Mahendra Shah

The incidence of fungal keratitis and endophthalmitis following penetrating keratoplasty

PURPOSE To determine the incidence of postkeratoplasty fungal endophthalmitis and keratitis at the New York Eye and Ear Infirmary. To determine whether there is a relationship between culture-positive corneoscleral donor material and postoperative infection. METHODS The microbiologic records of corneoscleral donor rims submitted for culture following penetrating keratoplasty at the New York Eye and Ear Infirmary between January 1998 and January 2003 were reviewed. The incidence of rim cultures positive for fungi was tabulated. Clinical outcome measures were recorded for each patient receiving corneal donor tissue. RESULTS Of 2466 donor corneoscleral rims cultured during the study period, 344 were positive for microbial growth (13%). Of those rims with positive cultures, 28 (8.6%) were positive for fungus. All fungi cultured were Candida species. Four of the 28 recipient eyes (14%) who received contaminated donor material went on to develop postkeratoplasty fungal infections. There were no cases of fungal infection in any postkeratoplasty patients in the absence of contaminated donor rims during the study period. Overall, there was a 0.16% incidence of fungal infection (4/2466) following penetrating keratoplasty. There were 18 positive donor rims identified in the first 4 years of the study, but there were 10 cases in the last 10 months of the study. CONCLUSIONS The overall incidence of fungal infection following penetrating keratoplasty is low, but all cases in our study were associated with positive rim cultures. Whether prophylactic antifungal therapy would be of any benefit in the presence of a positive corneoscleral rim culture has not yet been determined.

Bacteria size determination by elastic light scattering

Light extinction and angular scattering measurements were performed on three species of bacteria with different sizes and shapes ( Pseudomonas aeruginosa, Staphylococcus aureus, and Bacillus subtilis). The Gaussian ray approximation of anomalous diffraction theory was used to determine the average bacteria size from transmission measurements. A rescaled spectra combining multiple angular data was analyzed in the framework of the Rayleigh-Gans theory of light scattering. Particle shape and size distribution is then obtained from the rescale spectra. Particle characteristics (size and/or shape) retrieved from both methods are in good agreement with size and shape measured under scanning electron microscopy. These results demonstrate that light scattering may be able to detect and identify microbial contamination in the environment.

An in vitro resistance study of levofloxacin, ciprofloxacin, and ofloxacin using keratitis isolates of Staphylococcus aureus and Pseudomonas aeruginosa

PURPOSE We compared levofloxacin with ciprofloxacin and ofloxacin using the in vitro susceptibilities of Staphylococcus aureus (SA) and Pseudomonas aeruginosa (PA) keratitis isolates. DESIGN Retrospective, clinical laboratory study of antibiotic susceptibility among keratitis isolates. PARTICIPANTS Keratitis isolates from 200 patients with either SA or PA keratitis. METHODS Minimum inhibitory concentrations (MICs) were determined for levofloxacin, ofloxacin, and ciprofloxacin for 93 SA keratitis isolates (68 fluoroquinolone-resistant and 25 susceptible, as determined by disk diffusion) and 107 PA keratitis isolates (13 fluoroquinolone-resistant and 94 susceptible). National Committee for Clinical Laboratory Standards susceptibilities were determined and analyzed statistically. Time kill studies were determined for fluoroquinolone-susceptible and -resistant isolates to all antibiotics at 8 microg/ml. The killing rates were determined by regression, and the colony count decreases were analyzed. MAIN OUTCOME MEASURES The susceptibilities and potencies of levofloxacin, ciprofloxacin, and ofloxacin to SA and PA were determined from the MICs. Time kill studies determined the killing rates and decreases in colony counts. RESULTS The fluoroquinolone-resistant SA susceptibilities to levofloxacin, ofloxacin, and ciprofloxacin were only 22%, 10%, and 3%, respectively. The fluoroquinolone-susceptible SA were 100% susceptible to all antibiotics, with levofloxacin demonstrating the best potency. The fluoroquinolone-resistant PA were resistant to all antibiotics. The fluoroquinolone-susceptible PA isolates were highly susceptible to levofloxacin, ofloxacin, and ciprofloxacin, with ciprofloxacin demonstrating the highest potency. For fluoroquinolone-susceptible SA and PA, the time kill studies determined that the killing rates and decreases in colony counts were equivalent for all three antibiotics tested. The time kill studies demonstrated no colony count decreases for the fluoroquinolone-resistant SA and PA. CONCLUSIONS Taken together, our susceptibility and time kill data failed to demonstrate convincing differences in the susceptibility of SA and PA keratitis isolates to levofloxacin, ciprofloxacin, and ofloxacin. In general, bacterial isolates that were resistant to ciprofloxacin and ofloxacin were also resistant to levofloxacin.

Microbiological Spectrum and Antibiotic Sensitivity in Endophthalmitis: A 25-Year Review

PURPOSE To identify the spectrum and susceptibility pattern of pathogens responsible for culture-positive endophthalmitis referred to a single institution and investigate possible trends in both pathogens and antibiotic sensitivities over the past 25 years. DESIGN A retrospective, laboratory-based study of consecutive microbiological isolates. PARTICIPANTS A total of 988 consecutive culture-positive endophthalmitis isolates from 911 eyes. METHODS All culture-positive endophthalmitis isolates collected from 1987 to 2011 were identified. Susceptibility rates to a variety of antibiotics were calculated. Chi-square test for trend was used to detect changes in spectrum or susceptibility over time. MAIN OUTCOME MEASURES Microbial spectrum and susceptibility pattern over time. RESULTS A total of 988 isolates were identified from 911 eyes. The average patient age was 67 ± 18 years, and 55% of the patients were female. The most prevalent pathogens were coagulase-negative staphylococcus (39.4%), followed by Streptococcus viridans species (12.1%) and Staphylococcus aureus (11.1%). Gram-negative organisms and fungi accounted for 10.3% and 4.6% of all isolates, respectively. With the exception of 2 isolates, Enterococcus faecium and Nocardia exalbida, all the other 725 (99.7%) gram-positive bacteria tested were susceptible to vancomycin. Of the 94 gram-negative organisms tested against ceftazidime, 2 were of intermediate sensitivity and 6 were resistant. For 8 antibiotics, increasing microbial resistance over time was observed: cefazolin (P = 0.02), cefotetan (P = 0.006), cephalothin (P<0.0001), clindamycin (P = 0.04), erythromycin (P<0.0001), methicillin/oxacillin (P<0.0001), ampicillin (P = 0.01), and ceftriaxone (P = 0.006). For 3 antibiotics, increasing microbial susceptibility was observed: gentamicin (P<0.0001), tobramycin (P = 0.005), and imipenem (P<0.0001). CONCLUSIONS Coagulase-negative staphylococcus remains the most frequently identified cause of endophthalmitis. Vancomycin and ceftazidime seem to be excellent empiric antibiotics for treating endophthalmitis. Although a statistically significant trend toward increasing microbial resistance against a variety of antibiotics, including cephalosporins and methicillin, was observed, a significant trend toward decreasing microbial resistance against aminoglycosides and imipenem also was detected.

Molecular characterization, biofilm analysis and experimental biofouling study of Fusarium isolates from recent cases of fungal keratitis in New York State

BackgroundTo characterize Fusarium isolates from recent cases of fungal keratitis in contact lens wearers, and to investigate fungal association with MoistureLoc solution.MethodsWe studied six fungal isolates from recent cases of keratitis in New York State. The isolates were characterized by nucleotide sequencing and phylogenetic analyses of multiple genes, and then typed using minisatellite and microsatellite probes. Experimental fungal biofilm formation was tested by standard methods. MoistureLoc solutions were tested in biofouling studies for their efficacy in elimination of Fusarium contamination.ResultsFusarium solani – corneal ulcers (2 isolates), lens case (1 isolate), and F. oxysporum – corneal ulcer (1 isolate), eye (1 isolate), were recovered from five patients. An opened bottle of MoistureLoc solution provided by a patient also yielded F. solani. Two distinct genotypes of F. solani as well as of F. oxysporum were present in the isolated strains. Remarkably, F. solani strains from the lens case and lens solution in one instance were similar, based on phylogenetic analyses and molecular typing. The solution isolate of F. solani formed biofilm on contact lenses in control conditions, but not when co-incubated with MoistureLoc solution. Both freshly opened and 3-month old MoistureLoc solutions effectively killed F. solani and F. oxysporum, when fungal contamination was simulated under recommended lens treatment regimen (4-hr). However, simulation of inappropriate use (15 – 60 min) led to the recovery of less than 1% of original inoculum of F. solani or F. oxysporum.ConclusionTemporary survival of F. solani and F. oxysporum in MoistureLoc suggested that improper lens cleaning regimen could be a possible contributing factor in recent infections.

In vitro effectiveness of 13 agents in otomycosis and review of the literature

Many agents have been recommended for treating otomycosis, but no preparation has been widely accepted. To compare the effectiveness of many recommended preparations, we performed an in vitro study using 15 species of fungi and yeast cultured from patients presenting with otomycosis during the past year. By measuring zones of inhibition, we assessed the effectiveness of aqueous Merthiolate®, Burows solution (2%), VoSol® HC, VoSol® plain, Cortisporin® suspension, clotrimazole 1%, Mycostatin®, amphotericin B, ethanol 95%, miconazole, tolnaftate 1%, natamycin, and flucytosine.

Normal ocular flora in healthy eyes from a rural population in Sierra Leone

Purpose To identify the normal conjunctival flora in a rural population in Sierra Leone and compare the normal ocular flora of a developing country with that of the developed world. Methods Conjunctival swabs obtained from healthy right eyes of 276 residents of Masungbo, Sierra Leone, were analyzed for growth of microorganisms. Results The most commonly isolated organisms from conjunctival swabs of healthy eyes were coagulase-negative Staphylococcus (28.6%), fungus (26.0%), Staphylococcus aureus (19.9%), Gram negatives other than Pseudomonas/Haemophilus (9.8%), Nocardia/Actinomyces (6.5%), and Pseudomonas aeruginosa (6.2%). Conclusions There is a high prevalence of fungal and bacterial colonization of healthy eyes in a rural population of Sierra Leone. A relationship may exist between resident normal flora and the etiology of ocular infections.

Native fluorescence and excitation spectroscopic changes in Bacillus subtilis and Staphylococcus aureus bacteria subjected to conditions of starvation

Fluorescence emission and excitation spectra were measured over a 7-day period for Bacillus subtilis (Bs), a spore-forming, and Staphylococcus aureus (Sa), a nonspore-forming bacteria subjected to conditions of starvation. Initially, the Bs fluorescence was predominantly due to the amino acid tryptophan. Later, a fluorescence band with an emission peak at 410 nm and excitation peak at 345 m, from dipicolinic acid, appeared. Dipicolinic acid is produced during spore formation and serves as a spectral signature for detection of spores. The intensity of the 410-nm band continued to increase over the next 3 days. The Sa fluorescence was predominantly from tryptophan and did not change over time. In 6 of the 17 Bs specimens studied, an additional band appeared with a weak emission peak at 460 cm and excitation peaks at 250, 270, and 400 nm. The addition of beta-hydroxybutyric acid to the Bs or the Sa cultures resulted in a two-order of magnitude increase in the 460-nm emission. The addition of Fe2+ quenched the 460 emission, indicating that a source of the 460-nm emission was a siderophore produced by the bacteria. We demonstrate that optical spectroscopy-based instrumentation can detect bacterial spores in real time.

Recurrent bleb infections

AIM To report the patient characteristics, causative organisms, and clinical outcomes in patients with recurrent bleb related ocular infections. METHODS The medical records of all patients diagnosed with bleb related ocular infection at the New York Eye and Ear Infirmary over a 10 year period were reviewed. Recurrent bleb infection was defined as at least two episodes of bleb purulence with or without associated intraocular inflammation separated by a quiescent period of at least 3 months. RESULTS Recurrent bleb infections developed in 12 eyes of 12 patients (10 men, two women) a mean of 16.3 (SD 17.9) months (range 3–51 months) after the initial infection. Two patients developed a third episode 3 and 20 months, respectively, after the second infection, yielding a total of 14 recurrent infection episodes. Recurrent infection developed after trabeculectomy in 11 eyes (adjunctive 5-fluorouracil, nine eyes; mitomycin C, one eye; no antifibrosis agent, one eye) and following cataract extraction with inadvertent bleb formation in one eye. Four (36.4%) of the filtered eyes had undergone trabeculectomy at the inferior limbus. The mean follow up time from filtering surgery to the first bleb related infection was 28 months for the nine patients treated with 5-fluorouracil and 14 months for the single patient treated with mitomycin C. 11 (78.6%) cases had a documented bleb leak in the 4 week period before or at the time of recurrent infection. Topical, prophylactic antibiotics had been used in 7/14 (50%) cases. The same organism was cultured from the initial and recurrent infections in 2/14 (14.3%) cases. CONCLUSION Eyes that have been successfully treated for bleb related infection remain at risk for recurrent infection. No apparent correlation exists between organisms responsible for the initial and recurrent infections. The increased rate of recurrent bleb related infection in patients receiving adjunctive 5-fluorouracil compared to mitomycin C may have been related to the longer follow up of the 5-fluorouracil eyes.

COLLETOTRICHUM GRAMINICOLA : A NEW CORNEAL PATHOGEN

Purpose We report the first case of an ocular infection with the fungus Colletotrichum graminicola causing keratitis in a 27-year-old man. Methods Twenty-four months after a postoperative course complicated by recurrent fungal keratitis requiring two penetrating keratoplasties. two anterior chamber washouts, a conjunctival flap, and medical treatment with topical natamyein, intracameral amphotericin B, and oral fluconazole. Results The patient has shown no signs of fungal recurrence despite a failed corneal graft. Conclusions C. graminicola is a new corneal pathogen and should be included in the differential diagnosis of mycotic keratitis.