Acanthamoeba castellanii
Pathogen Safety Data Sheets: Infectious Substances – Acanthamoeba castellanii
SECTION I – INFECTIOUS AGENT
Name: Acanthamoeba castellanii
Agent type: Parasite
Family: Acanthamoebidae
Genus: Acanthamoeba
Species: castellanii
Synonym or cross-reference
Also known as Acanthamoeba castellani and Acanthamoeba sp. ATCC 30011 1. Acanthamoeba castellanii is a causative agent of Acanthamoeba keratitis and granulomatous amoebic encephalitis.
Characteristics
A. castellanii is a free-living amoeba 2. The reference genome sequence for castellanii, NEFF-v1, is based on the Neff strain and contains a linear double-stranded DNA genome of approximately 42 Mbp with a G+C content of 58% 2. Morphological characteristics vary depending on environmental conditions 3 4. A. castellanii can adopt an oval-shaped or elongated infective trophozoite form, measuring approximately 15 to 50 μm, or a dormant, stress-resistant, double-walled cyst form measuring 10 to 25 μm 3 4.
Properties
Acanthamoeba species are ubiquitous in the environment and have been isolated from numerous aqueous-associated sources (e.g., public water supplies, swimming pools, bottled water, surface water samples), air-conditioning units, sewage, compost, sediments, soil, vegetables, air, and contact lenses and their cases 3 5. They have also been recovered from hospitals, dialysis units, and eye wash stations 5. Its biphasic life cycle allows for this prevalent distribution in the environment 3 5.
The vegetative trophozoite form exists when environmental conditions are favourable for growth; in this form, A. castellanii moves slowly using acanthopodia (spine-like hyaline protrusions) and feeds on other microbes, including bacteria, algae, yeasts, or small organic particles 3 5. Under adverse environmental conditions, such as extreme temperatures, pH, osmolarity, nutrient depletion, and desiccation, A. castellanii can differentiate into a metabolically inactive cyst. Its double-walled structure enables A. castellanii to survive under hostile conditions for prolonged periods 3. Cysts are airborne, which may help spread the species into the environment. A. castellanii reproduces asexually by binary fission 3 5.
Some bacteria and viruses ingested by trophozoites are able to evade digestion. For example, Mollivirus 6, adenovirus 7, Lausannevirus 8, murine norovirus 9, and Medusavirus 10 can replicate within trophozoites. Bacteria including Legionella pneumophila 11 12, Protochlamydia amoebophila 13, Porphromonas gingivalis, Prevotella intermedia 14, Bacillus anthracis 15, Vibrio cholerae 16, and Mycobacterium spp. 17 18 are able to survive and grow within A. castellanii trophozoites. Depending on conditions and pathogenicity of the microorganisms and the amoebae, the interactions vary leading to lysis of the microorganisms or amoebae, intake of microorganisms without any changes, incubation and growth of the microorganisms within the amoebae, or increase in virulence of the microorganisms 19. Thus, both trophozoites and cysts may serve as reservoirs for a number of pathogenic microorganisms, leading to co-infection 19.
Section II – Hazard identification
Pathogenicity and toxicity
Several Acanthamoeba species, including A. castellanii, have been reported to cause Acanthamoeba keratitis (AK) 5 20. Onset of symptoms may range from a few days to several weeks, depending on the inoculum size and/or the extent of corneal trauma, and clinical signs include foreign body sensation in the eye, redness, itching, pain, photophobia, epiphora, edema, and blurred vision or loss of vision 5 21 22. A ring-like stromal infiltrate appears in approximately 50% of patients in the advanced stage of Acanthamoeba keratitis 3. Usually only one eye is affected 23 24. Acanthamoeba sclerokeratitis (ASK) is an uncommon complication of AK and presumably has an immune-mediated origin 3. ASK is estimated to occur in 14–16% of AK cases and is characterized by severely reduced vision and severe pain, which may lead to necrotizing scleritis and corneal perforation 25. The prognosis of ASK is generally poor and occasionally requires enucleation to alleviate severe ocular pain 25. A. castellanii cysts may resist drug treatment and persist within the host for prolonged periods. Recurrent infection occurs in approximately 10% of cases 26, and has been reported up to 5 years after initial infection 27. Early diagnosis and treatment are necessary to prevent permanent ocular damage and vision loss 5 28.
A. castellanii can also cause granulomatous amoebic encephalitis (GAE), particularly in immunocompromised or otherwise debilitated individuals 4 5. Onset of GAE is slow and follows a chronic course characterized by several weeks to months of headache, low-grade fever, stiff neck, mental state abnormalities, nausea, vomiting, lethargy, visual disturbances, and focal neurologic deficits, depending on the topographic site of lesions, followed by loss of consciousness, hemiparesis, seizures, and coma in later stages 5 29. Although rare, GAE has been reported in otherwise healthy individuals. For example, an otherwise healthy Taiwanese farmer presented with general weakness and difficulty of defecation and urination two weeks after aspiration of muddy water, and subsequently developed nausea, vomiting, severe headache, and gradual altered consciousness and emotional agitation during hospitalization 30. Despite antibiotic treatment, he developed hypotension, bradycardia, and myoclonus. The patient recovered following antimicrobial, corticosteroid, and diuretic therapy, although slow response to time, place, and person orientation was noted. The mortality rate of Acanthamoeba central nervous system (CNS) infection may exceed 90% 31.
Cutaneous manifestations of A. castellanii are characterized by hard erythematous nodules and/or skin ulcerations, including the presence of firm papulonodules with purulent drainage which develop into non-healing indurated ulcerations, and demonstrate Acanthamoeba trophozoites and cysts 5 20. In otherwise healthy individuals, these infections are very rare and are typically self-limiting. However, in immunocompromised individuals, disseminated infection may follow, leading to fatal outcomes 5. The reported mortality rate for Acanthamoeba cutaneous infection without CNS involvement is approximately 73% in AIDS patients, while that for cutaneous infection with concomitant CNS infection is 100% 20.
A rare cause of osteomyelitis of a bone graft of the mandible associated with A. castellanii infection was reported in a 32-year-old prediabetic woman 32. A. castellanii sinusitis was reported in an HIV-infected patient and was characterized by sinus congestion, epistaxis, nasal crusting, and frontal headache 33. Acanthamoeba spp. also appear to be a vector in transmission of bacterial pathogens to susceptible hosts; approximately 24% of Acanthamoeba spp. isolates from clinical and environmental sources appear to carry intracellular bacteria 34.
A case of acute, hemorrhagic, necrotizing meningoencephalitis was reported in a dog; A. castellanii was isolated from the canine’s lung and kidneys, which were presumed to be the primary sites of lesions 35. A. castellanii has also been isolated from cornea swab samples from cats with keratitis 36. Disseminated disease associated with Acanthamoeba spp. infection has been reported in dogs 37 38 39 and horses 40.
Epidemiology
Acanthamoeba spp. occur worldwide and are ubiquitous in the environment 3 5. A. castellanii has been isolated from the nasal mucosa of approximately 4% of healthy individuals 41, whereas A. castellanii genotype T4 antibody prevalence rates of over 85% were detected in 114 healthy individuals from 37 countries 42.
A. castellanii genotype T4 has been implicated in AK and GAE worldwide 5. The estimated annual incidence of AK is 0.1 to 14.9 cases per 100,000 contact lens wearers 3. An outbreak in the United States linked to use of a particular contact lens solution affected approximately 158 people in 2007 43. GAE incidence is rare. Less than 200 cases have been reported worldwide, making accurate diagnosis and treatment difficult 31 44.
Use of contact lenses and ocular trauma are risk factors for AK 23; over 93% of reported AK cases were associated with use of contact lenses 22 23 45. Improper usage, such as extended wear and poor hygiene during lens storage and handling, is usually a contributing factor 23.
Immunocompromised individuals are predisposed to GAE 4 5 46. GAE is generally associated with individuals with comorbidities, including malignancies, Hodgkin’s disease, systemic lupus erythematosus, diabetes, renal failure, cirrhosis, tuberculosis, skin ulcers, and HIV infection 20. Predisposing factors include alcoholism, drug abuse, steroid treatment, cancer chemotherapy, radiotherapy, organ transplantation, and excessive use of antibiotics 20 47.
Host range
Natural host(s): Humans 3 5, dogs 35 48, and cats 36 53.
Other host(s): Pigs 46, hamsters 49, mice, and rats 50 are notable experimentally infected hosts.
Infectious dose
Unknown. However, infectious doses (ID50) were experimentally determined in mice infected with A. castellanii strain HN-3 by intranasal exposure; an ID50 of 811 organisms was determined with brain invasion as an endpoint response, whereas the ID50 in an acute meningoencephalitis model was 2,483 organisms 51. A median lethal dose (LD50) of 5,276 amebae was also determined 56.
Incubation period
Varies from days in cases of Acanthamoeba keratitis to several weeks to months in cases of GAE, cutaneous lesions, and sinusitis 20.
Communicability
The preferred mode of transmission is contact of A. castellanii with mucous membranes or damaged skin 23 52 53 54. In immunocompromised individuals, A. castellanii can enter via skin lesions or inhalation of cysts and cause cutaneous or lower respiratory tract infections, respectively 23. Transmission by ingestion of contaminated water has been reported in otherwise healthy individuals 30, while transmission by direct contact with intact skin has resulted in cutaneous infection 55. Transmission by injection was reported in a peripheral stem cell transplant recipient 56.
Section III – Dissemination
Reservoir: None.
Zoonosis: None.
Vectors: None.
Section IV – Stability and viability
Drug susceptibility/Drug resistance
Diamidines (e.g., hexamidine 57, pentamidine 57 58, propamidine); biguanides (e.g., chlorhexidine 57, polyhexamethylene biguanide); azoles (e.g., clotrimazole 57, voriconazole 57, itraconazole 59); amphotericin B 57; neomycin 60; flucytosine and miltefosine 57 61 have been used in the treatment of Acanthamoeba infections and were effective against A. castellanii in vitro.
High resistance to polyhexamethylene biguanide was described in clinical isolates in Taiwan 62, whereas strains resistant to erythromycin, chloramphenicol, and oligomycin were reported in the United States 63. Treatment failure with rifampin and ketoconazole was reported in an HIV-infected patient with A. castellanii infection 33. Generally, Acanthamoeba cysts are more resistant to drugs than trophozoites 20.
Susceptibility to disinfectants
Treatment with povidone-iodine resulted in a reduction of viable trophozoites and, to a lesser degree, cysts 64 65. Formalin (10%) 66 treatment and sodium hypochlorite (2.5%) treatment for 15 minutes showed cysticidal effects 67. Ethanol (60-70%) 67 68 and isopropanol (20%) 66 68 are effective against trophozoites and moderately effective against cysts. Treatment with peracetic acid (0.2%) at 55°C for 10 minutes, ortho-phthalaldehyde (0.55%) for 10 minutes, and hydrogen peroxide (7.5%) for 20 minutes were effective against Acanthamoeba cysts 67.
Physical inactivation
Acanthamoeba cysts can be inactivated by moist heat treatment at 65°C for 15 minutes 67 69. A. castellanii ATCC 50370 trophozoites may be completely inactivated by cold atmospheric gas plasma (CAP) for 2 minutes, whereas complete inactivation of A. castellanii cysts was reported after 4 minutes of exposure to CAP 70.
Survival outside host
A. castellanii is ubiquitous in the environment and has been isolated from soil, fresh and marine water, and air samples 4 5 71. A. castellanii cysts, and the viable microbes they may contain, can survive for prolonged periods 17 72 73. Acanthamoeba cysts can survive for several years 74 75.
Section V – First aid/medical
Surveillance
A. castellanii can be detected in patient samples using culture-based methods and PCR 3 52. Patient samples can be examined microscopically to detect trophozoites and/or cysts. Experimental diagnostic techniques including autofluorescence signatures of pathogens are under investigation 76.
Note: The specific recommendations for surveillance in the laboratory should come from the medical surveillance program, which is based on a local risk assessment of the pathogens and activities being undertaken, as well as an overarching risk assessment of the biosafety program as a whole. More information on medical surveillance is available in the Canadian Biosafety Handbook (CBH).
First aid/treatment
Treatment for AK usually involves topical application of a biguanide (e.g., polyhexamethylene biguanide, chlorhexidine gluconate) in combination with a diamidine (e.g., hexamidine) 26 77. Duration of treatment is usually about 6 weeks 26, but may last up to 26 months 78. Surgical intervention may be required. GAE has been treated successfully using various combinations of drugs including antibiotics such as pentamidine, cotrimoxazole, propamidine isethionate, azoles, amphotericin B, flucytosine, rifampin, azithromycin, amikacin, and anticancer drugs such as miltefosine, phenothiazines and thioridazine 26 46 79.
Note: The specific recommendations for first aid/treatment in the laboratory should come from the post-exposure response plan, which is developed as part of the medical surveillance program. More information on the post-exposure response plan can be found in the CBH.
Immunization
No vaccine currently available.
Note: More information on the medical surveillance program can be found in the CBH, and by consulting the Canadian Immunization Guide.
Prophylaxis
No known post-exposure prophylaxis.
Note: More information on prophylaxis as part of the medical surveillance program can be found in the CBH.
Section VI – Laboratory hazard
Laboratory-acquired infections
None reported to date.
Note: Please consult the Canadian Biosafety Standard (CBS) and CBH for additional details on requirements for reporting exposure incidents. A Canadian biosafety guideline describing notification and reporting procedures is also available.
Sources/specimens
Corneal scrapings 3, corneal biopsy specimens 3, cerebrospinal fluid 5, brain tissue 5, saliva 42, nasal and skin biopsy specimens 33.
Primary hazards
Exposure of mucous membranes/skin to infectious material 23 52, inhalation of airborne or aerosolized infectious material 23, and autoinoculation with infectious material 56 are the primary exposure hazards associated with A. castellanii.
Special hazards
A. castellanii is capable of harbouring a variety of microbes, including some that are pathogenic to humans and animals 6 7 8 9 10 11 12 13 14 15 16 17 18.
Section VII – Exposure controls/personal protection
Risk group classification
A. castellanii is a Risk Group 2 Human Pathogen and Risk Group 2 Animal Pathogen 80 81.
Containment requirements
Containment Level 2 facilities, equipment, and operational practices outlined in the CBS for work involving infectious or potentially infectious materials, animals, or cultures.
Protective clothing
The applicable Containment Level 2 requirements for personal protective equipment and clothing outlined in the CBS are to be followed. The personal protective equipment could include the use of a labcoat and dedicated footwear (e.g., boots, shoes) or additional protective footwear (e.g., boot or shoe covers) where floors may be contaminated (e.g., animal cubicles, PM rooms), gloves when direct skin contact with infected materials or animals is unavoidable, and eye protection where there is a known or potential risk of exposure to splashes.
Note: A local risk assessment will identify the appropriate hand, foot, head, body, eye/face, and respiratory protection, and the personal protective equipment requirements for the containment zone and work activities must be documented.
Other precautions
A biological safety cabinet (BSC) or other primary containment devices to be used for activities with open vessels, based on the risks associated with the inherent characteristics of the regulated material, the potential to produce infectious aerosols, the handling of high concentrations of regulated materials, or the handling of large volumes of regulated materials.
Use of needles and syringes to be strictly limited. Bending, shearing, re-capping, or removing needles from syringes to be avoided, and if necessary, performed only as specified in standard operating procedures (SOPs). Additional precautions are required with work involving animals or large scale activities.
Additional information: For diagnostic laboratories handling primary specimens that may contain Acanthamoeba castellanii, the following resources may be consulted:
Section VIII – Handling and storage
Spills
Allow aerosols to settle. Wearing personal protective equipment, gently cover the spill with absorbent paper towel and apply suitable disinfectant, starting at the perimeter and working towards the centre. Allow sufficient contact time before clean up (CBH).
Disposal
All materials/substances that have come in contact with the regulated materials to be completely decontaminated before they are removed from the containment zone or standard operating procedures (SOPs) to be in place to safely and securely move or transport waste out of the containment zone to a designated decontamination area / third party. This can be achieved by using decontamination technologies and processes that have been demonstrated to be effective against the regulated material, such as chemical disinfectants, autoclaving, irradiation, incineration, an effluent treatment system, or gaseous decontamination (CBH).
Storage
The applicable Containment Level 2 requirements for storage outlined in the CBS are to be followed. Primary containers of regulated materials removed from the containment zone to be labelled, leakproof, impact resistant, and kept either in locked storage equipment or within an area with limited access.
Section IX – Regulatory and other information
Canadian regulatory context
Controlled activities with A. castellanii require a Human Pathogens and Toxins licence issued by the Public Health Agency of Canada.
The following is a non-exhaustive list of applicable designations, regulations, or legislations:
- Human Pathogen and Toxins Act and Human Pathogens and Toxins Regulations
- Health of Animals Act and Health of Animals Regulations
Last file update: May 2023
Prepared by: Centre for Biosecurity, Public Health Agency of Canada.
Disclaimer
The scientific information, opinions, and recommendations contained in this Pathogen Safety Data Sheet have been developed based on or compiled from trusted sources available at the time of publication. Newly discovered hazards are frequent and this information may not be completely up to date. The Government of Canada accepts no responsibility for the accuracy, sufficiency, or reliability or for any loss or injury resulting from the use of the information.
Persons in Canada are responsible for complying with the relevant laws, including regulations, guidelines and standards applicable to the import, transport, and use of pathogens in Canada set by relevant regulatory authorities, including the Public Health Agency of Canada, Health Canada, Canadian Food Inspection Agency, Environment and Climate Change Canada, and Transport Canada. The risk classification and related regulatory requirements referenced in this Pathogen Safety Data Sheet, such as those found in the Canadian Biosafety Standard, may be incomplete and are specific to the Canadian context. Other jurisdictions will have their own requirements.
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