Figure 3

Gut microinjection confirms the roles TLR2 and the microbiome in MAC 101 and Mma20 proliferation. (A) Schematic of the gut microinjection procedure. (B) Robotic gut microinjection. The injection site is the area of the anterior bulb of the gut. Successful injection was characterized by the absence of extra-intestinal leakage of bacteria. (C,D) Representative fluorescence images showing three situations after 24 hours of gut microinjection of MAC 101 (C) and Mma20 (D). Representative images are tlr2 wild-type larvae under germ-free (GF) condition. Bacteria are shown in magenta. Situation 1 represents clearance of bacteria, Situation 2 represents retention of bacteria within the gut, and Situation 3 represents dissemination of bacteria outside the gut. Scale bar: 100 µm for full-body images; 25 µm for zooms. (E,I) Frequency of each situation after 24 hours of gut microinjection of MAC 101 (E) and Mma20 (I). (F,J) Total bacterial burden calculated by combining situations 2 and 3 for MAC 101 (F) and Mma20 (J). (G,K) Bacterial burden of situations 2 for MAC 101 (G) and Mma20 (K). (H,L) Bacterial burden of situations 3 for MAC 101 (H) and Mma20 (L). For MAC 101 infection, the data from the GF tlr2^+/+ (n=103) group, GF tlr2^-/- (n=79) group, CONVD tlr2^+/+ (n=92) group and CONVD tlr2^-/- (n=110) group are based on three independent experiments. For Mma20 infection, the data from GF tlr2^+/+ (n=79) group, GF tlr2^-/- (n=72) group, CONVD tlr2^+/+ (n=59) group and CONVD tlr2^-/- (n=62) group are based on three independent experiments. Statistical significant difference was determined by one-way ANOVA, red arrows point to the median, ns, non-significant, *, P < 0.05, **, P < 0.01, ***, P < 0.001, ****, P < 0.0001.