Variations in irradiation energy and rose bengal concentration for photodynamic antimicrobial therapy of fungal keratitis isolates.

The purpose is to assess the efficacy of rose bengal photodynamic antimicrobial therapy (PDAT) using different irradiation energy levels and photosensitizer concentrations for the inhibition of fungal keratitis isolates. Seven different fungi (Aspergillus fumigatus, Candida albicans, Curvularia lunata, Fusarium keratoplasticum, Fusarium solani, Paecilomyces variotii, and Pseudallescheria boydii) were isolated from patients with confirmed infectious keratitis. Experiments were performed in triplicate with suspensions of each fungus exposed to different PDAT parameters including a control, green light exposure of 5.4 J/cm2, 2.7 J/cm2 (continuous and pulsed), and 1.8 J/cm2 and rose bengal concentrations of 0.1%, 0.05%, and 0.01%. Plates were photographed 72 h after experimentation, and analysis was performed to assess fungal growth inhibition. PDAT using 5.4 J/cm2 of irradiation and 0.1% rose bengal completely inhibited growth of five of the seven fungal species. Candida albicans and Fusarium keratoplasticum were the most susceptible organisms, with growth inhibited with the lowest fluence and minimum rose bengal concentration. Fusarium solani, Pseudallescheria boydii, and Paecilomyces variotii were inhibited by lower light exposures and photosensitizer concentrations. Aspergillus fumigatus and Curvularia lunata were not inhibited by any PDAT parameters tested. Continuous and pulsed irradiation using 2.7 J/cm2 produced similar results. Rose bengal PDAT successfully inhibits the in vitro growth of five fungi known to cause infectious keratitis. Differences in growth inhibition of the various fungi to multiple PDAT parameters suggest that susceptibilities to PDAT are unique among fungal species. These findings support modifying PDAT parameters based on the infectious etiology.

Photoactivated chromophore-corneal cross-linking accelerates corneal healing in fungal keratitis: an updated meta-analysis.

AIM: To determine the effectiveness and safety of photoactivated chromophore-corneal cross-linking (PACK-CXL) adjuvant in infectious keratitis by April 5, 2022. METHODS: We searched randomized controlled trials (RCTs) comparing standard antibiotic treatment (SAT) plus PACK-CXL to SAT in infectious keratitis in Embase, MEDLINE with PubMed, Web of Science, and Cochrane Library. We independently screened and extracted data using predesigned tables. Cochrane's risk-of-bias tool was utilized to examine the quality of RCTs. A random-effects model was employed to determine the overall effect size of the meta-analyses. Grading of Recommendations, and Assessment, Development and Evaluations (GRADE) was also performed to examine the quality of evidence. RESULTS: Seven eligible RCTs with 283 patients were acquired. Adjuvant PACK-CXL reduced the time needed to perform corneal healing in fungal keratitis (- 1.33 months; 95% CI, - 1.83 to - 0.42, I2 = 0%, P < 0.05) as compared to SAT alone. The risks of adverse events were not significantly different both in fungal and bacterial keratitis. Due to the substantial heterogeneity among studies, such as population, the type and severity of infectious keratitis, drug regimens of SAT, PACK-CXL protocol, and the judgment of subjective outcomes, the evidence grade was low. CONCLUSION: Adjuvant PACK-CXL accelerates fungal keratitis healing as compared to SAT alone. But more rigorous RCTs are required to determine the clinical effectiveness and safety.

Zwitterionic silver nanoparticle based antibacterial eye drops for efficient therapy of bacterial keratitis.

Inefficient biofilm clearance and the risk of drug resistance pose significant challenges for antibiotic eye drops in the treatment of bacterial keratitis (BK). Recently, silver nanoparticles (AgNPs) have emerged as promising alternatives to antibiotics due to their potent antibacterial activity and minimal drug resistance. However, concerns regarding the potential biotoxicity of aggregated AgNPs in tissues have limited their practical application. In this study, polyzwitterion-functionalized AgNPs with excellent dispersion stability in the ocular physiological environment were chosen to prepare antibacterial eye drops. Zwitterionic AgNPs were synthesized using a copolymer, poly(sulfobetaine methacrylate-co-dopamine methacrylamide) (PSBDA), as a stabilizer and a reducing agent. The resulting antibacterial eye drops, named ZP@Ag-drops, demonstrated outstanding biocompatibility in in vitro cytotoxicity tests and in vivo rabbit eye instillation experiments, attributed to the zwitterionic PSBDA surface. Furthermore, the ZP@Ag-drops exhibited strong antibacterial activity against multiple pathogenic bacteria, particularly in penetrating and eradicating biofilms, due to the synergistic bactericidal effect of the released Ag+ and reactive oxygen species (ROS). Importantly, in vivo BK rabbit models showed that the ZP@Ag-drops effectively inhibited corneal infection and prevented ocular tissue damage, surpassing the therapeutic effect of commercial levofloxacin eye drops (LEV-drops). Overall, this study presents a promising alternative option for the effective treatment of BK using antibacterial eye drops.

Multifunctional Polymer Vesicles for Synergistic Antibiotic-Antioxidant Treatment of Bacterial Keratitis.

As an acute ophthalmic infection, bacterial keratitis (BK) can lead to severe visual morbidity, such as corneal perforation, intraocular infection, and permanent corneal opacity, if rapid and effective treatments are not available. In addition to eradicating pathogenic bacteria, protecting corneal tissue from oxidative damage and promoting wound healing by relieving inflammation are equally critical for the efficient treatment of BK. Besides, it is very necessary to improve the bioavailability of drugs by enhancing the ocular surface adhesion and corneal permeability. In this investigation, therefore, a synergistic antibiotic-antioxidant treatment of BK was achieved based on multifunctional block copolymer vesicles, within which ciprofloxacin (CIP) was simultaneously encapsulated during the self-assembly. Due to the phenylboronic acid residues in the corona layer, these vesicles exhibited enhanced muco-adhesion, deep corneal epithelial penetration, and bacteria-targeting, which facilitated the drug delivery to corneal bacterial infection sites. Additionally, the abundant thioether moieties in the hydrophobic membrane enabled the vesicles to both have ROS-scavenging capacity and accelerated CIP release at the inflammatory corneal tissue. In vivo experiments on a mice model demonstrated that the multifunctional polymer vesicles achieved efficient treatment of BK, owing to the enhanced corneal adhesion and penetration, bacteria targeting, ROS-triggered CIP release, and the combined antioxidant-antibiotic therapy. This synergistic strategy holds great potential in the treatment of BK and other diseases associated with bacterial infections.

In Situ Hybridization of Polymeric Curcumin to Arginine-Derived Carbon Quantum Dots for Synergistic Treatment of Bacterial Infections.

Effective infectious keratitis treatment must eliminate the pathogen, reduce the inflammatory response, and prevent persistent damage to the cornea. Infectious keratitis is generally treated with broad-spectrum antibiotics; however, they have the risk of causing corneal epithelial cell damage and drug resistance. In this study, we prepared a nanocomposite (Arg-CQDs/pCur) from arginine (Arg)-derived carbon quantum dots (Arg-CQDs) and polymeric curcumin (pCur). Partial carbonization of arginine hydrochloride in the solid state by mild pyrolysis resulted in the formation of CQDs, which exhibited enhanced antibacterial activity. pCur was formed by the polymerization of curcumin, and further crosslinking reduced its cytotoxicity and improved antioxidative, anti-inflammatory, and pro-proliferative activities. The pCur in situ conjugated with Arg-CQDs to form the Arg-CQDs/pCur nanocomposite, which showed a minimum inhibitory concentration of ca. 10 µg mL-1, which was >100-fold and >15-fold lower than that of the precursor arginine and curcumin, respectively, against Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa. The Arg-CQDs/pCur nanocomposite with combined antibacterial, antioxidative, anti-inflammatory, pro-proliferative properties, and long-term retention on cornea enabled synergistic treatment of bacterial keratitis. In a rat model, it can effectively treat P. aeruginosa-induced bacterial keratitis at a concentration 4000-fold lower than the commercially used drug, Sulmezole eye drops. Arg-CQDs/pCur nanocomposites have great potential for application in antibacterial and anti-inflammatory nanoformulations for clinical use to treat infectious diseases.

Antimicrobial Drug-Drug Interactions in the Treatment of Infectious Keratitis.

PURPOSE: Infectious keratitis is a serious disease requiring immediate, intensive, and broad-spectrum empiric treatment to prevent vision loss. Given the diversity of organisms that can cause serious corneal disease, current guidelines recommend treatment with several antimicrobial agents simultaneously to provide adequate coverage while awaiting results of microbiology cultures. However, it is currently unknown how the use of multiple ophthalmic antimicrobial agents in combination may affect the efficacy of individual drugs. METHODS: Using a panel of 9 ophthalmic antibiotics, 3 antifungal agents, and 2 antiacanthamoeba therapeutics, fractional inhibitory concentration testing in the standard checkerboard format was used to study 36 antibiotic-antibiotic combinations, 27 antibiotic-antifungal combinations, and 18 antibiotic-antiacanthamoeba combinations against both Staphylococcus aureus and Pseudomonas aeruginosa for synergistic, additive, neutral, or antagonistic drug-drug interactions. RESULTS: We demonstrate that while most combinations resulted in no change in antimicrobial efficacy of individual components, the combination of erythromycin + polyhexamethylene biguanide was found to be antagonistic toward P. aeruginosa . Conversely, 18 combinations toward S. aureus and 15 combinations toward P. aeruginosa resulted in additive or synergistic activity, including 4 with improved activity toward both species. CONCLUSIONS: Understanding how drug-drug interactions may affect drug efficacy is critical to selecting the appropriate combination therapy and improving clinical outcomes of this blinding disease.

Protective effects of resolvin D1 in Pseudomonas aeruginosa keratitis.

PURPOSE: Here, we explored the protective effects of resolvin D1 (RvD1) in Pseudomonas aeruginosa (PA) keratitis. METHODS: C57BL/6 (B6) mice were used as an animal model of PA keratitis. Plate counting and clinical scores were used to assess the severity of the infection and the therapeutic effects of RvD1 in the model. Myeloperoxidase assay was used to detect neutrophil infiltration and activity. Quantitative PCR (qPCR) was used to examine the expression of proflammatory and anti-inflammatory mediators. Immunofluorescence staining and qPCR were performed to identify macrophage polarization. RESULTS: RvD1 treatment alleviated PA keratitis severity by decreasing corneal bacterial load and inhibiting neutrophil infiltration in the mouse model. Furthermore, RvD1 treatment decreased mRNA levels of TNF-α, IFN-γ, IL-1ß, CXCL1, and S100A8/9 while increasing those of IL-1RA, IL-10, and TGF-ß1. RvD1 treatment also reduced the aggregation of M1 macrophages and increased that of M2 macrophages. RvD1 provided an auxiliary effect in gatifloxacin-treated mice with PA keratitis. CONCLUSION: Based on these findings, RvD1 may improve the prognosis of PA keratitis by inhibiting neutrophil recruitment and activity, dampening the inflammatory response, and promoting M2 macrophage polarization. Thus, RvD1 may be a potential complementary therapy for PA keratitis.

Chitosan-poly(lactide-co-glycolide)/poloxamer mixed micelles as a mucoadhesive thermo-responsive moxifloxacin eye drop to improve treatment efficacy in bacterial keratitis.

A mucoadhesive self-assembling polymeric system was developed to carry moxifloxacin (M) for treating bacterial keratitis (BK). Chitosan-PLGA (C) conjugate was synthesized, and poloxamers (F68/127) were mixed in different proportions (1: 5/10) to prepare moxifloxacin (M)-encapsulated mixed micelles (M@CF68/127(5/10)Ms), including M@CF68(5)Ms, M@CF68(10)Ms, M@CF127(5)Ms, and M@CF127(10)Ms. The corneal penetration and mucoadhesiveness were determined biochemically, in vitro using human corneal epithelial (HCE) cells in monolayers and spheroids, ex vivo using goat cornea, and in vivo via live-animal imaging. The antibacterial efficacy was studied on planktonic biofilms of P. aeruginosa and S. aureus (in vitro) and Bk-induced mice (in vivo). Both M@CF68(10)Ms and M@CF127(10)Ms demonstrated high cellular uptake, corneal retention, muco-adhesiveness, and antibacterial effect, with M@CF127(10)Ms exhibiting superior therapeutic effects in P. aeruginosa and S. aureus-infected BK mouse model by reducing the corneal bacterial load and preventing corneal damage. Therefore, the newly developed nanomedicine is promising for clinical translation in treating BK.

The Antibacterial Efficacy of High-Fluence PACK Cross-Linking Can Be Accelerated.

Purpose: To determine whether high-fluence photoactivated chromophore for keratitis cross-linking (PACK-CXL) can be accelerated. Methods: Solutions of Staphylococcus aureus and Pseudomonas aeruginosa with 0.1% riboflavin were prepared and exposed to 365 nm ultraviolet (UV)-A irradiation of intensities and fluences from 9 to 30 mW/cm2 and from 5.4 to 15.0 J/cm2, respectively, representing nine different accelerated PACK-CXL protocols. Irradiated solutions and unirradiated controls were diluted, plated, and inoculated on agar plates so that the bacterial killing ratios (BKR) could be calculated. Additionally, strains of Achromobacter xylosoxidans, Staphylococcus epidermidis, and Stenotrophomonas maltophilia were exposed to a single accelerated PACK-CXL protocol (intensity: 30 mW/cm2, total fluence: 15.0 J/cm2). Results: With total fluences of 5.4, 10.0, and 15.0 J/cm2, the range of mean BKR for S. aureus was 45.78% to 50.91%, 84.13% to 88.16%, and 97.50% to 99.90%, respectively; the mean BKR for P. aeruginosa was 69.09% to 70.86%, 75.37% to 77.93%, and 82.27% to 91.44%, respectively. The mean BKR was 41.97% for A. xylosoxidans, 65.38% for S. epidermidis, and 78.04% for S. maltophilia for the accelerated PACK-CXL protocol (30 mW/cm2, 15 J/cm2). Conclusions: The BKR of high-fluence PACK-CXL protocols can be accelerated while maintaining a high, but species-dependent, BKR. The Bunsen to Roscoe law is respected in fluences up to 10 J/cm2 in S. aureus and P. aeruginosa, whereas fluences above 10 J/cm2 show strain dependence. Translational Relevance: The high-fluence PACK-CXL protocols can be accelerated in clinical practice while maintaining high levels of BKR.

Susceptibility pattern of bacterial isolates in equine ulcerative keratitis: Implications for empirical treatment at a university teaching hospital in Sydney.

Corneal ulceration is a common ophthalmic condition in horses. It is frequently caused by trauma to the corneal surface, followed by secondary infection by commensal or pathogenic organisms including Streptococcus equi subspecies zooepidemicus, Pseudomonas aeruginosa and Staphylococcus spp. Emerging antimicrobial resistance amongst these organisms has raised the need for appropriate antimicrobial therapy selection, to optimise treatment efficacy while minimising further antimicrobial resistance. Medical records of 38 horses presented at the University Veterinary Teaching Hospital Camden for ulcerative keratitis between 2010 and 2020 were reviewed to identify those with positive bacterial cultures and antimicrobial susceptibility profiles (13/38). Common susceptibility patterns were identified and used to guide the empirical treatment of equine bacterial corneal ulcers. Pseudomonas spp. (64.3%), Streptococcus equi subspecies zooepidemicus (14.3%) and Actinobacillus spp. (14.3%) were most commonly identified. Susceptibility to amikacin, gentamicin and ciprofloxacin was observed in 100%, 66.7% and 85.7% Pseudomonas spp. isolates respectively. Resistance to polymyxin B and neomycin occurred in 85.7% and 71.4% of Pseudomonas spp., respectively. All Streptococcus equi subspecies zooepidemicus organisms in this study were susceptible to ceftiofur, cephalexin, penicillin and ampicillin, while they were all resistant to gentamicin, neomycin, enrofloxacin and marbofloxacin. Predominating in this study, Pseudomonas spp. maintained overall aminoglycoside susceptibility despite some emerging resistance, and good fluoroquinolone susceptibility. High resistance to Polymyxin B could have arisen from its common use as first-line therapy for bacterial corneal ulcers. Although further research is required, these new findings about predominant bacteria in equine corneal ulceration in the Camden region and their antimicrobial susceptibility patterns can be used to guide the empirical treatment of bacterial corneal ulcers in horses.

A Literature-Based Review and Analysis of the Pharmacodynamics of the Dose Frequency of Topical 0.3% Ciprofloxacin and 0.3% Ofloxacin in the Day-1 Treatment of Bacterial Keratitis.

Purpose: To determine the appropriate dose frequency for the second-generation fluoroquinolones (2FQs), ciprofloxacin 0.3% and ofloxacin 0.3%, in the day-1 treatment of bacterial keratitis (BK) based on the corneal concentrations achievable and required Minimum Inhibitory Concentration90 (MIC90) of common BK isolates. Methods: Literature-based ocular MIC90 required to treat bacterial isolates of BK patients were determined for each fluoroquinolone. Published corneal concentrations for each 2FQ, and the drop regimens used to reach these concentrations, were then analyzed to determine the relationship between the corneal 2FQ concentration and the amount of drug applied per hour and the total amount applied. Results: Significant relationships were found to exist for corneal concentrations of both ciprofloxacin and ofloxacin and the amount of drug applied per hour (both P = 0.005), and the total amount of drug applied (P = 0.003 and P = 0.0004, respectively). Derived ciprofloxacin drops/hour corneal concentrations agreed well with both a literature-based regimen and the manufacturers' day-1 drop regimen for various MIC90. Derived ofloxacin drops per hour indicated a higher rate than that suggested by the manufacturer. Conclusions: Both a literature-based and the manufacturers' drop regimens for the day-1 treatment of BK using 0.3% ciprofloxacin have a pharmacodynamic basis, which is related to the required MIC90 of commonly encountered isolates in BK. Dose frequency for 0.3% ofloxacin should be in line with the manufacturers' maximum suggested drop regimen. Commonly suggested drop regimens below these recommendations for either FQ may need to be revised in view of these findings.

Therapeutic effect of Atractylenolide I on Aspergillus fumigatus keratitis by affecting MyD88/ NF-κB pathway and IL-1ß, IL-10 expression.

PURPOSE: Atractylenolide I (AT-I) is a natural sesquiterpene with anti-inflammatory effects. The purpose of this study was to research the anti-inflammatory effect of AT-I on Aspergillus fumigatus(A. fumigatus) keratitis in mice. METHODS: Cytotoxicity test and cell scratch test were used to determine the therapeutic concentrations of corneal infections. In vivo and in vitro studies, mouse cornea and human corneal epithelial cells (HCECs) infected with A. fumigatus were treated with AT-I or dimethyl sulfoxide (DMSO). Then, to analyze the effect of AT-I on inflammatory response, namely neutrophil or macrophage recruitment and the expression of cytokines involving MyD88, NF-κB, interleukin 1ß (IL-1ß) and interleukin 10 (IL-10). To study the effects of the drug, the techniques used include slit-lamp photography, immunofluorescence, myeloperoxidase (MPO) detection, quantitative real-time polymerase chain reaction (QRT-PCR), and western blot. At the same time, in order to explore the combined effect of the drug and natamycin, slit-lamp photographs and clinical scores were used to visually display the disease process. RESULTS: No cytotoxicity was observed under the action of AT-I at a concentration of 800 µM. In mouse models, AT-I significantly suppressed inflammatory responses, reduced neutrophil and macrophage recruitment, and decreased myeloperoxidase levels early in infection. Studies have shown that AT-I may reduce the levels of IL-1ß and IL-10 by inhibiting the MyD88/ NF-κB pathway. The drug combined with natamycin can increase corneal transparency in infected mice. CONCLUSION: AT-I may inhibit MyD88 / NF-κB pathway and the secretion of inflammatory factors IL-1 ß and IL-10 to achieve the therapeutic effect of fungal keratitis.

Cell Wall Destruction and Internal Cascade Synergistic Antifungal Strategy for Fungal Keratitis.

Fungal keratitis is one of the most common blindness-causing diseases, but clinical antifungal treatment remains a challenge. The fungal cell wall and biofilm matrix which severely confine the drug preparation are the critical obstructive factors to therapeutic effects. Herein, we report ethylenediaminetetraacetic acid (EDTA) modified AgCu2O nanoparticles (AgCuE NPs) to disrupt the cell wall and then eradicate C. albicans through the internal cascade synergistic effects of ion-released chemotherapy, chemodynamic therapy, photodynamic therapy, and mild photothermal therapy. AgCuE NPs exhibited excellent antifungal activity both in preventing biofilm formation and in destroying mature biofilms. Furthermore, AgCuE NP based gel formulations were topically applied to kill fungi, reduce inflammation, and promote wound healing, using optical coherence tomography and photoacoustic imaging to monitor nanogel retention and therapeutic effects on the infected murine cornea model. The AgCuE NP gel showed good biosafety and no obvious ophthalmic and systemic side effects. This study suggests that the AgCuE NP gel is an effective and safe antifungal strategy for fungal keratitis with a favorable prognosis and potential for clinical translation.

The clinical treatment of bacterial keratitis: A review of drop instillation regimes.

Bacterial keratitis (BK) presentations are often treated using the commercially available second-generation fluoroquinolones ciprofloxacin 0.3% and ofloxacin 0.3% as monotherapy. The guidelines available for instillation regimes are often not supported by data from clinical studies. This review examines the peer-reviewed clinical studies and compared treatment failure rates for ciprofloxacin 0.3% and ofloxacin 0.3% for BK in relation to Day-1 drop-regimes. From the statistical analysis, this review derived evidence-based clinically applicable minimum drop-regimes for the treatment of BK on Day-1. Lower numbers of drops of ciprofloxacin on Day-1 were significantly associated with increased treatment failure rates (p < 0.002). The derived minimum number of drops on Day for ciprofloxacin on Day-1 was 47 drops, and for ofloxacin 24 drops. The mean number of drops used in the clinical studies was significantly lower than the manufacturers' recommended Day-1 regimes for both ciprofloxacin (p = 0.0006) and ofloxacin (p = 0.048). From Day-3 to -6 of treatment the drop rates for ciprofloxacin relative to recommended rates were higher, and for ofloxacin lower (p = 0.014). The findings of this review were then compared with a representative sample of published guidelines and case studies to determine the validity of applying those drop-regimes in clinical practice. Although the manufacturers' suggested minimum drop-regimes on Day-1 were significantly different (120 drops ciprofloxacin, 34 drops ofloxacin, p < 0.0001), many of the published guidelines suggested the same drop-regime for both fluoroquinolones. The suggested drop numbers on Day-1 for ciprofloxacin in these guidelines and case studies were significantly less than those used in the clinical studies (p = 0.043). Increased treatment failure rates for ciprofloxacin are associated with lower drop numbers on Day-1. The Day-1 dosing rates for ciprofloxacin and ofloxacin should be considered separately, and the regimes suggested in published guidelines and case studies may need be re-considered in light of the findings of this review.

Clinical Outcomes of Rose Bengal Mediated Photodynamic Antimicrobial Therapy on Fungal Keratitis with Their Microbiological and Pathological Correlation.

PURPOSE: To report the clinical outcome of Photodynamic Antimicrobial Therapy (PDAT) with Rose Bengal (RB) used as an early adjuvant therapy in patients with fungal keratitis and their microbiological and pathological correlation. METHODS: Patients with microbiologically confirmed fungal keratitis underwent PDAT-RB along with topical natamycin 5% drops hourly and oral ketoconazole 200 mg twice a day. This was performed by applying rose bengal (0.1%) to the de-epithelialized cornea for 30 minutes, followed by irradiation with a 6 mW/cm2 custom-made green LED source for 15 minutes (5.4 J/cm2). The corresponding fungal isolates were tested in vitro using PDAT-RB and corneal buttons were evaluated for correlation. RESULTS: Following informed consent, seven patients (male-5, female-2, mean age 47.7 years) with fungal keratitis were recruited. There were 3 cases each of Fusarium and Aspergillus flavus and 1 case of Acremonium sp. The average vertical and horizontal diameters of the corneal infiltrate were 4.12 ± 0.55 and 3.99 ± 1.19 mm, respectively. The average depth of corneal involvement was 283 ± 75.27µ as measured by anterior segment OCT. The clinical resolution was achieved in the cases with Fusarium keratitis with an average time of 39 days. Three cases of A. flavus and a single patient with Acremonium keratitis worsened and needed therapeutic keratoplasty (TPK) for resolution. Post-TPK, the corneal tissues grew A. flavus in one out of three cases and Acremonium sp. in one case. In vitro PDAT-RB experiment was performed on the corresponding fungal isolates grown from the corneal scraping. PDAT-RB produced clear inhibition of Fusarium and Acremonium sp. with no effect on the growth of A. flavus. Histopathologically, 2 out of 4 (50%) corneal buttons showed fungal filaments. CONCLUSIONS: While the in vitro and in vivo results of PDAT-RB matched for Fusarium sp. and Aspergillus flavus keratitis being favourable in the former and non-favourable in the latter, these results were discrepant in Acremonium sp.

Secondary Amine Pendant ß-Peptide Polymers Displaying Potent Antibacterial Activity and Promising Therapeutic Potential in Treating MRSA-Induced Wound Infections and Keratitis.

Interest in developing antibacterial polymers as synthetic mimics of host defense peptides (HPDs) has accelerated in recent years to combat antibiotic-resistant bacterial infections. Positively charged moieties are critical in defining the antibacterial activity and eukaryotic toxicity of HDP mimics. Most examples have utilized primary amines or guanidines as the source of positively charged moieties, inspired by the lysine and arginine residues in HDPs. Here, we explore the impact of amine group variation (primary, secondary, or tertiary amine) on the antibacterial performance of HDP-mimicking ß-peptide polymers. Our studies show that a secondary ammonium is superior to either a primary ammonium or a tertiary ammonium as the cationic moiety in antibacterial ß-peptide polymers. The optimal polymer, a homopolymer bearing secondary amino groups, displays potent antibacterial activity and the highest selectivity (low hemolysis and cytotoxicity). The optimal polymer displays potent activity against antibiotic-resistant bacteria and high therapeutic efficacy in treating MRSA-induced wound infections and keratitis as well as low acute dermal toxicity and low corneal epithelial cytotoxicity. This work suggests that secondary amines may be broadly useful in the design of antibacterial polymers.

PACK Cross-Linking as Adjuvant Therapy Improves Clinical Outcomes in Culture-Confirmed Bacterial Keratitis.

PURPOSE: We recently showed the positive clinical effects of combining accelerated corneal cross-linking (PACK-CXL) with antibiotic treatment in patients with presumed bacterial keratitis. In this study, we compare the impacts of a combined PACK-CXL/standard antibiotic treatment (PACK-ABX group) with standard antibiotic treatment alone (ABX group) in patients with culture-confirmed bacterial keratitis. METHODS: We reviewed patients with moderate and severe bacterial keratitis and confirmed bacterial cultures. Clinical outcomes were compared for standard antibiotic treatment alone, before the initiation of PACK-CXL, and after adjuvant use of PACK-CXL. RESULTS: A total of 47 eyes of 47 patients were included: 26 eyes in the PACK-ABX group and 21 eyes in the ABX group. Pathogens, baseline demographics (besides age), and clinical parameters were similar between the 2 groups. The PACK-ABX patients had better final uncorrected visual acuity [mean difference 0.57 Logarithm of the Minimum Angle of Resolution, 95% Confidence Interval (CI): 0.16-0.99, P = 0.07] and best-corrected visual acuity (mean difference 0.70 Logarithm of the Minimum Angle of Resolution, 95% CI: 0.23-1.16, P = 0.04), shorter reepithelialization time (mean difference 9.63 days, 95% CI: 3.14-16.12, P = 0.004), and reduced number of clinic visits (mean difference 4.8 meetings, 95% CI: 1.4-8.2, P = 0.007) and need for tectonic grafts (0 vs. 33.3%, P = 0.002). A multivariate analysis controlling for age, sex, ulcer size, and Gram stain showed that PACK-ABX treatment remained significantly associated with reepithelialization time (ß = 14.5, P = 0.001). CONCLUSIONS: In our study, PACK-CXLs addition to the standard of care in cases of culture-proven bacterial keratitis had a positive effect on the final visual acuity and time to resolution, compared with the standard-of-care treatment.

Hybrid derivative of cathelicidin and human beta defensin-2 against Gram-positive bacteria: A novel approach for the treatment of bacterial keratitis.

Bacterial keratitis (BK) is a major cause of corneal blindness globally. This study aimed to develop a novel class of antimicrobial therapy, based on human-derived hybrid host defense peptides (HyHDPs), for treating BK. HyHDPs were rationally designed through combination of functional amino acids in parent HDPs, including LL-37 and human beta-defensin (HBD)-1 to -3. Minimal inhibitory concentrations (MICs) and time-kill kinetics assay were performed to determine the concentration- and time-dependent antimicrobial activity and cytotoxicity was evaluated against human corneal epithelial cells and erythrocytes. In vivo safety and efficacy of the most promising peptide was examined in the corneal wound healing and Staphylococcus aureus (ATCC SA29213) keratitis murine models, respectively. A second-generation HyHDP (CaD23), based on rational hybridization of the middle residues of LL-37 and C-terminal of HBD-2, was developed and was shown to demonstrate good efficacy against methicillin-sensitive and methicillin-resistant S. aureus [MIC = 12.5-25.0 µg/ml (5.2-10.4 µM)] and S. epidermidis [MIC = 12.5 µg/ml (5.2 µM)], and moderate efficacy against P. aeruginosa [MIC = 25-50 µg/ml (10.4-20.8 µM)]. CaD23 (at 25 µg/ml or 2× MIC) killed all the bacteria within 30 min, which was 8 times faster than amikacin (25 µg/ml or 20× MIC). After 10 consecutive passages, S. aureus (ATCC SA29213) did not develop any antimicrobial resistance (AMR) against CaD23 whereas it developed significant AMR (i.e. a 32-fold increase in MIC) against amikacin, a commonly used treatment for BK. Pre-clinical murine studies showed that CaD23 (0.5 mg/ml) achieved a median reduction of S. aureus bioburden by 94% (or 1.2 log10 CFU/ml) while not impeding corneal epithelial wound healing. In conclusion, rational hybridization of human-derived HDPs has led to generation of a potentially efficacious and safe topical antimicrobial agent for treating Gram-positive BK, with no/minimal risk of developing AMR.

Combined Phialemonium curvatum and Acanthamoeba Keratitis: The Importance of Early Diagnosis and Specific Therapy.

PURPOSE: To report the clinical and confocal findings of a unique case of combined Phialemonium curvatum and Acanthamoeba keratitis and to highlight the role of the prompt diagnosis and specific medical treatment in preserving visual function. METHODS: A case report and literature review. RESULTS: A 54-year-old woman presented with a 3-day history of visual impairment, photophobia, and ocular pain in her right eye. Her best corrected visual acuity was 0.4 Logarithm of the Minimum Angle of Resolution scale, and the slit-lamp examination showed whitish corneal stromal infiltrate with satellite lesions. In vivo confocal microscopy evidenced Acanthamoeba cysts and fungal hyphae that resulted P. curvatum in the culture examination. The intensive medical treatment was started with topical 0.02% polyhexamethylene biguanide, voriconazole 1%, and moxifloxacin hydrochloride 0.5%. Progressive improvement of clinical and confocal pictures was registered with a complete recovery of visual function after 1 month. CONCLUSIONS: This is the first case report of combined P. curvatum and Acanthamoeba keratitis. The fast diagnosis with in vivo confocal microscopy allowed early and intensive specific treatment with recovery of corneal infection.

Cationic Polyelectrolyte Nanocapsules of Moxifloxacin for Microbial Keratitis Therapy: Development, Characterization, and Pharmacodynamic Study.

Microbial keratitis (MK) is a vision-threatening disease and the fourth leading cause of blindness worldwide. In this work, we aim to develop moxifloxacin (MXN)-loaded chitosan-based cationic mucoadhesive polyelectrolyte nanocapsules (PENs) for the effective treatment of MK. PENs were formulated by polyelectrolyte complex coacervation method and characterized for their particle size, surface charge, morphology, mucoadhesive property, in-vitro and ex-vivo release, ocular tolerance, and antimicrobial efficacy studies. The pharmacodynamic study was conducted on rabbit eye model of induced keratitis and it is compared with marketed formulation (MF). Developed PENs showed the size range from 230.7 ± 0.64 to 249.0 ± 0.49 nm and positive surface charge, spherical shape along with appropriate physico-chemical parameters. Both in-vitro and ex-vivo examination concludes that PENs having more efficiency in sustained release of MXN compared to MF. Ocular irritation studies demonstrated that no corneal damage or ocular irritation. The in-vivo study proved that the anti-bacterial efficacy of PENs was improved when compared with MF. These results suggested that PENs are a feasible choice for MK therapy because of their ability to enhance ocular retention of loaded MXN through interaction with the corneal surface of the mucous membrane.