SvpA, a novel surface virulence-associated protein required for intracellular survival of Listeria monocytogenes

A previously unknown protein, designated SvpA (surface virulence-associated protein) and implicated in the virulence of the intracellular pathogen Lis teria monocytogenes, was identified. This 64 kDa protein, encoded by svpA, is both secreted in culture supernatants and surface-exposed, as shown by i mmunogold labelling of whole bacteria with an anti-SvpA antibody. Analysis of the peptide sequence revealed that SvpA contains a leader peptide, a pre dicted C-terminal transmembrane region and a positively charged tail resemb ling that of the surface protein ActA, suggesting that SvpA might partially reassociate with the bacterial surface by its C-terminal membrane anchor. An allelic mutant was constructed by disrupting svpA in the wild-type strai n L028. The virulence of this mutant was strongly attenuated in the mouse, with a 2 log decrease in the LD50 and restricted bacterial growth in organs as compared to the wild-type strain. This reduced virulence was not relate d either to a loss of adherence or to a lower expression of known virulence factors, which remained unaffected in the svpA mutant. It was caused by a restriction of intracellular growth of mutant bacteria. By following the in tracellular behaviour of bacteria within bone-marrow-derived macrophages by confocal and electron microscopy studies, it was found that most svpA muta nt bacteria remained confined within phagosomes, in contrast to wild-type b acteria which rapidly escaped to the cytoplasm. The regulation svpA was ind ependent of PrfA, the transcriptional activator of virulence genes in L. mo nocytogenes. In fact, SvpA was down-regulated by MecA, ClpC and ClpP, which are highly homologous to proteins of Bacillus subtilis forming a regulator y complex controlling the competence state of this saprophyte. The results indicate that: (i) SvpA is a novel factor involved in the virulence of L. m onocytogenes, promoting bacterial escape from phagosomes of macrophages, (i i) SvpA is, at least partially, associated with the surface of bacteria; an d (iii) SvpA is PrfA-independent and controlled by a MecA-dependent regulat ory network.