Acanthamoeba keratitis: improving the Scottish diagnostic service for the rapid molecular detection of Acanthamoeba species

Abstract

Acanthamoeba species are responsible for causing the potentially sight-threatening condition, Acanthamoeba keratitis, which is commonly associated with contact lens use. In this report, we highlight the challenges faced using conventional laboratory identification methods to identify this often under-reported pathogen, and discuss the reasons for introducing the first national service in Scotland for the rapid and sensitive molecular identification of Acanthamoeba species. By comparing culture and molecular testing data from a total of 63 patients (n580 samples) throughout Scotland presenting with ocular eye disease, we describe the improvement in detection rates where an additional four positive cases were identified using a molecular assay versus culture. The testing of a further ten patients by confocal imaging is also presented. This report emphasizes the importance of continuing to improve clinical laboratory services to ensure a prompt, correct diagnosis and better prognosis, in addition to raising awareness of this potentially debilitating opportunistic pathogen. INTRODUCTIONAcanthamoeba keratitis is an uncommon but sightthreatening disease of the cornea caused by free-living protozoan amoebae belonging to the genus Acanthamoeba (Lorenzo- Morales et al., 2013). This painful, often unilateral eye disease may eventually progress to blindness and is associated with contact lens use but can also be diagnosed in non-contact lens wearers. Acanthamoeba species can be found in a variety of different environments including recreational and drinking water, soil and air (Gianinazzi et al., 2009;Legarreta et al., 2013;Rivera et al., 1991). Trophozoites and highly robust cysts can persist for years and are often resistant to drug treatments.Global increases of Acanthamoeba keratitis occurred during the 1970s and 1980s with links to contact lens use in over 85% of cases, which is consistent with recent data from America and Canada (Stehr-Green et al., 1989;Fraser et al., 2012;Page & Mathers, 2013). In addition, noncommercial saline solutions have been implicated as potential risk factors (Newton et al., 1986). The UK experienced an increase in cases during the 1990s which coincided with increased use of soft contact lenses (Illingworth et al., 1995).There is a lack of recent robust data on the prevalence of Acanthamoeba keratitis in Scotland, but based on a study by Seal et al. (1999), the incidence was 149 per million soft contact lens wearers compared with much lower incidence rates of around 18 to 21 per million in the rest of the UK (Radford et al., 2002). Presently, there are approximately 3.7 million contact lens wearers in the UK (British Contact Lens Association, http://www.bcla.org.uk/).An early, accurate diagnosis is crucial for achieving a better prognosis. However, conventional techniques to detect Acanthamoeba species may lack sensitivity, which is likely to result in the true prevalence being underestimated. The challenges of an early diagnosis are compounded by the ability of Acanthamoeba to mimic non-...

In Vitro Evaluation of the Inhibitory Effect of Topical Ophthalmic Agents on Acanthamoeba Viability

“…The results obtained in this study for chlorhexidine and PHMB are consistent with previously published studies, with PHMB demonstrating superior activity to chlorhexidine 31,36. The standard topical PHMB used in treating AK is 0.02% (200 μg/ml), which is nearly 20 times the mean MCC for PHMB found in this study; the finding agrees with the general favorable in vitro sensitivities and clinical outcome with PHMB compared to other antiamoebic drugs 15. However, Sunada et al35 did not find that their Acanthamoeba isolates had high in vitro susceptibility to PHMB.…”

“…The main limitation of this study is that findings from in vitro sensitivities do not always correlate with clinical outcome 15. There are many factors to explain this.…”

“…In the United Kingdom, the diagnosis of AK is not standardized and depends largely upon individual clinics and hospitals 15. A variety of methods can be used with culture of a corneal scrape on 2.5% nonnutrient agar that has been overlaid with a lawn of Escherichia coli , the most common method utilized and considered the gold standard.…”

Acanthamoeba keratitis: Molecular typing of Acanthamoeba species directly from ocular tissue

“…In recent years, advances in laboratory techniques have improved the diagnosis of AK which is crucial, especially as reports of this disease within the UK have increased over the past ten years ( Carnt et al, 2020 ). These include the implementation of molecular tests such as real-time PCR assays ( Rivière et al, 2006 ; Alexander et al, 2015 ) and conventional PCR assays ( Schroeder et al, 2001 ; Booton et al, 2002 ; Di Cave et al, 2009 ). These rapid and sensitive molecular assays provide in-depth data by amplifying the 18S ribosomal ribonucleic acid (rRNA) small subunit region known as DF3 (diagnostic fragment 3, approximately 240 nucleotides long).…”

Queratitis por Acanthamoeba sp. en usuarios de lentes de contacto: casos clínicos

“…Otro factor que predispone a la infección es el antecedente de traumatismo y posterior exposición al parásito. En este trabajo se presentan tres casos de QA asociada al uso de LC siendo el segundo reporte de casos de QA a nivel país, el primer caso reportado hace referencia a una mujer usuaria de LC que desarrolló la infección después de almacenar sus LC en agua de canilla (12)(13) .…”