Beclomethasone effects on Mm infection progression and bacterial dissemination. (A–C). Bacterial burden (A), number of bacterial clusters (B) and the average size of bacterial clusters (C) were determined at 1, 2, 3 and 4 dpi following intravenous Mm injection (28 hpf) and treatment with vehicle or 25 µM beclomethasone, started at 2 h before infection. Significant increases due to the beclomethasone treatment were observed for all parameters at 4 dpi. For the number of bacterial clusters, the increase was also significant at 3 dpi. Statistical analysis was performed by two-way ANOVA with Tukey’s post hoc test. Each data point represents a single larva and the means ± s.e.m. of data accumulated from three independent experiments are shown in red. Statistical significance is indicated by: ***P<0.001; ****P<0.0001. (D) Effect of beclomethasone on dissemination of Mm by hindbrain ventricle injection. Hindbrain infections were performed at 28 hpf, and at 24 hours post infection (hpi), a significantly increased percentage of larvae with disseminated Mm infection was detected in the beclomethasone-treated group compared to the vehicle group. Statistical analysis was performed by two-tailed t-test. Values shown are the means ± s.e.m. of three independent experiments with a total sample size of 27 in the vehicle-treated group and 31 in the beclomethasone-treated group. Statistical significance is indicated by: *P<0.05. (E) Representative images of embryos with and without dissemination of the infection upon hindbrain injection of Mm. Scale bar = 200 μm.
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