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Background-Microbiological investigations of vitreous fluid (VF) and aqueous humour (AH) specimens have often failed to detect the infecting agent in infectious endophthalmitis, resulting in a clinical dilemma regarding therapy. In this study, the polymerase chain reaction (PCR) was evaluated in the diagnosis of bacterial and Propionibacterium acnes endophthalmitis. Methods-58 intraocular specimens (30 VF and 28 AH) from 55 cases of endophthalmitis and 20 specimens (14 VF and 6 AH) as controls from non-infective disorders were processed for microbiological investigations. Nested PCR directed at the 16S rDNA using universal primers for eubacterial genome was done. PCR for P acnes was performed on specimens microbiologically negative by conventional techniques but eubacterial genome positive. Results-Of the 20 controls from noninfective cases, one (5%) was positive using eubacterial primers and none with P acnes primers. PCR for eubacterial genome showed 100% correlation with 20 (34.5%) bacteriologically positive specimens. Eubacterial genome, was detected in 17 (44.7%) of 38 bacteriologically negative specimens and nine (52.9%) out of the 17 were positive for P acnes genome. Among the 21 eubacterial PCR negative specimens, seven were fungus positive. By inclusion of PCR, microbiologically positive specimens increased from 46.5% to 75.8%. PCR on AH was as sensitive as that on VF for the detection of both eubacterial and the P acnes genome. Conclusion-PCR performed on AH and VF is a reliable tool for the diagnosis of bacterial and P acnes endophthalmitis particularly in smear and culture negative specimens.

Eales' disease, first described by the British ophthalmologist Henry Eales in 1880, is characterized by three overlapping stages of venous inflammation (vasculitis), occlusion, and retinal neovascularization. Diagnosis is mostly clinical and requires exclusion of other systemic or ocular conditions that could present with similar retinal features. In recent years, immunological, molecular biological, and biochemical studies have indicated the role of human leukocyte antigen, retinal autoimmunity, Mycobacterium tuberculosis genome, and free radical-mediated damage in the etiopathogenesis of this disease. However, its etiology appears to be multifactorial. The management depends on the stage of the disease and consists of medical treatment with oral corticosteroids in the active inflammatory stage and laser photocoagulation in the advanced retinal ischemia and neovascularization stages.

Chronic suppurative lacrimal canaliculitis is an important cause of ocular surface discomfort. Treatment with topical antibiotics is often inadequate and surgical treatment by canaliculotomy and canalicular curettage has been the mainstay of treatment in the past. The role of canalicular antibiotic irrigation has been inadequately studied. We report the clinical features, microbiological profile and treatment outcome in a series of 12 patients with suppurative lacrimal canaliculitis. Two patients had Actinomyces infection, five had Nocardia infection and seven patients had polymicrobial infection. Three patients had resolution of canaliculitis on combination broad-spectrum topical antibiotic therapy using ciprofloxacin and fortified cefazolin. In nine patients, topical antibiotic therapy was combined with canalicular irrigation using fortified cefazolin. All patients had excellent resolution of canaliculitis without the need for surgical treatment. Availability of broad-spectrum antibiotics and canalicular irrigation may offer an alternative to surgery in the management of suppurative lacrimal canaliculitis.

Purpose: To standardize in-vitro antifungal susceptibility testing by agar dilution method to find out the minimum inhibitory concentration (MIC) of amphotericin B, fluconazole and ketoconazole on ocular fungal isolates. Methods: A total of 180 ocular fungal isolates (130 filamentous fungi and 50 yeasts) were included. The antifungal drugs such as amphotericin B (0.0625-8 µg/mL), fluconazole (0.2-819.6 µg/mL) and ketoconazole (0.025-6.4 µg/mL) were incorporated in doubling dilutions in the yeast nitrogen base medium. The MIC was determined as the lowest concentration of the antifungal drug preventing growth of macroscopically visible colonies on drug containing plates when there was visible growth on the drug -free control plates. Results: All 50 ocular isolates of yeast were susceptible to amphotericin B, while two (4%) and five (10%) strains were resistant to fluconazole and ketoconazole respectively. Of the 130 filamentous fungi tested, six (4.6%) were resistant to amphotericin B, 49 (37.7%) and 10 (7.6%) were resistant to fluconazole and ketoconazole respectively. Percentile 50 (MIC 50) and Percentile 90 (MIC 90) for all the three antifungal agents were calculated. Aspergillus niger, Aspergillus terreus and Candida krusei were found to be resistant to fluconazole and ketoconazole. Conclusion:This technique was found to be reliable, cost effective and easy to perform with consistent results.

Use of polymerase chain reaction in detection of Mycobacterium tuberculosis complex DNA from vitreous sample of Eales' disease

Background:The objective of the study was the determination of the incidence of culture-proven postoperative endophthalmitis and probable sources of infection.Materials and Methods:It was a prospective study on the microbiology, incidence and probable sources of infection in patients with postoperative infectious endophthalmitis carried out in a tertiary care eye hospital. Consecutive patients diagnosed with postoperative infectious endophthalmitis during the years 2000-2007 were investigated for the causative infective agent and possible sources of infection. The surgical data and microbiological data including the investigations performed to trace the source were recorded in a specific formatted form and were gathered and compiled for analysis.Results:Data of analysis showed that 98 (0.042%) out of 2,31,259 patients who underwent intra-ocular surgery developed infectious endophthalmitis. Among these, 70 (0.053%) occurred after cataract, 10 (0.5%) after penetrating keratoplasty (PK) and 18 (0.018%) following other types of intra-ocular surgeries. The predominant infectious agents isolated were bacteria (89.7%), with equal proportions of Gram-positive and Gram-negative bacteria. Polymicrobial infection was noted in four and fungi in seven patients. Occurrence of postoperative endophthalmitis was sporadic and not related to any specific part of period in a year. Sources of infection were donor corneal rim in six post-PK patients and phaco probe in one who had postphacoemulsification endophthalmitisConclusions:Overall incidence of postoperative endophthalmitis over an 8-year period was quite low. The sources of infection could be established in six post-PK endophthalmitis patients and in a postcataract surgery.

We evaluated the efficacy of autologous expanded corneal epithelial cell transplants derived from harvested limbal biopsy cultured on a thermoreversible polymer (Mebiol Gel) for the management of unilateral limbal stem cell deficiency (LSCD). Corneal limbal biopsies from 12 rabbits were cultured on a thermoreversible polymer Mebiol Gel at 37 degrees C. Cells were harvested from the dishes after 3 weeks by reducing temperature to 4 degrees C. Autologous transplantation was undertaken to reconstruct the experimentally induced limbal stem cell deficiency in the rabbit eyes. The corneas of both eyes of all rabbits were harvested later for molecular studies. Reparative surgery was a total success in seven rabbits, partial success in two, and failure in three eyes. Histology of the seven successful eyes showed the successful growth of the corneal epithelium. Immunohistochemistry and reverse transcriptase polymerase chain reaction showed the cornea phenotype and stem cell-associated markers in the limbus of the seven successful eyes, indicating the homing of these cells into limbus. In the three failure cases and in the two control rabbit eyes, used in the study, histology showed presence of goblet cells and vascularization in the stroma with abortive formation of corneal epithelium. Our results suggest that transplantation of autologous limbal epithelial cells grown in thermoreversible polymer Mebiol Gel may restore a nearly normal ocular epithelial surface in eyes with unilateral LSCD.