Methicillin resistance and biofilm production of Staphylococcus epidermidis isolates from infectious and normal flora conjunctiva.

PURPOSE: Coagulase-negative staphylococci have been reported to be the most frequent cause of bacterial postoperative endophthalmitis. Biofilm formation is the major virulence factor of Staphylococcus epidermidis and is often associated with methicillin resistance. This study aims at evaluating the presence of biofilm-producing and methicillin resistance genes (mecA) in S. epidermidis. METHODS: S. epidermidis isolated from clinically infected sites (group 1) and from normal human conjunctiva (group 2) were studied. All the isolates were tested for their ability to produce biofilm by the conventional Christensen´s method and the presence of mecA by PCR using the 22-mer oligonucleotides as primers. RESULTS: In total 20 isolates from group 1 and 22 from group 2 were studied. Biofilm and mecA were detected in 15 (75 %) and in 14 (70 %) in group 1 as compared to 8 (36.3 %) and 4 (18.2 %) in group 2 (p = 0.016). Simultaneously, biofilm production and presence of mecA genes were observed in 13/20 (65.0 %) in group 1, and 4/22 (18.2 %) in group 2 (p = 0.002). Multi-resistance was observed in 55 % in group 1 and 9 % in group 2 (p = 0.002); 57 % of the biofilm-producing strains was multi-resistant in contrast to none of the non-producing strains. In all multi-resistant strains, biofilm production was seen. CONCLUSIONS: Biofilm formation capacity was widely distributed, particularly among mecA (+) S. epidermidis strains, which also displayed a high diversity of antibiotic resistance profiles.

Effect of antibiotic prophylaxis on Coagulase-negative Staphylococcus virulence factor profiles in patients undergoing cataract surgery.

PURPOSE: In this prospective study, multiplex polymerase chain reaction (PCR) was used to identify genes encoding virulence factors (ica, atlE and mecA) in Coagulase-negative Staphylococcus (CNS) isolates from the ocular microbiota of patients undergoing cataract surgery and to investigate possible changes in the CNS profile due to antibiotic prophylaxis. METHODS: Between 09/2011 and 08/2013, patients undergoing cataract surgery were recruited at the Department of Ophthalmology, National University of Asuncion, Paraguay. In the eye to be operated on, patients received moxifloxacin 0.5 % eye drops four times at the day before surgery and a last drop 1 hour before surgery (T1). The other eye remained as control (T0). Conjunctival swabs were taken from both eyes 1 hour after the last drop. The presence of genes encoding biofilm formation (ica and atlE) and methicillin resistance (mecA) was detected by a multiplex PCR. RESULTS: Of the 162 patients (162 study eyes, 162 fellow eye as control group), 87 (53.7 %) eyes were positive for CNS at T0 yielding 96 CNS isolates; 70 eyes (43.2 %) were positive at T1 yielding 77 CNS isolates. For this study, 43 CNS isolates (44.8 %) from T0 and 45 (64.3 %) from T1 were used. Of the total isolates, 81.8 % (72/88) had at least one virulence factor gene (37/43 from T0 and 35/45 from T1) (p = 0.314). Simultaneous detection of ica and atlE genes was higher in T0 (58.0 %) than T1 (46.7 %), but the difference was not significant (p = 0.28). CONCLUSION: A high frequency of genes encoding virulence factors was observed in the coagulase-negative Staphylococcus isolates. The use of moxifloxacin did not significantly modify the CNS virulence factor profiles.

Comparing Different Decontamination Procedures in Harvesting Human Donor Cornea.

PURPOSE: To evaluate the effects of current hygiene standards for the enucleation of postmortem eyes by investigating the number of microorganisms during subsequent steps of decontamination and tissue processing. MATERIALS AND METHODS: This prospective, non-randomized cohort study includes 184 postmortem eyes of 92 human donors. Enucleation was performed, according to an ophthalmic surgical procedure. Two groups were generated as follows: right eyes were allocated to group A, left eyes were allocated to group B. In group A, a mucosal disinfectant was used accessorily. Conjunctival smears were taken to examine germ load in both groups before any intervention, in group A after mucosal disinfection, in both groups after transportation of the whole globes in transport fluid, and in both groups after a bath in 0.75% povidone iodine solution for at least 3 minutes just before preparation of the corneoscleral disc. Smears were sent to the local microbiologic laboratory in an aseptic package for testing. RESULTS: All smears showed multiple contaminations (n = 184/184 eyes) before treatment with povidone iodine solution. Contamination was in both groups significantly prevented using the treatment strategy of an at least 3-minute bath in 0.75% povidone iodine solution (n = 1/184 eyes; p < 0.01) As a side effect, almost every eye of group A and none of group B showed brown iodine staining corresponding to corneal epithelial erosion. CONCLUSIONS: An aseptic setting for donor enucleation similar to a surgical procedure seems not to influence the outcome of germ colonization. The most effective step to decontaminate donor eyes is to use 0.75% povidone iodine solution for at least 3 minutes.

Clinical and epidemiological characteristics of infectious keratitis in Paraguay.

To describe the clinical and epidemiological characteristics of patients with severe infectious keratitis in Asunción, Paraguay between April 2009 and September 2011. All patients with the clinical diagnosis of severe keratitis (ulcer ≥2 mm in size and/or central location) were included. Empiric treatment consisted of topical antibiotics and antimycotics; in cases of advanced keratitis, fortified antibiotics were used. After microbiological analysis, treatment was changed if indicated. In total 48 patients (62.5 % males, 25 % farmers) were included in the analysis. A central ulcer was found in 81.3 % (n = 39). The median delay between onset of symptoms and time of first presentation at our institution was 7 days (range 1-30 days). Fungal keratitis was diagnosed in 64.5 % (n = 31) of patients, of which Fusarium sp. (n = 17) was the most common. Twenty-one patients (43.8 %) reported previous trauma to the eye. The globe could be preserved in all cases. While topical therapy only was sufficient in most patients, a conjunctival flap was necessary in six patients suffering from fungal keratitis. The high rate of fungal keratitis in this series is remarkable, and microbiological analysis provided valuable information for the appropriate treatment. In this setting, one has to be highly suspicious of fungal causes of infectious keratitis.

Intravitreal voriconazole: in vitro safety-profile for fungal endophthalmitis.

INTRODUCTION: Fungal endophthalmitis is a rare but sight-threatening disease. Despite an expanding range of fungal pathogens, there are only few therapeutic agents for its treatment available. Voriconazole is a second-generation synthetic triazole with a broad action against yeasts and molds. The current study investigates the safety of Voriconazole for intravitreal application in a cell culture model. METHODS: Primary human retinal pigment epithelium cells (RPE) and primary human optic nerve head astrocytes were treated with concentrations of Voriconazole ranging from 25 microg/mL to 10 mg/mL. Possible toxic effects and IC50 were evaluated after 24 hours and under conditions of oxidative stress. By treating the RPE cell lines with tumor-necrosisfactor alpha (TNF-alpha), lipopolysaccharides (LPS), and interleukin-6 (IL-6) the effects of Voriconazole on cellular viability under conditions of inflammation were investigated. Toxicity was evaluated by colorimetric measuring the inhibition of RPE cell proliferation (MTT). Additionally cell viability was quantified by a microscopic live-dead-assay. RESULTS: Concentrations <250 microg/mL Voriconazole had no influence neither on RPE nor on optic nerve head astrocytes cell proliferation and cell viability when administered for 24 hours and under oxidative stress. When preincubated with tumor-necrosis-factor alpha, lipopolysaccharides and interleukin-6 for 24 hours and subsequently treated with Voriconazole at concentrations up to 250 microg/mL for 24 hours no significant decrease in proliferation and viability was observed. CONCLUSIONS: This study showed that no significant toxicity existed for Voriconazole in vitro on primary RPE and optic nerve head astrocytes when administered in therapeutic concentrations up to 250 microg/mL.

Bacterial contamination of ocular surface and needles in patients undergoing intravitreal injections.

PURPOSE: To evaluate potential sources of bacterial contamination during intravitreal (IVT) injection procedures. METHODS: Patients scheduled for IVT injection were asked to enroll in the study at the California Vitreoretinal Center (Menlo Park, CA) and the Vantage Eye Center (Salinas, CA) between October 2004 and April 2005. A total of 104 patients participated in the study, with a total of 118 IVT injection procedures performed on 107 eyes. Standard microbiological techniques were used to culture, identify, and quantify bacterial contamination of injection needles and the bulbar conjunctiva at the injection site in patients undergoing IVT injections. The main outcomes measured were type and quantity of bacterial isolates. RESULTS: Two (2%) of 114 needles collected were contaminated with bacteria. The prevalence of bacterial contamination of the injection site on the bulbar conjunctiva was 43% before prophylaxis on the day of the injection with topical antibiotics and povidone-iodine, with a statistically significant reduction to 13% after prophylaxis (P < 0.0001). Coagulase-negative Staphylococcus, the most common bacterium isolated from the ocular surface, was isolated from both culture-positive needles. CONCLUSIONS: IVT injection needles became contaminated with bacteria during the injection procedure. Although the contamination rate was low, this supports a mechanism of postinjection endophthalmitis in which there is direct inoculation of ocular surface flora into the vitreous cavity by the injection needle.

Effects of intravitreal corticosteroid in the treatment of Staphylococcus aureus-induced experimental endophthalmitis.

PURPOSE: To investigate whether intravitreal injection of dexamethasone in addition to antibiotics can minimize intraocular tissue injury caused by Staphylococcus aureus endophthalmitis. METHODS: Albino rabbits were infected with an intravitreal injection of 1000 colony-forming units of S. aureus. The rabbits were randomized to receive no treatment (control group; n = 2), intravitreal vancomycin and amikacin (n = 5), or a combination of intravitreal vancomycin, amikacin, and dexamethasone (n = 5) 20 hours following inoculation of bacteria. All rabbits except for the control group also received intravenous imipenem every 8 hours for 4 days. The eyes were evaluated by clinical examination, electroretinogram (ERG), and histologic studies. RESULTS: Eyes treated with intravitreal dexamethasone demonstrated less inflammation on clinical examination compared with eyes that received antibiotics alone. The ERG responses of eyes that received both intravitreal antibiotics and steroid were significantly better at 45 hours, 7 and 14 days following inoculation (P < 0.05) compared to eyes that received antibiotics alone. Histologic studies 14 days following infection demonstrated less tissue destruction for eyes treated with dexamethasone. CONCLUSION: Compared to intravitreal antibiotics alone, intravitreal corticosteroids may improve visual outcome of S. aureus endophthalmitis by reducing inflammation and preserving electrophysiologic retinal function.

Clinical results of topical fluconazole for the treatment of filamentous fungal keratitis.

BACKGROUND: Fungal keratitis is a rare but serious condition that may result in loss of vision. The potentially poor prognosis might be due to a delay in diagnosis and/or to limited treatment options. The aim of this study is to evaluate the clinical outcome of patients treated with topical fluconazole 0.2% for the treatment of filamentous fungal keratitis. METHODS: Retrospective case series. A chart review of all patients evaluated at the Ophthalmology Department of the Hospital de Clínicas in Paraguay from January 1997 to December 2000 identified 25 cases of fungal keratitis. Among these cases, one patient discontinued the treatment and another received amphotericin as the first line drug, resulting in 23 cases available for data analysis. Twelve patients were treated with topical fluconazole 0.2% alone (Group I) and 11 patients received a combination of topical fluconazole 0.2% and oral ketoconazole 200 mg twice daily (Group II). RESULTS: Sixteen of 23 cases showed resolution of the keratitis, 9/12 (75%) in Group I and 7/11 (64%) in Group II. Seven patients (30%) did not respond to medical treatment and required a surgical procedure to preserve eye integrity. Superficial and small ulcers have a significantly better prognosis than do lrge and deep ulcers. Concomitant oral ketoconazole (400 mg/day) did not improve the clinical results. CONCLUSIONS: Our findings of study suggest that topical fluconazole 0.2% is a safe and effective antifungal drug for the management of filamentous mycotic keratitis, particularly in cases that are not severe. The addition of oral ketoconazole to topical fluconazole did not improve the clinical outcome.

Comparison of preoperative conjunctival bacterial flora in patients undergoing glaucoma or cataract surgery.

PURPOSE: To assess differences in conjunctival bacterial flora between patients undergoing glaucoma and cataract surgery. PATIENTS AND METHODS: A prospective study comparing conjunctival bacterial cultures obtained from 339 patients undergoing either cataract (n = 258) or glaucoma (n = 81) surgery. All cultures were acquired during the preoperative visit, approximately three to seven days prior to surgery. The culture samples were inoculated onto blood and chocolate agar, as well as blood culture broth media. All bacterial isolates were identified and statistical analyses were performed to determine if there were differences in flora between the eyes undergoing cataract versus glaucoma surgery. RESULTS: Two hundred fifteen of 258 eyes (83%) undergoing cataract surgery were found to have positive bacterial growth, compared with 62 of 81 eyes (77%) of those undergoing glaucoma surgery (P = 0.2246). Coagulase-negative Staphylococci, the most common bacterial isolate, was cultured from 167 eyes (65%) in the cataract group and 42 (52%) in the glaucoma group (P = 0.0514). Among all bacterial isolates, only Corynebacterium species was found to be statistically different between the two patient groups with 92 (36%) and 11 (14%) eyes testing positive in the cataract and glaucoma groups, respectively (P = 0.0003). CONCLUSIONS: There was no statistically significant difference in the proportion of conjunctival culture samples testing positive for bacterial growth in eyes undergoing glaucoma surgery compared with those undergoing cataract surgery. Glaucoma medications, or their preservatives, do not appear to significantly alter conjunctival flora. Techniques used for endophthalmitis prophylaxis prior to cataract surgery are likely appropriate for glaucoma surgery as well.

Three-day application of topical ofloxacin reduces the contamination rate of microsurgical knives in cataract surgery: a prospective randomized study.

PURPOSE: To determine the rate of contamination of microsurgical knives during cataract surgery and the benefit of a 3-day versus a 1-hour preoperative application of topical ofloxacin in reducing the contamination rate. DESIGN: Prospective, randomized controlled trial. PARTICIPANTS: Seventy-eight eyes of 75 patients were randomly assigned to control (39 eyes) or study groups (39 eyes). METHODS: All patients from both groups received 0.3% topical ofloxacin 1 hour before surgery, 5% povidone-iodine (PVI) scrub of the periorbital area, and 2 drops of PVI onto the ocular surface preoperatively. The patients in the study group also received ofloxacin 4 times a day for 3 days before surgery. MAIN OUTCOME MEASURES: Microsurgical knives were placed in blood culture broth media immediately after the incision had been made. The number of positive cultures and types of bacteria isolated were determined. RESULTS: Ten of 39 knives (26%) in the control group were found to be positive for bacterial growth compared with only 2 of 39 (5%) in the study group (P = 0.028). CONCLUSIONS: The initial paracentesis incision frequently results in contamination of the microsurgical knife and may serve as a mechanism for introducing bacteria from the ocular surface into the anterior chamber. The application of topical ofloxacin for 3 days before surgery significantly reduces the contamination rate of the microsurgical knives, compared with a preoperative application of ofloxacin given 1 hour before surgery.

The challenge of determining aqueous contamination rate in anterior segment intraocular surgery.

PURPOSE: To determine aqueous contamination rate in anterior segment intraocular surgery using two different techniques of obtaining aqueous fluid and to assess whether a 3-day application of topical 0.3% ofloxacin reduces this contamination rate compared with a 1-hour application. DESIGN: Randomized clinical trial. METHODS: One hundred and thirty-three eyes of 130 patients undergoing anterior segment intraocular surgery were randomized to either control (64 eyes received topical ofloxacin 1 hour before surgery) or study groups (69 eyes received topical ofloxacin four times a day for 3 days before surgery in addition to 1 hour preoperatively). Eyes in both groups received a periorbital iodine scrub and two drops of topical 5% iodine. Aqueous fluid was obtained at the beginning and conclusion of surgery using a cannula passed through a paracentesis or a needle passed through clear cornea. The aqueous, cannula, and needles were inoculated in blood culture media broth and bacterial growth was identified. RESULTS: Overall, eight of 89 aqueous samples (9%) obtained using a cannula at the beginning of surgery were culture-positive. Similarly, six of 41 aqueous samples (15%) obtained through a needle through clear cornea at the beginning of surgery showed contamination. At the conclusion of surgery, nine of 112 samples (8%) showed positive cultures. There was no difference in the aqueous contamination rates between the control and study groups. CONCLUSIONS: Despite the use of a needle to obtain aqueous fluid at the beginning of surgery before creating a paracentesis, the aqueous contamination rate remained higher than that found at the conclusion of surgery. A 3-day application of topical ofloxacin before surgery did not reduce the anterior chamber aqueous contamination rate relative to a 1-hour application.

Ocular alterations in patients with cutaneous and mucocutaneous leishmaniasis.

Leishmaniasis is a parasitosis with high prevalence and increasing epidemiologic relevance. Ocular manifestations of visceral leishmaniasis (kala-azar) have been well studied, but less is known about ocular alterations in cutaneous and mucocutaneous leishmaniasis (CL/MCL). Fifty-five patients with CL/MCL and a seronegative control group of 39 were examined ophthalmologically in Paraguay. CL/MCL was diagnosed clinically by detection of the parasite, with serological methods and/or intradermal reaction.