Ana Luisa Hofling-Lima

Shifting trends in in vitro antibiotic susceptibilities for common ocular isolates during a period of 15 years.

PURPOSE To assess the in vitro susceptibility of the most common ocular bacterial isolates to several antibiotics and verify changing trends in the antibiotic susceptibility in a 15-year period. DESIGN Experimental study. METHODS All cultures positive for Staphylococcus aureus, coagulase-negative Staphylococcus (CNS), Streptococcus sp, and Pseudomonas sp in conjunctival (n = 4,585) and corneal (n = 3,779) samples from patients seen at the Federal University of São Paulo from 1985 to 2000 were evaluated. Cultures were performed in liquid and solid media, and susceptibility tests were done to amikacin, gentamicin, neomycin, tobramycin, ciprofloxacin, norfloxacin, ofloxacin, cephalothin, and chloramphenicol. RESULTS Amikacin and neomycin showed an improvement of their sensitivity during the study period (88%-95% and 50%-85%, respectively) for corneal and conjunctival samples. Gentamicin and tobramycin revealed a decrease of sensitivity in time, from 95% to less than 80% in corneal and conjunctival samples. Ciprofloxacin, norfloxacin, and ofloxacin had good sensitivity to all evaluated bacteria, better in conjunctiva (95%) than in cornea (90%). Sensitivity of S. aureus to cephalothin decreased during the study but was still 98% for CNS. Chloramphenicol had good sensitivity to S. aureus (85% in corneal and 92%in conjunctival samples), CNS (87% and 88.5%), and Streptococcus sp (95% and 96%). CONCLUSIONS Gentamicin, tobramycin, and cephalothin decreased their in vitro susceptibility to all tested pathogens. The fluoroquinolones remained a good choice in the treatment of ocular infections, with high suscep-tibility to all pathogens tested. Chloramphenicol also revealed an increase in its susceptibility to all bacteria evaluated.

The Use of Vital Dyes in Ocular Surgery

Vital dyes have advanced diagnosis and surgical technique in various specialties, including oncology, gastroenterology, and ophthalmology. In ocular surgery vital dyes are widely used in cataract and vitreoretinal surgery. Worldwide, intra-operative use of trypan blue during cataract surgery has enhanced visualization of the anterior capsule during capsulorrhexis, and patent blue has been recently licensed in Europe for cataract surgery. For chromovitrectomy, the vital dyes indocyanine green, infracyanine green, and brilliant blue stain the internal limiting membrane, and trypan blue and triamcinolone acetonide help visualize epiretinal membranes and vitreous, respectively. Intra-operative vital dyes are finding uses in corneal, glaucoma, orbit, strabismus, and conjunctival surgery. We provide a summary of current knowledge of the use of vital dyes in ocular surgery. We review the properties of dyes, techniques of application, indications, and complications in ocular surgery. Vital dyes represent an expanding area of research, and novel dyes deserve further investigation.

Corneal infections after implantation of intracorneal ring segments.

Purpose: To report risk factors, clinical course, and outcome in patients with infectious keratitis following implantation of intracorneal ring segments (ICRS). Methods: The records of 8 patients with culture-proven infectious keratitis after ICRS (Ferrara® or Intacs®) implantation were retrospectively reviewed. Age, gender, corneal findings, ocular abnormalities, the condition that led to ICRS implantation, immediate prior use of a contact lens, elapsed time between implantation and the onset of symptoms, previous medications, and systemic disorders were noted. Results: Culture-positive infectious keratitis developed in 7 eyes of 7 patients (2 men and 5 women) with a mean age of 35 years who underwent Ferrara implantantion for the treatment of keratoconus and in a 29-year-old man who underwent Intacs implantation for correction of low myopia. Contact lens use, diabetes, and trauma were factors possibly associated with the risk of infection in three cases. Microorganisms, identified in all cases, included Staphylococcus aureus, Streptococcus viridans, Streptococcus pneumoniae, Pseudomonas sp, Nocardia sp, Klebsiella sp, and Paecylomices sp. Onset of symptoms of infection varied from less than 1 week to 22 months postoperatively, depending on the infecting organism. Conclusions: Infectious keratitis following ICRS implantation is a sight-threatening complication for which early recognition and rapid institution of appropriate treatment may result in a better visual outcome.

Genotyping of 44 Isolates of Fusarium solani, the Main Agent of Fungal Keratitis in Brazil

ABSTRACT In a retrospective study performed over 6 years in Brazil, Fusarium solani was found to be the most common species causing mycotic keratitis. The genetic diversity of 44 isolates from 39 patients was assessed by enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) and PCR restriction fragment length polymorphism (PCR-RFLP) fingerprinting. ERIC-PCR was more discriminatory than PCR-RFLP for differentiating the strains. By combining of the results of both techniques, we identified 40 genotypes. Molecular typing revealed a high genomic heterogeneity of the strains of F. solani studied.

Infectious post-LASIK crystalline keratopathy caused by nontuberculous mycobacteria.

Purpose. To report three cases of infectious crystalline keratopathy caused by non-tuberculous mycobacteria after LASIK surgery. Methods. Interventional case reports and literature review. Results. Infectious keratitis with clinical features of crystalline keratopathy after LASIK is described. Culture revealed Mycobacterium chelonae from the corneal scrapings of the three patients, all of whom underwent medical and surgical (debridement) treatment. Conclusions. Mycobacteria may cause infectious crystalline keratopathy after LASIK. The presence of crystalline keratopathy in patients that underwent LASIK must be considered an indicator of nontuberculous mycobacteria infection. Microbiologic work-up of a corneal specimen is required for the institution of appropriate therapy.

Acute endophthalmitis following intravitreal bevacizumab (Avastin) injection.

Purpose:To report two cases of acute endophthalmitis following intravitreal bevacizumab injection.Methods:Two patients with exudative age-related macular degeneration were treated sequentially with an intravitreal injection of bevacizumab and developed signs of severe but painless infectious endophthalmitis 2 days later. Vitreous samples were obtained, followed by the injection of vancomycin 1 mg/0.1 ml and ceftazidime 2.25 mg/0.1 ml. Pulsed-field gel electrophoresis (PFGE) was used to determine whether the isolated microorganisms were the same.Results:Coagulase-negative staphylococci were identified and isolated from the vitreous specimen of both patients. PFGE revealed different patterns of banding, excluding that interpatient contamination occured.Conclusions:Infectious endophthalmitis is a potential complication of intravitreal bevacizumab injection.

Detection and Gram Discrimination of Bacterial Pathogens from Aqueous and Vitreous Humor Using Real-Time PCR Assays

PURPOSE To develop and apply real-time PCR protocols to the detection and classification of the Gram status of bacterial pathogens in aqueous and vitreous humor collected from clinically suspected intraocular infections. METHODS The analytical specificity of two PCR assays, SYBR Green 16S rDNA-Based Universal PCR (SGRU-PCR), and a Multiplex Gram-Specific TaqMan-Based PCR (MGST-PCR), was determined with 31 clinically important pathogens, including 20 Gram-positive and 11 Gram-negative. Analytical sensitivity was determined with a 10-fold dilution of Staphylococcus epidermidis and Escherichia coli DNA. Assays were further tested on aqueous (n = 10) and vitreous humor (n = 11) samples collected from patients with clinically diagnosed intraocular infections. RESULTS DNA was amplified from all control bacterial isolates when using SGRU-PCR. MGST-PCR correctly classified the Gram status of all these isolates. The SGRU-PCR limit of detection of S. epidermidis and E. coli DNA was 100 fg/μL (E = 0.82 and 0.86; r(2) = 0.99) and for MGST-PCR, 1 pg/μL (E = 0.66 and 0.70; r(2) = 0.99. For clinical intraocular samples, positivity of culture was 47.6% and for real-time PCR assays, 95.2%. Gram classification was achieved in 100% of MGST-PCR-positive samples. Among microbiologically negative samples, real-time PCR assays were positive in 90% of cases. The false-positive rate in control aqueous was 3.2%, and control samples of vitreous were negative. CONCLUSIONS The real-time PCR assays demonstrated good correlation, with culture-proven RESULTS With the use of these methods, bacterial detection was improved from 47.6% to 95.3%, demonstrating them to be sensitive, rapid tests for diagnosis of bacterial endophthalmitis.

Comparative analysis of the nuclear lens opalescence by the Lens Opacities Classification System III with nuclear density values provided by Oculus Pentacam: a cross-section study using Pentacam Nucleus Staging software

PURPOSE To compare the clinical classification of cataract using the Lens Opacities Classification System (LOCS) III with the mean values of lens density provided by the Pentacam Scheimpflug System in nuclear cataracts. METHODS One hundred and one eyes from 101 patients with age-related nuclear cataract were submitted to clinical examination for lens grading score using LOCS III. According to LOCS III, nuclear opalescence was divided in six groups. Patients were evaluated by the Pentacam Scheimpflug System for the mean lens density using the Pentacam lens densitometry program (PLDP), the Pentacam Nucleus Staging (PNS) mean value and the PNS cataract grading score. RESULTS A positive correlation between the mean values of lens density and LOCS III classification, considering groups 1 to 5, could be noticed with PLDP and PNS mean value. The mean values between the groups were similar using the PLDP and the PNS mean value. However, when the PNS cataract grading score was evaluated, there was low correspondence with LOCS III classification. CONCLUSION Pentacam Scheimpflug device offers an objective measure of the lens nuclear density on nuclear cataracts. PLDP and the PNS mean value were both useful to evaluate age-related nuclear cataract up to LOCS III group 5.

Microbial profile and antibiotic susceptibility of culture-positive bacterial endophthalmitis

PurposeTo assess the distribution of microorganisms isolated from patients with bacterial endophthalmitis and their antimicrobial susceptibility.MethodsRetrospective analysis of medical and microbiological records of patients with suspected diagnosis of endophthalmitis. The following information was assessed: number of presumed and culture-positive endophthalmitis cases, source of infection, microbiological result (aqueous and/or vitreous culture and Gram staining), microbial characterization and distribution, and antimicrobial susceptibility.ResultsA total of 107 (46%) of 231 patients with bacterial endophthalmitis showed positive results by gram stain or culture. Of these, 97 (42%) patients were positive for culture only. Most of them (62%) were secondary to a surgical procedure (postoperative), 12% were posttraumatic and 26% were secondary to an unknown source or the data were unavailable. A total of 100 microorganisms were isolated (38 aqueous and 67 vitreous samples) from the 97 culture-positive cases (91% were gram-positive and 9% were gram-negative). Coagulase-negative Staphylococcus(CoNS) (48%) were the most frequently isolated, followed by Stretococcus viridans(18%), and Staphylococcus aureus(13%). The antimicrobial susceptibility for CoNS was as follows: amikacin—91.6%, cephalothin—97.9%, ceftriaxone—50%, ciprofloxacin—62.5%, chloramphenicol—91.8%, gatifloxacin—79.5%, gentamicin—72.9%, moxifloxacin—89.5%, ofloxacin—70.8%, oxacillin—58.3%, penicillin—33.3%, tobramycin—85.4%, and vancomycin—100%.ConclusionGram-positive bacteria were the major causes of infectious endophthalmitis in this large series, usually following surgery. CoNS was the most common isolate. Of interest, susceptibility to oxacillin and fourth-generation quinolones was lower than previously published.

An Outbreak of Keratitis Caused by Mycobacterium immunogenum

ABSTRACT From 8 October to 12 November 2003, 36 patients underwent surgical correction of myopia in a São Paulo, Brazil, clinic. Five patients had clinical signs of infectious keratitis, and a Mycobacterium species with previously unreported patterns determined by PCR restriction enzyme analysis of the hsp65 gene and PCR restriction enzyme analysis of the 16S-23S rRNA internal transcribed spacer (ITS) was isolated from corneal scrapings from four of these patients. Subsequent evaluation by phenotypic tests and partial sequencing of the hsp65, sodA, rpoB, and 16S rRNA genes and the ITS supported the species identification as a variant of Mycobacterium immunogenum. The source of infection was not determined. The outbreak was caused by a single clone, as evidenced by identical pulsed-field gel electrophoresis and enterobacterial repetitive intergenic consensus-PCR profiles. This is the first report of an outbreak where this species was isolated from infected tissues.