Number of bacteria and time of co-incubation with bacteria required for the development of Acanthamoeba keratitis

Abstract

Purpose : Recently, much interest has been focused on bacteria extracted from Acanthamoeba strains isolated from patients with Acanthamoeba keratitis. We hypothesized that bacteria may be a factor contributing to the development of Acanthamoeba keratitis and that the virulence of Acanthamoeba may depend on the time of coexistence with bacteria. We investigated interactions between Acanthamoeba and Pseudomonas aeruginosa for the development of keratitis in rabbit corneas.

Methods : Acanthamoeba castellanii (ATCC50492) and Pseudomonas aeruginosa (PAO-1) were used. Two densities of P. aeruginosa (high; 1×10⁸/ml, low; 3×10⁵/ml) and two durations of co-incubation (long; 6 h, short 2 h) of Acanthamoeba with 1×10⁸/ml of P. aeruginosa were tested. Acanthamoeba alone or Acanthamoeba co-incubated with P. aeruginosa was inoculated into rabbit corneas. After inoculation of the microorganisms, levofloxacin eye drops were administered. Clinical score of the cornea was evaluated after inoculation.

Results : Inoculation of Acanthamoeba alone did not produce keratitis during a 5-day observation period. Rabbit corneas inoculated with Acanthamoeba co-incubated with low-density P. aeruginosa followed by topical levofloxacin treatment were clear with few infiltrates and the corneas remained clear. Corneas inoculated with Acanthamoeba co-incubated with high-density P. aeruginosa followed by levofloxacin treatment developed severe keratitis, and clinical scores were significantly higher compared to high-density P. aeruginosa alone followed by levofloxacin treatment (scores 7, 9.6, 8.5versus 3, 3.5, 3.25 on days 1- 3, all p<0.01). Using high-density P. aeruginosa, long (6 h) co-incubation time of Acanthamoeba with P. aeruginosa resulted in more severe keratitis compared to short (2 h) co-incubation time (scores , 9.7, 12.7, 12.1, 9.8, 8.7 versus 7, 9.6, 8.5, 6.9, 5.6 on days 1-5, all P<0.01).

Conclusions : These results suggest that the presence of bacteria is essential and a critical number of bacteria is required for the development of Acanthamoeba keratitis. The time of coexistence with bacteria may be an important determinant of the severity of Acanthamoeba keratitis.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.