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In vivo detection of Staphylococcus aureus endocarditis by targeting pathogen-specific prothrombin activation

Abstract

Coagulase-positive Staphylococcus aureus (S. aureus) is the major causal pathogen of acute endocarditis, a rapidly progressing, destructive infection of the heart valves. Bacterial colonization occurs at sites of endothelial damage, where, together with fibrin and platelets, the bacteria initiate the formation of abnormal growths known as vegetations. Here we report that an engineered analog of prothrombin could be used to detect S. aureus in endocarditic vegetations via noninvasive fluorescence or positron emission tomography (PET) imaging. These prothrombin derivatives bound staphylocoagulase and intercalated into growing bacterial vegetations. We also present evidence for bacterial quorum sensing in the regulation of staphylocoagulase expression by S. aureus. Staphylocoagulase expression was limited to the growing edge of mature vegetations, where it was exposed to the host and co-localized with the imaging probe. When endocarditis was induced with an S. aureus strain with genetic deletion of coagulases, survival of mice improved, highlighting the role of staphylocoagulase as a virulence factor.

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Figure 1: Identification of coagulase-positive endocarditis in a mouse model using a fluorescent prothrombin analog.
Figure 2: The mechanism underlying fluorescent prothrombin localization during vegetation formation.
Figure 3: Noninvasive imaging of coagulase-positive S. aureus endocarditis via AF680-ProT.
Figure 4: Comparison of AF680-ProT signal in FMT-CT studies of endocarditis in mice infected with S. aureus Newman or coagulase-deficient S. aureus Newman.
Figure 5: PET-CT imaging of coagulase-positive S. aureus endocarditis with 64Cu-DTPA-ProT.

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Acknowledgements

We would like to thank Y. Fisher-Jeffes for review of the manuscript and C. Vinegoni, Z. Mueller and J. Sullivan for help with imaging and data analysis. We would also like to thank T. Foster for materials and helpful suggestions. This work was funded in part by grants from the US National Institutes of Health F32-HL094010, K99-HL094533 (P.P.); F32DK083868 (J.P.); R01-HL096576, R01-HL095629 (M.N.); R01-EB006432, T32-CA79443, R24-CA92782, P50-CA86355 (R.W.) and R37-HL071544, R01-HL038779 (P.E.B.) and a Deutsche Herzstiftung grant (F.L.).

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P.P. designed experiments, collected and analyzed the data, and wrote the manuscript. J.-L.F. developed the endocarditis model. J.P. conducted the in situ hybridization experiments. B.M. acquired the PET-CT data and fused images from different modalities. Y.I. and E.A. performed the histology experiments. E.K. synthesized, characterized and optimized the PET reporter. F.L., F.K.S. and M.J.P. labeled leukocytes and analyzed data. P.W. performed optical imaging experiments and analyzed data. P.F.-P. modeled the chelator in the PET version of the probe. O.S. made knockout bacteria. P.P., A.A.M., H.K.K., T.M.H. and P.E.B. designed the imaging probe and performed in vitro characterization. M.N. and R.W. designed experiments, supervised the project, developed in vivo imaging strategies and systems, reviewed, analyzed and discussed data, and wrote the manuscript. All authors edited the manuscript.

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Correspondence to Matthias Nahrendorf or Ralph Weissleder.

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The authors declare no competing financial interests.

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Supplementary Figures 1–7, Supplementary Table 1 and Supplementary Methods (PDF 895 kb)

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Panizzi, P., Nahrendorf, M., Figueiredo, JL. et al. In vivo detection of Staphylococcus aureus endocarditis by targeting pathogen-specific prothrombin activation. Nat Med 17, 1142–1146 (2011). https://doi.org/10.1038/nm.2423

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