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Jorge Maestre, Edith Perez, Martha Diaz, Eduardo Alfonso, Darlene Miller; Molelcular Characterization of Virulence Genes Associated with MRSA Keratitis isolates. Invest. Ophthalmol. Vis. Sci. 2012;53(14):6140.
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To identify and characterize the molecular spectrum and frequency of virulence genes associated with MRSA isolates recovered from keratitis.
Conventional and multiplex PCR assays were run to verify the presence of the mecA gene and determine isolate origin (healthcare (HCA) vs community (CA) associated via SCCmec Cassette type profiling of 63 randomly selected isolatesrecovered from patients diagnosed with MRSA keratitis within the last 9 years.Separate molecular assays were run to confirm the presence of the Panton Valentin Leukocidin (PVL) necrotizing toxin gene and to genotype the global quorum sensing accessory gene regulator (agr) locus ( agr types I, III = CA, SCCmec IV, PVL+, USA300-800;agr type II= HCA, SCCmec II, PVL-,USA100, vancomycin tolerance/resistance). SCCmec profiles, PVL genes and agrprofiles were correlated with presenting diagnosis, antibiotic exposures and vancomycin e of MICs.
The mecA virulence gene was confirmed in 82.5% (N=52/63) percent of the MRSA isolates identified by conventional microbiology testing. The 11 (17.5%) mecA negative/phenotypic MRSA isolates were resistant to methicillin by another mechanism and were less likely to be surgery related (0 vs 15.4%), and or exposed to antibiotics (42.9% vs 73.8%). Both HCA-MRSA (SCCmec type II, 88.3%) and CA-MRSA (SCCmec types IV, V, 11.7%) were documented among the 43 (82.7%) typable strains. Clinical diagnosis included keratitis, NOS(N=52, 82,5), post-pk (N=8, 12.7%)and other (N=3, 4.8%). All identified post-pk and contact lens wearers (N=10-15.9%) were documented in the mecA positive group. The PVL toxin gene was documented in 5.8% (N=3) of the mecA positive isolates and 18.2% (N=2) of the mecA negative isolates with an overall positive rate of 7.9% (N=5/63) . 82.7% (N=43) of the 52 confirmed MRSA isolates and 63.4% (N=7) of the mecA negative isolates were agr genotype II. Overall 79.4% (N=50/63) were arg type II, 17.4% (N=11/63), agr type I, and 3.2%, agr type III. Agreement between SCCmec II and agr II was 93.0%. Vancomycin MIC values ranged was 0.5-4 ug/ml with the MIC90=2 ug/ml at the higher limits of susceptible for both mecA positive (96.1% of isolates inhibited) and mecA negative (100% inhibited). Inhibition by agr types: I (90.9%), II (98%), III (100%).
MRSA isolates recovered from keratitis are predominantly healthcare related (SCCmec II), PVL negative and harbor the quorum sensing genotype for vancomycin heteroresistance (agr II). This combination of virulence genes may impact efficacy and selection of antibiotic prophylaxis and therapeutic management of patients presenting with MRSA keratitis.
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