The trophozoites can occur in two forms: ameboid and flagellate (Table 50-1, Figure 50-1). The size ranges from 7 to 35 µm. The diameter of the rounded forms is usually 15 µm. There is a large, central karyosome and no peripheral nuclear chromatin. The cytoplasm is somewhat granular and contains vacuoles. The ameboid form organisms change to the transient, pear-shaped flagellate form when they are transferred from culture or teased from tissue into water and maintained at a temperature of 27° to 37° C. These flagellate forms do not divide, but when the flagella are lost, the ameboid forms resume reproduction. Cysts are generally round, measuring from 7 to 15 µm with a thick double wall.

TABLE 50-1

Free-Living Amebae Causing Disease in Humans

	Acanthamoeba			Balamuthia	Naegleria
Morphology^a	Trophozoite Cyst Keratitis Acanthamoeba in brain			Trophozoite Cyst Balamuthia in brain	Trophozoite Flagellated form
Disease parameter	Granulomatous amebic encephalitis (GAE)	Acanthamoeba keratitis	Cutaneous lesions Sinusitis	GAE	Primary amebic meningoencephalitis (PAM)
General disease description	Chronic, protracted, slowly progressive CNS infection (may involve lungs); generally associated with individuals with underlying diseases	Painful, progressive, sight-threatening corneal disease; patients generally immunocompetent	Most common in patients with AIDS, with or without CNS involvement; those receiving immunosuppressive therapy for organ transplantation	Chronic, protracted, slowly progressive CNS infection (may involve lungs); generally associated with individuals with underlying diseases	Rare, but nearly always fatal infection; migration of amebae to brain through olfactory nerve; symptoms can mimic bacterial meningitis; death usually occurs 3-7 days after onset of symptoms; clinical suspicion based on history critical
Entry into body	Olfactory epithelium, respiratory tract, skin, sinuses	Corneal abrasion	Skin, sinuses, respiratory tract	Olfactory epithelium, skin, respiratory tract	Olfactory epithelium
Incubation period	Weeks to months	Days	Weeks to months	Weeks to months	Days
Clinical symptoms	Confusion, headache, stiff neck, irritability	Blurred vision, photophobia, inflammation, corneal ring, pain	Skin lesions, nodules, sinus lesions, sinusitis	Slurred speech, muscle weakness, headache, nausea, seizures	Headache, nausea, vomiting, confusion, fever, stiff neck, seizures, coma
Disease pathology	Focal necrosis, granulomas	Corneal ulceration	Granulomatous reaction in skin, inflammation	Multiple necrotic foci, inflammation, cerebral edema	Hemorrhagic necrosis
Diagnostic methods	Brain biopsy, CSF smear/wet prep, culture, indirect immunofluorescence on tissue,^b PCR^b	Corneal scrapings or biopsy, stain with calcofluor white, culture, confocal microscopy	Skin lesion biopsy, culture, indirect immunofluorescence of tissue^a	Brain biopsy, culture on mammalian cells, indirect immunofluorescence of tissue^a	Brain biopsy, CSF wet prep, culture, indirect immunofluorescence of tissue,^a PCR^a

^a Acanthamoeba in brain and Balamuthia in brain courtesy Dr. Govinda Visvesvara, Centers for Disease Control and Prevention.

^b Indirect immunofluorescence on tissue and PCR methods available from Centers for Disease Control and Prevention.

Figure 50-1 *Naegleria fowleri, Acanthamoeba* spp. Diagram of trophozoites and cysts *(upper row)*. Flagellate and cyst forms of *Naegleria fowleri; (lower row)* trophozoite and cyst of *Acanthamoeba* spp. (Illustration by Sharon Belkin; from Garcia LS: *Diagnostic medical parasitology,* ed 5, Washington, DC, 2007, ASM Press.)

Pathogenesis and Spectrum of Disease

Primary amebic meningoencephalitis (PAM) caused by N. fowleri is an acute, suppurative infection of the brain and meninges (Figure 50-2). With extremely rare exceptions, the disease is rapidly fatal in humans. The period between organism contact and onset of symptoms such as fever, headache, and rhinitis varies from a few days to 2 weeks. Early symptoms include vague upper respiratory tract distress, headache, lethargy, and occasionally olfactory problems. The acute phase includes sore throat; a stuffy, blocked, or discharging nose; and severe headache. Progressive symptoms include pyrexia, vomiting, and stiffness of the neck. Mental confusion and coma usually occur approximately 3 to 5 days before death, which is usually caused by cardiorespiratory arrest and pulmonary edema.

Figure 50-2 *Naegleria fowleri* in brain tissue. Hematoxylin and eosin stain. Note the large karyosome.

PAM resembles acute bacterial meningitis, and these conditions may be difficult to differentiate. Unfortunately, if the CSF Gram stain is interpreted incorrectly as a false positive, the resulting antibacterial therapy has no impact on the amebae and the patient will usually die within a few days.

Laboratory Diagnosis

Acanthamoeba Spp.

General Characteristics

Unlike N. fowleri, Acanthamoeba spp. do not have a flagellate stage in the life cycle, only the trophozoite and cyst. Several species of Acanthamoeba cause granulomatous amebic encephalitis (GAE), primarily in immunosuppressed, chronically ill, or otherwise debilitated individuals. These patients usually have no relevant history involving freshwater exposure. Acanthamoeba spp. also cause amebic keratitis, and it is estimated that the incidence in the United States may be one to two cases per million contact lens users. Apparently, the incidence of Acanthamoeba keratitis in the United Kingdom is 15 times higher than that in the United States and 7 times higher than that in Holland.

Motile organisms have spine-like pseudopods; there is a wide organism size range (25 to 40 µm), with the average diameter of the trophozoites being 30 µm. The nucleus has the typical large karyosome, similar to that found in N. fowleri. This morphologic characteristic can be seen on a wet preparation.

The cysts are usually round with a single nucleus, also having the large karyosome as in the trophozoite nucleus. The double wall is usually visible, with the slightly wrinkled outer cyst wall and what has been described as a polyhedral inner cyst wall. This cyst morphology is identifiable in organisms cultured on agar plates.

Pathogenesis and Spectrum of Disease

GAE

Meningoencephalitis caused by Acanthamoeba spp. may present as an acute suppurative inflammation of the brain and meninges similar to N. fowleri infection. The incubation period of GAE is unknown; several weeks or months are probably necessary to establish disease. The clinical course tends to be subacute or chronic and is usually associated with trauma or underlying disease, not as a result of swimming. GAE may present with symptoms of confusion, dizziness, drowsiness, nausea, vomiting, headache, lethargy, stiff neck, seizures, and sometimes hemiparesis. Unlike PAM caused by N. fowleri, both trophozoites and cysts are found throughout the tissue. Also, dissemination to other tissues such as the liver, kidneys, trachea, and adrenals can occur in immunocompromised individuals; or additional unusual sites also include the ear and necrotic bone from a bone graft of the mandible. Some patients, especially those with AIDS, can develop erythematous nodules, chronic ulcerative skin lesions, or abscesses.

Keratitis

Acanthamoeba spp. also cause keratitis and corneal ulceration. Clinicians need to consider acanthamoebic infection in the differential diagnosis of eye infections that are not responding to bacterial, fungal, or viral therapy. These infections are often due to direct exposure of the eyes to contaminated materials or solutions. Use of contact lenses is the leading risk factor for keratitis. Conditions that are linked with disease include the use of home-made saline solutions, poor contact lens hygiene, and corneal abrasions. A contact lens can act as a mechanical vector for transport of amebae present in the storage case onto the cornea. Subsequent multiplication and invasion of the tissue may occur. Decreased corneal sensation has contributed to the misdiagnosis of Acanthamoeba keratitis as herpes simplex keratitis. Acanthamoeba keratitis may be present as a secondary or opportunistic infection in patients with herpes simplex keratitis. Unfortunately, as a result, treatment can be delayed for 2 weeks to 3 months.

Laboratory Diagnosis

Routine Methods

The most effective culture approach uses non-nutrient agar plates with Page’s saline and an overlay growth of Escherichia coli on which the amebae feed. Tissue stains are also effective, and cysts isolated from cultures can be stained with Gomori’s silver methenamine, periodic acid-Schiff, and calcofluor white. Identification of Acanthamoebae in ocular samples and other tissues can be difficult, even for trained laboratory professionals; in histologic preparations, the organisms appear similar to keratoplasts, as well as neutrophils and monocytes. It has been estimated that up to 70% of clinical Acanthamoeba keratitis cases are misdiagnosed as viral keratitis. Also, the average time to diagnosis of keratitis attributable to Acanthamoeba infection can average 2.5 weeks longer for non–contact lens wearers than for contact lens users.

Other Methods

CSF or bronchoalveolar lavage fluid cytospin preparations can be used to look for amebae in patients with GAE or respiratory symptoms. The characteristic morphology of the Acanthamoeba trophozoites, such as the prominent nucleolus, contractile vacuole, and cytoplasmic vacuoles, can be seen more easily using trichrome or hematoxylin and eosin stains on fixed preparations after cytocentrifugation.

In the differential diagnosis of GAE, other space-occupying lesions of the central nervous system (CNS) (e.g., tumor, abscess, fungal infection) must also be considered. Predisposing conditions include Hodgkin’s disease, diabetes, alcoholism, pregnancy, and corticosteroid therapy. Organisms have also been found in the adrenal gland, brain, eyes, kidneys, liver, pancreas, skin, spleen, thyroid gland, and uterus.

In infections caused by Acanthamoeba spp., periodic acid-Schiff stains the cyst wall red and methenamine silver stains the cyst black. Normally, Naegleria and Acanthamoeba isolates are identified to the species level by a reference laboratory, such as the Centers for Disease Control and Prevention, using indirect fluorescent antibody procedures with a monoclonal or polyclonal antibody.

Therapy

Disseminated Infections

Trophozoites and cysts of Acanthamoeba isolates vary in their sensitivity to antimicrobial agents. They are sensitive in vitro to ketoconazole, pentamidine, hydroxystilbamidine, paromomycin, 5-fluorocytosine, polymyxin, sulfadiazine, trimethoprim-sulfamethoxazole, azithromycin, and extracts of medicinal plants, especially, to combinations of these drugs. In vitro testing confirms strain and species differences in sensitivity.

Acanthamoeba Keratitis

Prompt treatment is essential. Patients should be seen by an ophthalmologist immediately. Various prescription eye medications are available.

Balamuthia Mandrillaris

General Characteristics

Balamuthia mandrillaris is uncommon and was thought to be a harmless soil organism, with no relevance for infecting mammals. However, since the appearance of B. mandrillaris was first seen at the San Diego Wild Animal Park in a gibbon that died of meningoencephalitis, a number of primates, as well as dogs, sheep, and horses, have died of CNS infection caused by this organism. Approximately 100 cases of human amebic encephalitis worldwide have been identified with about half of the cases diagnosed within the United States. Death can occur from a week to several months after the onset of symptoms. Patients eventually die with a massive CNS infection. Genotyping studies indicate that lethal infections caused by B. mandrillaris are due to a single species with a global distribution.

The life cycle is similar to that of Acanthamoeba spp.; like Acanthamoeba spp., Balamuthia does not have a flagellated stage in the life cycle. Both trophozoites and cysts are found in CNS tissue, and their sizes are similar to those of Acanthamoeba trophozoites and cysts. It is difficult to differentiate Balamuthia from Acanthamoeba spp. in tissue sections under a light microscope. Using electron microscopy, the cysts are characterized by having three layers in the cyst wall: an outer wrinkled ectocyst, a middle structure–less mesocyst, and an inner thin endocyst. Under light microscopy, they appear to have two walls: an outer irregular wall and an inner round wall. In some cases, Balamuthia trophozoites in tissue sections appear to have more than one nucleolus in the nucleus. In such cases, it may be possible to distinguish Balamuthia amebae from Acanthamoeba organisms on the basis of nuclear morphology, because Acanthamoeba trophozoites have only one nucleolus.

Pathogenesis and Spectrum of Disease

The disease is very similar to GAE caused by Acanthamoeba spp. The clinical presentation is subacute or chronic and is usually not associated with swimming in freshwater. No characteristic clinical symptoms, laboratory findings, or radiologic indicators have been found to be diagnostic for GAE. Whether single or multiple, the lesions in the brain involve mainly the cerebral cortex and subcortical white matter. Symptoms include headache, nausea, vomiting, fever, visual disturbances, dysphagia, seizures, and hemiparesis. The clinical course ranges from a few days to several months. In immunocompetent hosts, an inflammatory response occurs; however, with rare exceptions, these patients also tend to die with severe CNS disease.

Laboratory Diagnosis

B. mandrillaris does not grow well on E. coli–seeded non-nutrient agar plates. However, these organisms can be cultured in mammalian cell cultures using monkey kidney cells and MRC, HEp-2, and diploid macrophage cell lines. Using human brain microvascular endothelial cells, B. mandrillaris has been cultured postmortem from brain and CSF from a case of granulomatous amebic meningoencephalitis. A cell-free growth medium is also commercially available. Although serum antibodies have been identified in infections, laboratory testing is not routinely available.

Therapy

In vitro studies indicate that B. mandrillaris is susceptible to pentamidine isethiocyanate and that patients may benefit from this treatment. Other studies indicate that ketoconazole, propamidine isethionate, clotrimazole, and certain biguanides have amebicidal activity.

Trichomonas Vaginalis

General Characteristics

Infection is acquired primarily through sexual intercourse, hence the need to diagnose and treat asymptomatic males. The organism is capable of survival for extended periods in a moist environment such as damp towels and underclothes; however, this mode of transmission is thought to be very rare. Infection with Trichomonas vaginalis occurs worldwide. It is estimated that 5 million women and 1 million men in the United States have trichomoniasis, with an estimated 7.4 million new cases occurring annually. The prevalence of trichomoniasis worldwide is estimated to be more than 170 million cases, which does not include the number of asymptomatic cases that remain untreated. In North America, more than 8 million new cases are reported yearly, with an estimated rate of asymptomatic cases being as high as 50%. Trichomoniasis is the primary non-viral sexually transmitted disease worldwide. Infection with T. vaginalis has major health consequences for women, including complications in pregnancy, association with cervical cancer, and predisposition to HIV infection.

The life cycle of T. vaginalis has a single trophozoite stage, and is very similar in morphology to other trichomonads (Table 50-2). The trophozoite is 7 to 23 µm long and 5 to 15 µm wide. The axostyle is usually obvious and protrudes through the bottom of the organism, whereas the undulating membrane ends halfway down the side of the trophozoite. There are a large number of granules evident along the axostyle.

TABLE 50-2

Characteristics of Trichomonas vaginalis

Shape and size	Pear-shaped, 7-23 µm long (average, 13 µm); width, 5-15 µm
Motility	Jerky, rapid
Number of nuclei and visibility	1; not visible in unstained mounts
Number of flagella (usually difficult to see)	3-5 anterior, 1 posterior
Other features	Seen in urine, urethral discharge, and vaginal smears; undulating membrane extends length of body; no free posterior flagellum; axostyle easily seen
Infective stage	Trophozoite
Usual location	Vagina (male, urethra)
Striking clinical findings	Leukorrhea, pruritus vulvae (thin white urethral discharge in male)
Other sites of infection	Urethra (prostate in male)
Stage usually recovered during clinical phase	Trophozoite only—no cyst
	Trichomonas, Giemsa stain

Pathogenesis and Spectrum of Disease

Growth of the organism results in inflammation and large numbers of trophozoites in the tissues and the secretions. As the acute infection becomes more chronic, the purulent discharge diminishes, with a decrease in the number of organisms. Symptoms such as vaginal or vulval pruritus and discharge are often sudden and occur during or after menstruation as a result of the increased vaginal acidity. Symptoms include vaginal discharge (42%), odor (50%), and edema or erythema (22% to 37%). Complaints also include dysuria and lower abdominal pain.

From 25% to 50% of infected women may be asymptomatic and have a normal vaginal pH of 3.8 to 4.2 and normal vaginal flora. Even in the carrier form, about 50% of women will become symptomatic during the following 6 months.

Although vaginitis is the most common finding in women with trichomoniasis, other complications include distention of a fallopian tube with pus, endometritis, infertility, low birth weight, and cervical erosion. There is also an increased association with HIV transmission and cervical dysplasia.

Dysuria, often the earliest symptom, occurs in about 20% of women with vaginal trichomoniasis. Infected males may be asymptomatic, or the infection may be self-limited, persistent, or result in recurring urethritis. In nonspecific urethritis, T. vaginalis has been detected in 10% to 20% of subjects and in 20% to 30% of those whose sexual partners had vaginitis. Once established, the infection persists for an extended period in females but only for about 10 days or less in males. T. vaginalis is the cause of 11% of all cases of non–gonococcal urethritis in males.

Respiratory distress has been reported in a full-term, normal male infant with T. vaginalis with severe respiratory problems following delivery. A wet preparation of thick, white sputum demonstrated few leukocytes and motile flagellates, which were identified as T. vaginalis. This study supports previous data confirming that the organism may cause neonatal pneumonia.

Laboratory Diagnosis

Humans are the only natural host for T. vaginalis, and organisms reside in the vagina and prostate; they usually do not survive outside the urogenital tract. The parasites feed on the mucosal surface of the vagina, where bacteria and leukocytes are abundant. The preferred pH for good parasitic growth in females is slightly alkaline or acidic (6.0 to 6.3 optimal), not the normal pH (3.8 to 4.2) of the healthy vagina. The organisms can also be recovered in urine, in urethral discharge, or after prostatic massage. Often, the organisms are recovered in centrifuged urine sediment from both male and female patients.

Wet Mounts

The identification of T. vaginalis is often based on the examination of wet preparations of vaginal and urethral discharges, urine, and prostatic secretions. This examination must be performed within 10 to 20 minutes after sample collection; if not, organisms lose motility and may not be identified. Several specimens may need to be examined for detection of the organisms. The sensitivity associated with wet mount examinations varies between 40% and more than 80%. Often, the percent detection from this procedure is quite low with limited sensitivity and specificity.

Stained Smears

Giemsa or Papanicolaou stain can be used. However, atypical cellular changes can be misinterpreted, particularly on the Papanicolaou smear. The organisms are routinely missed on Gram stains. The number of false-positive and false-negative results reported on the basis of stained smears strongly suggests that confirmation should be accomplished by observation of motile organisms either from the direct wet mount or from appropriate culture media.

Culture

A convenient plastic envelope method has been developed, which allows immediate examination and culture in one self-contained system. This system is commercially available as the InPouchTV (BIOMED Diagnostics, San Jose, Calif), which serves as the specimen transport container, the growth chamber during incubation, and the “slide” during microscopy. Once it is inoculated, it requires no opening for examination, and positive growth will occur within 5 days. The sensitivity of this system is reported to be superior to those of other available culture methods.

Antigen Detection

Several diagnostic tests have been developed, including the XenoStrip-Tv (Xenotope Diagnostics, Inc., San Antonio, Tex) and the OSOM Trichomonas Rapid Test (Sekisui Diagnostics, Houston, TX), both of which are more sensitive than the wet mount.

Molecular Diagnostics

The use of polymerase chain reaction (PCR) methods has led to improvements in T. vaginalis detection; nonviable organisms and cells and target sequences can also be detected. In addition to various culture and transport options, there are several other products available, including the Affirm VPIII probe from Becton Dickinson (Cockeysville, Md), the Quik-Tri/Can latex agglutination from Pan Bio InDX, Inc (Baltimore, Md), and the T.VAG DFA from Chemicon (Temecula, Calif). Depending on the patient population, client base, number of requests, and cost, one or more of these options may be appropriate for a particular diagnostic laboratory.

Therapy

Metronidazole is recommended for the treatment of urogenital trichomoniasis, although resistance to both metronidazole and other 5-nitromidazoles has been reported. It is also recommended that all sexual partners be treated simultaneously to avoid immediate reinfection. Metronidazole-resistant T. vaginalis has been implicated in an increased number of cases; unfortunately, this drug is currently the only drug approved for the treatment of trichomoniasis in the United States. Tinidazole has also been used for therapy.

Pentatrichomonas Hominis

Pentatrichomonas hominis derives its name on the basis of the morphologic structure of the trophozoite. The organism has five anterior flagella and a parabasal body. The organism is recovered worldwide and is considered to be nonpathogenic although it has been isolated from patients with diarrhea.

General Characteristics

There is no known cyst stage. The trophozoite resides in the large intestine where it feeds on bacteria. The organism resembles Trichomonas vaginalis, measuring 5 to 15 µm in length and 7 to 10 µm in width with an undulating membrane and an axostyle. The organism’s undulating membrane extends the entire length of the body, differentiating it from Trichomonas vaginalis.

Pathogenesis and Spectrum of Disease

Although the organism is considered nonpathogenic, it is the most commonly identified flagellate other than G. lamblia and Dientamoeba fragilis. The organism may be associated with infections in warm climates.

Laboratory Diagnosis

Therapy

Because the organism is nonpathogenic, it is important to differentiate it from Trichmonas vaginalis. However, if identified, no treatment is necessary.

Toxoplasma Gondii

Toxoplasma gondii is a protozoan parasite that infects most species of warm-blooded animals, including humans. Members of the cat family, Felidae, are the only known definitive hosts for the sexual stages of T. gondii and serve as the main reservoirs of infection. Cats become infected with T. gondii through carnivorism or by ingestion of oocysts. Outdoor cats are much more likely to become infected than domestic cats that are confined indoors. After tissue cysts or oocysts are ingested by the cat, organisms are released and invade epithelial cells of the cat small intestine, where they undergo an asexual cycle followed by a sexual cycle with the formation of oocysts, which are excreted in the feces. The uninfective oocyst takes 1 to 5 days after excretion to become infective. Cats shed oocysts for 1 to 2 weeks and large numbers may be shed, often more than 100,000 per gram of feces. Oocysts survive in the environment for several months to more than 1 year and are resistant to disinfectants, freezing, and drying. However, they are killed by heating to 70° C for 10 minutes. The life cycle in the cat takes approximately 19 to 48 days after infection with the oocysts but only 3 to 10 days after the ingestion of meat infected with cysts (e.g., a mouse) (Figure 50-3).

General Characteristics

There are three infectious stages of T. gondii: the tachyzoites (in groups or clones), the bradyzoites (in tissue cysts), and the sporozoites (in oocysts from cat feces). Tachyzoites rapidly multiply in any cell of the intermediate host and in epithelial cells of the definitive host (cats). Bradyzoites are found within the tissue cysts and usually multiply very slowly; the cyst may contain few to hundreds of organisms, and intramuscular cysts may reach 100 µm in size. The tissue cysts can be found in visceral organs such as the lungs, liver, and kidneys; however, they are more prevalent in the brain, eyes, and skeletal and cardiac muscle. Intact tissue cysts can persist for the life of the host and do not cause an inflammatory response.

Tachyzoites are crescent-shaped and are 2 to 3 µm wide by 4 to 8 µm long (Table 50-3). One end tends to be more rounded than the other. Giemsa is the stain of choice; the cytoplasm stains pale blue, and the nucleus stains red and is situated toward the broad end of the organism.

TABLE 50-3

Morphology of Toxoplasma gondii Stages Found in Humans

Stage	Description
Tachyzoites	Tachyzoites are crescent shaped and are 2-3 µm wide by 4-8 µm long. One end tends to be more rounded than the other. Giemsa is stain of choice; cytoplasm stains pale blue, and nucleus stains red and is situated toward broad end of organism. Tachyzoites are usually seen in early, more acute phases of infection. Tachyzoites rapidly multiply in any cell of the intermediate host (many animals and humans) and in nonintestinal epithelial cells of the definitive host (cats).
Bradyzoites Bone marrow, leukemia patient Bradyzoites in tissue	Bradyzoites are found within tissue cysts and multiply very slowly; cyst may contain few to hundreds of organisms, and intramuscular cysts may reach 100 µm in size. Although tissue cysts are seen in visceral organs such as lungs, liver, and kidneys, they are more common in brain, eyes, and skeletal and cardiac muscle.

Stage

Description

Tachyzoites

Tachyzoites are crescent shaped and are 2-3 µm wide by 4-8 µm long. One end tends to be more rounded than the other. Giemsa is stain of choice; cytoplasm stains pale blue, and nucleus stains red and is situated toward broad end of organism. Tachyzoites are usually seen in early, more acute phases of infection. Tachyzoites rapidly multiply in any cell of the intermediate host (many animals and humans) and in nonintestinal epithelial cells of the definitive host (cats).

Bradyzoites

Bone marrow, leukemia patient

Bradyzoites in tissue

Bradyzoites are found within tissue cysts and multiply very slowly; cyst may contain few to hundreds of organisms, and intramuscular cysts may reach 100 µm in size. Although tissue cysts are seen in visceral organs such as lungs, liver, and kidneys, they are more common in brain, eyes, and skeletal and cardiac muscle.

Cysts are formed in chronic infections, and the bradyzoites within the cyst wall are strongly periodic acid-Schiff positive. During the acute phase, there may be groups of tachyzoites that appear to be cysts; however, they are not strongly periodic acid-Schiff positive and have been termed pseudocysts.

Pathogenesis and Spectrum of Disease

As the tachyzoites actively grow, increase in number, and eventually rupture from the cell, they invade adjacent cells. This process creates additional lesions. Once the cysts are formed, the process becomes quiescent, with little or no multiplication and spread. In the immunocompromised or immunodeficient patient, a cyst rupture or primary exposure to the organisms often leads to lesions. The organisms can be disseminated via the lymphatic system and the bloodstream to other tissues.

Toxoplasmosis can be categorized into four groups: (1) acquired in the immunocompetent patient; (2) acquired or reactivated in the immunodeficient patient; (3) congenital; and (4) ocular.

Diagnosis and their interpretations may differ for each clinical category.

Immunocompetent Individuals

In almost all cases, no clinical symptoms are seen during the acute infection. However, 10% to 20% of these patients with acute infection may develop painless cervical lymphadenopathy and a flulike illness. This presentation is self-limited with symptoms resolving within weeks to months. Acute visceral manifestations are seen in rare cases. The majority of these patients remain asymptomatic or consider they have experienced nothing more than a common cold.

Immunocompromised Individuals

Infections in the compromised patient can lead to severe complications (Table 50-4). Underlying conditions that may impact the disease outcome include Hodgkin’s disease, non-Hodgkin’s lymphomas, leukemia, solid tumors, collagen vascular disease, organ transplantation, and AIDS. In the immunocompromised patient, the CNS is primarily involved, but these patients may also have myocarditis or pneumonitis. More than 50% of these patients will show altered mental state, motor impairment, seizures, abnormal reflexes, and other neurologic sequelae. Toxoplasmosis in patients being treated with immunosuppressive drugs may be due to either newly acquired or reactivated latent infection.

TABLE 50-4

People at Risk for Severe Toxoplasmosis

Category	Comments
Infants born to mothers who are first exposed to Toxoplasma infection several months before or during pregnancy	Mothers who are first exposed to Toxoplasma more than 6 months before becoming pregnant are not likely to pass infection to their children
Persons with severely weakened immune systems	Infection that occurred at any time during life can reactivate in an immunocompromised individual

In transplant recipients, the disease presentation depends on prior exposure to T. gondii by the donor and recipient, the type of organ involved, and the patient’s level of immunosuppression. Reactivation of a latent infection or an acute primary infection acquired directly from the transplanted organ can lead to severe disease. Stem cell transplant (SCT) recipients are particularly susceptible to severe toxoplasmosis, primarily attributable to reactivation of a previously acquired latent infection.

Before the use of highly active antiretroviral therapies (HAARTs), Toxoplasma encephalitis (TE) was a life-threatening opportunistic infection among patients with AIDS and was usually fatal if not treated. In AIDS patients with reactivated latent infections, psychiatric manifestations of T. gondii are seen, including altered mental status (60%) with delusions, auditory hallucinations, and thought disorders. T. gondii enhances HIV-1 replication within reservoir host cells and, at the same time, HIV-1 undermines acquired immunity to the parasite, promoting reactivation of chronic toxoplasmosis.

Congenital Infections

Congenital infection results when the mother acquires a primary infection during pregnancy. The majority of patients remain asymptomatic during the acute infection. However, congenital infections may be severe if the mother becomes infected during the first or second trimester. At birth or soon thereafter, symptoms in these infants may include retinochoroiditis, cerebral calcification, and occasionally hydrocephalus or microcephaly. Because treatment of the mother may reduce the severity of disease in the infant, prompt and accurate diagnosis is mandatory. Many infants who are asymptomatic at birth will subsequently develop symptoms of congenital toxoplasmosis; however, treatment may help prevent subsequent sequelae. Central nervous system involvement may not appear until several years later.

Ocular Infections

Ocular toxoplasmosis, an important cause of chorioretinitis, may be the result of congenital or acquired infection, acquired infection being more common than congenital infection. Patients with congenital infection may be asymptomatic until the second or third decade; at that point, cysts may rupture with lesions and develop in the eye. Chorioretinitis is characteristically bilateral in patients with congenital infection but is often unilateral in individuals with acute acquired T. gondii infection.

Laboratory Diagnosis

The most common method of diagnosis for toxoplasmosis is serologic testing for T. gondii– specific antibodies. Other procedures include PCR; examination of biopsy specimens, buffy coat cells, or cerebrospinal fluid; or isolation of the organism in tissue culture or in laboratory animals. It is important to remember that many individuals have been exposed to T. gondii and may have cysts within the tissues. Recovery of organisms from tissue culture or animal inoculation may be misleading, because the organisms may be isolated but may not be the etiologic agent of disease. However, two situations in which organism detection may be very significant are (1) tachyzoites in smears and/or tissue cultures inoculated from cerebrospinal fluid and (2) tachyzoites in patients with acute pulmonary disease and the demonstration of intracellular and extracellular tachyzoites in Giemsa-stained smears of bronchoalveolar lavage (BAL) fluid.

When laboratory personnel decide to initiate Toxoplasma-specific antibody testing or switch to a different antibody detection kit, the user must carefully review the manufacturer’s package insert and published literature for information on the sensitivity and specificity rates.

An in-laboratory comparison of kits should be performed, using positive and negative samples confirmed by a toxoplasmosis reference laboratory.

The serologic diagnosis of toxoplasmosis is very complex and has been discussed extensively in the literature (Wilson & McAuley, 2003); a number of additional procedures include enzyme immunoassays, enzyme-linked immunosorbent assays (ELISAs), direct agglutination, an immunosorbent agglutination assay, an indirect immunofluorescence assay (IFA), immunocapture, and immunoblot tests.