Affiliation:
1. The Philips Institute of Oral and Craniofacial Molecular Biology
2. Department of Microbiology and Immunology
3. Center for the Study of Biological Complexity
4. Department of Adult Health Nursing, Virginia Commonwealth University, Richmond, Virginia 23298
Abstract
ABSTRACT
Streptococcus sanguinis
is an important cause of infective endocarditis. Previous studies have identified lipoproteins as virulence determinants in other streptococcal species. Using a bioinformatic approach, we identified 52 putative lipoprotein genes in
S. sanguinis
strain SK36 as well as genes encoding the lipoprotein-processing enzymes prolipoprotein diacylglyceryl transferase (
lgt
) and signal peptidase II (
lspA
). We employed a directed signature-tagged mutagenesis approach to systematically disrupt these genes and screen each mutant for the loss of virulence in an animal model of endocarditis. All mutants were viable. In competitive index assays, mutation of a putative phosphate transporter reduced in vivo competitiveness by 14-fold but also reduced in vitro viability by more than 20-fold. Mutations in
lgt
,
lspA
, or an uncharacterized lipoprotein gene reduced competitiveness by two- to threefold in the animal model and in broth culture. Mutation of
ssaB
, encoding a putative metal transporter, produced a similar effect in culture but reduced in vivo competiveness by >1,000-fold. [
3
H]palmitate labeling and Western blot analysis confirmed that the
lgt
mutant failed to acylate lipoproteins, that the
lspA
mutant had a general defect in lipoprotein cleavage, and that SsaB was processed differently in both mutants. These results indicate that the loss of a single lipoprotein, SsaB, dramatically reduces endocarditis virulence, whereas the loss of most other lipoproteins or of normal lipoprotein processing has no more than a minor effect on virulence.
Publisher
American Society for Microbiology
Subject
Molecular Biology,Microbiology
Cited by
53 articles.
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