Species identification, antifungal susceptibility profiles and biofilm formation attributes of Rhodotorula isolates from ocular infections.

BACKGROUND: Members of genus Rhodotorula are widely distributed in nature and have been traditionally considered non-pathogenic. Last few decades have seen the yeast as an emerging pathogen. We observed increase in numbers of Rhodotorula isolates from ocular infections in last few years, thus this prospective study was planned. OBJECTIVES: To identify the species of Rhodotorula isolates from ocular infections. To know the antifungal susceptibilities and study the biofilm formation attributes of the isolates. MATERIALS AND METHODS: Rhodotorula isolates were speciated using conventional methods, Matrix Assisted Laser Desorption and Ionisation - Time of Flight (MALDI- TOF) and sequencing of ITS region of ribosomal DNA. Antifungal susceptibility testing (AFST) was done using disc diffusion and E-test. Biofilm formation was studied using XTT [2,3-bis (2-methoxy-4-nitro-5-sulfo-phenyl)-2H-tetra-zolium-5-carboxanilide] assay. RESULTS: Twenty four isolates (92.3%) were identified as R. mucilaginosa and two as R. Minuta. AFST showed high MICs against Fluconazole, Amphotericin-B, Caspofungin, Micafungin and Flucytosine; MIC distribution from low to very high against Voriconazole, Itraconazole and Natamycin; and very low MICs against Posaconazole 57.7% of isolates were strong biofilm producers, 23.1% were moderate, and 19.2% were non producers. CONCLUSIONS: This is the first prospective study on species distribution, antifungal susceptibility and biofilm production attributes of Rhodotorula isolates from ocular infections; also first time demonstrating the utility of proteomics based MALDI-TOF in diagnosing Rhodotorula up to species level. The study has shown high MICs against the conventional azoles, Amphotericin-B and Flucytosine. However, low MICs against Posaconazole and Natamycin give a hope for their possible therapeutic use.

Chlamydia and ocular adnexal lymphomas: An Indian experience.

CHLAMYDIA AND OCULAR ADNEXAL LYMPHOMAS: AN INDIAN EXPERIENCE: Ocular adnexal lymphomas (OALs) are a heterogeneous group of malignancies, majority being extranodal mucosa-associated lymphoid tissue (MALT) type. Different geographical regions have reported association of Chlamydia with OALs (MALT type). In India, role of Chlamydia in OALs remains unexplored. The aim of this study was to detect Chlamydia and to correlate with clinicopathological features of OALs in India. The clinicopathological features of 41 OAL cases were studied prospectively. Chlamydia DNA was detected by genus specific PCR amplifying major outer membrane protein (MOMP) gene followed by DNA sequencing. Chlamydia immunoexpression was evaluated by immunofluorescence and immunohistochemistry. The results were correlated with clinicopathological features including follow-up and survival. Chlamydia genome was detected in 3/41 (7.3%) OAL cases by PCR. Direct sequencing revealed C. trachomatis in 3 positive cases. Immunofluorescence and immunohistochemistry showed Chlamydia antigen in 5/41 and 1/41 cases respectively. Immunofluorescence demonstrated higher sensitivity than immunohistochemistry. A significant association was observed between Chlamydia positivity and orbital location (P=0.05). Follow-up revealed relapse in 2 Chlamydia positive cases (P=0.056). Our results demonstrate for the first time presence of C. trachomatis genome in 7.3% OAL cases in India. As no other reports are documented, more detailed studies from different regions within India are needed to explore status of Chlamydia in OALs.

Clinical implications of microbial biofilms in chronic rhinosinusitis and orbital cellulitis.

BACKGROUND: Discovery of sessile mode of microbial existence (Biofilm state) focussed much interest, during the recent years, on the study of biofilms in many recurring and chronic infections. However, the exact role of microbial biofilms in chronic rhinosinusitis and orbital cellulitis were not elucidated earlier. The purpose of the present study was to look for the adherent property and biofilm producing ability of the clinical isolates in chronic rhinosinusitis and orbital cellulitis, and to look for the effects of antimicrobial agents on these biofilms by colorimetric assay and ultrastructural analysis. METHODS: Organisms were isolated and identified from various clinical samples in patients with chronic sinusitis and orbital cellulitis. Antimicrobial sensitivity testing was carried out by the standard protocol. Biofilms were developed; quantified and antimicrobial drug perfusion through the biofilm model was evaluated by the earlier devised procedure. Electronmicroscopic study of the biofilm was performed by the recommended technique. RESULTS: Of the total of 70 clinical samples processed, 48 i.e. 68.5 % grew bacteria and 13 i.e.(18.6 %) fungi. Staphylococcus aureus (20), S epidermidis (16) and Pseudomonas aeruginosa (6) accounted for the majority of the bacterial isolates. Aspergillus flavus (8), however was the commonest amongst the fungi. A total of 40 bacteria and 8 fungi could be tested for biofilm production. Eighteen (45 %) of the 40 bacterial isolates and 4(50 %) out of the 8 A flavus isolates were found to be biofilm producers. In vitro adherence testing revealed that majority i.e. 16 (88.8 %) of the 18 biofilm positive bacteria were adherent to artificial surfaces. Antimicrobial drug perfusion through the biofilm model was poor. Antimicrobial treatment was totally ineffective against strong biofilm producers, whose electron microscopic picture was quite similar to that observed for biofilm producers without any antimicrobial pre-treatment. CONCLUSIONS: Filamentous fungi, like bacteria were capable of forming biofilms, which could be one of the important virulence factors in determining the pathogenic potential of these organisms in causing chronic rhinosinusitis and orbital cellulitis.

Clinical Profile, Antibiotic Resistance and Outcomes in Bacterial Endophthalmitis: Coagulase-Negative Staphylococcus Endophthalmitis as Compared to Other Organisms.

BACKGROUND: To evaluate the clinical profile, outcomes and antibiotic resistance in bacterial endophthalmitis. METHODS: This was a post-hoc analysis of a study conducted at a tertiary centre, where 60 consecutive cases of culture-proven bacterial endophthalmitis were included prospectively. Group 1 included coagulase-negative Staphylococcus endophthalmitis (CNSE), while group 2 included the remaining cases. Clinical features, antibiotic resistance and visual outcomes were compared. Visual acuity >3/60 at six months of follow-up was defined as a good visual outcome. RESULTS: Group 1 had 31 cases, while group 2 had 29. Group 2 included 12 gram-positive and 17 gram-negative isolates. Among the groups, group 2 had more patients with presenting visual acuity below hand motions close to the face (25 vs. 12, p<0.001), poor visual outcomes (26 vs. 3, p<0.001) and retinal detachment (RD) (10 vs. 2, p=0.007). Pseudomonas was most commonly resistant to antibiotics, and ceftazidime (p=0.005) and cefazolin (p=0.009) resistance were higher in group 2 isolates. In group 1, five isolates were resistant to any one of the antibiotics, whereas in group 2, 13 isolates were resistant to any one of the antibiotics (p=0.024). CONCLUSIONS: In the current study, eyes in the group of endophthalmitis caused by CNSE achieved better visual acuities at the last follow-up compared to eyes with endophthalmitis caused by other bacteria. Antibiotic resistance in isolates other than CNSE is a cause of concern.

Comparative evaluation of topical versus intrastromal voriconazole as an adjunct to natamycin in recalcitrant fungal keratitis.

OBJECTIVE: To compare the efficacy of topical voriconazole and topical natamycin with that of intrastromal voriconazole and topical natamycin in patients with recalcitrant fungal keratitis. DESIGN: Randomized clinical trial. PARTICIPANTS: Forty eyes of 40 patients with fungal keratitis (positive smear or culture results or both) larger than 2 mm, involving up to two thirds of the stromal depth, and not responding to topical natamycin therapy for 2 weeks were recruited. INTERVENTION: The patients were randomized to receive either topical 1% voriconazole therapy (n = 20) or intrastromal injections of voriconazole 50 µg/0.1 ml (n = 20). The patients in both groups continued topical natamycin 5% every 4 hours until the ulcer healed. MAIN OUTCOME MEASURES: Primary outcome measure was best spectacle-corrected visual acuity (BSCVA) 3 months after intervention, and secondary outcome measures were time to healing and the size of the scar. RESULTS: The patients in both groups had comparable baseline parameters. The mean BSCVA after treatment was 1.295 ± 0.5 logarithm of the minimum angle of resolution (logMAR) units in the topical group and 1.692 ± 0.29 logMAR units in the intrastromal group. The visual acuity after treatment was significantly better in the topical voriconazole group (P = 0.008). Nineteen patients receiving topical voriconazole and 16 patients who were given intrastromal voriconazole healed with therapy. CONCLUSIONS: Topical voriconazole seems to be a useful adjunct to natamycin in fungal keratitis not responding to topical natamycin. Intrastromal injections did not offer any beneficial effect over topical therapy.

Coinfection with Acanthamoeba and Pseudomonas in contact lens-associated keratitis.

PURPOSE: To report coinfection with Acanthamoeba and Pseudomonas aeruginosa in a case with contact lens-associated keratitis. CASE REPORT: A 20-year-old woman presented to the emergency department of our hospital with a 4-day history of progressively increasing pain, redness, photophobia, mucopurulent discharge, and diminution of vision in her right eye. She was being treated for contact lens-related Pseudomonas keratitis in another hospital before presentation. Gram stain of corneal scrapings revealed gram-negative bacilli. Both Gram stain and 10% KOH wet mount showed the presence of Acanthamoeba cysts. Microbiological cultures obtained from contact lenses and contact lens storage case showed the presence of Pseudomonas aeruginosa and Acanthamoeba. Topical therapy was started in the form of hourly gentamycin 1.3%, cefazolin 5%, chlorhexidine 0.02%, propamidine 0.1%, polymyxin B 30,000 IU eye drops, and neosporin (neomycin, bacitracin, polymyxin) eye ointment four times a day. Symptomatic improvement was observed within 48 hours, along with a decrease in the density of infiltrates and a reduction in the anterior chamber reaction. Repeat corneal scrapings on day 10 showed Acanthamoeba but no bacilli. Progressive resolution of the infiltrate was noted during the next few days. Epithelialization was complete by day 24, following which the amoebicidal therapy was tapered during the next 4 weeks. Complete resolution of keratitis was achieved after 7 weeks of treatment. CONCLUSIONS: Both P. aeruginosa and Acanthamoeba are potentially devastating causes of microbial keratitis. Our case highlights the importance of considering the possibility of a concurrent infection in cases with contact lens-related keratitis.

Evaluation of liposomal amphotericin B for the treatment of fungal keratitis in a tertiary eye care hospital.

Purpose: To evaluate the efficacy of liposomal amphotericin B (L-AMB) for the treatment of fungal keratitis. Methods: Patients with fungal keratitis confirmed by potassium hydroxide (KOH) smear and/or confocal microscopy were administered topical L-AMB and randomized into three groups treated with three different formulations. The medication was administered two hourly till clinical improvement was achieved, followed by six hourly till complete resolution. The outcome measures were time to clinical improvement, resolution of epithelial defect, stromal infiltrate, hypopyon, extent and density of corneal opacity, neovascularization, and best corrected visual acuity (BCVA) at 3 months. Results: Mean age of the patients was 46.6 ± 14.8 years, and trauma with vegetative matter was the most common predisposing factor. Aspergillus flavus (36%) was the most common fungus cultured, followed by Fusarium (23%). Mean time to clinical improvement, time to resolution of epithelial defect, mean time to resolution of infiltrate, and time to resolution of hypopyon were 3.45 ± 1.38, 25.35 ± 8.46, 37.97 ± 9.94, and 13.33 ± 4.90 days, respectively, and they were comparable among the three groups. There was a significant difference between treatment failure and success cases in terms of days of presentation (P < 0.01), size of the epithelial defect (P-value 0.04), and infiltrate size at presentation (P-value 0.04). At 3 months follow-up, no statistically significant difference was noted in BCVA and mean scar size among groups. Conclusion: L-AMB in a gel form is an effective antifungal agent that promotes the healing of fungal ulcers with notably least vascularization and better tolerance. Trial registration number: CTRI/2020/04/024550.

Central Line-Associated Bloodstream Infections: Effect of Patient and Pathogen Factors on Outcome.

Introduction: Patients on central lines are often having multiple morbidities, and invasive devices provide a niche for biofilm formation, which makes central line-associated bloodstream infections (CLABSIs), a serious concern in health-care settings, as the infections difficult to treat. In this study, we evaluated the common bacteria causing CLABSI, and various patient and pathogen factors affecting the clinical outcome. Methods: In the prospective observational study, patients diagnosed with CLABSI were recruited. Extensive clinical, microbiological, and other laboratory workup was done, and observations were recorded. Congo red agar method, tube test, and microtiter plate assay were used for eliciting the biofilm-forming attributes of the bacterial pathogens. Results: Klebsiella pneumoniae was responsible for 48% of CLABSI, followed by Coagulase-negative Staphylococci (16%) and Staphylococcus aureus and Acinetobacter baumannii (12% each). Fifty-six percent of the isolates produced biofilms. The median (interquartile range) duration of hospital stay till death or discharge was 30 (20, 43) days. The all-cause mortality was 44%. Patients having a deranged liver function on the day of diagnosis (P value for total bilirubin 0.001 and for aspartate transaminase 0.02), and those infected with multidrug-resistant organisms (P value = 0.04) had significantly poor prognosis. The difference in the demographic, clinical, laboratory profile, and outcome of patients infected with biofilm producers and nonproducers was not found to be statistically significant. Conclusion: The study throws light on various host and pathogen factors determining the cause and outcome of CLABSI patients. To the best of our knowledge, this is the first study trying to decipher the role of biofilm formation in the virulence of pathogens and the prognosis of CLABSI.

Ocular Chlamydia trachomatis infections in patients attending a tertiary eye care hospital in north India: a twelve year study.

BACKGROUND & OBJECTIVES: Ocular infection with Chlamydia trachomatis is a major public health problem in densely populated countries like India. The true prevalence of such infections is uncertain due to insufficient data available from India. The aim of this study was to do a retrospective analysis of C. trachomatis eye infections in patients attending the outpatient department of Dr Rajendra Prasad Centre for Ophthalmic Sciences, All India Institute of Medical Sciences, New Delhi, over a period of 12 years. METHODS: From 1997 to 2008, the Chlamydia laboratory received conjunctival swabs from 1281 consecutive patients for C. trachomatis detection after thorough clinical examination. Specimens were subjected to direct fluorescent antigen detection assay using monoclonal antibody based commercial kit to detect the presence of C. trachomatis antigen. RESULTS: Antigen positivity varied between 22-28 per cent. Children below 11 yr and people above the age of 60 yr showed comparatively higher antigen positivity (25.7 and 27.8%, respectively). As compared to males significantly (P<0.05) higher number of females in the age group of 31-60 yr were positive for C. trachomatis antigen. Patients with the clinical diagnosis of follicular/allergic conjunctivitis and trachoma showed higher rate of antigen positivity. INTERPRETATION & CONCLUSIONS: Northern India having dry and arid climatic conditions in most parts of the year was considered in the past as one of the trachoma hyper-endemic foci. The study indicated that laboratory proven C. trachomatis eye infection still persisted in this part of the country throughout the study period of 12 years.

Moxifloxacin resistance: intrinsic to antibiotic or related to mutation?

PURPOSE: To evaluate and compare the pattern of resistance to topical moxifloxacin of ocular isolates recovered during comparable periods within 2 years. METHODS: Records of 611 consecutive cases of bacterial corneal ulcers that presented to our center between April and August 2007 were compared with records of 417 cases that presented in April and August 2006. Records of culture and sensitivity to cefazolin, tobramycin, gatifloxacin, and moxifloxacin at the time of presentation were analyzed. Parameters recorded included total number of cases in each period, culture-positive cases, organisms isolated, and sensitivity by Kirby-Bauer disk-diffusion test. The data from 2007 was compared with those from 2006 using χ test. RESULTS: Culture-positive rate was 70.2% in 2007 compared with 65.0% in 2006 (p = 0.143). The percentage of culture-positive cases resistant to moxifloxacin was 5.46% in 2007 compared with 5.42% in 2006 (p = 0.977). There was no significant difference between resistance against cefazolin during the two periods (p = 0.895). Resistance to gatifloxacin and tobramycin was significantly higher in samples tested in 2007 as compared with those tested in 2006 (gatifloxacin, p = 0.037; tobramycin, p = 0.031). CONCLUSIONS: Resistance to moxifloxacin did not change significantly during a period of 2 years in our study. The static resistance pattern to moxifloxacin may indicate that such resistance to moxifloxacin is intrinsic in the antibiotic rather than new resistance stemming from mutations.

Evaluation of 0.3% gatifloxacin hydrochloride in decontamination of donor corneas.

PURPOSE: To evaluate demographic, clinical, and microbiological profile of eye donors and efficacy of 0.3% gatifloxacin hydrochloride in microbial decontamination of donor corneas. METHODS: About 513 donors and 1,026 corneas received at National Eye Bank of a tertiary care hospital during 1-year period were analyzed prospectively in this randomized clinical trial. The donor eyes were graded and treated with 5% povidone-iodine, 0.4% amikacin sulphate, and 0.3% gatifloxacin hydrochloride. The parameters evaluated were death enucleation time (DET), grading of donor corneas, microbiological profile of culture organisms, and their sensitivity to various antibiotics. RESULTS: Mean DET was 6.29±5.7 hours. Forty one percent eyes were optical grade corneas and the majority of donors (38.5%) had accidental deaths. Good grade eyes were maximum with DET of <1 hour and were comparable between 0-6 hours and 6-12 hours. About 57.6% (591/1026) eyes were culture positive; most common organisms were Pseudomonas spp (53%) and Coagulase-negative Staphylococci (24%). Culture positivity reduced significantly after treatment with povidone iodine and amikacin (P=0.002, right eye; P=0.004; left eye) and decreased further with use of gatifloxacin (P=0.001). Pseudomonas (93%), Coagulase-negative Staphylococci (96.3%), Staphylococcus aureus (90.5%), enterococci and gram-negative bacilli were sensitive to gatifloxacin. Pseudomonas spp which were multidrug-resistant were sensitive to polymyxin-B. CONCLUSIONS: Gatifloxacin hydrochloride in addition to amikacin sulphate is beneficial for donor eye decontamination. Polymyxin-B may be used for multidrug-resistant Pseudomonas spp.

Acanthamoeba keratitis: Experience from a tertiary eye care center in North India.

The free-living amebae of genus Acanthamoeba are an important cause of microbial keratitis. The clinical appearance of Acanthamoeba keratitis (AK) usually mimics viral or fungal keratitis. Thus, microbiological workup plays a significant role in the diagnosis and timely treatment of such cases. We report a retrospective case series of seven culture-confirmed AK cases from a tertiary eye care center in North India. Various risk factors and triggers of infection, clinical presentations, microbiological findings, and management of AK are elucidated.

Real-life comparison of three intravitreal antibiotic drug regimens in endophthalmitis.

Purpose: Real-life comparison of three intravitreal drug regimens used in cases of endophthalmitis at a tertiary care center in India. Methods: In this prospective, comparative study, patients of bacterial endophthalmitis were grouped according to intravitreal antibiotic drug regimens into Group 1 (ceftazidime and vancomycin), Group 2 (piperacillin + tazobactam and vancomycin), and Group 3 (imipenem and vancomycin). Forty-eight hours after injection nonresponding/worsening patients underwent vitrectomy. Vitreous samples were subjected to microbiological and pharmacokinetic tests. Results: A total of 64 patients were included and divided into Group 1: 29, Group 2: 20, and Group 3: 15 cases. Also, 75% of patients were post-surgical endophthalmitis, whereas 25% were post-traumatic. Improvement in vision (V90-0) and vision at 3 months (V90) were comparable between the three groups. Visual recovery was poorer in post-traumatic cases. In post-surgical cases, visual recovery was poorer in those presenting beyond 72 h of onset of symptoms (P = 0.0002). Polymerase chain reaction (PCR) positivity (66%) was higher than BACTECTM (33%) and culture (14%). Antibiotic resistance was comparable amongst the three groups. Most patients (62/64) further underwent vitrectomy. Ceftazidime and vancomycin achieved vitreous concentrations more than the minimum inhibitory concentration (MIC) at 48 h after the first injection. Conclusion: The choice of antibiotics did not affect the rate of vitrectomy and final vision in a real-life scenario. Ceftazidime and vancomycin can still be used as first-line intravitreal antibiotics owing to their comparable microbial sensitivity profile and adequate ocular bioavailability.

Spectrum of bacterial keratitis in North India: A retrospective analysis of six years at a tertiary care eye center.

Purpose: To analyze the pattern of bacterial pathogens causing infective keratitis and their resistance to the recommended antibiotics over six years. Methods: It was a retrospective study of 9,357 cases of bacterial keratitis from January 2015 to December 2020, at a tertiary care ophthalmic center. A total of 9,547 corneal specimens were obtained from the study subjects. Demographic details of the patients, pathogenic bacteria isolated, and their antimicrobial susceptibility were noted and analyzed. Results: Bacterial pathogens were identified in 23.52% of the specimens. The most common isolates were coagulase-negative Staphylococci (60.75%), followed by Pseudomonas aeruginosa (14.23%), Staphylococcus aureus (13.92%), gram negative bacilli of the family Enterobacterales (8.64%), Streptococcus spp. (1.72%), Acinetobacter spp. (0.13%), and other non-fermenting gram-negative bacilli (0.57%). In Staphylococci, 55-80% of isolates were resistant to erythromycin, and 40-70% to fluoroquinolones, while no resistance was observed against vancomycin. 40-60% of isolates of P. aeruginosa were resistant to cephalosporins, 40-55% to fluoroquinolones, and 30-60% to aminoglycosides. Also, 40-80% of isolates of Enterobacterales were resistant to cephalosporins, and 50-60% to fluoroquinolones. Most gram-negative isolates were susceptible to carbapenems and polymyxin B. Conclusion: To the best of our knowledge, our study is the largest compilation of microbiological profile of bacterial keratitis from North India. It highlights the current trend of the bacterial pathogens that cause infectious keratitis. Staphylococci and Pseudomonas were found to be the most common pathogens. Increased resistance was seen against some of the commonly prescribed empirical antibiotics. Such evidence is useful for restructuring the empirical prescription practices from time to time.

Targeted metagenomics using next generation sequencing in laboratory diagnosis of culture negative endophthalmitis.

To study the feasibility of 16S rRNA metagenomics using next generation sequencing (NGS) along with broad range PCR assay for 762 bp region of 16S rRNA gene with Sanger's sequencing, in microbial diagnosis of culture negative endophthalmitis. Vitreous fluid from 16 culture negative and one culture positive endophthalmitis patients, admitted to a tertiary care hospital were processed for targeted metagenomics. NGS of 7 variable regions of 16S rRNA gene was done using Ion Torrent Personal Genome Machine (PGM). Sequence data were analyzed using Ion Reporter software using QIIME and BLSATN tools and Greengenes and NCBI-Genbank databases. Bacterial genome sequences were detected in 15 culture negative and culture positive vitreous specimens. The sequence reads varied between 25,245-540,916 with read length between 142bp-228bp and coverage depth was 41.0X and 81.2X. Operational taxonomic unit (OTUs) of multiple bacterial genera and species were detected in 13 culture negative vitreous specimens and OTUs of a single bacterial species were detected in 2 culture negative and 1 culture positive specimens; one negative specimen had no bacterial DNA. Maximum numbers of OTUs detected by NGS for a bacterial species from any vitreous specimen was the one which was detected and identified by Sanger's sequencing in broad range PCR. All the bacteria were belonging to clinically relevant species. Broad range PCR with sequencing failed to identify bacteria from 5 of the 16 (31.25%) culture negative vitreous specimens. Metagenomics could detect and identify bacterial pathogens in 15 of the 16 culture negative vitreous specimen's up to species level. With rapidly decreasing cost, metagenomics has a potential to be used widely in endophthalmitis diagnosis, in which culture negativity is usually high.

Role of therapeutic contact lens following Descemet's stripping automated endothelial keratoplasty: A randomized control trial.

Purpose: Therapeutic contact lenses (TCL) are known to help in epithelial healing and decreasing pain after various corneal surgeries. However, literature lacks any data describing their use following Descemet's stripping automated endothelial keratoplasty (DSAEK) where intraoperative epithelial debridement is commonly performed. Here we study the efficacy and safety of TCL in patients undergoing DSAEK. Methods: In this prospective, randomized, controlled clinical trial. 40 eyes of 40 patients of pseudophakic bullous keratopathy undergoing DSAEK were enrolled and randomized into two groups, control (no TCL) and test (TCL). Primary outcome was time taken for epithelial healing and secondary outcomes were postoperative pain score, graft attachment, best spectacle-corrected visual acuity, and endothelial cell loss at 3 months. Results: Average time taken for epithelial healing was 3.35 ± 0.49 days in the test group and 4.95 ± 1.05 days in the control group (P < 0.001). Average pain scores in first operative week were significantly lower in the test group as compared to control (P < 0.001). Graft detachment occurred in eight patients in control group and two in test group (P = 0.03). Both rebubbling rates and average endothelial cell loss at 3 months were higher in the control group with P = 0.07 and 0.06 respectively. No contact lens-related adverse effects were noted during the study period. Conclusion: Use of TCL in DSAEK leads to faster epithelial healing and lesser postoperative pain. In addition, it may also contribute to lower rebubbling rates and endothelial cell loss.

Determination of surgical outcomes with a novel formulation of intrastromal natamycin in recalcitrant fungal keratitis: A pilot study.

PURPOSE: To evaluate the outcomes of water-soluble intrastromal natamycin (IS-NTM) as an adjunct therapy for recalcitrant fungal keratitis. METHODS: This was a prospective interventional pilot study in the setting of a tertiary eye-care center. Twenty eyes of 20 consecutive patients with microbiologically proven recalcitrant fungal keratitis (ulcer size >2 mm, depth >50%, and not responding to topical NTM for 2 weeks) were recruited. The selected patients were injected with a novel composition of IS-NTM (10 ug/0.1 mL, soluble natamycin) prepared aseptically in the ocular pharmacology department. All the patients continued using topical NTM suspension 5% 4-hourly until the ulcer healed. Repeat injections were undertaken after 72 h depending on the clinical response and all the patients were followed till 6 months. RESULTS: The mean age of the patients was 40.42 ± 10.09 years. The mean duration of the presentation was 20.8 ± 5.1 days. The most commonly isolated organisms were Aspergillus sp. (12/20, 60%) and Fusarium sp. (8/20, 40%). No patient had iatrogenic perforation or precipitate formation after IS-NTM injection. The overall cure rate with IS-NTM was 95% (19/20 patients). The number of patients who healed with the 1st, 2nd, and 3rd injection was 13, 5, and 1, respectively. One (5%) had no response to treatment and was subjected to penetrating keratoplasty. The average time taken for the resolution of the epithelial defect, stromal infiltrates, and hypopyon was 34 ± 5.2 days, 35.3 ± 6.4 days, and 15 ± 2.5 days. Healing with deep vascularization and cataract was noted in 6/19 eyes (31%) and 13/19 eyes (68.42%), respectively. CONCLUSION: Intrastromal injection of a novel formulation of NTM holds a promising role as adjunctive therapy to topical NTM in the management of recalcitrant filamentous fungal keratitis. The preliminary results are encouraging and further studies are required to validate the results.

Evaluation of various preservation media for storage of donor corneas.

PURPOSE: To compare the physical and microbiological characteristics of McCarey-Kaufman (MK), Cornisol, and Optisol-GS media and evaluate the outcomes of keratoplasty performed using corneas stored in these three media. METHODS: The study involved 60 donor corneas which were distributed in 3 groups: MK, Cornisol, and Optisol-GS. Corneas in these groups were further analyzed based on the type of keratoplasty performed (full thickness versus endothelial keratoplasty). At baseline, the endothelial cell density and death to preservation time of donor corneas were recorded. Following keratoplasty, patients were evaluated on day 1, at 1 month, 3 months, and 6 months follow-up. Outcomes were assessed in terms of corrected distance visual acuity (CDVA), endothelial cell density, percentage endothelial cell loss, and corneal thickness. The storage media were also assessed for their physical quality and their microbiological characteristics. RESULTS: Physical characteristics of all three media were found to be within normal limits. Mean CDVA was comparable among the 3 groups at 6-month follow-up. The absolute endothelial cell count values were significantly lower for corneas stored in MK medium (1873.7 ± 261.1 cells/mm2) compared to the Cornisol (2085.0 ± 230.3 cells/mm2) and Optisol-GS media [(2180.3 ± 217.2 cells/mm2) (P = <0.001)]. Corneas stored in Optisol-GS medium were significantly thinner at 1-month follow-up with no significant difference at 6 months (P = 0.66). CONCLUSION: Optisol-GS and Cornisol media were found to preserve endothelial cell density better and stabilize corneal thickness earlier as compared to the MK medium. However, the functional outcomes were comparable among the three groups.

Comparison of Safety and Efficacy of Intrastromal Injections of Voriconazole, Amphotericin B and Natamycin in Cases of Recalcitrant Fungal Keratitis: A Randomized Controlled Trial.

OBJECTIVE: To compare the safety and efficacy of intrastromal voriconazole (IS-VCZ), amphotericin B (IS-AMB) and natamycin (IS-NTM) as an adjunct to topical natamycin (NTM) in cases of recalcitrant fungal keratitis. DESIGN: Prospective randomized trial. SETTING: Tertiary eye centre. PARTICIPANTS: Sixty eyes of 60 patients with microbiologically proven recalcitrant fungal keratitis (ulcer size >2 mm, depth >50% of stroma, and not responding to topical NTM therapy for two weeks) were recruited. METHODS: patients were randomized into three groups of 20 eyes, each receiving ISVCZ 50ug/0.1 mL, ISAMB, 5ug/0.1 mL and ISNTM 10ug/0.1 mL (prepared aseptically in ocular pharmacology). The patients in all three groups continued topical NTM 5% every four hours until the ulcer healed. Primary outcome measure was time taken till complete clinical resolution of infection, and secondary outcome measure was best corrected visual acuity (BCVA) at six months. RESULTS: All three groups had comparable baseline parameters. The mean duration of healing was significantly better (p=0.02) in the ISNTM group (34±5.2 days) as compared to the ISVCZ group (36.1±4.8 days) and the ISAMB group (39.2±7.2 days). About 95%, 90% and 95% patients healed successfully in the ISVCZ, ISAMB and ISNTM groups, respectively. In terms of healing, deep vascularization was significantly greater in the ISAMB group (55%, p=0.02) when compared to the ISVCZ and ISNTM groups (31% and 26%, respectively). There were fewer repeat injections in the ISNTM group (7/20 vs 8/20 and 9/20 in the ISVCZ and ISNTM groups, respectively). CONCLUSION: Intrastromal injections are a safe and effective adjunct to conventional therapy in the management of recalcitrant fungal keratitis. ISNTM had a similar visual outcome with faster healing while ISAMB had a higher rate of deep vascularization after healing.

Post-operative endophthalmitis: Antibiogram & genetic relatedness between Pseudomonas aeruginosa isolates from patients & phacoemulsifiers.

BACKGROUND & OBJECTIVES: Though not frequently but there are reports showing phacoemulsifiers as a potent source of infection in post-operative cases of endophthalmitis. This study was carried out to find antibiogram and genetic relatedness between Pseudomonas aeruginosa isolates from a post-cataract surgery endophthalmitis outbreak (3 patients) and internal tubings of 5 phacoemulsifiers. METHODS: In vitro antimicrobial sensitivity patterns of the 8 bacterial isolates were observed. Genetic analysis of the bacterial isolates was done using random amplification of polymorphic DNA (RAPD) assay and PCR ribotyping. The resulting DNA band patterns were examined visually and by computer assisted analysis using unweighted pair group method. RESULTS: The three P. aeruginosa patient isolates were found to be different from the five phacoemulsifier isolates in sensitivity towards 3 antibiotics and by genetic analysis (33 and 44% homology by RAPD assay and PCR ribotyping). Two of the patient isolates shared 100 per cent genetic homology by RAPD assay and another pair shared 100 per cent homology by PCR ribotyping. The five isolates from phacoemulsifiers did not share significant genetic homology. There was significant genetic variation between bacterial isolates from patients and phaco emulsifiers. INTERPRETATION & CONCLUSION: Though the three P. aeruginosa isolates obtained from the patients were phenotypically similar and genetically close, they differed from the phaco-machine isolates both genetically, and in their antibiogram profile. However, the five phacoemulsifier isolates were genetically diverse though they shared the same antibiogram profile. Therefore the Ringer's lactate from phacomachines could not be conclusively proven to be the source of infection.