Identification and functional analysis of invasion associated locus B (IalB) in Bartonella species.

Bartonellosis is caused by the genus Bartonella. Bartonella is widely distributed in the ruminants, cats, dogs, rodents and other mammals including humans. At least 13 species or subspecies of Bartonella are zoonotic, and each species appears to be highly adapted to one or a limited number of reservoir animals in which it is asymptomatic, while it can be transmitted to humans in which a variety of clinical manifestations can be caused. It was reported that Bartonella henselae infection rate among domestic cats was high in nature, making it one of the leading, important, and easily neglected zoonotic diseases. The aims of this study were to identify the expression, localization, immunogenicity and functional mechanism of Bartonella virulence factor IalB. We found that recombinant IalB protein could react with the serum from infected reservoir hosts and anti-IalB polyclonal antibodies could react with different Bartonella species by western blot analysis. According to these results, we proposed that IalB protein and anti-IalB antibodies would be good candidates for diagnosis of Bartonella infection by antigen-based anti-IalB antibodies or antibody-based IalB antigen capture immunoassay, respectively. We also found that IalB had a putative 22-amino-acid signal sequence and little IalB was localized to the outer membrane of Bartonella birtlesii by electron microscopy assay. Incubation with anti-IalB polyclonal antibodies resulted in inhibition of the invasion of mouse erythrocytes by B. birtlesii. According to these results, we propose that IalB could be a secreted protein that facilitates Bartonella entry into erythrocytes. In conclusion, these results improve our understanding of IalB as a candidate for immunodiagnosis and how IalB affects Bartonella-erythrocyte entry.

Partial disruption of translational and posttranslational machinery reshapes growth rates of Bartonella birtlesii.

UNLABELLED: Specialization of bacteria in a new niche is associated with genome repertoire changes, and speciation in bacterial specialists is associated with genome reduction. Here, we tested a signature-tagged mutant library of 3,456 Bartonella birtlesii clones to detect mutants that could grow rapidly in vitro. Overall, we found 124 mutants that grew faster than the parental wild-type strain in vitro. We sequenced the genomes of the four mutants with the most rapid growth (formed visible colonies in only 1 to 2 days compared with 5 days for the wild type) and compared them to the parental isolate genome. We found that the number of disrupted genes associated with translation in the 124 rapid-growth clones was significantly higher than the number of genes involved in translation in the full genome (P < 10(-6)). Analysis of transposon integration in the genome of the four most rapidly growing clones revealed that one clone lacked one of the two wild-type RNA ribosomal operons. Finally, one of the four clones did not induce bacteremia in our mouse model, whereas infection with the other three resulted in a significantly lower bacterial count in blood than that with the wild-type strain. IMPORTANCE: Here, we show that specialization in a specific niche could be caused by the disruption of critical genes. Most of these genes were involved in translation, and we show that evolution of obligate parasitism bacteria was specifically associated with disruption of translation system-encoding genes.

Candidatus Neoehrlichia mikurensis in bank voles, France.

To further assess the geographic occurrence, possible vectors, and prevalence of Candidatus Neoehrlichia mikurensis, we analyzed spleen tissues from 276 voles trapped close to human settlements in France; 5 were infected with the organism. Sequencing showed the isolates carried the same genotype as the bacteria that caused disease in humans and animals elsewhere in Europe.

Identification of Bartonella Trw host-specific receptor on erythrocytes.

Each Bartonella species appears to be highly adapted to one or a limited number of reservoir hosts, in which it establishes long-lasting intraerythrocytic bacteremia as the hallmark of infection. Recently, we identified Trw as the bacterial system involved in recognition of erythrocytes according to their animal origin. The T4SS Trw is characterized by a multiprotein complex that spans the inner and outer bacterial membranes, and possesses a hypothetical pilus structure. TrwJ, I, H and trwL are present in variable copy numbers in different species and the multiple copies of trwL and trwJ in the Bartonella trw locus are considered to encode variant forms of surface-exposed pilus components. We therefore aimed to identify which of the candidate Trw pilus components were located on the bacterial surface and involved in adhesion to erythrocytes, together with their erythrocytic receptor. Using different technologies (electron microscopy, phage display, invasion inhibition assay, far western blot), we found that only TrwJ1 and TrwJ2 were expressed and localized at the cell surface of B. birtlesii and had the ability to bind to mouse erythrocytes, and that their receptor was band3, one of the major outer-membrane glycoproteins of erythrocytes, (anion exchanger). According to these results, we propose that the interaction between TrwJ1, TrwJ2 and band 3 leads to the critical host-specific adherence of Bartonella to its host cells, erythrocytes.

Strategies of exploitation of mammalian reservoirs by Bartonella species.

Numerous mammal species, including domestic and wild animals such as ruminants, dogs, cats and rodents, as well as humans, serve as reservoir hosts for various Bartonella species. Some of those species that exploit non-human mammals as reservoir hosts have zoonotic potential. Our understanding of interactions between bartonellae and reservoir hosts has been greatly improved by the development of animal models for infection and the use of molecular tools allowing large scale mutagenesis of Bartonella species. By reviewing and combining the results of these and other approaches we can obtain a comprehensive insight into the molecular interactions that underlie the exploitation of reservoir hosts by Bartonella species, particularly the well-studied interactions with vascular endothelial cells and erythrocytes.

Role of the spleen in Bartonella spp. infection.

Bartonella spp. are intra-erythrocytic pathogens of mammals. In this study, we investigated the role of the spleen, and other tissue and organs in Bartonella infection. Using an in vivo model of mice infection by Bartonella birtlesii, we detected accumulation of bacteria in the spleen, with transient infection of the liver, but failed to detect any bacteria in brain or lymph nodes. We then compared bacteraemia in normal Balb/C mice and in splenectomized mice. Bacteraemia in splenectomized mice was 10-fold higher than in normal mice and lasted 2 weeks longer. In conclusion, the spleen seems to retain and filter infected erythrocytes rather than to be a sanctuary for chronic Bartonella infection.

Vector competence of the tick Ixodes ricinus for transmission of Bartonella birtlesii.

Bartonella spp. are facultative intracellular vector-borne bacteria associated with several emerging diseases in humans and animals all over the world. The potential for involvement of ticks in transmission of Bartonella spp. has been heartily debated for many years. However, most of the data supporting bartonellae transmission by ticks come from molecular and serological epidemiological surveys in humans and animals providing only indirect evidences without a direct proof of tick vector competence for transmission of bartonellae. We used a murine model to assess the vector competence of Ixodes ricinus for Bartonella birtlesii. Larval and nymphal I. ricinus were fed on a B. birtlesii-infected mouse. The nymphs successfully transmitted B. birtlesii to naïve mice as bacteria were recovered from both the mouse blood and liver at seven and 16 days after tick bites. The female adults successfully emitted the bacteria into uninfected blood after three or more days of tick attachment, when fed via membrane feeding system. Histochemical staining showed the presence of bacteria in salivary glands and muscle tissues of partially engorged adult ticks, which had molted from the infected nymphs. These results confirm the vector competence of I. ricinus for B. birtlesii and represent the first in vivo demonstration of a Bartonella sp. transmission by ticks. Consequently, bartonelloses should be now included in the differential diagnosis for patients exposed to tick bites.

The Trw type IV secretion system of Bartonella mediates host-specific adhesion to erythrocytes.

Bacterial pathogens typically infect only a limited range of hosts; however, the genetic mechanisms governing host-specificity are poorly understood. The alpha-proteobacterial genus Bartonella comprises 21 species that cause host-specific intraerythrocytic bacteremia as hallmark of infection in their respective mammalian reservoirs, including the human-specific pathogens Bartonella quintana and Bartonella bacilliformis that cause trench fever and Oroya fever, respectively. Here, we have identified bacterial factors that mediate host-specific erythrocyte colonization in the mammalian reservoirs. Using mouse-specific Bartonella birtlesii, human-specific Bartonella quintana, cat-specific Bartonella henselae and rat-specific Bartonella tribocorum, we established in vitro adhesion and invasion assays with isolated erythrocytes that fully reproduce the host-specificity of erythrocyte infection as observed in vivo. By signature-tagged mutagenesis of B. birtlesii and mutant selection in a mouse infection model we identified mutants impaired in establishing intraerythrocytic bacteremia. Among 45 abacteremic mutants, five failed to adhere to and invade mouse erythrocytes in vitro. The corresponding genes encode components of the type IV secretion system (T4SS) Trw, demonstrating that this virulence factor laterally acquired by the Bartonella lineage is directly involved in adherence to erythrocytes. Strikingly, ectopic expression of Trw of rat-specific B. tribocorum in cat-specific B. henselae or human-specific B. quintana expanded their host range for erythrocyte infection to rat, demonstrating that Trw mediates host-specific erythrocyte infection. A molecular evolutionary analysis of the trw locus further indicated that the variable, surface-located TrwL and TrwJ might represent the T4SS components that determine host-specificity of erythrocyte parasitism. In conclusion, we show that the laterally acquired Trw T4SS diversified in the Bartonella lineage to facilitate host-restricted adhesion to erythrocytes in a wide range of mammals.

Insights in Bartonella host specificity.

The genus Bartonella comprises a unique group of emerging gram-negative, intracellular bacteria that can cause a long-lasting intraerythrocytic bacteremia in their reservoir hosts. In recent years, the widespread occurrence and diversity of these bacteria has been increasingly recognized. This has resulted in a dramatic expansion of the genus Bartonella to 24 currently described species or subspecies, among which at least half have been associated with human disease. Bartonella infections have been observed in virtually all species examined, extending from humans to carnivores, ungulates, rodents, lagomorphs, insectivores, and bats. Adaptation by Bartonellae to such a diverse range of mammals has resulted in host specificity, and all validated Bartonella species described to date are capable of parasitizing only a limited number of animal species. In this review, the possible mechanisms explaining the specificity of each Bartonella species for its reservoir host are discussed.

Transmission of Bartonella henselae by Ixodes ricinus.

Bartonella spp. are facultative intracellular bacteria associated with several emerging diseases in humans and animals. B. henselae causes cat-scratch disease and is increasingly associated with several other syndromes, particularly ocular infections and endocarditis. Cats are the main reservoir for B. henselae and the bacteria are transmitted to cats by cat fleas. However, new potential vectors are suspected of transmitting B. henselae, in particular, Ixodes ricinus, the most abundant ixodid tick that bites humans in western Europe. We used a membrane-feeding technique to infect I. ricinus with B. henselae and demonstrate transmission of B. henselae within I. ricinus across developmental stages, migration or multiplication of B. henselae in salivary glands after a second meal, and transmission of viable and infective B. henselae from ticks to blood. These results provide evidence that I. ricinus is a competent vector for B. henselae.

Cloning and analysis of a cDNA encoding a putative serine protease comprising two trypsin-like domains of Trichinella spiralis.

The cDNA encoding a putative serine protease, TsSerP, was cloned by degenerative polymerase chain reaction and screening of the cDNA library from Trichinella spiralis adult-newborn larvae stage. Sequence analysis revealed the presence of two trypsin-like serine protease domains flanking a hydrophilic domain, with the catalytic triad residue histidine in the alpha domain substituted by an arginine residue. Southern blots indicated that this was a single copy gene in the parasite genome. Northern blots demonstrated a single 2.3-kb transcript during the muscle larvae and adult stages of T. spiralis. The recombinant protein from the TsSerP beta domain (betaSerP) was produced but not recognised by T. spiralis-infected swine serum. An anti-betaSerP polyclonal serum detected a 69-kDa polypeptide in the soluble antigens of T. spiralis muscle larvae. Immunolocalisation analysis located TsSerP on the inner layer of the cuticle and oesophagus of the parasite, suggesting a potential role in its moulting and/or digestive functions.