Predominant Outer Membrane Antigens of Bartonella henselae

Abstract

A hallmark of Bartonella henselae is persistent bacteremia in cats despite the presence of a vigorous host immune response. To understand better the long-term survival of B. henselae in cats, we examined the feline humoral immune response to B. henselae outer membrane (OM) proteins in naturally and experimentally infected cats. Initially, a panel of sera (n ‫؍‬ 42) collected throughout North America from naturally infected cats was used to probe B. henselae total membranes to detect commonly recognized antigens. Twelve antigens reacted with sera from at least 85% of cats, and five were recognized by sera from all cats. To localize these antigens further, OMs were purified on discontinuous sucrose density step gradients. Each membrane fraction (OM, hybrid or inner membrane [IM]) contained less than 1% of the total malate dehydrogenase activity (soluble marker), indicating very little contamination by cytoplasmic proteins. FtsI, an integral IM cell division protein, was used to identify the low-density fraction ( ‫؍‬ 1.13 g/cm 3 ) as putative IM (<5% of the total FtsI localized to the high-density fraction) while lipopolysaccharide (LPS) and Pap31, a homolog of the Bartonella quintana heme-binding protein A (HbpA), defined the high-density fraction ( ‫؍‬ 1.20 g/cm 3 ) as putative OM. Additionally, little evidence of cross-contamination between the IM and OM was evident by two-dimensional gel electrophoresis. When purified OMs were probed with feline sera, antigenic proteins profiles were very similar to those observed with total membranes, indicating that many, but not all, of the immunoreactive proteins detected in the initial immunoblots were OM components. Interestingly, two-dimensional immunoblots indicated that B. henselae LPS and members of the Hbp family of proteins did not appear to stimulate an humoral response in any infected cats. Seven proteins were recognized by at least 70% of sera tested, but only three were recognized by all sera. Nanospray-tandem mass spectrometry was used to identify OM components, including the immunodominant OM proteins. Recognition of the nonimmunogenic nature of the major OM components, such as LPS, and identification of the predominant immunogens should elucidate the mechanisms by which B. henselae establishes persistent bacteremic infections within cats. Additionally, the common antigens may serve as potential feline vaccine candidates to eliminate the pathogen from its animal reservoir.

Intruders below the Radar: Molecular Pathogenesis of Bartonella spp

“…Interestingly, the antiapoptotic activity of these supernatants was weaker than the net effect of live B. henselae, hence confirming the role of additional factors that are known to act on the same phenotype and may account for an attenuation of GroEL potency during evolution (like the VirB/D4 T4SS). GroEL was further found to localize to the outer membrane not only of B. bacilliformis but also of B. henselae, and it is highly immunogenic in various Bartonella species (43,84,229,357,432). In addition, it was recognized that Bartonella GroEL harbors a C-terminal phenylalanine that is a general characteristic of outer membrane proteins, indicating that GroEL of Bartonella would be adapted to exposure on the bacterial surface (69,409).…”

“…In contrast to IalA and CtpA, IalB is an outer membrane protein based on its signal sequence and findings in B. henselae (84), thus seeming to be the natural candidate for the host-interacting component of the invasion-associated locus. Initial results that IalB of B. bacilliformis was located in the inner membrane were surprising for the authors of the study as well and could have been caused by suboptimal growth conditions (100) (31,36,118,136).…”

Proteomic and Immunoblot Analyses of Bartonella quintana Total Membrane Proteins Identify Antigens Recognized by Sera from Infected Patients

“…Culture-based diagnosis of Bartonella infection is even more difficult and time-consuming (21,24), and molecular biology techniques have little practical application outside the research lab. In most of the immunoscreens for diagnostic antigens in Bartonella workers have used pooled sera from a small number of patients without culture-proven infections or sera collected from experimentally infected small animals (8,13).…”

Bartonella henselae exists as a mosaic of different genetic variants in the infected host

“…Infected cats develop a relapsing bacteraemia that may persist for up to 2 years (Breitschwerdt & Kordick, 2000). Previous investigations have shown that B. henselae induces specific humoral and cell-mediated immune responses in naturally or experimentally infected hosts, including cats, humans and mice (Abbott et al, 1997;Arvand et al, 1998aArvand et al, , 2001aChenoweth et al, 2004;Guptill et al, 1999;Regnath et al, 1998). The mechanisms involved in the long-term survival of the bacteria despite vigorous host immune responses are not entirely understood.…”