Indian Journal of Pathology and Microbiology

: 2017  |  Volume : 60  |  Issue : 4  |  Page : 601--603

Opportunistic free: Living amoeba now becoming a usual pathogen?

Dnyaneshwari Purushottam Ghadage1, Archana Chintaman Choure1, Archana Bhimrao Wankhade2, Arvind Vamanrao Bhore1,  
1 Department of Microbiology, Smt. Kashibai Navale Medical College and General Hospital, Pune, Maharashtra, India
2 Department of Microbiology, CM Medical College, Kanchandur, Chhattisgarh, India

Correspondence Address:
Archana Chintaman Choure
Department of Microbiology, Smt. Kashibai Navale Medical College and General Hospital, Pune - 411 041, Maharashtra


Acanthamoeba species cause granulomatous Acanthamoeba encephalitis in immunocompromised patients. We report a case of acute purulent meningoencephalitis with a focal neurological deficit caused by Acanthamoeba species in a 2 years immunocompetent child.

How to cite this article:
Ghadage DP, Choure AC, Wankhade AB, Bhore AV. Opportunistic free: Living amoeba now becoming a usual pathogen?.Indian J Pathol Microbiol 2017;60:601-603

How to cite this URL:
Ghadage DP, Choure AC, Wankhade AB, Bhore AV. Opportunistic free: Living amoeba now becoming a usual pathogen?. Indian J Pathol Microbiol [serial online] 2017 [cited 2021 Apr 18 ];60:601-603
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Granulomatous acanthamoeba encephalitis (GAE) is a life-threatening infection caused by the free-living amoebae. Acanthamoeba species, Balamuthia species and Sappinia pedata are most commonly associated with GAE.[1] GAE cases are commonly seen in immunocompromised patients and with high mortality.[2]

 Case Report

A 2-year-old child came to paediatric outpatient department (OPD) on October 3, 2015 with chief complaints of, inability to stand and walk for the past 2 days. On examination, the patient was afebrile, conscious, oriented but irritable. In central nervous system examination, right leg muscle power was reduced to Grade III, plantar reflex was absent on both the sides and sixth cranial nerve (Abducens) palsy was seen. Rest systemic examination was within normal limit.

In history, his mother told that he was having high-grade fever during the past week of December 2014 and full month of January 15. The patient was treated outside on OPD basis. He responded to medication and became afebrile. However after a gap of 1 month, the mother noticed that he was taking help of hands for standing and unable to walk.

There were no history of diabetes mellitus, asthma, steroid intake, prolonged intake of antibiotics, protein energy malnutrition, trauma, burn, and surgery. Both parents and child was nonreactive to HIV antibodies. On thorough general examination, congenital lumbar sinus was found.

We received cerebrospinal fluid (CSF) for microbiological work-up. On wet mount trophozoite forms were seen [Figure 1]. It was approximately 20–30 μm. On Gram stain structures larger than pus cells were seen [Figure 2]. These were gram-negative nucleated cells with irregular size. After addition of Lacto Phenol Cotton Blue stain we could appreciate nucleus and nucleolus [Figure 3]. With these finding we were suspecting some free-living amoebic infection, so to make internal structures clear Giemsa stain of CSF was performed [Figure 4]. It showed trophozoites with nucleus and nucleolus. Culture done on nonnutrient medium with lawn of Escherichia coli. Depending on morphology we were suspecting Acanthamoeba species, and our findings were confirmed by JIPMER. For species identification, they had requested CSF for polymerase chain reaction (PCR), but unfortunately, subsequent lumbar puncture was dry tap. Hence, because of unavailability of CSF, we could not do speciation.{Figure 1}{Figure 2}{Figure 3}{Figure 4}

Peripheral blood smear examination was normal. In CSF cytology total nucleated cells were 30,000/mm,[3] lymphocytes 5%, and polymorphonucleocytes 95%. Liver function tests and kidney function tests were within normal limits. MRI of the brain with contrast findings– mild communicating hydrocephalus was noted with subtle meningeal enhancement in the left parietal region. MRI of the spine revealed infective process involving lumbar portions of spinal cord showing dural enhancement likely meningitis. The patient started with injection vancomycin, ceftriaxone and dexamethasone.


Acanthamoeba is having several species including Acanthamoeba castellanii, Acanthamoeba astronyxis, Acanthamoeba culberstoni, Acanthamoeba polyphaga, Acanthamoeba hatcheffi, Acanthamoeba rhysodes, Acanthamoeba divionensis, Acanthamoeba lugdunensis and Acanthamoeba lenticulata. These are associated with human disease. Acanthamoeba have been classified into 15 genotypes (T1–T15) and genotype T4 is associated with GAE.[2]

Life cycle of Acanthamoeba: it has two morphological forms cyst and trophozoite. Trophozoites are infective forms, although both cysts and trophozoites gain entry into the body through various means. Entry can occur through the eye, nasal passages, or ulcerated/broken skin. When it enters the respiratory system, or through the skin, it can invade the central nervous system by hematogenous dissemination causing GAE or disseminated disease or skin lesions in persons with compromised immune system.[1],[3]

Diagnosis of GAE requires a high index of suspicion by clinician and laboratory testing by expert microbiologist. Radiological investigations such as CT-Scan and MRI are not conclusive. Findings on radiological investigations are confusing with TB meningitis, neurocysticercosis, disseminated encephalomyelitis, and viral encephalitis.[4]

Diagnostic modalities available are Microscopy: This is the most common method used for diagnosis of GAE. Cyst and trophozoite forms are demonstrated by microscope in unstained or stained form. CSF, skin/sinus/lung biopsy, and brain tissue biopsy are the best samples. Wet mount shows motile trophozoites approximately 15–40 μm with spiny projections. In CSF sample, only trophozoites are seen cysts are usually absent. In histopathological examination of biopsy material after staining both forms can be seen. Various stains used are Giemsa, H and E, Trichrome, Wright, Grocott's GMS, CFW, etc.[3] Using electron microscope and immunohistochemistry we can identify genera. Culture - Acanthamoeba species are identified by three methods. These are axenic culture, xenic culture, and tissue culture. They grow best at 25°C–30°C in culture media inoculated with E. coli and other aerobic bacteria. Serological - GAE can present from sub-acute to chronic, lasting weeks to months. Detection of antibodies is considered reliable method. ITF, ELISA and Flow cytometry methods are used for it. Molecular diagnosis - It is an emerging modality for diagnosis of GAE. Nuclear and mitochondrial 18S rDNA, RNAse P genes are targeted. PCR analysis of nuclear 18S rDNA is used for identification of genotypes in Acanthamoeba species.


No single drug is effective; hence multidrug combinations are used to treat the patients. Drugs targeting different receptors and proteins are combined for the success of treatment. The commonly used antibiotics are pentamidine, cotrimoxazole, propamidine, isethionate, azoles, amphotericin-B, flucytosine, rifampin, azithromycin, amikacin, etc.[3] Timely diagnosis and early treatment can save the life of the patient.[3]

First case series report of acute pyogenic meningitis due to Acanthamoeba was published during 1965. In this report, three cases were of children age group, and one was an adult. All cases were immunocompetent but had a fatal outcome.[5] According to published English language literature, from the year 2000–2014 only four cases were reported in paediatric patients.[6] Out of four children, one had mandibular tuberculosis and died because of Acanthamoeba encephalitis. One child had Acute Lymphoblastic Leukemia and survived. Rest two cases were immunocompetent and survived.[6]


Our case is an immunocompetent 2 years male having congenital lumbar sinus. This could be a possible site of entry for the parasite. In 1½ year follow-up, the patient is doing well, so with a good outcome.

We really need to assess it and observe for a longer duration to fix the nature of pathogen, route of infection and mortality associated with it. Since in our case, some questions are unanswered.... our patient is residing in a city there was no exposure to fresh water and age is only 2 years so what could be the cause of infection??

It has been observed with the time that cases in immunocompetent individuals are rising, and hence, there is a possibility that previously considered opportunistic pathogen is turning out to be regular pathogen....???.


All authors thank department of paediatric for sending sample and providing clinical details of patient. We also thank Dr. Rakesh Singh sir and team from JIPMER for confirming the pathogen and lastly the two year old patient and parents for cooperation.

Financial support and sponsorship


Conflicts of interest

There are no conflicts of interest.


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3Parija SC, Dinoop K, Venugopal H. Management of granulomatous amebic encephalitis: Laboratory diagnosis and treatment. Trop Parasitol 2015;5:23-8.
4Parija SC. Textbook of Medical Parasitology: Protozoology & Helminthology. 3rd ed. New Delhi: All India Publishers & Distributors; 2006. Chapter 3, Page no 49-61.
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