Development of single-tube real-time PCR assay for the rapid detection of Aspergillus and Fusarium-The two most common causative agents in fungal keratitis.

BACKGROUND: To compare the performance of conventional, semi-nested and real-time panfungal ITS PCRs for diagnosing fungal keratitis (FK) and develop genus-specific real-time PCR for the most common aetiology of FK. METHODS: This multicentric study includes 232 corneal samples from suspected FK patients from four centres across India between November 2019 through August 2021. A total of 87 corneal buttons were included for the comparison of conventional, semi-nested and real-time ITS PCRs, of which 68 were from confirmed FK patients. Of these 87 samples, 44 (microscopy and culture positive for Aspergillus sp. and/or Fusarium sp.) were used for the standardisation of genus-specific real-time primers/probes. Subsequently, the best method showing highest sensitivity and specificity was validated in 188 samples. RESULTS: On Bayesian comparison, conventional ITS2 PCR showed best performance (sensitivity and specificity of 55.88% and 100%, respectively). Since, real-time ITS2 PCR was also considerably efficient (sensitivity and specificity of 51.47% and 84.21%, respectively) in comparison with the conventional PCR but faster, cost-effective, and less labor-intensive, ITS-2 real-time PCR is a suitable method that can be applied along with culture and microscopy. During validation, real-time PCR with genus-specific primers showed 61.76% and 91.18% sensitivity with specificity of 98.05% and 79.22%, respectively, for Aspergillus sp. and Fusarium sp. Aspergillus probe, Fusarium probe and duplex PCR showed sensitivity of 52.94%, 50% and 54.41% with specificity of 92.86%, 82.47% and 75%, respectively. No cross-reactivity of genus-specific PCRs was observed during standardisation. CONCLUSIONS: ITS-2 real-time PCR can be applied as an adjunct with conventional methods for the diagnosis of FK. The genus-specific duplex real-time PCRs are rapid which reduces the turnaround time (TAT) avoiding the need for sequencing.

Case Series: Mixed Infectious Keratitis by Pythium insidiosum and Fungal Species.

SIGNIFICANCE: This case series is the first to illustrate mixed infection from Pythium sp. and fungal species in corneal ulcer. PURPOSE: This case series aimed to alert all toward the possibility of both Pythium sp. and fungal species infection in case of nonresponding corneal ulcer treated with either antifungals or antipythium drugs alone. Increased suspicion of mixed infection in case of nonresponding fungal/ Pythium keratitis may facilitate early and prompt management. CASE REPORTS: Six patients presented with signs of either fungal or Pythium keratitis. They underwent ophthalmological examinations, smear examinations, cultures, and polymerase chain reaction (PCR). Therapeutic penetrating keratoplasty was performed in cases where symptoms worsened after treatment with either antifungal or antipythium drugs. The half corneal button (HCB) was shared for histopathological and microbiological examinations. In the first case, smear examination from corneal scraping (CS) revealed Pythium -like filaments, which were confirmed with PCR; however, Aspergillus nidulans grew in culture. In the second case, iodine-potassium iodide (IKI) staining was positive for Pythium ; however, PCR was positive for both Pythium and fungus, which was further confirmed by DNA sequencing. In the third case, IKI staining and HCB were positive for Pythium ; however, PCR was positive for fungus, which was identified as Candida saitoana with DNA sequencing. In the fourth case, Pythium grew in the CS culture; however, Candida sp. grew in the HCB culture. In the fifth case, Cladosporium sp. grew in culture from CS; however, Pythium insidiosum grew from the anterior chamber exudate after therapeutic penetrating keratoplasty. In the sixth case, smear examination revealed septate fungal filaments, and Cladosporium sp. grew in culture; however, HCB on histopathological examination showed features of Pythium keratitis. CONCLUSIONS: In unresponsive cases of Pythium or fungal keratitis, diagnostic modalities such as IKI and PCR should be implemented as a routine practice, in addition to smears and cultures.

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.

Challenges in management of microbial keratitis during COVID-19 pandemic related lockdown: a comparative analysis with pre pandemic data.

PURPOSE: To study the challenges of managing microbial keratitis(MK) during the COVID19 pandemic related lockdown and assess the outcomes of treatment at a tertiary cornea service. METHODS: Retrospective, non comparative study of electronic medical records of MK presenting to a network of four tertiary care cornea services. The medical history, presenting clinical features, microbiology work up and treatment outcomes were analyzed. The primary outcome measure was final outcome at last follow up. Secondary outcomes measures were non-compliance to treatment due to travel restrictions, therapeutic PKP not done due non availability of corneal tissues. Results- MK was noted in 330 eyes of 330 patients between April and May 2020. Of these 237(71.8%) were males. Median age was 45 years(IQR, 33-56). Low socioeconomic status noted in 102(30.9%). Patients travelling beyond the district from where the hospital was located comprised of 64.9%(n=214). At a median follow up of 32 days(IQR, 9-54), 118(35.8%) patients had resolved, with medical management, 73(22.1%) patients were under active treatment, 139(42.1%) were lost to follow up. Sixty-six patients(20%) were non-compliant to treatment of which 59 could not follow appointment schedule due to travel restrictions. Therapeutic PKP (TPK) was planned in 48/128 (37.5%) patients, but was performed in only 34/48 (70.8%) due to non-availability of donor corneas. CONCLUSIONS: Abnormal social circumstances due to the COVID pandemic and the ensuing impediments to travel for access to health care affected compliance to treatment of ocular emergencies such as microbial keratitis.

Tissue remodeling in adult vernal keratoconjunctivitis.

Our aim is to describe local tissue remodeling in a cohort of adult VKC patients. Male patients diagnosed with active VKC were enrolled in an open pilot study into two groups according disease onset: childhood classic VKC and adult VKC. Visual acuity and ocular surface clinical examination focusing on chronic inflammatory sequelae and impression cytology were performed in all enrolled subjects. Conjunctival imprints were processed for molecular, biochemical and immunofluorescent analysis for tissue remodeling (TGFß1,2,3 and αSMA) and epigenetic (DNMT3a, Keap1; Nrf2) markers as well as androgen receptors were investigated and compared between groups. Clinical assessment showed increased conjunctival scarring in adult VKC compared to classic VKC. Immunoreactivity for αSMA and expression of TGFß were higher in adult VKC group. Significantly higher levels of TGFß3 (3.44 ± 1.66; p < 0.05) were detected in adult VKC compared to childhood VKC, associated with an increasing trend of TGFß1 (1.58 ± 0.25) and TGFß2 (1.65 ± 0.20) isoforms levels. Molecular analysis showed a relative increase in tissue remodeling/fibrogenic transcripts (TGFß isoforms and αSMA) associated to a significant increase of selective epigenetic targets (DNMT3, Nrf2 and keap1) in adult VKC phenotype. Increased local conjunctival androgen receptors was detected in patients with adult variants compared to classic childhood VKC and healthy subjects. Finally, a direct correlation between TGFß and androgen receptor expression was also detected. A pro-fibrotic clinical and biomolecular trait was unveiled in adult variant of VKC, which causes ocular surface disease and visual impairment.

Elucidating the clinical, microbiological and molecular diagnostic aspects of Macrophomina phaseolina keratitis.

This study reports the clinico-microbiological features of Macrophomina phaseolina keratitis. Clinically diagnosed as microbial keratitis, six patients underwent microbiological evaluation. Fungal culture isolates from cornea were subjected to DNA sequencing of the ITS region, phylogenetic analysis and reconfirmation by polymerase chain reaction (PCR). Minimum inhibitory concentrations (MICs) of six antifungal drugs were determined by microbroth dilution method against the six isolates. All patients were treated with antifungals. Failed medical therapy necessitated therapeutic penetrating keratoplasty (TPK). Corneal buttons were processed for histopathology. In all patients, the corneal scraping showed septate hyaline fungal filaments. The BLAST analysis for ITS sequences of all six fungal isolates suggested M. phaseolina, however, when limited to sequences from type material, they matched M. pseudophaseolina. Phylogenetic analysis could not differentiate between these two species and clustered in a single clade. PCR assay of specific gene sequence [MpCal (calmodulin)] reconfirmed all isolates as M. phaseolina. The MICs of voriconazole and posaconazole were lowest (0.03 to 2 and 0.1 to 2µg/mL respectively) and all isolates were susceptible to natamycin. Except for case 1, which healed with a scar on treatment, all other cases worsened, despite medical treatment, necessitating TPK. Histopathology of 3 out of 4 buttons showed the presence of fungal filaments. While direct microscopic examination of corneal scrapings is helpful in diagnosis, identification of M. phaseolina in culture is challenging. Although MICs of commonly used antifungals are low response to medical therapy is not encouraging; patients may require TPK for resolution of infection in M. phaseolina keratitis.

DNA sequencing, phylogenetic analysis and specific PCR confirmed Macrophomina phaseolina keratitis in six patients. Although antifungal susceptibility showed the organisms to be susceptible to natamycin five patients did not respond to treatment and needed keratoplasty.

Assessment of genotypes, endosymbionts and clinical characteristics of Acanthamoeba recovered from ocular infection.

INTRODUCTION: Acanthamoeba is an emerging pathogen, infamous for its resilience against antiprotozoal compounds, disinfectants and harsh environments. It is known to cause keratitis, a sight-threatening, painful and difficult to treat corneal infection which is often reported among contact lens wearers and patients with ocular trauma. Acanthamoeba comprises over 24 species and currently 23 genotypes (T1-T23) have been identified. AIMS: This retrospective study was designed to examine the Acanthamoeba species and genotypes recovered from patients with Acanthamoeba keratitis (AK), determine the presence of endosymbionts in ocular isolates of Acanthamoeba and review the clinical presentations. METHODOLOGY: Thirteen culture-confirmed AK patients treated in a tertiary eye care facility in Hyderabad, India from February to October 2020 were included in this study. The clinical manifestations, medications and visual outcomes of all patients were obtained from medical records. The Acanthamoeba isolates were identified by sequencing the ribosomal nuclear subunit (rns) gene. Acanthamoeba isolates were assessed for the presence of bacterial or fungal endosymbionts using molecular assays, PCR and fluorescence in situ hybridization (FISH). RESULTS: The mean age of the patients was 33 years (SD ± 17.4; 95% CI 22.5 to 43.5 years). Six (46.2%) cases had AK associated risk factors; four patients had ocular trauma and two were contact lens wearers. A. culbertsoni (6/13, 46.2%) was the most common species, followed by A. polyphaga and A. triangularis. Most of the isolates (12/13) belonged to genotype T4 and one was a T12; three sub-clusters T4A, T4B, and T4F were identified within the T4 genotype. There was no significant association between Acanthamoeba types and clinical outcomes. Eight (61.5%) isolates harboured intracellular bacteria and one contained Malassezia restricta. The presence of intracellular microbes was associated with a higher proportion of stromal infiltrates (88.9%, 8/9), epithelial defect (55.6%, 5/9) and hypopyon (55.6%, 5/9) compared to 50% (2/4), 25% (1/4) and 25% (1/4) AK cases without intracellular microbes, respectively. CONCLUSIONS: Genotype T4 was the predominant isolate in southern India. This is the second report of T12 genotype identified from AK patient in India, which is rarely reported worldwide. The majority of the Acanthamoeba clinical isolates in this study harboured intracellular microbes, which may impact clinical characteristics of AK.

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.

Microbes of the human eye: Microbiome, antimicrobial resistance and biofilm formation.

BACKGROUND: The review focuses on the bacteria associated with the human eye using the dual approach of detecting cultivable bacteria and the total microbiome using next generation sequencing. The purpose of this review was to highlight the connection between antimicrobial resistance and biofilm formation in ocular bacteria. METHODS: Pubmed was used as the source to catalogue culturable bacteria and ocular microbiomes associated with the normal eyes and those with ocular diseases, to ascertain the emergence of anti-microbial resistance with special reference to biofilm formation. RESULTS: This review highlights the genetic strategies used by microorganisms to evade the lethal effects of anti-microbial agents by tracing the connections between candidate genes and biofilm formation. CONCLUSION: The eye has its own microbiome which needs to be extensively studied under different physiological conditions; data on eye microbiomes of people from different ethnicities, geographical regions etc. are also needed to understand how these microbiomes affect ocular health.

Rabbit models of dry eye disease: Current understanding and unmet needs for translational research.

Dry eye disease (DED) is emerging as an eye health pandemic, affecting millions worldwide. The development of novel drugs, drug delivery systems, and targeted therapies for addressing the inflammation in DED necessitates progress in experimental models of DED. Animal models of DED have been created for simulating the two clinically described forms of DED: lacrimal insufficiency and the evaporative DED models. Although most DED models have relied upon rodents, the larger eye size and longer life span of rabbits and the closer resemblance to human lacrimal glands, render rabbits a promising near-ideal model for studying DED. Since the first rabbit DED model was described, numerous modifications including the use of topical epitheliotoxic drugs, neural abolition, activated lymphocytes injection, and surgical dacryoadenectomy have been introduced. The stability of these models, whether short-term or long-term, accordingly guides their experimental or therapeutic utility. A rabbit autoimmune dacryoadenitis model has successfully simulated DED signs and features of lacrimal gland inflammation, as observed in Sjogren's syndrome, that improved with mesenchymal stem cell therapy. This review summarizes the comparative microscopic anatomy of rabbit and human lacrimal glands, various existing rabbit DED models and their respective suitability for understanding pathogenetic mechanism of DED or for experimental drug testing. Also, the insights gained from animal models in dry eye management is described along with the future perspectives. There is still a pressing need of developing rabbit models for studying the pathogenesis of complex ocular surface changes in evaporative and aqueous deficiency DED other than autoimmune dacryoadenitis.

Development of antibiotic resistance in the ocular Pseudomonas aeruginosa clone ST308 over twenty years.

Corneal infection caused by a bacteria Pseudomonas aeruginosa is common cause of ocular morbidity. Increasing antibiotic resistance by ocular P. aeruginosa is an emerging concern. In this study the resistome of ocular isolates of Pseudomonas aeruginosa clone ST308 isolated in India in 1997 (PA31, PA32, PA33, PA35 and PA37) and 2018 (PA198 and PA219) were investigated. All the isolates of ST308 had >95% nucleotide similarity. The isolates from 2018 had larger genomes, coding sequences, accessory and pan genes compared to the older isolates from 1997. The 2018 isolate PA219 was resistant to all antibiotics except polymyxin B, while the 2018 isolate PA198 was resistant to ciprofloxacin, levofloxacin, gentamicin and tobramycin. Among the isolates from 1997, five were resistant to gentamicin, tobramycin and ciprofloxacin, four were resistant to levofloxacin while two were resistant to polymyxin B. Twenty-four acquired resistance genes were present in the 2018 isolates compared to 11 in the historical isolates. All isolates contained genes encoding for aminoglycoside (aph(6)-Id, aph(3')-lIb, aph(3″)-Ib), beta-lactam (blaPAO), tetracycline (tet(G)), fosfomycin (fosA), chloramphenicol (catB7), sulphonamide (sul1), quaternary ammonium (qacEdelta1) and fluoroquinolone (crpP) resistance. Isolate PA198 possessed aph(3')-VI, rmtD2, qnrVC1, blaOXA-488, blaPME-1, while PA219 possessed aadA1, rmtB, qnrVC1, aac(6')-Ib-cr, blaTEM-1B, blaVIM-2, blaPAO-1, mph(E), mph(A), msr(E). In both recent isolates qnrVC1 was present in Tn3 transposon. In 219 blaTEM-1 was carried on a transposon and blaOXA-10 on a class 1 integron. There were no notable differences in the number of single nucleotide polymorphisms, but recent isolates carried more insertions and deletions in their genes. These findings suggest that genomes of P. aeruginosa ocular clonal strains with >95% nucleotide identity isolated twenty years apart had changed over time with the acquisition of resistance genes. The pattern of gene mutations also varied with more insertions and deletions in their chromosomal genes which confer resistance to antibiotics.

Linezolid shows high safety and efficacy in the treatment of Pythium insidiosum keratitis in a rabbit model.

Efficacy and safety of three antibiotics (Linezolid-LZ, 0.2%; Azithromycin-AZ, 1%; Tigecycline-TG, 1%) were determined in the treatment of Pythium insidiosum keratitis in rabbits. Infection of right eye of 38 rabbits was induced by standard intracorneal injection of P. insidiosum zoospores (left eye, intracorneal saline). Corneal infection developed in all right eyes. One hourly eye drops of one of the three antibiotics was instilled in both eyes (3 groups of 12 rabbits each) except in controls. Half of the rabbits in each group received intracorneal injection of the respective antibiotic after 4 days of starting eye drops. Clinical scoring of eyes was done over next 3 weeks. The reduction in scores post-treatment was significant for each drug (LZ: p < 0.025, AZ: p < 0.025, TG: p < 0.01). Scores with LZ (median change of 3) was significantly (p = 0.013) higher than TG (median change of 2) and comparable (p = 0.06) to AZ (median change of 3). Reduction in clinical scores in eyes receiving intracorneal antibiotics was not significantly different from the eyes that did not receive intracorneal antibiotics (p = 0.73). While no adverse effect of LZ was seen in the control corneas, 66-100% of rabbits showed reaction to AZ and TG. Histopathology showed severe inflammation in all infected corneas and intraocular extension in some of the rabbits with poor response. The success rate was noted to be 16.7%, 25% and 50% in AZ, TG and LZ respectively (p = 0.45). LZ demonstrated superior efficacy and safety and can be considered for trial in human disease.

Alterations in the conjunctival surface bacterial microbiome in bacterial keratitis patients.

Microbial keratitis is an infectious disease of the eye, in which the cornea is inflamed. Under severe conditions, keratitis can lead to significant loss of vision and enucleation of the eye. Ocular trauma is the major risk factor causing keratitis and microorganisms viz., bacteria, fungi, viruses are the causative agents. The current study characterized the conjunctival bacterial microbiomes of healthy individuals and individuals with bacterial keratitis (BK) and assessed whether ocular microbiome dysbiosis is prevalent in BK patients. Ocular bacterial microbiomes were generated from the conjunctival swabs of healthy controls (HC-SW) and conjunctival swabs (BK-SW) and corneal scrapings (BK-CR) of BK patients using V3-V4 amplicon sequencing and data analysed using QIIME and R software. The Alpha diversity indices, diversity and abundance of different phyla and genera, heat map analysis, NMDS plots and inferred functional pathway analysis clearly discriminated the bacterial microbiomes of conjunctival swabs of healthy controls from that of BK patients. Preponderance of negative interactions in the hub genera were observed in BK-CR and BK-SW compared to HC-SW. In addition, a consistent increase in the abundance of pathogenic bacteria, as inferred from published literature, was observed in the conjunctiva of BK patients compared to HC and this may be related to causing or exacerbating ocular surface inflammation. This is the first study demonstrating dysbiosis in the ocular bacterial microbiome of conjunctiva of bacterial keratitis patients compared to the eye of healthy controls. The bacterial microbiome associated with the corneal scrapings of keratitis individuals is also described for the first time.

Risk Factors for Contact Lens-Related Microbial Keratitis and Associated Vision Loss in a South Indian Population.

OBJECTIVES: To identify risk factors associated with contact lens-related microbial keratitis (CL-MK) and subsequent vision loss in a south Indian population. METHODS: A retrospective study of medical records at the LV Prasad Eye Institute in Hyderabad, India, of patients diagnosed with CL-MK and of controls who had no history of corneal inflammation during contact lens wear was undertaken. Variables such as demographic data, contact lens wear details, duration of the event, visual acuity, epithelial defect and infiltrate size, and microbiology of the cornea during the event were collected. Differences between cases and controls were analyzed using parametric and nonparametric tests. Logistic regression was used to calculate the odds ratio (OR) and associated 95% confidence intervals in univariate and multivariate analyses for cases vs. controls and for factors associated with vision loss. RESULTS: One hundred sixty-seven cases of CL-MK and 153 controls were included in the analyses. Risk factors associated with the greatest increased OR for CL-related MK were: being in professional employment vs. a student (OR=3.9), disposing lenses yearly versus monthly or biweekly (OR=2.2), and any overnight wear (OR=2.8). Risk factors associated with vision loss were: high myopia (OR=3.6), severe vs. mild severity (OR=16.0), and hypopyon (OR=4.3). CONCLUSIONS: Identification of these risk factors may help inform safe contact lens wear habits and management strategies.

Risk factors, clinical features and outcomes of Neisseria keratitis.

OBJECTIVE: To describe the clinical features, risk factors and outcomes of Neisseria keratitis. METHODS: This is a retrospective observational study wherein medical records of cases with microbiologically proven Neisseria keratitis were reviewed. Data pertaining to the underlying predisposing factors, clinical characteristics of the corneal ulcer, antibiotic susceptibility of the Neisseria species isolate from the corneal scraping, the treatment given, and outcomes were collected and analyzed. RESULTS: Medical records of 60 patients (60 eyes) with Neisseria keratitis were reviewed. Among the causes of poor ocular surface as predisposing factor, vernal keratoconjunctivitis (n = 6 eyes), along with use of topical corticosteroids (n = 18 eyes) was the most common. The ulcer was characterized by a central infiltrate (31/60, 51.7%) involving up to the mid-stroma (43/60, 71.7%). Of the forty-four (73.3%) eyes with pure Neisseria keratitis, 31 eyes (72.1%) resolved with medical therapy alone while five eyes (11.6%) underwent therapeutic penetrating keratoplasty and in two (4.6%) eyes evisceration was performed. The other 5/44 (11.6%) patients were lost to follow-up. Resolution with medical therapy was found to be similar in cases with pure infection and mixed infection (p = 0.58). CONCLUSIONS: Neisseria keratitis most commonly causes a mild form of keratitis and is often associated with the poor ocular surface or prior steroid use. In most cases medical therapy is sufficient for complete resolution of the keratitis.

ENTEROCOCCUS ENDOPHTHALMITIS: Clinical Settings, Antimicrobial Susceptibility, and Management Outcomes.

PURPOSE: To report the clinical presentation and management outcome of patients with endophthalmitis caused by Enterococcus species and to report the susceptibility profile of the isolates. METHODS: Twenty-nine cases with culture-proven Enterococcus endophthalmitis from January 2005 to May 2018 underwent vitrectomy/vitreous biopsy, intravitreal antibiotic with or without additional procedures. The undiluted vitreous was subjected to microbiologic evaluation. A favorable anatomical outcome was defined as preservation of the globe, absence of hypotony, attached retina, and absence of active inflammation at the last visit. Favorable visual outcome was final visual acuity ≥20/400. RESULTS: There were 24 men (82.8%). Mean age at presentation was 32.89 ± 25.25 years (median 24 years). Inciting event was open globe injury in 18 (62%), endogenous in 5 (17.24%), postcataract surgery in 3 (10.34%), postscleral buckling in 2 (6.89%), and postkeratoplasty in 1 (3.44%). Enterococcus casseliflavus was the commonest species isolated (14/29, 48.27%) followed by E. faecalis (9/29, 31%). Susceptibility to vancomycin was seen in 27/29 isolates (93%). Visual acuity was ≤20/400 in all eyes at presentation and ≥20/400 in 10/29 cases (34.48%) at final visit. Anatomical success was seen in 18/29 eyes (62%). Corneal involvement was high at 24/29 eyes (82.75%). CONCLUSION: Enterococcus is not an uncommon organism in the setting of endophthalmitis after open globe injury. Resistance to vancomycin is rare. Multidrug resistance pattern is restricted to E. faecalis. Visual outcome is poor despite early and appropriate therapy due to inherent organism virulence.

Multidrug-Resistant Escherichia coli Canaliculitis.

Escherichia coli canaliculitis is an exceptionally rare organism to cause primary canaliculitis. The present case describes unilateral canaliculitis refractory to conventional therapy with a significant history of recurrent culture proven E. coli urinary tract infection. Microbiological analysis revealed E. coli bacilli and histological examination showed goblet cell metaplasia, subepithelial edema with acute and chronic inflammatory infiltrate. The possibility of an endogenous infection or autoinoculation secondary to urinary tract infection cannot be ruled out.

Unusual microbiological presentations in polymicrobial post-operative endophthalmitis and their clinical correlations.

PURPOSE: To report the clinical course, microbiological spectrum and visual outcomes of three cases of post-operative endophthalmitis caused by unusual combination of micro-organisms. METHODS: Retrospective review of medical charts at a tertiary eye care centre over a period of 1 year, of subjects with post-operative endophthalmitis and more than one microbiological isolate. RESULTS: We report 3 cases with unusual combination of microorganisms. In case 1, two organisms (Burkholderia cepacia and Aeromonas veronii, from the vitreous cavity and capsular bag, respectively) with an identical antibiotic sensitivity pattern were found, while in case 2, the organisms (Streptococcus pneumoniae and Sphingomonas paucimobilis, from cornea and vitreous cavity, respectively) isolated had different sensitivity patterns. In case 3, two different strains of the same organism (Enterococcus faecalis) were found. Cases 1 and 2 achieved good anatomical and visual outcomes, while in case 3, vision remained poor despite a good anatomical outcome. CONCLUSION: Unusual combinations of organisms in post-operative endophthalmitis can introduce unique clinical characteristics and should specifically be considered in atypical clinical presentations, poor response to standard therapy and unusual recurrence patterns.

ROLE OF EARLY VITRECTOMY IN THE TREATMENT OF FUNGAL ENDOPHTHALMITIS.

PURPOSE: To study whether immediate pars plana vitrectomy at the time of sampling vitreous for culture alter the visual and structural outcomes in fungal endophthalmitis. METHODS: Surgical results of 66 consecutive patients with culture-positive fungal endophthalmitis attending a tertiary eye care hospital in eastern India were analyzed. Based on the timing of vitrectomy, they were divided into 2 groups: Group 1-immediate vitrectomy (n = 31) and Group 2-diagnostic vitrectomy alone (n = 35) as the first-line intervention. The demographics, clinical features, and visual and structural outcomes were compared between the groups. RESULTS: The proportion of postoperative (54.83 vs. 51.42%), traumatic (29.03 vs. 34.28%), and endogenous (16.12 vs. 14.28%) endophthalmitis and clinical signs such as pain (54.83 vs. 60%), lid edema (48.38 vs. 48.57%), conjunctival congestion (87.09 vs. 94.28%), and corneal involvement (87.09 vs. 80%) in Groups 1 and 2, respectively, were comparable. Logarithm of the minimum angle of resolution vision improvement in Group 1 was significant (P = 0.0278) though there was no statistically significant difference in posttreatment vision between the groups (P = 0.322). Phthisis was seen in 25.8% in Group 1 and 25.7% in Group 2 (P = 0.993). Aspergillus species was the most common fungal pathogen isolated. Early vitrectomy plus intravitreal antifungals in a subset of clinically suspected fungal endophthalmitis resulted in statistically significant visual improvement over diagnostic vitrectomy plus intravitreal antifungals (P = 0.013). CONCLUSION: A strong clinical suspicion that translates into early vitrectomy plus intravitreal antifungal antibiotics leads to favorable visual and structural outcomes. A long wait till microbiological confirmation to institute antifungal therapy may result in poorer outcome.