Update on Acanthamoeba Keratitis: Diagnosis, Treatment, and Outcomes

Home , Acanthamoeba keratitis, Keratitis

REVIEW

Nicholas J. R. Maycock, BSc (Hons), MBBS, FRCOphth, and Rakesh Jayaswal, MBChB, FRCOphth, FRCS(Ed)

THE PATHOGEN Purposes: A literature review to describe the current diagnosis and Acanthamoeba spp. are ubiquitous, free-living proto- management of Acanthamoeba keratitis. zoans that have been isolated from a variety of habitats Results: Acanthamoeba is a ubiquitous protozoan: 8 species, 5 including air, soil, dust, water, and the nasopharyngeal 7,8 genotypic classes have been reported to cause keratitis. It is potentially mucosa of healthy individuals. They are known to thrive a sight-threatening infection, and there is often a poor prognosis in ponds, swimming pools, hot tubs, and CL solutions and because of a significant delay in diagnosis and frequently a lack of exist in either an active trophozoite or dormant cystic form effective medical management. Main risk factors are contact lens (Fig. 1). Over 20 unique species of Acanthamoeba have been wear, poor hygiene, and contact with contaminated water. Current identified with 8 reported to cause keratitis, the most common methods of diagnosis include corneal scrapings for histopathologic of which are Acanthamoeba castellani and Acanthamoeba 8,9 analysis, tissue culture, confocal microscopy, and polymerase chain polyphaga. There are 15 genotypic classes designated T1 to 10 reaction (PCR), each are reviewed in turn. Treatment options include T15 with T3, 4, 5, 6, and 11 documented as causing AK. medical (biguanides, diamidines, and corticosteroids) and surgical Acanthamoeba has 2 stages to its life cycle (Fig. 2), an (epithelial debridement, amniotic membrane transplant, and penetrat- active trophozoite stage that exhibits growth and a dormant ing keratoplasty). cystic stage with minimal metabolic activity. The trophozoite is capable of slow locomotion via pseudopodia.11,12 It feeds Conclusions: Early diagnosis and treatment are required to on yeast, small bacteria, other protozoans, and in the cornea, effectively manage this condition. keratocytes.13 They reproduce asexually via binary fission and m Key Words: Acanthamoeba, keratitis, review article, cornea are 25 to 50 m in size. The cyst is the dormant form and is 15 to 30 mm in size. (Cornea 2016;35:713–720) Encystment occurs under adverse conditions such as food deprivation, desiccation, and changes in temperature and pH. It allows the organism to survive extreme conditions, and cysts are canthamoeba keratitis (AK) is a potentially sight- resistant to biocides, chlorination, antibiotics, and low temper- Athreatening infection of the ocular surface that is atures.12 Excystment occurs when the trophozoite emerges from produced by several amebas of the genus Acanthamoeba. the cyst under favorable environmental conditions. There is First described in the 1970’sasanocularpathogen,1 it a seasonal trend toward disease onset in the summer months.14,15 causes a chronic keratitis that is often refractory to traditional antibiotic therapy. There is often a poor prognosis because of a significant delay in diagnosis and Epidemiology frequently a lack of effective medical treatment. AK has Acanthamoeba keratitis is most common in CL wearers, been increasing in prevalence in recent years; causes seem with reported rates in the range of 1 to 33 cases per million to be multifactorial but outbreaks have been recorded CL wearers.16 The wide variation is thought to be because of because of the use of certain contact lenses (CL) and their the prevalence of CL use, the contamination of domestic and – cleaning solutions.2 6 New and improved methods of swimming pool water, the amebicidal efficacy of CL cleaning diagnosis and treatment are needed to effectively manage systems, the use of reusable soft CLs, and the use of this condition. diagnostic tests for AK.17,18

Received for publication September 17, 2015; revision received January 10, Pathogenesis 2016; accepted January 14, 2016. Published online ahead of print March 18, 2016. Our understanding of how Acanthamoeba invades 8,19 From the Eye Department, Queen Alexandra Hospital, Portsmouth, Hants, the cornea comes from animal studies. Adhesion is an United Kingdom. important ﬁrst step and has been shown to be controlled by The authors have no funding or conﬂicts of interest to disclose. a number of adhesion proteins and cell surface molecules. Reprints: Nick Maycock, Department of Eye, Queen Alexandra Hospital, Portsmouth, Hants PO6 3LY, United Kingdom (e-mail: nickmaycock@ This leads to phagocytosis and release of enzymes yahoo.com). and toxins such as ecto-ATPase, neuraminidase, superox- Copyright © 2016 Wolters Kluwer Health, Inc. All rights reserved. ide dismutase, elastase, protease, phospholipases,

Cornea Volume 35, Number 5, May 2016 www.corneajrnl.com | 713

Copyright Ó 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited. Maycock and Jayaswal Cornea Volume 35, Number 5, May 2016

multipurpose solution and hydrogen peroxide solution still contract AK. It has been shown that most commercially available multipurpose solutions are ineffective against Acanthamoeba,31–35 suggesting other factors are at play and further research is required. Other documented causes include swimming and showering with CLs in situ, inadequate CL disinfection, exposure to contaminated water, and corneal trauma. Most cases occur in CL wearers, and AK prevalence can be reduced by strict contact lens hygiene. Typically, amebas inhabit the lens case via tap water, rapidly multiply if the case is not cleaned properly and regularly, and then infect the eye via the CLs.36

DIAGNOSIS It is important to remember and consider AK in all cases of CL keratitis, especially where onset is insidious and features are atypical as early diagnosis and treatment is essential to ensure a good visual outcome.37–41 Clinical features are often mistaken for fungal or herpes simplex FIGURE 1. Pictures of Acanthamoeba cyst and trophozoite. infection, which may lead to a delay in diagnosis and appropriate treatment. It is important to remember that AK is polymicrobial or coinfected with herpes simplex virus in glycosidases, and acanthaporin.20 The resulting stromal 10% to 23% of cases.13 degradation allows deep penetration into the cornea. Acanthamoeba trophozoites are able to penetrate Descemet membrane, but intraocular infection rarely occurs because Clinical Features of the intense neutrophil response in the anterior The disease is usually unilateral and should be consid- chamber.21 ered in any case of keratitis or trauma contaminated with soil CL use causes microtrauma to the epithelium and an or water. There is a spectrum of disease with a progression of upregulation of glycoproteins. Soft lenses have a more symptoms and signs from epithelial to stromal disease: the adherent surface than hard lenses, which allows the tropho- pain is often severe and disproportionate to the clinical zoite to bind and thereby gain access to the corneal surface. signs, whereas some may be pain free. Photophobia, pain, Trophozoites bind to glycoproteins on the corneal epithelium, epiphora, and reduced visual acuity are commonly the first triggering the release of a number of cytopathic factors reported symptoms. including mannose-induced protein 133. Corneal epithelial Early findings include an epitheliopathy with punctate destruction and apoptosis allow invasion of the stroma. The keratopathy, pseudodendrites (Fig. 3), epithelial or subepi- release of multiple proteases allows the organism to penetrate thelial infiltrates, and perineural infiltrates. Tu et al38,42 deep into the stroma,12,22–24 causing direct cytolysis, phago- described 5 levels of AK severity: epitheliitis, epitheliitis cytosis, and apoptosis.8 They cluster around corneal nerves with radial neuritis, anterior stromal disease, deep stromal leading to radial keratoneuritis,25 and it is thought that keratitis, or ring infiltrate. The characteristic ring infiltrate is trophozoite-derived proteases contribute to nerve damage only seen in approximately 50% of patients (Fig. 4). In the and the severe pain associated with AK.8 early stage, it can easily be confused with Herpes simplex A thorough understanding of the immune response is keratitis, whereas in the advanced stage, the infection important for management of the condition. Topical cortico- resembles the clinical picture of a fungal keratitis or a corneal steroids, which are widely used in ophthalmology, are known ulcer.20 Perineural infiltrates are highly suggestive for AK and to suppress the activity of neutrophils and macrophages have been reported in up to 63% of cases at 6 weeks.13 in vivo. Interestingly, they have been shown to accelerate Although certain features are pathognomic for the condition, trophozoite excystment and proliferation, which may poten- they are not always present, and the clinician must therefore tially increase their susceptibility to antiamebic therapy.13 have a high index of suspicion in cases where the history and However, caution must be urged in their use in any infectious other features are suggestive. keratitis because of the potential risk of worsening As the disease progresses, ring infiltrates, ulceration, the condition. and a secondary sterile anterior uveitis with hypopyon are common. Perineural infiltrates regress, but the condition may lead to iris damage with a permanently dilated pupil, abscess Risk Factors formation, scleritis, glaucoma, cataract, corneal melt, and The main risk factor is CL wear.3,13,26–30 Although lens perforation. Advanced disease with perforation can lead to hygiene is often poor, patients who regularly disinfect with a cataract, which is a poor prognostic sign. Bacterial

Copyright Ó 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited. Cornea Volume 35, Number 5, May 2016 Diagnosis, Treatment, and Outcomes of AK

FIGURE 2. Life cycle of Acanthamoeba spp. in humans (from Centers for Disease Control 2010: http://www.cdc.gov/parasites/ acanthamoeba/biology.html). Adaptations are themselves works protected by copyright. So in order to publish this adaptation, authorization must be obtained both from the owner of the copyright in the original work and from the owner of copyright in the translation or adaptation. superinfection should always be considered, especially if Immunohistological staining with monoclonal antibodies there is no response to treatment. has improved detection, but techniques are invasive, time- consuming, and reliant on the technician’s skill. Consequently, they are often delayed until there is a high index of suspicion or Culture when there has been no response to treatment. Repeat testing is Amebic cultures should be included in any corneal required if initial investigations are negative, and it has limited scrape in suspected infective keratitis. Acanthamoeba feeds use in assessing treatment response. The effectiveness of readily on inactivated E. coli agar plates, and cultures need to isolating Acanthamoeba in cultures has been reported as be reviewed daily under a light microscope for trails that between 30% and 60%,37,46 and it may require weeks to obtain indicate migration of Acanthamoeba. Cultures are often used final results from the laboratory.47 Therefore, clinicians must be in conjunction with a smear slide or small lamellar disc primarily guided by the history and clinical findings. biopsy for microscopy. A number of stains are used for the detection of cysts: Lactophenol-cotton blue, acridine orange, calcofluor white, silver, immunoperoxidase, and hematoxylin Confocal Microscopy and eosin.20,43 Acanthamoeba has been cultured from CLs, Confocal microscopy allows examination of corneal lens cases, and lens cleaning solutions when corneal biopsies structures at a cellular level in real time.46–53 Previously were negative.12,21,44,45 It is imperative that clinicians ask limited by low resolution, newer systems and computer patients to surrender their most recent CLs and lens cases for software allow a more rapid and detailed examination and microbiological investigation. permit monitoring of the disease course. It is noninvasive and

Copyright Ó 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited. Maycock and Jayaswal Cornea Volume 35, Number 5, May 2016

Polymerase Chain Reaction Polymerase chain reaction (PCR) can identify the presence of amebic DNA in a tissue sample. The process is technical and expensive38,42 but has significant potential diagnostic properties. However, a diagnosis of AK is not confirmed unless there is a corresponding positive tissue sample: in 2 UK National Surveys, a tissue diagnosis was only obtained in 177/349 cases (50%).26,55 A frequently used PCR technique involves amplification of a fragment of the 18S recombinant RNA gene and was first described by Schroeder et al56 in 2001. Several studies have reported the use of real-time PCR to diagnose AK.57–61 Real- time PCR can quantitate DNA at the site of infection making it possible to make an accurate diagnosis, start a tailored antiamebic regimen early, and monitor treatment success.

PREVENTION It is important to minimize risk factors for Acantha- moeba infection. Strict CL and storage case hygiene are essential with the use of appropriate disinfecting solutions; no overnight wear of CLs, no topping-off of solutions, and no homemade saline. It is also important to avoid contact with contaminated water such as swimming or showering with CL in situ. Any corneal trauma, however trivial, is a potential portal for infection and must be minimized.

TREATMENT FIGURE 3. Clinical features of Acanthamoeba keratitis: early disease showing linear epitheliopathy and pseudodendrite. Medical Acanthamoeba trophozoites are sensitive to a number has a spatial resolution of 2 to 4 mm, allowing the analysis of of available medications: antibiotics, antiseptics, antifungals, individual corneal cells and the inflammatory response.54 and antiprotozoals, including metronidazole, antivirals, and Because of a small field of view, it is limited in its ability antineoplastic therapies. Acanthamoeba cysts may lead to to confirm a complete absence of Acanthamoeba cysts.47 prolonged or resistant infection as most of the above treat- There is some concern that the high sensitivity and specificity ments are ineffective, and only those that are cysticidal will values for the diagnosis of AK (.90%) may lead to a number stand any chance of success. Cysts have shown a high of false-positive or false-negative results.13 resistance in vitro to imidazoles and antifungals such as voriconazole. The diamidines and biguanides are the most effective cysticidal antiamebics, and their use has been well documented.13,41

Biguanides Polyhexamethylene biguanide (PHMB) 0.02% to 0.06% (200–600 mg/mL) and chlorhexidine 0.02% to 0.2% (200–2000 mg/mL) are commonly used biguanides.41,62 They disrupt the cytoplasmic membrane and damage cell compo- nents and respiratory enzymes. PHMB and chlorhexidine have low minimal cysticidal concentrations of 2 mg/mL, although there have been some reports of clinically resistant organisms.63,64 Both have low levels of corneal epithelial toxicity,13 which support their use as ﬁrst-line therapy. When used in combination with diamidines, there is evidence of a synergistic effect.13,65 PHMB is most frequently used at a concentration of 0.02% but can be increased to 0.06% in FIGURE 4. Clinical features of Acanthamoeba keratitis: classic unresponsive or severe infection. Similarly, chlorhexidine is ring inﬁltrate. commenced at 0.02% but increased to 0.2% if required.

Copyright Ó 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited. Cornea Volume 35, Number 5, May 2016 Diagnosis, Treatment, and Outcomes of AK

Diamidines Resistant Organism Propamidine isethionate 0.1% (1000 mg/mL) and Persistent keratitis is because of the presence of viable hexamidine 0.1% (1000 mg/mL) are commonly used diami- organisms in the cornea despite treatment with a biguanide dines. Their antimicrobial effect comes from their ability to and diamidine. It is important to exclude bacterial superin- alter the cell membrane structure and its permeability. They fection or concurrent herpes infection in such cases. In penetrate into the cytoplasm causing denaturation of cyto- addition, it is crucial to differentiate those cases that are plasmic proteins, and enzymes and have been shown to be caused by persistent inflammation rather than effective against both trophozoite and cyst. They are well Acanthamoeba infection. tolerated but prolonged therapy may lead to ocular surface A persistent epithelial defect may be the first evidence toxicity. Hexamidine has been shown to have a faster of a resistant organism. Other causes include epithelial amebicidal effect against both cyst and trophozoites. toxicity from an intensive treatment regimen or severe anterior segment inflammation, both will usually settle spontaneously if adequately controlled. Repeated cultures Treatment Regimens are necessary to determine whether it is a persistent Acan- The goals of therapy are (1) the removal of Acantha- thamoeba infection or cross-infection. Unfortunately, both moeba cysts and trophozoites from the corneal tissue and (2) confocal microscopy and PCR are unable to distinguish resolution of the host inflammatory response. It is thought that between viable or nonviable Acanthamoeba cysts, leaving early intensive therapy is more effective as the cysts have not culture of corneal scrapes or biopsies as the definitive way of had time to penetrate deep into the corneal tissue or mature identifying persistent culture-positive disease. Repeated cor- fully.13 Failure to kill or eradicate all cysts will result in neal cultures must be performed to identify the persistence of recurrence of the disease. viable Acanthamoeba and any possible resistance. If unsuc- Patients are usually commenced on a biguanide and cessful, a lamellar keratectomy removing the infected corneal a diamidine as combination therapy. For the first 48 hours tissue may be therapeutic and diagnostic.13 these are given hourly (day and night) before being reduced to In vitro drug sensitivity testing is another possible hourly (daytime only) for a number of days or weeks. It is option in resistant cases. Recently, a patient with severe AK important that the treatment regimen is reduced and tailored responded to personalized treatment with voriconazole after to each clinical case to minimize any epithelial toxicity. The specific sensitivities to this drug were assayed from amebas in aim is to reduce therapy to 4 times a day, but patients may the patient’s cornea.73 need to be on therapy for up to 6 months.55 Further research is required to assess the effectiveness of available treatments and different regimens; despite low MMC’s and reports of Surgical success, there has been some variation in outcomes.13,66,67 Extensive epithelial debridement of the affected area Extracorneal manifestations of AK may be present in may be therapeutic as (1) it promotes penetration of topical both early and late disease and indicate a poor prognosis. therapy74 and (2) if performed early when the pathogen is still Treatment with oral nonsteroidal anti-inflammatory medica- intraepithelial, it may aid in its removal.13 The debrided tion, high-dose systemic steroids, or other systemic immuno- epithelium is sent for histology, culture and sensitivities. suppressive drugs such as cyclosporine may need to be Since the introduction of biguanides as medical therapy, continued for several months13,68 to control the inflammation PK is not recommended as a means of removal of organisms and eradicate the pathogen. Further work is required to better from the cornea.13,34,69,75 Therapeutic keratoplasty should understand this aspect of the disease and find new and therefore be reserved for (1) corneal perforation that does improved treatments. not respond to repeat gluing, (2) significant cataract, or (3) severe corneal abscess.13,73 Corneal grafts for AK should be kept to the minimum size required because of the risk of Controversy of Topical Steriods rejection with large grafts and the potential need for repeat The use of topical corticosteroids remains controversial grafting. Recurrence in the early postoperative period has in AK. They are usually not required in cases that are reduced significantly since the introduction of biguanides.13,76 diagnosed early and responsive to antiamebic drugs. How- Late recurrence several months after surgery may still occur.13 ever, when there is significant anterior segment inflammation, PK is used to improve vision in patients with scarred steroids may facilitate a rapid resolution of symptoms but corneas or irregular astigmatism after treated/quiescent AK must be used judiciously because of the potential risk of infection. The long-term outcome is good in these pa- worsening the condition by damping down the host inflam- tients.13,69,75,77 There is evidence to suggest that outcomes matory response. It is essential that anti-amoebic therapy is are better if surgery is reserved for visual rehabilitation rather continued for a number of weeks after the steroids have been than therapeutic removal of infected tissue.78 stopped to eradicate the disease.13,69–71 The condition may be Anterior segment inflammation should be treated considered cured once the eye has been free of inflammation with systemic and topical immunosuppression. Antiamebic after 4 weeks of tapered biguanide monotherapy.13 Particular therapy should be used preoperatively and continued post- care must be taken as recent evidence suggests that steroid use operatively to maximize elimination of the organism may result in increased pathogenicity of the amebas.72 Further and minimize any risk of recurrence. Care should be taken work is required to investigate this phenomenon. to use regimens that minimize epithelial toxicity, and

Copyright Ó 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited. Maycock and Jayaswal Cornea Volume 35, Number 5, May 2016 treatment should be continued until culture results confirm suggest any difference in prognosis between the Acantha- elimination of the organism. If culture results are positive, it is moeba subspecies.10 sensible to continue antiamebic therapy and steroids for 6 months, as recurrence has been documented up to 3 months after successful grafting.13 If the culture results are negative, it THE FUTURE? is assumed all organisms have been eliminated and anti- AK remains a challenging condition to diagnose and amebic therapy is usually discontinued after several weeks. treat. Early diagnosis and treatment are reliant on clinical In addition, bipedicle conjunctival flaps and cryopre- signs, which may be complicated by secondary coinfection served amniotic membrane grafts can restore ocular surface and compounded by the poor sensitivity of culture or integrity and provide metabolic and mechanical support for histology. Confocal microscopy has been recommended, but corneal healing in patients with AK.79 However, in large it is thought the sensitivity and specificity of this technique corneal perforations, penetrating keratoplasty remains the are too low to rely on those for a definitive diagnosis. only effective surgical option. Detection of Acanthamoeba DNA via PCR is highly sensitive and specific but needs further research before uptake is more widespread. The development of a more streamlined tech- Other Procedures nique and commercial kit would be a major step forward. Photorefractive surgery has recently been described in If the disease is severe with recurrent scleritis and AK. Kandori et al66 reported 4 cases of AK that developed recalcitrant to standard therapies, it may be necessary to try large corneal stromal abscesses after treatment with topical more than one immunosuppressive agent. Igras and Murphy86 therapy. These were removed using laser phototherapeutic reported a case of 1 patient with severe AK who required keratectomy: final acuities were between 20/16 and 20/25 treatment with 4 immunosupressives to bring it under control. with no disease recurrence. Given the minimal published data Potential regimens of the future may incorporate the use of on the use of this procedure in AK, it is difficult to draw firm multiple immunosuppressants to limit disease spread. conclusions from this limited case series. Other potential areas for further research include Cross-linking (CXL) is another relatively new treatment genetic therapy, new markers, new targets for drug therapy, option. In vitro studies80,81 have shown no amebicidal effect and stem cell research. Three genes, heat shock protein 70, of riboflavin combined with UVA exposure, but some clinical actin-I, and elongation factor-1 alpha have been shown to case reports suggest a more promising picture. Garduño- have varying expression and functions during Acanthamoeba Vieyra et al82 administered CXL to a patient in Mexico T4 genotype differentiation.87 In addition, it has recently been instead of topical medical therapies. A significant improve- shown that interleukin 17A mediates a protective effect from ment was noted after 24 hours, with symptoms resolving after Acanthamoeba infection in the cornea. This is in sharp 3 months. After 5 months, 20/20 vision was reached. Khan contrast with other corneal infections such as herpes and et al83 have since reported 3 similar cases that responded Pseudomonas aeruginosa keratitis where interleukin 17A equally well to CXL, with all ulcers closing within 7 weeks. exacerbates corneal pathology and inflammation.38,42 Further In subsequent PK surgery for scarring, no organisms were understanding and more studies of these different pathways detected in excised tissue. It is possible that the collagen- may be useful in the design of an efficient new therapeutic stabilizing effect prevents further tissue damage and isolates strategy in AK. and prevents reproduction of the amebas. Autophagy inhibitors are under investigation as poten- Further studies are required to investigate in more detail tial future treatments. Moon et al88 evaluated the inhibitors 3- the properties of CXL and how it might work in AK and other methyladenine, LY294002, wortmannin, bafilomycin A, and infectious keratitis. There is an apparent lack of data to chloroquine and assessed their ability to reduce Acanthamoe- suggest CXL works consistently. In addition, there are no ba encystment. In combination with PHMB, they have been significant data to show whether the standard Dresden shown to have low cytopathic effects on human corneal cells protocol is more effective than a pulsed approach, and this and high cytopathic effects on Acanthamoeba cells. requires more investigation and research. Although individual Research suggests that certain bacteria are required for case report results seem promising, there are no formal Acanthamoeba to cause keratitis.89 Nakagawa et al89 showed clinical trials thus far to recommend incorporation into that pretreatment of Acanthamoeba with levofloxacin mark- standard practice. edly reduced disease activity, suggesting another potential, and readily available, treatment avenue. It has been shown that treatment of an Acanthamoeba PROGNOSIS neurotrophic ulcer with a topical regenerating agent The most important factors associated with outcome are [CACICOL20 (1 drop on alternate days) for 8 weeks] led the disease severity at presentation and the time taken to start to rapid resolution of the epithelial defect.90 These agents effective therapy.13,84,85 A delay of more than 3 weeks is require further investigation and randomized controled associated with a worse prognosis, and very few patients do studies to assess their efficacy and potential place in the badly if diagnosed and treated within this period.13 Chew treatment armamentarium. et al15 showed that visual acuity of worse than 20/50 at the A better understanding of the disease process at the time of diagnosis and stromal involvement had a significantly molecular level is essential if we are to provide a faster, more worse prognosis. There are no reports in the literature to reliable diagnosis and a more effective treatment. All these

Copyright Ó 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited. Cornea Volume 35, Number 5, May 2016 Diagnosis, Treatment, and Outcomes of AK new and emerging treatments have the potential to radically 25. Alizadeh H, Niederkorn JY, McCulley JP, et al. Acanthamoeba keratitis. In: improve the management of Acanthamoeba keratitis but Cornea Fundamentals. Diagnosis and Management. 2005;1:1115–1122. 26. Radford CF, Minassian DC, Dart JK. Acanthamoeba keratitis in England require further investigation. Clinical practice has changed and Wales: incidence, outcome and risk factors. Br J Ophthalmol. 2002; little in recent years, and more studies and statistical analysis 86:536–542. are required to validate these potential new treatments. The 27. Kilvington S, Gray T, Dart J, et al. Acanthamoeba keratitis: the role of condition still presents significant challenges for the clinician, domestic tap water contamination in the UK. Invest Ophthalmol Vis Sci. – and new additions to the management armamentarium would 2004;45:165 169. 28. Joslin CE, Tu EY, McMahon TT, et al. Epidemiological characteristics of be very welcome. a Chicago-area Acanthamoeba keratitis outbreak. Am J Ophthalmol. 2006;142:212–217. 29. Jeong HJ, Lee SJ, Kim JH, et al. Acanthamoeba keratopathogenicity REFERENCES of isolates from domestic tap water in Korea. Exp Parasitol. 2007;117: 1. Naginton J, Watson PG, Playfair TJ, et al. Amoebic infection of the eye. 357–367. Lancet. 1974;2:1537–1540. 30. Dart JK, Radford CF, Minassian D, et al. Risk factors for microbial 2. Verhelst D, Koppen C, Van Looveren J, et al. Contact lens-related keratitis with contemporary contact lenses: a case control study. corneal ulcers requiring hospitalization: a 7-year retrospective study in Ophthalmology. 2008;115:1647–1654. Belgium. Acta Ophthalmol Scand. 2006;84:522–526. 31. Zannetti S, Fiori PI, Pinna A, et al. Susceptibility of Acanthamoeba 3. Joslin CE, Tu EY, Shoff ME, et al. The association of contact lens solution castellanii to contact lens disinfecting solutions. Antimicrob Agents and Acanthamoeba keratitis. Am J Ophthalmol. 2007;144:169–180. Chemother. 1995;39:1596–1598. 4. Por YM, Mehta JS, Chua JL, et al. Acanthamoeba keratitis associated 32. Hiti K, Walochnik J, Faschinger C, et al. Microwave treatment of with CL wear in Singapore. Am J Ophthalmol. 2009;148:7–12.e2. contact lens cases contaminated with Acanthamoeba. Cornea. 2001;20: 5. Verani J, Lorick SA, Yoder JS, et al. National outbreak of Acanthamoeba 467–470. keratitis associated with use of CL solution, United States. Emerg Infect 33. Hiti K, Walochnik J, Haller-Schober EM, et al. Viability of Acantha- Dis. 2009;15:1236–1242. moeba after exposure to multi-purpose disinfecting contact lens solution 6. Tu EY, Joslin CE. Recent outbreaks of atypical contact lens-related and two hydrogen peroxide systems. Br J Ophthalmol. 2002;86: keratitis: what have we learned? Am J Ophthalmol. 2010;150: 144–146. 602–608. 34. Hammersmith KM. Diagnosis and management of Acanthamoeba 7. Niederkorn JY, Alizideh H, Leher JP, et al. The pathogenesis of keratitis. Curr Opin Ophthalmol. 2006;17:327–331. Acanthamoeba keratitis. Microbes Infect. 1999;1:437–443. 35. Shoff M, Joslin CE, Tu EY, et al. Efficacy of contact lens systems against 8. Clarke DW, Niederkorn JY. The pathophysiology of Acanthamoeba recent clinical and tap water Acanthamoeba isolates. Cornea. 2008;27: keratitis. Trends Parasitol. 2006;22:175–180. 713–719. 9. Centers for Disease Control and Prevention 2010. Available at: http:// 36. Walochnik J, Scheikl U, Haller-Schober EM. Twenty years of acantha- www.cdc.gov/parasites/acanthamoeba/biology.html. Accessed September moeba diagnostics in Austria. J Eukaryot Microbiol. 2015;62:3–11. 13, 2015. 37. Bacon AS, Fraser DG, Dart JK. A review of 72 consecutive cases of 10. Booton GC, Joslin CE, Shoff M, et al. Genotypic identification of Acanthamoeba keratitis, 1984–1992. Eye (Lond). 1993;7:719–725. Acanthamoeba spp. isolates associated with an outbreak of Acantha- 38. Tu EY, Joslin CE, Sugar J, et al. Prognostic factors affecting visual moeba keratitis. Cornea. 2009;28:673–676. outcome in Acanthamoeba keratitis. Ophthalmology. 2008;115:1998– 11. Bowers B, Korn ED. The fine structure of Acanthamoeba castellani I. 2003. The trophozoite. J Cell. Biol. 1968;39:95–111. 39. Szentmáry N, Goebels S, Matoula P, et al. Acanthamoeba keratitis— 12. Marciano-Cabral F, Cabral G. Acanthamoeba as agents of disease in a rare and often late diagnosed disease. Klin Monbl Augenheilkd. 2012; humans. Clin Microbiol Rev. 2003;16:273–307. 229:521–528. 13. Dart JKG, Saw VPJ, Kilvington S. Acanthamoeba keratitis: diagnosis 40. Bouheraoua N, Gaujoux T, Goldschmidt P, et al. Prognostic factors and treatment update 2009. Am J Ophthalmol. 2009;148:487–499. associated with the need for surgical treatments in Acanthamoeba 14. McAllum P, Bahar I, Kaiserman I, et al. Temporal and seasonal trends in keratitis. Cornea. 2013;32:130–136. Acanthamoeba keratitis. Cornea. 2009;28:7–10. 41. Lorenzo-Morales J, Martín-Navarro CM, López-Arencibia A, et al. 15. Chew HF, Yildiz EH, Hammersmith KM, et al. Clinical outcomes and Acanthamoeba keratitis: an emerging disease gathering importance prognostic factors associated with Acanthamoeba keratitis. Cornea. worldwide? Trends Parasitol. 2013;29:181–187. 2011;30:435–441. 42. Tu YE, Joslin CE, Sugar J, et al. The relative value of confocal 16. Centers for Disease Control and Prevention 2012. Available at: http:// microscopy and superficial corneal scrapings in the diagnosis of www.cdc.gov/parasites/acanthamoeba/epi.html. Accessed September 13, Acanthamoeba keratitis. Cornea. 2008;27:764–772. 2015. 43. Sharma S, Athmanathan S, Ata-Ur-Rasheed M, et al. Evaluation of 17. Ibrahim YW, Boase DL, Cree IA. Factors affecting the epidemiology of immunoperoxidase staining technique in the diagnosis of Acanthamoeba Acanthamoeba keratitis. Ophthalmic Epidemiol. 2007;14:53–60. keratitis. Indian J Ophthalmol. 2001;49:181–186. 18. Watson S, Dart JKG. Acanthamoeba keratitis. In: Johnson GJ, 44. Moore MB, McCulley JP. Acanthamoeba keratitis associated with Minassian D, Weale RA, et al, eds. The Epidemiology of Eye Disease. contact lenses: six consecutive cases of successful management. Br J London, United Kingdom: Arnold; 2003:200–205. Ophthalmol. 1989;73:271–275. 19. Clarke DW, Niederkorn JY. The immunobiology of Acanthamoeba 45. Armstrong M. The pathogenesis of human Acanthamoeba infection. keratitis. Microbes Infect. 2006;8:1400–1405. Infect Dis Rev. 2000;2:65–73. 20. Lorenzo-Morales J, Khan N, Walochnik J. An update on Acanthamoeba 46. Parmar DN, Awwad ST, Petroll WM, et al. Tandem scanning confocal keratitis: diagnosis, pathogenesis and treatment. Parasite. 2015;22:10. corneal microscopy in the diagnosis of Acanthamoeba keratitis. Oph- 21. Illingworth CD, Cook SD, Karabatsas CH, et al. Acanthamoeba keratitis: thalmology. 2006;113:538–547. risk factors and outcome. Br J Ophthalmol. 1995;79:1078–1082. 47. Shiraishi A, Uno T, Oka N, et al. In vivo and in vitro laser confocal 22. Hadas E, Mazur T. Proteolytic enzymes of pathogenetic and non- microscopy to diagnose Acanthamoeba keratitis. Cornea. 2010;29:861–865. pathogenetic strains of Acanthamoeba spp. Trop Med Parasitol. 1993;44: 48. Cavanagh HD, Petroll WM, Alizadeh H, et al. Clinical and diagnostic use 197–200. of confocal microscopy in patients with corneal disease. Ophthalmology. 23. Cao Z, Jefferson DM, Panjwani M Role of carbohydrate-mediated 1993;100:1444–1454. adherence in cytopathogenetic mechanisms of Acanthamoeba. J Biol 49. Bohnke M, Masters BR. Confocal microscopy of the cornea. Prog Retin Chem. 1998;273:15838–15845. Eye Res. 1999;18:553–628. 24. Cho JH, Na BK, Song CY. Purification and characterisation of an 50. Kaufman SC, Musch DC, Belin MW, et al. Confocal microscopy: extracellular serine protease from Acanthamoeba castellani. IUBMB Life. a report by the American Academy of Ophthalmology. Ophthalmology. 2000;50:209–214. 2004;111:396–406.

Copyright Ó 2016 Wolters Kluwer Health, Inc. Unauthorized reproduction of this article is prohibited. Maycock and Jayaswal Cornea Volume 35, Number 5, May 2016

51. Kobayashi A, Sakurai M, Shirao Y, et al. In vivo confocal microscopy in 71. Park DH, Palay DA, Daya SM, et al. The role of corticosteroids in the the acute phase of corneal inflammation. Ophthalmic Surg Lasers management of Acanthamoeba keratitis. Cornea. 1997;16:277–283. Imaging. 2003;34:433–436. 72. Robaei D, Carnt N, Minassian DC, et al. The impact of topical 52. Kobayashi A, Sugiyami K, Huang AJ. In vivo confocal microscopy in corticosteroid use before diagnosis on the outcome of Acanthamoeba patients with central cloudy dystrophy of Francois. Arch Ophthalmol. keratitis. Ophthalmology. 2014;121:1383–1388. 2004;122:1676–1679. 73. Arnalich-Montiel F, Martín-Navarro CM, Alió JL, et al. Successful 53. Kanavi MR, Javadu M, Yazdani S, et al. Sensitivity and specificity of monitoring and treatment of intraocular dissemination of acanthamoeba. confocal scan in the diagnosis of infectious keratitis. Cornea. 2007;26: Arch Ophthalmol. 2012;130:1474–1475. 782–786. 74. Sun X, Zhang Y, Li R. Acanthamoeba keratitis: clinical characteristics 54. Heidelberg Retina Tomograph 2 Operating Instructions of Software and management. Ophthalmology. 2006;113:412–416. Version 1.1. Dossenheim, Germany: Heidelberg Engineering GmbH; 2004. 75. Awwad ST, Parmar DN, Heilman M, et al. Results of penetrating 55. Radford CF, Lehmann OJ, Dart JK. Acanthamoeba keratitis: multi-centre keratoplasty for visual rehabilitation after Acanthamoeba keratitis. Am J survey in England 1992–1996. National Acanthamoeba Keratitis Study Ophthalmol. 2005;140:1080–1084. Group. Br J Ophthalmol. 1998;82:1387–1392. 76. Awwad ST, Petroll WM, McCulley JP, et al. Updates in Acanthamoeba 56. Schroeder JM, Booton GC, Hay J, et al. Use of subgenic 18S ribosomal keratitis. Eye Contact Lens. 2007;33:1–8. DNA PCR and sequencing for genus and genotype identification of 77. Larkin DF, Kilvington S, Dart JK. Treatment of AK with polyhexa- Acanthamoeba from humans with keratitis and from sewage sludge. methylene biguanide. Ophthalmology. 1992;99:185–191. J Clin Microbiol. 2001;39:1903–1911. 78. Robaei D, Carnt N, Minassian DC, et al. Therapeutic and optical 57. Rivière D, Szczebara FM, Berjeaud JM, et al. Development of real-time keratoplasty in the management of Acanthamoeba keratitis: risk PCR assay for quantification of Acanthamoeba trophozoites and cysts. factors, outcomes, and summary of the literature. Ophthalmology. J Microbiol Methods. 2006;64:78–83. 2015;122:17–24. 58. Qvarnstrom Y, Visvesvara GS, Sriram R, et al. Multiplex real-time PCR 79. Abdulhalim BE, Wagih MM, Gad AA, et al. Amniotic membrane graft to assay for simultaneous detection of Acanthamoeba spp., Balamuthia conjunctival flap in treatment of non-viral resistant infectious keratitis: mandrillaris and Naegleria fowleri. J Clin Microbiol. 2006;44:3589–3595. a randomised clinical study. Br J Ophthalmol. 2015;99:59–63. 59. Thompson PP, Kowalski RP, Shanks RM, et al. Validation of real-time 80. Kashiwabuchi RT, Carvalho FR, Khan YA, et al. Assessing efficacy of PCR for laboratory diagnosis of Acanthamoeba keratitis. J Clin Micro- combined riboflavin and UV-A light (365 nm) treatment of Acantha- biol. 2008;46:3232–3236. moeba trophozoites. Invest Ophthalmol Vis Sci. 2011;52:9333–9338. 60. Kandori M, Inoue T, Takamatsu F, et al. Two cases of Acanthamoeba 81. del Buey MA, Cristóbal JA, Casas P, et al. Evaluation of in vitro efficacy keratitis diagnosed only by real-time polymerase chain reaction. Cornea. of combined riboflavin and ultraviolet a for Acanthamoeba isolates. Am J 2010;29:228–231. Ophthalmol. 2012;153:399–404. 61. Itahashi M, Higaki S, Fukuda M, et al. Utility of real-time polymerase 82. Garduño-Vieyra L, Gonzalez-Sanchez CR, Hernandez-Da Mota SE. chain reaction in diagnosing and treating Acanthamoeba keratitis. Ultraviolet-a light and riboflavin therapy for Acanthamoeba keratitis: Cornea. 2011;30:1233–1237. a case report. Case Rep Ophthalmol. 2011;2:291–295. 62. Lim N, Goh D, Bunce C, et al. Comparison of polyhexamethylene 83. Khan YA, Kashiwabuchi RT, Martins SA, et al. Riboflavin and biguanide and chlorhexidine as monotherapy agents in the treatment of ultraviolet light a therapy as an adjuvant treatment for medically Acanthamoeba keratitis. Am J Ophthalmol. 2008;145:130–135. refractive Acanthamoeba keratitis: report of 3 cases. Ophthalmology. 63. Kilvington S, Hughes R, Byas J, et al. Activities of therapeutic agents 2011;118:324–331. and myristamidopropyl dimethylamine against Acanthamoeba isolates. 84. Bacon AS, Dart JK, Ficker LA, et al. Acanthamoeba keratitis. The value Antimicrob Agents Chemother. 2002;46:2007–2009. of early diagnosis. Ophthalmology. 1993;100:1238–1243. 64. Perez-Santonja JJ, Kilvington S, Hughes R, et al. Persistently culture 85. Claerhout I, Goegebuer A, van Den BC, et al. Delay in diagnosis and positive Acanthamoeba keratitis: in vivo resistance and in vitro sensitiv- outcome of Acanthamoeba keratitis. Graefes Arch Clin Exp Ophthal. ity. Ophthalmology. 2003;110:1593–1600. 2004;242:648–653. 65. Hay J, Kirkness CM, Seal DV, et al. Drug resistance and Acanthamoeba 86. Igras E, Murphy C. Use of multiple immunosuppressive agents in keratitis: the quest for alternative antiprotozoal chemotherapy. Eye recalcitrant Acanthamoeba scleritis. BMJ Case Rep. 2015;2015: (Lond). 1994;8:555–563. bcr2014208536. 66. Kandori M, Inoue T, Shimabukuro M, et al. Four cases of Acanthamoeba 87. Abedkhojaseh H, Niyyati M, Rezaei S, et al. Identifying differentially keratitis treated with phototherapeutic keratectomy. Cornea. 2010;29: expressed genes in trophozoites and cysts of Acanthamoeba T4 1199–1202. genotype: implications for developing new treatments for Acanthamoeba 67. Elder MJ, Kilvington S, Dart JK. A clinicopathological study of in vitro keratitis. Eur J Protistol. 2015;51:34–41. sensitivity testing and Acanthamoeba keratitis. Invest Ophthalmol Vis 88. Moon EK, Kim SH, Hong Y, et al. Autophagy inhibitors as a potential Sci. 1994;35:1059–1064. antiamoebic treatment for Acanthamoeba keratitis. Antimicrob Agents 68. Lee GA, Gray TB, Dart JK. Acanthamoeba sclerokeratitis: treatment with Chemother. 2015;59:4020–4025. systemic immunosuppression. Ophthalmology. 2002;109:1178–1182. 89. Nakagawa H, Hattori T, Koike N, et al. Investigation of the role of 69. Illingworth CD, Cook SD. Acanthamoeba keratitis. Surv Ophthalmol. bacteria in the development of Acanthamoeba keratitis. Cornea. 2015;34: 1998;42:493–508. 1308–1315. 70. Radford CF, Bacon AS, Dart JK, et al. Risk factors for Acanthamoeba 90. Mateo A, Abadía B, Calvo P, et al. Treatment of Acanthamoeba keratitis in contact lens users: a case-control study. BMJ. 1995;310: neurotrophic corneal ulcer with topical matrix therapy. J Ophthalmic 1567–1570. Inflamm Infect. 2015;5:18.