Figure 1.
Mutation in ropB results in truncation of the RopB protein and abrogation of SpeB expression.
(A) Schematic representation of the RopB protein expressed by GAS strains 5448 and 5628. In 5628, white region represent homology to 5448, shaded region indicates unique protein sequence in 5628. (B) Western blot for SpeB using overnight culture supernatants. (C) SpeB activity assay using azocasein substrate.
Table 1.
Primers used for sequencing the ropB locus (including speB), the covR/S locus, luxS, rofA and ropA.
Figure 2.
RopB mutation does not affect in vitro growth, hemolytic activity, extracellular DNase activity or resistance to neutrophil killing.
(A) OD600 of in vitro grown bacterial cultures in THB. (B) Hemolysis of red blood cells by bacterial culture supernatants. (C) Degradation of calf thymus DNA by bacterial culture supernatants, run on a 1% agarose gel. (D) Bacterial survival after 30 min incubation with human neutrophils.
Figure 3.
Microarray analysis emphasizes the down-regulation of the SpeB operon.
The figure shows the top 10 genes up- and down-regulated in the ropB mutant bacteria compared to WT when the bacteria are inoculated in vivo (P<0.05). The values plotted represent log ratios (log mutant/WT), and genes co-clustered in operons are highlighted (black bars, SpeB operon; white bars, SLS operon).
Figure 4.
RopB and SpeB-negative bacteria show reduced virulence in systemic infection models.
(A) Intraperitoneal infection of C57BL6/J mice with 5×107 cfu of GAS strains with 5% mucin. (B) Intravenous challenge of C57BL6/J mice with 2×108 cfu of GAS strains.