Depletion of Gr1⁺- cells reduces WITS diversity loss and prevents the cecal population crash between day 1 and day 2 p.i.

<p>(A) Schematic overview of the experimental setup. Starting with streptomycin-pretreatment one day prior to infection with the <i>S.</i> Tm^WT WITS cocktail in a dilution of 1∶7000, Gr1⁺-cells (mostly neutrophilic granulocytes) were depleted by daily i.p. injection of 100 µg of anti-mouse-Gr1-antibody. The control group was injected with an equal volume of PBS. (B) Successful reduction in Ly6C⁺ Ly6G⁺ cells in the murine blood was verified daily using a flow cytometry assay. A representative blot is given in panel B. (C) The cecal <i>S.</i> Tm population size was monitored in the Gr1⁺-cell depleted mice and the control group and organ loads and cecum pathology of respective animals were assessed. (D) CD18 immunofluorescent staining of cecal section from Gr1⁺-cell depleted mice and PBS treated control groups confirmed reduction of granulocytes in the course of the experiment. Scale bars: 50 µm (E) Quantification of luminal granulocytes from tissue sections. (F) At day 4 p.i. animals were sacrificed and the population composition in cecal contents were analyzed by rtqPCR. Organ pathogen loads and cecum pathology at day 4 p.i. indicate systemic pathogen spread and high intestinal inflammation. ns = not significant (P≥0.05), * P<0.05, ** P<0.01, *** P<0.001, Mann-Whitney U test. Grey lines indicate detection limits. Blue numbers indicate the median of the evenness indices of single mice.