk factors include having:
a. Multiple sexual partners
b. A history of other sexually transmitted infections
c. Previous episodes of Trichomoniasis
d. Unprotected sex or sex without a condom.
- T. vaginalis primarily infects the squamous epithelium of the genital tract and
vagina and in the male urethra and prostate; replicates by binary fission.
- exist in trophozoite form--→ does not survive in the external environment;
survive outside human body in wet condition for more than 3 hours.
 CLINICAL MANIFESTATIONS
• Incubation Period: 4-28 days.
• The acute inflammation caused by the parasite results in the characteristic liquid
vaginal secretions, “frothy” greenish or yellow in color, that cover the
mucosa down to the urethral orifice, vestibular glands, and clitoris.
• The vaginal secretions are very irritating; cause intense itchiness and burning
sensation.
• associated with an increased incidence of postpartum endometritis.
• 2% of cases is so-called strawberry cervix described as punctate hemorrhages of the
cervix on speculum examination.
• In males may be latent and essentially asymptomatic. In some cases, it is responsible
for an irritating persistent and recurring urethritis.
 COMPLICATIONS

• Is usually in pregnant women

1. Premature delivery (usually before 37th week)
2. Low birth weight baby
3. Premature rupture of the membrane
4. It can transmit the infection to the baby as the baby passes the birth
canal.
In men:
Moat common: prostatitis
 CLINICAL MANIFESTATIONS

• IN FEMALES:
➢ Watery , foul-smelling “frothy, greenish vaginal discharge
➢ Vulvar itching and burning
➢ Vaginal and cervical mucosa
➢ Tender, reddened, eroded and petechial hemorrhages may be present.
➢ Dyspareunia (painful sexual intercourse)
IN MALES:
➢ Maybe asymptomatic and serve as a carrier
➢ In symptomatic cases prostate, seminal vesicles and urethra may be infected
➢ Discharge thin white urethral discharge
➢ Dysuria ( painful urination)
LIFE CYCLE
LIFE CYCLE OF T.
VAGINALIS
 DIAGNOSIS

Sites sampled to establish a diagnosis of Trichomonas vaginalis:

In women:
• Swab taken from the posterior vaginal fornix at the time of speculum examination; vaginal or
urethral secretions or discharge
• Self-taken vaginal swabs produce similar results to clinician-taken samples when using nucleic acid
amplification tests (NAATs) for diagnosis.
In men:
• Prostatic secretion or semen
• First-void urine, using NAATs.
 DIAGNOSIS

I. Wet Mount Microscopy

a. KOH wet mount
b. Saline wet mount
- is the quickest and most inexpensive way to diagnose trichomoniasis, but the sensitivity of this
technique is low at 50 to 70%.
- sensitivity is highest in women presenting with vaginal discharge.
• KOH wet mount
- done to get the characteristic odor that is Whiff test; production of amines will produce a “fishy”
odor
• Saline wet mount
- to detect the Trichomonas organism showing characteristic pear-shaped with their flagella
and very active motility.
• Detection of vaginal pH – another form of diagnostic method to determine the vaginal pH which is
greater than 4.5 (normal vaginal pH)
 DIAGNOSIS

II. Culture
- has higher sensitivity than microscopy; can detect infection in men.
- was considered the gold standard and it takes 2-5 days.
III. Pap smear may also show trichomonads (sensitivity 60%; specificity 95%).
IV. Molecular Detection
Nucleic Acid Amplification Tests (NAATs)
- recognized as having a much greater sensitivity and specificity,
especially in men and asymptomatic women.
- becoming the current ‘gold standard’.
- can detect T. vaginalis ribosomal RNA in vaginal or endocervical
swabs and urine samples from women and men.
Sensitivities: 88 – 97%; Specificities: 98 – 99%
 DIAGNOSIS

Note:
- Too soon NAAT testing after treatment can result a false positive; due to
detection of remnant trichomonad DNA.
- 2 – 3 weeks post treatment most remnant DNA has cleared.
 Metronidazole or tinidazole

- to be given 2 g as a single dose. (WHO & US CDC)

- Reported cure rate range from 86% to 100%.

TREATMENT -Sexual partners must be treated concomitantly to prevent

reinfection.

- If treatment fails and reinfection is ruled out, a seven-day regimen of

500 mg metronidazole three times a day may be considered.

- If either this regimen fails, a 2 g daily dose for 5 days of either

metronidazole or tinidazole can be used.
- In pregnancy, metronidazole remains the drug of choice for
trichomoniasis.
Sex partners should be treated simultaneously and intercourse should
be avoided till treatment is completed.
 Lactating women:

TREATMENT Metronidazole: withhold breastfeeding during

treatment and for 12 – 24 hrs after last dose.

Tinidazole: interruption of breastfeeding during

treatment and 3 days after the last dose.
 EPIDEMIOLOGY

• Trichomonas infection occurs worldwide and estimate of 170 to 190 million are infected with
Trichomonas.
• Prevalence is higher among women of child-bearing age.
• 5 to 20% of women and 2 to 12% of men in developed countries are infected.
• Higher prevalence is associated with greater frequency of sexual intercourse with multiple
partners and with commercial sex workers.
• is often associated with other sexually transmitted infections.
• In the U. S. study, 70% of male partners of women with trichomoniasis were likewise
infected and the majority of the infected male partners were asymptomatic (77%).
• In the Philippines, the prevalence among sex workers varies according to the method of diagnosis
used. 15% studied with microscope and 37% studied with culture. One study surveyed 421 male
sex workers and there were no positive cases among them based on microscopy.
• In Zimbabwe and South Africa, trial participants diagnosed with trichomoniasis were more likely to
test positive for HIV in their next visit.
• Perinatal transmission of HIV was likewise more likely if the mother had vaginal infections.
 PREVENTION AND CONTROL

• Prevention is best achieved by reducing the risk of exposure, like:

• Limiting the number of sexual partners, and proper use of protective devices such as condoms and
spermicidal foams
• To prevent “pingpong” or recurrent infections, there should be simultaneous treatment of sexual
partners.
• Prompt follow-up of patients and their contacts,
health and sex education about venereal disease are also important.