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M. E. Zegans, K. C. Cady, J. Wagner, J. H. Hammond, D. Murphy, R. T. Allar, A. A. Borquez, G. A. O'Toole; Bacteriophage DMS3 Inhibits Biofilm Formation and Swarming of Pseudomonas aeruginosa in a CRISPR Dependent Manner. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4486. doi: https://doi.org/.
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Bacterial biofilms are associated with pathogenic infections and antibiotic resistance. The gram negative bacterium Pseudomonas aeruginosa (Pa) is able to form biofilms on a variety of medical implants, including contact lenses and is an important corneal pathogen. Bacteriophages play an important role in modifying bacterial biology and virulence, but also have therapeutic potential in treating bacterial infection.
All genetic, biochemical and microbiologic assays were performed following previously published protocols. The Pseudomonas aeruginosa PA14 (PA14) strain was used in all experiments other than those involving clinical isolates. Ocular and non ocular clinical isolates from Dartmouth Medical School and the Campbell Laboratory at the University of Pittsburgh were also tested. Biofilm assays were incubated in minimal M63 medium supplemented 0.4% arginine at 37°C.
We observed that lysogenic infection of PA14 with bacteriophage DMS3 results in inhibition of biofilm formation and swarming motility, both important bacterial group behaviors. This inhibition requires the clustered regular interspaced short palindromic repeats (CRISPR) region in the host. CRISPRs and CRISPR associated genes (cas) are found in most Bacteria and Archaea. Mutating or deleting certain cas genes and 1 of the 2 CRISPR’s in this region restored biofilm formation and swarming in DMS3 lysogens. By PCR, cas genes were present in 50% of ocular Pa clinical isolates examined and infection with DMS3 inhibited binding of PA14 to contact lenses.
We have demonstrated that lysogenic bacteriophage infection of PA14 with DMS3 can inhibit biofilm formation and swarming and presented evidence that the host CRISPR region is required for this phenotype. The cas genes were commonly found in ocular isolates indicating that this system is conserved among clinically relevant isolates. These findings have important implications in developing antimicrobial strategies utilizing bacteriophage.
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