Abstract
Purpose: :
While P. aeruginosa adhesion to contact lenses has been studied, published literature has been confusing or even contradictory. Bacterial gene expression is regulated in response to prevailing environmental conditions. Thus, this study tests the hypothesis that bacteria express multiple phenotypes when interacting with the surface of a contact lens, possibly relating to the confusion in this field.
Methods: :
Mian’s minimal media containing 5x106cfu/ml bacteria was pumped through a flow chamber containing a mounted contact lens for 1 h at a shear rate of 0.3 s-1. The lens was positioned under a microscope to enable attachment dynamics to be visualized and quantified. The role of flagella was studied using fliC (flagellin) mutants of PAO1 or PAK compared to respective wild-types, and using PA103 which lacks flagella. Swimming motility was studied using PAK motAB that retains flagella but cannot swim. PAO1 pilA-was used to study the role of pili. LPS was examined using O-antigen mutants: PAO1::gmd (lacks A-band), PAO1::wpmB (lacks B-band), and PAO1::rmlC (lacks both A and B-band).
Results: :
Individual bacteria expressed different attachment phenotypes within the experimental system. Some approached the lens surface and adhered, while others became "tethered" to the lens surface. A subgroup of each later detached. Still others approached the surface but did not attach. PAO1 attached at a rate of 360 cells/mm2-min, 40 times faster than PAO1 fliC- (8 cells/mm2-min). A similar difference was seen for PAK (220 cells/mm2-min) compared to its fliC mutant (13 cells/mm2-min). Interestingly, PAK motAB- was also defective in adhesion (8 cells/mm2-min). Accordingly, nonmotile PA103 attached slowly at 4 cells/mm2-min. There was no attachment difference between PAO1 and PAO1 pilA-. The PAO1 parent of the LPS mutants adhered at 760 cells/mm2-min, slightly faster than its mutants: PAO1::gmd, at 550; PAO1::wpmb, at 310; and PAO1::rmlC, at 580 cells/mm2-min. Tethered bacteria were not seen for motAB-, fliC-, or PA103, but all other mutants displayed similar phenotypes to wild-type.
Conclusions: :
Swimming motility is required for efficient adhesion to contact lenses, but the presence of flagella (appendages that mediate swimming) is not sufficient. Neither pili, nor specific LPS O-antigens are needed. Swimming motility was required for the tethering phenotype. Mechanisms for the involvement of swimming are to be elucidated, but may include promoting access to the lens surface, a direct role in adhesion, or a role in regulating the expression of unidentified adhesion factors.
Keywords: contact lens • pseudomonas • microbial pathogenesis: experimental studies