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Background: The genus Acanthamoeba is a free-living opportunistic protozoan parasite, which widely distributed in soil and fresh water. Acanthamoeba keratitis, which causes a sight-threating infection of the cornea, is going to rise in Iran and worldwide. The aim of this study was to compare direct microscopy, culture and PCR for detection of Acanthamoeba spp. in clinical samples and to determine the genotypes of Acanthamoeba spp. by sequencing 18SrRNA gene. Methods: Among patients clinically suspected to AK referred to a tertiary ophthalmology center at Mashhad, northeastern Iran. During 2017-18, twenty corneal scrapes specimens obtained. The samples were divided into three parts, subjected to direct microscopic examination, culture onto non-nutrient agar and PCR technique. Sensitivity, specificity, accuracy and likelihood ratio were evaluated. Results: Among 20 persons clinically suspected to amoebic keratitis, 13(69.2%) patients definitely diagnosed as Acanthamoeba keratitis. Wearing contact lens, eye trauma due to foreign particle and swimming in fresh water were the main predisposing factors. Most of patients suffered from pain and photophobia. Corneal ring infiltration and epithelial defect were common clinical sings. Direct examination had the lowest sensitivity and sensitivity of both Nelson-PCR and JDP-PCR methods were equal and highest. In addition, the results of sequencing identified that all strains belonged to T4 genotype. Conclusion: Amoebic keratitis is a sporadic parasitic keratitis, which is mainly seen in contact lens user in Mashhad. PCR based on 18S ribosomal DNA with JDP primers is a reliable and highly sensitive method for diagnosis of Acanthamoeba keratitis in clinically suspected cases.

Purpose: To describe two cases of Acanthamoeba keratitis diagnosed and treated at the epithelial stage of disease and to underscore the importance of early diagnosis on prognosis. Observations: Case 1 is a 28-year-old male who developed Acanthamoeba keratitis after prolonged contact lens wear. Case 2 is a 43-year-old male with poor contact lens hygiene who was initially misdiagnosed and treated for herpetic keratitis. Both cases presented with epitheliopathy and were successfully treated with corneal epithelial debridement and topical anti-amoebic therapy, with complete avoidance of deeper extension of infection and associated complications. Conclusion and importance: Epithelial stage Acanthamoeba keratitis represents a critical window of opportunity to achieve rapid cure. Acanthamoeba epitheliopathy may be mistaken for other conditions such as herpetic keratitis, contact lens overwear, or dry eye. Given worsening prognosis following delayed diagnosis, it is important for clinicians to be suspicious of Acanthamoeba keratitis in all contact lens wearers who develop elevated epitheliopathy.

Background: Acanthamoeba keratitis (AK) is a vision-threatening disease, usually associated with contact lens (CL) wear. As overnight orthokeratology (OOK) is increasingly used to control myopia, we have found incidence of OOK-associated AK is increasing. This study aimed to investigate the clinical presentation and visual outcomes of OOK-associated AK. Methods: Demographic characteristics, clinical features, and treatment outcomes were collected by reviewing the medical charts of CL-associated AK patients (n = 35) diagnosed at Taipei Veterans General Hospital from 2001 to 2016. Cases were OOKassociated AK patients (n = 13), and controls were all other CL-associated AK patients (n = 22). Student t tests and chi-square tests were used to compare cases and controls. Linear regression analyses were used to identify factors associated with the final visual outcome in CL-associated AK. Results: OOK-associated AK accounted for half of all CL-associated AK after 2010. OOK-associated AK patients and other CL-associated patients had similar best-corrected logarithm of the minimum angle of resolution visual acuity (BCLVA) before treatment (1.10 ± 0.75 vs 1.13 ± 0.76, p = 0.893), but OOK-associated AK patients were younger (17.15 ± 3.21 vs 26.36 ± 12.81 years, p = 0.004), had less severe disease (ring infiltration, 0% vs 31.82%, p = 0.023), and had better post-treatment BCLVA (0.06 ± 0.15 vs 0.51 ± 0.95, p = 0.041). Multiple linear regression analysis showed that better BCLVA after treatment in CL-associated AK was associated with initial presentation without ring infiltration (p = 0.002) but not with OOK use itself (p = 0.793). Twenty-six of 35 CL-associated AK patients had final BCLVA equal to or better than 0.10 (Snellen visual acuity of 6/7.5). All 13 OOK-associated AK cases were treated with chlorhexidine 0.02% ± voriconazole 1% ± oral voriconazole, and 12 of these patients had final BCLVA equal to or better than 0.10. Conclusion: Most CL-associated AK patients had satisfactory visual outcomes. Half of AK at our hospital is OOK-associated since 2010. Early diagnosis and correct treatment may be the reason why OOK-associated AK patients had better vision prognosis. [ABSTRACT FROM AUTHOR]

Acanthamoeba keratitis (AK) is a rare infectious disease and accurate diagnosis has remained arduous as clinical manifestations of AK were similar to keratitis of viral, bacterial, or fungal origins. In this study, we described the production of a polyclonal peptide antibody against the adenylyl cyclase-associated protein (ACAP) of A. castellanii, and evaluated its differential diagnostic potential. Enzyme-linked immunosorbent assay revealed high titers of A. castellaniispecific IgG and IgA antibodies being present in low dilutions of immunized rabbit serum. Western blot analysis revealed that the ACAP antibody specifically interacted with A. castellanii, while not interacting with human corneal epithelial (HCE) cells and other causes of keratitis such as Fusarium solani, Pseudomonas aeruginosa, and Staphylococcus aureus. Immunocytochemistry (ICC) results confirmed the specific detection of trophozoites and cysts of A. castellanii co-cultured with HCE cells. The ACAP antibody also specifically interacted with the trophozoites and cysts of 5 other Acanthamoeba species. These results indicate that the ACAP antibody of A. castellanii can specifically detect multiple AK-causing members belonging to the genus Acanthamoeba and may be useful for differentially diagnosing Acanthamoeba infections. [ABSTRACT FROM AUTHOR]

Contact lens usage has contributed to increased incidence rates of Acanthamoeba keratitis (AK), a serious corneal infection that can lead to blindness. Since symptoms associated with AK closely resemble those incurred by bacterial or fungal keratitis, developing a diagnostic method enabling rapid detection with a high degree of Acanthamoeba-specificity would be beneficial. Here, we produced a polyclonal antibody targeting the carboxylesterase (CE) superfamily protein secreted by the pathogenic Acanthamoeba and evaluated its diagnostic potential. Western blot analysis revealed that the CE antibody specifically interacts with the cell lysates and conditioned media of pathogenic Acanthamoeba, which were not observed from the cell lysates and conditioned media of human corneal epithelial (HCE) cells, Fusarium solani, Staphylococcus aureus, and Pseudomonas aeruginosa. High titers of A. castellanii-specific antibody production were confirmed sera of immunized mice via ELISA, and these antibodies were capable of detecting A. castellanii from the cell lysates and their conditioned media. The specificity of the CE antibody was further confirmed on A. castellanii trophozoites and cysts co-cultured with HCE cells, F. solani, S. aureus, and P. aeruginosa using immunocytochemistry. Additionally, the CE antibody produced in this study successfully interacted with 7 different Acanthamoeba species. Our findings demonstrate that the polyclonal CE antibody specifically detects multiple species belong to the genus Acanthamoeba, thus highlighting its potential as AK diagnostic tool. [ABSTRACT FROM AUTHOR]

Members of the protozoan genus Acanthamoeba are free-living amoebas and the primary causative agent of amoebic keratitis and granulomatous encephalitis. Amoebic keratitis mainly occurs in individuals who wear soft contact lenses following trauma and exposure to contaminated sources. As the survival, growth, and multiplication of this protozoan increases in biofilms, which is the microbial accumulation, several cases of amoebic keratitis have been reported as co-infections with bacteria, especially Pseudomonas aureginosa. Pseudomonas improves the binding of this protozoan to soft lens surfaces by its special surface enzymes, resulting in Acanthamoeba keratitis and sometimes mixed infection. Biofilm accumulates microorganisms, including bacteria and fungi, on living and non-living surfaces. The firm attachment of these microbial accumulations to the surface can cause various diseases and resistance to treatment. The critical point is that bacteria have less destructive effects in the plantonik state, and when the specific signals of these microbiome cause the recall of other microorganisms and their interaction, they lead to problems. Unfortunately, the ocular defense barrier, which includes the eyelids, lacrimal glands, and tears, cannot kill microbial agents when microorganisms accumulate and adhere firmly to the lens surfaces. Due to the great importance of AK, especially in medical contact lens users, biofilm formation is an important issue that has been considered in recent years. Biofilm formation-induced drug resistance, which is caused by the disproportionate density of microorganisms on the inner and outer surfaces of the biofilm. There are several ways to prevent the formation of biofilms on the surfaces of contact lenses, including the use of antimicrobial peptides and various antibiotics and coating them on the inner surface of contact lenses to slow-release of antibiotics and prevent biofilm formation. In the present review article, we aimed to introduce biofilms as one of the important risk factors for developing Acanthamoeba keratitis and prevention ways of biofilms formation. [ABSTRACT FROM AUTHOR]

Infection with Acanthamoeba spp. may result in granulomatous amoebic encephalitis and Acanthamoeba keratitis. Water is an important habitat where Acanthamoeba species thrive. Therefore, studying the occurrence of this free-living amoeba in water sources will help understand the infection dynamics. The aim of the study was to survey and report on the presence of Acanthamoeba spp. in water resources from the Ordu and Giresun provinces in Black Sea. Acanthamoeba spp. was found in 1/17 natural spring water samples from Ordu and in 2/18 from Giresun. Acanthamoeba species were not detected in any of the investigated tap water samples. Sequencing of the (SSU) rDNA gene resulted in the identification of haplotype I (Acanthamoeba genotype: KJ094684). T4 (8.6%) was the only isolated genotype in both Ordu and Giresun provinces. This is the first report of Acanthamoeba T4 genotype in natural spring water resources in the Black Sea. The occurrence of Acanthamoeba species in natural spring water sources should be considered as a potential risk for human infection, especially to highrisk populations. [ABSTRACT FROM AUTHOR]

Acanthamoeba is a genus of free‐living amoebae, pervasively found in the environment. Most of its pathogenic species are the causative agent of sight‐threatening Acanthamoeba keratitis and fatal granulomatous amoebic encephalitis. Despite the advancements in the field of chemotherapy, treating Acanthamoeba infections is still challenging due to incomplete knowledge of the complicated pathophysiology. In case of infection, the treatment regimen for the patients is often ineffective due to delayed diagnosis, poor specificity, and side‐effects. Besides the resistance of Acanthamoeba cysts to most of the drugs, the recurrence of infection further complicates the recovery. Thus, it is necessary to develop an effective treatment which can eradicate these rare, but serious infections. Based on various computational and in vitro studies, it has been established that the synthetic scaffolds such as heterocyclic compounds may act as potential drug leads for the development of antiamoebic drugs. In this review, we report different classes of synthetic compounds especially heterocyclic compounds which have shown promising results against Acanthamoeba. Moreover, the antiamoebic activities of synthetic compounds with their possible mode of actions against Acanthamoeba, have been summarized and discussed in this review. [ABSTRACT FROM AUTHOR]

Abstract Background Free-living amoebae of the genus Acanthamoeba are cosmopolitan, widely distributed protozoans that cause a severe, vision-threatening corneal infection known as Acanthamoeba keratitis (AK). The majority of the increasing number of AK cases are associated with contact lens use. Appropriate eye hygiene and effective contact lens disinfection are crucial in the prevention of AK because of the lack of effective therapies against it. Currently available multipurpose contact lens disinfection systems are not fully effective against Acanthamoeba trophozoites and cysts. There is an urgent need to increase the disinfecting activity of these systems to prevent AK infections. Synthesized nanoparticles (NPs) have been recently studied and proposed as a new generation of anti-microbial agents. It is also known that some plant metabolites, including tannins, have anti-parasitic activity. The aim of this study was to evaluate the anti-amoebic activity and cytotoxicity of tannic acid-modified silver NPs (AgTANPs) conjugated with selected multipurpose contact lens solutions. Methods The anti-amoebic activities of pure contact lens care solutions, and NPs conjugated with contact lens care solutions, were examined in vitro by a colorimetric assay based on the oxido-reduction of alamarBlue. The cytotoxicity assays were performed using a fibroblast HS-5 (ATCC CRL-11882) cell line. The results were statistically analysed by ANOVA and Student-Newman-Keuls test using P < 0.05 as the level of statistical significance. Results We show that the NPs enhance the anti-Acanthamoeba activities of the tested contact lens solutions without increasing their cytotoxicity profiles. The activities are enhanced within the minimal disinfection time recommended by the manufacturers. Conclusions The conjugation of the selected contact lens solutions with AgTANPs might be a novel and promising approach for the prevention of AK infections among contact lens users.

A 28-year-old woman with a history of trauma to her right eye 2 months prior reported experiencing a stinging sensation and tearing in the morning since the injury occurred and the need to occasionally use an eye patch. Three days before presentation she had been prescribed a therapeutic contact lens (CL) with the diagnosis of a corneal epithelial defect. She described significant pain despite the CL. There was a corneal lesion with haze at the base surrounded by corneal edema. Corneal confocal images revealed hyperreflective cystic lesions that suggested Acanthamoeba keratitis (AK). However, the lesion healed within 10 days and the results of cultures taken before the initiation of treatment proved to be negative. The history of trauma and CL wear, the presence of severe pain, corneal findings, and the confocal microscopy detection of cysts led to a suspicion of AK in a differential diagnosis, but the final diagnosis was recurrent epithelial erosion based on the negative culture results, quick response to treatment, and the possibility of similar confocal findings in a healing epithelium. Since AK may cause loss of vision, suspicion should require that samples be obtained for microbiological study and close follow-up of the clinical course until a final diagnosis can be achieved.

A 35-year old soft contact lens wearer with a proven bilateral Acanthamoeba keratitis developed a nodular scleritis. Based on the stepladder approach described by Iovieno et al. [Ophthalmology. 2014 Dec;121(12):2340–7], nonsteroidal anti-inflammatory drugs, methylprednisolone, and later azathioprine were added to the antiamoebic treatment. Unfortunately, there was further deterioration and an endophthalmitis developed. Unbearable pain and concerns of spread to the brain urged an enucleation. Histopathological examination confirmed Acanthamoeba cysts in the cornea, sclera, retina, choroid, and vitreous body. As a side effect of the immunosuppressive treatment, the patient developed myopathy, pulmonary aspergillosis, and an avascular necrosis of the hip. Scleritis is a devastating complication of Acanthamoeba keratitis with a poor prognosis and a high enucleation rate. Acanthamoeba sclerokeratitis is, due to cyst-free biopsies, mostly assigned to an immune-mediated mechanism, justifying the use of immunosuppressive treatment. Scleritis in our case contributed to the extracorneal spread of Acanthamoeba. Our case is the first documented extracorneal spread of Acanthamoeba without previous surgery. Extracorneal spread of Acanthamoeba should be considered, even in the case of false-negative biopsies. We strongly recommend serial sections of the retrieved scleral specimen in case of negative histopathological examination to exclude an infection. Even when an immune-mediated scleritis is suspected, systemic immunosuppressive treatment should always be used with the greatest caution. Awareness of the side effects and monitoring by an experienced physician is mandatory.

The Aсanthamoeba keratitis is a rare, but very dangerous disease of cornea; its development is connected with contact lens wearing, and corneal microtraumas with contamination of particles of the soil or water. The disease has no pronounced distinguishing clinical signs therefore it is difficult to distinguish them from herpetic or fungal keratitis. Nevertheless, it can be suspected on the basis of anamnestic data (use of contact lenses, corneal injury with contamination by the soil or water), uneven strong pain syndrome, ring-shaped infiltration of cornea, absence of effect of traditional therapy. Isolation of Acanthamoeba by bioculture method, morphological research of corneal bioptat (by light and luminescent microscopy), gene diagnostics, in – vivo confocal microscopy of the cornea can give additional help in the diagnosis. Also perspective diagnostic methods of Acanthamoeba keratitis are given. Nevertheless, so far there is no conventional technique of diagnosis of this disease.

To the best of our knowledge, this is the second reported case of Acanthamoeba keratitis (AK) as a result of scleral lens use and the first case of AK associated with Maxim scleral lens use (Accu-Lens, Inc., Lakewood, CO, USA). A 22-year-old male scleral lens user presented at the department of ophthalmology at Gazi University Hospital complaining of painful corneal opacities and erosion in the cornea of right eye. A real-time polymerase chain reaction assay (Primerdesign, Southampton, UK) was performed, and Acanthamoeba spp. DNA was amplified on the corneal specimen. A topical antimicrobial treatment was prescribed, and the symptoms had improved significantly at the 2-week follow-up. Contact lens wearers always run the risk of developing AK, even with gas-permeable scleral contact lenses. Therefore, AK must be considered as an important differential diagnosis in patients who use scleral contact lenses.

Purpose: Radial keratoneuritis (RK) is a common feature of Acanthamoeba keratitis (AK). In vivo confocal microscopy (IVCM) is noninvasive and provides real-time images for the diagnosis of corneal diseases by allowing the visualization of corneal structures and morphologies of living organisms at the cellular level. Images of AK with RK obtained using commercial white light IVCM devices have not been frequently evaluated. In the present study, a white light IVCM device was used to evaluate the corneal findings and describe spatial changes in the corneal nerves at different depths in cases of early-stage AK with RK. Methods: In this retrospective, observational study, white light IVCM images focused on RK were evaluated for Acanthamoeba cysts/trophozoites, corneal deposits, and altered corneal nerves, with special emphasis on three-dimensional spatial changes in the corneal nerves at different depths. Results: Seventeen eyes of 17 patients exhibiting early-stage AK with RK were included in the study. Acanthamoeba cysts/trophozoites were observed in the corneal epithelium of 13 eyes and stroma of 7 eyes. Alterations in the corneal nerve morphology and density were observed from the basal epithelial layer to the stromal layer in 12 eyes. Acanthamoeba trophozoites were attached to the corneal stromal nerves in five eyes. Conclusion: These findings suggest that white light IVCM can identify consistent corneal findings, particularly spatial changes in the corneal nerves, in cases of early-stage AK with RK.

Purpose: To report and analyze the outcomes of therapeutic deep anterior lamellar keratoplasty (DALK) in patients with advanced Acanthamoeba keratitis (AK). Methods: Medical records of microbiologically confirmed AK, underwent DALK from 2004 to 2017, were reviewed and the data related to early and late outcome including complications were retrieved. Outcome of cases with largest diameter of infiltrate ≥8 mm at the time of surgery (advanced keratitis) were analyzed and compared with those with less severe keratitis (infiltrate size less than 8 mm). Results: Out of 23 patients of AK in whom DALK was performed, ten (43.4%) patients had advanced keratitis. Mean age of these patients was 38.7 ± 8.6 years (range, 25 to 56). Median visual acuity at presentation was 2.78 (IQR, 1.79–3.0) that improved to 1.79 (IQR, 0.70–2.78) postoperatively. Early complications included recurrence of AK in 2 (20%), Descemet's membrane detachment in 5 (50%), and persistent epithelial defect in 3 (30%) cases. Overall, 6 (60%) grafts failed, whereas 4 (40%) patients had clear graft at their last follow-up. Median follow-up of these cases was 5 months (IQR, 1.4–11.4). One graft developed stromal rejection, which resolved with increased dose of corticosteroids. In comparison, DALK performed for less severe keratitis (N = 13) had 1 (7.6%) recurrence and 2 (15.8%) grafts failure (OR, 8.25). The probability of one-year graft survival and eradication of infection was 32% and 74.1%, respectively, in advanced cases compared to 91.6% and 83.9% in less severe cases. Conclusion: Outcome of DALK in advanced Acanthamoeba keratitis is less favorable compared to those carried out for less severe keratitis cases.

This study aims to describe how in-vivo confocal microscopy (ICVM) results improved diagnosis and treatment in three patients with complex corneal disorders at a single institution. Case one was a 36-year-old woman contact lens wearer referred to the hospital eye service (HES) by her community optician for a suspected corneal ulcer in her left eye. The case demonstrated that where laboratory cell culture was inconclusive, IVCM imaging improved diagnosis and more importantly adjusted the initial treatment till the complete resolution of the case. Case two was a shared-care 66-year-old keratoconus patient under a complex immunosuppression regime who had developed a recent series of post-surgical complications of fungal origin and was experiencing eye pain. IVCM was able to differentiate between an immune-mediated response and fungal keratitis and guide the clinicians towards an optimized treatment. Case three was a long-standing dry eye disease in a 64-year-old woman diagnosed with primary Sjögren’s syndrome where previous treatments failed to improve her symptomatology. IVCM was crucial for prescribing allogeneic serum eyedrops by anticipating early immune changes in the sub-basal corneal nerve plexus. In-vivo confocal microscopy can be an essential non-invasive imaging technique for improving clinicians’ diagnostic precision by adding a layer of certainty that other techniques may lack. Additionally, IVCM allows adjustment of the treatment accordingly, by instantly following any pathologic changes at the cellular level.

Acanthamoeba keratitis (AK) is a blinding corneal infection caused by the protozoan Acanthamoeba. The long-term course of AK suggests the host immunity could not kill Acanthamoeba rapidly. The immune status is still unclear in the late stage of AK. The comparative transcriptome analysis was made based on the bulk RNA sequencing of cornea tissues from AK patients and donors. Differentially expressed genes and enriched signaling pathways were calculated. CIBERSORT algorithm was used for immune infiltration analysis of cornea tissue between AK and normal controls. A total of 2668 differentially expressed genes, including 1477 upregulated genes and 1191 downregulated genes, were detected. Gene Ontology analysis revealed that the pathways were significantly enriched in leukocyte migration, regulation of T-cell activation, the external side of plasma membrane, collagen-containing extracellular matrix, immune receptor activity, and cytokine binding. The Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that the pathways were significantly enriched in the cytokine–cytokine receptor interaction, hematopoietic cell lineage, and Staphylococcus aureus infection pathway. The immune infiltration profiles varied little between AK and normal controls. Compared with normal tissue, cornea tissue of AK contained a higher proportion of M0 macrophages and CD8 T cells, while resting memory CD4 T cells contributed to a relatively lower portion (p < 0.05). Finally, the expression levels of cell markers and SLAMF7/STAT6 pathway were confirmed by histopathology examinations, RT-qPCR, and Western blot.

Acanthamoeba keratitis (AK) is a rare but severe corneal infection caused by the free-living amoeba, Acanthamoeba , which is ubiquitously present in the environment. This condition predominantly affects contact lens wearers but can also occur in non-lens users, particularly those exposed to contaminated water or with compromised immune systems. AK is characterized by progressive corneal inflammation, epithelial defects, and ulceration, which can lead to significant visual impairment or blindness if not promptly diagnosed and treated. This review aims to provide a comprehensive overview of AK by synthesizing current knowledge on its epidemiology, risk factors, pathophysiology, clinical manifestations, diagnostic approaches, and therapeutic strategies. The review also highlights preventive measures and public health strategies to reduce the incidence of this debilitating condition. A detailed examination of existing literature was conducted, focusing on the global incidence of AK, demographic trends, and various risk factors such as contact lens use, environmental exposures, and immunity status. The review also delves into the pathophysiology of Acanthamoeba infection, the host immune response, and the challenges in distinguishing AK from other forms of infectious keratitis. Therapeutic strategies, including medical and surgical interventions, are analyzed, along with emerging treatments. The global incidence of AK has increased, particularly among contact lens users, due to poor hygiene practices and environmental exposures. Early diagnosis remains challenging, often leading to delayed treatment and poorer outcomes. Biguanides and diamidines are the mainstays of medical therapy, with surgical options considered in advanced cases. Emerging therapies, such as photodynamic therapy and antimicrobial peptides, show promise in enhancing treatment outcomes. AK poses a significant threat to ocular health due to its potential for severe visual impairment and the complexities associated with its diagnosis and treatment. Early recognition, appropriate management, and public health initiatives focused on prevention are crucial for improving patient outcomes. Ongoing research and a collaborative approach among healthcare providers are essential to advancing the understanding and management of AK., Competing Interests: Conflicts of interest: In compliance with the ICMJE uniform disclosure form, all authors declare the following: Payment/services info: All authors have declared that no financial support was received from any organization for the submitted work. Financial relationships: All authors have declared that they have no financial relationships at present or within the previous three years with any organizations that might have an interest in the submitted work. Other relationships: All authors have declared that there are no other relationships or activities that could appear to have influenced the submitted work., (Copyright © 2024, Garg et al.)

Topic: To provide an overview on the incidence of Acanthamoeba keratitis (AK)., Clinical Relevance: Although being a sight-threatening cause of infectious keratitis, a comprehensive assessment of the incidence of AK is lacking., Methods: Incidence of AK was computed as the number of eyes with AK per health care center, per year (annualized center incidence [ACI]). Two meta-analytical ratios also were calculated: (1) the ratio of eyes with AK to the count of eyes with nonviral microbial keratitis (MK) and (2) the ratio of eyes with AK to the overall population (i.e., the total number of people in a nation or region, as indicated by the authors in each study). Center was defined as the health care facility where the study took place. Actual and projected estimates of the number of eyes with AK in years were calculated multiplying the ratio of eyes with AK to the total population and the corresponding population estimates, sourced from the United Nations Population Prospects., Results: Overall, 105 articles were included, published between 1987 and 2022. The total number of eyes identified was 91 951, with 5660 eyes affected by AK and 86 291 eyes affected by nonviral MK. The median ACI was 1.9 eyes with AK per health care center per year (95% confidence interval [CI], 1.5-2.6 eyes), with no statistically significant differences among continents. The ratio of eyes with AK to the total number of eyes with MK was 1.52% (95% CI, 1.03%-2.22%), whereas the ratio of eyes with AK in relationship to the entire population was estimated at 2.34 eyes per 1 000 000 people (95% CI, 0.98-5.55 per 1 000 000 people). The projected increase in the numbers of eyes with AK indicated an increase of 18.5% (n = 15 355 eyes with AK) in 2053 and 25.5% (n = 16 253 eyes with AK) in 2073, compared with the baseline of 2023 (n = 12 953 eyes with AK)., Discussion: Acanthamoeba keratitis emerged as a relatively low-incident disorder, and no significant differences in terms of its incidence were found among different continents., Financial Disclosure(s): Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article., (Copyright © 2024 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Purpose: To determine the prevalence and reasons for delays in diagnosis in patients with Acanthamoeba keratitis (AK) presenting to Wilmer Eye Institute, Baltimore, Maryland. Methods: This retrospective study analysed all patients with culture‐positive AK seen between 2012 and 2019 at a tertiary referral centre. Patient demographic information, clinical history, risk factors, symptom duration, referral patterns, slit lamp examination findings, visual acuity and need for surgery were collected. Results: The study included 45 eyes of 43 patients. On average, patients were symptomatic for 52.6 days before culture collection. Thirty‐one percent of patients were diagnosed within 28 days of symptom onset while 69% were diagnosed after 28 days. Before presentation to a tertiary care centre, 69% of patients were evaluated by an ophthalmologist outside of this institution and 27% were evaluated by a provider other than an ophthalmologist. AK was most commonly misdiagnosed as herpetic keratitis, occurring in 38% of patients. The strongest risk factor for AK was contact lens use. Only 11% of patients presented with the classic ring infiltrate and 82% had pain. Patients with an early versus late diagnosis had a mean Snellen visual acuity (VA) of 20/224 versus 20/296 at presentation (p = 0.33) and a mean Snellen VA of 20/91 versus 20/240 at final visit (p = 0.07). 11% of patients required a therapeutic penetrating keratoplasty. Conclusion: Delayed diagnosis of AK in our cohort occurred due to a misdiagnosis as herpetic keratitis, non‐specific clinical signs including the lack of pain in a number of patients, and a delay in referral to a tertiary care centre. Any contact lens wearer with an atypical keratitis should be referred promptly for Acanthamoeba cultures. [ABSTRACT FROM AUTHOR]

Purpose: To report our experience with oral miltefosine (MLT) as an adjunct treatment for progressive Acanthamoeba keratitis (AK). Methods: Retrospective case series of all patients who underwent treatment with oral MLT for AK at Bascom Palmer Eye Institute from 2017 to 2020. Results: Six females from 16 to 55 years old, with a microbiologic diagnosis of Acanthamoeba, were treated with MLT and standard medical treatment. Four of the six cases deteriorated after initiating treatment and three required a therapeutic keratoplasty. Two patients improved after 1 week of MLT and optical penetrating keratoplasty was performed after clinical resolution. Microbiologic culture of corneal buttons was negative in all cases. All corneal grafts remain clear at last follow-up with best-corrected visual acuity of 20/40 or better. Conclusion: Oral MLT may be a viable adjunctive therapy for recalcitrant AK; however, its use may be associated with a severe inflammatory reaction. Further studies are needed to evaluate its efficacy and variable clinical response. [ABSTRACT FROM AUTHOR]

Background: Acanthamoeba spp. are one of the free-living amoeba that spread worldwide causing keratitis. Owing to the increase in the use of lenses, whether for medical or cosmetic purposes, the incidence of disease increases every year. Contamination of the lenses with the Acanthamoeba trophozoites or cysts may lead to eye infection and cause sight-threatening keratitis in human. We isolated Acanthamoeba spp. from new lenses, used lenses, and contact lens disinfecting solutions and identified them based on morphological characteristics and molecular test. Methods: New and used lenses and contact lens disinfecting solutions were cultured on monogenic media. Light and scanning electron microscope was used to identify Acanthamoeba spp. morphological features. Genotype identification was also evaluated using PCR sequencing of 18S rRNA gene specific primer pair JDP1 and JDP2. Results: A hundred samples were examined, 29 (29%) were infected with Acanthamoeba spp. That belonged to two strains of Acanthamoeba (Acanthamoeba 41 and Acanthamoeba 68). 18S rRNA of the Acanthamoeba 41 had 99.69% sequence identity to Acanthamoeba castellanii clone HDU-JUMS-2, whereas Acanthamoeba 68 had 99.74% similar pattern to that of Acanthamoeba sp. isolate T4 clone ac2t4 that are morphologically identified as Acanthamoeba polyphaga. The obtained data revealed that the isolated strains belong to T4 genotype that was evolutionarily similar to strains isolated in Iran. Conclusions: Cosmetic lenses and disinfectant solutions are a major transmissible mode for infection. This genotype is common as the cause of Acanthamoeba keratitis. To avoid infection, care must be taken to clean the lenses and their preservative solutions and prevent contamination with the parasite. [ABSTRACT FROM AUTHOR]

Purpose: To determine the awareness regarding contact lens related ocular complications among opticians of Lahore and to find out the relationship between the awareness and their educational qualification. Study Design: Cross sectional survey. Place and Duration of Study: University of Lahore from April 2020 - December 2020. Methods: This survey included opticians of different areas of Lahore, Pakistan. Two hundred and eleven opticians responded to a pre-tested, structured close ended questionnaire. Data comprised of demographics and contact lens selling experience. Information regarding awareness about contact lens related complications was also sought. The data were analyzed using SPSS 25.0. Results: A total of 211 opticians participated in the survey. All subjects were males. Fifty four (25.6%) subjects were from the age group of 15 to 30 years, 122 (57.8%) were from the age group of 31 to 45 and 35 (16.6%) were from the age group of 46 to 60 years. Only 4.7% had contact lens dispensing diploma. Thirty seven percent patients had contact lens selling experience of 2 to 10 years. Similar percentage had more than 10 years of experience. Ten percent had less than one year of experience. Only 54 participants out of 211 had knowledge about dry eye, 9 knew about Acanthamoeba keratitis and Bacterial keratitis and only 2 were aware of corneal opacity. Conclusion: Majority of the opticians of Lahore are unaware of the contact lenses related complications. Sale of cosmetic lenses for fashion purpose should be discouraged and banned if not prescribed by optometrist/ ophthalmologist. [ABSTRACT FROM AUTHOR]

Background: Acanthamoebae are a causative agent of Acanthamoeba keratitis (AK) in immunocompetent individuals. Since access to propamidine isethionate (Brolene®) as a first-line treatment has been limited in recent years, in the current study, we examined the effects of pentamidine isethionate against trophozoite and cyst forms of Acanthamoeba. Methods: This experimental study was conducted in the Department of Medical Parasitology and Mycology, School of Public Health, Tehran University of Medical Sciences, Tehran, Iran, during 2019-2020. Pentamidine isethionate at concentrations of 50, 100, 200, 400, 600, 800, and 1000 µM were tested against trophozoites and cyst stages of T4 genotype, at 24- and 48-hour incubation period, and the viability was determined by trypan blue staining. In addition, the cytotoxic effect of the drug was examined in Vero cells using the 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Results: The 50% inhibitory concentration (IC50) of pentamidine isethionate on trophozoite after 24 and 48h were 97.4 µM and 60.99 µM. These results on cyst after 24 and 48h were 470 µM and 175.5 µM, respectively. In MTT assay, the drug showed an inhibitory effect on Vero cell growth with IC50 values of 115.4 µM and 87.42 µM after 24h and 48h, respectively. Conclusion: Pentamidine isethionate exhibited an inhibitory effect on trophozoite and cyst. Given that the trophozoicidal activity of the drug is in the safe dose, it could be suggested as an alternative in patients with AK; however, further investigation is needed in an animal model to confirm the data. [ABSTRACT FROM AUTHOR]

Acanthamoeba keratitis (AK), the vision-threatening disease caused by the amphizoic, potentially parasitic amoebae is growing threat for public health in Poland and worldwide. The report presents the case of 70-year-old man with severe keratitis admitted to an Ophthalmology Clinic. Before admission, the patient had been treated for 6 months with antibacterial and antifungal drugs in other units, without improvement in the eye condition. The use of in vivo confocal microscopy and in vitro cultivation allowed diagnosis to be verified and AK successfully treated. Awareness of the threat to public health caused by Acanthamoeba spp is still insufficient. If there is failure in response to first line therapy, AK should be taken into account,despite the lack of identified risk factors. In vitro monitoring of amoebic strain can be helpful for prognosis of the course of the corneal disease. Improvement in duration from first symptoms until proper diagnosis is decisive for better treatment efficacy. [ABSTRACT FROM AUTHOR]

This study aimed to develop a miltefosine-eluting contact lens (MLF-CL) device that would allow sustained and localized miltefosine release for the treatment of Acanthamoeba keratitis. MLF-CLs were produced in three different miltefosine doses by solvent-casting a thin miltefosine-polymer film around the periphery of a methafilcon hydrogel, which was then lathed into a contact lens. During seven days of in vitro testing, all three formulations demonstrated sustained release from the lens at theoretically therapeutic levels. Based on the physicochemical characterization of MLF-CLs, MLF-CL’s physical properties are not significantly different from commercial contact lenses in terms of light transmittance, water content and wettability. MLF-CLs possessed a slight reduction in compression modulus that was attributed to the inclusion of polymer-drug films but still remain within the optimal range of soft contact lenses. In cytotoxicity studies, MLF-CL indicated up to 91% viability, which decreased proportionally as miltefosine loading increased. A three-day biocompatibility test on New Zealand White rabbits revealed no impact of MLF-CLs on the corneal tissue. The MLF-CLs provided sustained in vitro release of miltefosine for a week while maintaining comparable physical features to a commercial contact lens. MLF-CL has a promising potential to be used as a successful treatment method for Acanthamoeba keratitis.

Acanthamoeba spp. are among the most worldwide prevalent protozoa. It is the causative agent of a disease known as Acanthamoeba keratitis, a painful and severe sight-threatening corneal infection that can lead to blindness. In recent years, the prevalence of Acanthamoeba keratitis has rapidly increased, growing its importance to human health. This systematic review aims to assess the distribution of Acanthamoeba sp. genotypes causing keratitis around the world, considering the sample collected type and the used identification method. Most of the cases were found in Asia and Europe. Not surprisingly, the T4 genotype was the most prevalent worldwide, followed by T3, T15, T11, and T5. Furthermore, the T4 genotype contains a higher number of species. Given the differences in pathology, susceptibility to treatment, and clinical outcome between distinct genotypes, it is essential to genotype isolates from Acanthamoeba keratitis cases to help to establish a better correlation between in vitro and in vivo activities, resulting in better drug therapies and successful treatment in cases of this important ocular infection. [ABSTRACT FROM AUTHOR]

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Efficacious treatments against Acanthamoeba Keratitis (AK) is challenging, often ineffective and linked to the intragenotype variation in the drug efficacy. Increased oxygen can facilitate host response and can inhibit some organisms. Herein, we report the effect of increased oxygen concentrations on Acanthamoeba spp. growth and its effect on ROS (reactive oxygen species) production. The exposition to pure oxygen could reduce cell growth by at least 60% for Acanthamoeba castellanii Neff, Acanthamoeba polyphaga, and Acanthamoeba griffini. The increase in ROS production confirming that oxygen cell's growth inhibition was due to oxidative stress. Further studies are needed to determine oxygen saturation level, time of oxygen exposition, and number of sessions needed to eliminate the parasite. [ABSTRACT FROM AUTHOR]

Purpose: In cases of corneal opacity with vascularization and peripheral thinning, traditional keratoplasty techniques have several risks and drawbacks. We report the results of a two-step surgical strategy consisting in performing a large diameter tectonic lamellar keratoplasty (TLK) to restore appropriate corneal thickness and an avascular recipient bed, followed by central optical PK within the lamellar graft at a later date. Methods: This single-institution study analyzes the results of 7 eyes of 7 patients who received PK after large diameter TLK. All patients were affected by deep post-infectious corneal opacity with persistent stromal vascularization and peripheral thinning. The main outcomes measured were graft survival, visual acuity, refraction, and endothelial cell density. Results: TLK was performed in all cases with 10/10.1-mm diameters. After a mean interval of 14 months, central PK was performed with a median host-graft diameter of 7.75/8.25 mm. Mean follow-up after PK was 52 months. At last follow-up, 6/7 (86%) grafts were clear. Endothelial rejection occurred in 5/7 (71%) eyes, with one patient having multiple episodes and subsequent graft failure. At 2 years, all patients had a visual acuity ≥ 20/40, with an average refractive astigmatism of 3.75 diopters. Conclusion: Optical PK within a previous TLK is a safe and efficient technique for treating deep corneal opacity associated with extensive vascularization and peripheral thinning. Extended follow-up period is necessary to assess endothelial cell loss and long-term efficacy of the procedure. [ABSTRACT FROM AUTHOR]

[Display omitted] Although rare, amoebic keratitis (AK) is a disease caused by Acanthamoeba spp. that can lead to blindness. The drugs currently available for its treatment are very toxic, which has motivated the investigation for more effective and safe therapeutic options. In this study, the in vitro activity of ß-caryophyllene (BCP) was exploited taking into account its action against other protozoans as well as its well-known healing and anti-inflammatory properties (aspects relevant for the AK pathogenesis). On the other hand, high volatilization and oxidation phenomena are found for this compound, which led to its incorporation into nanoemulsions (NEs). Two emulsifying agents were tested, resulting in monodisperse systems with reduced droplet size (<265 nm) and high surface charge (positive and negative for NEs prepared with cetrimonium bromide −CTAB and Phosal® 50+, respectively). NEs prepared with CTAB were shown to be more stable after long-term storage at 4 and 25 °C than those prepared with Phosal®. Pure BCP, at the highest concentration (500 µM), resulted in a level of inhibition of Acanthamoeba trophozoites equivalent to that of reference drug (chlorhexidine). This activity was even greater after oil nanoencapsulation. The reduced droplet size could improve the interaction of the oil with the microorganism, justifying this finding. Changes in surface charge did not impact the activity. Positively charged NEs improved the interaction and retention of BCP in the cornea and thus should be prioritized for further studies. [ABSTRACT FROM AUTHOR]

Acanthamoeba castellanii , a free-living amoeba, can be pathogenic to humans causing a corneal infection named Acanthamoeba keratitis (AK). The mannose-binding protein (MBP) is well established as the major factor related to Acanthamoeba pathogenesis. However, additional factors that participate in the adhesion process and protect trophozoites from cytolytic effects caused by host immune responses remain unknown. Ectonucleotidases, including 3′-nucleotidase/nuclease (3′-NT/NU), a bifunctional enzyme that was recently reported in A. castellanii , are frequently related to the establishment of parasitic infections. We verified that trophozoites can hydrolyze 3′-AMP, and this activity is similar to that observed in other protists. The addition of 3′-AMP increases the adhesion of trophozoites to LLC-MK2 epithelial cells, and this stimulation is completely reversed by DTT, an inhibitor of ecto-3′-nucleotidase activity. Lesions in corneal cells caused by AK infection may elevate the extracellular level of 3′-AMP. We believe that ecto-3′-nucleotidase activity can modulate the host immune response, thus facilitating the establishment of parasitic infection. This activity results from the generation of extracellular adenosine, which can bind to purinergic receptors present in host immune cells. Positive feedback may occur in this cascade of events once the ecto-3′-nucleotidase activity of trophozoites is increased by the adhesion of trophozoites to LLC-MK2 cells. [ABSTRACT FROM AUTHOR]

To report clinical findings and prognostic factors for visual and morphological outcomes in patients with Acanthamoeba keratitis (AK). Single-center, retrospective, longitudinal study of 51 cases of AK diagnosed by real-time polymerase chain reaction (RT-PCR) between March 2010 and October 2022. The primary outcome was the final best corrected visual acuity (BCVA). Poor visual outcome was defined as a final BCVA ≥ 1 logMAR unit, while good visual outcome was defined as a final BCVA < 1 logMAR unit. Eyes from these two groups were compared, regarding demographic and initial clinical variables, anti- Acanthamoeba treatment used, and complications of the disease. Early diagnosis was defined as ≤ 14 days from symptom onset to diagnostic confirmation and initiation of Acanthamoeba medical treatment. Multivariable logistic regression was used to determine predictors of poor visual outcome. A total of 51 eyes from 46 patients diagnosed with AK, all contact lens (CL) wearers, were included in this study. Average follow-up was 39.0 ± 30.2 [total range 14–120] months. Thirty-one eyes (60.8 %) presented good visual outcome, with a lower baseline age (30.5 ± 9.0 vs. 42.3 ± 15.8; p = 0.020), better initial BCVA (0.8 ± 0.7 logMAR units vs. 1.3 ± 0.9 logMAR units; p = 0.047), higher rate of early diagnosis (45.2 % vs. 5.6 %; p = 0.004), and higher rate of therapeutic epithelial debridement (64.5 % vs. 10 %; p < 0.001). 20 eyes (39.2 %) presented poor visual outcome, with 12 eyes undergoing evisceration/enucleation (23.5 %). These 20 eyes presented a higher rate of complications (90 % vs. 61.3 %; p = 0.031). In multivariable analysis, early diagnosis of AK (OR 19.78; 95 % CI 2.07–189.11; p = 0.010) and therapeutic epithelial debridement (OR 19.02; 95 % CI 3.27–110.57; p = 0.001) were associated with a good visual outcome. In the present study, poor visual outcome was present in 39 % of affected eyes. Early AK diagnosis (≤14 days from symptom onset) and therapeutic epithelial debridement were associated with good final visual outcome. [ABSTRACT FROM AUTHOR]

PurposeTo determine risk factors and clinical signs that may differentiate between bacterial, fungal, and acanthamoeba keratitis among patients presenting with presumed infectious keratitis.DesignHospital-based cross-sectional study.MethodsWe examined the medical records of 115 patients with laboratory-proven bacterial keratitis, 115 patients with laboratory-proven fungal keratitis, and 115 patients with laboratory-proven acanthamoeba keratitis seen at Aravind Eye Hospital, Madurai, India, from 2006-2011. Risk factors and clinical features of the 3 organisms were compared using multinomial logistic regression.ResultsOf 95 patients with bacterial keratitis, 103 patients with fungal keratitis, and 93 patients with acanthamoeba keratitis who had medical records available for review, 287 (99%) did not wear contact lenses. Differentiating features were more common for acanthamoeba keratitis than for bacterial or fungal keratitis. Compared to patients with bacterial or fungal keratitis, patients with acanthamoeba keratitis were more likely to be younger and to have a longer duration of symptoms, and to have a ring infiltrate or disease confined to the epithelium.ConclusionsRisk factors and clinical examination findings can be useful for differentiating acanthamoeba keratitis from bacterial and fungal keratitis.