Characterization of a chimeric FH expressed by Tg mice, and infection experiments with Tg and normal mice.

<p>(<b>A</b>) SDS-PAGE and western blot analysis of purified chimeric FH (chim. FH), derived from mouse and human FH. This FH was compared with pure human Y402 FH (hFH) and pure mouse FH (mFH). Streptococcal protein Rib <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003323#ppat.1003323-StlhammarCarlemalm1" target="_blank">[81]</a> was included as negative control. The blot was probed with antiserum to human FH. Bound antibodies were detected with radiolabeled protein G followed by autoradiography. (<b>B</b>) Binding of the M5 protein to immobilized FH proteins. The proteins used were those shown in panel A. The wells of microtiter plates were coated with the protein indicated, using 0.1 µg protein per well, with protein Rib as negative control. The immobilized proteins were analyzed for ability to bind biotinylated pure M5 protein, added at the concentrations indicated, and bound M protein was detected by the addition of radiolabeled streptavidin. (<b>C</b>) Comparison of bacterial growth in wild type mice and Tg mice. The mice (n = 8 for wt and n = 8 for Tg) were subjected to invasive infection with a sublethal dose of the <i>S. pyogenes</i> M5 strain and sacrificed after 18 h, followed by determination of bacterial counts in the spleen. (<b>D</b>) Survival of wild type (wt) and Tg mice after infection with M5 bacteria. The mice (n = 7 for wt and n = 10 for Tg) were subjected to invasive infection with an ∼LD₉₀ dose of the <i>S. pyogenes</i> M5 strain and survival was recorded regularly, as indicated. (<b>E</b>) Survival of wild type (wt) and Tg mice after infection with M5 bacteria. The mice (n = 12 for wt and n = 12 for Tg) were subjected to invasive infection, as in D, but with a lower bacterial dose. (<b>F</b>) Growth of the <i>S. pyogenes</i> M5 strain in wild type mice injected with pure human Y402 FH (2×100 µg) or with PBS. The mice (n = 6 for FH and n = 6 for PBS) received FH or PBS 4 h before the infection and mixed with the infecting bacteria. Bacterial counts in spleens were analyzed, as in (C). (<b>G</b>) Growth of the <i>S. pyogenes</i> M5 strain in wild type mice injected with normal human serum (NHS) or normal mouse serum (NMS). The mice (n = 8 for NHS and n = 7 for NMS) were injected with two 200 µl doses of serum, one dose given 4 h before the infection and one mixed with the infecting bacteria. Analysis as in (C).