May 2004
Volume 45, Issue 13
ARVO Annual Meeting Abstract  |   May 2004
Translocation of Pseudomonas aeruginosa Across Intact Corneal Epithelium In vitro Requires Twitching Motility
Author Affiliations & Notes
  • S.J. Fleiszig
    School of Optometry,
    Univ of California–Berkeley, Berkeley, CA
  • I. Alarcon
    Dept of Microbiology & School of Optometry,
    Univ of California–Berkeley, Berkeley, CA
  • Footnotes
    Commercial Relationships  S.J. Fleiszig, None; I. Alarcon, None.
  • Footnotes
    Support  R01–EY11221
Investigative Ophthalmology & Visual Science May 2004, Vol.45, 4640. doi:
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      S.J. Fleiszig, I. Alarcon; Translocation of Pseudomonas aeruginosa Across Intact Corneal Epithelium In vitro Requires Twitching Motility . Invest. Ophthalmol. Vis. Sci. 2004;45(13):4640.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract: : Purpose: Contact lens wear is the most common predisposing factor for corneal infection caused by Pseudomonas aeruginosa (PA). The aim of this study was to begin to explore the strategies used by bacteria to cross the corneal epithelium in the absence of full thickness epithelial injury. Methods: Rabbit corneal epithelial cell layers were grown to confluence on 3 µm pore–sized filters in vitro, and the apical chamber was then inoculated for 6 h using 106 CFU of invasive PA strain PAK. At hourly time points, viable counts were taken from both the upper and lower chambers, and the transepithelial resistance (TER) was compared to HBSS–inoculated (negative control) and EGTA treated (positive control) cells. The role of twitching motility in bacterial translocation across the cells was explored by comparing the wild–type bacteria to pilU (twitching) mutant bacteria, and pilA (pili) mutant bacteria. Results: Bacteria began to appear in the lower chamber after 1 h and by 8 h the upper/lower chamber numbers approached equilibration. Unexpectedly, the TER across infected cell layers remained above baseline, and similar to uninoculated cells, throughout the 8 h assay. The pilA and pilU mutants were markedly attenuated in their ability to translocate across the cells compared to the wild type PAK (> 2 log reduction, p = 0.01 in both cases). Conclusions: Invasive PA, which possess the capacity to invade individual corneal epithelial cells, can translocate multilayered corneal epithelium in vitro without reducing TER. These results suggest that the tight junction barrier between cells, required for maintaining TER, is not disturbed during the process. Whether or not intracellular travel is used for translocation is yet to be determined; however, the data obtained with mutant bacteria demonstrate a yet to be defined role for twitching motility.

Keywords: Pseudomonas • cornea: epithelium • microbial pathogenesis: experimental studies 

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