Browsing by Author "Patel, Naiya"
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Item Efficacy of Contact Lens Care Solutions Against Neutrophil Enhanced Biofilms(2015-01-26) Hinojosa, Jorge A.; Patel, Naiya; Zhu, Meifang; Robertson, Danielle M.PURPOSE: To evaluate currently available chemically preserved and peroxide-based lens care products (LCPs) antimicrobial efficacies against neutrophil enhanced biofilms in reference strains of S.auerus(SA), S. marcescens (SM), S. maltophilia(ST), and P.aeruginosa(PA). METHODS: Lotrafilcon B lenses were inoculated with 2.5 x 107 CFUs of American Type Culture Collection (ATCC) reference strains of either SA, SM, ST, or PA and 8.3x106 cells of human neutrophils harvested by whole blood centrifugation. Lenses were then incubated over night under conditions that facilitate biofilm formation. Lenses were then disinfected with either multipurpose contact lens solutions (MPS): Biotrue (BT), PureMoist (PM) or hydrogen peroxide-based lens care systems (HPB): ClearCare (CC), PeroxiClear (PC) according to specific manufacture guidelines. Antimicrobial activity was then quantified by quantitative culturing (colony-forming units) and by fluorescence confocal microscopy (FCM) using LIVE/DEAD BacLight Bacterial Viability Kit (Molecular Probes, Eugene, OR). RESULTS: Mean colony growth counts for each bacterial strain + neutrophils were: SA= 1.44 x 107 (CFU/ml), SM= 2.56x109 (CFU/ml), ST= 1.17x1010 (CFU/ml), PA= 3.43x109 (CFU/ml). After treatment, SA exhibited a 7-log reduction with all LCPs (P=0.609); SM exhibited 9-log reduction for all LCPs except BT, which showed a 7-log reduction (P < 0.001); ST exhibited a 9-log to 10-log reduction for all LCPs (P=0.040); PA showed a 9-log reduction for all LCPs (P=1.000). FCM with viability staining revealed the presence many unviable bacteria still adhered to the lens surface after treatment with LCPs. CONCLUSIONS: Although all LCPs meet FDA criterion of a 3-log reduction minimum even against neutrophil enhanced biofilms, FCM revealed that many remnants still remain adhered on the lens surface after just one use. Further deposition of bacterial products from continued used could enhance biofilm formation and promote a host inflammatory response, both of which could precipitate into a corneal infiltrative event (CIE). In addition, mean colony growth counts showed that SA exhibited the lowest growth on lens surfaces, which may explain why SA is more commonly associated with sterile CIEs.Item Polymorphonuclear Leukocyte Enhancement of Bacterial Biofilms on Contact Lens Surfaces(2015-01-26) Patel, Naiya; Hinojosa, Jorge A.; Zhu, Meifang; Robertson, Danielle M.Contact lens-wear represents a leading risk factor for the development of infectious keratitis, which can result in significant vision loss. It is well established that bacterial colonization of the posterior lens surface represents the initial event in the pathogenesis of lens-related infection. Using an invasive clinical isolate, our prior work has shown that Pseudomonas aeruginosa biofilm formation on contact lens surfaces is dramatically accelerated in the presence of dying neutrophils. The goal of this study was to investigate the capacity of five FDA test strains, all commonly associated with contact lens-related infiltrative events, to form biofilms on contact lens surfaces in the presence of neutrophil-derived cellular debris. Neutrophils were obtained from healthy, human volunteers by venipuncture and isolated using Ficoll gradient separation. Unworn Lotrafilcon B silicone hydrogel contact lenses were incubated overnight in one of five reference strains from the American Type Culture Collection with or without neutrophils at a 1:1 bacteria:neutrophil ratio. Test strains included: Pseudomonas aeruginosa, Staphylococcus epidermidis, Staphylococcus auerus, Stenotrophomonas maltophilia, and Serratia marcescens. Adherent bacteria were visualized using scanning electron microscopy (SEM) or stained using a BacLight live/dead assay followed by laser scanning confocal microscopy. The number of viable bacteria adherent to the lens surface was also quantified by standard colony counts. Live/dead staining showed greater numbers of viable bacteria adherent to lens surfaces when cultured in the presence of neutrophils. Colony counts confirmed a higher number of viable bacteria for four of the five test strains: S. auerus (p<0.001), S. maltophilia (p<0.001), P. aeruginosa (p=0.030), and S. marcescens (p<0.001). This effect was not evident with S. epidermidis (p=0.659). SEM showed similar findings. This is the first study to demonstrate the ability of these reference strains to form biofilms on contact lens surfaces in the presence of neutrophils. These findings suggest that, in the setting of intense inflammation under the lens, common contact lens-related pathogens possess the capacity to colonize and resist clearance by the innate immune system. Further studies are needed to correlate these findings with disease in an animal model.