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.

Molecular characterization of drug-resistant and drug-sensitive Aspergillus isolates causing infectious keratitis.

Purpose: To study the susceptibilities of Aspergillus species against amphotericin B in infectious keratitis and to find out if drug resistance had any association with the molecular characteristics of the fungi. Materials and Methods: One hundred and sixty Aspergillus isolates from the corneal scrapings of patients with keratitis were tested for susceptibilities to amphotericin B by broth microdilution method. These included Aspergillus flavus (64 isolates), A. fumigatus (43) and A. niger (53). Fungal DNA was extracted by glass bead vertexing technique. Polymerase chain reaction (PCR) assay was standardized and used to amplify the 28S rRNA gene. Single-stranded conformational polymorphism (SSCP) of the PCR product was performed by the standard protocol. Results: Of the 160 isolates, 84 (52.5%) showed low minimum inhibitory concentration (MIC) values (≤ 1.56 μg/ml) and were designated as amphotercin B-sensitive. Similarly, 76 (47.5%) had high MICs (≥ 3.12 μg/ml) and were categorized as amphotericin B-resistant. MIC50 and MIC90 values ranged between 3.12-6.25 μg/ml and 3.12-12.5 μg/ml respectively. A. flavus and A. niger showed higher MIC50 and MIC90 values than A. fumigatus. The SSCP pattern exhibited three extra bands (150 bp, 200 bp and 250 bp each) in addition to the 260 bp amplicon. Strains (lanes 1 and 7) lacking the 150 bp band showed low MIC values (≤ 1.56 μg/ml). Conclusion: A. niger and A. flavus isolates had higher MICs compared to A. fumigatus, suggesting a high index of suspicion for amphotericin B resistance. PCR-SSCP was a good molecular tool to characterize Aspergillus phenotypes in fungal keratitis.

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.

Phenotypic and plasmid pattern analysis of Staphylococcus epidermidis in bacterial keratitis.

BACKGROUND: Staphylococcus epidermidis, a commensal of the conjunctival sac has been incriminated as the commonest etiological agent of bacterial keratitis. However, the pathogenic potential of this commensal organism is not clearly known. AIM: To determine any phenotypic, molecular markers of S. epidermidis pathogenicity in bacterial keratitis. MATERIALS AND METHODS: A total of 382 corneal ulcer isolates of S. epidermidis and 87 S. epidermidis isolates from healthy eyes (controls) were studied. Speciation, biotyping and antibiotic sensitivity testing were performed by conventional methods. Tube slime and adherence tests were carried out by recommended techniques. Plasmid analysis was conducted by a standard protocol. STATISTICAL ANALYSIS: Chi-square test was employed for calculations. RESULTS: Out of 382 corneal ulcer isolates (Pathogens) 284 (74.3%) belonged to biotypes I and II. Slime was detected in 164 (42.9%) of 382 pathogens vs. 21 (24.1%) of 87 controls (P<0.001). Sixty-five (39.6%) of 164 slime positive isolates were multidrug-resistant as compared to only 49 (22.4%) of 218 slime negative isolates (P<0.001). A significantly higher number i.e, 73.1% (120/164) of slime-producers possessed a 21 Kb plasmid in contrast to only 53.2% (116/218) of nonslime-producers (P<0.001). Presence of this plasmid had a statistical correlation of low significance with multidrug resistance (P=0.04). One hundred and seventy-two (45.0%) of 382 pathogens and 24 (27.6%) of the 87 controls were adherent to artificial surfaces (P=0.003) and the majority of the adherent organisms (99/172, 57.6%) were slime producers (P<0.001). CONCLUSIONS: Slime was associated with multidrug resistance in corneal ulcer isolates of S. epidermidis. The 21 Kb plasmid could determine virulence as it was responsible for slime production and adherence.