Version May 2024
INTRODUCTION — Free-living amebas are environmental protozoan parasites with worldwide distribution. They exist in nature without the need for a host; they are not well adapted to parasitism and do not require a vector for transmission to humans or animals [1].
Four genera of amebas cause disease in humans: Naegleria (only N. fowleri), Acanthamoeba (several species), Balamuthia (only B. mandrillaris), and Sappinia (only S. pedata) [2,3]. All of these species cause central nervous system (CNS) infections; several species of Acanthamoeba may cause localized extra-CNS infections in immunocompetent hosts or disseminated infections in immunocompromised hosts.
Members of the genus Prototheca are also environmental pathogens that are rare causes of human infection; Prototheca are classified as green algae, which are unicellular eukaryotes that do not have chlorophyll.
Issues related to infections caused by free-living amebas and members of the genus Prototheca will be reviewed here.
CENTRAL NERVOUS SYSTEM INFECTIONS — Two distinctive clinical syndromes associated with free-living amebas are well recognized: primary amebic meningoencephalitis (PAM) and granulomatous amebic encephalitis (GAE).
Primary amebic meningoencephalitis — PAM is an acute hemorrhagic meningoencephalitis caused by N. fowleri. It is characterized by a fulminant course with clinical and laboratory features that resemble acute bacterial meningitis.
Naegleria fowleri
Epidemiology — N. fowleri is a thermophilic ameboflagellate protozoan parasite globally distributed in water and soil that lives in temperatures above 30°C and tolerates temperatures up to 45°C. The life cycle of N. fowleri includes three stages: the infective trophozoite, a transient non-dividing flagellate stage, and the cystic stage (figure 1) [4].
N. fowleri has been isolated from a variety of fresh- and warm water sources worldwide (lakes, ponds, rivers, streams, irrigation canals, hot springs, unchlorinated swimming pools, spas, aquaria, and sewage) but not from seawater [1,5-7]. History of recreational water activities is the most common risk factor for infection [1,8]. Activities such as swimming, diving, jumping, splashing, use of watercrafts, waterskiing, surfing, exposure to hot springs, and facial contact with mud have been reported in PAM cases [9]. Isolation of the ameba in the environment where cases have occurred is common [10]. Tap water exposures [11], including during ritual purification and nasal irrigation [12-14], have also been described. Asymptomatic children and patients with PAM may harbor the ameba in the nasal mucosa [15]. There is concern that global warming and changes in the ecosystems N. fowleri inhabit may lead to more cases worldwide [12,16,17]. Although no major changes in the incidence of PAM have been reported from the United States [18], numerous cases are being reported from the Indian subcontinent, mostly among elderly male patients not engaged in recreational activities [19].
PAM was initially reported in Australia in 1965; since then, it has been reported in more than 16 countries [20]. The total number of cases is unknown; one review included 381 cases [21]. In another series of well-documented PAM cases from the United States, 111 cases were reported. The mean patient age was 12 years (range 8 months to 66 years); 79 percent were males, and 62 percent were children. No evidence of immunosuppression was identified, and 87 percent of cases occurred during the summer [20]. Water exposure occurred in 81 percent of cases, including exposure to lakes and ponds in 74 percent. No clear source was identified in 19 percent of cases.
Exposure is more common than disease. There is a high prevalence of positive serology [22,23], while the risk of infection is approximately 2.6 cases per million exposures [24]. The reasons for the low prevalence of disease in humans despite frequent exposure are unknown.
Transmission to humans occurs primarily through inhalation of infested water. Activities that increase contact of the nasal mucosa with infested water predispose humans to infection. Trophozoites penetrate the olfactory mucosa, cross the cribriform plates, and ultimately reach the olfactory bulb. Water exposure in patients with tympanic membrane ruptures may also lead to PAM [25]. Cases of infection with no clear history of exposure have also been reported [20].
The risks and benefits should be weighed carefully when an organ from a patient with PAM is considered for transplantation. Thus far, acquisition of primary amebic encephalitis from solid organ transplantation has not been observed. In a comprehensive evaluation of 21 transplant recipients of solid organs donated by five patients who died of PAM, there was no evidence of PAM among the recipients [26]. However, extra–central nervous system (CNS) dissemination of N. fowleri infection may occur and might pose a risk for disease transmission via transplantation [26].
Clinical manifestations — PAM is an acute meningoencephalitis associated with an extremely high mortality rate. The case-fatality rate in one series including 111 cases was 99 percent [20]. In that study, the mean incubation period was 5 days (range 1 to 7 days), the mean time from onset of symptoms to death was 5.3 days (range 1 to 12 days), and the mean time from exposure to death was 9.9 days (range 6 to 17 days). In another series including 142 patients in the United States (median age 12 years), only 27 percent were diagnosed pre-mortem; 3 patients survived [27].
Common clinical manifestations include high fever, severe headache, photophobia, nausea, vomiting, behavioral abnormalities, seizures, and altered mental status. Smell and taste abnormalities may occur early and may reflect the migration of the organism through the olfactory mucosa. Physical examination demonstrates meningeal signs (Kerning and Brudzinski signs) and cranial nerve palsies indicating uncal herniation [2]. The progression of the disease is very rapid, with profound mental alteration and severe cranial hypertension leading to herniation and death in a matter of few days. Cardiac involvement with arrhythmia has also been reported [2].
Laboratory findings demonstrate leukocytosis with neutrophilia. Examination of the cerebrospinal fluid (CSF) demonstrates a yellowish-white to grayish color initially, which turns into frankly hemorrhagic fluid as the disease progresses [2]. The CSF white blood cell count ranges from 300 to 26,000 cells per mm3 with marked polymorphonuclear predominance; CSF red blood cells are usually seen, ranging from 250 to 24,600 cells per mm3. Glucose is usually markedly consumed in the CSF, and the protein concentration is usually elevated. CSF pressure is usually very high, ranging from 300 to 600 mm H20 [2]. Studies for bacterial pathogens are usually negative, including Gram stain and culture. However, secondary bacterial infections can be introduced by the ameba; this was illustrated in a report of Burkholderia cepacia (a soil-water transmitted pathogen) infection of the CSF in a patient with PAM [16].
Diagnosis — Clinicians should have a high index of suspicion for PAM in the setting of acute meningoencephalitis with negative results for bacteria and common viruses, which should prompt the search for amebas in the CSF. History of exposure increases the likelihood of PAM, but it is not always obtained.
The diagnosis of PAM is generally established via observation of motile trophozoites on examination of a centrifuged CSF wet mount preparation immediately after obtaining the sample. A phase-contrast microscope is useful to optimize visualization of the ameba. Trophozoites tend to immobilize when exposed to diluted fluid and may resemble leukocytes. Giemsa or trichrome stains help in identifying morphologic features of the trophozoites: 10 to 25 microns in size with a single nucleus and a centrally located nucleolus with no peripheral chromatin (picture 1 and picture 2) [4].
A multiplex polymerase chain reaction (PCR) facilitates for rapid detection; it can identify the deoxyribonucleic acid (DNA) of the major free-living amebas in CSF in about five hours, although this diagnostic methodology is not widely available [28]. N. fowleri can be isolated in non-nutrient agar coated with enteric bacteria. Serology is not useful for diagnosis.
Imaging findings on computed tomography (CT) and magnetic resonance imaging (MRI) are nonspecific. Brain edema, basilar meningeal enhancement, hydrocephalus, and areas of cerebral infarction have been described (image 1) [29]. Lesions in the brain tend to be located in the orbitofrontal and temporal lobes, base of the brain, cerebellum, and upper spinal cord [30]. A purulent exudative inflammation along the leptomeninges and extensive necrosis and hemorrhage of the parenchyma are characteristic pathological findings (picture 3) [31].
Brain tissue can be examined for trophozoites, and brain sections can be stained with immunofluorescent anti-N. fowleri antibodies [32].
Differential diagnosis — PAM must be distinguished from acute bacterial meningitis. The clinical manifestations and CSF findings of these two entities are similar. Epidemiological exposure may be useful in differentiating the two conditions. In general, patients with PAM receive treatment for bacterial meningitis before the definitive diagnosis is established. (See "Clinical features and diagnosis of acute bacterial meningitis in adults".)
Treatment — The optimal approach to treatment of PAM due to N. fowleri is uncertain; no studies evaluating the efficacy of single-drug or combination-drug regimens have been performed [33]. The rarity of the disease, delay in diagnosis, fulminant clinical course, and the difficulties in making a rapid diagnosis have hampered the evaluation of drug regimens. In theory, the best drug regimen should include an amebicidal drug (or a combination of drugs) with good in vitro activity that is capable of crossing the blood-brain barrier.
Based on articles published following well-documented survivors as well as extrapolation from experience with treatment of infection due to Acanthamoeba and Balamuthia infection, we suggest the following combination of drugs (in addition to steroids to control cerebral edema) [2,34-36]:
●Conventional amphotericin B (1.5 mg/kg/day intravenously [IV] +/- intrathecally)
●Rifampin (rifampicin; 10 mg/kg/day orally in three divided doses or every 24 hours)
●Fluconazole (10 mg/kg/day IV or orally)
●Miltefosine (<45 kg: 50 mg orally twice daily; ≥45 kg: 50 mg orally three times daily)
●Azithromycin (500 mg IV or orally)
The optimal duration of therapy is uncertain; reports range from 9 to 30 days.
Amphotericin B deoxycholate demonstrates the most favorable in vitro activity (with minimum inhibitory concentrations [MICs] in the range of 0.018 to 1 mcg/mL) and is considered the drug of choice. Administration of amphotericin B is warranted as soon as the diagnosis is considered [37]. Liposomal amphotericin B is less effective in animal models and has higher MICs than the conventional preparation [38-40].
Other active drugs in vitro include the azoles (fluconazole, voriconazole, ketoconazole, posaconazole, and itraconazole), rifampicin, miltefosine, and azithromycin [4]. The in vitro activity of rifampicin is uncertain [34]. Variations in virulence factors and sensitivity to antimicrobials have been postulated to explain clinical outcomes [33].
Approximately 11 survivors of PAM are reported in the literature [12,33,35,41,42]. In most of these cases, an early diagnosis was made, and treatment with amphotericin B was initiated promptly; the most frequently used regimen was amphotericin B plus rifampicin (10 mg/kg/day). Synergy of amphotericin B with azoles (fluconazole 10 mg/kg/day) and rifampin has also been described. In one case, intrathecal administration of amphotericin B and miconazole was used [34].
The use of miltefosine for treatment of PAM has been extrapolated from favorable clinical experience with its use in the treatment of Balamuthia mandrillaris and Acanthamoeba spp infections [43]. (See 'Acanthamoeba' below and 'Balamuthia mandrillaris' below.)
Azithromycin has both animal and in vitro efficacy against N. fowleri and appears to be synergistic when administered with amphotericin B [39,44].
Posaconazole has been found to inhibit growth of N. fowleri in vitro within 12 hours [45], and experimental murine infection has been cured with posaconazole, suggesting that in the future this drug may replace fluconazole as the azole of choice; further study is needed to confirm these findings.
Granulomatous amebic encephalitis — GAE is a rare subacute-chronic infection of the CNS caused by Acanthamoeba species, B. mandrillaris, and S. pedata.
Acanthamoeba — Acanthamoeba is the most common ameba found in nature and largely causes opportunistic infections in immunocompromised hosts. Predisposing conditions include diabetes, alcoholism, cirrhosis, human immunodeficiency virus (HIV) infection, chronic renal failure, systemic lupus, malignancy, chemotherapy, and organ transplantation including hematopoietic stem cell transplantation (mostly allogenic transplant recipients but also solid organ transplantation) [46,47]. Of more than 150 cases of GAE reported in the literature, 11 have been in immunocompetent hosts [48]. No cases of GAE acquired by solid organ transplantation have been described.
Epidemiology — Acanthamoeba can be found in soil (beach sands, flowerpot soil), air, and in a number of water environments rich in biofilms (brackish water, sewage, humidifiers, heating hospital environments, dental and dialysis units, contact lenses) [5]. Modes of transmission include inhalation of cysts carried by wind through the respiratory tract or direct skin contact followed by hematogenous spread [1]. No seasonality has been reported, and usually no history of water or soil exposure is elicited from GAE patients [4]. Acanthamoeba have been found in the nasal mucosa of both patients and asymptomatic individuals, suggesting a nasal route of spread [2,4].
The Acanthamoeba life cycle includes two stages: a vegetative trophozoite and a cyst (figure 2). Seventeen different genotypes have been identified, from T1 to T17; the majority of human cases are associated with the genotype T4, followed by genotypes T1, T10, and T12 [48]. A unique case associated with genotype T2 in an HIV-infected patient has also been reported [49].
Other infections can occur simultaneously with Acanthamoeba, including Legionella spp, Vibrio cholerae, B. cepacia, Listeria monocytogenes, and Mycobacterium spp [5,50]. A fatal case of multiple protozoan infections (Acanthamoeba spp, B. mandrillaris, and Toxoplasma gondii) in an HIV-infected patient has also been reported [51].
Clinical manifestations — GAE consists of a prolonged clinical course characterized by weeks or months of worsening headache, low-grade fever, visual disturbances, behavioral abnormalities, and focal neurologic deficits depending on the topographic location of lesions [2]. The incubation period is usually not known as it is often difficult to determine when exposure took place. Focal neurologic lesions mimicking toxoplasma encephalitis have been reported among patients with HIV [52]. Unlike patients with toxoplasmic encephalitis, these patients do not respond to conventional antitoxoplasmic therapy. As the disease progresses, intracranial pressure, seizures, loss of consciousness, coma, and death ensue.
Lumbar puncture is generally contraindicated for patients who present with a focal lesion associated with elevated intracranial pressure. For patients in whom the CSF can be obtained, nonspecific findings are observed, including mild pleocytosis with lymphocytic predominance, high protein concentration, and low or normal glucose concentration [53]. Acanthamoeba trophozoites are seldom seen in a CSF sample [1], but Giemsa stain of the CSF sediment may demonstrate trophozoites [54].
Diagnosis — Brain tissue is needed to confirm the diagnosis. Staining with hematoxylin-eosin demonstrates both trophozoites and cysts. Trophozoites are seen in the perivascular space; their size is 15 to 30 microns, and they have a large nucleolus with a centrally located nucleolus (picture 4) [53]. Thick-walled cysts (10 to 25 microns usually with a double layer) can be seen in tissue sections (picture 5). In patients with concomitant skin or pulmonary lesions, tissue samples from these organs may also demonstrate trophozoites [55]. Indirect immunofluorescence and immunoperoxidase stains are helpful for identifying Acanthamoeba in brain and other tissues [53,56].
Tissue samples can be cultured in non-nutrient agar coated by enteric bacteria [4]. PCR methods applied to brain or other tissues provide a more rapid diagnosis; however, availability of these methods is limited [28,57,58].
Brain CT scans and MRI show single or multiple space-occupying lesions with ring enhancement (image 2) [59]. Temporal and parietal lobes are most commonly affected.
Postmortem examination of the brain demonstrates significant edema and multiple hemorrhagic areas [4]. Microscopic examination reveals extensive necrosis and the presence of multinucleated giant cells. Granulomas can be seen in immunocompetent patients, and an abundance of both trophozoites and cysts located in perivascular areas are characteristic findings (picture 6) [4].
Differential diagnosis — The differential diagnosis of GAE is broad and includes pathogens that cause focal neurologic lesions in immunosuppressed patients. Brain biopsy is needed to establish the diagnosis of GAE. The differential diagnosis includes:
●Tuberculosis – Clinical manifestations of CNS tuberculosis include seizure or headache; occasionally, hemiplegia or signs of raised intracranial pressure are observed. Diagnostic tools include CSF examination and radiography. (See "Central nervous system tuberculosis: An overview".)
●Nocardiosis – CNS nocardiosis consists of parenchymal abscess that can occur in any region of the brain. Signs and symptoms are nonspecific; they may include fever, headache, meningismus, seizure, and/or focal neurologic deficits. The diagnosis requires isolation of the organism from a clinical specimen. (See "Nocardia infections: Epidemiology, clinical manifestations, and diagnosis".)
●Bacterial brain abscess – Clinical manifestations of brain abscess may include headache, focal neurologic deficits, and seizures. The diagnosis consists of imaging (MRI or CT) and lumbar puncture in the absence of mass effect. (See "Pathogenesis, clinical manifestations, and diagnosis of brain abscess".)
●Aspergillosis – CNS aspergillosis may occur in the setting of local extension of disease from the paranasal sinuses or via disseminated infection. Clinical manifestations include seizures or focal neurologic signs. Imaging findings include ring-enhancing lesions or cerebral infarction. Confirmation of the diagnosis requires biopsy. (See "Epidemiology and clinical manifestations of invasive aspergillosis".)
●Cryptococcosis – Clinical manifestations of cryptococcal meningoencephalitis may be acute or chronic and may include fever, headache, lethargy, and altered mental status. A lumbar puncture is necessary to establish the diagnosis. (See "Epidemiology, clinical manifestations, and diagnosis of Cryptococcus neoformans meningoencephalitis in patients with HIV".)
●Histoplasmosis – Clinical manifestations of CNS histoplasmosis include headache, altered mental status, and cranial nerve deficits. CSF examination demonstrates pleocytosis with a preponderance of lymphocytes; histopathology characteristically demonstrates granulomatous inflammation. (See "Diagnosis and treatment of disseminated histoplasmosis in patients without HIV".)
●Toxoplasmosis – Clinical manifestations of toxoplasmosis include headache, altered mental status, and confusion. The diagnosis is established via detection of the organism in a biopsy specimen. (See "Toxoplasmosis in patients with HIV".)
●Cysticercosis – Clinical manifestations of cysticercosis include seizures and headache. The diagnosis is largely based on clinical presentation and radiographic imaging. (See "Cysticercosis: Clinical manifestations and diagnosis".)
●Primary CNS lymphoma – Clinical manifestations of primary CNS lymphoma include focal neurologic deficits, neuropsychiatric symptoms, and signs of elevated intracranial pressure. The diagnosis is established via biopsy. (See "Primary central nervous system lymphoma: Clinical features, diagnosis, and extent of disease evaluation".)
●Balamuthia encephalitis – Clinical manifestations of Balamuthia encephalitis include fever, headache, seizure, cranial nerve dysfunction, or localized motor deficits. The diagnosis is established via brain biopsy. (See 'Balamuthia mandrillaris' below.)
Treatment — The optimal approach to treatment is uncertain [60]; therefore, combination regimens are preferred over single-drug regimens. We favor empiric treatment with a combination of miltefosine, fluconazole, and pentamidine isethionate. Trimethoprim-sulfamethoxazole, metronidazole, and a macrolide (azithromycin or clarithromycin) can be added to this regimen as well.
A number of drugs have in vitro activity and have resulted in success for a few patients when used alone or in combination: rifampicin, azoles (fluconazole, itraconazole, voriconazole), pentamidine isethionate, sulfadiazine, flucytosine, azithromycin, miltefosine, and caspofungin [53,61-68]. Variations in susceptibility and virulence may account for poor clinical responses [33].
Acanthamoeba are not inhibited in vitro by amphotericin B [66], although cure with amphotericin B plus rifampicin has been reported in one patient [69]. Cure using miltefosine monotherapy has also been described [67]. The benefit of hyperbaric oxygen is uncertain; its use in combination with pentamidine isethionate, miltefosine, fluconazole, trimethoprim-sulfamethoxazole, and metronidazole was successful in one patient [70].
Corifungin, a water-soluble polyene, has amebicidal in vitro activity against both the cyst and trophozoite of Acanthamoeba castellanii and may be a promising therapeutic agent [71].
Single cerebral lesions should be resected if possible [69].
Balamuthia mandrillaris — The life cycle is summarized in the figure (figure 3).
Epidemiology — B. mandrillaris infection was initially isolated from the brain of a baboon that died of meningoencephalitis at the San Diego Zoo in 1986 [72]. A serologic test was developed from this isolate and subsequently used to establish a posthumous diagnosis for 16 human cases of meningoencephalitis due to an unknown ameba (including cases from the United States, Mexico, Peru, Venezuela, and Australia).
The organism was further isolated and identified in 1993 [73]. Subsequently, about 200 cases have been reported from all continents except from Africa [74-77]. Most cases are reported in the Western hemisphere, with the highest concentration in South America and the United States [78-90]:
●In a review of 109 cases between 1974 and 2016 in the United States, males accounted for 68 percent of cases, the median age was 36 years (4 months to 91 years), and 55 percent were of Hispanic origin. The state where exposure occurred was documented in only 30 cases; California had the highest number of cases (12), followed by Arizona (4). A total of 85 percent of patients reported a history of exposure to soil, and 66 percent reported that they had been swimming prior to the onset of the illness [91].
●In a 2011 review of 35 cases in the United States, most came from the southern states: 15 from California, 6 in Arizona, 6 in Texas, and 3 from Mississippi [74]. Males were affected 2.5 times more frequently than females, and 55 percent of patients were younger than 15 years old. Subsequently, 9 additional cases of GAE due to Balamuthia have been described in the United States: 3 cases in California plus 1 in Texas, 1 in North Carolina, 1 in Arizona, 1 in Arkansas, 1 in Florida, and 1 in Colorado [51,92-99].
●In a retrospective study of 30 Peruvian patients, the mean age was 21 years, 50 percent were below 15 years of age, 67 percent were males, and none had evidence of immunosuppression. All patients had cutaneous lesions on the face and 73 percent of them had evidence of CNS involvement by the time of diagnosis [100].
The disease appears to occur more frequently among patients of Hispanic origin; possible explanations include genetic susceptibility or environmental exposure [74,101].
Balamuthia is most commonly isolated from soil [102-106]. In two reports, the ameba has been isolated from environmental sources near patients [103,106]. Infection has been correlated with activities including agricultural exposure, desert motorcycling, dirt biking, swimming, and gardening [74]. The disease may occur in immunocompetent as well as in immunosuppressed patients. Transmission via organ donation has been described in at least three clusters, including one asymptomatic donor [107-109].
Clinical manifestations — Three patterns of clinical presentation have been described: some patients develop an initial skin lesion followed by development of the neurologic manifestations in weeks or months, while others present with CNS involvement [91]; in addition, one case of purely cutaneous Balamuthia infection has been observed [110].
Early recognition of the skin lesion may facilitate diagnosis and treatment prior to development of CNS involvement. In some cases, the skin lesion may disappear in the absence of treatment; in such circumstances, subsequent CNS involvement still develops.
The frequency of the skin lesion varies according to the country of origin of the case. In a review of 109 B. mandrillaris cases in the United States between 1974 and 2016, a skin lesion was observed in 6 percent of cases [91]. In contrast, among 50 cases in Peru between 1990 and 2012, cutaneous involvement was present in 90 percent of cases [111]. A similar pattern has been observed in China and other parts of the world [112,113].
Patients with cutaneous involvement commonly present with a nonulcerated asymptomatic plaque, either single or with occasional satellite lesions (picture 7) [75,77,114,115]. A history of previous trauma can be elicited in many cases. The most common location is the central face over the nose; other locations described are the knee, thigh, chest, and elbow. Occasionally, the border is raised, giving the appearance of an annular lesion. The diameter may be one to several centimeters, and the color may vary from skin toned to slightly red with a violaceous hue. In some cases, the border is ill defined, suggesting soft tissue infiltration. If untreated, tissue infiltration may progress to involve the central face. Sensation is preserved, a sign that permits differentiation from tuberculoid leprosy.
CNS symptoms are due to thrombotic angiitis, leading to hemorrhage, infarction, and necrosis [79]. Patients who present with neurologic involvement commonly have fever and malaise [74]. Initial symptoms may include regression, unilateral headaches, focal seizures, cranial nerve dysfunction, or localized motor deficits [78,116]. Signs of increased intracranial pressure (diffuse headache, nausea, and vomiting) may be present, and later meningeal signs may develop, followed by progressive loss of consciousness (from stupor to coma). This progression generally occurs over 2 to 12 weeks.
A case series from the United States (109 cases over a period of 32 years) reported a median length of time from possible exposure to death of 24 days and median duration of hospital stay of 21 days. Most (99 percent) had encephalitis. Concomitant cutaneous and CNS lesions occurred in only 6 percent of cases. An immunosuppressive conditions was present in 39 percent of patients. Most patients presented with fever and focal neurologic manifestations. CSF showed findings suggestive of aseptic meningitis. Brain images demonstrated multifocal lesions (23 percent), edema (27 percent), and enhancing lesions (29 percent) [91].
Diagnosis — Recognition of the characteristic skin lesion on the central face facilitates early treatment and potential cure of an otherwise fatal disease. The differential diagnosis for cutaneous lesions involving sites other than the face is broader, and a high index of suspicion is required. Common histopathologic characteristics include the presence of a diffuse granulomatous reaction in the reticular dermis in the absence of epidermal changes, ill-defined granulomas, surrounding infiltrate rich in lymphocytes and plasma cells, and abundant multinucleated giant cells inside and outside granulomas (picture 8) [35]. Definitive diagnosis requires visualization of a trophozoite; these are present in 60 to 75 percent of cases and are scarce in number. In addition, trophozoites may be easily confused with histiocytes; visualization of the nucleus and nucleolus of the amoeba may be a distinguishing clue. The trophozoites are occasionally seen in a lacunar space and they can have either an oval or an irregular morphology, with a bubbly cytoplasm. In some cases, the nucleus may contain more than one nucleolus and may show a clear-cut central space in the nucleolus. This finding is often reported as a nuclear perforation and is a feature that distinguishes B. mandrillaris from Acanthamoeba [100].
Direct immunofluorescence or immunoperoxidase staining should be performed if possible [73,117]. These are species-specific assays with high sensitivity for detection of the ameba in tissue; they can be performed at the United States Centers for Disease Control and Prevention. A PCR technique for detection of amebic DNA material in human tissue has been developed based on a probe consisting of a primer pair from sequence data of the Balamuthia mitochondrial 16S ribosomal ribonucleic acid (rRNA) gene [118].
In the setting of CNS involvement, the diagnosis is much more challenging due to a broader differential diagnosis. Clinical clues include a hemorrhagic spinal tap and presence of subacute meningoencephalitis not responding to appropriate antibiotic therapy. CSF may demonstrate lymphocytic pleocytosis with mild to severe elevation of protein concentration and normal to low glucose concentration [116]. Balamuthia is rarely seen in the CSF. Real-time PCR on CSF can provide results in five hours, although this is not available in many areas [28]. Metagenomic next-generation sequencing has the potential to be incorporated into the initial workup of encephalitis, even if an amebic origin is unsuspected [99]. In one case, next-generation sequencing of blood and spinal fluid established the diagnosis, while in another case, thymine-adenine cloning using eukaryotic primers in paraffin-fixed brain tissue was used to confirm the diagnosis [119,120].
Imaging of CNS by CT or MRI typically demonstrates multiple lesions ranging from small, solid lesions to large, nodular lesions with ring enhancement (image 3) [121]. Intralesional hemorrhage is an important radiological clue.
Brain biopsy typically demonstrates granulomas with foamy macrophages and multinucleated giant cells accompanied by lymphocytes [78,117]. Amebae have a predilection for location in the vascular walls of capillaries and venules, and vascular damage may be present as tissue hemorrhage. Neutrophils may predominate, and amebas may be seen. Direct immunofluorescence or immunoperoxidase staining should also be performed if possible, as with skin biopsies [117].
Treatment — The optimal approach to treatment is uncertain; experience is based on a limited number of case reports [122-126]. Treatment with multiple drugs for a prolonged duration is warranted.
In the United States, six surviving patients have received a combination of pentamidine, flucytosine, fluconazole, and a macrolide (clarithromycin or azithromycin) plus any of the following: sulfadiazine, miltefosine, thioridazine, or liposomal amphotericin B [124-126]. Subsequently, another described a survivor treated with a number of potentially active antimicrobials, making it difficult to determine the role of each one of these drugs or in combination; the last combination included fluconazole, azithromycin, trimethoprim-sulfamethoxazole, and miltefosine [92].
In Peru, five surviving patients have been treated with a combination of fluconazole or itraconazole, in addition to albendazole and miltefosine [122]. Three patients had CNS involvement at the time of presentation, and two had cutaneous involvement only. Among 109 patients in the United States between 1974 and 2016, 10 patients survived [91]. The medications received were available for nine patients, including at least one each who was treated with azithromycin, clarithromycin, fluconazole, flucytosine, sulfadiazine, pentamidine, or miltefosine [91].
For purely cutaneous Balamuthia infection, one patient was treated with azithromycin, fluconazole, sulfadiazine, and a short period of miltefosine [110] while four patients in China were treated with the veterinarian drug diminazene [127]. In those four cases, clinical cure was achieved in one case with medications alone, and in two cases combining the drug and surgical excision of the lesion; the last patient died from liver failure after receiving the medication.
Wide resection of the skin lesion has been used as an additional therapy in some Peruvian cases. A similar approach may be useful for patients with single CNS lesions, followed by medical therapy [126].
Sappinia pedata — The genus Sappinia comprises two species, S. pedata and S. diploidea. The life cycle of these amebas involves a vegetative trophozoite and a cystic form; animal feces appear to be a necessary component of the life cycle [128]. A PCR assay can discriminate between the two species of Sappinia and can be incorporated into already-existing multiplex PCR methods to detect Acanthamoeba spp, B. mandrillaris, and N. fowleri, allowing for rapid detection and prompt initiation of therapy [129,130].
Only one human case of GAE due to Sappinia has been reported (attributed to S. diploidea) [131]. The patient was a previously healthy 38-year-old male farmer with no evidence of immunosuppression and who engaged in handling livestock; he presented with headache, vomiting, loss of consciousness, and blurry vision following a sinus infection. A single 2 cm focal lesion located in the posterior left temporal lobe was found and resected. The pathological examination demonstrated trophozoites (40 to 60 microns in diameter) with two nuclei (picture 9). The infection was likely acquired by inhalation [131].
The patient was treated with a combination of azithromycin (250 mg/day for 31 weeks), intravenous pentamidine isethionate (300 mg/day for 6 weeks), itraconazole (200 mg/day), and flucytosine (2.75 g four times a day for 25 weeks) with complete recovery.
ACANTHAMOEBA INFECTION OUTSIDE THE CENTRAL NERVOUS SYSTEM — Acanthamoeba can cause infection outside of the central nervous system (CNS); these include localized infections involving the skin, the nasopharyngeal area, and keratitis. Disseminated infections have also been reported.
Cutaneous infection — Acanthamoeba cutaneous infection is a rare clinical presentation seen mostly in immunocompromised hosts. It was initially described in an HIV-infected patient, but it has been associated with other immunosuppressive conditions; it has also been reported in healthy hosts [64,132,133]. The differential diagnosis is wide and includes viral, mycobacterial, fungal, and bacterial infections.
One report of five culture-confirmed cases collected over 11 years in a single institution in Peru noted that 80 percent were male, the mean age was 28 years, the clinical course was prolonged (duration of symptoms of approximately four months), and freshwater exposure was noted in two cases [133]. Three patients were HIV infected, had only cutaneous involvement, and died of opportunistic infections; two patients had no evidence of immunosuppression and developed granulomatous amebic encephalitis (GAE) [133]. Cutaneous lesions began as papules or nodules mostly in the lower limbs and evolved to necrotic ulcers in four patients. Other lesions included infiltrative plaques and cellulitis-like lesions. Lesions may be painful or painless. Pathological examination demonstrated an intense inflammatory reaction in the dermis with histiocyte predominance. Plasmocytes and neutrophils were also present, and Acanthamoeba trophozoites were abundant [133].
The optimal approach to treatment of localized cutaneous infections is uncertain. Regimens that have been tried with success include twice-daily applications of topical chlorhexidine gluconate and 2% ketoconazole cream, combined with systemic therapy including intravenous pentamidine isethionate, an azole (itraconazole or ketoconazole), and flucytosine [64,134]. Other successful combination regimens have included miltefosine, flucytosine, voriconazole, sulfadiazine [135], liposomal amphotericin B plus voriconazole [66], and miltefosine alone [136].
Nasopharyngeal infection — Acanthamoeba infection of the nasal mucosa and paranasal sinuses is a very rare condition that has been described primarily in severely immunosuppressed HIV-infected patients [137]. The disease is characterized by a chronic course with nasal discharge and sinusitis unresponsive to antimicrobials. Concomitant skin lesions are common [137]. Findings on the physical examination include purulent nasal secretion, hard brown crusts, and erosion of the nasal septum. Computed tomography (CT) imaging demonstrates mucosal thickening. The diagnosis is established via tissue biopsy stained with hematoxylin-eosin demonstrating a large number of trophozoites.
Medical treatment has been largely unsuccessful; few patients have responded. Management should consist of surgical excision combined with medical treatment. Drugs that have been administered successfully in one patient include flucytosine, pentamidine, amphotericin B, rifampicin, and ketoconazole [138].
Keratitis — Acanthamoeba may infect the cornea; keratitis most typically occurs among immunocompetent contact lens wearers who do not adhere to recommended cleaning procedures or as a result of direct inoculation following trauma.
Epidemiology — The main risk factor for acquiring amebic keratitis is wearing contact lenses for long periods of time [139]. Acanthamoeba has been isolated from contact lens cases, from which it is seeded into the eye, penetrating the corneal stroma and establishing chronic infection [33]. Use of nonsterile tap water in preparation of contact lens solutions is a well-recognized route of acquisition. In addition, contaminated commercial contact lens solutions have been associated with transmission in the United Kingdom and the United States [140,141]. Factors associated with amebic keratitis include use of recalled contact lens solutions, daily use of soft contact lenses, use of all-in-one (also call multi-purpose) lens solutions, showering while wearing contact lenses, and poor contact lens hygiene [142].
Clinical manifestations — Clinical manifestations include conjunctival hyperemia, tearing, foreign body sensation, pain, and photophobia. Usually a single eye is affected, but bilateral involvement has been observed. Epithelial irregularities and pseudodendritic epithelial lesions are considered early lesions [143]. Ring-shaped stromal infiltrates are characteristic late-stage lesions of Acanthamoeba keratitis [1].
Diagnosis — Trophozoites can be visualized via staining of cornea scrapings with fluorescent dye calcofluor [144]. Acanthamoeba keratitis can also be diagnosed by confocal microscopy [145-147]. Cornea scrapings can be cultured and molecular methods can be applied to differentiate species and quantify the amebic load. In general, polymerase chain reaction (PCR) methods are more sensitive than special stains and cultures [148]. Corneal biopsies can be cultured and examined microscopically for the presence of trophozoites and cysts. Four genotypes are responsible for most of the infections (T4, T3, T6, and T11) [149].
Treatment — Treatment should be instituted promptly, although the optimal approach is uncertain [33,150]; prolonged infection is a recognized risk factor for treatment failure. Combination therapy should be administered, including polyhexamethylene biguanide (0.02% PHMB) or biguanide-chlorhexidine in combination with propamidine (0.1%) or hexamidine (0.1%) [149,151]. Initially, drops should be applied frequently, with subsequent taper over three to four weeks. Debridement may also be warranted. Patients with poor response may benefit from topical riboflavin plus irradiation with ultraviolet light or topical voriconazole [152,153].
Challenges in treatment of Acanthamoeba keratitis include ameba encystation and development of resistance to medication. Prognostic factors for achieving best visual acuity response include good initial visual acuity, infection related to swimming, absence of epithelial defect, treatment with chlorhexidine, and no receipt of steroids [154]. Early use of topical steroid prior to initiation of specific antimicrobial therapy has been associated with poorer outcome [146].
Chronic refractory cases may require corneal transplantation. Unresponsive cases may require enucleation [33].
Acanthamoeba keratitis that is associated with an ipsilateral scleritis typically has poor clinical outcomes. Management with anti-inflammatory/immunosuppressive treatment is usually effective in reducing both scleral inflammation and symptoms and possibly reduces the likelihood of enucleation [155].
Disseminated infection — Acanthamoeba can cause disseminated infection in immunocompromised hosts [1]. The skin and lungs are most frequently affected, with or without CNS involvement [156]. Pulmonary involvement can be bilateral with patchy infiltrates [156]. The diagnosis is made by observing trophozoites and cysts in tissue samples. This condition is usually diagnosed postmortem. The optimal approach to treatment is uncertain; combination therapy is warranted. (See 'Treatment' above.)
PROTOTHECA — Human protothecosis is an extremely uncommon infectious disease caused by achlorophyllous algae of the family Chlorellaceae, genus Prototheca; it most commonly affects immunocompromised hosts [157,158]. The organisms are microscopic single-cell structures that divide asexually by endosporulation and binary fission. Five species are known, of which two affect humans: P. wickerhamii and P. zopfii [157,159]. The pathogenesis is not known.
Protothecosis is widely distributed across the globe. The algae are found in soil, decaying plants and water, and certain food items, and they also exist as a saprophyte in human skin, nails, and respiratory and digestive tracts [160]. The organism can infect humans as well as other mammals, such as cows (commonly causing mastitis), deer, dogs, and cats.
Epidemiology — Protothecosis can be acquired exogenously or, more rarely, endogenously in previously colonized patients. In most cases, the source of infection is exogenous contact with contaminated soil or water, commonly occurring after traumatic inoculation. Infection has also been described as a complication following surgery [161,162].
Protothecosis occurs most commonly among patients who are immunosuppressed, such as those who are on corticosteroids, have a hematologic malignancy, or have received an organ transplant. Patients who have other chronic underlying conditions (such as diabetes mellitus, alcoholism, or chronic peritoneal dialysis) are also at risk of infection, as are patients with intravascular catheters [157,163-165]. Neutrophil dysfunction increases the risk for infection, although neutropenia does not appear to be an important risk factor [157,166]. HIV infection does not appear to be an important risk factor for protothecosis [157,166].
In patients with intact immunity, the infection usually follows an indolent clinical course. Individuals whose occupation may predispose to contact with Prototheca species include rice farmers, fisherman, aquarium staff, and handlers of raw seafood [157]. Most cases are reported in patients >30 years of age, though all ages may be affected.
Clinical manifestations — The incubation period for Prototheca infection ranges from 10 days to 4 months. The disease can follow an acute or chronic clinical course and can present as localized or disseminated disease. Three clinical forms are recognized: cutaneous disease, olecranon bursitis, and disseminated disease.
●Cutaneous disease – Cutaneous involvement is the most common clinical presentation of protothecosis, accounting for more than half of cases; most occur in association with immunosuppression. The most commonly affected sites include exposed areas such the arms, legs, face, and neck. On the extremities, the infection has a predilection for distal areas such as fingers, wrist, dorsum of the hands, and forearms [167,168].
The typical manifestation consists of a single vesiculobullous and ulcerative lesion associated with purulent discharge and crusting. A number of other skin lesions have also been described, including translucent papules, erythematous plaques, verrucous plaques, nodules, and abscesses as well as hypopigmented and atrophic lesions (picture 10). A case of an eczematous plaque has been described [169]. Cutaneous disease usually consists of a single lesion, although cases of multiple lesions have been described. In addition, cases of disseminated disease may be associated with initial cutaneous involvement [163].
●Olecranon bursitis – Patients with olecranon bursitis due to Prototheca infection are usually immunocompetent, and there is frequently a history of previous trauma to the elbow. The predilection of infection for the olecranon bursa is not fully understood; the association may be explained in part by predisposition of this site to trauma. The clinical course is usually protracted, with symptoms appearing weeks after trauma. Characteristic features include local pain, tenderness, erythema, swelling, and mass formation with serosanguineous discharge [159,170,171].
●Disseminated disease – Disseminated disease occurs in association with immunosuppression or catheter use [164,165,172,173]. Involved organs include the skin, subcutaneous tissues, gastrointestinal tract, peritoneum, blood, and spleen [157,159,173,174]. Algemia presenting with fever and sepsis syndrome mimicking bacteremia has also been described [175], as has eosinophilic meningitis [176].
In addition to these clinical presentations, tenosynovitis following sclerosing therapy of varicose veins and chronic meningitis have also been reported [162,177].
Diagnosis — The diagnosis of protothecosis should be considered in patients in relevant risk groups, such as those who are immunosuppressed or have an indwelling intravascular catheter, and whose infection (cutaneous disease with suppurative or ulcerative lesions, olecranon bursitis, or disseminated disease) is not responsive to empiric antibacterial therapy.
The diagnosis of protothecosis is established based on microbiologic tests and/or direct identification in tissue specimens [157].
In the microbiology laboratory, Prototheca species can be isolated using a number of different culture media including Sabouraud dextrose agar (incubated at 25 to 35°C), beef infusion broth, blood agar, or brain infusion agar. Prototheca can be aerobic or microaerophilic and usually grow in three to five days; microscopic confirmation using lactophenol cotton blue demonstrates sporangia with endospores. Once isolated in culture, the organism can be identified morphologically via wet slide preparation with calcofluor white, which typically demonstrates sporangia (mother cells; 3 to 30 microns), from which approximately 2 to 20 endospores (9 to 11 microns) develop; the grouping of endospores forms a morula with a characteristic daisy shape (picture 11). P. wickerhamii has more symmetrical morula forms and smaller sporangia than P. zopfii [157]. Use of matrix-assisted laser desorption ionization–time of flight mass spectrometry (MALDI-TOF MS) for diagnosis of P. wickerhamii has been described in a case report [163].
Skin biopsy typically demonstrates pseudoepitheliomatous hyperplasia with granuloma formation [159]. The infiltrate may also include a variety of inflammatory cells, including lymphocytes, neutrophils, and eosinophils. Histopathological examination of the bursa demonstrates similar findings, with granuloma formation and predominance of giant cells and plasma cells. Prototheca are not observed readily via hematoxylin-eosin preparations; to enhance their features, special staining procedures such as periodic acid-Schiff and Gomori methenamine silver stains are needed (picture 11).
Differential diagnosis — The differential diagnosis of protothecosis includes:
●Bacterial infection – A number of bacterial pathogens may cause cutaneous lesion(s), olecranon bursitis, or systemic infection associated with intravascular catheters. Bacterial infections may be distinguished from protothecosis in that bacteria grow readily with routine culture techniques and generally respond clinically to antibacterial therapy. (See "Clinical manifestations of Staphylococcus aureus infection in adults".)
●Fungal infection (chromoblastomycosis, blastomycosis, cryptococcosis, paracoccidioidomycosis, coccidioidomycosis, and histoplasmosis) – Fungal pathogens may cause cutaneous lesion(s), olecranon bursitis, or systemic infection, particularly in immunocompromised hosts. Fungal infections are diagnosed via culture and/or histopathology. (See related topics.)
●Tuberculosis – Tuberculosis may present with extrapulmonary manifestations including cutaneous lesion(s), olecranon bursitis, or disseminated infection. The diagnosis is established via isolation in culture and supported by acid-fast smear and/or histopathology findings. (See "Clinical manifestations, diagnosis, and treatment of miliary tuberculosis".)
●Mycobacterium marinum infection – M. marinum is a mycobacterial infection typically associated with soft tissue or joint involvement in the setting of water exposure; the diagnosis is established via isolation in culture and supported by acid-fast smear and/or histopathology findings. (See "Soft tissue infections following water exposure".)
●Sporotrichosis – Sporotrichosis is a fungal infection that is typically associated with soft tissue involvement following inoculation of the organism from soil; the diagnosis is established via isolation in culture and supported by histopathology findings. (See "Clinical features and diagnosis of sporotrichosis".)
●Leishmaniasis – Leishmaniasis causes a spectrum of mucocutaneous disease; the diagnosis requires demonstration of the parasite in a clinical specimen by histology, culture, or polymerase chain reaction. (See "Cutaneous leishmaniasis: Clinical manifestations and diagnosis".)
Treatment — The optimal approach to treatment of protothecosis is uncertain; clinical data are limited, and the rarity of the infection precludes rigorous study.
In general, management of cutaneous disease should consist of surgical resection (for a single lesion or small number of lesions) and/or debridement (for necrotic ulcerative lesions). Subsequent systemic therapy is warranted for patients with persistent or deep infection. The optimal regimen is uncertain; options for systemic therapy include a combination of intravenous amphotericin B with oral tetracycline [178] or oral therapy with an azole antifungal agent (such as itraconazole, fluconazole, voriconazole, or posaconazole) [157]. The optimal duration of therapy is uncertain and should be guided by clinical and microbiologic response; in various reports, the duration of therapy ranges from days to weeks [157].
In general, management of olecranon bursitis should consist of bursectomy; failure of serial drainage has been described [175]. If bursectomy cannot be performed, local instillation of amphotericin B deoxycholate could be performed (1 mg amphotericin B in 1 mL of 5% dextrose in water) weekly for six weeks [179,180]. The optimal subsequent management is uncertain; administration of oral itraconazole, 200 mg once or twice daily, for at least two months is reasonable [181].
In general, management of disseminated protothecosis consists of intravenous amphotericin B, preferably given as a lipid formulation at a dosage of 3 to 5 mg/kg daily. One case report reported successfully treating cutaneous P. wickerhamii infection in an immunosuppressed patient with liposomal amphotericin and itraconazole [182]. Intravascular catheters, if present, must be removed. The duration of treatment depends on clinical and microbiologic responses; typically, weeks of treatment are required.
SUMMARY AND RECOMMENDATIONS
●Free-living amebas are environmental protozoan parasites with worldwide distribution. Four genera cause disease in humans: Naegleria, Acanthamoeba, Balamuthia, and Sappinia. All these species cause central nervous system (CNS) infections, but several species of Acanthamoeba may cause localized infections outside the CNS (in immunocompetent hosts) or disseminated infections (in immunocompromised hosts). (See 'Introduction' above.)
●Two CNS clinical syndromes are associated with free-living amebas: primary amebic meningoencephalitis (PAM) caused by Naegleria fowleri and granulomatous amebic encephalitis (GAE) caused by Acanthamoeba spp, Balamuthia mandrillaris, and Sappinia diploidea. (See 'Central nervous system infections' above.)
●Primary amebic meningoencephalitis is an acute hemorrhagic meningoencephalitis that occurs primarily in immunocompetent hosts; it resembles acute bacterial meningitis both clinically and in laboratory parameters. N. fowleri lives in freshwater environments. Transmission to humans occurs primarily through inhalation of infested water, and activities that increase contact of the nasal mucosa with infested water predispose to infection. The diagnosis is established by visualization of amebas in the cerebrospinal fluid (CSF). Few survivors have been reported; therefore, we favor prompt initiation of amphotericin B (1.5 mg/kg/day) as soon as the diagnosis is considered; adjunctive agents include fluconazole, rifampicin, azithromycin, and miltefosine. (See 'Primary amebic meningoencephalitis' above.)
●Acanthamoeba infection is transmitted via inhalation of cysts through the respiratory tract or by direct skin inoculation followed by hematogenous spread. GAE associated with Acanthamoeba almost exclusively occurs in immunocompromised hosts and is characterized by a subacute clinical presentation. The diagnosis is established by brain biopsy. Few survivors have been reported. The optimal approach to treatment is uncertain; combination regimens are preferred over single-drug regimens. We favor empiric treatment with a combination of miltefosine, fluconazole, and pentamidine isethionate. Trimethoprim-sulfamethoxazole, metronidazole, and a macrolide (azithromycin or clarithromycin) can be added to this regimen as well. (See 'Acanthamoeba' above.)
●Balamuthia mandrillaris is transmitted via contact with soil, and infection can occur among immunocompetent hosts. Two patterns of clinical presentation have been described: some patients develop an initial skin lesion followed by development of the neurologic manifestations in weeks or months, while others present with CNS involvement. (See 'Balamuthia mandrillaris' above.)
●Early recognition of the Balamuthia skin lesion may facilitate diagnosis and treatment prior to development of CNS involvement; the lesion consists of an asymptomatic nonulcerated plaque, most commonly on the central face over the nose (picture 7); the second most common location is in the knee area. The skin biopsy usually demonstrates a granulomatous reaction; amebae may be observed and confirmed by immunologic staining or polymerase chain reaction. Balamuthia CNS disease is more difficult to diagnose and has a broader differential diagnosis; brain biopsy may be required. The optimal approach to treatment is uncertain; treatment with multiple drugs for a prolonged duration is warranted. (See 'Balamuthia mandrillaris' above.)
●Acanthamoeba can cause infection outside the CNS; manifestations include cutaneous infection, nasopharyngeal infection, and keratitis. Keratitis usually occurs among immunocompetent contact lens wearers who do not adhere to proper cleaning procedures or as a result of direct inoculation following trauma. Clinical manifestations include conjunctival hyperemia, tearing, foreign body sensation, pain, and photophobia. Combination topical therapy should be administered, including polyhexamethylene biguanide or biguanide-chlorhexidine in combination with propamidine or hexamidine. Debridement may also be warranted. (See 'Acanthamoeba infection outside the central nervous system' above.)
●Human protothecosis is an uncommon infectious disease caused by achlorophyllous algae; it most commonly affects immunocompromised hosts. Clinical manifestations include cutaneous disease, olecranon bursitis, and disseminated disease. The diagnosis is established based on microbiologic tests and/or direct identification in tissue specimens. The optimal approach to treatment is uncertain and consists of antifungal therapy with or without surgery. (See 'Prototheca' above.)
ACKNOWLEDGMENT — The UpToDate editorial staff acknowledges Dr. Karin Leder, who contributed to earlier versions of this topic review.
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