Binding of Bartonella henselae to extracellular molecules: Identification of potential adhesins
Introduction
Bartonella henselae, an opportunistic and emerging pathogen, carried by cats, is the etiologic agent of cat-scratch disease (CSD), a persistent necrotizing inflammation of the lymph nodes that affects immunocompetent humans [1], [2], [3]. B. henselae infection in AIDS patients, or other immune-compromised hosts, causes severe and potentially fatal infections such as bacillary angiomatosis (BA), peliosis hepatitis (hepatis) and bacteremia [1], [4], [5]. However, despite its emerging status, little is known about the molecules involved in the bacterium adherence to and invasion of host cells.
Initiation of B. henselae infection requires a wound (trauma) to the skin, suggesting involvement of extracellular matrix (ECM) molecules. Bacterial adherence, a critical first step in the infection process, involves surface components (adhesins) that recognize and bind the host cell surface and ECM molecules including fibronectin (Fn), collagen (Cn), and heparin sulfate-containing proteoglycans. Fn in particular, binds specifically to several ECM molecules including heparin (Hep) and is reported as the main mediator for bacterial adherence [6], [7], [8], [9], [10]. Recent studies show that for many bacterial pathogens, the interaction with ECM molecules is a critical step in bacterial adherence to and invasion of host cells, particularly in wound-associated infections [11], [12], [13], [14].
We characterized in this study, the binding activity of B. henselae to ECM molecules in general and to Fn in particular. Ligand blotting, and binding assays using exogenous Fn, Fn fragments, rabbit anti-Fn and anti-B. henselae antibodies revealed that B. henselae has surface proteins that interact with both immobilized and soluble Fn as well as with host cells and host cells Fn. B. henselae bound specifically the Hep- and gelatin-binding domains of Fn. Glycosaminoglycans (GAGs) such as Hep and dextran sulfate (DxS-500) also may participate in the bacterium interactions with Fn and vitronectin (Vn).
Section snippets
B. henselae binds to immobilized and soluble ECM proteins
Fig. 1, Fig. 2 shows that B. henselae binds significantly (P<0.01) to Fn, Cn-type IX and X. By comparison, the levels of binding to Cn-type IV and laminin were significantly lower (P<0.01) but greater than that for BSA control. No binding was observed for the negative control fetuin. As expected, the positive control Staphylococcus aureus strain Phillips known to bind to Cn-type I, II and III, bound to all proteins tested with the exception of BSA and fetuin controls. The isogenic S. aureus
Discussion
Bacteria display several distinct and alternative adhesins for attachment to host cells. These adhesins are for the most part expressed under different environmental and host conditions or even at different host surfaces. Furthermore, some of these adhesins may be multifaceted or may function in distinct kinetics steps to promote tighter binding of bacteria to host cells. Hence, several adhesins acting collectively may define where a pathogen will adhere, colonize and begin to cause an
Bacteria, culture conditions and reagents
B. henselae genome sequencing B. henselae Houston-1 strain (ATCC # 49882) was grown on tryptic soy plates supplemented with 5% sheep blood in a humidified atmosphere at 37 °C under 5% CO2. E. coli JM109 (Stratagene, La Jolla CA), S. aureus Phillips, and its isogenic collagen adhesin-deficient mutant strain (Can-mutant PH100) (both generous gifts from Dr. Patti; Inhibitex Inc.), were grown on Luria-Bertani (LB) broth (supplemented with gentamicin (10 μg/ml for PH100 only)). The intact human plasma
Acknowledgments
We thank Dr. Joseph Patti (Inhibitex Inc.) for supplying the S. aureus strains and Mrs. Betty Handlin for helping with the figures. This work was partially funded by grant from the Noble Foundation, Ardmore, Oklahoma.
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