May 2007
Volume 48, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2007
The Corneal Epithelial Cell as a Professional Phagocyte? Intracellular Trafficking and Survival of Pseudomonas aeruginosa Involves the Type III Secretion System
Author Affiliations & Notes
  • A. A. Angus
    University of California, Berkeley, Berkeley, California
    Graduate Group In Microbiology,
  • A. D. Ackerman
    University of California, Berkeley, Berkeley, California
    Graduate Group in Infectious Diseases and Immunity,
  • D. K. Augustine
    University of California, Berkeley, Berkeley, California
    Graduate Group In Microbiology,
  • S. M. J. Fleiszig
    University of California, Berkeley, Berkeley, California
    Graduate Program in Vision Sciences,
  • Footnotes
    Commercial Relationships A.A. Angus, None; A.D. Ackerman, None; D.K. Augustine, None; S.M.J. Fleiszig, None.
  • Footnotes
    Support NIH Grant EY11221 to S.M.J.F. and NSF Graduate Research Program Award to A.A.A.
Investigative Ophthalmology & Visual Science May 2007, Vol.48, 1101. doi:https://doi.org/
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      A. A. Angus, A. D. Ackerman, D. K. Augustine, S. M. J. Fleiszig; The Corneal Epithelial Cell as a Professional Phagocyte? Intracellular Trafficking and Survival of Pseudomonas aeruginosa Involves the Type III Secretion System. Invest. Ophthalmol. Vis. Sci. 2007;48(13):1101. doi: https://doi.org/.

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

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Abstract

Purpose:: Pseudomonas aeruginosa invades corneal epithelial cells during infection, but little is known about the events subsequent to invasion.

Methods:: Live cell imaging was performed using phase contrast microscopy of cells inoculated with ~108 cfu/ml P. aeruginosa. Real-time and time-lapse video was recorded 5, 7 and 10-hours post-infection. Intracellular bacterial viability was quantified 4 and 8 hours post infection using antibiotic survival assays. The role of the Type Three Secretion System (T3SS) was explored since structural components and secreted effectors of this system have been implicated in affecting host cell function.

Results:: Invasive strains of P. aeruginosa were found to cause membrane blebbing at later time points. Bacteria were localized within these blebs wherein they demonstrated rapid flagellar mediated motility. Mutants lacking the entire T3SS (exsA-) and those deficient in specific components of the secretion apparatus (pcsC- and popB-) did not cause blebbing. For strain PA103, mutants lacking all known effector genes (exoU-exoT-) retained the ability to cause blebs. In contrast, strain PAO1 lost blebbing capacity when all known effector genes were mutated (exoS-exoT-exoY-). Mutants which lost bleb forming capacity all localized to spacious intracellular vesicles adjacent to the nucleus. Wild type bacteria and mutants which caused blebbing could survive and replicate intracellularly. Loss of blebbing capacity (with trafficking to paranuclear vesicles) coincided with lack of ability to survive within cells, except for the popB mutant. This mutant, which secretes T3SS effectors but cannot translocate them across host cell membranes, caused a marked increase in the number of paranuclear vesicles as compared to other non blebbing mutants and was able to survive and grow within them.

Conclusions:: The T3SS of Pseudomonas aeruginosa influences intracellular bacterial trafficking and enables bacteria to survive and replicate within corneal epithelial cells. In its absence, bacteria are trafficked to paranuclear vesicles and are killed. Thus, human corneal epithelial cells can kill bacteria after internalization. Therefore the T3SS plays two roles in evading this innate defense strategy; one involving known effectors and manipulation of intracellular trafficking, and the other conferring protection against killing within paranuclear vesicles.

Keywords: bacterial disease • cornea: epithelium • Pseudomonas 
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