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To evaluate the efficacy of CXL in treating fungal keratitis as an adjuvant therapy.Detailed clinical examination microbiological investigation was performed. Twenty fungal keratitis patients were recruited and randomized into two groups: group 1 (n= 11, standard antifungal), group 2 (n=9, corneal collagen crosslinking with standard antifungal). Corneal scraping and tear samples collected were subjected to real-time PCR targeting ITS, TLR analysis and cytokine analysis.The mean time for complete resolution of ulcer for group 2 was significantly shorter compared to group 1 and the final mean BCVA was better for group 2. Expression of IL-1β, IL-8, IFN-γ significantly decreased immediately post CXL in group 2 patients. Significant downregulation of TLR 6, TLR-3, TLR-4 was observed 3-days post CXL compared to group 1 patients.Adjuvant effect of CXL was significant in treating fungal keratitis compared to standalone antifungal treatment.

A young 33 year old male presented with non-resolving corneal infiltrate for 2 month duration in the right eye. KOH/ Calcoflour wet mount revealed sparsely septate fungal hyphae. Post therapeutic penetrating keratoplasty 3 doses of intracameral voriconazole(100μg/0.1ml) was administered suspecting recurrence. Fungal culture revealed non sporulating mould on SDA. PCR based DNA sequencing targeting the ITS region identified the fungal isolate as Mortierella wolfii (M. wolfii) belonging to zygomycetes. To the best of our knowledge, this is the first report of human fungal keratitis caused by M. wolfii.

An 18-year-old girl who had undergone deep anterior lamellar keratoplasty in her left eye for keratoconus a year back presented with inferonasal graft dehiscence with intact host Descemet's membrane and intact anterior chamber after sustaining blunt injury a week prior. The graft was sutured to the host bed, and complete resolution of graft edema was seen in 4 weeks. One year later, she underwent cataract surgery with foldable intraocular lens implantation. At her final follow-up, the best-corrected visual acuity in her left eye was 20/40 with a clear corneal graft and a stable posterior chamber intraocular lens implantation.

Acanthamoeba keratitis (AK) is a rare but vision-threatening keratitis that is usually unilateral; however, it has been reported to be bilateral with the use of contact lenses or orthokeratology.1 ...

Purpose: To assess the clinical and microbiological characteristics of nontuberculous mycobacterial (NTM) keratitis and to evaluate their response to medical therapy. Methods: Sixteen patients of NTM keratitis were retrospectively reviewed from May 2014 to May 2019. Laboratory diagnosis were made using Ziehl-Nielsen acidfast staining, routine culture method of isolation of nontuberculous mycobacteria and further identification of species by PCR (polymerase chain reaction)-based DNA sequencing targeting the heat shock protein-65 (hsp-65) gene. Results: Sixteen patients of microbiologically proven NTM keratitis were included. The average age at the time of presentation was 43.56 years (range, 24–73 years). The mean duration of symptoms was 2.23 months. The commonest risk factor was injury with organic material (43.7) followed by ocular surgery (25%). The majority of the nontuberculous mycobacteria were Mycobacterium abscessus (87.6%) followed by M. fortuitum (6.2%) and M. chelonae (6.2%). The in vitro sensitivity showed maximum sensitivity to Amikacin (AMK; 100%) followed by Azithromycin (AZM; 85.7%), and Clarithromycin (CLR; 85.7%). Out of a total of 16 patients, 12 (75%) had total success with medical therapy while 4 (25%) required surgical intervention. Conclusion: This study is focused on rapid and reliable identification of NTM keratitis through PCR-based identification method to enable effective medical management. The antibiotic susceptibility testing of different subspecies of NTM further reduced the need for surgical intervention. The effective role of AMK either alone or in combination with macrolide antibiotics is also highlighted in this study.

Chatterjee Amit,1,2 Shalini Muralikumar,3 Sargunam Janaki,4 Meena Lakshmipathy,5 Kulandai Lily Therese,4 Vetrivel Umashankar,3 Prema Padmanabhan,5 Janakiraman Narayanan1 1Department of Nanobiotechnology, Vision Research Foundation, Sankara Nethralaya campus, Chennai, Tamil Nadu, India; 2School of Chemical and Biotechnology, SASTRA University, Tanjore, Tamil Nadu, India; 3Centre for Bioinformatics, Vision Research Foundation, Sankara Nethralaya campus, Chennai, Tamil Nadu, India; 4L&T Microbiology Research Centre, Vision Research Foundation, Sankara Nethralaya campus, Chennai, Tamil Nadu, India; 5Department of Cornea, Medical Research Foundation, Sankara Nethralaya campus, Chennai, Tamil Nadu, India Background: Fungal keratitis is a major cause of corneal blindness accounting for more than one-third of microbiologically proven cases. The management of fungal keratitis is through topical or systemic antifungal medications alone or in combination with surgical treatment. Topical medications such as natamycin and voriconazole pose major challenges due to poor penetration across the corneal epithelium. To address the issue various carrier molecules like nanoparticles, lipid vesicles, and cell penetrating peptides were explored. But the major drawback such as non-specificity and lack of bioavailability remains.Purpose: In this study, we have attempted to design corneal specific cell penetrating peptide using subtractive proteomic approach from the published literature and tried to improve its bioavailability through gelatin hydrogel delivery system.Material and Methods: Using subtractive proteomic approach two peptides VRF005 and VRF007 were identified on the basis of solubility, cell permeability and amphipathicity. The peptides were modeled for three-dimensional structure and simulated for membrane penetration. The peptides were characterized using circular dichroism spectroscopy, dynamic light scattering and native polyacrylamide gel electrophoresis. Further uptake studies were performed on primary corneal epithelial cells and the stability was analyzed in corneal epithelial tissue lysates. Insilico prediction of peptides showed it to have antifungal activity which was further validated using colony forming assay and time killing kinetics. The duration of antifungal activity of peptide was improved using gelatin hydrogel through sustained delivery.Results: VRF005 and VRF007 showed α-helical structure and was within the allowed region of Ramachandran plot. The simulation study showed their membrane penetration. The peptide uptake was found to be specific to corneal epithelial cells and also showed intracellular localization in Candida albicans and Fusarium solani. Peptides were found to be stable up to 2 hours when incubated with corneal epithelial tissue lysate. Dynamic light scattering, and native polyacrylamide gel electrophoresis revealed aggregation of peptides. VRF007 showed antifungal activity up to 24 hour whereas VRF005 showed activity up to 4 hours. Hence gelatin hydrogel-based delivery system was used to improve the activity. Actin staining of corneal epithelial cells showed that the cells were attached on gelatin hydrogel.Conclusion: We have designed corneal specific cell penetrating peptides using subtractive proteomic approach. Bioavailability and delivery of peptide was enhanced using gelatin hydrogel system. Keywords: cell-penetrating peptide, gelatin hydrogel, CD spectroscopy, subtractive proteomic approach

PurposeTo report clinical profile and compare management options for Pythium keratitis.MethodRetrospective interventional study of 46 patients diagnosed as Pythium keratitis by PCR DNA sequencing from January 2014 to July 2017. Interventions were categorised into medical management (MM) (topical azithromycin and linezolid with oral azithromycin at presentation), surgery (S) (therapeutic penetrating keratoplasty, TPK), surgical adjunct (SA) (cryotherapy±alcohol with TPK) and medical adjunct (MA) (MM after TPK).ResultsPrimary treatment included MM (1 eye), SA (3 eyes) and S (42 eyes). Recurrence occurred in 27/43 eyes (MM+S group). Second surgery (S) was required in 11 eyes (TPK-2), with additional procedures (SA) in 10 eyes and evisceration in five eyes. 8/43 eyes received MA after TPK-1. One eye required TPK-3. Recurrence occured in all eyes that received MA (100%) and in 28 of 54 TPKs (51.8%) (TPK 1+2+3) in 42 eyes. Recurrence was noted in 1/14 (7.1%) that underwent SA.ConclusionThe currently available and recommended treatment for Pythium keratitis is surgical by means of a TPK and in worse cases evisceration. In our study, MM/MA measures showed no benefit with recurrence or worsening of infection requiring resurgery. Almost 50% of TPKs had a recurrence requiring resurgery. However, adjunctive procedures during TPK appear to have additional benefit with low risk of recurrence and could be included as routine care.

The purpose of the study was to report a case of ulcerative keratitis caused by an unusual algae Prototheca wickerhamii in a diabetic patient. This study design was a case report. A 46-year-old male, who was a known diabetic for 3 years, had an injury to the left cornea with the sparks of fire from wielding at work that developed into an ulcerative keratitis over a period of next 3 months as the patient was not on any medication. Corneal scraping culture report and Vitek 2 system investigation result confirmed it to be a P. wickerhamii infection. The patient was started on intensive topical 1% voriconazole and 5% natamycin for 1 month and with no improvement subsequently underwent penetrating keratoplasty. No recurrence of infection postoperatively was noted. This opportunistic algae rarely known to cause human eye infections is so far reported in either patients with severe systemic immunosuppression causing posterior segment eye involvement or as postcorneal surgery infections. We report an ulcerative keratitis by P. wickerhamii in a diabetic patient post corneal trauma with no prior ocular surgery.

PURPOSE To report the long-term outcome of corneal collagen cross-linking (CXL) for progressive keratoconus in pediatric patients. METHODS "Epithelium-off" CXL was performed in pediatric eyes with progressive keratoconus. Spectacle-corrected distance visual acuity (CDVA), retinoscopy, topography, and tomography were documented preoperatively and postoperatively at 3 months, 6 months, 1 year, and annually thereafter. RESULTS A total of 377 eyes of 336 pediatric patients aged 8 to 18 years with progressive keratoconus underwent CXL. Of these, 194 eyes had a follow-up beyond 2 years and up to 6.7 years. At last follow-up, there was significant improvement in mean CDVA from 0.33 ± 0.22 to 0.27 ± 0.19 logMAR (P ≤ 0.0001), reduction in mean topographic astigmatism from 7.22 ± 3.55 to 6.13 ± 3.28 D (P = 0.0001), mean flattening of 1.20 ± 3.55 diopters in maximum keratometry (Kmax) (P = 0.0002), and mean corneal thinning of 31.1 ± 36.0 μm (P < 0.0001) after CXL. The mean change in Kmax was most significant in moderately advanced keratoconus (average keratometry 48-53 diopters). Central cones showed more corneal flattening than peripheral cones. Stabilization or flattening of Kmax was seen in 85% of eyes at 2 years and in 76% after 4 years. Stabilization or improvement of CDVA was seen in 80.1% of eyes at 2 years and in 69.1% after 4 years. CONCLUSIONS CXL remains effective in stabilizing keratoconus for longer than 2 years in a majority of pediatric eyes. Flattening of Kmax was greater in moderately advanced keratoconus and central cones. Long-term follow-up beyond 4 years, however, revealed that a few eyes showed features suggestive of reversal of the effect of CXL.

To report clinical profile and compare management options for Pythium keratitis.Retrospective interventional study of 46 patients diagnosed as Pythium keratitis by PCR DNA sequencing from January 2014 to July 2017. Interventions were categorised into medical management (MM) (topical azithromycin and linezolid with oral azithromycin at presentation), surgery (S) (therapeutic penetrating keratoplasty, TPK), surgical adjunct (SA) (cryotherapy±alcohol with TPK) and medical adjunct (MA) (MM after TPK).Primary treatment included MM (1 eye), SA (3 eyes) and S (42 eyes). Recurrence occurred in 27/43 eyes (MM+S group). Second surgery (S) was required in 11 eyes (TPK-2), with additional procedures (SA) in 10 eyes and evisceration in five eyes. 8/43 eyes received MA after TPK-1. One eye required TPK-3. Recurrence occured in all eyes that received MA (100%) and in 28 of 54 TPKs (51.8%) (TPK 1+2+3) in 42 eyes. Recurrence was noted in 1/14 (7.1%) that underwent SA.The currently available and recommended treatment for Pythium keratitis is surgical by means of a TPK and in worse cases evisceration. In our study, MM/MA measures showed no benefit with recurrence or worsening of infection requiring resurgery. Almost 50% of TPKs had a recurrence requiring resurgery. However, adjunctive procedures during TPK appear to have additional benefit with low risk of recurrence and could be included as routine care.

We report a rare case of deep anterior lamellar keratoplasty (DALK) neovascularization managed with combination of subconjunctival bevacizumab and argon laser photocoagulation. A 24 year old male underwent Deep anterior lamellar keratoplasty for corneal stromal opacity following presumed viral keratitis. Deep corneal neovascularization was observed postoperatively which was successfully managed using a combination of subconjunctival bevacizumab and argon laser photocoagulation within one week of DALK. The neovascularization resolved by 3 months and at 2 years follow up, patient maintained good visual acuity of 6/12 Snellen's without recurrence of vascularization. A combination of bevacizumab and argon laser may be an effective approach to manage neovascularisation in the immediate postoperative phase (Post DALK) and improve graft survival.