Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Amebic meningoencephalitides and keratitis: challenges in diagnosis and treatment Govinda S. Visvesvara Centers for Disease Control and Prevention, Atlanta, Georgia, USA Correspondence to Govinda S. Visvesvara, PhD, Centers for Disease Control and Prevention, Roybal Campus, M.S.-D66, 1600 Clifton Road, Atlanta, GA 30333, USA Tel: +1 404 718 4159; e-mail: [email protected] This article is dedicated to the memory of Dr Frederick L. Schuster, a good friend and an often collaborator. Current Opinion in Infectious Diseases 2010, 23:590–594 Purpose of review Acanthamoeba spp., Balamuthia mandrillaris, and Naegleria fowleri, although freeliving amebae, also cause devastating diseases in humans leading to death. Acanthamoeba spp. and B. mandrillaris cause granulomatous amebic encephalitis, cutaneous and nasopharyngeal as well as disseminated infection. Acanthamoeba also causes a vision-threatening infection of the cornea, Acanthamoeba keratitis, principally in contact lens wearers. N. fowleri causes an acute, fulminating infection of the central nervous system, primary amebic meningoencephalitis, in healthy children and young adults who indulge in aquatic activities in fresh water. This review focuses on the recent developments in the diagnosis and treatment and clinical management of the diseases caused by these amebae. Recent findings Development of a multiplex real-time PCR test has made it possible to simultaneously detect all the three free-living amebae in a sample. It is a rapid assay with a short turnaround time of just 4–5 h. An early diagnosis would be helpful in initiating potentially effective treatment. A recent study reported exciting results indicating that loading of rokitamycin in chitosan microspheres improves and prolongs the in-vitro antiAcanthamoeba activity of the drug. Summary Diagnoses of these infections are challenging and antimicrobial therapy is empirical, which often results in fatalities. Further research is needed to explore the possibility of a better drug delivery system that crosses the blood–brain barrier and effectively reach the central nervous system. Keywords Acanthamoeba keratitis, Acanthamoeba spp., Balamuthia mandrillaris, granulomatous amebic encephalitis, Naegleria fowleri, primary amebic meningoencephalitis Curr Opin Infect Dis 23:590–594 ß 2010 Wolters Kluwer Health | Lippincott Williams & Wilkins 0951-7375 Introduction Free-living amebae belonging to the genera Acanthamoeba, Balamuthia, and Naegleria cause infections of the central nervous system (CNS) of humans [1–5]. Acanthamoeba and Naegleria are ubiquitous in nature and occur worldwide. They feed on bacteria and multiply by binary fission. The environmental niche of Balamuthia is not well known. It has been isolated from soil and dust [1,3,5,6] and it is possibly present in water. Balamuthia does not feed on bacteria but may feed on small amebae and/or small yeasts. Several species of Acanthamoeba cause a chronic granulomatous infection leading to death, principally in immunocompromised persons, whereas the only known species of Balamuthia (B. mandrillaris) causes a chronic granulomatous infection in both immunocompromised and 0951-7375 ß 2010 Wolters Kluwer Health | Lippincott Williams & Wilkins immunocompetent persons. Additionally, both Acanthamoeba spp. and B. mandrillaris cause disseminated infections including skin, sinuses, lungs, prostate, and uterus [1–3]. Acanthamoeba spp. also causes an infection of the eye, Acanthamoeba keratitis, principally in contact lens wearers [1,2,5,7 –9]. Among the 40 species that belong to the genus Naegleria, only one species, Naegleria fowleri, causes an acute, fulminating and a necrotizing hemorrhagic meningoencephalitis called primary amebic meningoencephalitis (PAM) in otherwise healthy children and young adults [1,4,5]. Additionally, another ameba belonging to the genus Sappinia (Sappinia pedata) has been identified as a cause of encephalitis in a human [10]. Although, hundreds of infections have been caused by Acanthamoeba spp., B. mandrillaris, and N. fowleri, only a few persons have survived these infections because of delay in diagnosis due to lack of knowledge, and empirical treatment due to lack of optimal therapy. The basic DOI:10.1097/QCO.0b013e32833ed78b Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited. Amebic meningoencephalitides and keratitis Visvesvara 591 biology of the amebae and available information on the therapy used will be discussed. Acanthamoeba spp. Acanthamoeba has two stages in its life cycle; a trophozoite stage in which the ameba feeds and reproduces by binary fission, and a double-walled cyst stage that is resistant to environmental pressures [1,2,11], probably due to the presence of complex carbohydrates in the cyst wall [12]. As many as 24 species are included in three groups based on morphology and 16 sequence types based on sequencing of the 18S rRNA gene [2,13]. Although several different species and several genotypes cause infections, the most prevalent amebae that cause infections are included in group II and sequence type T4 [2]. Therapy used for systemic Acanthamoeba infections Although a number of antimicrobials have been used in the past to treat CNS as well as nasopharyngeal infections no drug has been found to be useful. A few patients have recovered after a combination of multiple drugs including fluconazole, itraconazole, ketoconazole, pentamidine isethionate, trimethoprim–sulfamethoxazole (co-trimoxazole), sulfadiazine, and 5-fluorocytosine (flucytosine), but the outlook for most patients remains hopeless especially because of HIV/AIDS and other opportunistic infections [2,14,15]. Recently, however, several patients recovered after treatment with a combination of drugs partly because of early diagnosis [16–21]. Treatment of cutaneous infections without spreading to CNS has fared better. Some successful treatment regimens have included topical chlorhexidine and ketoconazole cream followed by pentamidine isethionate, 5-fluorocytosine, and itraconazole [1,2,14,18,22]. A number of antimicrobials have been tested in vitro, against different Acanthamoeba isolates [2,23]. For example, miltefosine (hexadecylphosphocholine) was found to be effective in vitro against Acanthamoeba isolates [24,25] and was found to be useful in systemic infections [21] and amebic keratitis [26]. In a recent article, Walochnik et al. [27] have demonstrated that Acanthamoeba penetrates the recently developed organ system culture of a skin equivalent within 24 h and that miltefosine at 50 mmol/l concentration neutralizes Acanthamoeba in such a system and that it is nontoxic to mammalian cells. In vivo, even a 6% (150 mmol/l) solution is well tolerated by human skin [27]. Miltefosine is also stable for at least 1 year and it is also much less toxic than chlorhexidine gluconate, an anti-Acanthamoeba drug used currently for Acanthamoeba keratitis. Mechanisms of efficacy are probably due to inhibition of lipid remodeling, phospholipid biosynthesis, intracellular sig- naling, targeting of cellular membranes, and/or induction of apoptosis [27,28]. Further, a number of factors such as the virulence of the amebae, the infectious dose, time at which therapeutic intervention started, the immune status of the host, and the use of steroids during the course of the infection contribute to the success or otherwise of the intervention program. Delivery of drugs to the affected areas, specifically the brain Many of the drugs that are currently used to target the infectious agents do not cross the blood–brain barrier; hence suitable antiamebic concentrations are unavailable in the CNS to neutralize the amebae. An effort has been made recently to mitigate this by using inert polymers such as chitosan microspheres as vehicles for antimicrobials. Chitosan particles loaded with pharmaceuticals could be useful in transporting the drug to the ocular surface or to the brain when sprayed into the nasal epithelium. Loading of rokitamycin in chitosan microspheres improves and prolongs the anti-Acanthamoeba activity when compared with the free drug [29]. Further research has shown that the chitosan derivative, diethylaminomethyl(diethyldimethylene ammonium) nmethylchitosan has better properties in regard to size, mucoadhesiveness, and in-vitro release of rokitamycin and thus is more suitable for ocular or nasal administration. Additionally, this chitosan derivative had no toxic effect in vitro to human umbilical vein endothelial cells [30]. Treatment for Acanthamoeba keratitis Prognosis for Acanthamoeba keratitis is better and has considerably improved during the past few years. This is probably due to improvement in diagnosis, better awareness, and understanding of the infection by the eye care providers (especially with regard to the association of Acanthamoeba keratitis and contact lenses) resulting in the timely initiation of antimicrobials. Therefore, prospects for the recovery are excellent. The drawback however is the possibility of conversion of the trophic stage to the cyst stage (reviewed in [9]), leading to the recurrence of infection. The cyst, because of its double wall and the presence of cellulose in the inner wall, is resistant to drugs (reviewed in [6,12]). The drugs currently recommended for the treatment of Acanthamoeba keratitis are two biguanides, polyhexamethylene biguanide (PHMB) and chlorhexidine, and two diamidines, propamidine isethionate (Brolene; Sanofi-Aventis, Paris, France) and hexamidine (Desomedine; Chauvin-Blache, Montpellier, France): PHMB (0.02%) or chlorhexidine (0.02%) along with a diamidine (either 0.1% propamidine or 0.1% hexamidine). These are used hourly as drops day and night for 48 h initially followed by hourly drops by Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited. 592 Antimicrobial agents day only for another 72 h. Because of epithelial toxicity this schedule is reduced to 2 h by day 5 for 3–4 weeks and then tapered off [6]. Although use of topical corticosteroids is controversial, it is necessary to control corneal inflammation, severe pain, and scleritis [6]. Some of the earlier studies also included topical neomycin although it has no activity against cysts of Acanthamoeba. Recent in-vitro testing of voriconazole, a triazole compound, showed some activity against the trophozoites of several Acanthamoeba isolates indicating that this triazole may be of use therapeutically [25]. Indeed, it has been shown that use of 1% voriconazole in three eyes of two patients as a second-line treatment of Acanthamoeba keratitis cleared the infection which was previously unresponsive to treatment with chlorhexidine and hexamidine [31]. Balamuthia mandrillaris B. mandrillaris was isolated in 1986 from the brain of a pregnant mandrill baboon that died of encephalitis. It was initially described as a leptomyxid ameba but was later recognized as distinct and different from the leptomyxids and was described as a new genus and species, B. mandrillaris [1,3,5]. An immunofluorescent test developed using an antibody made in rabbits against this ameba identified a number of human infections as caused by B. mandrillaris, although they were previously erroneously identified as either due to Acanthamoeba or an unknown ameba. B. mandrillaris causes granulomatous amebic encephalitis (GAE) similar to that caused by Acanthamoeba and can occur at any time of the year. Diagnosis is difficult and most cases are recognized postmortem [1,3,5]. B. mandrillaris has only two stages in its life cycle. The trophozoite is pleomorphic, large and measures from 12 to 60 mm. The cysts are more or less round and measure 12– 30 mm. Both the trophozoites and cysts are uninucleate and contain a large, centrally located dense nucleolus. Occasionally, the nucleus of trophozoites, especially in infected tissue sections, may have two or three nucleoli. Cysts appear to be double walled with a wavy outer wall and a round inner wall, just like those of Acanthamoeba, when viewed under an optical microscope. Ultrastructurally, however, they are tripartite with an outer thin and irregular ectocyst, an inner thick endocyst, and a middle amorphous fibrillar mesocyst [1,3]. Treatment for Balamuthia infections Of the more than 100 cases of balamuthiasis that have been reported, only seven patients have survived [1,3,5,32–35,36,37,38]. Of these seven, four patients were treated with a combination of pharmaceuticals including pentamidine isethionate, sulfadiazine, azithro- mycin/clarithromycin, flucytosine, and fluconazole. This regimen was based on the efficacy of some of these drugs after they were tested in vitro [3]. Pentamidine had to be discontinued because of toxicity [34,35,36,37]. Some of the patients also received thioridazide or trifluoperazine for a short time. A macrolide antibiotic like azithromycin was initially started but was switched to clarithromycin because of poor penetration of the former into the cerebrospinal fluid (CSF) [34]. Currently, four more patients, three in the USA and one in Australia, have been treated successfully. Two of the US patients were treated with miltefosine, which has been shown to have some activity in vitro against Balamuthia [25]. The regimen included fluconazole 400 mg intravenously (i.v.) twice daily (b.i.d.), pentamidine 380 mg i.v. daily (q.d.), sulfadiazine 150 mg four times daily (q.i.d.), flucytosine 2000 mg orally (p.o.) q.d., and miltefosine 150 mg p.o. q.d. Azithromycin that was given previously was discontinued because of toxicity (M. Volmer, K. Kokko, personal communication). An Australian patient with balamuthiasis also received i.v. pentamidine 300 mg q.d., oral azithromycin 600 mg q.d., oral itraconazole 200 mg b.i.d., oral sulfadiazine 1.5 mg q.i.d., oral fluconazole 1 g q.d., and corticosteroid weaned early. This patient underwent a complete excision of the brain abscess, which may be a significant factor according to the doctor who treated this patient (Doyle, personal communication). Thus, it appears that if balamuthiasis can be recognized early in its course, interventional therapy can be instituted that may result in medical cure. Naegleria fowleri N. fowleri is also known as an ameboflagellate because of the presence of a transitory, pear-shaped, nonfeeding, nondividing flagellate stage in the life cycle in addition to ameboid trophic and dormant cyst stages. The trophozoite is a slug-like ameba, feeds on Gram-negative bacteria and reproduces by binary fission. It exhibits rapid sinusoidal movement by producing anterior hemispherical bulges or lobopodia. The posterior end or uroid is sticky and often has several trailing filaments to which bacteria may adhere prior to being ingested. The trophozoite measures 10–25 mm and is characterized by a single nucleus with a prominent, centrally placed nucleolus that stains densely with chromatic dyes. The cytoplasm contains numerous mitochondria, ribosomes, food vacuoles, and a contractile vacuole. The trophozoite transforms into a flagellate stage when changes occur in its environmental niche such as during a rainstorm that might perturb the ionic concentration of the surrounding environment. The flagellate usually has two flagella at the anterior end but three or four flagella may be seen occasionally. It ranges in length from 10 to 16 mm and possesses a single nucleus with a large nucleolus. The flagellate lacks a cytostome and hence cannot feed Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited. Amebic meningoencephalitides and keratitis Visvesvara 593 and reverts into the ameboid form. The trophozoite transforms into a resistant cyst during adverse conditions such as scarcity of food supply or its habitat becomes dry. The cyst is usually spherical, has a single nucleus with a large nucleolus and two walls with a thick endocyst and a closely apposed thinner ectocyst. The wall has pores flush with its surface that may not be readily seen [1,4]. N. fowleri has been isolated from fresh water, thermal discharges of power plants, heated swimming pools, hot springs, hydrotherapy and remedial pools, aquaria, sewage, and even from nasal passages and throats of healthy individuals. N. fowleri is thermophilic and can tolerate temperatures of up to 458C; hence, they proliferate during summer months when the ambient temperature is high. Therefore, cases of PAM occur in the hot summer months when large numbers of young people frequent lakes, ponds, inadequately chlorinated swimming pools, and other warm fresh water bodies for recreational activities [1,4,39]. Amebae in the water enter the nasal passages, penetrate the nasal epithelium, and migrate along the olfactory nerves to the brain. PAM symptoms include headache, neck stiffness, and abrupt fever after exposure to fresh water. Because the disease has a short incubation period, is acute and fulminating, early diagnosis and initiation of antimicrobial therapy are critical for the survival of the patient [1–3,40]. Treatment for primary amebic meningoencephalitis The drug of choice in treating PAM is the antifungal antibiotic amphotericin B (AMB). In-vitro testing indicates that N. fowleri is highly sensitive to the drug, with a minimal amebicidal concentration of 0.026– 0.078 mg/ml [4]. However, only a few patients have survived this infection probably because of delayed diagnosis and delay in the initiation of treatment. In one well documented case, a patient who recovered from PAM was treated with high doses of AMB simultaneously both i.v. and intrathecally (i.t.): AMB (i.v. at 1.5 and i.t. at 1.5 mg/kg of body weight/day), and i.v. and i.t. miconazole (i.v. at 350 and i.t. at10 mg/m2 of body surface area/ day), and oral rifampin (10 mg/kg of body weight/day) [40]. N. fowleri was isolated from the CSF of the patient and in-vitro testing of the isolate indicated a synergistic or additive effect of AMB and miconazole. Rifampin was ineffective [40]. Resistance to AMB has not been reported [4]. A drawback in the use of the AMB is the risk of kidney impairment [4,40]. Antifungal drugs in addition to AMB have been tested against Naegleria isolates in vitro and in mice infected with the amebae. Schuster et al. [25] examined the in-vitro activity of miltefosine and voriconazole and found them to be active against N. fowleri at 40 mg/ml, a dose achievable in the CSF. Recently, Kim et al. [41] examined the in-vitro and in-vivo activity of AMB, miltefosine, and chlorpromazine, and found chlorpromazine to be the most active of the three compounds tested in vivo: 75% of mice infected with N. fowleri survived PAM, whereas only 55 and 40% of mice treated with miltefosine and amphotericin B, respectively, survived, indicating that chlorpromazine may be an effective drug for use in cases of PAM. Conclusion Although progress has been made in the treatment of amebic encephalitides, these diseases continue to have a high mortality. This is partly due to difficulty in diagnosis, lack of familiarity of the caregivers with the agents, initiation of therapy late in the course of infection, and lack of good therapeutic agents. Acanthamoeba keratitis is more readily recognized and the infection responds favorably to cationic disinfectants. Fast-acting and efficient drugs and effective delivery systems to reach the CNS are urgently needed. Acknowledgements The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Department of Health and Human Services or the Centers for Disease Control and Prevention. References and recommended reading Papers of particular interest, published within the annual period of review, have been highlighted as: of special interest of outstanding interest Additional references related to this topic can also be found in the Current World Literature section in this issue (p. 635). 1 Visvesvara GS, Moura H, Schuster FL. Pathogenic and opportunistic freeliving amoebae: Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri, and Sappinia diploidea. FEMS Immunol Microbiol 2007; 50:1–26. 2 Khan NA. Acanthamoeba spp. In: Khan NA, editor. Emerging protozoan pathogens. New York, NY: Taylor and Francis; 2008. pp. 3–69. 3 Schuster FL, Visvesvara GS. Balamuthia mandrillaris. In: Khan NA, editor. Emerging protozoan pathogens. New York, NY: Taylor and Francis; 2008. pp. 71–118. 4 Marciano-Cabral F, Cabral G. Naegleria fowleri. In: Khan NA, editor. Emerging protozoan pathogens. New York, NY: Taylor and Francis; 2008 . pp. 119–152. 5 Schuster FL, Visvesvara GS. Free-living amoebae as opportunistic and nonopportunistic pathogens of humans and animals. Int J Parasitol 2004; 34:1001–1027. 6 Niyyati M, Lorenzo-Morales J, Rezaeian M, et al. Isolation of Balamuthia mandrillaris from urban dust, free of known infectious involvement. Parasitol Res 2009; 106:279–281. Verani JR, Lorick LA, Yoder JS, et al. National outbreak of Acanthamoeba keratitis associated with use of a contact lens solution, United States. Emerg Infect Dis 2009; 15:1236–1242. A detailed descriptive epidemiologic investigation that highlighted the use of a particular multipurpose contact lens solution for the outbreak of Acanthamoeba keratitis in the United States in 2007 resulting in the recall of that particular contact lens solution. 7 8 Panjwani N. Pathogenesis of Acanthamoeba keratitis. Ocul Surf 2010; 8:70– 79. A nice review highlighting the molecular mechanisms involved in the pathogenesis of Acanthamoeba keratitis. Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited. 594 Antimicrobial agents 9 Dart JKG, Saw VPJ, Kilvington S. Acanthamoeba keratitis: diagnosis and treatment update 2009. Am J Ophthalmol 2009; 148:487–499. This is an excellent review of the available information on Acanthamoeba keratitis including drug therapy. 10 Qvarnstrom Y, Da Silva AJ, Schuster FL, et al. Molecular confirmation of Sappinia pedata as causative agent of amebic encephalitis. J Infect Dis 2009; 199:1139–1142. 11 Sriram R, Shoff M, Booton G, et al. Survival of Acanthamoeba cysts after dessiccation for more than 20 years. J Clin Microbiol 2008; 46:4045– 4048. 12 Dudley R, Jarroll EL, Khan NA. Carbohydrate analysis of Acanthamoeba castellanii. Exp Parasitol 2010; 122:338–343. 13 Corsaro D, Venditti D. Phylogenetic evidence for a new genotype of Acanthamoeba (Amoebozoa, Acanthamoebida). Parasitol Res 2010; 107:233– 238. 14 Marciano-Cabral F, Cabral G. Acanthamoeba spp. as agents of disease in humans. Clin Microbiol Rev 2003; 16:273–307. 15 Martinez MS, Gonzalez-Mediero G, Santiago P, et al. Granulomatous amebic encephalitis in a patient with AIDS. Isolation of Acanthamoeba sp. group II from brain tissue and successful treatment with sulfadiazine and fluconazole. J Clin Microbiol 2000; 38:3892–3895. 16 Saxena A, Mittal S, Burman P, Garg P. Acanthamoeba meningitis with successful outcome. Indian J Pediatr 2009; 76:1063–1064. 17 Petry F, Torzewski M, Bohl J, et al. Early diagnosis of Acanthamoeba infection during routine cytological examination of cerebrospinal fluid. J Clin Microbiol 2006; 44:1903–1904. 18 Walia R, Montoya JG, Visvesvara GS, et al. A case of successful treatment of Acanthamoeba infection in a lung transplant recipient. Transplant Infect Dis 2007; 9:51–54. 19 Fung KT-T, Dhillon AP, McLaughlin J, et al. Cure of Acanthamoeba cerebral abscess in a liver transplant patient. Liver Transpl 2008; 14:308– 312. 20 Lackner P, et al. Acute granulomatous Acanthamoeba encephalitis in an immunocompetent patient. Neurocrit Care 2010; 12:91–94. 21 Aichelburg AC, Walochnik J, Assadian O, et al. Successful treatment of disseminated Acanthamoeba sp. infection with miltefosine. Emerg Infect Dis 2008; 14:1743–1746. 22 Slater CA, Sickel J, Visvesvara GS, et al. Brief report: successful treatment of disseminated Acanthamoeba infection in an immunocompromised patient. N Engl J Med 1994; 331:85–87. 23 Roberts CW, Henriquez FL. Drug target identification, validation, characterization and exploitation for treatment of Acanthamoeba (species) infection. Exp Parasitol 2010; 126:91–96. 27 Walochnik J, Obwaller A, Gruber F, et al. Anti-Acanthamoeba efficacy and toxicity of miltefosine in an organotypic skin equivalent. J Antimicrob Chemother 2009; 64:539–545. A nice experimental study delineating the activity of miltefosine in destroying Acanthamoeba that had penetrated an organ culture of skin as well as demonstrating the stoarge stability of miltefosine over 1 year at 48C. 28 McBride J, Mullen AB, Carter C, Roberts CW. Differential cytotoxicity of phospholipid analogues to pathogenic Acanthamoeba species and mammalian cells. J Antimicrob Chemother 2007; 60:521–525. 29 Rassu G, Gavini E, Mattana A, Giunchedi P. Improvement of antiamoebic activity of rokitamycin loaded in chitosan microspheres. Open Drug Deliv J 2008; 2:38–43. 30 Rassu G, Gavini E, Jonassen H, et al. New chitosan derivatives for the preparation of rokitamycin loaded microspheres designed for ocular or nasal administration. J Pharm Sci 2009; 98:4852–4865. Further work by these authors indicates that chitosan derivatives have better properties including smaller size, microadhesiveness, and in-vitro release of drugs thus making them more suitable for ocular and nasal administration of drugs to treat infections of the eye and brain. 31 Bang S, Edell E, Eghrari AO, Gottsch JD. Treatment with voriconazole in 3 eyes with resistant Acanthamoeba keratitis. Am J Ophthalmol 2010; 149:66–69. 32 Bakardjiev A, Azimi PH, Ashouri N, et al. Amebic encephalitis caused by Balamuthia mandrillaris: report of four cases. Pediatr Infect Dis 2003; 22:447–452. 33 Galarza M, Cuccia V, Sosa FP, Monges JA. Pediatric granulomatous cerebral amebiasis: a delayed diagnosis. Pediatr Neurol 2002; 26:153–156. 34 Deetz TR, Sawyer TH, Billman G, et al. Successful treatment of Balamuthia amoebic encephalitis. Clin Infect Dis 2003; 37:1304–1312. 35 Jung S, Schelper RL, Visvesvara GS, Chang HT. Balamuthia mandrillaris meningoencephalitis in an immunocompetent patient. Arch Pathol Lab Med 2004; 128:466–468. 36 Schuster FL, Yagi S, Gavali S, et al. Under the radar: Balamuthia amebic encephalitis. Clin Infect Dis 2009; 48:879–887. An important study, dealing with more than 3500 cases of encephalitis during a span of 9 years, exhorts the clinicians to be aware of the signs and symptoms of the balamuthias including neuroimaging findings, CSF composition, and the ethnicity of the patients. 37 Cary LC, Maul E, Potter C, et al. Balamuthia mandrillaris meningoencephalitis; survival of a pediatric patient. Pediatrics 2010; 125:e699–e703. 38 Bravo FG, Cabrera J, Gottuzo E, Visvesvara GS. Cutaneous manifestations of infection by free-living amebas. In: Tropical dermatology, Tyring SK, Lupin O, Henggae UR, editors. Philadelphia: Elsevier; 2006. pp. 49–55. 24 Walochnik J, Duchene M, Seifert K, et al. Cytotoxic activities of alkylphosphocholines against clinical isolates of Acanthamoeba spp. Antimicrob Agents Chemother 2002; 46:695–701. 39 Yoder JS, Eddy BA, Visvesvara GS, et al. The epidemiology of primary amoebic meningoencephalitis, 1962–2008. Epidemiol Infect 2010; 138: 968–975. Emphasizes the development of science-based risk-reduction messages and strategies for the prevention of primary amebic meningoencephalitis. 25 Schuster FL, Guglielmo J, Visvesvara GS. In vitro activity of miltefosine and voriconazole on clinical isolates of free-living amebas: Balamuthia mandrillaris, Acanthamoeba spp., and Naegleria fowleri. J Eukaryot Microbiol 2006; 53:121–126. 40 Seidel JS, Harmatz P, Visvesvara GS, et al. Successful treatment of primary amebic meningoencephalitis. N Engl J Med 1982; 306:346–348. 26 Seal DV. Acanthamoeba keratitis update: incidence, molecular epidemiology and new drugs for treatment. Eye 2003; 17:893–905. 41 Kim J-H, Jung S-Y, Lee Y-J, et al. Effect of therapeutic chemical agents in vitro and on experimental meningencephalitis due to Naegleria fowleri. Antimicrob Agents Chemother 2008; 52:4010–4016. Copyright © Lippincott Williams & Wilkins. Unauthorized reproduction of this article is prohibited.