Characterization of Bartonella bacilliformis flagella and effect of antiflagellin antibodies on invasion of human erythrocytes

Abstract

Bartonella bacilliformis is the etiologic agent of Oroya fever in humans. Flagellum-mediated motility has been postulated as a major virulence factor for invasion of host cells. To address this hypothesis, we purified and characterized flagella from strain KC584 and then assessed their role in human erythrocyte association and invasion. Electron microscopy of the flagellar preparation showed a high concentration of filaments with a mean wavelength of 800 nm. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoblot analysis, and KBr density gradient centrifugation indicated that the flagellar filament is composed of a polypeptide of 42 kDa. The flageHlin is partially (ca. 50%) resistant to treatment with trypsin. The first 17 amino acid residues of the N terminus of the mature flagellin protein are GAAILTNDNAMDALQDL and show approximately 46% sequence identity to the residues of the N termini of two Caulobacter crescentus flagellin proteins. A monospecific polyclonal antibody to the flagellin protein was generated, and its specificity was verified by both immunoblot and immunogold analyses. Human erythrocyte invasion assays performed with bartonellae 4962 on July 16, 2020 by guest http://iai.asm.org/ Downloaded from

The Afipia toolbox and its use to isolate flagellar mutants

“…4f and h). A polyclonal antibody to flagella from B. bacilliformis (Scherer et al, 1993), which is closely related to A. felis, did not recognize Afipia flagellae, indicating substantial diversity in their flagella amino acid sequences. DNA was isolated and sequenced outward using primer Kan-2 FP01 for the transposon-flanking regions.…”

“…The antibodies CSD11 directed against the flagellin of Afipia (courtesy of Mr William Bibb, formerly Division of Bacterial and Mycotic Diseases, Centers for Disease Control and Prevention, Atlanta, GA) and a rabbit antiserum raised to a mixture of formaldehyde-fixed A. felis, Afipia broomeae, Afipia clevelandensis, Afipia genospecies 1, 2 and 3 were used in this study. Polyclonal anti-Bartonella bacilliformis flagellae was from Dr Michael Minnick, University of Montana, Missoula (Scherer et al, 1993).…”

Persistence ofBartonellaspp. stealth pathogens: from subclinical infections to vasoproliferative tumor formation

“…Some species of Bartonella spp. such as Bartonella schoenbuchensis , B. bacilliformis (Benson et al, 1986;Scherer et al, 1993), and Bartonella clarridgeiae (Kordick et al, 1997;Sander et al, 2000b) express flagella, polymeric rod-like structures that are important for bacterial motility. However, these structures may extend several micrometers from the bacterial outer membrane and thereby serve as ideal pathogen-associated molecular patterns for PRRs.…”

Involvement of host cell tyrosine phosphorylation in the invasion of HEp-2 cells by Bartonella bacilliformis

“…Recently, a two-gene locus was characterized from B. bacilliformis that is associated with the ability to invade human erythrocytes [3]. Further, the polar £agella of Bartonella [4] and deformin [5] have also been implicated in the invasion of erythrocytes.…”