Fig 2: Depletion of host macrophages alters the outcome of GBS infection in a cadD-dependent manner.A Conceptual diagram of methods utilized in these studies. Pregnant mice were intraperitoneally injected on embryonic day 12.5 and 14.5 with either anti-F4/80 antibody to deplete host macrophages, or an isotype control antibody. Mice were infected with GBS on embryonic day 13.5 and sacrificed 2 days post-infection. Vaginal tissues were collected from three separate dams for each experimental condition for analyses. Uterine tissues were collected from four separate dams for each experimental condition for analyses. For decidual, placental, amnion, and fetal tissues, each individual data point indicates an independent fetal-placental unit. B Flow cytometry confirms that macrophages were depleted within the decidua and placenta at the maternal-fetal interface. C Placental units were analyzed by immunohistochemical techniques using a polyclonal antibody to stain specifically for GBS (indicated by the brown stain). Micrographs are representative of three biological replicates which were analyzed in a blinded fashion. Microscopical imaging at ×100 magnification reveals bacterial invasion of the reproductive tract in pregnant animals infected with WT GBS (GB112), the ?cadD mutant (?cadD), and the complemented isogenic ?cadD mutant (?cadD:C). Micrographs were analyzed in a blinded fashion and are representative of three biological replicates. D Bacterial burden within reproductive tissues was evaluated by quantitative culture (Log CFU/mg) in response to macrophage depletion via anti-F4/80 injections (+) compared to isotype control treatments (-) lacking macrophage depletion. Significant changes in burden were observed in WT GB112-infected mice between the isotype control and the anti-F4/80 treatment groups in the decidua (*P = 0.020), placenta (*P = 0.0008), amnion (*P = 0.0312), and fetus (*P = 0.0266) by one-tailed Mann–Whitney U analysis. These results were also observed in the cohort of animals infected with the complemented mutant in the decidua (*P = 0.0385), placenta (*P = 0.0130), amnion (*P = 0.0011), and fetus (*P = 0.0242) by one-tailed Mann–Whitney U test, but not in the animals infected with the isogenic ?cadD mutant (NS = not significant).