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Infection and Immunity, July 2009, p. 2719-2729, Vol. 77, No. 7
0019-9567/09/$08.00+0     doi:10.1128/IAI.00617-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Proteomics-Based Identification of Anchorless Cell Wall Proteins as Vaccine Candidates against Staphylococcus aureus ^,

Eva Glowalla,^1, Bettina Tosetti,^1, Martin Krönke,^1,2 and Oleg Krut^1,2*

Institute for Medical Microbiology, Immunology and Hygiene, University of Cologne, Cologne, Germany,¹ Center of Molecular Medicine Cologne, Medical Center, University of Cologne, Cologne, Germany²

Received 21 May 2008/ Returned for modification 13 August 2008/ Accepted 3 April 2009

Staphylococcus aureus is an important human pathogen with increasing clinical impact due to the extensive spread of antibiotic-resistant strains. Therefore, development of a protective polyvalent vaccine is of great clinical interest. We employed an intravenous immunoglobulin (IVIG) preparation as a source of antibodies directed against anchorless S. aureus surface proteins for identification of novel vaccine candidates. In order to identify such proteins, subtractive proteome analysis (SUPRA) of S. aureus anchorless cell wall proteins was performed. Proteins reacting with IVIG but not with IVIG depleted of S. aureus-specific opsonizing antibodies were considered vaccine candidates. Nearly 40 proteins were identified by this preselection method using matrix-assisted laser desorption ionization—time of flight analysis. Three of these candidate proteins, enolase (Eno), oxoacyl reductase (Oxo), and hypothetical protein hp2160, were expressed as glutathione S-transferase fusion proteins, purified, and used for enrichment of corresponding immunoglobulin Gs from IVIG by affinity chromatography. Use of affinity-purified anti-Eno, anti-Oxo, and anti-hp2160 antibodies resulted in opsonization, phagocytosis, and killing of S. aureus by human neutrophils. High specific antibody titers were detected in mice immunized with recombinant antigens. In mice challenged with bioluminescent S. aureus, reduced staphylococcal spread was measured by in vivo imaging. The recovery of S. aureus CFU from organs of immunized mice was diminished 10- to 100-fold. Finally, mice immunized with hp2160 displayed statistically significant higher survival rates after lethal challenge with clinically relevant S. aureus strains. Taken together, our data suggest that anchorless cell wall proteins might be promising vaccine candidates and that SUPRA is a valuable tool for their identification.

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* Corresponding author. Mailing address: Institute for Medical Microbiology, Immunology and Hygiene, University of Cologne, Goldenfelsstrasse 19-21, 50935 Cologne, Germany. Phone: 49-(0)221-478 321 03. Fax: 49-(0)221-478 321 34. E-mail: oleg.krut{at}uni-koeln.de

Published ahead of print on 13 April 2009.

Supplemental material for this article may be found at http://iai.asm.org/.

Editor: J. L. Flynn

E.G. and B.T. contributed equally to this work.

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Infection and Immunity, July 2009, p. 2719-2729, Vol. 77, No. 7
0019-9567/09/$08.00+0     doi:10.1128/IAI.00617-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.