Session Type: 30min ePoster Review
Session Title: 30min ePoster Review
Authors(s): D.J. Vaca (1), A. Thibau (1), M.S. Leisegang (2), J. Malmström (3), L. Happonen (3), V.A.J. Kempf (1)
Authors Affiliations(s): (1) Institute for Medical Microbiology and Infection Control, University Hospital, Goethe University Frankfurt, Germany, (2) Institute for Cardiovascular Physiology, Goethe University Frankfurt, Germany, (3) Division of Infection Medicine, Lund University, Sweden
Background:
Bartonella henselae is a facultative intracellular Gram-negative bacterium responsible for cat scratch disease and vascular proliferations (bacillary angiomatosis) in humans. The trimeric autotransporter adhesin (TAA) Bartonella adhesin A (BadA) is a major pathogenicity factor of B. henselae, mediating bacterial adherence to endothelial cells (ECs) presumably via binding to extracellular matrix (ECM) proteins. In this research, we aimed to study and describe the interaction between BadA and fibronectin (FN) in the bacteria-host cell adhesion process.
Methods:FN knockout ECs were generated (LentiCRISPR) and exposed to B. henselae strains (wild type and BadA deficient) via in vitro infection models. Using an immunosorbent assay, B. henselae strains were exposed to proteolytic FN fragments to define the BadA binding sites in FN. As a proof of concept of the importance of the specific FN regions for BadA binding, inhibition assays were performed using bacteria and heparin in competition for FN binding. The specific BadA-FN interaction was further analysed using mass spectrometry of cross-linked peptides (XL-MS).
Results:The importance of BadA and FN interaction in the bacteria-host cell adhesion process was confirmed after the observation of reduced bacterial binding to ECs when using BadA or FN deficient conditions an in vitro infection model. Furthermore, binding experiments in an immunosorbent format between B. henselae and FN fragments showed higher BadA affinity to the heparin-binding domains in FN. The importance of these specific FN sites was confirmed by bacterial binding inhibition to FN in competition assays with heparin. Finally, the XL-MS analysis reported the exact protein-protein interactions occurring within the described FN sites.
Conclusions:We demonstrated that BadA and FN interaction plays a significant role in B. henselae adhesion to ECs. Moreover, the two heparin-binding domains of the FN molecule were identified as important BadA binding sites and the exact protein-protein interacting peptides were detected. This new information gives insight into the use of TAA-targeting specific peptides to treat bacterial infections by a new class of antibiotics (“anti-ligands”).
Keyword(s): Bacteria-host adhesion, Trimeric autotransporter adhesins, Extracellular matrix proteinCOI Institutional Grants: Yes
COI Other: This research was supported by the Viral and Bacterial Adhesin Network Training (ViBrANT) Program funded by the European Union’s HORIZON 2020 Research and Innovation Program under the Marie Sklodowska-Curie Grant Agreement No 765042, and by the Robert Koch-Institute, Berlin, Germany (Bartonella consiliary laboratory, 1369-354).
Session Type: 30min ePoster Review
Session Title: 30min ePoster Review
Authors(s): D.J. Vaca (1), A. Thibau (1), M.S. Leisegang (2), J. Malmström (3), L. Happonen (3), V.A.J. Kempf (1)
Authors Affiliations(s): (1) Institute for Medical Microbiology and Infection Control, University Hospital, Goethe University Frankfurt, Germany, (2) Institute for Cardiovascular Physiology, Goethe University Frankfurt, Germany, (3) Division of Infection Medicine, Lund University, Sweden
Background:
Bartonella henselae is a facultative intracellular Gram-negative bacterium responsible for cat scratch disease and vascular proliferations (bacillary angiomatosis) in humans. The trimeric autotransporter adhesin (TAA) Bartonella adhesin A (BadA) is a major pathogenicity factor of B. henselae, mediating bacterial adherence to endothelial cells (ECs) presumably via binding to extracellular matrix (ECM) proteins. In this research, we aimed to study and describe the interaction between BadA and fibronectin (FN) in the bacteria-host cell adhesion process.
Methods:FN knockout ECs were generated (LentiCRISPR) and exposed to B. henselae strains (wild type and BadA deficient) via in vitro infection models. Using an immunosorbent assay, B. henselae strains were exposed to proteolytic FN fragments to define the BadA binding sites in FN. As a proof of concept of the importance of the specific FN regions for BadA binding, inhibition assays were performed using bacteria and heparin in competition for FN binding. The specific BadA-FN interaction was further analysed using mass spectrometry of cross-linked peptides (XL-MS).
Results:The importance of BadA and FN interaction in the bacteria-host cell adhesion process was confirmed after the observation of reduced bacterial binding to ECs when using BadA or FN deficient conditions an in vitro infection model. Furthermore, binding experiments in an immunosorbent format between B. henselae and FN fragments showed higher BadA affinity to the heparin-binding domains in FN. The importance of these specific FN sites was confirmed by bacterial binding inhibition to FN in competition assays with heparin. Finally, the XL-MS analysis reported the exact protein-protein interactions occurring within the described FN sites.
Conclusions:We demonstrated that BadA and FN interaction plays a significant role in B. henselae adhesion to ECs. Moreover, the two heparin-binding domains of the FN molecule were identified as important BadA binding sites and the exact protein-protein interacting peptides were detected. This new information gives insight into the use of TAA-targeting specific peptides to treat bacterial infections by a new class of antibiotics (“anti-ligands”).
Keyword(s): Bacteria-host adhesion, Trimeric autotransporter adhesins, Extracellular matrix proteinCOI Institutional Grants: Yes
COI Other: This research was supported by the Viral and Bacterial Adhesin Network Training (ViBrANT) Program funded by the European Union’s HORIZON 2020 Research and Innovation Program under the Marie Sklodowska-Curie Grant Agreement No 765042, and by the Robert Koch-Institute, Berlin, Germany (Bartonella consiliary laboratory, 1369-354).
University Hospital, Goethe University of Frankfurt
Session Type: 30min ePoster Review
Session Title: 30min ePoster Review
Authors(s): D.J. Vaca (1), A. Thibau (1), M.S. Leisegang (2), J. Malmström (3), L. Happonen (3), V.A.J. Kempf (1)
Authors Affiliations(s): (1) Institute for Medical Microbiology and Infection Control, University Hospital, Goethe University Frankfurt, Germany, (2) Institute for Cardiovascular Physiology, Goethe University Frankfurt, Germany, (3) Division of Infection Medicine, Lund University, Sweden
Background:
Bartonella henselae is a facultative intracellular Gram-negative bacterium responsible for cat scratch disease and vascular proliferations (bacillary angiomatosis) in humans. The trimeric autotransporter adhesin (TAA) Bartonella adhesin A (BadA) is a major pathogenicity factor of B. henselae, mediating bacterial adherence to endothelial cells (ECs) presumably via binding to extracellular matrix (ECM) proteins. In this research, we aimed to study and describe the interaction between BadA and fibronectin (FN) in the bacteria-host cell adhesion process.
Methods:FN knockout ECs were generated (LentiCRISPR) and exposed to B. henselae strains (wild type and BadA deficient) via in vitro infection models. Using an immunosorbent assay, B. henselae strains were exposed to proteolytic FN fragments to define the BadA binding sites in FN. As a proof of concept of the importance of the specific FN regions for BadA binding, inhibition assays were performed using bacteria and heparin in competition for FN binding. The specific BadA-FN interaction was further analysed using mass spectrometry of cross-linked peptides (XL-MS).
Results:The importance of BadA and FN interaction in the bacteria-host cell adhesion process was confirmed after the observation of reduced bacterial binding to ECs when using BadA or FN deficient conditions an in vitro infection model. Furthermore, binding experiments in an immunosorbent format between B. henselae and FN fragments showed higher BadA affinity to the heparin-binding domains in FN. The importance of these specific FN sites was confirmed by bacterial binding inhibition to FN in competition assays with heparin. Finally, the XL-MS analysis reported the exact protein-protein interactions occurring within the described FN sites.
Conclusions:We demonstrated that BadA and FN interaction plays a significant role in B. henselae adhesion to ECs. Moreover, the two heparin-binding domains of the FN molecule were identified as important BadA binding sites and the exact protein-protein interacting peptides were detected. This new information gives insight into the use of TAA-targeting specific peptides to treat bacterial infections by a new class of antibiotics (“anti-ligands”).
Keyword(s): Bacteria-host adhesion, Trimeric autotransporter adhesins, Extracellular matrix proteinCOI Institutional Grants: Yes
COI Other: This research was supported by the Viral and Bacterial Adhesin Network Training (ViBrANT) Program funded by the European Union’s HORIZON 2020 Research and Innovation Program under the Marie Sklodowska-Curie Grant Agreement No 765042, and by the Robert Koch-Institute, Berlin, Germany (Bartonella consiliary laboratory, 1369-354).
Session Type: 30min ePoster Review
Session Title: 30min ePoster Review
Authors(s): D.J. Vaca (1), A. Thibau (1), M.S. Leisegang (2), J. Malmström (3), L. Happonen (3), V.A.J. Kempf (1)
Authors Affiliations(s): (1) Institute for Medical Microbiology and Infection Control, University Hospital, Goethe University Frankfurt, Germany, (2) Institute for Cardiovascular Physiology, Goethe University Frankfurt, Germany, (3) Division of Infection Medicine, Lund University, Sweden
Background:
Bartonella henselae is a facultative intracellular Gram-negative bacterium responsible for cat scratch disease and vascular proliferations (bacillary angiomatosis) in humans. The trimeric autotransporter adhesin (TAA) Bartonella adhesin A (BadA) is a major pathogenicity factor of B. henselae, mediating bacterial adherence to endothelial cells (ECs) presumably via binding to extracellular matrix (ECM) proteins. In this research, we aimed to study and describe the interaction between BadA and fibronectin (FN) in the bacteria-host cell adhesion process.
Methods:FN knockout ECs were generated (LentiCRISPR) and exposed to B. henselae strains (wild type and BadA deficient) via in vitro infection models. Using an immunosorbent assay, B. henselae strains were exposed to proteolytic FN fragments to define the BadA binding sites in FN. As a proof of concept of the importance of the specific FN regions for BadA binding, inhibition assays were performed using bacteria and heparin in competition for FN binding. The specific BadA-FN interaction was further analysed using mass spectrometry of cross-linked peptides (XL-MS).
Results:The importance of BadA and FN interaction in the bacteria-host cell adhesion process was confirmed after the observation of reduced bacterial binding to ECs when using BadA or FN deficient conditions an in vitro infection model. Furthermore, binding experiments in an immunosorbent format between B. henselae and FN fragments showed higher BadA affinity to the heparin-binding domains in FN. The importance of these specific FN sites was confirmed by bacterial binding inhibition to FN in competition assays with heparin. Finally, the XL-MS analysis reported the exact protein-protein interactions occurring within the described FN sites.
Conclusions:We demonstrated that BadA and FN interaction plays a significant role in B. henselae adhesion to ECs. Moreover, the two heparin-binding domains of the FN molecule were identified as important BadA binding sites and the exact protein-protein interacting peptides were detected. This new information gives insight into the use of TAA-targeting specific peptides to treat bacterial infections by a new class of antibiotics (“anti-ligands”).
Keyword(s): Bacteria-host adhesion, Trimeric autotransporter adhesins, Extracellular matrix proteinCOI Institutional Grants: Yes
COI Other: This research was supported by the Viral and Bacterial Adhesin Network Training (ViBrANT) Program funded by the European Union’s HORIZON 2020 Research and Innovation Program under the Marie Sklodowska-Curie Grant Agreement No 765042, and by the Robert Koch-Institute, Berlin, Germany (Bartonella consiliary laboratory, 1369-354).