Incidence of, risk factors for, and combined mechanism of late-onset open-angle glaucoma after vitrectomy.

PURPOSE: To estimate the incidence of and identify the risk factors for late-onset open-angle glaucoma (OAG) after uncomplicated pars plana vitrectomy (PPV). METHODS: All patients who underwent PPV at the Edward Harkness Eye Institute between January 1998 and January 2004 had at least 6 months of follow-up and did not have preexisting glaucoma or reason for secondary development of glaucoma were included. Retrospective cohort and matched case-control study designs were used. RESULTS: Of 285 vitrectomized eyes that met enrollment criteria, 11.6% (n = 33) developed glaucoma after vitrectomy. In the matched case-control analysis, the only variable that had a statistically significant association with the development of OAG was cataract extraction (CE), as compared with phakic status at the last follow-up (odds ratio = 16.4; 95% confidence interval, 2.1-127.4; P = 0.007). There was no difference in OAG development between eyes that had CE before or at the time of PPV and those that had it after PPV. The overall incidence of OAG development after PPV among all eyes, phakic eyes, and nonphakic eyes was 11.6%, 1.4%, and 15.0%, respectively. The difference in incidence between phakic and nonphakic eyes was statistically significant (P = 0.001). CONCLUSION: Lens extraction is a strong risk factor for the development of late-onset OAG after uncomplicated PPV. While the overall incidence of OAG development after PPV is substantial, it is more so among eyes that have had CE. The absence of substantial OAG incidence in phakic patients points toward a combined mechanism for late-onset post-PPV OAG involving PPV and CE at any time. Preoperative PPV counseling should include the risk of glaucoma development in addition to cataract development and the connection between the two. Patients who have undergone PPV, and especially those who also had CE in the same eye, should be carefully monitored for glaucoma.

Anti-retinal pigment epithelium antibodies in acute exudative polymorphous vitelliform maculopathy: a new hypothesis about disease pathogenesis.

OBJECTIVE: To investigate the etiology of acute exudative polymorphous vitelliform maculopathy (AEPVM) in a patient with metastatic melanoma, undiagnosed at initial examination, by testing for autoimmune mechanisms. METHODS: Serum samples were obtained from a 50-year-old man with AEPVM and metastatic unknown primary melanoma during the acute stage and 3 years later when subretinal fluid had resolved and melanoma was in remission (AEPVM convalescent stage). Western immunoblots using both serum samples against human donor retinal extract and cultured primary human retinal pigment epithelium (RPE) cell extract were performed to identify antiretinal and anti-RPE antibodies. Serum samples from 5 unaffected participants were tested as controls. Protein identification was performed using 2-dimensional gel electrophoresis and mass spectrometry and was then confirmed by blotting against purified protein. RESULTS: Western immunoblots of the patient's serum against human donor retinal extract and RPE cell extract demonstrated several antiretinal antibodies, as well as anti-RPE antibodies against a 26-kDa protein that was identified as peroxiredoxin 3 (PRDX3). Serum reactivity against PRDX3 was greatly decreased in the convalescent-stage serum sample compared with the acute-stage serum sample, while results of retinal extract Western immunoblots remained essentially unchanged. Five separate serum samples from participants without AEPVM had no autoantibodies against PRDX3. CONCLUSIONS: Paraneoplastic autoimmune reaction against RPE, with PRDX3 as the putative antigen, may be a cause of AEPVM. This is the first report to date linking a human RPE disease with anti-RPE antibodies against a heretofore undetermined putative protein. Testing for RPE autoantibodies may be useful in exploring the pathogenesis of other presumed RPE-related diseases.

Use of a multiplex molecular beacon platform for rapid detection of methicillin and vancomycin resistance in Staphylococcus aureus.

Drug resistance, particularly vancomycin and methicillin resistance, in Staphylococcus aureus continues to emerge as a significant public health threat in both the hospital and community settings. In addition to the limited treatment options, S. aureus strains acquire and express numerous virulence factors that continue to increase its ability to cause a wide spectrum of human disease. As a result, empirical treatment decisions are confounded and there is a heightened need for a diagnostic test (or assay) to rapidly identify antibiotic resistance and specific virulence determinants and indicate the appropriate treatment. To that end we developed a platform using multiplex molecular beacon probes with real-time PCR for the rapid detection of drug resistance-determining genes and virulence factors in S. aureus. In this study, we demonstrate the specificity and sensitivity of our platform for detection of the genes conferring methicillin (mecA) and vancomycin (vanA) resistance as well as a gene encoding the virulence factor Panton-Valentine leucocidin (lukF) in S. aureus isolates.

Comparative sequencing of the serine-aspartate repeat-encoding region of the clumping factor B gene (clfB) for resolution within clonal groups of Staphylococcus aureus.

Molecular techniques such as spa typing and multilocus sequence typing use DNA sequence data for differentiating Staphylococcus aureus isolates. Although spa typing is capable of detecting both genetic micro- and macrovariation, it has less discriminatory power than the more labor-intensive pulsed-field gel electrophoresis (PFGE) and costly genomic DNA microarray analyses. This limitation hinders strain interrogation for newly emerging clones and outbreak investigations in hospital or community settings where robust clones are endemic. To overcome this constraint, we developed a typing system using DNA sequence analysis of the serine-aspartate (SD) repeat-encoding region within the gene encoding the keratin- and fibrinogen-binding clumping factor B (clfB typing) and tested whether it is capable of discriminating within clonal groups. We analyzed 116 S. aureus strains, and the repeat region was present in all isolates, varying in sequence and in length from 420 to 804 bp. In a sample of 36 well-characterized genetically diverse isolates, clfB typing subdivided identical spa and PFGE clusters which had been discriminated by whole-genome DNA microarray mapping. The combination of spa typing and clfB typing resulted in a discriminatory power (99.5%) substantially higher than that of spa typing alone and closely approached that of the whole-genome microarray (100.0%). clfB typing also successfully resolved genetic differences among isolates differentiated by PFGE that had been collected over short periods of time from single hospitals and that belonged to the most prevalent S. aureus clone in the United States. clfB typing demonstrated in vivo, in vitro, and interpatient transmission stability yet revealed that this locus may be recombinogenic in a primarily clonal population structure. Taken together, these data show that the SD repeat-encoding region of clfB is a highly stable marker of microvariation, that in conjunction with spa typing it may serve as a DNA sequence-based alternative to PFGE for investigating genetically similar strains, and that it is useful for analyzing collections of isolates in both long-term population-based and local epidemiologic studies.

Multilocus sequence typing is a reliable alternative method to DNA fingerprinting for discriminating among strains of Candida albicans.

Multilocus sequence typing (MLST) has emerged as a powerful new DNA-typing tool for the evaluation of intraspecies genetic relatedness. This method relies on DNA sequence analysis of nucleotide polymorphisms in housekeeping genes and has shown a high degree of intraspecies discriminatory power for bacterial and fungal pathogens. However, the results of the MLST scheme for Candida albicans have heretofore never been formally compared to those of other established typing techniques. To assess the value of MLST relative to those of other DNA fingerprinting tools for discriminating among strains of C. albicans, we applied it to a previously well-characterized set of 29 C. albicans isolates evaluated by the random amplified polymorphic DNA (RAPD), multilocus enzyme electrophoresis (MLEE), and Ca3 Southern hybridization probe techniques. MLST identified three clusters of genetically related isolates, with 82.3% direct concordance with MLEE, 82.7% with RAPD analysis, and 86.2% with the Ca3 Southern hybridization technique. When MLST was applied to a subset of 22 isolates of unrelated origins, it identified 21 independent diploid sequence types (DSTs), resulting in a discriminatory power of 99.6%. These DSTs were 96.9, 99.6, and 99.6% concordant with the genotypes identified by RAPD analysis, MLEE, and Ca3 Southern hybridization, respectively. These results demonstrate that MLST is a highly effective technique that performs at least comparably to other established DNA fingerprinting techniques.

spa typing method for discriminating among Staphylococcus aureus isolates: implications for use of a single marker to detect genetic micro- and macrovariation.

Strain typing of microbial pathogens has two major aims: (i). to index genetic microvariation for use in outbreak investigations and (ii). to index genetic macrovariation for use in phylogenetic and population-based analyses. Until now, there has been no clear indication that one genetic marker can efficiently be used for both purposes. Previously, we had shown that DNA sequence analysis of the protein A gene variable repeat region (spa typing) provides a rapid and accurate method to discriminate Staphylococcus aureus outbreak isolates from those deemed epidemiologically unrelated. Here, using the hypothesis that the genetic macrovariation within a low-level recombinogenic species would accurately be characterized by a single-locus marker, we tested whether spa typing could congruently index the extensive genetic variation detected by a whole-genome DNA microarray in a collection of 36 isolates, which was recovered from 10 countries on four continents over a period of four decades, that is representative of the breadth of diversity within S. aureus. Using spa and coa typing, pulsed-field gel electrophoresis (PFGE), and microarray and multilocus enzyme electrophoresis (MLEE) data in molecular epidemiologic and evolutionary analyses, we determined that S. aureus likely has a primarily clonal population structure and that spa typing can singly index genetic variation with 88% direct concordance with the microarray and can correctly assign isolates to phylogenetic lineages. spa typing performed better than MLEE, PFGE, and coa typing in discriminatory power and in the degree of agreement with the microarray at various phylogenetic depths. This study showed that genetic analysis of the repeat region of protein A comprehensively characterizes both micro- and macrovariation in the primarily clonal population structure of S. aureus.