Unless the trophozoites and cysts match the size requirements, they are unlikely to be E. hartmanni. Definitive

identification relies on examination of permanent stained smears and measurements made with the calibrated

microscope.

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ENDOLIMAX NANA:

General Characteristics:

Endolimax nana, one of the smaller nonpathogenic amebae, has a worldwide distribution and is seen as frequently as E.

coli. E. nana has the same life cycle stages as E. dispar and the other nonpathogenic amebae. The trophozoite usually

measures 6 to 12 μm in diameter (normal range, 8 to 10 μm) .

Figure 8 A to C, Trophozoites of Entamoeba hartmanni. D and E, Cysts of E. hartmanni.

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 Although rarely seen, motility is sluggish and nonprogressive with blunt, hyaline pseudopods. In the permanent

stained smear, normally no peripheral chromatin is seen on the nuclear membrane, and the karyosome is large,

with either a central or an eccentric location in the nucleus (see Figures 9 and 10). E. nana shows more nuclear

variation than any of the other amebae, and occasionally E. nana can mimic D. fragilis or E. hartmanni. The

cytoplasm may have small vacuoles containing ingested debris or bacteria, but it also may appear relatively clean.

Cysts usually measure 5 to 10 μm in diameter (normal range, 6 to 8 μm). Cysts as large as 14 μm have been seen.

The cyst is usually oval to round, with the mature cyst containing four nuclei. The nuclei typically have no

peripheral chromatin and are somewhat evenly distributed in the cyst. Occasionally, very small, slightly curved

chromatoidal bars are present. The two-nucleated stage is not commonly seen, and frequently both trophozoites

and cysts are present in clinical specimens.

Figure 9 A to C, Trophozoites of Endolimax nana. D and E, Cysts of E. nana.

Figure 10 A, Endolimax nana trophozoite. B, E. nana

cyst, iodine stain. C, E. nana cyst. D, E. nana cyst

Travers, Sioux Falls, S.D.)

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Epidemiology:

 Transmission occurs through the ingestion of mature cysts from contaminated food or water. The cysts of E.

nana are less resistant to desiccation than those of E. coli. E. nana is also found in warm, moist climates and in

other areas with a low standard of personal hygiene and poor sanitary conditions. Pathogenesis and Spectrum of

Disease E. nana is considered nonpathogenic and does not cause disease.

Laboratory Diagnosis:

Although cysts sometimes can be seen in a wet preparation, definitive identification of E. nana relies on

examination of permanent stained smears.

Iodamoeba butschlII

General Characteristics:

 Iodamoeba bütschlii, one of the nonpathogenic amebae, has a worldwide distribution. Generally, the acquisition rate

for this organism is not as high as that for E. coli and E. nana. The life cycle stages of I. bütschlii are exactly the same

as those of E. nana. The trophozoite varies from 8 to 20 μm in diameter and has fairly active motility in a fresh stool

preparation . The cytoplasm is granular, containing numerous vacuoles with ingested debris and bacteria. The cytoplasm

is more vacuolated than in E. nana trophozoites. The nucleus has a large karyosome, which can be either centrally

located or eccentric (Figures 11 and 12).

A B C

Figure 11 A, Trophozoites of Iodamoeba bütschlii. B and C, Cysts of I. bütschlii

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On the permanent stained smear, the nucleus may appear to have a halo, and chromatin granules fan out around the

karyosome. If the granules are on one side, the nucleus may appear to have a “basket nucleus” arrangement of chromatin,

more commonly seen in the cyst stage. The trophozoites of I. bütschlii and E. nana may appear similar and are difficult to

differentiate at the species level, even on the permanent stained smear. Both organisms are considered nonpathogenic. E.

nana is recovered in clinical specimens much more frequently than is I. bütschlii. I. bütschlii cysts are round to oval.

The glycogen vacuole is so large that occasionally the cyst collapses on itself. Because nuclear multiplication does not

occur in the cyst form, the mature cyst contains a single nucleus. The cysts measure approximately 5 to 20 μm in diameter

and are rarely confused with those of other amebae (see Figures 11 and 12).

Epidemiology:

 Transmission of I. bütschlii occurs through the ingestion of mature cysts from contaminated food or water. This

organism is also found in warm, moist climates and in other areas with a low standard of personal hygiene and poor

sanitary conditions.

Pathogeneis and Spectrum of Disease:

 I. bütschlii is considered nonpathogenic and does not cause disease.

Figure 12 A, Iodamoeba bütschlii trophozoites. B, I. bütschlii cyst. C, I. bütschlii cyst. D, I. bütschlii cyst.

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Laboratory Diagnosis

 Although I. bütschlii cysts sometimes can be seen in a wet preparation, definitive identification relies on the

examination of permanent stained smears. Therapy Specific treatment is not recommended for I. bütschlii.

Prevention (E. hartmanni, E. nana, I. bütschlii):

Prevention depends on adequate disposal of human excreta and improved personal hygiene, preventive measures that

apply to most of the intestinal protozoa.

Blastocystis hominis:

General Characteristics:

Now generally considered a causative agent of intestinal disease. The current recommendation is to report the presence of

B. hominis and quantitate from the permanent stained smear (i.e., rare, few, moderate, many, packed); this information

may be valuable in helping to assess the pathogenicity of the organism in the individual

Patient/ B. hominis consists of four major forms. The cyst form is the most recently described form of the life cycle

stages. Thick-walled cysts are thought to be responsible for external transmission through the fecal-oral route; thin-walled

cysts are thought to cause autoinfection. Cysts can vary in shape but are mostly ovoid or spherical. The central vacuole

form (also referred to as the central body form) is the most common form found in clinical stool samples. The large

central vacuole can occupy most of the cellular volume. The amoeboid form is rarely seen. The granular form can be seen

in cultures of B. hominis.

Epidemiology:

Infection with B. hominis is acquired by the fecal-oral

route from infective forms contained in the feces. The organisms can be ingested in contaminated food and drink or

acquired from fomites or through various sexual practices that may include accidental ingestion of fecal organisms. As

with E. histolytica, flies and cockroaches can be responsible for mechanical transmission. Human-tohuman and animalto-human transmission are probably more common than suspected. B. hominis is a common intestinal parasite of humans

and animals, with a worldwide distribution. Depending on the geographic location, it may be detected in 1% to 40% of

fecal specimens. B. hominis may be the most common parasite found in the intestinal tract. Pathogenesis and Spectrum of

Disease B. hominis can cause diarrhea, cramps, nausea, fever, vomiting, abdominal pain, and urticaria and may require

therapy. A possible relationship between B. hominis and intestinal obstruction and perhaps even infective arthritis has

been suggested. In patients with other underlying conditions, the symptoms may be more pronounced. The incidence of

this organism appears to be higher than suspected in stools submitted for parasite examination. In symptomatic patients in

whom no other etiologic agent has been identified, B. hominis should certainly be considered the possible pathogen. It has

been suggested that proteases of genetic subtype 3 could be considered a virulence factor responsible for protein

degradation and subsequent pathogenesis.

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Laboratory Diagnosis:

Routine Methods. Routine stool examinations are very effective in recovering and identifying B. hominis; the permanent

stained smear is the procedure of choice, because examination of wet preparations may not easily reveal the organism. If

the fresh stool is rinsed in water before fixation (for the concentration method), B. hominis organisms, other than the

cysts, are destroyed, and a false-negative report may result. The organisms should be quantitated in the report (i.e., rare,

few, moderate, or many). It is also important to remember that other possible pathogens should be adequately ruled out

before a patient is treated for B. hominis.

Antigen Detection. Fecal immunoassays to detect B.hominis antigen have been developed but are not yet commercially

available. The technique currently used is the enzyme-linked immunosorbent assay (ELISA).

Antibody Detection. ELISA and fluorescent antibody tests have been developed to detect serum antibody to B. hominis

infections. A strong antibody response is consistent with the ability of this organism to cause symptoms. Also,

demonstration of serum antibody production both during and after B. hominis symptomatic disease is immunologic

evidence for the pathogenic role for this protozoan, although it may take 2 years or longer with chronic infections to

develop a serologic response.

Therapy:

Although clinical evidence is limited, studies have been done on the in vitro susceptibility of B. hominis to

numerousdrugs. Currently, metronidazole (Flagyl) appears to be the most appropriate drug. Diiodohydroxyquin

(Yodoxin) also has been effective, and dosage schedules for these two drugs are as recommended for other intestinal

protozoa. The development of new drug sensitivity assays may improve researchers’ ability to evaluate the activities of

various drugs against this organism.

Prevention:

 Prevention requires improved personal hygiene and sanitary conditions, in addition to proper disposal of fecal material.

FLAGELLATES:

The Mastigophora, or flagellates, have specialized locomotor organelles called flagella; these are long, thin, cytoplasmic

extensions that may vary in number and position, depending on the species. Different genera of flagellates may live in the

intestinal tract, the bloodstream, or various tissues.

 Four common species of flagellates are found in the intestinal tract: Giardia lamblia, Dientamoeba fragilis, Chilomastix

mesnili, and Pentatrichomonas hominis (Figures 48-16 to 48-23). Several other smaller, nonpathogenic flagellates, such

as Enteromonas hominis and Retortamonas intestinalis (see Figure 48-16) G. lamblia and D. fragilis are the flagellates

considered pathogenic. D. fragilis has been associated with diarrhea, nausea, vomiting, and other nonspecific intestinal

complaints. Trichomonas vaginalis is pathogenic but occurs in the urogenital tract. Trichomonas tenax is occasionally

found in the mouth and may be associated with poor oral hygiene.

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Giardia lamblia:

General Characteristics:

 G. lamblia is the most common cause of intestinal infection worldwide.

Other than B. hominis, G. lamblia (also called G. duodenalis and G. intestinalis) is probably the most common protozoan

organism identified in individuals in the United States. It causes symptoms ranging from mild diarrhea, flatulence, and

vague abdominal pains to acute, severe diarrhea to steatorrhea and a typical malabsorption syndrome. Various

documented waterborne and food-borne outbreaks have occurred during the past several years. A number of animals may

serve as reservoir hosts for G. lamblia. Differentiation of flagellates is based on overall shape, numbers, an arrangements

of flagella. Both the trophozoite and cyst stages are included in the life cycle of G. lamblia. Trophozoites divide by means

of longitudinal binary fission, producing two daughter trophozoites. The organism is found most commonly in the crypts

in the duodenum. Trophozoites are the intestinal dwelling stage and attach to the epithelium of the host villi by means of

the ventral disk. The attachment is substantial and results in disk “impression prints” when the organism detaches from

the surface of the epithelium.Trophozoites may remain attached to or may detach from the mucosal surface. Because the

epithelial surface sloughs off the tip of the villus every 72 hours, the trophozoites apparently detach at that time. G.

lamblia trophozoites are teardrop shaped and have been described as “someone looking at you”. Cyst formation takes

place as the organisms move down through the jejunum after exposure to biliary secretions. The trophozoites retract the

flagella into the axonemes, the cytoplasm becomes condensed, and the cyst wall is secreted (see Figures 13 to 14).

Figure 13 A

to C,

Trophozoites

of Giardia

lamblia. D to

F, Cysts of

G. lamblia.

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the cyst matures, the internal structures are doubled, so that when excystation occurs, the cytoplasm divides, producing

two trophozoites. Excystation occurs in the duodenum or appropriate culture medium.

Epidemiology:

 Transmission of G. lamblia occurs by ingestion of viable cysts. Although contaminated food or drink may be the

source, intimate contact with an infected individual may also result in transmission of the organism. This organism is

found more frequently in children or in groups living in close quarters. Outbreaks have been associated with poor

sanitation facilities or sanitation breakdowns, as evidenced by infections of travelers and campers. Limited information is

available on seasonal variations in giardiasis. Some data suggest an association with the cooler, wetter months of the year,

which may implicate environmental conditions as advantageous to cyst survival.

Certain occupations may place an individual at risk for infection, such as sewage and irrigation workers, who may be

exposed to infective cysts. In situations in which young children are grouped together, such as in nursery schools, an

increased incidence of exposure and subsequent infection of both children and staff members may be seen. A high

incidence of giardiasis occurs in patients with immunodeficiency syndromes, particularly in those with common variable

hypogammaglobulinemia.

Giardiasis is the most common cause of diarrhea in these patients and may be associated with mild to severe villus

atrophy An estimated 200 million people in Asia, Africa, and Latin America have symptomatic infections. In the United

States, approximately 20,000 cases are reported yearly. However, an estimated 2 million cases may occur annually.

Figure 14 A, Giardia lamblia trophozoite. B, G. lamblia trophozoite, iodine stain. C, G. lamblia cysts

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Pathogenesis and Spectrum of Disease:

 The incubation period for giardiasis ranges from approximately 12 to 20 days. Giardiasis may not be recognized as the

cause, because the infection mimics acute viral. enteritis, bacillary dysentery, bacterial or other food poisonings, acute

intestinal amebiasis, or “traveler’s diarrhea” (toxigenic Escherichia coli). However, the type of diarrhea plus the lack of

blood, mucus, and cellular exudate is consistent with giardiasis.

Asymptomatic Infection. Although the parasites in the crypts of the duodenal mucosa may reach very highnumbers, they

may not cause a pathologic condition. The organisms feed on the mucous secretions and do not penetrate the mucosa.

Although organisms have been seen in biopsy material obtained from inside the intestinal mucosa, others have been seen

attached to the epithelium.

Intestinal Disease. For unknown reasons, symptomatic patients may have irritation of the mucosal lining, increased

mucus secretion, and dehydration. The onset may be accompanied by nausea, anorexia, malaise, lowgrade fever, and

chills, in addition to a sudden onset of explosive, watery, foul-smelling diarrhea. Other symptoms include epigastric pain,

flatulence, and diarrhea with increased amounts of fat and mucus in the stool but no blood. Weight loss often

accompanies these symptoms. Although some speculate that the organisms coating the mucosal lining may act to prevent

fat absorption, this does not completely explain the prevention of the uptake of other substances normally absorbed at

other intestinal levels. Severe malabsorption has also been linked with isolated levothyroxine malabsorption, leading to

severe hypothyroidism and secondary impairmen of pancreatic function. In both cases, treatment with metronidazole led

to complete remission of symptoms. Occasionally the gallbladder is involved, resulting in gallbladder colic and jaundice.

G. lamblia also has been identified in bronchoalveolar lavage fluid.

Chronic Disease. The acute phase often is followed by a subacute or chronic phase. Symptoms include recurrent, brief

episodes of loose, foul-smelling stools and possibly increased distention and foul flatus. Between episodes of mushy

stools, the patient may have normal stools or may be constipated. Abdominal discomfort includes marked distention and

belching with a rotten-egg taste. Chronic disease must be differentiated from amebiasis; disease caused by other intestinal

parasites (e.g., D. fragilis, Cryptosporidium spp., Cyclospora cayetanensis, Isospora belli, Strongyloides stercoralis);

inflammatory bowel disease; and irritable colon. On the basis of symptoms such as upper intestinal discomfort, heartburn,

and belching, giardiasis must also be differentiated from duodenal ulcer, hiatal hernia, and gallbladder and pancreatic

disease.

Laboratory Diagnosis:

Routine Methods. Routine stool examinations are normally recommended for the recovery and identification of

intestinal protozoa. However, in the case of G. lamblia, because the organisms are attached securely to the mucosa by

means of the sucking disk, a series of five or six stool samples may be examined without recovering the organism. The

organisms also tend to be passed in the stool on a cyclic basis. The Entero-Test capsule can be helpful for recovering the

organisms, as can the duodenal aspirate. Although cysts often can be identified on the wet stool preparation, many

infections may be missed without examination of a permanent stained smear. If material from the string test or mucus

from a duodenal aspirate is submitted, it should be examined as a wet preparation for motility; however, motility may be

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represented by nothing more than a slight flutter of the flagella, because the organism is caught up in the mucus. After

diagnosis, the positive specimen can be preserved as a permanent stain.

Antigen Detection. The development of fecal immunoassays to detect Giardia antigen in stool has dramatically

improved the sensitivity seen with the routine O&P examination. The ELISA has been used to detect Giardia antigen in

feces. Fluorescent methods with monoclonal antibodies have also proven extremely sensitive and specific in detecting G.

lamblia in fecal specimens. Other products are available as a cartridge format that uses an immunochromatographic strip–

based detection system for G. lamblia and/or Cryptosporidium spp. Any antigen detection system should always be

reviewed for compatibility with stools submitted in preservatives rather than fresh specimens. Some limitations exist on

the use of kits for organisms in the genus Entamoeba. However, commercial reagent kits for detecting Giardia and

Cryptosporidium spp. can be used with formalin-based stool preservatives or with fresh or frozen specimens. Many of

these cartridge format tests provide an answer within 10 minutes and are equal to or better than other immunoassays with

regard to sensitivity and specificity. Many of these newer methods are being used to test patients suspected of having

giardiasis or those who may be involved in an outbreak. The detection of antigen in stool or visual identification of

organisms by using monoclonal antibody reagents indicates current infection. The value of these detection assays as

rapid, reliable immunodiagnostic procedures has been emphasized by the increase in Giardia infections and the greater

awareness of particular incidences (e.g., nursery school settings). Because the organisms are shed so sporadically, use of a

fecal immunoassay does not eliminate the need to analyze multiple stool specimens for sensitive detection of G. lamblia;

a minimum of two stools should be tested. If the first specimen is negative, it may represent a false negative.

Histology. Trophozoites are detectable in the duodenum and proximal jejunum; however, mucosal invasion generally has

been found in areas where necrosis or mechanical trauma was present.

Apparently, patients with giardiasis also have reduced mucosal surface areas compared with control patients.

Nucleic Acid-Based Techniques: Currently, there are no molecular-based assays commercially available for the

detection of G. lamblia.

Prevention:

 The most effective practice for preventing the spread of infection in the child care setting is thorough hand washing by

the children, staff members, and visitors. Filtration systems have also been recommended, although they have certain

drawbacks, such as clogging.

Chilomastix mesnili:

General Characteristics

The C. mesnili trophozoite is pear shaped, measuring 6 to 24 μm long and 4 to 8 μm wide. It has a single nucleus and a

distinct oral groove, or cytostome (mouth), close to the nucleus. Flagella are difficult to see without obvious motility in a

wet preparation. The morphology can be

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seen on the permanent stained smear; the cytostome may be visible in some trophozoites. The cysts are pear or lemon

shaped and range from 6 to 10 μm long and 4 to 6 μm wide . They have a single nucleus and a typical curved cytostomal

fibril, called the shepherd’s crook. The cyst’s definitive morphology can be seen on a permanent stain.

Epidemiology:

C. mesnili tends to have a cosmopolitan distribution, although it is found more frequently in warm

climates.Transmission occurs through ingestion of infective cysts.

Pathogeneis and Spectrum of Disease:

C. mesnili is considered nonpathogenic and does not cause disease.

Laboratory Diagnosis

 Although cysts sometimes can be seen in a wet preparation, definitive identification of C. mesnili relies on

examination of permanent stained smears.

Therapy

 Specific treatment is not recommended for C. mesnili. Because these nonpathogenic organisms are acquired

through fecal-oral contamination.

Prevention:

Prevention depends on adequate disposal of human excreta and improved personal hygiene, preventive

measures that apply to most of the intestinal protozoa.

Dientamoeba fragilis

General Characteristics:

 D. fragilis was described in 1918. D. fragilis tends to be common in some pediatric populations, and the incidence is

higher for patients under 20 years of age in some studies. Some speculate that D. fragilis may be infrequently recovered

and identified; a low incidence or absence from survey studies may be due to poor laboratory techniques and a general

lack of knowledge about the organism. The D. fragilis trophozoite is characterized as having one nucleus (20% to 40%) or

two nuclei (60% to 80%). The nuclear chromatin usually is fragmented into three to five granules, and normally no

peripheral chromatin is seen on the nuclear membrane. The cytoplasm is usually vacuolated and may contain ingested

debris and some large, uniform granules. The cytoplasm can also appear uniform and clean with few inclusions. Size and

shape vary considerably among organisms, even on a single smear(Figure 15)

Pathogenesis and Spectrum of Disease:

 D. fragilis has been associated with a wide range of symptoms. Case reports of children infected with D.

fragilis reveal a number of symptoms, including intermittent diarrhea, abdominal pain, nausea, anorexia,

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malaise, fatigue, poor weight gain, and unexplained eosinophilia. The most common symptoms in patients

infected with this parasite appear to be intermittent diarrhea and fatigue. In some patients, both the organism

and the symptoms persist or reappear until appropriate treatment is initiated.

Laboratory Diagnosis:

Routine Methods. Diagnosis of D. fragilis infections depends on proper collection and processing techniques

(a minimum of three fecal specimens). Although the survival time for this parasite has been reported as 24 to 48

hours in the trophozoite form, the survival time in terms of morphology is limited, and stool specimens must be

examined immediately or preserved in a suitable fixative soon after defecation. It is particularly important to

examine permanent stained smears of stool with an oil immersion objective (×100).These trophozoites have

been recovered in formed stool; therefore, a permanent stained smear must be prepared for every stool sample

submitted for examination. Organisms seen in direct wet mounts may appear as refractile, round forms; the

nuclear structure cannot be seen without examination of the permanent stained smear.

Antibody Detection. On indirect immunofluorescence assay, serum samples from patients with confirmed D.

fragilis infections showed positive titers, and all matched controls had positive titers ranging from 20 to 160.

However, these tests are not routinely used, nor are the reagents commercially available.

Therapy:

Clinical improvement has been seen in adults receiving tetracycline, and symptomatic relief has been observed

in children receiving diiodohydroxyquin, metronidazole, or tetracycline.

Prevention:

Fecal-oral transmission has not been documented; therefore, it is difficult to speculate about preventive

measures. However, if transmission does occur from ingestion of certain helminth eggs, the appropriate hygiene

and sanitary measures to prevent contamination with fecal material are appropriate.

Figure 15Trophozoites of Dientamoeba fragilis

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Pentatrichomonas hominis:

 P. hominis is probably the most commonly identified flagellate, other than G. lamblia and D. fragilis. P. hominis has

been recovered from all parts of the world, in both warm and temperate climates, and is considered nonpathogenic and

noninvasive. It is not known to have a cyst stage P. hominis trophozoites live in the cecum and feed on bacteria. The

trophozoite measures 5 to 15 μm long and 7 to 10 μm wide. It has a pyriform membrane, which aid identification of the

organism. The undulating membrane extends the entire length of the body, in contrast to that seen in the pathogen T.

vaginalis (on which the membrane extends halfway down the body).