FIG 4.
HbpA expressed from the Austrian strains binds Hb and Hb-Hp at the bacterial surface. (A) WC ELISAs were used to assess surface exposure of HbpA in the strains indicated. Bacteria were grown overnight in low-iron PGTH medium (2 μM EDDA), cell densities were normalized by OD600, and the cultures were used to coat microtiter plates. Expression and surface exposure of HbpA were measured by anti-HbpA antibody, with the secondary antibody used as a negative control. Abs, absorbance. (B) Binding to Hb and Hb-Hp was measured by incubation of cultures bound to microtiter plates with either Hb or Hb-Hp followed by detection of the bound hemoprotein with anti-Hb or anti-Hp antibody, respectively. Hp binding (with anti-Hp detection) was used as a negative control. Results show the mean and standard deviation from at least three experiments. For Hb and Hb-Hp binding (B), all comparisons between each strain and its corresponding deletion mutant were statistically significant (P < 0.0005) by unpaired t test using GraphPad Prism v9.1.2.