Relationship between cytotoxicity and corneal epithelial cell invasion by clinical isolates of Pseudomonas aeruginosa

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Abstract
We have reported that some strains of Pseudomonas aeruginosa can enter corneal epithelial cells during experimental murine eye infection and when the cells are cultured in vitro. Following invasion, both the host cell and the intracellular bacteria can remain viable for up to 24 h. Others have reported that toxin-mediated damage of epithelial cells contributes to the pathogenesis of P. aeruginosa keratitis. To clarify the relationship between cell invasion and cytotoxicity, fourteen P. aeruginosa isolates were compared for their capacity to enter epithelial cells and for their ability to induce cytotoxicity. Bacterial invasion was quantified by gentamicin survival assays both in vivo and in vitro. Cytotoxicity was examined qualitatively by trypan blue exclusion assays and quantitatively by chromium release assays in vitro. A significant inverse correlation was found between the ability to induce cytotoxicity and epithelial cell invasion as measured by gentamicin survival assays. Both cytotoxic and noncytotoxic strains were identified among corneal and noncorneal isolates; all isolates that were not cytotoxic were capable of epithelial cell invasion. Efficient host cell invasion could not be demonstrated for cytotoxic strains; however, the gentamicin survival assay relies upon host cells retaining viability in order to yield useful results, and this may limit the effectiveness of this assay for testing epithelial cell invasion by cytotoxic strains. Since all of the corneal isolates that were tested were virulent in vivo, the results show that there are at least two different types of P. aeruginosa-induced disease, one caused by strains that are cytotoxic and the other involving bacteria that can enter epithelial cells and survive intracellularly without killing the host cell.