**5.6 Corneal collagen cross-linking (riboflavin with ultraviolet-A irradiation)**

Corneal collagen cross-linking (CXL) has been found successful in halting the progression of keratoconus by using riboflavin and UV-A light. In recent years, role of CXL in infectious keratitis is investigated in several studies with conflicting results on the efficacy of CXL in infectious keratitis [107–113]. Specifically the term photoactivated chromophore cross-linking (PACK-CXL) is used for CXL to treat infectious keratitis [108].

CXL may act in cases of fungal keratitis by a direct antifungal effect and by halting the ongoing melting, thus helping to avoid emergency keratoplasty [109–111]. Said et al. found that although PACK-CXL did not shorten the time to corneal healing, it prevented corneal melting [107]. PACK-CXL is found to be useful in fungal keratitis [108–110]. Abbouda et al. reported halting of corneal melting with PACK-CXL in one case while the other developed perforation [112]. The safety of CXL is of concern because the ultraviolet (UV) -A could damage intraocular structures. Spoerl et al. analyzed the expected damage compared with acceptable damage thresholds. During standard CXL of a cornea with a 400-μm thickness, the irradiances of the UV light reaching the iris, lens, and retina are less than the damage thresholds, and only the microbes, the corneal endothelium, and the keratocytes are at risk [113]. Minor complications after CXL, like transient limbitis and a transient increase in the size of the hypopyon in the first 24 h after CXL reported to be regress subsequently [107].

#### **5.7 Rose Bengal photodynamic therapy**

Photodynamic therapy (PDT) has been used in treatment of choroidal neovascularization in age-related macular degeneration, corneal neovascularization and in infectious keratitis due to Acanthamoeba [114]. PDT involves the activation of photosensitizers using light of varying wavelengths. Rose Bengal photodynamic therapy (RB-PDT) involved a photochemical process using Rose Bengal, excited with green light (wavelength: 500–550 nm) to generate reactive oxygen species (ROS), which, react with various intracellular components to cause cell death. In an in vitro study, Arboleda et al. have demonstrated RB PDT to be successful in fungal keratitis [115].

In a pilot clinical study by Naranjo et al., RB-PDAT was performed in 18 patients with progressive infectious keratitis unresponsive to standard medical therapy. RB-PDAT was considered successful in 13 individuals, defined as control of infection without the need for a therapeutic PK [116]. Amescua G et al. in an vitro and in a case study evaluated the efficacy of rose bengal photodynamic antimicrobial therapy (PDAT). They found that Riboflavin CXL demonstrated no inhibition of fungal isolate growth, whereas rose bengal PDAT inhibited fungal isolate growth within the irradiation zone. In addition, a case with resistant fusarium keratitis was treated successfully [117].

## **6. Future perspective**

#### **6.1 New targets in immunology**

In a study, the role of vitamin D receptor (VDR) in innate immunity being discovered, may be a new target of treatment that can be explored for FK [118]. Liposomes-encapsulated mannan extracts from *C. albicans* stimulate the production of antibodies protective against candidiasis in mice [119]. Probiotics, such as *L. rhamnosus*, *L. acidophilus*, *L. pyogenes*, *L. casei* GG and Bifidobacterium, reported to be protective from candidiasis by eliciting protective immune and non-immune responses in mice [120]. These experimental studies may further facilitate researches to develop fungal keratitis vaccine and use of probiotics in ocular surface for diseases prevention.

#### **6.2 Ocular novel drug delivery system**

Recently, many efforts have been made to improve topical ocular drug delivery by designing various novel drug delivery systems (NDDS), including liposomes, nanoparticles, nanoemulsions, nanosuspensions, micelles, nanofibers, etc.

Several in vitro and in vivo experimental studies have reported encouraging results with NDDS. In a study, the liposomal formulation of the antifungal drug voriconazole found to exhibit a sustained drug release profile, and an 8-fold increase in the amount of drug retained in the cornea after 1 hour of exposure compared with the conventional suspension formulation [121]. The nanoparticle formulation of amphotericin B showed a sustained and controlled drug release for *Fungal Keratitis: Recent Advances in Diagnosis and Treatment DOI: http://dx.doi.org/10.5772/intechopen.98411*

up to 11 hours, while the conventional drug formulation (0.15%) released the entire drug in only 4 hours. Nanoparticle formulation has also shown better pharmacokinetic properties, including 1.5-fold increase in half-life compared to the conventional solution formulation [122]. The microemulsion formulations of fluconazole showed a controlled release profile, releasing 50–80% of the drug in 12 hours, compared to the conventional drug solution, which released almost the entire drug in the first 6 hours [123]. In future, these newer formulations can be very useful in management of fungal keratitis.

### **6.3 Antimicrobial peptides**

Antimicrobial peptides (AMPs) have significant potential for use as antimicrobial agents for ocular or other infections [124]. AMPs, also known as host defense peptides, are naturally produced, small, cationic, amphiphilic peptides ranging in length from 12 to 50 amino acids. They are present on the surfaces of the eyes and in tears. More than 500 AMPs have been reported, including large molecules (RNases and S100A proteins); small peptides α and β defensins in human cationic antibacterial protein (CAP) 18, and α 37 amino acids; proteins like lysozyme and peptidoglycan recognition protein with significant bactericidal activity. The cations carried by AMPs can bind to the anion surface of the bacterial plasma membrane, causing the perforation of cell membrane and subsequently microbial death. AMPs also prevent microbial adhesion to and access into host cells and cause digestion of fungal cell wall by lysozyme [124, 125].

In vitro studies have shown AMPs Pc-C and Pc-E reduced binding of *Aspergillus fumigatus* to cells; CAP37 inhibits candida infection by fungicidal activity [124, 125]. Wu et al. evaluated in vivo application of synthetic β-sheet forming peptide (IKIK) 2-NH2 and (IRIK) 2-NH2 for treatment of FK in comparison with amphotericin B [126]. It was found that topical solutions of the designed peptides were safe, and as effective as the clinically-used Amphotercin B. Many other AMPs such as Clavanin A, Chitinase 3-like 1, and CXCL 10 and S100 proteins may have role in prevention of infection.
