Parasitology Research

, Volume 114, Issue 12, pp 4431–4439 | Cite as

Assessment of blood–brain barrier penetration of miltefosine used to treat a fatal case of granulomatous amebic encephalitis possibly caused by an unusual Balamuthia mandrillaris strain

  • Sharon L. Roy
  • Jane T. Atkins
  • Rosemaria Gennuso
  • Danny Kofos
  • Rama R. Sriram
  • Thomas P. C. Dorlo
  • Teresa Hayes
  • Yvonne Qvarnstrom
  • Zuzana Kucerova
  • B. Joseph Guglielmo
  • Govinda S. Visvesvara
Original Paper


Balamuthia mandrillaris, a free-living ameba, causes rare but frequently fatal granulomatous amebic encephalitis (GAE). Few patients have survived after receiving experimental drug combinations, with or without brain lesion excisions. Some GAE survivors have been treated with a multi-drug regimen including miltefosine, an investigational anti-leishmanial agent with in vitro amebacidal activity. Miltefosine dosing for GAE has been based on leishmaniasis dosing because no data exist in humans concerning its pharmacologic distribution in the central nervous system. We describe results of limited cerebrospinal fluid (CSF) and serum drug level testing performed during clinical management of a child with fatal GAE who was treated with a multiple drug regimen including miltefosine. Brain biopsy specimens, CSF, and sera were tested for B. mandrillaris using multiple techniques, including culture, real-time polymerase chain reaction, immunohistochemical techniques, and serology. CSF and serum miltefosine levels were determined using a liquid chromatography method coupled to tandem mass spectrometry. The CSF miltefosine concentration on hospital admission day 12 was 0.4 μg/mL. The serum miltefosine concentration on day 37, about 80 h post-miltefosine treatment, was 15.3 μg/mL. These are the first results confirming some blood–brain barrier penetration by miltefosine in a human, although with low-level CSF accumulation. Further evaluation of brain parenchyma penetration is required to determine optimal miltefosine dosing for Balamuthia GAE, balanced with the drug’s toxicity profile. Additionally, the Balamuthia isolate was evaluated by real-time polymerase chain reaction (PCR), demonstrating genetic variability in 18S ribosomal RNA (18S rRNA) sequences and possibly signaling the first identification of multiple Balamuthia strains with varying pathogenicities.


Balamuthia Granulomatous Encephalitis Miltefosine 


Conflict of interest

The authors declare that they have no conflicts of interest.


The findings and conclusions in this report are those of the authors and do not necessarily represent the official position of the Centers for Disease Control and Prevention.


  1. Ahmad AF, Andrews PW, Kilvington S (2011) Development of a nested PCR for environmental detection of the pathogenic free-living amoeba Balamuthia mandrillaris. J Eukaryot Microbiol 58:269–271. doi: 10.1111/j.1550-7408.2011.00541.x CrossRefPubMedGoogle Scholar
  2. Booton GC, Carmichael JR, Visvesvara GS, Byers TJ, Fuerst P (2003a) Genotyping of Balamuthia mandrillaris based on nuclear 18S and mitochondrial 16S rRNA genes. Am J Trop Med Hyg 68:65–69PubMedGoogle Scholar
  3. Booton GC, Carmichael JR, Visvesvara GS, Byers TJ, Fuerst PA (2003b) Identification of Balamuthia mandrillaris by PCR assay using the mitochondrial 16S rRNA gene as a target. J Clin Microbiol 41:453–455PubMedCentralCrossRefPubMedGoogle Scholar
  4. Bravo FG, Alvarez PJ, Gotuzzo E (2011) Balamuthia mandrillaris infection of the skin and central nervous system: an emerging disease of concern to many specialties in medicine. Curr Opin Infect Dis 24:112–117. doi: 10.1097/QCO.0b013e3283428d1e CrossRefPubMedGoogle Scholar
  5. Cary LC, Maul E, Potter C, Wong P, Nelson PT, Given C 2nd, Robertson W Jr (2010) Balamuthia mandrillaris meningoencephalitis: survival of a pediatric patient. Pediatrics 125:e699–e703. doi: 10.1542/peds.2009-1797 CrossRefPubMedGoogle Scholar
  6. Centers for Disease Control and Prevention (2010) Balamuthia mandrillaris transmitted through organ transplantation—Mississippi, 2009. MMWR Morb Mortal Wkly Rep 59:1165–1170Google Scholar
  7. Centers for Disease Control and Prevention (2013) Investigational drug available directly from CDC for the treatment of infections with free-living amebae. MMWR Morb Mortal Wkly Rep 62:666Google Scholar
  8. Deetz TR, Sawyer MH, Billman G, Schuster FL, Visvesvara GS (2003) Successful treatment of Balamuthia amoebic encephalitis: presentation of two cases. Clin Infect Dis 37:1304–1312CrossRefPubMedGoogle Scholar
  9. Dorlo TPC, Hillebrand MJX, Rosing H, Eggelte TA, de Vries PJ, Beijnen JH (2008a) Development and validation of a quantitative assay for the measurement of miltefosine in human plasma by liquid chromatography-tandem mass spectrometry. J Chromatogr B Analyt Technol Biomed Life Sci 865:55–62. doi: 10.1016/j.jchromb.2008.02.005 CrossRefPubMedGoogle Scholar
  10. Dorlo TP, van Thiel PP, Huitema AD, Keizer RJ, de Vries HJ, Beijnen JH, de Vries PJ (2008b) Pharmacokinetics of miltefosine in Old World cutaneous leishmaniasis patients. Antimicrob Agents Chemother 52:2855–2860. doi: 10.1128/AAC.00014-08 PubMedCentralCrossRefPubMedGoogle Scholar
  11. Dorlo TP, Balasegaram M, Beijnen JH, de Vries PJ (2012a) Miltefosine: a review of its pharmacology and therapeutic efficacy in the treatment of leishmaniasis. J Antimicrob Chemother 67:2576–2597. doi: 10.1093/jac/dks275 CrossRefPubMedGoogle Scholar
  12. Dorlo TP, Huitema AD, Beijnen JH, de Vries PJ (2012b) Optimal dosing of miltefosine in children and adults with visceral leishmaniasis. Antimicrob Agents Chemother 56:3864–3872. doi: 10.1128/AAC.00292-12 PubMedCentralCrossRefPubMedGoogle Scholar
  13. Food and Drug Administration (2015) Impavido/Miltefosine—label and approval history. In: Drugs@FDA. Accessed 01 July 2015.
  14. Dunnebacke TH, Schuster FL, Yagi S, Booton GC (2004) Balamuthia mandrillaris from soil samples. Microbiology 150:2837–2842CrossRefPubMedGoogle Scholar
  15. Eibl H, Unger C (1990) Hexadecylphosphocholine: a new and selective antitumor drug. Cancer Treat Rev 17:233–242CrossRefPubMedGoogle Scholar
  16. Finnin PJ, Visvesvara GS, Campbell BE, Fry DR, Gasser RB (2007) Multifocal Balamuthia mandrillaris infection in a dog in Australia. Parasitol Res 100:423–426CrossRefPubMedGoogle Scholar
  17. Foreman O, Sykes J, Ball L, Yang N, De Cock H (2004) Disseminated infection with Balamuthia mandrillaris in a dog. Vet Pathol 41:506–510CrossRefPubMedGoogle Scholar
  18. Huang ZH, Ferrante A, Carter RF (1999) Serum antibodies to Balamuthia mandrillaris, a free-living amoeba recently demonstrated to cause granulomatous amoebic encephalitis. J Infect Dis 179:1305–1208CrossRefPubMedGoogle Scholar
  19. Jaruratanasirikul S, Hortiwakul R, Tantisarasart T, Phuenpathom N, Tussanasunthornwong S (1996) Distribution of azithromycin into brain tissue, cerebrospinal fluid, and aqueous humor of the eye. Antimicrob Agents Chemother 40:825–826PubMedCentralPubMedGoogle Scholar
  20. Jung S, Schelper RL, Visvesvara GS, Chang HT (2004) Balamuthia mandrillaris meningoencephalitis in an immunocompetent patient: an unusual clinical course and a favorable outcome. Arch Pathol Lab Med 128:466–468PubMedGoogle Scholar
  21. Kiderlen AF, Radam E, Tata PS (2009) Assessment of Balamuthia mandrillaris-specific serum antibody by flow cytometry. Parasitol Res 104:663–670. doi: 10.1007/s00436-008-1243-6 CrossRefPubMedGoogle Scholar
  22. Kucerova Z, Sriram R, Wilkins PP, Visvesvara GS (2011) Identification of antigenic targets for immunodetection of Balamuthia mandrillaris. Clin Vaccine Immunol 18:1297–1301. doi: 10.1128/CVI.05082-11 PubMedCentralCrossRefPubMedGoogle Scholar
  23. Marschner M, Kötting J, Eibl H, Unger C (1992) Distribution of hexadecylphosphocholine and octadecyl-methyl-glycero-3-phosphocholine in rat tissues during steady-state treatment. Cancer Chemother Pharmacol 31:18–22CrossRefPubMedGoogle Scholar
  24. Martínez DY, Seas C, Bravo C, Legua P, Ramos C, Cabello AM, Gotuzzo E (2010) Successful treatment of Balamuthia mandrillaris amoebic infection with extensive neurological and cutaneous involvement. Clin Infect Dis 51:e7–e11. doi: 10.1086/653609 CrossRefPubMedGoogle Scholar
  25. Nau R, Sörgel F, Eiffert H (2010) Penetration of drugs through the blood-cerebrospinal fluid/blood-brain barrier for treatment of central nervous system infections. Clin Microbiol Rev 23:858–883. doi: 10.1128/CMR.00007-10 PubMedCentralCrossRefPubMedGoogle Scholar
  26. Niyyati M, Lorenzo-Morales J, Rezaeian M, Martin-Navarro CM, Haghi AM, Maciver SK, Valladares B (2009) Isolation of Balamuthia mandrillaris from urban dust, free of known infectious involvement. Parasitol Res 106:279–281. doi: 10.1007/s00436-009-1592-9 CrossRefPubMedGoogle Scholar
  27. Petersen CA, Greenlee MHW (2011) Neurologic manifestations of Leishmania spp. infection. J Neuroparasitology 2: doi:  10.4303/jnp/N110401
  28. Qvarnstrom Y, Visvesvara GS, Sriram R, da Silva AJ (2006) Multiplex real-time PCR assay for simultaneous detection of Acanthamoeba spp., Balamuthia mandrillaris, and Naegleria fowleri. J Clin Microbiol 44:3589–3595PubMedCentralCrossRefPubMedGoogle Scholar
  29. Qvarnstrom Y, Nerad TA, Visvesvara GS (2013) Characterization of a new pathogenic Acanthamoeba species, A. byersi n. sp., isolated from a human with fatal amoebic encephalitis. J Eukaryot Microbiol 60:626–633. doi: 10.1111/jeu.12069 CrossRefPubMedGoogle Scholar
  30. Reed RP, Cooke-Yarborough CM, Jaquiery AL, Grimwood K, Kemp AS, Su JC, Forsyth JRL (1997) Fatal granulomatous amoebic encephalitis caused by Balamuthia mandrillaris. Med J Aust 167:82–84PubMedGoogle Scholar
  31. Schuster FL, Visvesvara GS (1996) Axenic growth and drug sensitivity studies of Balamuthia mandrillaris, an agent of amebic meningoencephalitis in humans and other animals. J Clin Microbiol 34:385–388PubMedCentralPubMedGoogle Scholar
  32. Schuster FL, Dunnebacke TH, Booton GC, Yagi S, Kohlmeier CK, Glaser C, Vugia D, Bakardjiev A, Azimi P, Maddux-Gonzalez M, Visvesvara GS (2003) Environmental isolation of Balamuthia mandrillaris associated with a case of amebic encephalitis. J Clin Microbiol 41:3175–3180PubMedCentralCrossRefPubMedGoogle Scholar
  33. Schuster FL, Glaser C, Honarmand S, Maguire JH, Visvesvara GS (2004) Balamuthia amebic encephalitis risk, Hispanic Americans. Emerg Infect Dis 10:1510–1512CrossRefPubMedGoogle Scholar
  34. Schuster FL, Guglielmo BJ, Visvesvara GS (2006a) In-vitro activity of miltefosine and voriconazole on clinical isolates of free-living amoebas: Balamuthia mandrillaris, Acanthamoeba spp., and Naegleria fowleri. J Eukaryot Microbiol 53:121–126CrossRefPubMedGoogle Scholar
  35. Schuster FL, Honarmand S, Visvesvara GS, Glaser CA (2006b) Detection of antibodies against free-living amoebae Balamuthia mandrillaris and Acanthamoeba species in a population of patients with encephalitis. Clin Infect Dis 42:1260–1265CrossRefPubMedGoogle Scholar
  36. Schuster FL, Yagi S, Gavali S, Michelson D, Raghavan R, Blomquist I, Glastonbury C, AW B e, Scharnhorst D, Reed SL, Kuriyama S, Visvesvara GS, Glaser CA (2009) Under the radar: Balamuthia amebic encephalitis. Clin Infect Dis 7:879–887. doi: 10.1086/597260 CrossRefGoogle Scholar
  37. U.S. Food and Drug Administration (2014) Impavido (Miltefosine). Drugs@FDA—FDA approved drug products. Accessed 27 March 2015
  38. Visvesvara GS, Martinez AJ, Schuster FL, Leitch GJ, Wallace SV, Sawyer TK, Anderson M (1990) Leptomyxid ameba, a new agent of amebic meningoencephalitis in humans and animals. J Clin Microbiol 28:2750–2756PubMedCentralPubMedGoogle Scholar
  39. Visvesvara GS, Schuster FL, Martinez AJ (1993) Balamuthia mandrillaris, N. G., N. Sp., agent of amebic meningoencephalitis in humans and animals. J Eukaryot Microbiol 40:504–514CrossRefPubMedGoogle Scholar
  40. Visvesvara GS, Roy S, Maguire JH (2011) Pathogenic and opportunistic free-living amebae: Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri, and Sappinia pedata. In: Guerrant RL, Walker DH, Weller PF (eds) Tropical infectious diseases—principles, pathogens, & practice, 3rd edn. Elsevier, Churchill Livingstone, Philadelphia, pp 707–713CrossRefGoogle Scholar
  41. Wadhone P, Maiti M, Agarwal R, Kamat V, Martin S, Saha B (2009) Miltefosine promotes IFN-gamma-dominated anti-leishmanial immune response. J Immunol 182:7146–7154. doi: 10.4049/jimmunol.0803859 CrossRefPubMedGoogle Scholar
  42. World Health Organization (2011) Miltefosine (Inclusion)—adults and children. 18th expert committee on the selection and use of essential medicines. World Health Organization. Accessed 27 March 2015

Copyright information

© Springer-Verlag Berlin Heidelberg (outside the USA) 2015

Authors and Affiliations

  • Sharon L. Roy
    • 1
  • Jane T. Atkins
    • 2
  • Rosemaria Gennuso
    • 2
  • Danny Kofos
    • 2
  • Rama R. Sriram
    • 1
  • Thomas P. C. Dorlo
    • 3
    • 4
  • Teresa Hayes
    • 5
  • Yvonne Qvarnstrom
    • 6
  • Zuzana Kucerova
    • 1
  • B. Joseph Guglielmo
    • 7
  • Govinda S. Visvesvara
    • 1
  1. 1.Division of Foodborne, Waterborne and Environmental Diseases, National Center for Emerging and Zoonotic Infectious DiseasesCenters for Disease Control and PreventionAtlantaUSA
  2. 2.Methodist Children’s HospitalSan AntonioUSA
  3. 3.Department of Pharmacy and PharmacologySlotervaart Hospital—The Netherlands Cancer InstituteAmsterdamThe Netherlands
  4. 4.Utrecht Institute for Pharmaceutical SciencesUtrecht UniversityUtrechtThe Netherlands
  5. 5.Department of PathologyMethodist HospitalSan AntonioUSA
  6. 6.Division of Parasitic Diseases and Malaria, Center for Global HealthCenters for Disease Control and PreventionAtlantaUSA
  7. 7.Department of Clinical Pharmacy, School of PharmacyUniversity of California, San FranciscoSan FranciscoUSA

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