Fig. 3: TEM analysis of E. coli W SslE aggregations.

a Accumulation of SslE aggregates on the side of glass tubes containing overnight cultures grown in LB media at pH 5.0 or 7.0. Arrows indicate Congo red-stained ring containing SslE. b–g SslE localisation upon secretion from E. coli W strains assessed by immunoelectron microscopy. Bacteria were reacted with primary α-rSslE antibodies and secondary gold-labelled antibodies on carbon-coated nickel grids and then negatively stained with uranyl acetate. b, f E. coli W ΔsslE mutant washed in citrate-phosphate buffer at pH 5.0 or 4.0, respectively. c Wild-type W strain washed in citrate-phosphate buffer at pH 5.0 with α-rSslE antibodies reacting with the bacterial surface and secreted fibrous material. d Image highlighting clustering of antibodies to an extracellular aggregate. e Analysis of total and clustered (≤25 nm between particles) α-rSslE antibodies reacting with aggregates in the extracellular milieu of wild-type and ΔsslE mutant bacteria at pH 5.0. ^***P < 0.001; verses ΔsslE mutant by two-tailed Student’s test and error bars are the standard error of the mean. g ΔsslE::sslE W strain washed in citrate-phosphate buffer at pH 4.0, again with α-rSslE antibodies reacting with the bacterial surface and secreted fibrous material. Scale bar is equivalent to 200 nm. h TEM analysis of negatively stained rSslE aggregates. Two major species are observed. i Smaller filaments have an average width of ∼4.5 nm and are coated with globular structures of ~4.5 nm in diameter. Scale bar, 20 nm. j Schematic describing the overall dimensions of the smaller SslE fibre species. k Larger species of SslE aggregates appear as ~20–40 nm wide structures with variable lengths. Scale bar, 20 nm.