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Bartonella henselae, B. quintana, and B. bacilliformis: historical pathogens of emerging significance.

Bartonella species were virtually unrecognized as modern pathogens of humans until the last decade. However, identification of Bartonella species as the agents of cat-scratch disease, bacillary angiomatosis, urban trench fever, and possible novel presentations of Carrion's disease has left little doubt of the emerging medical importance of this genus of organisms. The three primary human pathogenic bartonellae, Bartonella bacilliformis (Carrion's disease), B. henselae (cat-scratch disease), and B. quintana (trench fever), present noteworthy comparisons in the epidemiology, natural history, pathology, and host-microbe interaction that this review will briefly explore.

Animals↗

Use of rpoB gene analysis for detection and identification of Bartonella species.

Identification of Bartonella species is of increasing importance as the number of infections in which these bacteria are involved increases. To date, these gram-negative bacilli have been identified by various serological, biochemical, and genotypic methods. However, the development of alternative tools is required, principally to circumvent a major risk of contamination during sample manipulation. The aim of our study was to investigate the possible identification of various Bartonella species by comparison of RNA polymerase beta-subunit gene (rpoB) sequences. This approach has previously been shown to be useful for the identification of members of the family Enterobacteriaceae (C. M. Mollet, M. Drancourt, and D. Raoult, Mol. Microbiol. 26:1005-1011, 1997). Following PCR amplification with specific oligonucleotides, a 825-bp region of the rpoB gene was sequenced from 13 distinct Bartonella strains. Analysis of these sequences allowed selection of three restriction enzymes (ApoI, AluI, and AflIII) useful for discerning the different strains by PCR-restriction fragment length polymorphism (PCR-RFLP) analysis. To confirm the potential value of such an approach for identification of Bartonella, the rpoB PCR was then applied to 94 clinical samples, and the results obtained were identical to those obtained by our reference PCR method. Twenty-four isolates were also adequately identified by PCR-RFLP analysis. In all cases, our results were in accordance with those of the reference method. Moreover, conserved regions of DNA were chosen as suitable primer targets for PCR amplification of a 439-bp fragment which can be easily sequenced.

Animals↗

Serodiagnosis of cat scratch disease: response to Bartonella henselae in children and a review of diagnostic methods.

In this study, sera from 116 children without clinical evidence of cat scratch disease (CSD) and sera from 19 children with CSD were investigated by two different immunofluorescent assays (test A and test B) to evaluate the seroprevalence of Bartonella henselae antibodies in this population. Antibodies against Bartonella henselae were found in noninfected children only in low titers: 13% and 3.5% of serum samples were positive by test A and test B, respectively. IgG titers as determined by test A ranged from 1:64 to 1:256, whereas test B did not yield titers >1:64. Of the 19 children with CSD, 7 (37%) and 5 (26%) had low antibody titers (1:64-1:256) on admission by test A and test B, respectively. In such cases, IgM antibodies against Bartonella henselae and/or a titer increase in the following weeks are required to prove suspected CSD. The results indicate that the seroprevalence of Bartonella henselae antibodies in children without evidence of CSD is low, and that the results may differ depending on the tests being used. Low antibody titers, however, were found not only in healthy children but also among patients with CSD, which could be indicative of the beginning or of the end of an illness. Currently, only few data are available concerning the seroprevalence of antibodies to Bartonella spp. among children. Although children are often affected by CSD, the specificity of most tests has been evaluated by investigating blood donors, who are usually adults. Several different serological tests are used for the diagnosis of CSD, including the immunofluorescence assay, the enzyme-linked immunosorbent assay, and Western blot (WB) analysis. The sensitivities of different IFAs range from 14 to 100%, depending on the antigen used, the cut-off chosen, and the test procedures. The current diagnostic value of different serological tests for diagnosis of CSD is reviewed.

Adolescent↗

In vitro susceptibilities of Rickettsia and Bartonella spp. to 14-hydroxy-clarithromycin as determined by immunofluorescent antibody analysis of infected vero cell monolayers.

The in vitro susceptibilities of Rickettsia akari, Rickettsia conorii, Rickettsia prowazekii, Rickettsia rickettsii, Bartonella elizabethae, Bartonella henselae and Bartonella quintana to different concentrations of clarithromycin, 14-hydroxy-clarithromycin (the primary metabolite of clarithromycin) and tetracycline in Vero cell cultures, were determined by enumeration of immunofluorescently-stained bacilli. The extent of antibiotic-induced inhibition of foci was recorded for each dilution of antibiotic and compared with an antibiotic-negative control. Based upon MIC data, clarithromycin alone is highly active against all three Bartonella spp., R. akari and R. prowazekii, while 14-hydroxy-clarithromycin is active against R. conorii, R. prowazekii and R. rickettsii. Further testing is warranted in animal models and human clinical trials, to examine the activity of both clarithromycin and its primary metabolite and to define further the role of clarithromycin in therapy, particularly of infections caused by obligate intracellular bacteria such as Rickettsia and Bartonella spp.

Animals↗

In vitro susceptibility of Bartonella species to 17 antimicrobial compounds: comparison of Etest and agar dilution.

OBJECTIVES: In vitro susceptibility testing of 31 Bartonella spp. strains including 21 Bartonella henselae isolates was performed for 17 antimicrobial agents (telithromycin, four macrolides, five fluoroquinolones, five aminoglycosides, doxycycline and rifampicin). METHODS: MICs were determined by agar dilution and Etest using chocolate agar containing 5% defibrinated sheep blood as assay medium. Longer incubation periods of 3-5 days in a humid atmosphere with 5% CO(2) were required until bacterial growth became visible and MICs could be read. RESULTS: The ketolide telithromycin was the most active agent exhibiting the lowest MICs. The Bartonella spp. were also highly susceptible to macrolides, particularly clarithromycin, and to doxycycline and rifampicin, with MICs of <or=0.12 mg/L. Gatifloxacin, gemifloxacin and moxifloxacin were the most potent fluoroquinolones, with MICs ranging from 0.06 to 2 mg/L. Netilmicin was the most active agent among the aminoglycosides. Etest MICs correlated well with MICs determined by agar dilution. CONCLUSIONS: Telithromycin, macrolides, doxycycline and rifampicin were the most effective agents against Bartonella spp. Our data confirm that Etest may be a reliable method for determining susceptibility of Bartonella spp.

Agar↗

Clinical impact of persistent Bartonella bacteremia in humans and animals.

Bartonella spp. are emerging vector-borne pathogens that cause persistent, often asymptomatic bacteremia in their natural hosts. As our knowledge progresses, it appears that chronic infection may actually predispose the host to mild, insidious nonspecific manifestations or induce, in selected instances, severe diseases. Persistent asymptomatic bacteremia is most common in animals that serve as the main reservoir for the specific Bartonella. In humans, these organisms are B. bacilliformis and B. quintana. Other Bartonella species, for which humans are not the natural reservoir, tend to cause persistent bacteremia only in immunodeficient individuals. In some of these individuals, endothelial cell proliferation may create lesions such as bacillary angiomatosis or bacillary peliosis. In cats, bacteremia of variable level and continuity may last for years. Some strains of B. henselae may induce clinical manifestations, including fever, mild neurological signs, reproductive disorders, whereas others do not induce clinically obvious disease. Reproductive disorders have also been reported in mice experimentally infected with B. birtlesii. Finally, canids constitute the most interesting naturally occurring animal model for the human disease. Like immunocompetent people, healthy dogs only occasionally demonstrate long-term bacteremia when infected with Bartonella spp. However, some dogs develop severe clinical manifestations, such as endocarditis, and the pathologic spectrum associated with Bartonella spp. infection in domestic dogs is rapidly expanding and resembles the infrequently reported clinical entities observed in humans. In coyotes, persistent bacteremia is more common than in domestic dogs. It will be of interest to determine if coyotes develop clinical or pathological indications of infection.

Animals↗

Differentiation of Bartonella species by a microimmunofluorescence assay, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and Western immunoblotting.

Bartonella species can be differentiated by microimmunofluorescence assay, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and immunoblotting with murine polyclonal antisera to Bartonella henselae, B. quintana, B. elizabethae, and B. bacilliformis. A pairwise comparison on the basis of SDS-PAGE protein profiles demonstrated similarity values for proteins of different Bartonella species ranging from 28.6 to 86.4%. Antigenic relationships revealed by immunoblotting with murine antisera were equivalent to those of proteins observed by SDS-PAGE. A dendrogram obtained on the basis of protein bands of SDS-polyacrylamide gels showed that Bartonella species could be divided into three groups. B. bacilliformis was distinct from all other Bartonella species; B. grahamii, B. taylorii, B. doshiae, and B. vinsonii formed a cluster, as did B. henselae, B. quintana, B. elizabethae, and B. clarridgeiae. These relationships were consistent with those revealed by parsimony trees derived from 16S rRNA and gltA gene sequencing. SDS-PAGE analysis showed that 120-, 104-, 85-, 71-, 54-, 47-, 40-, 33-, 30-, and 19-kDa proteins were present in all species, with the 54-kDa protein being the most dominant. Proteins with a molecular mass of less than 54 kDa allow the differentiation of species and are a possible target for future species-specific antibodies and antigens.

Bacterial Typing Techniques↗

Bartonella (Rochalimaea) quintana infections.

Bartonella (formerly Rochalimaea) quintana is the etiological agent of trench fever, a disease extensively reported during the World Wars. Recent molecular biology approaches have allowed dramatic extension of the spectrum of Bartonella infections. B. quintana is now also recognized as an etiological agent of fever and bacteremia, endocarditis, bacillary angiomatosis, and chronic lymphadenopathy. Human immunodeficiency virus-infected patients and/or homeless people are the most vulnerable to infection. Poverty and louse infestation were the main epidemiological factors associated with B. quintana infections during wartime. Although poverty and chronic alcoholism have been associated with modern cases of trench fever and bacteremia due to B. quintana in Europe and the United States, vectors for B. quintana have not been clearly identified and B. quintana has not been isolated from modern-day lice. Microscopic bacillary angiomatosis lesions are characterized by tumor-like capillary lobules, with proliferating endothelial cells. In vitro experiments have shown that B. quintana survives within endothelial cells and stimulates cell proliferation. These observations, together with the finding that lesions may regress when antibiotic therapy is administered, strongly suggest that B. quintana itself stimulates angiogenesis. Bartonella infections are characterized by a high frequency of relapses after brief courses of antibiotic therapy. It is to be noted that in vitro, although Bartonella species are highly susceptible to antibiotics, only the aminoglycosides have proved to be bactericidal. However, the most effective antibiotic regimen for Bartonella infections remains to be established.

Angiomatosis, Bacillary↗

Adhesion to and invasion of cultured human cells by Bartonella bacilliformis.

Bartonella bacilliformis was tested for its ability to adhere to and invade tissue culture cell monolayers. The parasite was able to efficiently bind and penetrate human dermal fibroblasts, human laryngeal epithelium, and human umbilical vein endothelial cells. Exposure of the organism to immune serum prepared against a crude Bartonella extract containing cell wall and membranous material resulted in decreased ability of the parasite to invade host cells. There was also an overall reduction in the invasiveness of bartonellae and total host cell association when human laryngeal epithelial cells and human umbilical vein endothelial cells were preexposed to cytochalasin D, indicating an active involvement of host cells in the uptake of bartonellae. Transmission electron microscopy revealed the presence of bartonellae inside and outside intracellular vacuoles. These data suggest that a surface-associated factor is involved in the invasion process and that internalization of the parasite by host cells involves a microfilament-dependent process similar to phagocytosis.

Adhesins, Bacterial↗

Prevalence of Bartonella species in domestic cats in The Netherlands.

Cats have been shown to provide the only known reservoir of Bartonella henselae, the causative agent of cat scratch disease. To determine the prevalence of Bartonella bacteremia and antibodies in Dutch cats, blood samples from 113 cats from shelters (sheltered cats), 50 pet cats, and 25 specific-pathogen-free (SPF) cats were analyzed. Culture and subsequent PCR-restriction fragment length polymorphism (RFLP) analysis of the 16S-23S rRNA intergenic region and 16S rRNA gene PCR-hybridization assays revealed a prevalence of Bartonella bacteremia in 22% of the sheltered cats and showed no bacteremia in the SPF cats. Three spacer RFLP types were found: types A, B, and G, with type B being predominant over types A and G. An important finding was the existence of mixtures of different Bartonella species. Bartonella DNA was detected in 7 of 27 DNA extracts from fleas combed from the sheltered cats (26%). Seropositivity was 50% for sheltered cats and 56% for pet cats, as determined by a B. henselae enzyme-linked immunoassay.

Animals↗

Pericardial effusion in a homeless man due to Bartonella quintana.

Bartonella quintana may cause trench fever, endocarditis, bacillary angiomatosis, and chronic bacteremia, and a reemergence among homeless populations in cities has been noted. Pericarditis from Rickettsia conorii and Coxiella burnetii infection has been described, but there have been no reports of pericarditis due to Bartonella spp. We report a case of pericardial effusion due to Bartonella quintana in a homeless man, diagnosed on the basis of PCR detection of Bartonella quintana in a pericardial biopsy sample and a fourfold rise in antibody titers. The patient recovered within 2 weeks with antibiotics active against bartonellae.

Adult↗

Bartonella bacilliformis GroEL: effect on growth of human vascular endothelial cells in infected cocultures.

Bartonella are the only bacteria known to induce angioproliferative lesions of the human vasculature and liver during infection. Previous work from our lab suggests that GroEL participates in the mitogenic response observed in HUVEC cultures supplemented with the soluble fraction of Bartonella bacilliformis. Work in this study shows that exposure to high concentrations of the fraction is actually cytotoxic for HUVECs. To analyze this phenomenon, live B. bacilliformis-HUVEC cocultures were employed to study the effect of excess bacterial GroEL on the host cell during active infection. Four B. bacilliformis strains were generated to produce varying levels of GroEL. HUVEC cocultures with LSS100, a strain that synthesizes markedly greater quantities of GroEL relative to others, significantly accelerates apoptosis of the cocultured HUVECs relative to other strains. Acceleration of apoptosis can be inhibited by Z-VAD-FMK, a pan-caspase inhibitor. Time course data show that, at 18 h of infection, both LSS100 and control strains significantly inhibit spontaneous apoptosis of cocultured HUVECs, as previously reported for other Bartonella species. However, by 48 h, LSS100 significantly increases apoptosis of the host cell. We hypothesize that intracellular Bartonella GroEL functions as an Hsp60 analogue, a eukaryotic orthologue known to accelerate pro-caspase 3 activation by enhancing its vulnerability to upstream activator caspases. These data suggest another strategy whereby Bartonella may regulate host cell growth.

Apoptosis↗

Pathogenic mechanisms of Bartonella quintana.

Bartonella quintana is an epi- and intracellular gram-negative rod responsible for both acute and chronic clinical manifestations. We review the literature about pathogenic mechanisms of B. quintana and discuss our data. Our efforts to clarify Bartonella quintana pathogenesis run on two parallel tracks. The first one concerns interactions between Bartonella quintana and endothelial cells by evaluation and modulation of apoptosis, signal transduction pathways and inflammation. The second one concerns some biological activities of Bartonella quintana endotoxin on human whole blood and endothelium. The elucidation of the mechanisms regulating the inflammatory/proliferative pattern of chronic clinical manifestations of Bartonella quintana infections may offer a contribution for addressing the pathogenesis of intracellular bacterial persistence.

Apoptosis↗

Detection of Bartonella species from ticks, mites and small mammals in Korea.

We investigated the prevalence of Bartonella infections in ticks, mites and small mammals (rodents, insectivores and weasels) collected during 2001 through 2004, from various military installations and training sites in Korea, using PCR and sequence analysis of 16S rRNA, 23S rRNA and groEL heat shock protein genes. The prevalence of Bartonella spp. was 5.2% (n = 1,305 sample pools) in ticks, 19.1% (n = 21) in mesostigmatid mites and 13.7% (n = 424 individuals) in small mammals. The prevalence within the family Ixodidae was, 4.4% (n = 1,173) in Haemaphysalis longicornis (scrub tick), 2.7% (n = 74) in H. flava, 5.0% (n = 20) in Ixodes nipponensis, 11.1% (n = 9) in I. turdus, 33.3% (n = 3) in I. persulcatus and 42.3% (n = 26) in Ixodes spp. ticks. In rodents, the prevalence rate was, 6.7% (n = 373) in Apodemus agrarius (striped field mouse) and 11.1% (n = 9) in Eothenomys regulus (Korean red-backed vole) and in an insectivore,Crocidura lasiura, 12.1% (n = 33). Neither of the two weasels were positive for Bartonella spp. Phylogenetic analysis based on amino acid sequence of a portion of the groEL gene amplified from one A. agrarius spleen was identical to B. elizabethae species. We demonstrated the presence of Bartonella DNA in H. longicornis, H. flava and I. nipponensis ticks, indicating that these ticks should be added to the growing list of potential tick vectors and warrants further detailed investigations to disclose their possible roles in Bartonella infection cycles.

Animals↗

Prolonged Bartonella bacteremia in cats associated with cat-scratch disease patients.

Recent evidence supports a causal relationship between Bartonella (Rochalimaea) henselae, cat-scratch disease (CSD), and bacillary angiomatosis. Cats appear to be the primary reservoir. Blood from 19 cats owned by 14 patients diagnosed with CSD was cultured. Blood samples from cats owned by veterinary students (n = 25) having no association with CSD or bacillary angiomatosis were cultured as controls. Eighty-nine percent (17 of 19) of cats associated with CSD patients and 28% (7 of 25) of controls were bacteremic with Bartonella species (chi-square = 16.47; P < 0.001). Twenty-three isolates were characterized as B. henselae, while one isolate from the cat of a CSD patient appeared to be a new Bartonella species. Thirteen cats remained culture positive during the ensuing 12-month period. Our results support the conclusion that B. henselae is the predominant species involved in CSD and is transmitted by cats. The incidence of Bartonella bacteremia in control cats suggests that B. henselae bacteremia is prevalent among the domestic cat population in the United States.

Adolescent↗

Bartonella (Rochalimaea) infections: beyond cat scratch.

Five species of Bartonella have been found to infect humans, henselae, quintana, elizabethae, bacilliformis, and vinsonii. The most common of these in North America are Bartonella quintana and Bartonella henselae, the agents of trench fever, bacillary angiomatosis, and parenchymal peliosis, and in the case of B. henselae cat-scratch disease. B. bacilliformis is endemic in Peru and Ecuador, where it causes oroya fever or Carrion's disease. New methods of diagnosing Bartonella infections have resulted in increased recognition of the many manifestations of these infections. Early recognition is crucial, as these are potentially fatal opportunistic infections that usually respond rapidly to appropriate antimicrobial therapy.

AIDS-Related Opportunistic Infections↗

Is pyogenic granuloma associated with Bartonella infection?

Lobular capillary hemangioma and bacillary angiomatosis due to Bartonella infection share several clinical and histopathologic characteristics. We sought to determine whether lobular capillary hemangioma is caused by the same agent as bacillary angiomatosis. Forty-five pathology specimens with a histologic diagnosis of lobular capillary hemangioma obtained from patients with the same clinical diagnosis were tested by immunohistochemistry and polymerase chain reaction for the presence of DNA elements of Bartonella spp. None of the 45 lobular capillary hemangioma specimens tested positive for Bartonella spp. We conclude that lobular capillary hemangioma is not associated with Bartonella spp infection. Further research is required to determine the etiologic agent.

Angiomatosis, Bacillary↗

Detection and characterization of feline Bartonella henselae in the Czech Republic.

The aims of the study were to characterize isolates of Bartonella henselae and to determine the prevalence of bacteremic domestic cats in urban and suburban parts of Prague, Czech Republic. Five (18%) gram-negative fastidious bacterial single-cat isolates were recovered from 27 hemocultures incubated without previous freezing. Four of these isolates originated from flea infested stray cats (n=6) and one from a shelter cat without any ectoparasites (n=21). None of the 34 previously frozen specimens from flea free pet cats yielded any bacteria. All five isolates were catalase and oxidase negative. Their enzymatic activity, RFLP profile of citrate synthetase gene (gltA) and DNA-DNA hybridization results were typical of B. henselae. According to their PvuII and BglI ribotypes the isolates could be allocated to two homogeneous groups. Ribotype HindIII and RFLP of 16S-23S rRNA spacer region analysis gave unique profiles different from those of Bartonella quintana, Bartonella elizabethae and Bartonella clarridgeiae. The 16S rRNA type-specific amplification revealed an identical profile typical of B. henselae genotype II for all the cat isolates studied. Pulsed-field gel electrophoresis (PFGE) assigned a different profile to each of the isolates studied. Determination of the enzymatic activity, RFLP of gltA gene, RFLP of 16S-23S rRNA spacer region, and HindIII ribotype could be efficient tools for identification of B. henselae isolates. Ribotyping (PvuII, BglI), 16S rRNA typing and PFGE may be useful methods to prospect ecology and epidemiology of the agent.

Animals↗