Acanthamoeba keratitis - A review.

This is a comprehensive review after a thorough literature search in PubMed-indexed journals, incorporating current information on the pathophysiology, clinical features, diagnosis, medical and surgical therapy, as well as outcomes of Acanthamoeba keratitis (AK). AK is a significant cause of ocular morbidity, and early diagnosis with timely institution of appropriate therapy is the key to obtaining good outcomes. The varied presentations result in frequent misdiagnosis, and co-infections can increase the morbidity of the disease. The first line of therapy continues to be biguanides and diamidines, with surgery as a last resort.

Comparison of therapeutic effects between big-bubble deep anterior lamellar keratoplasty and penetrating keratoplasty for medically unresponsive Acanthamoeba keratitis.

PURPOSE: To compare the outcomes of big-bubble deep anterior lamellar keratoplasty (BB-DALK) and penetrating keratoplasty (PKP) in the management of medically unresponsive Acanthamoeba keratitis (AK). METHODS: This retrospective study included 27 eyes of BB-DALK and 24 eyes of PKP from a tertiary ophthalmology care centre. Glucocorticoid eye drops were subsequently added to the treatment plan 2 months postoperatively based on the evaluation using confocal laser scanning microscopy. The clinical presentations, best-corrected visual acuity (BCVA), postoperative refractive outcomes, graft survival, and Acanthamoeba recurrence were analyzed. RESULTS: The AK patients included in the study were in stage 2 or stage 3, and the percentage of patients in stage 3 was higher in the PKP group (P = 0.003). Clinical presentations were mainly corneal ulcers and ring infiltrates, and endothelial plaques, hypopyon, uveitis and glaucoma were more common in the PKP group (P = 0.007). The BCVA and the graft survival rate showed no statistically significant differences between the two groups at 1 year after surgery. However, 3 years postoperatively, the BCVA of 0.71 ± 0.64 logMAR, the graft survival rate of 89.5%, and the endothelial cell density of 1899 ± 125 cells per square millimeter in the BB-DALK group were significantly better than those of the PKP group (P = 0.010, 0.046, and 0.032, respectively). 3 eyes (11.1%) in the BB-DALK group and 2 eyes (8.3%) in the PKP group experienced Acanthamoeba recurrence, but the rates showed no statistically significant difference between the two groups (P = 1.000). In the PKP group, immune rejection and elevated intraocular pressure were observed in 5 and 6 eyes, respectively. CONCLUSION: Corneal transplantation is recommended for AK patients unresponsive to antiamoebic agents. The visual acuity and graft survival can be maintained after BB-DALK surgery. Acanthamoeba recurrence is not related to the surgical approach performed, whereas complete dissection of the infected corneal stroma and delayed prescribing of glucocorticoid eye drops were important to prevent recurrence.

Differentiation of acanthamoeba keratitis from other non-acanthamoeba keratitis: Risk factors and clinical features.

INTRODUCTION: Infectious Keratitis is one of the most common ocular emergencies seen by ophthalmologists. Our aim is to identify the risk factors and clinical features of Acanthamoeba Keratitis (AK). METHODS: This retrospective chart review study was conducted at King Khaled Eye Specialist Hospital, Riyadh, Saudi Arabia, and included all the microbial keratitis cases, male and female patients of all ages. The main outcome is the differentiation between various microbial keratitis types. RESULTS: We included 134 consecutive eyes of 126 persons. We had 24 cases of acanthamoeba keratitis, 22 bacterial keratitis, 24 fungal keratitis, 32 herpetic keratitis, and 32 bacterial co-infection. Contact lens wear was found in 33 eyes (24.6%). Among acanthamoeba keratitis patients, 73% were ≤ 39 years of age, and 73% were females (P <0.001). Also, in AK cases, epithelial defect was found in all cases (100%), endothelial plaques were found in 18 eyes (69.2%), 12 cases had radial keratoneuritis (46.2%), and ring infiltrate was found in 53.8% of AK cases. CONCLUSIONS: We determined the factors that increase the risk of acanthamoeba infection and the clinical characteristics that help distinguish it from other types of microbial keratitis. Our findings suggest that younger females and patients who wear contact lenses are more likely to develop acanthamoeba keratitis. The occurrence of epitheliopathy, ring infiltrate, radial keratoneuritis, and endothelial plaques indicate the possibility of acanthamoeba infection. Promoting education on wearing contact lenses is essential to reduce the risk of acanthamoeba infection, as it is the most significant risk factor for this infection.

Acanthamoeba Keratitis in China: Genotypic and Clinical Correlations.

Purpose: To investigate the relationship between Acanthamoeba genotypes, clinical manifestations, and outcomes in Acanthamoeba keratitis (AK) patients. Methods: This retrospective study included 159 culture-confirmed AK patients. Patients' data were collected, including demographics, initial diagnosis, treatments, and clinical features. The genotype of Acanthamoeba was identified through sequencing the Diagnostic Fragment 3 (DF3) region in the small ribosomal subunit RNA genes. The phylogenetic tree was constructed using the ClustalW model and maximum likelihood method. Cases with "poor outcome" were defined based on specific clinical criteria, including corneal perforation, keratoplasty, other eye surgery, duration of anti-amoebic therapy ≥8.0 months, and final visual acuity ≤20/80. "Better outcome" cases were the remainder. The correlation between T4 subtypes, clinical phenotypes, and clinical prognosis were further analyzed. Results: In this study, AK was primarily attributed to the T4A genotype, with a positive correlation between geographical and genetic distances. The primary clinical associated with T4 subtypes was deep stromal infiltration. Results was also showed a significant association between T4 subtypes and clinical outcomes (P = 0.021). Further analysis revealed that T4C was closely associated with a better prognosis (P = 0.040) and T4D with worse outcomes (P = 0.013). Conclusions: In China, AK was predominantly caused by the T4A subtype. Geographical distance positively correlated with genetic distance. Clinical prognosis varied among different subtypes, notably in T4C and T4D. Translational Relevance: This study demonstrated the association between T4 subtypes and clinical phenotypes, as well as the effects of T4 subtypes on clinical prognosis.

Use of In Vivo Corneal Confocal Microscopy to Guide Excimer Laser With Adjunctive Mitomycin C for Treatment of Recalcitrant Acanthamoeba Keratitis.

PURPOSE: The aim of this study was to describe 3 cases of recalcitrant Acanthamoeba keratitis (AK) that were successfully treated using in vivo corneal confocal microscopy (IVCM) to guide excimer laser ablation depth with adjunctive mitomycin C 0.02%. METHODS: Three patients diagnosed with AK did not respond to several weeks of intensive topical therapy with antiamoebic agents. The patient underwent phototherapeutic keratectomy with topical mitomycin C 0.02% application. The maximum stromal depth of cysts measured by IVCM was 80 µm, 100 µm, and 240 µm, and the stromal ablation depths were 80 µm, 100 µm, and 100 µm, respectively. RESULTS: In all 3 eyes, AK resolved after a single excimer laser application, and topical treatment was gradually discontinued within 6 weeks afterward. In 1 eye, penetrating corneal transplantation was performed 6 weeks after phototherapeutic keratectomy because of ongoing severe corneal pain. IVCM and histology of the corneal transplant did not reveal any Acanthamoeba cysts within the excised corneal button. No recurrence was observed during the follow-up period of 19 to 34 months. CONCLUSIONS: IVCM-guided phototherapeutic keratectomy with mitomycin C 0.02% seems to be a safe and successful approach for the treatment of AK, especially in cases of resistance to topical treatment. Corneal IVCM should be performed before laser application to measure cyst depth, determine ablation depth, and assess postoperative treatment success.

Clinical outcomes and prognostic factors in Acanthamoeba keratitis.

PURPOSE: To report clinical findings and prognostic factors for visual and morphological outcomes in patients with Acanthamoeba keratitis (AK). METHODS: 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. RESULTS: 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. CONCLUSIONS: 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.

The role of naturally acquired intracellular Pseudomonas aeruginosa in the development of Acanthamoeba keratitis in an animal model.

BACKGROUND: Acanthamoeba is an environmental host for various microorganisms. Acanthamoeba is also becoming an increasingly important pathogen as a cause of keratitis. In Acanthamoeba keratitis (AK), coinfections involving pathogenic bacteria have been reported, potentially attributed to the carriage of microbes by Acanthamoeba. This study assessed the presence of intracellular bacteria in Acanthamoeba species recovered from domestic tap water and corneas of two different AK patients and examined the impact of naturally occurring intracellular bacteria within Acanthamoeba on the severity of corneal infections in rats. METHODOLOGY/PRINCIPAL FINDINGS: Household water and corneal swabs were collected from AK patients. Acanthamoeba strains and genotypes were confirmed by sequencing. Acanthamoeba isolates were assessed for the presence of intracellular bacteria using sequencing, fluorescence in situ hybridization (FISH), and electron microscopy. The viability of the bacteria in Acanthamoeba was assessed by labelling with alkyne-functionalized D-alanine (alkDala). Primary human macrophages were used to compare the intracellular survival and replication of the endosymbiotic Pseudomonas aeruginosa and a wild type strain. Eyes of rats were challenged intrastromally with Acanthamoeba containing or devoid of P. aeruginosa and evaluated for the clinical response. Domestic water and corneal swabs were positive for Acanthamoeba. Both strains belonged to genotype T4F. One of the Acanthamoeba isolates harboured P. aeruginosa which was seen throughout the Acanthamoeba's cytoplasm. It was metabolically active and could be seen undergoing binary fission. This motile strain was able to replicate in macrophage to a greater degree than strain PAO1 (p<0.05). Inoculation of Acanthamoeba containing the intracellular P. aeruginosa in rats eyes resulted in a severe keratitis with increased neutrophil response. Acanthamoeba alone induced milder keratitis. CONCLUSIONS/SIGNIFICANCE: Our findings indicate the presence of live intracellular bacteria in Acanthamoeba can increase the severity of acute keratitis in vivo. As P. aeruginosa is a common cause of keratitis, this may indicate the potential for these intracellular bacteria in Acanthamoeba to lead to severe polymicrobial keratitis.

Oxidase enzyme genes are differentially expressed during Acanthamoeba castellanii encystment.

Acanthamoeba castellanii, a ubiquitous protozoan, is responsible for significant diseases such as Acanthamoeba keratitis and granulomatous amoebic encephalitis. A crucial survival strategy of A. castellanii involves the formation of highly resistant cysts during adverse conditions. This study delves into the cellular processes underpinning encystment, focusing on gene expression changes related to reactive oxygen species (ROS) balance, with a particular emphasis on mitochondrial processes. Our findings reveal a dynamic response within the mitochondria during encystment, with the downregulation of key enzymes involved in oxidative phosphorylation (COX, AOX, and NADHalt) during the initial 48 h, followed by their overexpression at 72 h. This orchestrated response likely creates a pro-oxidative environment, facilitating encystment. Analysis of other ROS processing enzymes across the cell reveals differential expression patterns. Notably, antioxidant enzymes, such as catalases, glutaredoxins, glutathione S-transferases, peroxiredoxins, and thioredoxins, mirror the mitochondrial trend of downregulation followed by upregulation. Additionally, glycolysis and gluconeogenesis are downregulated during the early stages in order to potentially balance the metabolic requirement of the cyst. Our study underscores the importance of ROS regulation in Acanthamoeba encystment. Understanding these mechanisms offers insights into infection control and identifies potential therapeutic targets. This work contributes to unraveling the complex biology of A. castellanii and may aid in combatting Acanthamoeba-related infections. Further research into ROS and oxidase enzymes is warranted, given the organism's remarkable respiratory versatility.

Advances in Contact Lens Care Solutions: PVP-I Disinfectant and HAD Wetting Agents From Japan.

ABSTRACT: Half of the individuals who wear contact lenses use reusable lenses that require proper care. Improper contact lens (CL) care and using inadequate disinfecting solutions can lead to lens contamination, CL-related microbial keratitis, and Acanthamoeba keratitis. Oxidative disinfecting solutions, such as hydrogen peroxide, show higher efficacy than multipurpose solutions. Povidone-iodine (PVP-I), an oxidative disinfectant used in ophthalmic surgery, has been proven to be safe and effective. The PVP-I system, a CL disinfecting solution developed in Japan, has demonstrated excellent antimicrobial and antiviral properties. Although CL discomfort does not have a risk of ocular disorders with poor visual prognosis, such as keratitis, CL discomfort can still lead to lens dropout and thus needs to be addressed. To mitigate CL discomfort, it is essential to use disinfecting solutions containing surfactants and wetting agents that improve wettability of the lens surface. A CL solution containing hyaluronic acid derivatives (HADs) as wetting agents that permanently adhere to the lens surface to improve wettability of the lens surface was developed in Japan. There is potential for HAD to be integrated into various solutions. This article reviews the efficacy of novel PVP-I-based disinfecting solution and HAD wetting agents.

The Orphan Drug for Acanthamoeba Keratitis (ODAK) Trial: PHMB 0.08% (Polihexanide) and Placebo versus PHMB 0.02% and Propamidine 0.1.

PURPOSE: To compare topical PHMB (polihexanide) 0.02% (0.2 mg/ml)+ propamidine 0.1% (1 mg/ml) with PHMB 0.08% (0.8 mg/ml)+ placebo (PHMB 0.08%) for Acanthamoeba keratitis (AK) treatment. DESIGN: Prospective, randomized, double-masked, active-controlled, multicenter phase 3 study (ClinicalTrials.gov identifier, NCT03274895). PARTICIPANTS: One hundred thirty-five patients treated at 6 European centers. METHODS: Principal inclusion criteria were 12 years of age or older and in vivo confocal microscopy with clinical findings consistent with AK. Also included were participants with concurrent bacterial keratitis who were using topical steroids and antiviral and antifungal drugs before randomization. Principal exclusion criteria were concurrent herpes or fungal keratitis and use of antiamebic therapy (AAT). Patients were randomized 1:1 using a computer-generated block size of 4. This was a superiority trial having a predefined noninferiority margin. The sample size of 130 participants gave approximately 80% power to detect 20-percentage point superiority for PHMB 0.08% for the primary outcome of the medical cure rate (MCR; without surgery or change of AAT) within 12 months, cure defined by clinical criteria 90 days after discontinuing anti-inflammatory agents and AAT. A prespecified multivariable analysis adjusted for baseline imbalances in risk factors affecting outcomes. MAIN OUTCOME MEASURES: The main outcome measure was MCR within 12 months, with secondary outcomes including best-corrected visual acuity and treatment failure rates. Safety outcomes included adverse event rates. RESULTS: One hundred thirty-five participants were randomized, providing 127 in the full-analysis subset (61 receiving PHMB 0.02%+ propamidine and 66 receiving PHMB 0.08%) and 134 in the safety analysis subset. The adjusted MCR within 12 months was 86.6% (unadjusted, 88.5%) for PHMB 0.02%+ propamidine and 86.7% (unadjusted, 84.9%) for PHMB 0.08%; the noninferiority requirement for PHMB 0.08% was met (adjusted difference, 0.1 percentage points; lower one-sided 95% confidence limit, -8.3 percentage points). Secondary outcomes were similar for both treatments and were not analyzed statistically: median best-corrected visual acuity of 20/20 and an overall treatment failure rate of 17 of 127 patients (13.4%), of whom 8 of 127 patients (6.3%) required therapeutic keratoplasty. No serious drug-related adverse events occurred. CONCLUSIONS: PHMB 0.08% monotherapy may be as effective (or at worse only 8 percentage points less effective) as dual therapy with PHMB 0.02%+ propamidine (a widely used therapy) with medical cure rates of more than 86%, when used with the trial treatment delivery protocol in populations with AK with similar disease severity. FINANCIAL DISCLOSURE(S): Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.

Diagnosing and monitoring the characteristics of Acanthamoeba keratitis using slit scanning and laser scanning in vivo confocal microscopy.

INTRODUCTION: Acanthamoeba keratitis (AK) is a serious and potentially blinding ocular infection caused by the free-living amoeba, Acanthamoeba. In vivo confocal microscopy (IVCM) is a non-invasive device which has been proven of great use to diagnose Acanthamoeba infections immediately. The aim of this review was to establish different patterns and signs of AK that appear on the IVCM both before and after treatment. METHODS: A systematic review of the literature from 1974 until September 2021 was performed using Embase and PubMed, following The Preferred Reporting Items for Systematic reviews and Meta-Analyses (PRISMA) guidelines. RESULTS: Twenty different signs of AK were observed using IVCM. The included studies used vastly different criteria to diagnose infections, ranging from just 1 to 13 of the signs, demonstrating the current lack of a standardised diagnosis of this infection using the IVCM. The appearance of double wall cysts, trophozoites, signet rings, target signs and clusters were shown to be pathognomonic to AK infections. Bright spots located in the corneal epithelium were demonstrated as non-reliable predictors of AK. The presence of cysts in clusters and single file can predict the need for corneal transplantation. The morphological changes in cysts using the IVCM following treatment were described as breaking down to hollow forms and occasionally surrounded by black cavities. Using this information, a visual guideline for identifying AK signs in diagnosis and follow-up using IVCM was created. CONCLUSION: Increased awareness of the different signs and patterns of AK that appear on the IVCM is crucial in order to correctly identify an infection and increase the potential of this device. Our guidelines presented here can be used, but further studies are needed in order to determine the relationship and aetiology of these signs and cellular changes on the IVCM both before and after anti-amoeba treatment.

Depth, size of infiltrate, and the microbe - The trio that prognosticates the outcome of infective keratitis.

PURPOSE: To analyze the influence of infiltrate size, depth, and organism on the outcome of microbial keratitis. DESIGN: Retrospective comparative study. METHODS: Medical records of patients with infective keratitis, who reported from January 2015 to December 2019 to a tertiary eye care center, were analyzed. Size and depth of ulcer at presentation were the factors used to group patients, and the influence on the outcome of the organism causing it was analyzed. Grouping was as follows: group A: ulcer size <6 mm/anterior to midstromal infiltrate, group B: ulcer < 6 mm/full-thickness infiltrate, group C: ulcer >6 mm/anterior to midstromal infiltrate, group D: ulcer > 6 mm/full-thickness infiltrate. Patients with viral keratitis or unidentified organism were excluded. Response to treatment and best-corrected visual acuity (BCVA) at the final follow-up were the outcome measures. RESULTS: In the study, 1117/6276 patients were included, with 60.8% patients in group A. A significant improvement in visual acuity was noted in groups A/B compared to groups C/D. Group A had the best response to medical management, irrespective of the organism. Higher risk for surgery was noted in group C compared to group B, with group A as the reference. Overall resolution with medical treatment was noted in 70% miscellaneous keratitis, 64.8% bacterial keratitis, 64.3% mixed keratitis, 62.5% acanthamoeba keratitis, 52.6% fungal keratitis, and 12.1% Pythium keratitis. Bacteria and acanthamoeba responded better to medical management than fungal keratitis, whereas Pythium had the highest risk for surgery. CONCLUSION: An interplay between virulence of the organism along with depth and size of the infiltrate determines the outcome of microbial keratitis.

Incidence of Acanthamoeba Keratitis in the United Kingdom in 2015: A Prospective National Survey.

PURPOSE: The aims of this study were to quantify the incidence of Acanthamoeba keratitis (AK) in the United Kingdom and investigate risk factors and management parameters. METHODS: This was a prospective population-based study from January to December 2015 through the British Ophthalmic Surveillance Unit. Data were collected on demographics, clinical features, and management. Incidence rates were calculated from estimates of population and contact lens (CL) user numbers. Statistical analysis compared annualized incidences per million and altered risk ratios for AK with the England and Wales 24 months 1997/1998 to 1998/1999 study. RESULTS: The study identified 124 AK cases, an overall incidence of 2.35 per million. CL wearers accounted for 108 of 124 cases (87%), in whom the AK incidence was 26.94 per million. Herpes keratitis was initially misdiagnosed in 25 of 124 cases (20.2%). The highest incidence of AK was among planned replacement soft CL (PRSCL) wearers (50.65 per million), 7-fold greater than for daily disposable CL (DDSCL) users (7.24 per million). There was a significant increase in AK incidence ( P < 0.001) compared with both 1997/1998 [risk ratio 1.92, 95% confidence interval (CI) 1.38-2.66] and 1998/1999 (risk ratio 2.13, 95% CI 1.52-2.98) together with a higher incidence per million CL users of 26.94 versus 21.14 (1997/1998) and 17.53 (1998/1999). CONCLUSIONS: This study provides the first published data on the nationwide incidence of AK in the United Kingdom. The findings confirm an increasing incidence of AK, particularly among contact lens wearers since 1997/1998 to 1998/1999. PRSCLs were identified as a significant risk factor compared with DDSCLs. Misdiagnosis and treatment delays remain an ongoing problem for patients with AK.

Cell death of Acanthamoeba castellanii following exposure to antimicrobial agents commonly included in contact lens disinfecting solutions.

Several antimicrobial agents are commonly included in contact lens disinfectant solutions including chlorhexidine diacetate (CHX), polyhexamethylene biguanide (PHMB) or myristamidopropyl dimethylamine (MAPD); however, their mode of action, i.e. necrosis versus apoptosis is incompletely understood. Here, we determined whether a mechanism of cell death resembling that of apoptosis was present in Acanthamoeba castellanii of the T4 genotype (NEFF) following exposure to the aforementioned antimicrobials using the anticoagulant annexin V that undergoes rapid high affinity binding to phosphatidylserine in the presence of calcium, making it a sensitive probe for phosphatidylserine exposure. The results revealed that under the conditions employed in this study, an apoptotic pathway of cell death in this organism at the tested conditions does not occur. Our findings suggest that necrosis is the likely mode of action; however, future mechanistic studies should be accomplished in additional experimental conditions to further comprehend the molecular mechanisms of cell death in Acanthamoeba.

Proteases of Acanthamoeba.

The members of genus Acanthamoeba are the etiological agent of uncommon but severe or even fatal opportunistic infections in human beings. The presence of different classes of intracellular and extracellular proteases including serine proteases, cysteine proteases, and metalloproteases has been well documented in environmental and clinical isolates of Acanthamoeba spp. However, the role of the proteolytic enzymes in physiological, biological, and pathological mechanisms of the amoeba remains partially investigated. Some attempts have been conducted using various methods to determine the profile of proteases (number, class, optimal conditions, and activity of the enzymes), and possible pathogenicity mechanism of the proteolytic enzymes (various protein substrate degradation, cytopathic effect on different cell lines). In some cases, it was attempted to correlate intracellular and extracellular protease profile with pathogenicity potential of strains. This review revealed that the protease profile of different strains of Acanthamoeba was extremely complex, therefore, further comprehensive studies with application of a combination of various methods may help to elucidate the role of the enzymes.

Steroids in the Management of Infectious Keratitis.

PURPOSE: To summarize the evidence base on the use of topical corticosteroids for infectious keratitis. METHODS: Narrative review. RESULTS: Infectious keratitis is a painful condition that often results in visually significant corneal stromal scarring, even when antimicrobial therapy is successful. Corticosteroids may reduce inflammation and subsequent scar formation and while relieving the acute ocular pain associated with a corneal ulcer. However, corticosteroids also reduce the host immune response, which could hinder the ability to clear infection. The safety and effectiveness of corticosteroids depends to a large part on the efficacy of the antimicrobials being used to treat the underlying infection. Randomized trials have found that corticosteroids are safe and effective for herpetic keratitis when used with appropriate antiviral therapy, and are safe for bacterial keratitis when used with broad spectrum topical antibiotics. The effectiveness of corticosteroids for bacterial keratitis has not been shown conclusively, although more advanced bacterial corneal ulcers may do better with corticosteroids. No randomized trials have assessed the safety and effectiveness of steroids for fungal or acanthamoeba keratitis. Animal studies suggest corticosteroids may be harmful in fungal keratitis, and observational human studies have found that steroids are harmful for fungal and acanthamoeba keratitis when started prior to anti-amoebics. CONCLUSIONS: Topical corticosteroids, when used as an adjunct to antimicrobial therapy, may be beneficial if the antimicrobial being used can effectively clear or suppress the infection, such as in bacterial and herpetic keratitis. Randomized trials would be helpful to further delineate the role of corticosteroids for infectious keratitis.

Acanthamoeba castellanii trophozoites escape killing by neutrophil extracellular traps using their 3'-nucleotidase/nuclease activity.

Acanthamoeba castellanii is a free-living amoeba that acts as an opportunistic pathogen for humans and is the pathogenic agent of Acanthamoeba keratitis (AK). A. castellanii may present as proliferative and infective trophozoites or as resistant cysts during their life cycle. The immune response against AK is still poorly explored; however, it is well established that macrophages and neutrophils play essential roles in controlling corneal infection during the disease outcome. The release of NETs is one of the innate immune strategies to prevent parasite infection, especially when neutrophils interact with microorganisms that are too large to be phagocytosed, which is the case for amoeba species. The present work demonstrated that A. castellanii trophozoites can trigger NET formation upon in vitro interaction with neutrophils. Using DNase as a control, we observed increased parasite survival after coinciding with neutrophils, which may be correlated with NET degradation. Indeed, A. castellanii trophozoites degrade the NET DNA scaffold. Molecular analysis confirmed the occurrence of a 3'-nucleotidase/nuclease (3'-NT/NU) in the A. castellanii genome. We also demonstrated that trophozoites exhibit significantly higher 3'-NT/NU activity than cysts, which cannot trigger NET release. Considering that previous studies indicated the pathological role of 3'-NT-/NU in parasite infection, we suggest that this enzyme may act as the mechanism of escape of A. castellanii trophozoites from NETs.

AI-Based Decision-Support System for Diagnosing Acanthamoeba Keratitis Using In Vivo Confocal Microscopy Images.

Purpose: In vivo confocal microscopy (IVCM) of the cornea is a valuable tool for clinical assessment of the cornea but does not provide stand-alone diagnostic support. The aim of this work was to develop an artificial intelligence (AI)-based decision-support system (DSS) for automated diagnosis of Acanthamoeba keratitis (AK) using IVCM images. Methods: The automated workflow for the AI-based DSS was defined and implemented using deep learning models, image processing techniques, rule-based decisions, and valuable input from domain experts. The models were evaluated with 5-fold-cross validation on a dataset of 85 patients (47,734 IVCM images from healthy, AK, and other disease cases) collected at a single eye clinic in Sweden. The developed DSS was validated on an additional 26 patients (21,236 images). Results: Overall, the DSS uses as input raw unprocessed IVCM image data, successfully separates artefacts from true images (93% accuracy), then classifies the remaining images by their corneal layer (90% accuracy). The DSS subsequently predicts if the cornea is healthy or diseased (95% model accuracy). In disease cases, the DSS detects images with AK signs with 84% accuracy, and further localizes the regions of diagnostic value with 76.5% accuracy. Conclusions: The proposed AI-based DSS can automatically and accurately preprocess IVCM images (separating artefacts and sorting images into corneal layers) which decreases screening time. The accuracy of AK detection using raw IVCM images must be further explored and improved. Translational Relevance: The proposed automated DSS for experienced specialists assists in diagnosing AK using IVCM images.