Balamuthia mandrillaris

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Balamuthia mandrillaris observed under light, scanning and transmission electron microscope Image credit: Gonzalez-Robles, Science Direct.

Classification

Eukaryota; Amoebozoa; Discosea; longamoebia; Balamuthiidae

Species

NCBI: [1]

Balamuthia mandrillaris

Description and Significance

Balamuthia mandrillaris, a free-living amoeba, is the causative agent of the rare yet fatal neurological condition known as granulomatous amoebic encephalitis (GAE). Discovered in 1986 within the brain of a deceased mandrill at the San Diego Wild Animal Park. The pathogen was successfully isolated and studied for the first time in 1993 by Govinda Visvesvara, paying tribute to his mentor William Balamuth for his amoebae research. (Cope et al.,2019)

B. mandrillaris primarily dwells in soil and poses a great threat to human health. This amoeba can invade the human body through open wounds or inhalation, and it has been isolated from soil samples. Believed to be found across temperate regions globally, evidence supporting this includes the detection of antibodies to the protist in the bloodstream of healthy individuals (Schuster et al.,2003).A comprehensive understanding of the ecology and pathogenesis of B. mandrillaris is essential for effective prevention and treatment strategies against infections caused by this amoeba (Baig, 2015).

Genome Structure

The genome of Balamuthia mandrillaris is characterized by its complexity and unique features. B. mandrillaris possesses a relatively large genome with multiple chromosomes. The specific number of chromosomes is not mentioned but it may vary between different strains. The genome is circular, which is a common feature among amoebas. The draft genome highlights the importance of reference genome sequencing for this pathogen, emphasizing the ongoing efforts to assemble a comprehensive and accurate genomic sequence (Greninger et al., 2015).

Cell Structure, Metabolism and Life Cycle

The cell structure of B. mandrillaris is characterized by a distinctive amoeboid form with a central granular endoplasm and a clear ectoplasm, enabling motility and phagocytosis. B. mandrillaris exhibits a unique multilayered cell wall, a distinguishing feature among free-living amoebas, which likely contributes to its resilience and virulence in causing infections (Khan & Siddiqui 2015).

B. mandrillaris is a heterotrophic amoeba, relying on external food sources for energy. The organism gains energy through phagocytosis, actively ingesting bacteria and other particles. As part of its metabolic activity, B. mandrillaris produces molecules such as adenosine triphosphate (ATP), reflecting its capacity for cellular energy generation during the trophozoite stage (Feldman & Yohannan 2019).

The life cycle of Balamuthia mandrillaris involves a trophic amoeboid stage and a cystic stage, both of which are infectious. During the trophic stage it actively feeds on bacteria for metabolic purposes. The trophozoite is pleomorphic and uninucleated, but rarely can be binucleated. The crucial transition occurs when the amoeba transforms into a dormant cyst, enabling its survival in adverse conditions. During the cyst stage, B. mandrillaris produces a protective outer ectocyst layer, and the cysts serve as the main infectious form responsible for the transmission of the pathogen (Khan & Siddiqui 2015).

Ecology and Pathogenesis

B. mandrillaris has been associated with amebic encephalitis, highlighting its presence in soil and freshwater environments, specifically in moist and warm regions. While its symbiotic relationships and biogeochemical significance are not explicitly detailed, its contributions to the environment involve its role as a free-living organism in microbial communities (Schuster et al., 2003). Understanding the ecology of B. mandrillaris will help solve its interactions within the environment and impacts on human health, as one can contract it during an activity like swimming.

The pathogenicity of B. mandrillaris begins in granulomatous amoebic encephalitis (GAE), particularly affecting immunocompromised hosts (Bhosale & Parija, 2021). The organism's ability to cause disease in humans is marked by its neurotropic nature, leading to severe neurological symptoms. Virulence factors contributing to its pathogenesis remain areas of active research. (Baig, 2014). B. mandrillaris is bigger than the size of human leukocytes, rendering phagocytosis impossible. Instead, the immune system tries to confine the amoeba at the point of entry (open wound), by initiating a type IV hypersensitivity reaction (Baig, 2015) Upon infiltration, the amoeba may induce a skin lesion or migrate to the brain, causing GAE,(Cope et al., 2019) which is often fatal. This granulomatous characteristic is mainly seen in immunocompetent patients, while immunocompromised patients display "perivascular cuffing" (Baig, 2014). Balamuthia-induced GAE manifests with symptoms like focal paralysis, seizures, and brainstem manifestations (Bhosale & Parija, 2021).

Diagnostic Evaluation of Fatal Balamuthia mandrillaris Image credit: Hawkins, Primates.

References

Schoch, C. L., Ciufo, S., Domrachev, M., Hotton, C. L., Kannan, S., Khovanskaya, R., Leipe, D., Mcveigh, R., O'Neill, K., Robbertse, B., Sharma, S., Soussov, V., Sullivan, J. P., Sun, L., Turner, S., & Karsch-Mizrachi, I. (2020). NCBI Taxonomy: a comprehensive update on curation, resources and tools. Database : the journal of biological databases and curation, 2020, baaa062. https://doi.org/10.1093/database/baaa062

Baig A (2014, December). Granulomatous amoebic encephalitis: ghost response of an immunocompromised host? J Med Microbiol, 63(12), 1763–1766. https://doi.org/10.1099/jmm.0.081315-0 PMID: 25239626

Baig AM (2015, April). Pathogenesis of amoebic encephalitis: Are the amoebas being credited to an 'inside job' done by the host immune response? Acta Tropica, 148, 72–76. https://doi.org/10.1016/j.actatropica.2015.04.022

Bhosale NK, Parija SC. (2021). Balamuthia mandrillaris: An opportunistic, free-living ameba – An updated review. Tropical Parasitology, 11(2), 78–88. https://doi.org/10.4103/tp.tp_36_21 PMID: 34765527

Cope JR, Landa J, Nethercut H, Collier SA, Glaser C, Moser M, Puttagunta R, Yoder JS, Ali IK, Roy SL (2019) The Epidemiology and Clinical Features of Balamuthia mandrillaris Disease in the United States, 1974–2016. Clinical Infectious Diseases 68(11): 1815–1822. https://doi.org/10.1093/cid/ciy813

Greninger AL, Messacar K, Dunnebacke T, Naccache SN, Federman S, Bouquet J, Mirsky D, Nomura Y, Yagi S, Glaser C, Vollmer M, Press CA, Kleinschmidt-DeMasters BK, Dominguez SR, Chiu CY (2015) Clinical metagenomic identification of Balamuthia mandrillaris encephalitis and assembly of the draft genome: the continuing case for reference genome sequencing. Genome Medicine 7(1): 113. https://doi.org/10.1186/s13073-015-0235-2

Schuster FL, Dunnebacke TH, Booton GC, Yagi S, Kohlmeier CK, Glaser C, Vugia D, Bakardjiev A, Azimi P, Maddux-Gonzalez M, Martinez AJ, Visvesvara GS (2003) Environmental Isolation of Balamuthia mandrillaris Associated with a Case of Amebic Encephalitis. J. Clin. Microbiol. 41(7): 3175–3180. https://doi.org/10.1128/JCM.41.7.3175-3180.2003

Siddiqui R, Khan NA (2015). "Balamuthia mandrillaris: Morphology, biology, and virulence". Trop. Parasitol. 5(1): 15–22. https://doi.org/10.4103/2229-5070.149888

Yohannan B, Feldman M (2019). Fatal Balamuthia mandrillaris encephalitis. Case Reports in Infectious Diseases 2019: 1–5. https://doi.org/10.1155/2019/9315756

Author

Page authored by Bella Readling, student of Prof. Bradley Tolar at UNC Wilmington.

This student page has not been curated.

Classification

Balamuthia mandrillaris is a single-celled, free-living amoeba commonly encountered in the natural environment. This organism falls within the Eukaryotic domain and is categorized within the Amoebozoa clade. Furthermore, B. mandrillaris is situated within the Amoebozoa phylum and is a part of the Discosea class. Additionally, it is a member of the Longamoebia order and it belongs to the Balamuthiidae family (1).

Introduction

B. mandrillaris, is a free-living amoeba found in soil and freshwater which is known to cause a deadly encephalitic infection in humans (2). The lethality of B. mandrillaris infection is notoriously high, with a reported fatality rate of >98% and very few survivors among published case reports (3)(4). The specific factors relating to infection/disease in humans are not well understood, such as whether underlying disease or genetic predisposition plays a role in its pathogenic propensity in humans (2)(5). In part due to the infrequency of reported infections, several of B. mandrillaris mechanisms as a pathogen are still not fully understood, with one such example being its ability to bind to several cells simultaneously (4).

Genome Structure

The overall genome size of B. mandrillaris is 41.6 kb (2). In 1990, it was discovered that the circular mitochondrial genome of B. mandrillaris contained two RNA genes, 18 transfer RNA genes, and 38 coding sequences (2). Other strains of B. mandrillaris have mitochondrial genomes between 39.8 to 42.8 kb, containing 33 protein coding genes, and as many as 14 transfer RNA genes (2). The mechanism of B. mandrillaris pathogenesis has been identified: the pathogen infects its targets by aiming for the 18S rRNA and 16S rRNA (2).

Cell Structure

B. mandrillaris has an unique cell structure because it undergoes a dormant cyst phase and a vegetative trophozoite phase (7). The vegetative trophozoite phase typically contains a single nucleus, but can be binucleated at times and “contains numerous mitochondria, ribosomes, endoplasmic reticulum, and divide by asexual reproduction that occurs by binary fission,” (7). When B. mandrillaris is in the vegetative trophozoite phase, it can extend filament-like structures through the use of pseudopodia: in this state, it is known to be shaped irregularly with an uneven surface (7). In the dormant cyst phase, the B. mandrillaris cell structure differs from that of the vegetative trophozoite phase. This is seen by spherical cysts, also with one nucleus, that are smaller in size by approximately 20-30 µm (7). Additionally, the dormant cyst phase of B. mandrillaris includes an inner wall that is the endocyst, a fibrillar middle layer known as mesocyst, and an outer wall that is the ectocyst (7). These B. mandrillariscysts have been shown to resist denaturing conditions due to their strong and stiff three-walled structure (7).

Metabolic Processes

B. mandrillaris contains ergosterol that protects the cell while allowing it to undergo biosynthesis (7). B. mandrillaris reproduces asexually when in the vegetative trophozoite phase. However, when presented with suboptimal conditions, the cell transforms into the dormant cyst phase for protection and does not reproduce (7). There is evidence that the pathogen exhibits protease activity (8). The pathogen’s protease activity is used to degrade the extracellular matrix of the brain tissue of its host (8). Additionally, ‘’B. mandrillaris’’ refuses to consume Gram-negative bacteria and is unable to grow on an agar medium that is covered in bacteria (2).

Ecology

B. mandrillaris is a free-living amoeba in the natural environment according to two reported isolations (4). The first isolation is from soil of a potted plant (4). In 2003, a fatal case of amebic encephalitis, a nervous system infection caused by amoeba in freshwater lakes and rivers, was studied to reveal ‘’B. mandrillaris’’ as the culprit. B. mandrillaris was found in the autopsied brain, and soil samples of the child victim’s home areas revealed the presence of B. mandrillaris in a potted plant (9). The second reported isolation is of balamuthia amebic encephalitis (BAE) in dogs that swam in pond water (4). ‘’B. mandrillaris’’ was found in the autopsies of both the dogs (9). However, B. mandrillarisfailed to be isolated from the pond water samples (4). The lack of ecological knowledge on ‘’B. mandrillaris’’ may be due to its slow growing nature, or because it is less abundant in the environment (4). Food sources for B. mandrillaris is not currently known in great detail (4). Studies have shown that ‘’B. mandrillaris’’ does not consume bacteria for growth, but utilizes bacteria to remain in the trophozoite stage of growth (4). It is known that ‘’B. mandrillaris’’ growth does particularly well on mammalian cells, specifically human and monkey (4). Additionally, B. mandrillaris growth was observed on Acanthamoeba trophozoites, but not on Acanthamoeba cysts (4). This may be due to the cellulose contained by Acanthamoeba cysts that Acanthamoeba trophozoites lack (4).

Pathology

B. mandrillaris antibodies are found in healthy populations; however, this bacteria can produce infection in both immunocompromised and immunocompetent individuals (4). B. mandrillaris can enter the body through breaks in the skin and/or through the respiratory tract. B. mandrillaris then invades the intracellular space (4). It is not yet entirely clear how B. mandrillaris invades the central nervous system through the bloodstream. Research has found that the blood-brain barrier (BBB) separates blood from the central nervous system, protecting it (4). An essential aspect of infection due to B. mandrillaris is its ability to cross this layer, but the specific methodology of this crossing is not yet understood (4). Research supports one methodology that may be how B. mandrillaris crosses the BBB: interleukin-6 (IL-6) is a protein that increases the permeability of BBB by modulating adhesion molecule expression (4). It was found that B. mandrillaris mediated a release of IL-6, reducing the permeability of the BBB, and therefore allowing it to cross (4). In addition, one study confirmed that B. mandrillaris efficiently kills host cells by engaging specific host cell signaling pathways to engulf and penetrate (4). The spherical shape of B. mandrillaris trophozoites can bind and penetrate multiple mammalian cells at a time, though the specifics of this mechanism are not yet known (4).B. mandrillaris can also kill their host cell by lysis (4). Specific phospholipases have been found in B. mandrillaris which hydrolyze phospholipids, causing membrane dysfunction of the host cell (4).

B. mandrillaris can also degrade the extracellular matrix (ECM) of the basal lamina around blood vessels in the brain (4). Degradation of this ECM leads to destruction of the central nervous system (4). The ECM contains types of collagen that are difficult to degrade; however, B. mandrillaris has protease properties that are able to cleave the types of collagen in the ECM (4). Additional protease properties revealed elastase which has the ability to degrade elastin, also found in the ECM (4). In these ways, B. mandrillaris can invade the ECM, the BBB, and the central nervous system. Infection of B. mandrillaris leads to balamuthia amoebic encephalitis (BAE) (4). This is a chronic disease that can last from 3 months to up to 2 years and disproportionately infects the young and the elderly (4). This disease presents with side effects including headaches, nausea, seizures, and sinus infection (10). It almost always leads to a fatal outcome in as little as weeks, and diagnosis is often post-mortem (10). BAE has not only been seen in humans, but in other mammals as well (4).

Current Research

While contracting B. mandrillaris is rare, current research is focused on providing a clear diagnosis and effective treatment. One of the earliest symptoms of B. mandrillarisare skin lesions. Researchers compared skin lesions of various biopsies to provide physicians accurate characteristics to look for (11). Some characteristics being ill-defined tuberculoid granulomas in a reaction pattern paired with many giant cells (11). The description coupled with an laboratory analysis, using an immunofluorescence method (4) or PCR-based assay (12), would allow time for treatment before the breakdown of the central nervous system.The proposed treatment is currently broad spectrum microbials (13). Due to treatment’ limited success against Balamuthia amoebic encephalitis, researchers tested a range of antiparasitic drugs against B. mandrillaris. None of the compounds were able to inhibit its growth (14). In the process, single RNA-sequencing of B. mandrillaris was used to locate portions of the genome that could be targeted by drug screenings. This enables future studies to pair structure guided drug discoveries with the sequenced genomes to develop new treatments (14).

References

[1] Schoch, C. L., Ciufo, S., Domrachev, M., Hotton, C. L., Kannan, S., Khovanskaya, R., Leipe, D., Mcveigh, R., O'Neill, K., Robbertse, B., Sharma, S., Soussov, V., Sullivan, J. P., Sun, L., Turner, S., & Karsch-Mizrachi, I. (2020). NCBI Taxonomy: a comprehensive update on curation, resources and tools. Database : the journal of biological databases and curation, 2020, baaa062. https://doi.org/10.1093/database/baaa062

[2] Bhosale N. & Parija S. (2021). Balamuthia mandrillaris: An opportunistic, free living ameba - an updated review. Tropical Parasitology, 78-88. https://doi.org/10.4103/tp.tp_36_21

[3] Jennifer R Cope, Janet Landa, Hannah Nethercut, Sarah A Collier, Carol Glaser, Melanie Moser, Raghuveer Puttagunta, Jonathan S Yoder, Ibne K Ali, Sharon L Roy, The Epidemiology and Clinical Features of Balamuthia mandrillaris Disease in the United States, 1974–2016, Clinical Infectious Diseases, Volume 68, Issue 11, 1 June 2019, Pages 1815–1822, https://doi.org/10.1093/cid/ciy813

[4] Matin, A., Siddiqui, R., Jayasekara, S., & Khan, N. (2008). Increasing importance of Balamuthia mandrillaris. American Society for Microbiology Journals, 435-448.

[5] Kum, S. J., Lee, S. J. H. W., Jung, S. J. H. W. H. R., Choe, S. J. H. W. H. R. M., Kim, S. J. H. W. H. R. M. S. P. (2019). Amoebic Encephalitis Caused by Balamuthia mandrillaris. Journal of Pathology and Translational Medicine, 53(5), 327–331. DOI: https://doi.org/10.4132/jptm.2019.05.14

[6]Siddiqui, R., & Khan, N. A. (2015). Balamuthia mandrillaris: Morphology, biology, and virulence. Tropical parasitology, 5(1), 15–22.

[7] Matin, A., Stins, M., Kim, K., & Khan, N. (2006). Balamuthia mandrillaris exhibits metalloprotease activities. FEMS Immunology & Medical Microbiology, Volume 47, Issue 1. https://doi.org/10.1111/j.1574-695X.2006.00065.x

[8] Schuster, F.L., Dunnebacke, T.H., Booton, G.C., Yagi, S., Kohlmeier, C.K., Glaser, C., Vugia, D., Bakardjiev, A., Azimi, P., Maddux-Gonzalez, M., Martinez, A.J., Visvesvara, G.S. (2003) Environmental isolation of Balamuthia mandrillaris associated with a case of amebic encephalitis. J Clin Microbiol. 41(7):3175-80.

[9] Finnin, P.J.,Visvesvara, G.S., Campbell, B.E., Fry, D.R., Gasser, R.B. (2006) Multifocal Balamuthia mandrillaris infection in a dog in Australia. Parasitol Res;100(2):423-6.

[10] Visvesvara, G.S., Moura, H., Schuster, F.L. (2007). Pathogenic and opportunistic free-living amoebae: Acanthamoeba spp., Balamuthia mandrillaris, Naegleria fowleri, and Sappinia diploidea. FEMS Immunology & Medical Microbiology. Vol. 50 1-26.

[11] Alvarez, P., Torres-Cabala, C., Gotuzzo, E., & Bravo, F. (2022). Cutaneous balamuthiasis: A clinicopathological study. JAAD International, 6:51-58.

[12] Booton, G.C., Carmichael, J.R., Visvesvara, G.S., Byers, T.J., & Fuerst, P.A. (2003). Identification of Balamuthia mandrillaris by PCR Assay Using the Mitochondrial 16S rRNA Gene as a Target. Journal of Clinical Microbiology, 41(1), 453-455.

[13] Sakusic A., Chen B., McPhearson K., Badi M., Freeman WD., Huang JF., Siegel JL., Jentoft ME., Oring JM., Verdecia J., Meschia JF., (2023) Balamuthia mandrillaris Encephalitis Presenting as a Symptomatic Focal Hypodensity in an Immunocompromised Patient. Open Forum Infectious Diseases, 10-3.

[14] Phan, I.Q, Rice, C.A, Craig, J. (2021) The transcriptome of Balamuthia mandrillaris trophozoites for structure-guided drug design. Sci Rep 11, 21664.

Author

Edited by Bella Patel, Arshis Marfatia, Max Skop, Sari Levine, June Gao, students of Jennifer Bhatnagar for BI 311 General Microbiology, 2015, Boston University.