2008 OMIG, Abstract 11

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Clinical isolates of P. aeruginosa harbor bacteriophages and CRISPR genes
Rory T. Allar¹, Kyle C. Cady¹, John H. Hammond¹, George A. O’Toole¹, Michael E. Zegans^1,2
1Department of Microbiology and Immunology, ²Department of Surgery (Ophthalmology) Dartmouth Medical School, Hanover, NH

Purpose:Bacteriophages exert a profound effect on bacterial ecology and pathogenesis. There has been little investigation of the role of bacteriophage during Pseudomonas aeruginosa (PA)infections of the ocular surface. CRISPR genes have recently been implicated in resisting bacteriophage infection through a proposed RNAi-like mechanism and we have found that CRISPR-bacteriophage interactions can alter bacterial biofilm formation
Methods:All genetic, biochemical and microbiologic assays were performed following previously published protocols. Clinical isolates were obtained from Dartmouth Medical School and the Campbell Lab at UPMC. 43 ocular isolates, 12 CF isolates, 25 sputum isolates and 16 urine isolates of PA were studied. The presence of phage was detected by the presence or absence of plaque formation on lawns of 3 PA lab strains (PA01, PA14 and PAK). PCR was performed on all clinical isolates using primers with homology to the cas1 gene, a highly conserved CRISPR-associated gene of PA14. Additional DNA sequencing was performed and analyzed in cas1 containing strains.
Results:

Sequence analysis of CRISPR regions in select cas1 containing strains demonstrated strong homology to the PA14 CRISPR region.
Conclusions:Clinical isolates of PA are often infected with bacteriophage and commonly contain a CRISPR region with homology to that of the lab strain PA14.  These results suggest that bacteriophages and bacterial responses to them may play an important role in the biology strains infecting the eye.

Disclosure code: N