A patient with granulomatous amoebic encephalitis caused by Balamuthia mandrillaris survived with two excisions and medication

Background Granulomatous amoebic encephalitis (GAE) is a rare central nervous system infection caused by the Balamuthia mandrillaris or Acanthamoeba species. Diagnosis is challenging because of the non-specific clinical presentation, cerebrospinal fluid analysis, and radiological features. There is no effective treatment for GAE to date. Case presentation A 54-year-old male was admitted to hospital after experiencing acute onset of numbness and weakness on his left limb. Due to the initial consideration of intracranial tumor, surgical removal of the right parietal lesion was performed. However, the patient had a headache accompanied by diplopia, difficulty walking and a new lesion was found in the left occipital-parietal lobe two weeks after the first operation. High-throughput next-generation sequencing (NGS) detected the presence of high copy reads of the B. mandrillaris genome sequence in the patient’s blood, cerebral spinal fluid (CSF), and brain tissue. Pathological investigation of the brain tissue showed granulomatous changes and amoebic trophozoite scattered around blood vessels under high magnification. The patient was re-operated due to developing progressive confusion caused by subfalcine herniation of the left cerebral hemisphere. The lesions of the right parietal lobe were obviously decreasing in size after the first surgery, and the lesions of the left occipital lobe and the sunfalcine herniation didn’t ameliorate two months after the second surgery. The patient was transferred to local hospital for continuous treatment with sulfamethoxazole and azithromycin. After five months of the second surgery, the patient showed good recovery with mild headache. Conclusions This is the first report of a patient with B. mandrillaris encephalitis initially confirmed by NGS and have experienced two excisions, responding favorably to the combination of surgeries and medications. Early surgical resection of intracranial lesions combined with drug treatment may offer the chance of a cure.

Encephalomyelitis Caused by Balamuthia mandrillaris in a Woman With Breast Cancer: A Case Report and Review of the Literature

Balamuthia mandrillaris is one cause of a rare and severe brain infection called granulomatous amoebic encephalitis (GAE), which has a mortality rate of >90%. Diagnosis of Balamuthia GAE is difficult because symptoms are non-specific. Here, we report a case of Balamuthia amoebic encephalomyelitis (encephalitis and myelitis) in a woman with breast cancer. She sustained trauma near a garbage dump 2 years ago and subsequently developed a skin lesion with a Mycobacterium abscessus infection. She experienced dizziness, lethargy, nausea and vomiting, inability to walk, and deterioration of consciousness. Next-generation sequencing of cerebrospinal fluid (CSF) samples revealed B. mandrillaris, and MRI of both brain and spinal cord showed abnormal signals. T-cell receptor (TCR) sequencing of the CSF identified the Top1 TCR. A combination of amphotericin B, flucytosine, fluconazole, sulfamethoxazole, trimethoprim, clarithromycin, pentamidine, and miltefosine was administrated, but she deteriorated gradually and died on day 27 post-admission.

Various brain-eating amoebae: the protozoa, the pathogenesis, and the disease

Among various genera of free-living amoebae prevalent in nature, some members are identified as causative agents of human encephalitis, in which Naegleria fowleri followed by Acanthamoeba spp. and Balamuthia mandrillaris have been successively discovered. As the three dominant genera responsible for infections, Acanthamoeba and Balamuthia work as opportunistic pathogens of granulomatous amoebic encephalitis in immunocompetent and immunocompromised individuals, whereas Naegleria induces primary amoebic meningoencephalitis mostly in healthy children and young adults as a more violent and deadly disease. Due to the lack of typical symptoms and laboratory findings, all these amoebic encephalitic diseases are difficult to diagnose. Considering that subsequent therapies are also affected, all these brain infections cause significant mortality worldwide, with more than 90% of the cases being fatal. Along with global warming and population explosion, expanding areas of human and amoebae activity in some regions lead to increased contact, resulting in more serious infections and drawing increased public attention. In this review, we summarize the present information of these pathogenic free-living amoebae, including their phylogeny, classification, biology, and ecology. The mechanisms of pathogenesis, immunology, pathophysiology, clinical manifestations, epidemiology, diagnosis, and therapies are also discussed.

The transcriptome of Balamuthia mandrillaris trophozoites for structure-guided drug design

Balamuthia mandrillaris, a pathogenic free-living amoeba, causes cutaneous skin lesions as well as granulomatous amoebic encephalitis, a ‘brain-eating’ disease. As with the other known pathogenic free-living amoebas (Naegleria fowleri and Acanthamoeba species), drug discovery efforts to combat Balamuthia infections of the central nervous system are sparse; few targets have been validated or characterized at the molecular level, and little is known about the biochemical pathways necessary for parasite survival. Current treatments of encephalitis due to B. mandrillaris lack efficacy, leading to case fatality rates above 90%. Using our recently published methodology to discover potential drugs against pathogenic amoebas, we screened a collection of 85 compounds with known antiparasitic activity and identified 59 compounds that impacted the growth of Balamuthia trophozoites at concentrations below 220 µM. Since there is no fully annotated genome or proteome of B. mandrillaris, we sequenced and assembled its transcriptome from a high-throughput RNA-sequencing (RNA-Seq) experiment and located the coding sequences of the genes potentially targeted by the growth inhibitors from our compound screens. We determined the sequence of 17 of these target genes and obtained expression clones for 15 that we validated by direct sequencing. These will be used in the future in combination with the identified hits in structure guided drug discovery campaigns to develop new approaches for the treatment of Balamuthia infections.

The application of shotgun metagenomics to the diagnosis of granulomatous amoebic encephalitis due to Balamuthia mandrillaris: a case report

Background Granulomatous amoebic encephalitis (GAE) is an infrequent and fatal infectious disease worldwide. Antemortem diagnosis in this condition is very difficult because clinical manifestations and neuroimaging are nonspecific. Case presentation A 60-year-old Japanese woman was admitted with a chief complaint of left homonymous hemianopsia. Brain-MRI showed extensive necrotizing lesions enhanced by gadolinium, in the right frontal lobe, right occipital lobe, and left parietal lobe. Epithelioid granulomas of unknown etiology were found in the biopsied brain specimens. Shotgun metagenomic sequencing using a next-generation sequencer detected DNA fragments of Balamuthia mandrillaris in the tissue specimens. The diagnosis of granulomatous amoebic encephalitis was confirmed using an amoeba-specific polymerase chain reaction and immunostaining on the biopsied tissues. Conclusions Shotgun metagenomics is useful for the diagnosis of central nervous system infections such as GAE wherein the pathogens are difficult to identify.

Free Living Amoebic Infections: Review

Infections caused by Naegleria fowleri , Acanthamoeba spp., and Balamuthia mandrillaris result in a variety of clinical manifestations in humans. These amoebae are found in water and soil worldwide. Acanthamoeba spp. and B. mandrillaris cause granulomatous amoebic encephalitis, which usually presents as a mass, while N. fowleri causes primary amoebic meningoencephalitis. Acanthamoeba spp. can also cause keratitis, and both Acanthamoeba spp. and B. mandrillaris can cause lesions in skin and respiratory mucosa. These amoebae can be difficult to diagnose clinically as these infections are rare and, if not suspected, can be misdiagnosed with other more common diseases. Microscopy continues to be the key first step in diagnosis but the amoeba can be confused with macrophages or other infectious agents if an expert in infectious disease pathology or clinical microbiology is not consulted. Although molecular methods can be helpful in establishing the diagnosis, these are only available in referral centers. Treatment requires combination of antibiotics and antifungals and, even with prompt diagnosis and treatment, mortality for neurological disease is extremely high.

Cytochrome P450 monooxygenase of Acanthamoeba castellanii participates in resistance to polyhexamethylene biguanide treatment

Acanthamoeba spp. are free-living parasites that can cause severe infections such as granulomatous amoebic encephalitis (GAE) and amoebic keratitis (AK). Polyhexamethylene biguanide (PHMB) is a topical application for AK treatment. However, PHMB is not entirely effective against all Acanthamoeba strains or isolates. The mechanisms by which Acanthamoeba protects itself against extreme drug conditions without encystation are still unknown. According to a previous study, cytochrome P450 monooxygenase (CYP450MO) plays an important role in the oxidative biotransformation of numerous drugs related to metabolism. In this study, a CYP450MO fragment was inserted into the pGAPDH-EGFP vector and transfected into Acanthamoeba castellanii. We found that CYP450MO-overexpressing Acanthamoeba had higher survival rates than those of the control cells after PHMB treatment. Moreover, we also found that encystation-related genes such as cellulose synthase I (CSI), encystation-mediating serine proteinase (EMSP), and autophagy-related protein 8 (ATG8) expression levels were not significantly different between Acanthamoeba transfected by pGAPDH-EGFP or pGAPDH-EGFP-CYP450MO. We suggest that Acanthamoeba transfected by pGAPDH-EGFP-CYP450MO may not induce encystation-related genes to resist PHMB treatment. In conclusion, these findings indicate that CYP450MO may be an additional target when PHMB is used for treatment of amoebic keratitis.

Discovery of Amoebicidal Compounds by Combining Computational and Experimental Approaches

Pathogenic and opportunistic free-living amoebae such as Acanthamoeba spp. can cause keratitis (AK), which may ultimately lead to permanent visual impairment or blindness. Acanthamoeba can also cause a rare but usually fatal granulomatous amoebic encephalitis (GAE). Current therapeutic options for AK require a lengthy treatment with nonspecific drugs that often are associated with adverse effects. Recent developments in the field led us to target cAMP pathways, specifically phosphodiesterase. Guided by computational tools we targeted the Acanthamoeba phosphodiesterase, RegA. Computational studies led to the construction and validation of a homology model followed by a virtual screening protocol guided by induced-fit docking and chemical scaffold analysis using our MBC chemical library. Subsequently, 18 virtual screening hits were prioritized for further testing in vitro against A. castellanii, identifying amoebicidal hits containing piperidine and urea imidazole cores. Promising activities were confirmed in the resistant cyst form of the amoeba and in additional clinical Acanthamoeba strains, increasing their therapeutic potential. Mechanism of action studies revealed that these compounds produce apoptosis through ROS-mediated mitochondrial damage. These chemical families show promise for further optimization to produce effective anti-Acanthamoebal drugs.