Figure 6. mosGCTLs interrupt the deposition of AMPs onto bacteria cells.

a,b, Determination of the interaction between mosGCTL-32 and E. coli cells by confocal microscopy (a) and flow cytometry (b). c, The interaction between mosGCTL-32 and bacterial cells was mediated by surface polysaccharides. (i) Elution assay. (ii) Competitive assay. All samples with mosGCTLs was determined by western blotting with an anti-V5 antibody. d,e, mosGCTLs interrupt the deposition of AMPs onto bacterial cells. The cells with AMPs were assessed using an immunofluorescence assay (d) or numbered using a flow cytometry assay (e). The E. coli cells (ST515 strain) were equipped with a GFP reporter with constitutive expression. f,g, The role of mosGCTLs in maintenance of commensal bacteria in the mosquito midguts. The mosGCTLs-silencing mosquitoes were fed with the Alexa 546-labelled Cec mixture (with an equal amount of Cec A, B and E). Mosquitoes inoculated with GFP dsRNA were used as negative controls. The midguts of fed mosquitoes were dissected at 4 h post oral feeding. (f) SYTO 16 green fluorescent nucleic acid dye was used to stain gut microbiome DNA. (g) The percentage (%) of Cec-coated cells was calculated as red-stained cells/total green-stained cells. The result was independently measured in the six different midguts. Data are represented as mean±s.d. in each group and analysed using the non-parametric Mann–Whitney test. a,d,f, The bacteria were imaged with the multiple track mode of a Zeiss LSM780 meta confocal microscopy. The scale bars represent 5 μm in a, b, and the right panel of f (×100), and represent 20 μm in the left panel of f (×40). a–g, The experiments were biologically repeated three times with similar results.