Polymorphonuclear Leukocyte Enhancement of Bacterial Biofilms on Contact Lens Surfaces

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.