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1.
Genomics ; 112(6): 4769-4776, 2020 11.
Article in English | MEDLINE | ID: mdl-32882326

ABSTRACT

The several virulence and drug-resistance mechanisms of Pseudomonas aeruginosa responsible for poor clinical outcomes in keratitis patients remain largely unknown. Here, we investigated the distribution of virulence factors and drug resistance by genes, mutations, efflux-pump systems of P. aeruginosa strains from keratitis patients with different clinical outcomes, included of whole-genome sequenced and annotated our five P. aeruginosa strains. Of the large number of virulence genes detected in all the genomes, MDR/XDR strains carry exoU and non-MDR strains carry exoS exotoxin of the type III secretion system, considered as main contributors of keratitis pathogenesis. However, several strain-specific virulence and resistance genes were detected in keratitis strains with poor outcome. Mainly, the flagellar genes fliC and fliD detected in the exoS carrying strains, reported to alter the host immune response, might impact the clinical outcome. This study may provide new insights into the genome of ocular strains and requires further functional studies.


Subject(s)
Keratitis/microbiology , Pseudomonas Infections/microbiology , Pseudomonas aeruginosa/genetics , Drug Resistance, Bacterial/genetics , Genome, Bacterial , Genomics , Humans , Microbial Sensitivity Tests , Virulence Factors/genetics
2.
Exp Eye Res ; 186: 107700, 2019 09.
Article in English | MEDLINE | ID: mdl-31233730

ABSTRACT

Corneal mycotic ulceration is predominantly due to Aspergillus and Fusarium solani infection in tropical countries. In this study, we examined the proteome profile of tear samples from A. flavus keratitis patients at various stages of infection. The proteome was profiled using 2D PAGE and the protein levels were quantified using 2D DIGE. Alpha-1-antitrypsin, apolipoprotein, haptoglobin, lactoferrin and albumin were up regulated while cystatin SA III precursor, lacrimal lipocalin precursor, lacritin precursor and Zinc alpha-2 glycoprotein (ZAG) were down regulated in tear fluid. In the case of ZAG all proteoforms were down regulated as the disease progressed from early to late stage of infection. Western blot analysis confirmed the results observed using DIGE. Further, there were no gender specific differences in the levels of ZAG expression in keratitis patient tear film. Published results show up regulation of ZAG in Fusarium keratitis patient tear indicating subtle changes in the early events of host response to these two fungal pathogens. We conclude that ZAG level could be used as an indicator of A. flavus or F. solani infection, even during the early stage of the disease.


Subject(s)
Aspergillosis , Aspergillus flavus , Eye Infections, Fungal , Keratitis , Proteome/metabolism , Seminal Plasma Proteins/metabolism , Tears/metabolism , Adolescent , Adult , Aged , Aspergillosis/metabolism , Aspergillosis/microbiology , Down-Regulation , Eye Infections, Fungal/metabolism , Eye Infections, Fungal/microbiology , Female , Humans , Keratitis/metabolism , Keratitis/microbiology , Male , Middle Aged , Young Adult , Zn-Alpha-2-Glycoprotein
3.
J Infect Dis ; 211(1): 130-4, 2015 Jan 01.
Article in English | MEDLINE | ID: mdl-25001461

ABSTRACT

Interleukin 17A (IL-17) production by peripheral blood neutrophils was examined in patients with fungal keratitis and in uninfected individuals in southern India, which has high levels of airborne Aspergillus and Fusarium conidia. Il17a gene expression and intracellular IL-17 were detected in all groups, although levels were significantly elevated in neutrophils from patients with keratitis. There were no significant differences in plasma IL-17 and IL-23 between patients with keratitis and uninfected individuals; however, combined data from all groups showed a correlation between the percentage IL-17 producing neutrophils and plasma IL-23, and between plasma IL-17 and IL-6 and IL-23.


Subject(s)
Eye Infections, Fungal/blood , Eye Infections, Fungal/microbiology , Interleukin-17/biosynthesis , Keratitis/blood , Keratitis/microbiology , Neutrophils/immunology , Adult , Aspergillosis/genetics , Aspergillosis/immunology , Aspergillosis/microbiology , Aspergillus/immunology , Case-Control Studies , Cohort Studies , Eye Infections, Fungal/immunology , Fusariosis/blood , Fusariosis/genetics , Fusariosis/immunology , Fusariosis/microbiology , Fusarium/immunology , Humans , India , Interleukin-17/blood , Interleukin-17/genetics , Interleukin-23/biosynthesis , Interleukin-23/blood , Interleukin-23/genetics , Interleukin-6/biosynthesis , Interleukin-6/blood , Interleukin-6/genetics , Interleukin-6/immunology , Keratitis/genetics , Keratitis/immunology , Middle Aged
4.
Cornea ; 42(1): 110-112, 2023 Jan 01.
Article in English | MEDLINE | ID: mdl-36036669

ABSTRACT

PURPOSE: Outcomes of Acanthamoeba keratitis are often worse in India than in the United States. The goal of this study was to determine whether antiamoebic susceptibility patterns were different when comparing Acanthamoeba isolates from India with those of the United States. METHODS: Acanthamoeba isolates were obtained from corneal scrapings of 43 patients with infectious keratitis seen at the Francis I. Proctor Foundation (N = 23) and Aravind Eye Hospital (N = 20) from 2008 through 2012 and plated on growth media. A previously described minimum cysticidal concentration (MCC) assay was performed by a single laboratory technician to assess susceptibility to 5 antiamoebic agents for all isolates. Testing was conducted in triplicate, with the median MCC chosen for analyses. RESULTS: The MCC (µg/mL) of polyhexamethylene biguanide was 6.25 [IQR 5.47-12.5] for Aravind isolates and 6.25 [IQR 6.25-9.375] for Proctor isolates ( P = 0.75), corresponding values were 6.25 [IQR 3.125-6.25] and 3.125 [IQR 3.125-9.375] for chlorhexidine ( P = 0.81), 2500 [IQR 2500-5000] and 5000 [IQR 1250-20,000] for voriconazole ( P = 0.25), 15.6 [IQR 15.6-39.0625] and 15.6 [IQR 15.6-31.25] for hexamidine ( P = 0.92), and 15.6 [IQR 7.81-15.6] and 15.6 [IQR 7.81-31.25] for propamidine ( P = 0.42). CONCLUSIONS: This study found no statistically significant differences in antiamoebic susceptibility of Indian versus US samples from Acanthamoeba keratitis clinical isolates. These findings suggest that differences in antiamoebic susceptibility are likely not responsible for differential outcomes in Acanthamoeba keratitis between the 2 locations.


Subject(s)
Acanthamoeba Keratitis , Acanthamoeba , Humans , Acanthamoeba Keratitis/drug therapy , Chlorhexidine/pharmacology , Voriconazole/pharmacology , California
5.
J Infect Dis ; 204(6): 942-50, 2011 Sep 15.
Article in English | MEDLINE | ID: mdl-21828275

ABSTRACT

BACKGROUND: Filamentous fungi of the genera Aspergillus and Fusarium are major causes of corneal ulcers in the United States and in the developing world and result in significant visual impairment and blindness. METHODS: RNA was extracted from 110 patients with corneal ulcers in southern India within 1 week of infection with either Fusarium solani or Aspergillus flavus, and gene expression was determined by quantitative polymerase chain reaction. Posttransplant corneas from later stage disease (>2 weeks after infection) were also examined. RESULTS: Expression of Dectin-1, Toll-like receptor 2 (TLR2), TLR4, TLR9, and NOD-like receptor protein (NLRP)3 messenger RNA was elevated >1000-fold compared with uninfected donor corneas, whereas Dectin-2 was constitutively expressed in uninfected corneas. Furthermore, interleukin 1ß (IL-1ß) expression was elevated >1000-fold, whereas IL-1α expression was not increased. Expression of IL-8, IL-17, and tumor necrosis factor α was also elevated. CD3(+)and CD4(+) T cells were detected in infected posttransplant corneas. Expression of IL-17 and interferon γ was elevated but not that of IL-4. There were no significant differences in the host response between Aspergillus- and Fusarium-infected corneas at any time point. CONCLUSIONS: There is a common innate and adaptive immune response to these filamentous fungi, which includes the generation of T-helper 1 and T-helper 17 cells.


Subject(s)
Aspergillus flavus/immunology , Cornea/immunology , Fusarium/immunology , Gene Expression Profiling , Keratitis/immunology , Mycoses/immunology , Adolescent , Adult , Aged , Aged, 80 and over , Female , Humans , India , Keratitis/microbiology , Male , Middle Aged , Mycoses/microbiology , Reverse Transcriptase Polymerase Chain Reaction , Th1 Cells/immunology , Th17 Cells/immunology , Young Adult
6.
Cornea ; 41(11): 1386-1391, 2022 Nov 01.
Article in English | MEDLINE | ID: mdl-34743096

ABSTRACT

PURPOSE: The objective of this study was to compare the clinical and microbiological profiles of culture-proven pure Corynebacterium keratitis with mixed infection and their antibiotic susceptibility patterns over a 2-year period. METHODS: A retrospective analysis of culture-proven cases of Corynebacterium keratitis over a 2-year period was performed in 3 different tertiary eye care centers. All isolates were tested for antibiotic susceptibility in vitro using the disc-diffusion method for 7 antibiotics. RESULTS: Altogether 108 cases were identified as culture-positive Corynebacterium keratitis in 3 tertiary eye care centers. Of these, 60.2% (n = 65) and 39.8% (n = 43) of cases were due to pure Corynebacterium and mixed infection, respectively. The mean duration of symptoms was 23.2 ± 29.6 days. In the mixed-infection group, fungus was identified as the coexistent pathogen in 22 cases (51.1%). Ocular surface disorder was the most common risk factor (33.9%) in Corynebacterium keratitis. The most frequently isolated species was Corynebacterium amycolatum (22.2%) in both groups. Therapeutic keratoplasty was performed in 8.3% of cases. There was no significant difference in the outcome between the 2 groups. Cefazolin resistance was seen in 13.9% of patients, and all isolates were susceptible to vancomycin. The resistance pattern showed emerging resistance toward fluoroquinolone because the isolates were resistant to gatifloxacin (58.3%), moxifloxacin (47.2%), ciprofloxacin (54.6%), and ofloxacin (45.4%). CONCLUSIONS: Ocular surface disorder is the most common risk factor in Corynebacterium keratitis. Although fluoroquinolones are commonly used as first-line therapy in microbial keratitis, the in vitro resistance pattern indicates that these are less likely to be effective in infection with Corynebacterium species.


Subject(s)
Coinfection , Eye Infections, Bacterial , Keratitis , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/therapeutic use , Cefazolin , Ciprofloxacin/therapeutic use , Coinfection/drug therapy , Corynebacterium , Eye Infections, Bacterial/drug therapy , Eye Infections, Bacterial/microbiology , Fluoroquinolones/pharmacology , Fluoroquinolones/therapeutic use , Gatifloxacin , Humans , Keratitis/drug therapy , Keratitis/microbiology , Microbial Sensitivity Tests , Moxifloxacin/therapeutic use , Ofloxacin/therapeutic use , Retrospective Studies , Tertiary Care Centers , Vancomycin/therapeutic use
7.
Exp Eye Res ; 92(6): 454-63, 2011 Jun.
Article in English | MEDLINE | ID: mdl-21396361

ABSTRACT

The proteomic profile of tear fluid is of fundamental interest in eye research. In this study we optimized the tear sample preparation method for two-dimensional (2D) analysis and determined the protein profile of tear fluid from healthy males and females. To find the most efficient method for tear sample preparation, four widely applied precipitation methods and ultrafiltration were compared. Of these, TCA precipitation & ultrafiltration resulted in efficient sample concentration and desalting. Use of a nonionic wetting agent, Tergitol NP7, in rehydration solution during isoelectric focusing improves protein separation in 2D gel electrophoresis considerably. Using this optimized method, tear protein profile was analyzed from healthy males and females. Of the thirty six tear proteins identified by LC-MS/MS, seven tear proteins were found to be significantly up regulated in the healthy female tear samples when compared to the male tear samples. These results indicate that the tear protein profile differs with respect to the sex. Mostly, the up regulated proteins are involved in the local immune defense; implying that there may be a sex difference in the ability to defend against infection at the anterior segment of the eyes of normal individuals.


Subject(s)
Eye Proteins/analysis , Proteome/analysis , Tears/chemistry , Electrophoresis, Gel, Two-Dimensional/methods , Fatty Alcohols/pharmacology , Female , Humans , Isoelectric Focusing , Male , Molecular Sequence Data , Peptide Fragments/analysis , Proteomics/methods , Sequence Analysis, Protein , Sex Factors , Tandem Mass Spectrometry
8.
Indian J Ophthalmol ; 69(5): 1068-1072, 2021 05.
Article in English | MEDLINE | ID: mdl-33913834

ABSTRACT

Purpose: Earlier our group has demonstrated the drug reservoir function of the human amniotic membrane (HAM) using stable moxifloxacin and fortified cefazolin ophthalmic formulations and found it as a suitable tool to deliver drugs for an extended duration. The purpose of this study was to evaluate the extended-release kinetics of voriconazole from the impregnated human amniotic membrane (HAM) in vitro. Methods: HAM buttons were incubated with freshly prepared 1% topical ophthalmic formulation of voriconazole for 5 different exposure time to investigate the ideal exposure time for the extended-release of voriconazole from HAM. The drug release kinetics was studied in simulated tear fluid for 5 weeks and the amount of voriconazole released at different intervals was estimated using high-performance liquid chromatography (HPLC) with photodiode array (PDA) detector. Results: There was a marginal increase in drug entrapment efficiency with increased drug exposure time but neither the drug entrapment nor the drug release was found to be statistically significant (P ≥ 0.5). Voriconazole was detectable even at 5 weeks. Conclusion: A sustained release of voriconazole was achieved up to 5 weeks, when voriconazole was incubated with amniotic membrane for all the studied drug soaking times. Thus, voriconazole impregnated amniotic membrane can be considered for the sustained delivery for its in fungal keratitis.


Subject(s)
Eye Infections, Fungal , Pharmaceutical Preparations , Amnion , Antifungal Agents/therapeutic use , Eye Infections, Fungal/drug therapy , Humans , Moxifloxacin , Voriconazole
9.
Transl Vis Sci Technol ; 10(5): 6, 2021 04 29.
Article in English | MEDLINE | ID: mdl-34003973

ABSTRACT

Purpose: We aimed to develop a novel chemical cross-linker treatment for keratoconus by reacting dicarboxylic acid spacer molecules and amine functional groups on protein structure of the tissue using carbodi-imide chemistry. We propose this as an alternative to conventional cross-linking treatment for keratoconus. Methods: The study involved optimization of the cross-linker formulation. Mechanical stiffness of ex vivo porcine and human corneas after application of the cross-linker was measured. Histochemical analysis was performed to record changes in gross morphology after cross-linker treatment on ex vivo porcine and human and in vivo rabbit corneas. Terminal deoxynucleotidyl transferase-mediated dUTP-X nick-end-labeling (TUNEL) staining was performed to study apoptotic effects of cross-linker. Cytotoxicity potential of cross-linker was evaluated by studying explant cultures for cellular outgrowth and immunostaining assays on porcine and human corneas after treatment. Results: We demonstrated a clinically relevant increase in stiffness in ex vivo experiments using porcine and human cornea without removal of corneal epithelium. Histological analysis showed no change in gross morphology of cornea and no evidence of apoptosis. In vivo treatment of rabbit eyes demonstrated initial thinning of corneal epithelium that recovered after seven days although with abnormal regularity of cells. Cellular outgrowth from corneal explant cultures after treatment further confirmed cell survival after treatment. Conclusions: This chemical cross-linking of corneal tissue has potential advantages over current therapeutic options including lower cytotoxicity to stromal cells than ultraviolet A treatment. Translational Relevance: The cross-linker has potential to become a treatment for keratoconus because it overcomes the need for procedures using specialized equipment and ensures accessibility to large populations.


Subject(s)
Keratoconus , Animals , Cornea , Cross-Linking Reagents , Humans , Keratoconus/drug therapy , Rabbits , Riboflavin , Swine , Ultraviolet Rays
10.
Indian J Ophthalmol ; 67(4): 472-475, 2019 04.
Article in English | MEDLINE | ID: mdl-30900576

ABSTRACT

Purpose: Our previous study demonstrated the drug reservoir function of human amniotic membrane (HAM) using stable moxifloxacin as a model drug. The purpose of the present study is to evaluate whether HAM can be used as a drug carrier for extended release of extemporaneous preparation of cefazolin. Methods: HAM Buttons (1 Control, 5 Test) were incubated in a freshly prepared (1 ml) sterile topical solution of cefazolin 5% (w/v) for 3 h and 24 h at two different temperatures. The groups were designated as follows: Group IA: Soaking duration 3 h at 4°C; Group IB: Soaking duration 3 h at room temperature; Group IIA: Soaking duration 24 h at 4°C; and Group IIB: Soaking duration 24 h at room temperature. The release kinetics of cefazolin from different groups of drug-laden HAM was studied for a period of 5 days. Samples were assayed for estimation of cefazolin content at different time intervals by High Performance Liquid Chromatography (HPLC) with Photodiode array (PDA) detector. Results: Three-hour cefazolin treatment with HAM at 4°C caused high drug entrapment (24%) compared to room temperature (11%; P < 0.005); however, the release kinetics was not significantly different between Group IA and IB as well as Group IIA and IIB up to the study period. Increase in drug treatment duration did not show increase in entrapment, but caused two-fold (IA Vs IIA) and 1.6-fold (IB Vs IIB) less drug entrapment at 4°C and room temperature, respectively. Conclusion: The results reveal that HAM may be a suitable drug carrier for extended delivery of fortified formulations without compromising stability.


Subject(s)
Amnion , Cefazolin/pharmacokinetics , Corneal Ulcer/drug therapy , Drug Carriers , Tears/chemistry , Anti-Bacterial Agents/pharmacokinetics , Cells, Cultured , Chromatography, High Pressure Liquid , Corneal Ulcer/metabolism , Corneal Ulcer/pathology , Drug Stability , Female , Humans , Ophthalmic Solutions
11.
Mol Vis ; 14: 500-7, 2008 Mar 12.
Article in English | MEDLINE | ID: mdl-18385783

ABSTRACT

PURPOSE: Mycotic keratitis is a major cause of corneal blindness in India. A proper understanding of the pathogenesis may help in refining the existing treatment. The purpose of this study is to examine the total tear protein profile of fungal keratitis patients, which may have a bearing on pathogenesis and disease progression. METHODS: Tear samples were collected from culture positive fungal keratitis patients. Tears from the uninfected fellow eye and from other healthy individuals served as controls. Two-dimensional electrophoresis (2DE) was used for the separation of fractionated tear proteins, and selected protein spots, which showed differential expressions, were identified using matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry. Wherever needed, tag sequencing of peptide fragments using post source decay (PSD) was done to confirm the identification. RESULTS: The glutaredoxin-related protein was expressed only in the tears of fungal keratitis patients. Six other normal tear proteins were present in both samples but with varied expression levels. Prolactin inducible protein and serum albumin precursor were upregulated in the infected samples. Cystatin S precursor, cystatin SN precursor, cystatin, and human tear lipocalin were downregulated in the infected samples. CONCLUSIONS: Tears can be used as a clinical source to study the proteomic responses in patients with fungal keratitis. The glutaredoxin-related protein is known to be produced by Aspergillus during oxidative stress conditions, and the presence of this protein in the tears of patients with mycotic keratitis indicates that this pathogen undergoes stress-related gene expression during infection.


Subject(s)
Aspergillosis , Eye Proteins/metabolism , Fusarium , Keratitis/metabolism , Keratitis/microbiology , Mycoses , Chromatography, Liquid , Electrophoresis, Gel, Two-Dimensional , Humans , Proteomics , Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization , Tandem Mass Spectrometry
12.
Arch Ophthalmol ; 125(6): 789-93, 2007 Jun.
Article in English | MEDLINE | ID: mdl-17562990

ABSTRACT

OBJECTIVE: To characterize the susceptibility of filamentous fungi isolated from keratitis to amphotericin B, natamycin, caspofungin acetate, itraconazole, voriconazole, and posaconazole. METHODS: Ninety isolates from fungal keratitis cases at Aravind Eye Hospital in South India were tested using macrobroth dilution for susceptibility to amphotericin B, natamycin, caspofungin, itraconazole, voriconazole, and posaconazole. The minimum inhibitory concentration (MIC) median and 90th percentile were determined. RESULTS: The 90 isolates included 41 Aspergillus species, 38 Fusarium species, and 11 others. The triazoles and caspofungin had the lowest MICs against Aspergillus species; voriconazole, amphotericin B, and posaconazole had the lowest MICs against Fusarium species, and none of the Fusarium species were inhibited by itraconazole or caspofungin. Amphotericin B had significantly lower MICs compared with natamycin, but after correcting for the typical prescription dose, natamycin was superior. CONCLUSION: No single agent was universally most effective, but voriconazole and other triazoles demonstrated the broadest spectrum. Itraconazole and caspofungin were not effective against Fusarium species. CLINICAL RELEVANCE: Fungal ulcers are commonly treated empirically; drugs are typically selected without regard to susceptibility data. The nonocular infectious disease literature suggests modern fungal susceptibility methods are clinically relevant, but ocular studies are limited. Our results suggest antifungal therapy might be tailored to individual organisms.


Subject(s)
Antifungal Agents/pharmacology , Aspergillus/drug effects , Corneal Ulcer/microbiology , Eye Infections, Fungal/microbiology , Fusarium/drug effects , Amphotericin B/pharmacology , Aspergillus/isolation & purification , Caspofungin , Echinocandins , Fusarium/isolation & purification , Humans , Itraconazole/pharmacology , Lipopeptides , Microbial Sensitivity Tests , Natamycin/pharmacology , Peptides, Cyclic/pharmacology , Prospective Studies , Pyrimidines/pharmacology , Triazoles/pharmacology , Voriconazole
13.
Am J Ophthalmol ; 144(1): 144-6, 2007 Jul.
Article in English | MEDLINE | ID: mdl-17601443

ABSTRACT

PURPOSE: To analyze cases with posttraumatic lenticular abscess and study the etiology, clinical presentation, management, and outcome. DESIGN: Retrospective case series. METHODS: Seventeen eyes of 17 patients with traumatic lenticular abscesses were managed with extracapsular cataract extraction after aspirating the abscess. RESULTS: The mean age of the patients was 40.3 years, and males constituted 82%. The mean time to presentation after injury was 14.35 days (range, one to 60 days), and the patients had a mean follow-up of 125.94 days (range, 21 to 300 days). Culture of the lenticular abscess revealed bacterial growth in eight cases (47%) and fungi in four cases (23.5%). In five (29.4%) cases, culture was negative. Staphylococcus epidermidis grew in seven cases (41%). Thirteen eyes (77%) had best-corrected visual acuity better than 20/120. CONCLUSIONS: Surgical removal of the abscess, with systemic and local antimicrobial treatment is effective in cases of posttraumatic intralenticular abscess.


Subject(s)
Abscess/etiology , Eye Infections, Bacterial/etiology , Eye Infections, Fungal/etiology , Eye Injuries, Penetrating/complications , Lens Diseases/etiology , Lens, Crystalline/injuries , Abscess/therapy , Adult , Anti-Bacterial Agents/therapeutic use , Cataract Extraction , Combined Modality Therapy , Eye Infections, Bacterial/therapy , Eye Infections, Fungal/therapy , Eye Injuries, Penetrating/therapy , Female , Humans , Lens Diseases/therapy , Lens Implantation, Intraocular , Male , Retrospective Studies , Treatment Outcome
14.
Cornea ; 26(2): 207-11, 2007 Feb.
Article in English | MEDLINE | ID: mdl-17251814

ABSTRACT

PURPOSE: To study the infiltrating cells and quantify the levels of matrix metalloproteinases (MMP-8, MMP-9) and tissue inhibitor of metalloproteinases (TIMP-1, TIMP-2) in the cornea, tear, and serum of patients with fungal keratitis. METHODS: Experimental study. Infected corneal tissue from 4 patients with fungal keratitis (group 1) scheduled to undergo therapeutic keratoplasty accounted for the histopathologic and cytospin smear analysis. Ten patients with fungal keratitis from group 2 served for the quantification of MMPs and TIMPs. Five patients with keratoconus undergoing penetrating keratoplasty and 5 cadaver corneas were chosen as controls for group 2. Corneal buttons obtained during keratoplasty, 15 to 20 microL of tears collected using the capillary flow method, and 3 mL of blood was obtained from patients with fungal keratitis and patients with keratoconus. Corneal button sections from group 1 were stained with hematoxylin and eosin and Grocott methenamine silver nitrate for the histopathologic studies and Giemsa staining for the cytospin smear analysis. Enzyme-linked immunosorbent assay was used for the quantification of total MMP-8, MMP-9, TIMP-1, and TIMP-2 in the corneal homogenates, tear, and serum samples of group 2. RESULTS: Corneal sections from group 1 revealed dense fungal filaments and a large proportion (91.4% +/- 38%) of polymorphonuclear leukocytes (PMNs). Significant elevation in the levels of MMP-8 and MMP-9 (P < 0.05) in the fungal keratitis corneas was observed in group 2 compared with the cadaver and keratoconus corneas. The ratio of MMP/TIMP was also higher in the fungal keratitis corneas. CONCLUSIONS: Infiltrating PMNs in the cornea of patients with fungal keratitis contributed to the increased activities of MMP-8 and MMP-9, thereby enhancing tissue destruction and derangement.


Subject(s)
Eye Infections, Fungal/enzymology , Keratitis/enzymology , Matrix Metalloproteinase 8/metabolism , Matrix Metalloproteinase 9/metabolism , Tissue Inhibitor of Metalloproteinase-1/metabolism , Tissue Inhibitor of Metalloproteinase-2/metabolism , Adult , Aged , Aged, 80 and over , Biomarkers/metabolism , Cornea/enzymology , Cornea/microbiology , Cornea/pathology , Enzyme-Linked Immunosorbent Assay , Eye Infections, Fungal/microbiology , Eye Infections, Fungal/pathology , Female , Follow-Up Studies , Humans , Keratitis/microbiology , Keratitis/pathology , Male , Middle Aged , Neutrophils/pathology , Severity of Illness Index , Tears/enzymology
15.
Indian J Ophthalmol ; 55(1): 27-31, 2007.
Article in English | MEDLINE | ID: mdl-17189883

ABSTRACT

AIM: To determine the cellular profile and cytokine levels in the tear fluid of fungal keratitis patients. MATERIALS AND METHODS: Tear samples were collected from six fungal keratitis patients (Group I) from active stages of the disease up to resolution. Tears collected from the following served as controls: uninfected fellow eye (Group II A) of Group I, patients undergoing cataract surgery (Group II B) and acute conjunctivitis (Group II C). The cellular profile was evaluated. Interleukines (IL-6, IL-8 and IL-1beta) were estimated using sandwich enzyme immunoassay. Statistical analysis was carried out using nonparametric two-sample median test. RESULTS: Polymorphonuclear leukocytes (PMN) were the predominant infiltrating cells in Group I. During the initial stages of fungal infection, levels of IL-6 and IL-8 in the tear samples were found to be significantly increased when compared with Group II A (P=0.019 for IL-6, P<0.001 for IL-8). This was also true for IL -8 (P=0.008) levels in Group I and Group II B). While IL-6 levels decreased significantly towards healing, IL-8 remained slightly elevated even after healing. These cytokines were at the base level in Group II A. Lymphocytes and PMN were present in equal proportions in Group II C, which showed elevated levels of cytokines but not to the extent of Group I. CONCLUSION: This horizontal study indicates that understanding the nature of the inflammatory response in the tears of fungal keratitis patients is of considerable interest and warrants further investigations.


Subject(s)
Cytokines/metabolism , Keratitis/microbiology , Keratitis/pathology , Mycoses , Neutrophil Infiltration , Tears/cytology , Tears/metabolism , Adult , Humans , Keratitis/metabolism , Lymphocytes/pathology , Male , Middle Aged , Pilot Projects
17.
Br J Ophthalmol ; 101(8): 1119-1123, 2017 08.
Article in English | MEDLINE | ID: mdl-28043985

ABSTRACT

BACKGROUND: Clinical outcomes in fungal keratitis vary between Fusarium and Aspergillus spp, therefore distinguishing between species using morphological features such as filament branching angles, sporulation along filaments (adventitious sporulation) or dichotomous branching may be useful. In this study, we assessed these three features within Heidelberg Retina Tomograph 3 in vivo confocal microscopy (IVCM) images from culture-positive Fusarium and Aspergillus spp keratitis participants. METHODS: Prospective observational cohort study in Aravind Eye Hospital (February 2011-February 2012). Eligibility criteria: age ≥18 years, stromal infiltrate ≥3 mm diameter, Fusarium or Aspergillus spp culture-positive. EXCLUSION CRITERIA: previous/current herpetic keratitis, visual acuity <6/60 in fellow eye, >80% corneal thinning. IVCM was performed and images analysed for branch angle, presence/absence of adventitious sporulation or dichotomous branching by a grader masked to the microbiological diagnosis. RESULTS: 98 participants were included (106 eligible, 8 excluded as no measurable branch angles); 68 were positive for Fusarium spp, 30 for Aspergillus spp. Mean branch angle for Fusarium spp was 59.7° (95% CI 57.7° to 61.8°), and for Aspergillus spp was 63.3° (95% CI 60.8° to 65.8°), p=0.07. No adventitious sporulation was detected in Fusarium spp ulcers. Dichotomous branching was detected in 11 ulcers (7 Aspergillus spp, 4 Fusarium spp). CONCLUSIONS: There was very little difference in the branching angle of Fusarium and Aspergillus spp. Adventitious sporulation was not detected and dichotomous branching was infrequently seen. Although IVCM remains a valuable tool to detect fungal filaments in fungal keratitis, it cannot be used to distinguish Fusarium from Aspergillus spp and culture remains essential to determine fungal species.


Subject(s)
Aspergillosis/diagnostic imaging , Eye Infections, Fungal/diagnostic imaging , Keratitis/diagnostic imaging , Adult , Aged , Aspergillus/isolation & purification , Corneal Ulcer/diagnostic imaging , Corneal Ulcer/microbiology , Female , Fusarium/isolation & purification , Humans , Keratitis/microbiology , Male , Microscopy, Confocal , Middle Aged , Young Adult
18.
Ophthalmology ; 113(4): 526-30, 2006 Apr.
Article in English | MEDLINE | ID: mdl-16581414

ABSTRACT

PURPOSE: To identify risk factors at diagnosis that can serve as prognostic indicators of primary treatment failure in cases of fungal keratitis. DESIGN: Prospective, nonrandomized, interventional, comparative study. PARTICIPANTS: A total of 115 consecutive patients with fungal keratitis treated at one center during a 6-month period. METHODS: Patients with a microscopic corneal ulcer smear that was positive for fungus were enrolled and treated with 5% natamycin monotherapy according to the protocol of the hospital. Treatment responses were assessed at the end of 4 weeks. The prognostic indicators were used in a Poisson model for multiple regression analysis to estimate the relative risk of the main prognostic variables. MAIN OUTCOME MEASURES: Response of the ulcer to treatment. RESULTS: Of the 115 patients analyzed in the study, 52 (45.2%) were treatment successes, 27 (23.5%) had slow-healing ulcers, and 36 (31.3%) were refractory to primary treatment. Multivariate analysis showed that the predictors of treatment failure were ulcers that exceeded 14 mm(2) (P = 0.009), the presence of hypopyon (P = 0.003), and identification of Aspergillus (P = 0.003). CONCLUSION: In patients with fungal keratitis treated with 5% natamycin monotherapy, larger ulcer size and infection with Aspergillus were predictors of a poor outcome.


Subject(s)
Antifungal Agents/therapeutic use , Corneal Ulcer/drug therapy , Corneal Ulcer/microbiology , Eye Infections, Fungal/drug therapy , Mycoses/drug therapy , Natamycin/therapeutic use , Adolescent , Adult , Aged , Child , Child, Preschool , Corneal Ulcer/diagnosis , Eye Infections, Fungal/diagnosis , Eye Infections, Fungal/microbiology , Female , Fungi/isolation & purification , Humans , Male , Middle Aged , Mycoses/diagnosis , Mycoses/microbiology , Prospective Studies , Risk Factors , Treatment Failure , Treatment Outcome
19.
Indian J Ophthalmol ; 54(1): 23-7, 2006 Mar.
Article in English | MEDLINE | ID: mdl-16531666

ABSTRACT

PURPOSE: To record the natural history of herpes simplex keratitis and estimate visual impairment in eyes of patients diagnosed with herpes simplex keratitis. MATERIALS: This was a retrospective, descriptive case study for five years of the eyes of patients with clinically diagnosed herpes simplex keratitis. These patients had presented to the cornea services and a cornea specialist carried out the ocular examination under slit-lamp magnification. The medical records were reviewed by the authors. Visual acuity at presentation and clinical details on examination were recorded and, diagnosis of the stage of disease given in the case record was noted. Statistical analysis was done using chi-square-test and Fisher exact test. RESULTS: Of a total of 212 patients (220 eyes), there were 144 males and 68 females; 118 eyes presented with stromal lesions, 44 eyes with epithelial lesions, 39 eyes with epithelial and stromal lesions and 19 eyes with endothelitis. The improvement in visual acuity to more than 20/40 in the four clinical groups was 23 out of 44 eyes with epithelial lesion (52.27%), 83 out of the 118 eyes with stromal lesions (70.33%), 23 out of 39 eyes with epithelial and stromal lesions (58.97%) and 8 out of the 19 eyes with endothelitis (42.10%). The mean improvement in visual acuity was 3.44 lines (SD +/- 4.06). CONCLUSION: Most patients had mild disease. The majority of the patients attained the final visual acuity of 6/12 or better in the affected eye. Thus, there was only a temporary visual morbidity with fairly good functional visual outcome in majority of the patients.


Subject(s)
Keratitis, Herpetic/complications , Vision, Low/etiology , Adolescent , Adult , Aged , Child , Child, Preschool , Cornea/pathology , Female , Humans , Infant , Keratitis, Herpetic/pathology , Male , Middle Aged , Retrospective Studies , Severity of Illness Index , Visual Acuity
20.
Ophthalmology ; 112(11): 1884-9, 2005 Nov.
Article in English | MEDLINE | ID: mdl-16157381

ABSTRACT

OBJECTIVE: To determine the incidence of acute-onset postcataract endophthalmitis, identify risk factors, and determine clinical outcomes in a tertiary eye care center in South India. DESIGN: Retrospective, interventional, observational case series. PARTICIPANTS: All patients who underwent cataract extraction from January 2002 to December 2003 at Aravind Eye Hospital, Madurai. METHODS: The records of patients with acute endophthalmitis after cataract surgery were reviewed. MAIN OUTCOME MEASURES: Acute-onset postcataract endophthalmitis, incidence rates, risk factors, organisms cultured, and visual acuity outcomes after treatment. RESULTS: During the study period, 36072 cataract surgeries were performed; these included 22294 cases of phacoemulsification and 9503 cases of extracapsular cataract extraction. A total of 19 eyes developed acute endophthalmitis after cataract surgery (average 2-year incidence, 0.05%). Ten cases were culture positive (average 2-year incidence, 0.03%). In multivariate analysis, increased risk of endophthalmitis was associated with intraoperative complications, such as posterior capsular rent and vitreous loss (relative risk [RR], 6.57; 95% confidence interval [CI], 1.5-28.5; P = 0.05) for all endophthalmitis cases, age >60 years for culture-positive endophthalmitis cases (RR, 6; 95% CI, 0.7-47.8; P = 0.04), and an extracapsular cataract extraction technique for culture-positive endophthalmitis cases (RR, 4.9; 95% CI, 1.2-19.3; P = 0.001). After follow-up of 37 days, 5 (29.41%) of the 17 patients achieved a best-corrected visual acuity (BCVA) of 20/40, and the remaining 7 (32.3%) had a BCVA better than 20/200. Of the culture-positive cases, Nocardia species was the most common organism isolated, accounting for more than half of these cases (6/10 [60%]). CONCLUSIONS: This study found the incidence of acute postoperative endophthalmitis after cataract surgery to be 0.05%. Extracapsular cataract extraction technique and the occurrence of intraoperative complications are major risk factors for developing endophthalmitis. Visual outcome after endophthalmitis was generally poor. Nocardia is a lesser-known but virulent organism causing endophthalmitis, the management of which still poses a therapeutic dilemma.


Subject(s)
Cataract Extraction , Endophthalmitis/epidemiology , Postoperative Complications/epidemiology , Acute Disease , Adult , Aged , Aged, 80 and over , Bacteria/isolation & purification , Endophthalmitis/microbiology , Female , Follow-Up Studies , Humans , Incidence , India/epidemiology , Male , Middle Aged , Postoperative Complications/microbiology , Retrospective Studies , Risk Factors , Time Factors , Treatment Outcome , Visual Acuity/physiology
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