Cytopathogenicity of Acanthamoeba isolates on rat glial C6 cell line.

The pathogenicity of Acanthamoeba isolates from keratitis patients (the Hamburg isolate from Germany, H-1 and a Philippine isolate, IB-1-7) as well as an environmental isolate, W4 was assayed in vitro using rat glial C6 cell line. Results indicate that both live amebae and cell-free supenatants from H-1 and IB-1-7 clones produced cytopathic effects (CPE) on rat glial C6 cells in a dose-and-time-dependent fashion. A dose of 10(5) cells/ml induced death and moderate areas of destruction of individual cells after 48 hours of incubation. Results of both free zone capillary electrophoresis and sodium dodecyl sulphate polyacrylamide gel electrophoresis suggest the release of amebic products to the culture medium that could at least partially explain the observed cytopathogenicity after 48 hours. Furthermore, results of SDS-PAGE indicate differences between the secretions of the isolates, with bands produced by the two ocular isolates that were not seen with the environmental isolates. That the secretions can produce a cytopathic effect (CPE) has been shown by the cytotoxicity assays using protein concentrations of the secretory products. Protein concentration of 0.30 microg/microl of culture supenatants from H-1 and IB-1-7 clones produced similar effects on the cell monolayers after 2 hours of incubation. This concentration caused the highest % cell death as measured by both trypan blue exclusion (TBE) and 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl-tetrazolium bromide (MTT) assays. In contrast, using W4 clone, corresponding concentrations of both trophozoites and culture supernatant did not cause significant cell death and cellular disintegration.

The pathogenicity of a Philippine isolate of Naegleria sp. in mice: effects of dose levels and routes of infection.

The pathogenicity of a Philippine isolate of Naegleria sp. was evaluated using 3-4 week-old mice as experimental animals. Results showed that only the massive doses of 10(6) and 10(7) amebae/mouse inoculated intranasally could successfully establish ameba infection in the brain and cause death after 2-6 days. The effect of the ameba on the mortality rate of inoculated mice was dose-dependent. The amebae were recovered in the brain when inoculated through intracerebral and intranasal routes and in the lungs, liver, and intestines when administered through intranasal and oral routes. By intraperitoneal inoculation, recovery of amebae was positive in all major organs except in the heart. Intravenous inoculation resulted to positive recovery in the lungs, spleen, liver, and heart. Infectivity of the ameba isolate in major organs was route-dependent.