Parasites and Diarrhea. I: Protozoans and Diarrhea
Rayhan Hashmey, Robert M. Genta, and A. Clinton WhiteJr.
Intestinal parasitism is extremely common, with
approximately 70% of all people harboring one or more
intestinal parasite. Parasitism and diarrhea are both hyperendemic in areas where sanitation is suboptimal. Many
clinicians assume that the identification of intestinal parasites in patients with diarrhea implies that the parasites
are the cause. This approach is frequently misguided.
Some intestinal parasites such as Giardia lamblia and Entamoeba histolytica certainly do cause diarrhea. Others, for
example Entamoeba coli and Ascaris lumbricoides, almost certainly do not. In addition, there are a number of other
organisms that have been associated with diarrheal illness in some cases, which may or may not be important
pathogens.
In this article, we will review the role ofprotozoans
as definite and possible causes of diarrhea. In Part 11, we
review the role of helminths in diarrhea.
Protozoans as Definite Causes of Diarrhea
Table 1 lists the major protozoan parasites that have
been associated with diarrhea. G. lamblia and E. histolytica are well-known causes of diarrheal illness while
Cryptosporidium, Isospora belli, microsporidia and Cyclospora
species have been increasingly recognized in association
with diarrhea, especially in patients with AIDS. Balantidium
coli is a well-recognized but less fiequent cause of diarrhea. Blastocystis horninis and Dientamoebafvagilis have also
been identified in diarrheal illness, but their causal role
is less clear. Other organisms such as Entamoeba dispar
almost certainly do not cause diarrhea.
Giardiasis
Organism and Life Cycle
G. lamblia is a binucleate, flagellated intestinal protozoan. Giardia are primitive eukaryotes and lack mitochondria.The life cycle is simple and consists of cyst and
trophozoite stages. The trophozoite, which measures
9-21 pm long, is tear-drop shaped and on stained smears
has a characteristic face-like appearance, due to the presence o f symmetrically placed nuclei with large
karyosomes.The dorsal surface is convex, while the ventral surface has a cytoskeletal disc made up of microtubules
linked by microribbons.'.* Four pairs of posteriorly
directed flagella help in 1ocomotion.Trophozoites multiply by longitudinal binary fission. Encystation occurs
in the small bowel and is facilitated by neutral pH and
bile salts. Ingesting as few as 1G25 mature cysts can establish infection. Excystation requires a low p H as well as
the presence of pancreatic enzymes.3
Epidemiology
G. lamblia is one of the major causes of parasitic diarrhea worldwide. In the developing world giardiasis is pandemic, with peak prevalence rates of up to 20% in
children less than 10 years of age.4 In the United States
G. lamblia is among the most frequently isolated intestinal parasites and has been identified in up to 7.2% of stool
specimens submitted for parasitologic examination.'a6
Giardiasis often occurs in the setting of waterborne outbreaks.' It is an important cause of chronic diarrhea in
travelers returning from developing countries.' Most
studies estimate infection rates of 1-3% in short-term visitors to endemic area^.^.^ In a large series from Switzer-
Table 1 Protozoan Parasites and Diarrhea
Dejnite Causes of Diarrhea
Causing bloody dnrrhea / dysentery
Entamoeba kistolytica
Balantidium coli
Causing watery diarrhea / malabsorption
Giardia lamblia
Cryptosporidium parvum
isospora belli
Cyclospora cayetanensis
Microsporidia
Possible Causes of Diarrhea
Rayhan Hashmey, MD, Robert M. Genta, MD, and A. Clinton
White Jr., MD: Departments of Medicine, Pathology, and
Microbiology and Immunology, Baylor College of Medicine,
Houston, Texas.
Reprint requests: A. Clinton White Jr., MD, Department of
Infectious Diseases, Baylor College of Medicine, Rm 561E.
One Baylor Plaza, Houston, TX 77030.
J Travel Med 1996;4:17-31.
Dientamoeba fragilis
Blastocystis hominis
Not Associated with Diarrhea
Entamoeba dispar
Entamoeba coli
Entamoeba hartmanni
iodamoeba biitschlii
Endolimax nana
Trichomonas horninis
18
land, G. lamblia was found in 0.4% of returning travelers
who had chronic diarrhea."'Travelers usually get infected
after consumption of contaminated water." Although
infrequent, several outbreaks of giardiasis have been associated with contaminated f ~ o d . ~ *Person-to-person
-'~
transmission is also important, especially in groups such
as children in day care centers, sexually active male
homosexuals, and institutionalized patient^.'^ These
groups may have very high cyst carriage rates, up to 50%
in some studies, and can transmit disease to friends and
family.'' Animal-to-human transmission has been postulated as a common source ofinfection but has only been
documented in a Canadian outbreak traced to beavers."
Pathology and Pathogenesis
The trophozoite adheres to the mucosal surface
using the ventral disc. This results in distortion of the
micmvih and disruption of the brush b ~ r d e r . ' ~ . ' ~ T
parhe
asite also elaborates several cytopathic substances including cysteine pmteinases, enterotoxin, and a surface binding
l e ~ t i n . ~ In
J ~addition,
,'~
infiltration of the lamina propria
with lymphocytes has been noted and may lead to villus atrophy. These processes contribute to the development of disaccharidase deficiencies commonly associated
with Giardia infection. Isozyme electrophoresis and
genetic studies have demonstrated considerable phenotypic and genotypic heterogeneity among G. lamblia
isolates. Analysis of sequential isolates suggests that this
heterogeneity is due to the h g h rate of genetic rearrangement within the Giardia genome.20Chronic infection has
been associated with hypogammaglobulinemia, proteincalorie malnutrition, previous gastrectomy, and use of
immunosuppressive medication.3~'*~19
Both systemic and
mucosal antibody production occurs in giardiasis.21Secretory IgA interferes with the adhesion mechanism of the
parasite and may help clear the infection. The greater
prevalence of disease in younger patients suggests a partial i m r n ~ n i t y . ~
Clinical Manifestations
The clinical manifestations of giardiasis are remarkably &verse,including asymptomatic carriage, acute diarrheal disease, severe chronic diarrhea with malabsorption,
and growth failure in
Patients with acute
symptomatic giardiasis complain of diarrhea, abdominal
cramps, bloating, flatulence, malaise, nausea, and anorexia.
Stools may be greasy or foul smelling. Fever and tenesmus are uncommon and point toward other pathogens
that are invasive. Extraintestinal manifestations have been
described in rare cases and include urticaria, reactive
arthritis, and biliary tract di~ease.'~
Chronic infection is
associated with profound malaise, lassitude, epigastric
discomfort, headaches, and steatorrhea. Patients who
develop recurrent diarrhea after treatment for Giardia
infection may have associated lactose intolerance rather
than relapse of their infection.
J o u r n a l o f T r a v e l M e d i c i n e , V o l u m e 4, N u m b e r 1
Diagnosis
The most common method for diagnosis of giardiasis is microscopic examination of the stools for trophozoites using either wet mount or trichrome ~ t a i n . ~ ~ T h r e e
consecutive stool specimens when properly examined
yield a diagnosis in 80-90% of cases.25Iodine staining is
used for better identification of the cysts.The stool sample may be preserved in 10% buffered formalin and
examined later using trichrome or iron hematoxylin
stains.The duodenal string test (Entero-Test) may increase
the yield, but this procedure is not well-tolerated by
patients. In selected patients, endoscopic aspiration and
biopsy may be considered. Giardia antigen detection
assays are commercially available and are highly sensitive
and specific.2h
Treatment and Prevention
Metronidazole is the most frequently used drug for
the treatment of giardiasis in the United States and is associated with a cure rate of 8c-90%." The adult dosage is
250 mg PO t.i.d. for 5-7 days; children should receive
5 mg/kg PO t.i.d. for 7 days. Quinacrine, which has a
comparable efficacy, is associated with far more side
effects and is no longer available in the United States.
Tinidazole is frequently used worldwide but is not available in the US. It has excellent efficacy when given in a
single oral dose of 2 g.28Furazolidone 25-50 mg P O q.i.d.
is available as a liquid suspension. Recommended duration of therapy is for 7 days. It is well-tolerated by children, but the efficacy is lower. Pregnant women may be
treated with paromomycin, a nonabsorbable aminoglycoside, which is effective in 60-70% of cases.Treatment
of asymptomatic cyst passers has been advocated in
developed countries and returning travelers. Such an
approach is futile in hyperendemic areas due to rapid reinfection.2' Travelers to developing countries should be
advised against the ingestion of untreated surface water.
Bringing water to boil is adequate to kill Giardia cysts,
though boiling for a few minutes may be required at
higher altitudes. Halogenation with iodine- or chlorinebased agents is generally effective.6 Person-to-person
transmission can be reduced by practicing strict handwashing. '()
Amebiasis (Entamoebahistolyfica and Entamoeba dispar)
Organisms and Life Cycle
Amebiasis is caused by infection with the enteric
protozoan E. histolytiru.The existence of pathogenic and
nonpathogenic strains of E. kistolytica has been postulated
for some time based on studies showing differences in
isozyme electrophoretic patterns (zymodemes).31,32 More
recently, genomic differences in DNA polymerase chain
reaction amplification products, rRNA probe studies,and
cloned genes have confirmed the existence of two distinct species that are morphologicallyidenti~al.~'~"'The
non-
Hashrney e t a l . . P a r a s i t e s a n d D i a r r h e a . I: P r o t o z o a n s a n d D i a r r h e a
pathogenic species, now named Entamoeba dispar, is only
associated with the asymptomatic carrier state, while E.
histolytica causes tissue invasion and symptomatic disease.
The genus Entamoebu also includes five other species
(E. coli, E. gingivulis, E. moshkovskii, E. hartmanni and E.
polecki] known to infect humans. None of these species
cause disease in humans.34
The life cycle of E. histolyticuincludes trophozoite and
cystic stages. Cysts are excreted into the environment and
can remain viable for several weeks.The mature cyst is
quadrinucleate and ranges in size fi-om 8.5-19 prn in diameter. Infection is acquired by ingestion of the cyst form.
Excystation occurs in the small bowel and results in eight
motile trophozoites ranging from 12-60 p.m in size.
Epidemiology
Both species have a worldwide distribution with
approximately 12% of the world’s population infected
with one of the
E. dispar has never been
associated with disease and does not elicit any specific
serologic response. In contrast, E. histolyticu causes serum
antiamebic antibody production in -90% of patients
with invasive disease.35Since E. histolytica and E. dispar
are morphologically identical, most of the data on prevalence includes both. In most areas E. dispav is 10 times
as common as E. histolyticu. Amebiasis is endemic in
countries such as Mexico, India,West and South Africa,
and Central and South America, where up to 50% of the
population may be infected. Serologic surveys in endemic
areas indicate that up to 25% of the population have had
prior asymptomatic infection with E. histolyti~a.~~
Epidemiologic risk factors include lower socioeconomic
status, migration from areas of high endemicity, institutionalization (especially in mentally retarded individuals)
and living in a communal setting. Patients with amebic
liver abscess have a high prevalence (>75%) of asymptomatic intestinal colonization with E. histolytica that
may result in a prolonged carrier state if inappropriately
treated.” Invasive amebic disease carries an annual worldwide mortality of 40,000-100,000.
The overall prevalence of intestinal amebiasis in the
United States was about 4% of clinical stool specimens,
but this appears to be decreasing.36In the southwestern
United States there is a higher prevalence of invasive
disease, especially among immigrants from Mexico. E.
histolytiu is not a common cause of traveler’s diarrhea. However, amebiasis can be a cause of chronic diarrhea in travelers returning from developing countries.” The risk of
acquiring infection is proportional to the duration of stay
in the endemic area. Ninety-five percent of travelers or
immigrants who develop liver disease will present within
5 months after returning from an area of endemicity.
Pathology and Pathogenesis
E. histolyticu trophozoites attach to the colonic
epithelium by means of a galactose inhibitable adherence
19
lectin (GIAL), a 260 kD surface protein.This adherence
lectin protects the parasite against complement-mediated
lysis by inhibiting the assembly of C 8 and C 9 components with the membrane attack ~ o m p l e x . ’ ~ ~The
~*~~’
cytolytic activity of E. histolyticu is dependent upon direct
contact to the target cell, binding by the GIAL, and the
presence of extracellular calcium ions.4”Proteolytic disruption of tissue planes is mediated by a variety of parasite enzyme^.^^^^^ Proteinases cause extracellular tissue
destruction and degradation of secretory 1gA.The spectrum of colonic lesions range from nonspecific thickening
of the mucosa to frank ~lceration.~’The
classic description of flask-shaped ulcers is seen in less than half of cases.
After invadmg the colonic epithelium, the amebic trophozoites can reach the liver via the ascending portal venous
system. In the liver, extracellular proteinases digest the
hepatic tissues, thereby resulting in focal areas of tissue
destruction that can coalesce into necrotic areas termed
liver abscesses. Immunity to amebic infection is incomplete, although patients cured of invasive disease rarely
have reinfection.
Clinical Manifestations
Noninvasive lumenal infection with E. histolytica
may result in asymptomatic infection or nonspecific gastrointestinal complaints such as bloating, cramps,decreased
appetite, and flatulence. Amebic colitis on the other
hand has a subacute onset characterized by bloody mucoid
diarrhea, abdominal pain, weight loss, bloating, tenesmus, cramps, and fever. Amebic colitis can occur in a
chronic nondysenteric syndrome with intermittent diarrhea, mucus, and abdominal ~ a i n . ~Complications
’.~~
of
intestinal infection include fulminant colitis, toxic megacolon, intestinal perforation, stricture formation, and
ameboma (annular lesion of the colon, usually near the
cecal area, inhstinguishable fi-om carcinoma of the
Chronic amebic colitis is clinically indistinguishable
from idiopathic inflammatory bowel disease. Because
corticosteroid therapy may result in perforation, stool
examination for trophozoites and serologic test for amebic disease should be performed prior to making a diagnosis of inflammatory bowel disease.
Extraintestinal disease most commonly presents as
amebic liver abscess. Patients present with fever and right
upper quadrant abdominal pain. Laboratory findings
include leukocytosis and elevated liver enzymes. Liver
abscesses occasionally rupture into the pleural space,
resulting in pleural efisions. Extraintestinal disease rarely
involves the pericardium, genitourinary system, and the
brain.
Diagnosis
The diagnosis of intestinal infection is made by
finding cysts or trophozoites in the stool. A saline wet
mount of the stool should be made kom a fi-esh stool specimen, as trophozoites disintegrate in 30 minutes at room
J o u r n a l of T r a v e l M e d i c i n e , V o l u m e 4 , N u m b e r 1
20
temperat~re.~'
If immediate evaluation cannot be performed, the specimen should be fixed in polyvinyl alcohol and examined later using trichrome o r iron
hematoxylin stains. Stool examination is positive in >85%
ofpatients when at least three stool specimens are examined on alternate days.46Hematophagous trophozoites
when seen are pathognomonic o f E. hi~tolytica.~'
Endoscopy with scraping or biopsy is a useful technique
in evaluation of difficult to diagnose cases.
In nonendemic areas, serum antiamebic antibody tests
may be useful in the diagnosis of invasive disease. Over
85% of patients with biopsy-proven disease have detectable
antibody by the seventh day of illness." In endemic
areas a positive serology is less helpful. U p to 25% of the
population may have serologic evidence of prior infect i ~ n Several
. ~ ~ new stool and serum antigen detection
techniques appear promising in the identification of
pathogenic versus nonpathogenic strain^.^'
The presence of an amebic liver abscess is suggested
by a history of fever and right upper quadrant pain,
hepatic tenderness on physical examination, and an elevation in liver enzymes. Diagnosis requires the demonstration of the abscess by ultrasonography or computerized
tomcgraphy scan.
Treatmenf
Amebicidal drugs have been divided into intralumenal agents (e.g., diloxanide furoate, paromomycin,
iodoquinol, and tissue agents metronidazole, tinidazole,
and emetine).48Ti~~ue
amebicides lull amebae in host tissues and organs, whereas the poorly absorbed lumenal
amebicides are active only in the intestinal l ~ m e n . ~ '
"E. histolytica" cysts found in the stool of asymptomatic cyst passers are usually those of E. dispar. Since this
parasite has never been shown to cause invasive infection, treatment is unnecessary. Serologic tests are negative and cysts are excreted in the stool for a limited
p e r i o d , usually 6-8 months. A n enzyme-linked
immunosorbent assay (ELISA) test to distinguish between
E. histolytica and E. dispar is being developed. Symptomatic
patients should be treated. In patients without evidence
of invasive disease, diloxanide furoate 500 mg P O t.i.d.
for I0 days is the therapy of choice. In the United States
this drug is available only from the Centers for Disease
Control and Prevention (CDC). Iodoquinol 650 mg
PO t.i.d. for 20 days is also effective but has bothersome
side effects and a longer duration of therapy. Paromomycin can be used as an alternative for lumenal or
mild colonic disease. Invasive disease should be treated
with metronidazole 750 mg P O t.i.d. for 10 days (tinidazole is also available outside the United States) followed
by a lumenal amebicidal agent in order to eradicate
intestinal disease.'') Tetracycline or erythromycin may
be used in patients unable to tolerate metronidazole, but
these drugs are not effective in hepatic disease. Dehy-
droemetine is available from the CDC. Due to the occurrence of serious side effects such as prolongation of the
Q-T interval, it should be administered in a monitored
hospital setting. Chloroquine may be added to metronidazole in extraintestinal disease. Fine needle aspiration
is indicated in left-sided abscesses and those unresponsive to medical therapy.
Cryptosporidiumpanrum
Organism and Life Cycle
The protozoan parasites of the genus Cryptosporidium are closely related to other coccidian parasites such
as TDxoplasma and Isospora. Although many species were
named based on presumed host-species specificity,there
is little host specificity for the strains infecting mammals.s1
Most organisms infecting mammals probably belong to
a single species, C.parvum.
Humans are infected after ingestion of oocysts (4-6
p,m in diameter).5'After excystation in the s m a l l intestines,
the motile sporozoites attach to the host cells via their
anterior end.They are enveloped by the microvilli and
subsequently develop along the lumenal surface of the
enterocyte within a parasitophorous vacuole separated
from the host cytoplasm by a dense organelle (intracellular but extracytoplasinic).The parasite enlarges to form
the trophozoite and divides forming meronts with four
to eight nierozoites. When nierozoites are released they
invade other host cells, and a portion of the merozoites
develop into the sexual forms (macrogamont and microgamont). M e r fertihzation the macrogamont develops into
an oocyst, which sporulates in situ. Most of the oocysts
are coated with a thick wall and are environmentallyhardy.
About 20% of the oocysts are thin-walled and may be
involved in an autoinfection cycle.
Epidemiology
The first human case of cryptosporidiosis was
reported as recently as 1976 and only seven human cases
were reported prior to 1982.52Thenumber of cases has
risen dramatically since the description ofAIDS. Subsequently, cryptosporidiosis has been identified in over 50
countries on all six inhabited continents in both normal
and immunocompromised hosts.s3
When clinical stool specimens are examined for
Crptosporidium oocysts, approximately 1-3% are positive
in studies from North America and Europe (range
0.1%-14.1%, mean of 1.6 and 2.6%, respectively). In
contrast, rates of 4.9-10.4% are typical in developing
countries (range 1.3-31 .5%).53.'4Seroprevalence rates,
which presumably reflect active or prior infection, range
from 25-35% in developed countries but up to 64%
among the urban indigent population in Latin
5.56
Cryptosporidiosis has been identified as a common
cause of diarrhea in developing c ~ u n t r i e s . ~Infec~~"~~
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tion is more common in children than in adults, but is
rare in breastfed infants.sy,6'Cryptosporidiosis appears to
be more prevalent in warm, wet month^.'^^."^ Cryptosporidiosis has also been frequently identified as a
cause of diarrhea in travelers returning from areas with
poor ~anitation.'~+'Theoocysts are very hardy and survive for prolonged periods in the environment. Cryptosporidiosis is associated with waterborne epidemics of
diarrheal
Chlorination of water supplies does
not appreciably kill the oocysts.
Initial descriptions of cryptosporidiosis in normal
hosts stressed animal contact;74however, epidemiologic
data have strongly supported only calf-to-human transmission."' Subsequent studies suggest that person-toperson transmission is also common, especially in urban
area^.^',^^ In developed countries, cryptosporidiosis is a
frequent cause of outbreaks of diarrhea in the context
of day-care center^.^^^^^^^' Person-to-person transmission
has also been documented in secondary cases in household c ~ n t a c t and
s ~ among
~ ~ ~ ~health care ~ o r k e r s . ~ ' T h e
high prevalence of disease among homosexual males
also suggests person-to-person transmission.*" Cryptosporidiosis is an increasingly recognized cause of diarrhea in travelers to developing countries. Cryptosporidium
was found to be the cause of diarrhea in 5% of travelers
to Leningrad and in short-term Peace Corps workers in
West Africa.*',''
T h e prevalence o f cryptosporidiosis is high
in immunocompromised hosts, particularly in patients
with AIDS. In the United States, 3-4% of patients
with AIDS have cryptosporidi~sis.'~
In contrast, in
Africa more than half of AIDS patients have cryptosporidi~sis.'~
Prevalence rates in Latin America are
intermediate.'s~x6
Pathology and Pathogenesis
Cvyptosporidium has been detected throughout the
gastrointestinal tract, in the biliary tract, and even in the
respiratory tract.shIntestinal changes include villus blunting, crypt hyperplasia, and infiltration of the lamina propria with polymorphonuclear leukocytes, lymphocytes,
The parasites are found within the
and plasma
microdus layer within parasitophorous vacuoles.Though
most patients exhibit normal duodenal villous architecture, high intensity infections, as evidenced by histologic
examination of biopsy specimens, were associated with
severe morphologic abnormalities."
The pathophysiology of diarrhea in cryptosporidiosis
is complex and likely involves both host and parasite factors. Current data suggest that site specific alteration in
the function of villus absorptive cells, as evidenced by
alteration in glucose-mediated sodium transport and
release by the host of proinflammatory cytokines as part
ofthe mucosal immune response, are the primary mech-
21
anism involved." Chloride dependent enterotoxic activity has also been described."
The host immune response in cryptosporidiosis has
not been clearly defined. Secretory antibodies have been
noted to interfere with sporozoite and merozoite attachment, but the development of specific serum and mucosal
In
antibodies in AIDS patients does not effect a
contrast, the cell mediated immune response, specifically C D 4 T cells and IFN-y, appear to play an important role in limiting infection.y4y6In patients with HIV,
the CD4 count is the best marker of the ability of the
immune system to clear infection."'
Dose response studies in healthy volunteers, using
one strain of C.parvum, have shown a low mean infective dose (ID5,) of 132 oocy~ts;'~
however, it is not
known whether immunocompromised individuals are
more susceptible to infection than normal hosts.The dose
of the inoculum did not influence the incubation period
or the severity of illness.
Clinical Manifestations
The clinical presentation of cryptosporidiosis varies
depending on the host response. In normal hosts, initial
symptoms follow an incubation period, usually 7-10
day^.^^.'^,^^^'' Onset is typically abrupt. The most commonly identified symptom is diarrhea, with watery stools
that may be voluminous. Mucus is usually not present.
Fecal leukocytes and blood are absent. Abdominal pain
and cramping were frequently noted in some series.74,"
Weight loss, anorexia, and flatulence have been noted.
Fever when noted is typically low grade. Nausea and vomiting are present only in a minority of patients. Respiratory symptoms have been frequently noted in some
series.5',6Z,99.100
In travelers and other immunocompetent hosts,
symptoms typically resolve in 10-1 4 days.64,74.76,"
However, C. parvum is among the most common causes of
diarrhea persisting for over 2 weeks, and chronic cases
lasting over 30 days are well described."" Oocysts are
excreted in the stool for 1-2 weeks after resolution of
symptoms.98Cryptosporidiosis has been associated with
malabsorption. Whether this reflects Cryptosporidium
causing malabsorption or merely the increased severity
and recognition of cryptosporidiosis in patients with malabsorption due to other causes is unclear. Asymptomatic shedding of oocysts has been noted in some
lo'
In immunocompromised hosts, disease onset is
usually i n s i d i o ~ s . ~ ~ * ~ ~Watery
. ~ ~ , ' " "diarrhea
J"~
may gradually increase over weeks to months. Weight loss and
profound dehydration are common. Abdominal pain
and cramping are also common. Symptoms typically
persist unless there is improvement in the immunologic
status.
22
Diagnosis
Since the symptoms of cryptosporidiosis are nonspecific, diagnosis usually depends on the demonstration
of oocysts in stool specimens. Considerable experience
i5 required to obtain reliable results.The sensitivity of stool
examination may be increased by concentration techniques or by staining. Sheather's sucrose floatation is the
most frequently used concentration te~hnique.'"~"'~
A number ofstaining techniques have been used for
Cryptosporidiurn.525637.103 Acid-fast staining is most commonly employed (Fig. 1). A number of acid-fast stains
are widely used, including modified Kenyoun, diniethyl
sulfoxide-modified stains, and fluorescent stains.'i)1,'iJ4 In
patients with diarrhea, oocysts can generally be identified by acid-fast staining without using concentration
techniques.
Monoclonal antibody based immunofluorescent
tests to detect oocysts in stool specimens are commercially available and demonstrate >90% sensitivity and
specificity.'"' In comparative studies, the immunofluorescence test is both more sensitive and more specific than
acid-fast staining technique^."'^^'"^ Widespread application of this test may be limited by the additional cost.
Tissue forms can be detected in biopsy specimens with
routine histologic stains including hematoxylin and eosin
or Giemsa.In8
Treatment
Fluid and electrolyte replacement are of primary
importance in all cases of diarrhea. Attention to nutrition may be critical since the immunosuppression associated with malnutrition may delay recovery."' In
compromised hosts, nonspecific antidiarrheal agents such
as loperamide or diphenoxylate may be required to limit
voluminous fluid losses. In a recent controlled clinical trial,
J o u r n a l o f Travel M e d i c i n e , Volume 4 , N u m b e r 1
the somatostatin analogue octreotide was no more effective than placebo.'"'
There are numerous reports of antiparasitic agents
used in cryptosporidiosis. Spiramycin, a poorly absorbed
macrolide antibiotic, was initially reported to affect
cryptosporidiosis in patients with AIDS in anecdotal
reports,"' but efficacy has not been demonstrated in controlled trials. "*
Paromomycin, a poorly absorbed aminoglycoside
antibiotic,"3 achieves effective concentrations along the
lumenal surface of the intestines, where the organism is
found in vivo."4 A small placebo-controlled trial demonstrated both clinical and parasitologic improvement with
paromomycin treatment in patients with AIDS."' Patients
experienced significant symptomatic improvement; however, a majority relapsed.
HIV-infected individuals should be advised not to
drink water directly from lakes or rivers. Currently there
are no recommendations regarding the use of household
tap water; however, persons who want to take independent precautions can reduce the risk of infection by
boiling water for 1 minute, by using"abso1ute" 1 micron
water filters,or by using high-quality bottled water.- Risk
of infection during travel to endemic areas can be reduced
by avoiding use of untreated surface and ground water
and by boiling water prior to use.
Isospora belli
1. belli is a coccidian parasite closely related to Cryptosporidium. The oocyst is elliptical and measures 22-33
by 10-15 p m in size.Within the oocyst are two sporo-
cysts, each ofwhich contains four sporozoites.The sporozoites are released in the intestines, invade the intestinal
epithelium, and undergo asexual (merogony) and sexual
(gametogony) division within cytoplasmic vacuoles of the
enterocyte."6 The oocysts are excreted prior to sporulation, which occurs outside the host.
Epidemiology
1. belli is widely distributed in the animal kingdom.
Figure 1 Oocysts of Cryptosporidiurn are visualized in a stool
specimen stained by the modified acid-fast stain (with permission from Dr. Lynn Garcia).
Infections have been described from nearly all the countries in the Americas, as well as in subSaharan Africa, the
Middle East,Asia and the Pacific islands,and E ~ r o p e . " ~ , " ~
Studies suggest an increased prevalence in tropical and
subtropical climates and in areas with poor sanitation.
Little is known about the prevalence of infection in
humans. In a limited number of studies employing adequate techniques, 0.1-1.8% of stools were positive."'
Prevalence among AIDS patients in the United States
is approximately 1%."' This may be an underestimate;
this organism was not routinely looked for, and cotrimoxazole is frequently prescribed in these patients. In
contrast the prevalence rate in less developed countries
is as high as 15%.K44,R5J20
The mechanisms of transmission are unknown, although spread from infected animals and humans is suspected.
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Pathology and Pathogenesis
Biopsy and autopsy specimens show villus atrophy,
crypt hyperplasia, and infiltration of the lamina propria
with inflammatory cells, in particular eosinophils."'J2'
All of the life cycle stages are visible within biopsy specimens.The mechanisms responsible for diarrhea are not
clear, but may relate to malabsorption, which has been
well described in both normal and compromised
Clinical Features
The clinical features of isosporiasis in immunocompetent hosts have been well defined by several large
series and from experimental and laboratory-acquired
infections.'1x~'22-'zs
After an incubation period (6-12
days in experimental and laboratory infections), patients
typically present with an acute onset of fever, headache,
and malaise. Gastrointestinal complaints include diarrhea
(both frequent and watery stools), abdominal pain, nausea, and vomiting. Initially the stools are watery, but
subsequently they may be fatty. Stool specimens do not
contain fecal leukocytes but frequently contain CharcotLeyden crystals and fat. Peripheral eosinophilia is fi-equent.
While symptoms are usually self limited, patients are often
ill for 6 weeks to 6 months. Even in apparently normal
hosts, a portion of patients go on to develop chronic diarrhea and malabsorption, which may persist for years.' l6.l2'
Patients with AIDS and isosporiasis present with an
insidious onset of diarrhea, weight loss, and abdominal
Fever and vomiting are less frequently noted.
Symptoms are usually present for months prior to diagnosis. Biliary disease,acalculous cholecystitis, and ulcerative colitis have been r e p ~ r t e d . ' ~ ~ . ' ' ~
Diagnosis
Diagnosis depends on the identification of organisms in stool or biopsy specimens.While the oocysts can
be identified in wet mounts of stool specimens, stool
examinations are frequently negative unless they are
concentrated. Zinc sulfate floatation has been highly
successful in some laboratories.'23Acid-fast stains appear
to increase the sensitivity of stool examination.'2y
Treatment
Isosporiasis responds promptly to therapy with
antifolate d r ~ g s . ' ~ " , ' ~ ' ,Trimethoprim
'~~,'~'
(160 mg) combined with sulfamethoxazole (800 mg) four times a day
for 10 days leads to prompt resolution of symptoms in
normal hosts and in patients with AIDS. For AIDS
patients, approximately half will relapse if not placed on
secondary prophylaxis. Both trimethoprim (TMP)/sulfamethoxazole (SMX) (160 mg/800 mg three times a
week) and sulfadoxine/pyrimethamine (500 mg/25 mg)
once weekly can prevent relapse.'30
23
Before its identification as a coccidian parasite, Cyclospora
had been referred to as cyanobacteria-like bodies (CLB)
and blue-green algae.132-'35
The appearance of oocysts is
similar to those of Cryptosporidium, but the size is slightly
larger (9-10 pm). Each oocyst contains two sporocysts,
each of which contain two sporozoites.I3'
Epidemiology
There is a wide geographic hstribution of Cyclospora.
Most of the reported cases have originated in tropical
countries such as India, Nepal, and South America. Cases
have been identified in travelers returning from the tropi c ~ . ' ~ ~ . ' "Gascon
~ ' ~ ' et al. found Cyclospora in 2.8% of
patients with traveler's diarrhea in their ~1inic.l~'
Transmission occurs via the oral route, and local outbreaks have
been traced to contaminated water or food supply. An
outbreak of relapsing diarrhea among 17 physicians and
three hospital employees in a Chicago hospital was traced
to a contaminated water s o ~ r c e . ' ~Most
~ ~ ~ 'outbreaks
~'
occur during the summer season.
Pathology and Pathogenesis
The site ofinfection is the s m a l l intestine.Endoscopic
biopsy specimens show a diffuse inflammatory response,
with an increase in intraepithelial leukocytes, mild to
severe villus atrophy, and crypt hyperplasia.'42Thepathogenesis of these lesions and the mechanism of diarrhea
are unclear; however, both invasion as well as toxin production have been p ~ s t u l a t e d . ' ~ ~
Clinical Presentation
The clinical presentation is similar to that of Cryptosporidium and Isospora infections. Clinically, patients
present with watery diarrhea, anorexia, and malaise.
Approximately half experience abdominal cramps and up
to a third have nausea, vomiting, and fever. Symptoms
often wax and wane.The duration of illness in immunocompetent patients is variable but typically lasts several
weeks (mean duration 43 days in one series).I3' In AIDS
patients, Cyclospora results in chronic intestinal infection
with prolonged diarrhea.137Duration of symptoms correlates with the presence of Cyclospora in the stool. Control groups either with other causes of diarrhea or
without symptoms only rarely have the organism in
their stools. C. cayetanensis may be associated with biliary tract disease.'43
Diagnosis and Treatment
Diagnosis requires a high index of suspicion. Diagnosis is dependent on microscopic demonstration of
morphologic characteristics,staining properties, and autofluorescence under ultraviolet light. In contrast to Cryptosporidium, staining is variable with the modified acid-fast
stain (Fig. 2). Oocysts autofluoresce when examined
under ultraviolet light and do not cross-react with
Cyclospora
immunologic tests specific for C r ~ p t o ~ p o r i d i u m . ' ~ ~ . ' ~ ~
Cyclospora cayetanensis, a coccidian parasite, has
Measurement of oocysts is recommended to distinguish
recently been identified as causing human infe~ti0n.l~' Cyclospora from Cryptosporidium.'44,'45Cotrimoxazole
24
Figure 2 A stool specimen demonstrating a variable uptake
of modified acid-fast stain by oocysts of Cyclospara (kindly provided by Dr. Charles Stager).
(TMP 160 mg/SMX 800 mg) PO b.i.d. for 5-7 days is
the treatment of choice, with recent studies reporting a
significant response to therapy and improvement in both
clinical and parasitologic parameter^.^'.'"^'^",'^'
Microsporidia (Enterocytozoon bieneusi and
Encephalitozoon intestinalis)
Organism and Life Cycle
Microsporidia are a group of obligate intracellular
protozoan parasites belonging to the phylum Microspora,
which consists of more than one hundred genera and over
a thousand
14' Human infection was first
reported in 1959 and thus far four genera have been implicated in human disease: Enterocytozoon sp. (E. bieneusz],
Encephalitozoon sp. (E. hellurn, E . canaliculi,E. intestinalis),
Nosema sp. ( N . connori, N. ocularurn, N. corneurn), and
Pleistophora sp.149a15fl
The major species associated with
enteric infection are Enterocytozoon bieneusi and Encephalitozoon intestinalis (previously called Septata intestinalis).
Microsporidia are primitive eukaryotic unicellular
parasites.They lack mitochondria and golgi membranes
and possess small prokaryotic-sized rRNA.'" They produce environmentally resistant spores that are characterized by coiled polar tubules. After entering the hosts,
the polar tubules inject the sporoplasm (nucleus and
cytoplasm) into the host cell.'48,149,152
Although there is
significant intergeneric variation, the life cycle has three
distinct phases: first, the spore or infective stage; second,
the proliferative stage during which the organism divides
by merogony (schizogony); and lastly, sporogony, the
spore-forming stage.
Epidemiology
Little is known about the epidemiology of human
microsporidiosis. Serologic surveys suggest that infection
with these organisms is widespread. Spread of the organ-
J o u r n a l o f T r a v e l M e d i c i n e , V o l u m e 4, N u m b e r 1
isms through environmental, person-to-person (fecal-oral,
aerosolized respiratory secretions), and animal-to-person
transmission have been postulated.'5"J53Although there
are few reported cases of microsporidial infection in
nonHIV patients, these organisms are increasingly being
recognized as etiologic agents of chronic disseminated disease in HIV-positive individuals. E. bieneusi and E. intestinalis have both been associated with chronic diarrhea in
AIDS patients. However, studies have not been unanimous in demonstrating this association. In one casecontrolled prospective study,the investigators did not find
a significant difference in the presence of microsporidia
on intestinal biopsy specimens when comparing HIV
infected patients with and without diarrhea.'54 In contrast, others have noted a clear association between infection and altered d-xylose absorption.'"* In the normal
host, microsporidia have been associated with traveler's
diarrhea.'55 It has been suggested that microsporidia
transiently infect immunocompetent individuals causing
disease only in immunosuppressed p a t i e n t ~ . ' ~ ~ , ' ~ ~ J ~ ~
Pathology and Pathogenesis
E. bieneusi infections generally appear to be limited
to the intestinal and biliary epithelium. The organisms
are primarily found in the tips of the villi. Meronts are
4-6 p m in diameter. E. bieneusi was initially identified
by electron m i c r o s ~ o p y .E.
~ ~bieneusi
~ ~ ' ~ infections
~
are limited to the intestinal and biliary epithelium. Histopathologic changes range from normal villus architecture to
severe d u s atrophy.Infiltration of the lamina propria with
mononuclear cells has been noted. Little is known about
t h e pathogenic basis o f diarrhea i n intestinal
microsporidiosis. Malabsorption of both carbohydrates
and fats has been d e m o n ~ t r a t e d . ~ " ~ ~ ' ~ ~ * ' ~ ~
E . intestinalis infects enterocytes, macrophages, fibroblasts, and endothelial cells of the lamina propria. In
contrast to E. bieneusi, widespread dissemination often
Clinical Presentation
Microsporidia have been associated with a self-liniited diarrheal illness in a few apparently immunocompetent patients. 155 Ophthalmologic involvement and
central nervous system disease have also been reported.
E. bieneusi appears to be associated with chronic diarrhea
in AIDS patients.Is4 In this group, the clinical presentation consists of diarrhea that is watery, without blood or
mucus, and of variable volume. Weight loss is common.
Since diagnosis was initially limited to patients who
underwent intestinal biopsy, self-limited cases and milder
cases may have been overlooked. Asymptomatic intestinal colonization by microsporidia has also been
reported. lS4 Microsporidial infection of the biliary mucosa
is associated with AIDS ~holangiopathy.'~'~'~
Clinical presentation of E. intestinalis resembles that of E. bieneusi,
except that extraintestinal disease is common.'6'
H a s h m e y e t a l . , P a r a s i t e s a n d D i a r r h e a . I: P r o t o z o a n s a n d D i a r r h e a
25
tions, and electron microscopy have been used to increase
sensitivity.
Treatment
There is no proven curative therapy currently available for E. bieneusi infections.As in all diarrheal illnesses,
attention to fluid and electrolyte management and nutrition are the mainstay of therapy. Symptomatic improvement may be achieved by the use of antidiarrheals.There
have been anecdotal reports of clinical responses to treatment with albendazole.16' Controlled trials are in progress.
E. intestinulis infection, on the other hand, shows a dramatic clinical and parasitologic response to albendazole
therapy.16'
Figure 3 Modified-trichrorne stain of a stool specimen showing microsporidial spores.
Diagnosis
T h e primary means of diagnosing intestinal
microsporidial infection is demonstration of the spores
in stool.The spores are small (1-3pm in diameter) and
not easily identified without special staining techniques.
Weber developed a modified trichrome stain that facilitates identification of spores in stool (Fig. 3).'62The
cysts can also be stained by fluorescent stains including
calcofluor white (Fig. 4) and indirect immunofluores~ e n c e . ' ~ Cytologic
',~~~
evaluation of readily obtainable
specimens (stool, duodenal aspirates, urine, sputum, nasal
discharge,and bronchoalveolar lavage) using light microscopic staining techniques such as Giemsa and modified
trichrome are becoming routine practice.I5' O n intestinal biopsy specimens, organisms stain with hematoxylin
and eosin. The spores also stain with toluidine blue,
brown-brenn or Giemsa stains. They are frequently
overlooked in routine sections due to their small size and
limited numbers. Thin plastic sections, touch prepara-
Balanfidium coli
B. coli is the only ciliate associated with human
infection. It has a widespread distribution,but is only rarely
found in human stool samples (<1%in all surveys).'65-167
Humans appear to be an incidental host with the primary reservoir in domestic and wild animals, including
pigs. The trophozoites are 30-150 p,m in length and
40-55 p,m wide, ovoid in shape, and actively motile in fi-esh
specimens.The cyst stage is 40-65 p,m in diameter.
The predominant site of infection is the colon,
especially the rectosigmoid. 167 Histologically, the lesions
resemble those seen with amebiasis, including flaskshaped ulcers with undermined edges and few neutrophils.The lesions may coalesce, involving most of the
colon. Most organisms are found at the periphery of
lesions. Occasionally, lesions may perforate.
Clinically many cases are a~ymptornatic.'"~'~~
A
chronic form has been described with loose bowel movements, abdominal pain, tenesmus, mucus in the stools, and
weight ~ o s s . Finally,
~ ~ ~ dysentery
J ~ ~
has been described in
a minority of patients. Patients present with sudden
onset of diarrhea with blood and/or leukocytes in the
stool. Abdominal pain and tenderness and weight loss
are frequent. Symptoms may last for months. Diagnosis
is by demonstration of the trophozoites in stools. Fresh
stools examined for motile trophozoites are probably more
sensitive than are preserved specimens.'66
Tetracyclines are recommended as treatment.'69
Metronidazole, diiodohydroxyquin, and paromomycin
also have some activity.'66a160
Protozoans as Possible Causes of Diarrhea
Dienfamoeba fragilis
Figure 4 Microsporial spores visualized under ultraviolet
light using a modified flurochrome calcofluor white stain (photograph kindly provided by Dr. Charles Stager).
D.fragilis is a flagellate, closely related to the trichomonads. It has been identified in most areas of the
world with a prevalence of 1.4-19% in stool specim e n ~ . An
~ ~ association
"
of D.fragi!is and Enterobius uermicularis (pinworm) has been noted, and some suggest that
they are transmitted t~gether.'~"
D.jagUis is considered
26
by some to be a cause of diarrheal illness. Most information on D.frugi/is-associated illness comes from retrospective studies. Several studies identified the organism
in stool samples of patients that lacked other recognized
pathogen~.'~+''~
In a recent survey, D.jiagilis was the only
parasite found in two-thirds of the cases.173It occurred
more frequently in children 5-9 years old. Eighty percent of the patients reported diarrheal symptoms. Anal
pr~ritis'~",'~'
and eosinophilia have been reported in a substantial portion of c a ~ e s . ' ~ ~ ~ ' ~ ~symptoms
T h e s e may in part
be due to concomitant pinworm infection.
None of these studies looked carefully for other
organisms that might have accounted for the symptoms
(e.g.,Esherichia coli, Stronnloides stercoralis, G. lamblia, Ctyptosporidium, and Isospora) . Furthermore, D.jiagilis has never
been shown to be invasive.While pathologic abnormalities have been described in infected patients (e.g., fibrosis
of the appendix),I7*other explanations seem more likely
than parasitism.Therapy with metronidazole, iodoquinol,
or paromomycin is effective, but eradicating the organism has only inconsistent effects on ~ y m p t o m s . ' ~ " , ' ~ ~
Blastocystis hominis
B. hominis is a common component of the human
intestinal flora. In the past, it was mistakenly believed to
be a yeast. Elegant studies, primarily by Zierdt and
coworkers, have conclusively demonstrated that B. hoininis
is a protozoan.'74
The role of t h s organism in the pathogenesis of diarrhea is contr~versial.'~~
B. hominis is present in 10-20%
of stools from both normal and ill populations. Several
series have been reported of patients with B. hominis in
their stools, the absence of other identified pathogens,
and gastrointestinal symptoms.'7"'7yA recent prospective
case-controlled study among tourists in Nepal failed to
show a difference in prevalence or heavy colonization
( > I 0 organisms per high power field) of B. hominis
between study subjects and controls.'X".'X'Symptoms
attributed to B. hominis include diarrhea, abdominal pain
and bloating, and anorexia. Duration of illness has been
variable. Eosinophilia has been noted in some cases.
Response to therapy with antiprotozoal agents has been
variable. Eradication of the organism does not correlate
well with clinical response.
N o t all commonly recognized pathogens were
sought in these studies, nor were noninfectious causes of
diarrhea excluded. O n careful evaluation, three groups
found that most patients with diarrhea and Blastocystis in
their stool have another explanation for their sympt o m ~ . ' ~ ' -Biopsy
' ~ ~ specimens have usually shown normal t i ~ s u e . " ~
Studies in gnotobiotic animals have
demonstrated colonization only when infected with
other stool organisms as well as B. hominis. Finally,
J o u r n a l o f T r a v e l Medicine, Volume 4 , Number 1
purportedly infected humans lacked a significant serologic response to the organism."'
Other Protozoa
A number of nonpathogenic protozoa may be identified by routine parasitologic examination of stool specimens. The most frequently encountered are Endolimax
nana and Entamoeba coli, each identified in approximately
4% of specimens. Others include Entamoeba hartmanrzi,
Iodamoeba butchlii and Trichomonas hominis.'
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