Intestinal Protozoa



Intestinal Protozoa



Objectives



1. Describe the basic life cycle, distinguishing morphologic characteristics, clinical disease (if pathogenic), laboratory diagnosis, and prevention for the organisms listed in the following table.


2. Define and identify the following parasitic structures: trophozoite, cyst, oocyst, spore, pseudopodia, flagella, cilia, chromatoidal bars, karysome, central vacuole cyst form, axoneme, cytosotome, spiral groove, undulating membrane, ventral disc, shepherd’s crook, axostyle, macronucleus, micronucleus, apical complex, sporocyst, sporozoite, spore, sarcocyst, and polar tubule.


3. Define life cycle processes, including merogony, gametogony, sporogony, schizogony, and the associated organism or organisms and associated stages.


4. Correlate the parasitic life cycles with the specific diagnostic stages for the organisms listed.


5. Distinguish pathogenic from nonpathogenic protozoa.



Parasites to Be Considered



*Currently, the name Entamoeba histolytica is used to designate true pathogens, and Entamoeba dispar is used to designate nonpathogens. Unless trophozoites containing ingested red blood cells are seen (E. histolytica), the two organisms cannot be differentiated based on the morphology revealed by permanent stained smears of fecal specimens. Immunoassay kits are available for identifying the E. histolytica/E. dispar group and for differentiating E. histolytica from E. dispar.


†Although some individuals have changed the species designation for the genus Giardia to G. intestinalis or G. duodenalis, no general agreement has been reached. Therefore, for this list we retain the name Giardia lamblia.


The protozoa are unicellular eukaryotic organisms, most of which are microscopic. They have a number of specialized organelles that are responsible for life functions and that allow further division of the group into classes. Most protozoa multiply by binary fission and are ubiquitous worldwide.


The important characteristics of the intestinal protozoa are presented in Tables 48-1 to 48-7. The clinically relevant intestinal protozoa are generally considered to be Entamoeba histolytica, Blastocystis hominis, Giardia lamblia, Dientamoeba fragilis, Balantidium coli, Isospora (Cystoisospora) belli, Cryptosporidium spp., Cyclospora cayetanensis, and the microsporidia. Nonpathogenic intestinal protozoa are listed in various figures and tables but are not discussed in detail.



TABLE 48-1


Intestinal Protozoa: Trophozoites of Common Amebae





































































Characteristic Entamoeba histolytica Entamoeba dispar Entamoeba hartmanni Entamoeba coli Endolimax nana Iodamoeba bütschlii
Size* (diameter or length) 12-60 µm (usual range, 15-20 µm); invasive forms may be > 20 µm Same size range as E. histolytica 5-12 µm (usual range, 8-10 µm) 15-50 µm (usual range, 20-25 µm) 6-12 µm (usual range, 8-10 µm) 8-20 µm (usual range, 12-15 µm)
Motility Progressive, with hyaline, finger-like pseudopodia; motility may be rapid Same motility as E. histolytica Usually nonprogressive Sluggish, nondirectional; blunt, granular pseudopodia Sluggish, usually nonprogressive Sluggish, usually nonprogressive
Nucleus (single) and visibility Difficult to see in unstained preparations Difficult to see in unstained preparations Usually not seen in unstained preparations Often visible in unstained preparation Occasionally visible in unstained preparations Usually not visible in unstained preparations
Peripheral chromatin (stained) Fine granules, uniform in size and usually evenly distributed; may have beaded appearance Fine granules, uniform in size and usually evenly distributed; may have beaded appearance Nucleus may stain more darkly than in E. histolytica, although morphology is similar; chromatin may appear as solid ring rather than beaded (trichrome) May be clumped and unevenly arranged on the membrane; may also appear as solid, dark ring with no beads or clumps Usually no peripheral chromatin; nuclear chromatin may be quite variable Usually no peripheral chromatin
Karyosome (stained) Small, usually compact; centrally located but may also be eccentric Small, usually compact; centrally located but may also be eccentric Usually small and compact; may be centrally located or eccentric Large, not compact; may or may not be eccentric; may be diffuse and darkly stained Large, irregularly shaped; may appear blotlike; many nuclear variations are common; may mimic E. hartmanni or Dientamoeba fragilis Large, may be surrounded by refractile granules that are difficult to see (“basket nucleus”)
Cytoplasm appearance (stained) Finely granular, “ground glass” appearance; clear differentiation of ectoplasm and endoplasm; if present, vacuoles are usually small Finely granular, “ground glass” appearance; clear differentiation of ectoplasm and endoplasm; if present, vacuoles are usually small Finely granular Granular, with little differentiation into ectoplasm and endoplasm; usually vacuolated Granular, vacuolated Granular, may be heavily vacuolated
Inclusions (stained) Noninvasive organism may contain bacteria; presence of red blood cells (RBCs) is diagnostic; presence of RBCs is the only characteristic that allows differentiation between pathogenic E. histolytica and nonpathogenic E. dispar Organisms usually contain bacteria; RBCs not present in cytoplasm May contain bacteria; no RBCs Bacteria, yeast, other debris Bacteria Bacteria


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*These sizes refer to wet preparation measurements. Organisms on a permanent stained smear may be 1 to 1.5 µm smaller as a result of artificial shrinkage.



TABLE 48-2


Intestinal Protozoa—Cysts of Common Amebae





























































Characteristic Entamoeba Histolytica/Entamoeba dispar Entamoeba hartmanni Entamoeba coli Endolimax nana Iodamoeba bütschlii
Size* (diameter or length) 10-20 µm (usual range, 12-15 µm) 5-10 µm (usual range, 6-8 µm) 10-35 µm (usual range, 15-25 µm) 5-10 µm (usual range, 6-8 µm) 5-20 µm (usual range, 10-12 µm)
Shape Usually spherical Usually spherical Usually spherical; may be oval, triangular, or other shapes; may be distorted on permanent stained slide because of inadequate fixative penetration Usually oval, may be round May vary from oval to round; cyst may collapse because of large glycogen vacuole space
Nucleus (number and visibility) Mature cyst: 4 nuclei
Immature cyst: 1-2 nuclei; nuclear characteristics difficult to see on wet preparation		 Mature cyst: 4 nuclei; Immature cyst: 1-2 nuclei (2-nucleated cysts very common) Mature cyst: 8 (occasionally 16 or more nuclei may be seen) Immature cysts with 2 or more nuclei are occasionally seen Mature cyst: 4 Immature cysts: 2 Very rarely seen and may resemble cysts of Enteromonas hominis Mature cyst: 1
Peripheral chromatin (stained) Peripheral chromatin present; fine, uniform granules, evenly distributed; nuclear characteristics may not be as clearly visible as in trophozoite Fine granules evenly distributed on the membrane; nuclear characteristics may be difficult to see Coarsely granular; may be clumped and unevenly arranged on membrane; nuclear characteristics not as clearly defined as in trophozoite; may resemble E. histolytica No peripheral chromatin No peripheral chromatin
Karyosome (stained) Small, compact, usually centrally located but occasionally may be eccentric Small, compact, usually centrally located Large, may or may not be compact and/or eccentric; occasionally may be centrally located Smaller than karyosome seen in trophozoites but generally larger than those of genus Entamoeba Larger, usually eccentric refractile granules may be on one side of karyosome (“basket nucleus”)
Cytoplasm, chromatoidal bodies (stained) May be present; bodies usually elongate, with blunt, rounded, smooth edges; may be round or oval Usually present; bodies usually elongate with blunt, rounded, smooth edges; may be round or oval May be present (less frequently than in E. histolytica); splinter shaped with rough, pointed ends Rare chromatoidal bodies present; occasionally small granules or inclusions seen; fine linear chromatoidals may be faintly visible on well-stained smears No chromatoidal bodies present; occasionally small granules may be present
Glycogen (stained with iodine) May be diffuse or absent in mature cyst; clumped chromatin mass may be present in early cysts (stains reddish brown with iodine) May or may not be present, as in E. histolytica May be diffuse or absent in mature cyst; clumped mass occasionally seen in mature cysts (stains reddish brown with iodine) Usually diffuse if present (stains reddish brown with iodine) Large, compact, well-defined mass (stains reddish brown with iodine)


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*Wet preparation measurements; in permanent stains, organisms usually are 1 to 2 µm smaller.



TABLE 48-3


Intestinal Protozoa—Trophozoites of Flagellates





























































Protozoa Shape and Size Motility Number of Nuclei and Visibility Number of Flagella (Usually Difficult to See) Other Features
Dientamoeba fragilis Shaped like amebae; 5-15 µm (usual range, 9-12 µm) Usually nonprogressive; pseudopodia are angular, serrated, or broad lobed and almost transparent Percentage may vary, but 40% of organisms have 1 nucleus and 60% have 2 nuclei; not visible in unstained preparations; no peripheral chromatin; karyosome is composed of a cluster of 4-8 granules Internal flagella; not visible Cytoplasm finely granular and may be vacuolated with ingested bacteria, yeasts, and other debris; may be great variation in size and shape on a single smear
Giardia lamblia Pear-shaped; length 10-20 µm; width, 5-15 µm “Falling leaf” motility may be difficult to see if organism is in mucus; slight flutter of flagella may be visible using low light (duodenal aspirate or mucus from Entero-Test capsule) 2; not visible in unstained mounts 4 lateral; 2 ventral, 2 caudal Sucking disk occupies one half to three fourths of ventral surface; pear-shaped front view, spoon-shaped side view
Chilomastix mesnili Pear-shaped; length 6-24 µm (usual range, 10-15 µm); width, 4-8 µm Stiff, rotary 1; not visible in unstained mounts 3 anterior, 1 in cytostome Prominent cytostome extending one third to one half the length of the body; spiral groove across ventral surface
Pentatrichomonas hominis Pear-shaped; length 5-15 µm (usual range, 7-9 µm); width 7-10 µm Jerky, rapid 1; not visible in unstained mounts 3-5 anterior, 1 posterior Undulating membrane extends the length of the body; posterior flagellum extends free beyond end of body
Trichomonas tenax Pear shaped; length 5-12 µm; average of 6.5-7.5 µm; width, 7-9 µm Jerky, rapid 1; not visible in unstained mounts 4 anterior, 1 posterior Seen only in preparations from mouth; axostyle (slender rod) protrudes beyond the posterior end and may be visible; posterior flagellum extends only halfway down the body; no free end
Enteromonas hominis Oval; 4-10 µm (usual range, 8-9 µm); width, 5-6 µm Jerky 1; not visible in unstained mounts 3 anterior, 1 posterior One side of the body is flattened; posterior flagellum extends free posteriorly or laterally
Retortamonas intestinalis Pear-shaped or oval; 4-9 µm (usual range, 6-7 µm); width, 3-4 µm Jerky 1; not visible in unstained mount 1 anterior, 1 posterior Prominent cytostome extends approximately one half the length of the body


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TABLE 48-4


Intestinal Protozoa—Cysts of Flagellates









































Protozoa Size Shape Number of Nuclei Other Features
Dientamoeba fragilis, Pentatrichomonas hominis, Trichomonas tenax No cyst stage      
Giardia lamblia 8-19 µm (usual range, 11-14 µm); width, 7-10 µm Oval, ellipsoidal, or may appear round 4; not distinct in unstained preparations; usually located at one end Longitudinal fibers in cysts may be visible in unstained preparations; deep staining median bodies usually lie across the longitudinal fibers. Shrinkage is common, with the cytoplasm pulling away from the cyst wall; “halo” effect may be seen around the outside of the cyst wall because of shrinkage caused by dehydrating reagents
Chilomastix mesnili 6-10 µm (usual range, 7-9 µm); width, 4-6 µm Lemon or pear shaped with anterior hyaline knob 1; not distinct in unstained preparations Cytostome with supporting fibrils, usually visible in stained preparation; curved fibril along side of cytostome, usually referred to as a “shepherd’s crook”
Enteromonas hominis 4-10 µm (usual range, 6-8 µm); width, 4-6 µm Elongate or oval 1-4; usually 2 lying at opposite ends of cyst; not visible in unstained mounts Resembles Endolimax nana cyst; fibrils or flagella usually not seen
Retortamonas intestinalis 4-9 µm (usual range, 4-7 µm); width, 5 µm Pear shaped or slightly lemon shaped 1; not visible in unstained mounts Resembles Chilomastix cyst; shadow outline of cytostome with supporting fibrils extends above nucleus; “bird beak” fibril arrangement


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TABLE 48-6


Morphologic Criteria Used to Identify Intestinal Protozoa (Coccidia, Blastocystis hominis)
































Protozoa Shape and Size Other Features
Cryptosporidium spp.
 C. parvum (humans and animals)
 C. hominis (humans)		 Oocyst generally round, 4-6 µm; each mature oocyst contains four sporozoites Oocyst, diagnostic stage in stool, sporozoites occasionally visible within oocyst wall; acid-fast positive using modified acid-fast stains; various other stages in life cycle can be seen in biopsy specimens taken from gastrointestinal tract (brush border of epithelial cells) and other tissues; disseminated infection well documented in compromised host; oocysts immediately infective (in both formed and/or watery specimens); nosocomial infections documented; use enteric precautions for inpatients.
Cyclospora cayetanensis Oocyst generally round, 8-10 µm; oocysts are not mature, no visible internal structure; oocysts may appear wrinkled Oocyst, diagnostic stage in stool; acid-fast variable using modified acid-fast stains; color range from clear to deep purple (tremendous variation); best results obtained with decolorizing solution consisting of 1% acid, 3% maximum; oocysts may appear wrinkled (like crumpled cellophane); mimic Cryptosporidium oocysts but are twice as large.
Isospora (Cystoisospora) belli Ellipsoidal oocyst; range 20-30 µm long, 10-19 µm wide; sporocysts rarely seen broken out of oocysts but measure 9-11µm Mature oocyst contains two sporocysts with four sporozoites each; usual diagnostic stage in feces is immature oocyst containing spherical mass of protoplasm (intestinal tract). Oocysts are modified acid-fast positive. Whole oocyst may stain pink, but just the internal sporocysts stain if the oocyst is mature.
Sarcocystis hominis
S. suihominis
S. bovihominis		 Oocyst thin-walled and contains two mature sporocysts, each containing four sporozoites; frequently thin oocyst wall ruptures; ovoid sporocysts each measure 10-16 µm long and 7.5-12 µm wide Thin-walled oocyst or ovoid sporocysts occur in stool (intestinal tract)
S. “lindemanni” Shapes and sizes of skeletal and cardiac muscle sarcocysts vary considerably Sarcocysts contain several hundred to several thousand trophozoites, each measuring 12-16µm long and 4-9µm wide. Sarcocysts may also be divided into compartments by septa, which are not seen in Toxoplasma cysts (tissue/muscle).
Blastocystis hominis Organisms are generally round, measure approximately 6-40 µm, and are usually characterized by a large, central body (looks like a large vacuole); this stage has been called the central body form The more amebic form can be seen in diarrheal fluid but is difficult to identify. The central body forms vary tremendously in size, even on a single fecal smear; this is the most common form seen. Routine fecal examinations may indicate a positive rate much higher than other protozoa; some laboratories report figures of 20% and higher.


TABLE 48-7


Microsporidia That Cause Human Infection




































































































Microsporidia Immunocompromised Patient Immunocompetent Patient Comments
Common      
Enterocytozoon bieneusi Chronic diarrhea; wasting syndrome, cholangitis, acalculous cholecystitis, chronic sinusitis, chronic cough, pneumonitis; cause of diarrhea in organ transplant recipients Self-limiting diarrhea in adults and children; traveler’s diarrhea; asymptomatic carriers Short-term culture only; three strains identified but not named; AIDS patients with chronic diarrhea (present in 5% to 30% of patients when CD4 lymphocyte counts are very low); pigs, nonhuman primates
Encephalitozoon hellem Disseminated infection; keratoconjunctivitis; sinusitis, bronchitis, pneumonia, nephritis, ureteritis, cystitis, prostatitis, urethritis Possibly diarrhea Cultured in vitro; detected in people with traveler’s diarrhea and co-infection with E. bieneusi; pathogenicity unclear; spores not reported yet from stool; psittacine birds
Encephalitozoon intestinalis Chronic diarrhea, cholangiopathy; sinusitis, bronchitis, pneumonitis; nephritis, bone infection, nodular cutaneous lesions Self-limiting diarrhea; asymptomatic carriers Cultured in vitro; formerly Septata intestinalis; AIDS patients with chronic diarrhea; dogs, donkeys, pigs, cows, goats
Encephalitozoon cuniculi Disseminated infection; keratoconjunctivitis, sinusitis, bronchitis, pneumonia; nephritis; hepatitis, peritonitis, symptomatic and asymptomatic intestinal infection; encephalitis Not described. Two HIV-serologically negative children with seizure disorder (suspect E. cuniculi infection) presumably were immunocompromised Cultured in vitro; wide mammalian host range
Uncommon      
Pleistophora sp. Myositis (skeletal muscle) Not described Tend to infect fish
Pleistophora ronneafiei Myositis Not described  
Trachipleistophora hominis Myositis; myocarditis keratoconjunctivitis; sinusitis Keratitis Cultured in vitro; AIDS patients
Trachipleistophora anthropophthera Disseminated infection; keratitis Not described AIDS patients
Anncaliia connori Disseminated infection Not described (Formerly Nosema connori); often infects insects; disseminated in infant with SCID
Anncaliia vesicularum Myositis Not described Formerly Brachiola vesicularum
Anncaliia algerae Myositis; nodular cutaneous lesions Keratitis (Formerly Nosema algerae or Brachiola algerae); cultured in vitro; skin nodules in boy with acute lymphocytic leukemia; found in arthropods
Nosema ocularum Not described Keratitis HIV–serologically negative individual
Vittaforma corneae Disseminated infection; urinary tract infection Keratitis (Formerly Nosema corneum); cultured in vitro; non-HIV patient
Microsporidium ceylonensis* Not described Corneal ulcer, keratitis HIV–serologically negative individual, autopsy
Microsporidium africanum* Not described Corneal ulcer, keratitis HIV–serologically negative individual, autopsy
Microsporidia (not classified)   Keratoconjunctivitis in a contact lens wearer  


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AIDS, Acquired immunodeficiency syndrome; HIV, human immunodeficiency virus; SCID, severe combined immunodeficiency


*Microsporidium is a collective generic name for microsporidia that cannot be classified



Amebae


The class Sarcodina, or Amebae, includes the organisms capable of movement by means of cytoplasmic protrusions called pseudopodia. This group includes free-living organisms, in addition to nonpathogenic and pathogenic organisms found in the intestinal tract and other areas of the body (see Tables 48-1 and 48-2). Occasionally, when fresh stool material is examined as a direct wet mount, motile trophozoites may be seen, as well as other, nonparasitic structures (Figure 48-1).


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Figure 48-1 Various structures that may be seen in stool preparations. 1, 2, and 4, Blastocystis hominis. 3 and 5 to 8, Various yeast cells. 9, Macrophage with nucleus. 10 and 11, Deteriorated macrophage without nucleus. 12 and 13, Polymorphonuclear leukocytes. 14 and 15, Pollen grains. 16, Charcot-Leyden crystals. (Modified from Markell EK, Voge M: Medical parasitology, ed 5, Philadelphia, 1981, WB Saunders.)


Entamoeba histolytica


General Characteristics


Living trophozoites (motile feeding stage) of E. histolytica vary in size from about 12 to 60 µm in diameter. Organisms recovered from diarrheic or dysenteric stools generally are larger than those in formed stool from an asymptomatic individual. The motility has been described as rapid and unidirectional. Although this characteristic motility is often described, amebiasis rarely is diagnosed on the basis of motility seen in a direct mount. The cytoplasm is differentiated into a clear outer ectoplasm and a more granular inner endoplasm.


E. histolytica has directional and progressive motility, whereas the other amebae tend to move more slowly and at random. However, motility is rarely seen even in a fresh wet mount from a patient with diarrhea or dysentery. The cytoplasm is generally more finely granular, and the presence of red blood cells (RBCs) in the cytoplasm is considered diagnostic for E. histolytica (Figure 48-2).


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Figure 48-2 Entamoeba histolytica trophozoite containing ingested red blood cells.

Permanent stained smears demonstrate accurate morphology compared with other techniques. When the organism is examined on a permanent stained smear (trichrome or iron-hematoxylin stain), the morphologic characteristics of E. histolytica/E. dispar are readily seen. The nucleus is characterized by evenly arranged chromatin on the nuclear membrane and a small, compact, centrally located karyosome (condensed chromatin). As mentioned, the cytoplasm usually is described as finely granular, with few ingested bacteria and scant debris in vacuoles. As stated previously, in organisms isolated from a patient with dysentery, RBCs may be visible in the cytoplasm, a feature diagnostic for E. histolytica (Figure 48-3). Most often, infection with E. histolytica is diagnosed on the basis of the organism’s morphology, without the presence of RBCs.


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Figure 48-3 1, Trophozoite of Entamoeba histolytica (note ingested red blood cells). 2, Trophozoite of Entamoeba histolytica/Entamoeba dispar (morphology does not allow differentiation between the two species). 3 and 4, Early cysts of E. histolytica/E. dispar. 5 to 7, Cysts of E. histolytica/E. dispar. 8 and 9, Trophozoites of Entamoeba coli. 10 and 11, Early cysts of E. coli. 12 to 14, Cysts of E. coli. 15 and 16, Trophozoites of Entamoeba hartmanni. 17 and 18, Cysts of E. hartmanni. (From Garcia LS: Diagnostic medical parasitology, ed 4, Washington, DC, 2001, ASM Press. Illustrations 4 and 11 by Nobuko Kitamura.)

As part of the life cycle, the trophozoites may condense into a round mass (precyst), and a thin wall is secreted around the immature cyst. Two types of inclusions may be found in this immature cyst: a glycogen mass and highly refractile chromatoidal bars (refractile chromatin structure) with smooth, rounded edges. As the cyst matures (metacyst) (see Figure 48-3; Figure 48-4), nuclear division occurs, with the production of four nuclei. Often chromatoidals may be absent in the mature cyst. Cyst morphology does not differentiate E. histolytica from E. dispar. Cyst formation occurs only in the intestinal tract; once the stool has left the body, cyst formation does not occur. The one-, two-, and four-nucleated cysts are infective and represent the mode of transmission from one host to another.


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Figure 48-4 Entamoeba histolytica/Entamoeba dispar cyst.


Epidemiology


Amebiasis is caused by infection with the true pathogen, Entamoeba histolytica. Recent evidence from molecular studies confirms the differentiation of pathogenic E. histolytica and nonpathogenic E. dispar (Figure 48-5) as two distinct species. E. histolytica is considered the etiologic agent of amebic colitis and extraintestinal abscesses (amebic liver abscess), whereas nonpathogenic E. dispar produces no intestinal symptoms and is not invasive in humans.


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Figure 48-5 Entamoeba dispar trophozoite; no ingested red blood cells are present.

Infection is acquired through the fecal-oral route from infective cysts contained in the feces. These cysts can be ingested in contaminated food or drink or contracted from fomites or various sexual practices that could include accidental ingestion of fecal organisms. Flies and cockroaches have been implicated as mechanical vectors of contaminated fecal material.


The infection occurs worldwide, particularly in areas with poor sanitation. It is estimated that E. histolytica infection kills more than 100,000 people each year.



Pathogenesis and Spectrum of Disease


The pathogenesis of E. histolytica is related to the organism’s ability to directly lyse host cells and cause tissue destruction. Amebic lesions show evidence of cell lysis, tissue necrosis, and damage to the extracellular matrix. Evidence indicates that E. histolytica trophozoites interact with the host through a series of steps: adhesion to the target cell, phagocytosis, and cytopathic effect. Numerous other parasite factors also play a role. From the perspective of the host, E. histolytica induces both humoral and cellular immune responses; cell-mediated immunity is the major human host defense against this complement-resistant cytolytic protozoan.


The presentations of disease are seen with invasion of the intestinal mucosa or dissemination to other organs (most often the liver) or both. However, it is estimated that a small proportion (2% to 8%) of infected individuals have invasive disease beyond the lumen of the bowel. Also, organisms may be spontaneously eliminated with no disease symptoms.



Asymptomatic Infection.

Individuals harboring E. histolytica may have either a negative or a weak antibody titer and negative stools for occult blood. They also may be passing cysts detectable by a routine ova and parasite (O&P) examination. However, these cysts cannot be morphologically differentiated from those of the nonpathogen, E. dispar. Although trophozoites may be identified, they will not contain any phagocytized RBCs and cannot be differentiated from E. dispar. Molecular analyses of organisms isolated from asymptomatic individuals generally indicate that the isolates belong to the nonpathogenic E. dispar. Generally, asymptomatic patients never become symptomatic and may excrete cysts for a short period. This pattern is seen in patients infected with either nonpathogenic or pathogenic organisms.



Intestinal Disease.

The incubation period varies from a few days to a much longer time; in an area where E. histolytica is endemic, it is impossible to determine exactly when exposure to the organism occurred. Normally, the incubation time ranges from 1 to 4 weeks. Although the exact mode of mucosal penetration is not known, microscopic studies suggest that amebae have enzymes that lyse. The enzymes are released from lysosomes on the surface of the amebae or from enzymes in the tissue released from ruptured organisms. Amebic ulcers often develop released the cecum, appendix, or adjacent portion of the ascending colon; however, they can also be found in the sigmoidorectal area. Other lesions may occur from these primary sites. Ulcers usually are raised, with a small opening on the mucosal surface and a larger area of destruction below the surface (i.e., flask shaped). The mucosal lining may appear normal between ulcers.


Invasive intestinal amebiasis has four clinical forms, all of which are generally acute: dysentery (bloody diarrhea), fulminating colitis, amebic appendicitis, and ameboma of the colon. Dysentery and diarrhea account for 90% of cases of invasive intestinal amebiasis. The severity of symptoms can range from asymptomatic to severe symptoms that mimic ulcerative colitis. Patients with colicky abdominal pain, frequent bowel movements, and tenesmus (a persistent feeling of needing to pass stool) may present with a gradual onset of disease. With the onset of dysentery, bowel movements are frequent (up to 10 per day). Although dysentery may last for months, it varies from severe to mild and may lead to weight loss and prostration. In severe cases, symptoms may begin very suddenly and include profuse diarrhea, fever, and dehydration with electrolyte imbalances.



Hepatic Disease.

Blood flow from the mesenteric veins surrounding the intestine returns blood, via the portal vein, to the liver, most commonly the upper right lobe. Amebae in the submucosa can be carried by the bloodstream to the liver. The onset of symptoms may be gradual or sudden; upper right abdominal pain and fever (38° to 39°C) are the most consistent findings. Although the liver may be enlarged and tender, liver function tests may be normal or slightly abnormal (jaundice is rare). The abscess can be visualized radiologically, sonically, or by radionuclear scan; most patients have a single abscess in the right lobe of the liver. The most common complication is rupture of the abscess into the pleural space. An abscess also can extend into the peritoneum and through the skin. Hematogenous spread to the brain, lung, pericardium, and other sites is possible.


Pyogenic and amebic liver abscesses are the two most common hepatic abscesses. The severity of a pyogenic abscess depends on the bacterial source and the patient’s underlying condition. An amebic abscess tends to be more prevalent in those with suppressed cell-mediated immunity, men, and younger individuals. E. histolytica cysts and trophozoites are found in the stools of only a few patients with liver abscess. About 60% of these patients have no intestinal symptoms or any history of dysentery.



Laboratory Diagnosis



Routine Methods.

The standard O&P examination is the recommended procedure for recovery and identification of E. histolytica in stool specimens. Microscopic examination of a direct saline wet mount may reveal motile trophozoites, which may contain RBCs. However, trophozoites with RBCs are found only in a limited number of cases. In many patients who do not present with acute dysentery, trophozoites may be present but do not contain RBCs, and the organisms may be pathogenic E. histolytica or nonpathogenic E. dispar. An asymptomatic individual may have few trophozoites and possibly only cysts in the stool. Although the concentration technique is helpful for demonstrating cysts, the most important technique for the recovery and identification of protozoan organisms is the permanent stained smear (normally stained with trichrome or iron-hematoxylin). A minimum of three specimens collected over not more than 10 days may be required for identification.


Sigmoidoscopy specimens may be very helpful for identifying organisms. At least six areas of the mucosa should be sampled. Smears from these areas should be examined after permanent staining. However, these specimens are not considered a substitute for the recommended minimum of three stool specimens submitted for O&P examination (direct, concentration, and permanent stained smear).


Liver aspirate material is rarely examined, and often the specimen was not collected properly. Aspirated material must be aliquoted into several different containers as it is removed from the abscess; amebae may be found only in the last portion of the aspirated material, theoretically material from the abscess wall, not necrotic debris from the abscess center.



Antigen Detection.

A number of enzyme immunoassay reagents are commercially available, and their specificity and sensitivity provide excellent options for the clinical laboratory. These tests can differentiate the E. histolytica/E. dispar group from the rest of the Entamoeba species, such as nonpathogenic Entamoeba coli or Entamoeba hartmanni. Other test reagents can distinguish between E. histolytica and E. dispar (Entamoeba histolytica II test, TechLab, Blacksburg, VA or Entamoeba CELESA path, Cellabs, Brookvale, Australia). These kits require fresh or frozen stool; fecal preservatives have been found to interfere with the Entamoeba spp. reagents. Because of the specificity of the immunoassay reagents, the laboratory can inform the physician whether the E. histolytica/E. dispar organisms seen in the stool specimen are pathogenic E. histolytica or nonpathogenic E. dispar. Without the use of these reagents, the only way to identify true pathogenic E. histolytica morphologically is to detect the rare presence of trophozoites containing ingested RBCs. If the laboratory does not use these reagents, the presence of E. histolytica/E. dispar should be reported to the physician, accompanied by commentary related to the newer information on pathogenicity. Depending on each state’s requirements, pathogenic E. histolytica generally is reported to the public health facility (county).



Antibody Detection.

Serologic testing for intestinal disease is rarely recommended unless the patient has true dysentery; even in these cases, the titer (e.g., indirect hemagglutination) may be low and thus difficult to interpret. A definitive diagnosis of intestinal amebiasis should not be made without demonstrating the presence of the organisms. In patients suspected of having extraintestinal disease, serologic tests are diagnostically more effective. Indirect hemagglutination and indirect fluorescent antibody tests have been reported positive with titers greater than or equal to 1 : 256 and greater than or equal to 1 : 200, respectively, in almost 100% of cases of amebic liver abscess. In the absence of STAT serologic tests for amebiasis (tests with very short turnaround times for results), the decision on diagnosis must be made on clinical grounds and on the basis of results of other diagnostic tests, such as scans.





Therapy


Two classes of drugs are used in the treatment of amebic infections: luminal amebicides, such as iodoquinol or diloxanide furoate, and tissue amebicides, such as metronidazole, chloroquine, or dehydroemetine. Because of the differences in drug efficacy, it is important that the laboratory report indicates whether cysts, trophozoites, or both are present in the stool specimen.






Hepatic Disease.

Metronidazole plus one of the luminal drugs should be used to treat hepatic disease. Some other combinations also can be used; some contain emetine, in which case the patient must be monitored very carefully for possible cardiotoxicity. The importance of using both luminal and tissue amebicides is emphasized in patients with amebic liver abscesses. Asymptomatic colonization may be present with the true pathogen, E. histolytica. In patients treated with metronidazole (tissue amebicide), generally a 100% clinical response to the hepatic lesions is seen; however, failure to eliminate the organism from the bowel can lead to second bouts with invasive disease and intestinal colonization. Also, these carriers constitute a public health hazard because of continued shedding of infective cysts.



Prevention


Humans are the reservoir host for E. histolytica, and infection can be transmitted to other humans, primates, dogs, cats, and possibly pigs. Accidental consumption of sewage-contaminated water provides another route of infection. Amebiasis is considered a zoonotic waterborne infection. The cyst stages are resistant to environmental conditions and can remain viable in the soil for 8 days at 28° to 34°C, for 40 days at 2° to 6°C, and for 60 days at 0°C. Cysts normally are removed by sand filtration or destroyed by 200 ppm of iodine, 5% to 10% acetic acid, or boiling. However, an asymptomatic carrier who is a food handler generally is thought to play the most important role in transmission. Proper disposal of contaminated feces is considered the most important preventive measure. Although vaccines have been discussed as a possibility for eliminating human disease, nothing currently is available.



Entamoeba coli


General Characteristics


The life cycle of E. coli is identical to that of E. dispar. After digestion of infective cysts, the organisms excyst in the intestinal tract and produce trophozoites. Cyst formation occurs as the gut contents move through the intestinal tract; the excreted cysts are the infective form that is transmitted to humans and some animals.


E. coli trophozoites are somewhat larger than those of E. histolytica and E. dispar and range from 15 to 50 µm in diameter (see Table 48-1; Figures 48-6 and 48-7; see Figure 48-3). Motility is sluggish with broad, short pseudopods. In wet preparations, differentiating nonpathogenic E. coli from pathogenic E. histolytica is almost impossible. On the permanent stained smear viewed at a higher magnification, the cytoplasm is granular with vacuoles containing bacteria, yeasts, and other food materials. The nucleus has a large blotlike karyosome that may be eccentric rather than centrally located. The chromatin on the nuclear membrane tends to be clumped and irregular. Although rare, if RBCs are present in the intestinal tract, E. coli may ingest them rather than bacteria.


image
Figure 48-6 A to C, Trophozoites of Entamoeba histolytica (note ingested red blood cells). D, Trophozoite of E. histolytica/E. dispar. E, Early cyst of E. histolytica/E. dispar. F to H, Trophozoites of Entamoeba coli. I and J, Cysts of E. coli.

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Aug 25, 2016 | Posted by in MICROBIOLOGY | Comments Off on Intestinal Protozoa

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