By Peter Malcolm Wallis, Brian R. Hammond
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Fresh TPS1 complete medium without the parasite and minimal essential medium (MEM) with 5% of calf serum were used as controls. 5 × 105 cells/mL to initiate the cultures for cellparasite interactions. For this purpose, effects of inocula of varying number of trophozoites ranging from 1 × 104 to 1 × 107 on HeLa and Vero cells at different intervals (0, 5, 18, 24, 48, 72 and 96 h) after incubation at 37°C were studied. 5 × 105 cells/mL • Corresponding author. In all the series, the experiments were terminated at different intervals (0 5, 18, 24, 48, 72 and 96 h) after incubation and the coverslips (2 for each interval in each series for the two cell systems studied), were removed from the tubes and processed for light and electron microscopic studies as described below.
The inability of Giardia trophozoites to undergo encystation in vitro has raised questions as to whether host related factors, such as gutassociated microorganisms or nutritive factors and stimuli from the small intestine, might play a role in this process. This model would be beneficial in many aspects of Giardia research, including biochemical analysis of the cyst and the cyst wall, development of giardicidal agents involved in blocking cyst wall formation, the testing of Kochs' postulates in regard to infectivity of cysts, and the investigation of questions related to nuclear division and cyst wall production.
The cyst with nonviable morphology (N) seen in Figure 3 was stained with the fluorogenic dye, PI, as shown in Figure 4, and fluoresced orange. Bar equals 5 microns for Figures 3 and 4. Figure 5. Bar equals 1 micron. Figures 6 and 7. SEMs of a Giardia culture after exposure to bile for three days. Giardia cysts (arrowheads) formed in vitro were seen scattered between flagellated trophozoites, which were attached to the substratum. At higher magnification, seen in Figure 7, the contrast between the flagellated trophozoite and the in vitro formed cyst was readily apparent.