Mesenchymal stem cell-based adjunctive therapy for Pseudomonas aeruginosa-induced keratitis: A proof-of-concept in-vitro study.

PURPOSE: Pseudomonas aeruginosa-induced keratitis is one of the most severe and challenging forms of corneal infection, owing to its associated intense inflammatory reactions leading to corneal necrosis and dense corneal scar with loss of vision. Since mesenchymal stem cells (MSCs) are reported to possess antimicrobial and immunomodulatory properties, they can be tested as an adjuvant treatment along with the antibiotics which are the current standard of care. This study aims to investigate the anti-bacterial and immunomodulatory roles of human bone marrow MSC-derived conditioned medium (MSC-CM) in P. aeruginosa-infected human corneal epithelial cells (HCECs) in vitro. METHODS: The effect of MSC-CM on the growth of clinical isolates of P. aeruginosa was evaluated by colony-forming unit assay. The expression of inflammatory cytokines (IL-6 and TNF-α) and an antimicrobial peptide (Lipocalin 2) in lipopolysaccharide-treated MSCs and HCECs was analyzed through ELISA. Corneal epithelial repair following infection with P. aeruginosa was studied through scratch assay. RESULTS: Compared to control (P. aeruginosa (5*105) incubated in DMEM (1 ml) at 37 °C for 16 h), MSC-CM significantly: i) inhibits the growth of P. aeruginosa (159*109 vs. 104*109 CFU/ml), ii) accelerates corneal epithelial repair following infection with P. aeruginosa (9% vs. 24% closure of the wounded area after 12 h of infection), and iii) downregulates the lipopolysaccharide-induced expression of IL-6, TNF-α and Lipocalin 2 in HCECs. A combination of MSC-CM with an antibiotic, Ciprofloxacin moderately regulated the expression of IL-6, TNF-α, and Lipocalin 2. CONCLUSION: MSC-CM holds promise as an adjunctive therapeutic approach for P. aeruginosa-induced corneal epithelial damage.

Proteomic landscape of extracellular vesicles in human retinal cells infected with Staphylococcus aureus and Pseudomonas aeruginosa: Role in endophthalmitis.

Extracellular Vesicles (EVs) have evolved as a promising entity for developing diagnostic and therapeutic biomarkers. We profiled global EV proteome of EVs from Human retinal cells (ARPE-19) infected with S. aureus and P. aeruginosa. EVs were isolated by ultracentrifugation and subjected to LC-MS/MS for proteome analysis. In S. aureus infection, sequest identified 864 proteins, of which 81 were differentially expressed in comparison to control. Similarly, in P. aeruginosa infection, of 516 proteins identified, 86 were differentially expressed. Additionally, 38 proteins were exclusive to infected sets. KEGG and Gene Ontology revealed crucial dysregulated pathways involving proteins such as complement cascades, annexins and calpain-2, all playing major role in the pathogenesis of the disease. This study provides insight into the global EV proteome of S. aureus and P. aeruginosa endophthalmitis with their functional correlation and distinctive pattern of expression. Calpain-2 and C8a are attractive biomarkers for bacterial endophthalmitis.

Antibiotic susceptibility in Endophthalmitis Management Study and intravitreal antibiotic practice trend in India-EMS Report #5.

AIM: Analyze antibiotic susceptibility in the Endophthalmitis Management Study (EMS) and compare it with the current intravitreal antibiotic practice trend of members of the Vitreoretinal Society of India (VRSI) practicing in India. METHODS: The microbiology work-up of undiluted vitreous included microscopy, culture-susceptibility, polymerase chain reaction (PCR), and next-generation sequencing (NGS). VRSI members were invited to the survey. The EMS conventional culture-susceptibility (PCR and NGS excluded) results were compared vis-a-vis gram-positive cocci (GPC), gram-negative bacilli (GNB), and less commonly used antibiotics with the current recommended intravitreal antibiotics. p < 0.05 was considered significant. RESULTS: Culture and positivity (culture + PCR/NGS) positivity was 28.8% and 56.1%, respectively. GPC was most susceptible to cefazolin, linezolid, and vancomycin; GNB was most susceptible to amikacin, ceftazidime, colistin, and imipenem. There was no susceptibility difference between cefazolin and vancomycin (p = 0.999) and between ceftazidime and imipenem (p = 1.0). Colistin was superior to ceftazidime (p = 0.047) against GNB. The GNB resistant to amikacin (n = 14) were equally susceptible to ceftazidime and colistin; resistant to ceftazidime (n = 16) were susceptible to colistin; and resistant to colistin (n = 7) were susceptible to ceftazidime. The preference of VRSI members (n = 231) practicing in India was a vancomycin-ceftazidime combination (82%), vancomycin for GPC (94%), ceftazidime for GNB (61%), and voriconazole for fungi (74%). CONCLUSION: In EMS, GPC had good susceptibility to vancomycin; GNB had good susceptibility to ceftazidime and colistin. Given the lower resistance of colistin, a vancomycin-colistin combination could be an alternative empiric treatment in post-cataract endophthalmitis in India.

Delayed infections following polybutylate-coated polyester (Ethibond) suture frontalis suspension surgery for severe blepharoptosis.

PURPOSE: To describe the incidence and management of delayed infections following frontalis sling suspension with polybutylate-coated polyester suture (Ethibond). METHODS: Retrospective, interventional case series of 177 eyes of 150 patients, who underwent frontalis suspension surgery with Ethibond (Johnson and Johnson, USA), at L V Prasad Eye Institute, Hyderabad, India, between January 2016 and February 2022. Patients were assessed for chronic secondary sling infection, defined as infection/suture granuloma occurring beyond 6 weeks post-surgery. All patients received postoperative oral antibiotics. The clinical profile, microbiological evaluation, antibiotic sensitivity patterns, and management outcomes were analyzed. RESULTS: Delayed infection following Ethibond sling suspension was noted in 14 eyes of 13 patients (7.9%). Of these, eight cases (61.5%) were females. The average time interval from surgery to presentation was 7.5 months (range: 2.5 months to 2.5 years). Eleven eyes underwent sling removal. Of the three who received initial antibiotic course, two eventually needed sling removal. Microbiological evaluation was available in seven eyes (50%), with Staphylococcus aureus as the most common organism. Of the six positive cultures, five (83.3%) were resistant to fluoroquinolones (FQs). CONCLUSION: Frontalis suspension with Ethibond has an 8% incidence of delayed infections, with Staphylococcus aureus as the most common organism. The authors recommend early sling removal in all patients with infection and recommend consideration of an alternative material in the event of future revision surgery.

Infectious crystalline keratopathy-clinical-microbiological profile and outcome of management.

PURPOSE: To describe the clinical features, management, and long-term outcome of Infectious crystalline keratopathy (ICK). METHODS: The medical records of clinically diagnosed and microbiologically proven cases of ICK were reviewed from January 2011 to December 2022. Clinical characteristics include the presence of whitish needle-like projections with branching, limited to anterior-mid stroma. Keratoplasty being the most common risk factor, graft-related microbial keratitis during the same period was also studied. The demography, clinical profile, microbiology, treatment, and outcome were analyzed, and compared with secondary graft infiltrate(GI). RESULTS: Medical records of 24 cases with ICK were reviewed. The mean age was 49.3 ± 20.1 years, with 15(62.5%) males. Prior keratoplasty was done in 18 (75%) cases, with a mean graft size of 10.1 ± 1.5 mm, and mean interval between the last graft and presentation was 9.7 ± 6.2 (3-90) months. In comparison to GI (n = 24), ICK patients (n = 18,75%) were less symptomatic, presented late (7.3 ± 6.5 days vs 16.3 ± 19.4, p = 0.003), using frequent topical steroids (> 3 times/day, p = 0.006), smaller infiltrate size < 4 mm (p = 0.008), central (p = 0.02), less associated with epithelial defect (p = 0.0001), hypopyon (p = of 0.0002), corneal perforation (p = 0.0006), and surgical management (p = 0.03). On microbiology, 22 (91.6%) ICK cases were culture positive, 14 (63.6%) gram-positive, 3 (13.6%) gram-negative, 2 (9%) mixed bacteria, and 3 (13.6%) fungus, comparable with GI. CONCLUSION: ICK affects poor ocular surfaces usually following keratoplasty with larger graft size, the use of steroids being the most common association, and it responds to medical management as compared to GI.

Proteome profiling of Extracellular Vesicles in Pseudomonas aeruginosa endophthalmitis: Prognostic and therapeutic significance in a mouse model.

Endophthalmitis is a sight-threatening infection and a serious consequence of complications during intraocular surgery or penetrating injury of which Pseudomonas aeruginosa is an important etiology. Extracellular vesicles (EVs) have evolved as a promising entity for developing diagnostic and therapeutic biomarkers due to their involvement in intracellular communication and pathogenesis of diseases. We aimed to characterise the protein cargo of extracellular vesicles, isolated from a murine (C57BL/6) model of P. aeruginosa endophthalmitis by LC-MS/MS at 24 h post infection (p.i). EVs were extracted by ultracentrifugation, characterized by Dynamic Light Scattering (DLS) and western blotting with tetraspannin markers, CD9 and CD81 and quantified by the ExoCet quantification kit. Multiplex ELISA was performed to estimate the levels of TNF-α, IL-6, IFN-γ and IL-1ß. Proteomic analysis identified 2010 proteins (FDR ≤0.01) in EVs from infected mice eyes, of which 137 were differentially expressed (P-value ≤ 0.05). A total of 101 proteins were upregulated and 36 were downregulated. Additionally, 43 proteins were exclusive to infection set. KEGG and Gene Ontology revealed, Focal adhesion, Phagosome pathway, Complement cascade and IL-17 signalling pathway are crucial upregulated pathways involving proteins such as Tenascin, caveolin 1, caveolin 2, glutamine synthetase, microtubule-associated protein, C1, C8 and IL-17. Tenascin and caveolins are known to suppress anti-inflammatory cytokines further exacerbating the disease. The result of this study provides insight into the global extracellular vesicle proteome of P. aeruginosa endophthalmitis with their functional correlation and distinctive pattern of expression and tenascin, caveolin 1 and caveolin 2 are attractive biomarkers for P. aeruginosa endophthalmitis.

Proteomic profiling of exosomes in a mouse model of Candida albicans endophthalmitis.

Exosomes play pivotal roles in intercellular communication, and pathophysiological functions. In this study, we aimed to understand the role of exosomal proteome derived from C. albicans infected mice (C57BL/6) eyeball. Exosomes were characterized by Dynamic Light Scattering and Western blot, quantified and subjected to LC-MS/MS and cytokine quantification by ELISA. The average size of exosomes was 170-200 nm with number of exosomes amounted to 1.42 × 1010 in infected set compared to control (1.24 × 109). Western blot was positive for CD9, CD63 and CD81 confirming the presence of exosomes. IL-6, IL1ß, TNF-α, and IFN-γ levels were significantly elevated in infected eye at 72 h.p.i. Proteomic analysis identified 42 differentially expressed proteins, of these 37 were upregulated and 5 were downregulated. Gene Ontology (GO) revealed enrichment of cell adhesion, cytoskeleton organisation, and cellular response proteins such as aquaporin-5, gasdermin-A, CD5 antigen-like, Catenin, V-ATPase, and vesicle associated protein. Additionally, KEGG pathway analysis indicated the association of metabolic and carbon signalling pathways with exosomes from C. albicans infected eye. The protein cargo in exosomes released during endophthalmitis with C. albicans seems to play a unique role in the pathogenesis of the disease and further validations with larger cohort of patients is required to confirm them as biomarkers.

Microbiological spectrum and antibiotic susceptibility patterns of non-viral conjunctivitis over 16 years at a tertiary eye care center in Southern India.

PURPOSE: To determine the trends in the microbial spectrum and antibacterial susceptibility patterns of non-viral conjunctivitis over 16 years. METHODS: Microbiology data were reviewed from 2006-2021 for all the patients with clinically and culture-proven infectious conjunctivitis. Conjunctival swabs and/or conjunctival scrapings were collected for microbiological workup, and the demographic and antibiotic susceptibility data were extracted from the EMR (Electronic Medical Record). For statistical analysis, χ2-test was performed. RESULTS: Of the 1711 patients, 814 (47.57%) were culture positive and 897 (52.43%) were culture negative. Of the total culture-proven conjunctivitis cases, 775/814 (95.20%) were bacteria, and 39/814 (4.80%) were fungi. Among these bacterial isolates, 75.74% were gram-positive bacteria, while 24.26% were gram negative. The predominant gram-positive pathogens isolated were S. epidermidis (16.7%), S. aureus (17.9%) (p < 0.05), and S. pneumoniae (18.2%), while Haemophilus spp. (36.2%) (p < 0.05) was the most often isolated gram-negative bacteria (36.2%), and Aspergillus spp. was the most commonly isolated fungus (50%). The susceptibility of gram-positive bacteria to cefazoline increased from 90.46 to 98% (p = 0.01), whereas the susceptibility for gatifloxacin decreased in both gram-positive (81-41%; p < 0.0001) and gram-negative bacteria (73-58%; p = 0.02). CONCLUSIONS: Increasing resistance of ocular isolates to mainstay antibiotics is a concern, and this data can assist healthcare practitioners in making informed choices regarding the treatment of ocular infections with ophthalmic antibiotics.

Herpes simplex virus keratitis: electronic medical records driven big data analytics report from a tertiary eye institute of South India.

OBJECTIVE: To describe the demographics and clinical profile of Herpes Simplex Virus (HSV) Keratitis in patients presenting to a multi-tier ophthalmology hospital network in South India. METHODS: We have reviewed the medical records of all patients having a clinical diagnosis of any form of HSV keratitis, seen between May 2012 and August 2020 across the L V Prasad Eye Institute network. All the further analyses of the groups were performed using the keywords used for making the diagnosis of HSV keratitis and the data were collected from the electronic medical record system. RESULTS: There were a total of 8308 (N = 8897 eyes) patients. Male: female ratio was 5368 (64.61%):2940 (35.39%). Unilateral involvement was in 7719 (92.91%) patients. The most common age group affected was between the third to fifth decades of life with 1544 (18.58%). 3708 (1.68%) eyes had mild visual impairment (< 20/70) while the rest of them had moderate to severe visual impairment as observed mainly (p ≤ 0.01) in Necrotizing stromal keratitis. 7314 (82.21%) eyes had normal intraocular pressure (10-21 mm Hg) while raised most commonly in keratouveitis (P ≤ 0.01). Epithelial Keratitis, Immune Stromal Keratitis, Endotheliitis, Neurotrophic keratopathy and Keratouveitis were observed in 1875 (17.22%) eyes, 5430 (61.03%) eyes, in 129(1.45%) eyes, 1188 (13.35%) eyes, 148 (1.66%) eyes and 256 (2.88%) eyes respectively. CONCLUSION: Based on our institute-based data, the most common type of HSV keratitis is Immune stromal keratitis followed by epithelial keratitis. Although not representative of the general population, this data provide useful insights related to HSV keratitis from India.

Molecular diagnosis of rhino-orbital mucormycosis in a COVID-19 setting.

PURPOSE: Mucormycosis is a severe fungal infection caused by species of the order Mucorales. Early and accurate diagnosis is a prerequisite in the management of the disease. In the present study, we evaluated and compared two PCR-based techniques for the diagnosis and identification of mucormycosis in patients with rhino-orbital mucormycosis (ROM) post-COVID-19. METHODS: Diagnosed clinically and radiologically, 25 patients of ROM were included in the study and endoscopically or blind collected nasal swabs or orbital tissues were submitted for microbiological evaluation (direct microscopy + culture) and PCR using primers targeting two different loci (ITS and 28S rDNA region) for diagnosis. All PCR products were further processed for species identification using Sanger sequencing whenever possible. RESULT: Of the 25 samples included in the study, 16 samples were positive for presence of fungal filaments by Smear suggestive of Mucorales sp., but only 7/25 grew in culture. ITS-based PCR was able to identify mucormycosis in 7/25 (28%) samples and 28S rDNA PCR showed positivity for 19/25 (76%) samples. Rhizopus oryzae was found to be the predominant species in our study. The sensitivity and specificity of 28S rDNA PCR compared to culture were found to be 85.71% and 27.78%, respectively, while for ITS-based PCR, they were 42.86% and 77.78%, respectively. CONCLUSIONS: 28S rDNA-based PCR is a reliable and sensitive method for early diagnosis of mucormycosis. Molecular techniques have shown a promising future to provide quick and effective treatment by accurately identifying the aetiologic agent.

Exosomes, extracellular vesicles and the eye.

Exosomes are a subset of extracellular vesicles which accommodate a cargo of bioactive biomolecules that generally includes proteins, nucleic acids, lipids, sugars, and related conjugates depicting the cellular environment and are known to mediate a wide array of biological functions, like cellular communication, cellular differentiation, immunomodulation, neovascularization, and cellular waste management. The exponential implication of exosomes in the pathological development and progression of various disorders including neurodegenerative diseases, cardiovascular diseases, and cancer has offered a tremendous opportunity for exploring their role in ocular conditions. Ocular diseases such as age-related macular disease, glaucoma, infectious endophthalmitis, diabetic retinopathy, autoimmune uveitis etc face various challenges in their early diagnosis and treatments due to contributing factors such as delay in the onset of symptoms, microbial identification, difficulty in obtaining samples for biopsy or being diagnosed as masquerade syndromes. Studies have reported unique exosomal cargos that are involved in successful delivery of miRNA or proteins to recipient cells to express desired expression or exploited as a diagnostic marker for various diseases. Furthermore, engineered exosomes can be used for targeted delivery of therapeutics and exosomes being natural nanoparticles found in all types of cells, host may not elicit an immune response against it. With the rapid advancement of opting personalized therapeutics, extending exosomal research to sight-threatening ocular infections can possibly advance the current diagnostic and therapeutic approaches. This review briefs about the current knowledge of exosomes in visual systems, advancements in exosomal and ophthalmic research, participation of exosomes in the pathogenesis of common ocular diseases, the challenges for exosomal therapies along with the future of this promising domain of research for diseases that fatally threaten billions of people worldwide.

Methicillin resistance in Staphylococcus aureus modulates the transcriptome and disease pathology in a murine model of endophthalmitis.

The ever-increasing incidence of methicillin-resistant strains of Staphylococcus aureus (MRSA) endophthalmitis is of particular concern as they are associated with poor outcomes. To compare the histology and whole transcriptome of Methicillin resistant (MRSA) and Methicillin-susceptible (MSSA) Staphylococcus aureus endophthalmitis in an experimental murine model. MRSA and MSSA endophthalmitis was induced in C57BL/6 mice and disease progression was scored clinically and histologically at 24 h p.i. Retinal changes were monitored by H&E, CD45, MPO and GFAP staining followed by retinal cell death evaluation. Whole Transcriptome was analysed using the SuperPrint G3 Mouse Gene Expression v2 chip. Differential gene expression analysis (Limma package, R) was done followed by enrichment of pathways (KEGG database). Increased corneal haze, diminished vitreous clarity and red reflex was observed in MRSA infected mice eye compared to MSSA (p = 0.04). Histological assessment also corroborated with increased disease severity in MRSA (p = 0.02). Although MRSA infected eye displayed higher CD45+ cells and greater GFAP intensity, the difference was not statistically significant. However, higher retinal cell death was found to be associated with the MRSA infection (p = 0.007). Our study also revealed that MRSA infection induces changes in host transcriptome (FC = 1.5, p = 0.05), revealing the involvement of several interleukins (IL-11,15,10,1ra), chemokines (CCL-11, CXCL-1), Interferon receptors, GM-CSF, M-CSF, MMPs, Neruopilin2 (NRP-2), Ubiquitin associated peptidase and apoptotic ligands. ErbB signalling, JAK-STAT, adipocytokine and Ras signalling were the top divergently enriched pathways. Our study confirms the differential host immune response triggered by MRSA infection in the eye. Our study may help to elucidate the mechanisms of pathogenesis and to identify additional candidate drug targets for the treatment of MRSA endophthalmitis.

Proteomic profiling of aspergillus flavus endophthalmitis derived extracellular vesicles in an in-vivo murine model.

Extracellular Vesicles (EVs) play pivotal roles in cell-to-cell communication, and are involved in potential pathological and physiological cellular processes. The aim of this study was to understand the proteomic cargo of these vesicles, in a murine model of Aspergillus flavus (AF) endophthalmitis. EVs were isolated from A. flavus infected C57BL/6 mice eyes by differential ultracentrifugation at 24 h post infection (p.i) and isolated EVs were characterized by Dynamic Light Scattering (DLS), Scanning Electron Microscopy (SEM), Exocet assay, and western blot. Proteomic profiling of EVs was then evaluated by mass spectrometry (LC-MS/MS) and compared it with control uninfected mice. The average size of the EVs were 180-280 nm by DLS and the number of EVs increased to 1.55 × 1010 in infected mice in comparison to EVs from uninfected eye (1.24 × 109). Western blot was positive for CD9, CD63, and CD81 confirming the presence of EVs. LC-MS/MS analysis, identified 81 differentially expressed proteins, of these 22 were up-regulated and 59 were down-regulated. Gene Ontology (GO) analysis revealed enrichment of lipid metabolism, protein complex binding, and transferase activity, and the proteins associated were Aquaporin-5, CD177 antigen, Solute carrier family-25, and Calcium/calmodulin-dependent protein kinase. Additionally, KEGG pathway analysis indicated that glucagon signalling, metabolic, and PPAR signalling pathway were significantly associated with EVs from A. flavus infected mice eyes. The protein cargo in EVs from A. flavus endophthalmitis provides new insights into the pathogenesis of fungal endophthalmitis and validation of these proteins can serve as diagnostic and/or prognostic biomarkers for patients with a clinical suspicion of fungal endophthalmitis. LAY SUMMARY: EVs play an important role in cell communication. In our study proteomic profiling of EVs isolated from A. flavus infected mice provided new insights into the understanding of the pathobiology of A. flavus endophthalmitis and validation of these proteins can serve as biomarkers.

CULTURE POSITIVITY RATES OF DEEP VITREOUS BIOPSY UNDER AIR VIS-À-VIS CONVENTIONAL ANTERIOR VITREOUS BIOPSY IN ENDOGENOUS ENDOPHTHALMITIS.

PURPOSE: To describe the comparison of the culture positivity rates of deep vitreous biopsy under air and conventional anterior vitreous biopsy in endogenous endophthalmitis. MATERIALS: A retrospective, consecutive, comparative series including cases of endogenous endophthalmitis from January 2014 to January 2021. They were divided into those where conventional anterior vitreous biopsy was taken and those where a deep biopsy was taken under air (DBA group). The culture positivity rate among the two groups was compared, and factors affecting the same were analyzed by a regression analysis. RESULTS: There were 140 eyes in the conventional anterior vitreous biopsy group and 44 eyes in the DBA group. Sex, age, duration of symptoms, underlying systemic illness, presenting vision in logMAR, and total number of vitreous interventions were comparable between the two groups. Visual outcome was better in patients undergoing DBA as compared with those who underwent conventional anterior vitreous biopsy , (1.28 ± 1.01 logMAR, 20/380 vs. 1.88 ± 1.33 logMAR, 20/1,500, P = 0.03). No case in the DBA group developed retinal detachment or any complication attributable to the technique. In the conventional anterior vitreous biopsy group, the culture positivity was seen in 43/140 samples (30.71%), and in the DBA group, it was noted in 29/44 samples (65.9%). Taking a DBA was the only factor that was significant both in bivariate (odds ratio 4.36, 95% confidence interval 2.12-8.95, P < 0.0001) and multivariate analysis (odds ratio 4.02, 95% confidence interval 3.1-7.43, P < 0.0001). CONCLUSION: Vitreous sampling can be performed from the deep vitreous cavity in endogenous endophthalmitis safely and effectively under air infusion, improving the culture positivity rates.

CLINICAL PRESENTATIONS, MICROBIOLOGY, AND FACTORS AFFECTING MANAGEMENT OUTCOMES IN LENS ABSCESS WITH CONCURRENT ENDOPHTHALMITIS.

PURPOSE: To describe the clinical presentations, microbiology, and factors affecting management outcomes in lens abscess with concurrent endophthalmitis. MATERIALS: A retrospective, consecutive, noncomparative series including cases of endophthalmitis with concurrent lens abscess from January 2017 to May 2021. RESULTS: This study included 102 eyes, predominantly male (71.6%). All cases were posttrauma. The mean age noted was 30.47 ± 19.51 years. Presenting vision was logMAR 3.02 ± 0.74 (median 3.5, Snellen 20/63245). A favorable anatomical outcome was seen in 63 eyes (61.8%), while a final favorable functional outcome was seen in 51 eyes (50%). The mean follow-up duration was 10.04 ± 10.87 months (median 4.5). Final vision was logMAR 2.13 ± 1.32 (median 2.7, Snellen 20/10023) (P < 0.0001). The mean follow-up duration was 10.04 ± 10.87 months (median 4.5). Increasing age (OR 1.04, P = 0.02), female sex (OR 7.91, P = 0.007), initial intervention of vitrectomy instead of limited vitreous biopsy (OR 11.72, P = 0.009), and a negative vitreous culture (OR 14.28, P = 0.0004) predicted a favorable anatomical outcome. Absence of a corneal infiltrate (OR 11.11, P = 0.003) and initial intervention of vitrectomy instead of a limited vitreous biopsy (OR 21.96, P < 0.0001) predicted a favorable functional outcome. Culture positivity was seen in 56.9% of the cases. Gram-positive organisms were predominant (n = 41) followed by Gram-negative organisms (n = 12) and fungi (n = 10). CONCLUSION: Lens abscess can present concurrently with traumatic endophthalmitis. Management should involve the complete removal of the crystalline lens along with a pars plana vitrectomy for optimal outcomes.

Retrobulbar Hyaluronidase Injection: Does It Reach the Eye?

PURPOSE: To determine if retrobulbar injection of hyaluronidase reaches the vitreous cavity, and to determine its concentration in the vitreous. METHODS: Prospective case-control study. Patients undergoing evisceration with implant for noninfective blind eyes were enrolled in the study. Before the evisceration, a retrobulbar injection of 3,000 IU of hyaluronidase (2 ml) was injected. Time from injection to in vivo sampling of posterior vitreous was noted. Vitreous samples from controls were obtained from patients undergoing vitrectomy for retinal detachment or diabetic retinopathy. Concentration of hyaluronidase was assessed in all 30 samples. An ELISA-based microtiter-technique was used to evaluate the activity of hyaluronidase by an avidin-peroxidase-based procedure using an ELISA reader. Incubations were carried out at room temperature and at 37°C. All the samples were analyzed in duplicates, and the mean of each sample was plotted on a scatter plot. RESULTS: Total of 30 vitreous samples were analyzed, of which 15 were controls and 15 were test samples. Of the 15 test samples, injection-to-sampling time was 0 to 20 minutes in 4 samples, 20 to 40 minutes in 6 samples, and 40 to 60 minutes in 5 samples. The highest concentration of hyaluronidase detected in control and test samples were 2.9 and 3.0 µg/ml, and the lowest concentration was 1.7 and 1.5 µg/ml (SD 0.3), respectively. There was no significant difference between control and test groups. CONCLUSION: Retrobulbar injection did not result in higher concentration of hyaluronidase in the posterior vitreous compared with controls when measured up to 60 minutes following injection.

Immunopathology of COVID-19 and its implications in the development of rhino-orbital-cerebral mucormycosis: a major review.

PURPOSE: To present a literature review on various immunopathologic dysfunctions following COVID-19 infection and their potential implications in development of rhino-orbital-cerebral mucormycosis (ROCM). METHODS: A literature search was performed via Google Scholar and PubMed with subsequent review of the accompanying references. Analogies were drawn between the immune and physiologic deviations caused by COVID-19 and the tendency of the same to predispose to ROCM. RESULTS: Sixty-two articles were reviewed. SARS-CoV-2 virus infection leads to disruption of epithelial integrity in the respiratory passages, which may be a potential entry point for the ubiquitous Mucorales to become invasive. COVID-19 related GRP78 protein upregulation may aid in spore germination and hyphal invasion by Mucorales. COVID-19 causes interference in macrophage functioning by direct infection, a tendency for hyperglycemia, and creation of neutrophil extracellular traps. This affects innate immunity against Mucorales. Thrombocytopenia and reduction in the number of natural killer (NK) cells and infected dendritic cells is seen in COVID-19. This reduces the host immune response to pathogenic invasion by Mucorales. Cytokines released in COVID-19 cause mitochondrial dysfunction and accumulation of reactive oxygen species, which cause oxidative damage to the leucocytes. Hyperferritinemia also occurs in COVID-19 resulting in suppression of the hematopoietic proliferation of B- and T-lymphocytes. CONCLUSIONS: COVID-19 has a role in the occurrence of ROCM due to its effects at the entry point of the fungus in the respiratory mucosa, effects of the innate immune system, creation of an environment of iron overload, propagation of hyperglycemia, and effects on the adaptive immune system.

Microbiological profile and antibiotic susceptibility trends in orbital cellulitis in India: an analysis over 15 years.

PURPOSE: To report the microbiologic profile and antibiotic susceptibility trends in orbital cellulitis. METHODS: Retrospective review of microbiology records of orbital cellulitis between 2005 and 2019. Orbital pus or conjunctival swab underwent culture for bacteria and fungi and antibiotic susceptibility testing for bacterial isolates. The microbiological profile and trends in antibiotic susceptibility were analyzed over the three study periods: 2005-2009, 2010-2014, and 2015-2019. RESULTS: Of the 203 patient samples, 189 (93%) were orbital pus samples, and 146 (72%) were culture positive. Organisms included bacteria (167/203, 82.3%), fungi (13/203, 6.4%), and mixed infection (25/203, 10.3%). Among bacteria, 79% were gram positive, with Staphylococcus aureus and Streptococcus species being commonest, and 21% were gram negative, with Pseudomonas aeruginosa and Enterobacteriaceae group being the commonest. Aspergillus flavus was the most common fungus isolated. Trend analysis revealed no change in the number of sterile cases and fungal cellulitis. Increase in gram positive bacteria was statistically significant (p = .0002) between 2005-2009 and 2015-2019. The increase in gram negative bacteria was statistically significant (p = .047) between all three time periods. Susceptibility patterns showed increasing trend of resistance to fluoroquinolones, that reached statistical significance for Ciprofloxacin, Moxifloxacin and Gatifloxacin (p < .05). Sterile sample was not found in any of the pediatric (0-16 years) cases (n = 55), compared to 28% in adults. CONCLUSION: There was a significant rise in gram positive and negative orbital infections over the 15 year period, with increased resistance to fluoroquinolones. Fungal cellulitis and sterile samples showed a steady trend. Orbital aspirate provides accurate detection of the causative organism.

Clinical profile of keratitis treated within 3 months of acute COVID-19 illness at a tertiary care eye centre.

PURPOSE: To report the spectrum of keratitis treated within 3 months of acute COVID-19 infection. METHODS: Retrospective, descriptive case series study of 19 eyes of 16 patients who presented at tertiary eye care centre in Southern India. RESULTS: Median age of the patients was 43(IQR 35-55.5) years. Majority (15/16, 93.75%) were males. Unilateral affliction was predominant (13/16, 81.25% patients). Nine had a history of hospitalization, five had received oxygen supplementation and five had been treated with steroids during COVID-19 illness. The median duration between COVID-19 diagnosis and the ocular symptoms in the eye was 29 (IQR 22-57) days. Microbiological diagnosis consisted of microsporidia in nine eyes of seven patients, fungus in six patients, Pythium in one patient, and herpes zoster ophthalmicus in one patient. One patient had neurotrophic keratitis. Therapeutic penetrating keratoplasty was performed in five patients, glue application in two patients and three were managed with tarsorrhaphy with/without amniotic membrane grafting or tenonplasty. There was medical and surgical cure in all patients. CONCLUSIONS: Microsporidia was the commonest cause of keratitis, followed by fungal infection. Majority of the microsporidia infections were keratoconjunctivitis. The fungal isolates identified were Aspergillus and Mucor species. All patients responded to conventional management guidelines with favourable outcomes.

Clinical profile and microbiological trends of therapeutic keratoplasty at a network of tertiary care ophthalmology centers in India.

OBJECTIVE: To describe the clinical profile and microbiology trends in patients undergoing therapeutic keratoplasty at a multi-tier ophthalmology hospital network in India. METHODS: This cross-sectional hospital-based study included 3147 eyes undergoing therapeutic keratoplasty between Jan 2016 and Dec 2020 (5 years period) as cases. The demographic data, clinical profile and microbiological analysis were collected using an electronic medical record system. RESULTS: Of a total of 13,625 eyes with microbial keratitis(non-viral), 3147 (23.1%) underwent therapeutic keratoplasty during the study duration. Majority of patients were males (68.35%), from a rural geography (49.89%) and in 51-60 years age bracket (23.74%). The mean age of the patients was 50.54 ± 15.83 years. Two-thirds of patients were from lower socio-economic strata (66.63%) with an agrarian background (36.51%). Of the 3,147 eyes, fungus (51.8%) was the most common indication of therapeutic keratoplasty followed by bacteria (16.87%) and parasite (1.27%). No organisms could be identified in about a third (29.33%) of the cases. Between 2016 and 2020, the trend of therapeutic keratoplasties for fungal infections steadily grew (39.9% vs 45.49%) while the bacterial infections showed a steady decline (23.15% vs 11.81%). CONCLUSIONS: Medical cure rate was seen in majority of those with microbial keratitis, and 23.1% eventually required management with therapeutic keratoplasty. Fungal keratitis was the most common indication for therapeutic keratoplasty. Male gender, rural setting, low socio-economic background and agricultural occupation are common risk factors for patients undergoing therapeutic keratoplasty in India.