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Bartonella henselae is a fastidious intraerythrocytic, gram-negative bacteria that causes cat scratch disease in humans. Ixodes ricinus has been confirmed to be a competent vector of B. henselae, and some indirect evidences from clinical cases and epidemiological studies also suggested that some other tick species, including Rhipicephalus sanguineus, may transmit the bacteria. B. henselae has been detected in R. sanguineus but no experimental investigations have been performed to evaluate the vector competency of this tick species regarding B. henselae transmission. To this end, this work aimed to assess the transstadial transmission of B. henselae between larvae and nymphs of R. sanguineus as well as transmission by nymphs infected at the larval stage. Four hundred B. henselae negative larvae were fed with B. henselae-infected blood by using an artificial membrane feeding system. After five days of feeding, B. henselae was detected by PCR in 57.1% (8/14) of engorged larval pools, 66.7% (4/6) of semi-engorged larval pools, and 66.7% (2/3) of larval feces pools. After molting, B. henselae DNA was also detected in 10% (1/10) of nymph pools, but not in tick feces. After a pre-fed step of nymphs infected at the larval stage on non-infected blood meal, B. henselae was detected by PCR in blood sample from the feeder, but no Bartonella colonies could be obtained from culture. These findings showed that B. henselae could be transstadial transmitted from R. sanguineus larvae to nymphs, and also suggest that these nymphs may retransmitted the bacteria through the saliva during their blood meal. This is the first study that validated the artificial membrane feeding system for maintaining R. sanguineus tick colony. It shows the possibility of transstadial transmission of B. henselae from R. sanguineus larvae to nymphs., Competing Interests: The authors have declared that no competing interests exist.

Bartonella henselae endocarditis mimicking systemic vasculitis has been reported in patients with valvulopathy. Herein, we describe a patient with B. henselae endocarditis involving a prosthetic pulmonic valve with positive anti-dsDNA antibodies misdiagnosed with systemic lupus erythematosus (SLE) based on the revised classification SLE criteria.

Bartonella henselae ( Bh ) is a Gram-negative rod transmitted to humans by a scratch from the common house cat. Infection of humans with Bh can result in a range of clinical diseases including lymphadenopathy observed in cat-scratch disease and more serious disease from persistent bacteremia. It is a common cause of blood-culture negative endocarditis as the bacterium is capable of growing as aggregates, and forming biofilms on infected native and prosthetic heart valves. The aggregative growth requires a trimeric autotransporter adhesin (TAA) called Bartonella adhesin A (BadA). TAAs are found in all Bartonella species and many other Gram-negative bacteria. Using Bh Houston-1, Bh Houston-1 ∆badA and Bh Houston-1 ∆badA /pNS2P Trc badA (a partial complement of badA coding for a truncated protein of 741 amino acid residues), we analyze the role of BadA in adhesion and biofilm formation. We also investigate the role of environmental factors such as temperature on badA expression and biofilm formation. Real-time cell adhesion monitoring and electron microscopy show that Bh Houston-1 adheres and forms biofilm more efficiently than the Bh Houston-1 ∆badA . Deletion of the badA gene significantly decreases adhesion, the first step in biofilm formation in vitro, which is partially restored in Bh Houston-1 ∆badA /pNS2P Trc badA . The biofilm formed by Bh Houston-1 includes polysaccharides, proteins, and DNA components and is susceptible to enzymatic degradation of these components. Furthermore, both pH and temperature influence both badA expression and biofilm formation. We conclude that BadA is required for optimal adhesion, agglutination and biofilm formation., Competing Interests: The authors declare no competing interests.

Background: The genus Bartonella includes fastidious, facultative intracellular bacteria mainly transmitted by arthropods and distributed among mammalian reservoirs. Bartonella spp. implicated as etiological agents of zoonoses are increasing. Apart from the classical Bartonella henselae, B. bacilliformis or B. quintana, other species (B. elizabethae, B. rochalimae, B. vinsonii arupensis and B. v. berkhoffii, B. tamiae or B. koehlerae, among others) have also been associated with human and/or animal diseases. Laboratory techniques for diagnosis (culture, PCR assays and serology) usually show lack of sensitivity. Since 2005, a method based on a liquid enrichment Bartonella alphaproteobacteria growth medium (BAPGM) followed by PCRs for the amplification of Bartonella spp. has been developed. We aimed to assess culture, molecular and serological prevalence of Bartonella infections in companion animal veterinary personnel from Spain., Methods: Each of 89 participants completed a questionnaire. Immunofluorescence assays (IFA) using B. vinsonii berkhoffii (genotypes I, II and III), B. henselae, B. quintana and B. koehlerae as antigens were performed. A cut-off of 1:64 was selected as a seroreactivity titer. Blood samples were inoculated into BAPGM and subcultured onto blood agar plates. Bartonella spp. was detected using conventional and quantitative real-time PCR assays and DNA sequencing., Results: Among antigens corresponding to six Bartonella spp. or genotypes, the lowest seroreactivity was found against B. quintana (11.2%) and the highest, against B. v. berkhoffii genotype III (56%). A total of 27% of 89 individuals were not seroreactive to any test antigen. Bartonella spp. IFA seroreactivity was not associated with any clinical sign or symptom. DNA from Bartonella spp., including B. henselae (n = 2), B. v. berkhoffii genotypes I (n = 1) and III (n = 2), and B. quintana (n = 2) was detected in 7/89 veterinary personnel. PCR and DNA sequencing findings were not associated with clinical signs or symptoms. No co-infections were observed. One of the two B. henselae PCR-positive individuals was IFA seronegative to all tested antigens whereas the other one was not B. henselae seroreactive. The remaining PCR-positive individuals were seroreactive to multiple Bartonella spp. antigens., Conclusions: High serological and molecular prevalences of exposure to, or infection with, Bartonella spp. were found in companion animal veterinary personnel from Spain. More studies using BAPGM enrichment blood culture and PCR are needed to clarify the finding of Bartonella PCR-positive individuals lacking clinical symptoms.

Bartonella henselae, the agent of cat scratch disease (CSD), appears to be a common organism responsible for lymphadenitis in both adults and children. There is a very low isolation rate for B. henselae from lymph nodes of patients with CSD. Our objective was to evaluate B. henselae viability in a large series of lymph nodes from patients with CSD. From January to November 2016, we analyzed lymph node biopsy samples from patients diagnosed with CSD. We used reverse transcription-quantitative polymerase chain reaction (RT-qPCR) to detect B. henselae RNA, as well as cultures, histological analyses, and fluorescence in situ hybridization (FISH). We tested 87 lymph nodes positive for B. henselae DNA but only 8 (9%) presented with B. henselae RNA. We did not find a significant difference for the pap threshold cycle (C T ) values between RNA-positive and RNA-negative lymph nodes (p = 0.5). Cultures, histological analyses, and FISH were negative for all the tested samples. We provide evidence that B. henselae are not or are rarely viable in most cases in the lymph nodes of patients with CSD.

Cardiovascular disease is still the major cause of death worldwide despite the remarkable progress in its prevention and treatment. Endothelial progenitor cells (EPCs) have recently emerged as key players of vascular repair and regenerative medicine applied to cardiovascular disease. A large amount of effort has been put into discovering the factors that could aid or impair the number and function of EPCs, and also into characterizing these cells at the molecular level in order to facilitate their therapeutic applications in vascular disease. Interestingly, the major cardiovascular risk factors have been associated with reduced number and function of EPCs. The bacterial contribution to cardiovascular disease represents a long-standing controversy. The discovery that Bartonella henselae can infect and damage EPCs revitalizes the enduring debate about the microbiological contribution to atherosclerosis, thus allowing the hypothesis that this infection could impair the cardiovascular regenerative potential and increase the risk for cardiovascular disease. In this review, we summarize the rationale suggesting that Bartonella henselae could favour atherogenesis by infecting and damaging EPCs, thus reducing their vascular repair potential. These mechanisms suggest a novel link between communicable and non-communicable human diseases, and put forward the possibility that Bartonella henselae could enhance the susceptibility and worsen the prognosis in cardiovascular disease.

Bartonella henselae (Rhizobiales: Bartonellaceae) is a Gram-negative fastidious bacterium of veterinary and zoonotic importance. The cat flea Ctenocephalides felis (Siphonaptera: Pulicidae) is the main recognized vector of B. henselae, and transmission among cats and humans occurs mainly through infected flea feces. The present study documents the use of a quantitative molecular approach to follow the daily kinetics of B. henselae within the cat flea and its excreted feces after exposure to infected blood for 48 h in an artificial membrane system. B. henselae DNA was detected in both fleas and feces for the entire life span of the fleas (i.e., 12 days) starting from 24 h after initiation of the blood meal.

Bartonella henselae, the causative agent of cat scratch disease, is increasingly recognized as a cause of culture-negative endocarditis. This report describes the first Japanese case, which was diagnosed after a prolonged culture of the excised aortic valve. High IgG and IgM titers to B. henselae pointed to a subacute course of the disease., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Bartonella henselae (Bh) is a worldwide distributed zoonotic pathogen. Depending on the immune status of the infected individual this bacterium can cause a wide spectrum of clinical manifestations, ranging from cat scratch disease (CSD) to bacillary angiomatosis (BA) and bacillary peliosis (BP). BA and BP are characterized by tumor-like lesions at the skin or in the inner organs, respectively. These structures display pathological sprouting of capillaries with enlarged and hyperproliferated vascular endothelial cells (ECs) that are frequently found in close association with bacteria. Here we review the cellular changes observed upon Bh infection of ECs in vitro and outline the role of the VirB type IV secretion system (T4SS) and its translocated effector proteins in the modulation of EC signalling cascades. The current model how this virulence system could contribute to the vasoproliferative activity of Bh is described.

The influence of in vitro passage on Bartonella henselae pathogenesis in cats has not been thoroughly evaluated. Our objective was to examine the bacterial kinetics and humoral immune responses in cats experimentally infected with three different in vitro passages of B. henselae F1, a genotype I strain of feline origin. The F1 strain was in vitro passaged 20 and 40 times, and each was inoculated into a group of 5 cats. The kinetics of bacteremia and the feline humoral immune response to bacterial antigens were compared to a previous study involving a group of six cats inoculated with the original F1 strain. Among the three groups of cats, the kinetics of bacteremia profiles and the humoral immune responses to B. henselae lysates were similar. The influence of passage on bacterial membrane proteins was examined. In vitro passage altered the expression of 4/17 (23.5%) bacterial membrane proteins and 6/15 (40%) bacterial membrane antigens. An association between poor seroreactivity to three lysate antigens (15-, 18- and 45kDa), prolonged bacteremia and decreased serum bactericidal activity was noted. Our data show that in vitro passage of B. henselae did not alter the kinetics of bacteremia, including the occurrence of relapsing bacteremia, in experimentally infected cats. This suggests that highly passaged strains may not be suitable for future vaccination studies. Furthermore, in vitro passage results in phenotypic and antigenic changes in the bacterial membrane protein profile, which warrants caution in the interpretation of studies involving passaged B. henselae strains.

Background: Most patients with infectious endocarditis (IE) due to Bartonella henselae have a history of exposure to cats and pre-existing heart valve lesions. To date, none of the reported patients have had a history of typical cat scratch disease (CSD) which is also a manifestation of infection with B. henselae., Case Presentation: Here we report the case of a patient who had CSD and six months later developed IE of the mitral valve caused by B. henselae., Conclusion: Based on this unique case, we speculate that CSD represents the primary-infection of B. henselae and that IE follows in patients with heart valve lesions.

Bartonella henselae isolates from different hosts display a marked genetic heterogeneity, as determined by pulsed-field gel electrophoresis (PFGE). The aim of the present study was to determine whether different genetic variants may coexist within the population of distinct B. henselae isolates and could be detected by PFGE. Three primary B. henselae isolates and the B. henselae reference strains ATCC 49793 and 49882 were subjected as single colony derived cultures in quadruplicate to PFGE analysis upon restriction with SmaI or NotI. Up to 4 fragment differences were found among the cultures obtained from each primary isolate, indicating the coexistence of genetic variants in the population of primary B. henselae isolates. The clonal relatedness of the genetic variants was confirmed by arbitrarily primed PCR and multi-locus sequence typing. In contrast to the primary isolates, no variants were detected among the single colony derived cultures of the high-passage ATCC strains. We hypothesized that the coexistence of different genetic variants may represent a feature that is restricted to primary or low-passage B. henselae isolates. The primary isolates were serially passed in vitro and then subjected as single colony derived cultures to PFGE analysis, which now revealed identical patterns among the quadruplicate cultures of each high-passage isolate. These results suggest that the population of a primary B. henselae isolate is composed of distinct genetic variants, which may disappear upon repeated passages on artificial culture media. Generation of genetic variants by B. henselae may represent an escape mechanism to circumvent the host specific immune responses.

Although there is evidence that endothelial cells are important targets for human pathogenic Bartonella species, the primary niche of infection is unknown. Here we elucidated whether human CD34+ hematopoietic progenitor cells (HPCs) internalize B. henselae and may serve as a potential niche of the pathogen. We showed that B. henselae does not adhere to or invade human erythrocytes. In contrast, B. henselae invades and persists in HPCs as shown by gentamicin protection assays, confocal laser scanning microscopy (CLSM), and electron microscopy (EM). Fluorescence-activated cell sorting (FACS) analysis of glycophorin A expression revealed that erythroid differentiation of HPCs was unaffected following infection with B. henselae. The number of intracellular B. henselae continuously increased over a 13-day period. When HPCs were infected with B. henselae immediately after isolation, intracellular bacteria were subsequently detectable in differentiated erythroid cells on day 9 and day 13 after infection, as shown by CLSM, EM, and FACS analysis. Our data provide, for the first time, evidence that a bacterial pathogen is able to infect and persist in differentiating HPCs, and suggest that HPCs might serve as a potential primary niche in Bartonella infections.

Bacteria of the genus Bartonella, a member of the Alphaproteobacteria, are fastidious, gram-negative, aerobic bacilli that comprise numerous species, subspecies, and subtypes. In human and veterinary medicine, species isolation remains a vital component of the diagnostic and therapeutic management of Bartonella infection. We describe a novel, chemically modified, insect-based liquid culture medium that supports the growth of at least seven Bartonella species. This medium will also support cocultures consisting of different Bartonella species, and it facilitated the primary isolation of Bartonella henselae from blood and aqueous fluid of naturally infected cats. This liquid growth medium may provide an advantage over conventional direct blood agar plating for the diagnostic confirmation of bartonellosis.

Bartonella henselae is a causative agent of cat scratch disease. We preliminarily tested four media for the bacterial growth, including agar plates with sheep, horse or rabbit blood, and chocolate agar. Of these media, rabbit blood and chocolate agar plate were found to be more excellent for the growth than the medium with sheep or horse blood. Blood samples from 60 domestic cats in Yamaguchi Prefecture were then cultured using 7% rabbit blood agar plates and BACTEC9050 (BD), automated blood culture microbial detection system. B. henselae was isolated from six of the 60 (10%) blood samples. Tiny colonies of B. henselae were visible on the agar medium after one week of culture at 35 degrees C in the 5% CO2 atmosphere. BACTEC 9050 detected B. henselae in one of the 10 blood samples and it took two weeks to detect the bacteria automatically, though gram stain failed to show organisms in the blood culture bottle. In conclusion, rabbit blood or chocolate agar and incubation of agar media more than one week and of BACTEC more than two weeks are recommended for the detection of B. henselae.

Bartonella henselae is an arthropod-borne zoonotic pathogen causing intraerythrocytic bacteraemia in the feline reservoir host and a broad range of clinical manifestations in incidentally infected humans. Remarkably, B. henselae can specifically colonize the human vascular endothelium, resulting in inflammation and the formation of vasoproliferative lesions known as bacillary angiomatosis and bacillary peliosis. Cultured human endothelial cells provide an in vitro system to study this intimate interaction of B. henselae with the vascular endothelium. However, little is known about the bacterial virulence factors required for this pathogenic process. Recently, we identified the type IV secretion system (T4SS) VirB as an essential pathogenicity factor in Bartonella, required to establish intraerythrocytic infection in the mammalian reservoir. Here, we demonstrate that the VirB T4SS also mediates most of the virulence attributes associated with the interaction of B. henselae during the interaction with human endothelial cells. These include: (i) massive rearrangements of the actin cytoskeleton, resulting in the formation of bacterial aggregates and their internalization by the invasome structure; (ii) nuclear factor kappaB-dependent proinflammatory activation, leading to cell adhesion molecule expression and chemokine secretion, and (iii) inhibition of apoptotic cell death, resulting in enhanced endothelial cell survival. Moreover, we show that the VirB system mediates cytostatic and cytotoxic effects at high bacterial titres, which interfere with a potent VirB-independent mitogenic activity. We conclude that the VirB T4SS is a major virulence determinant of B. henselae, required for targeting multiple endothelial cell functions exploited by this vasculotropic pathogen.

Bartonella henselae is a zoonotic pathogen that usually causes a self-limiting infection in immunocompetent individuals but often causes potentially life-threatening infections, such as bacillary angiomatosis, in immunocompromised patients. Both diagnosis of infection and research into the molecular mechanisms of pathogenesis have been hindered by the absence of a suitable liquid growth medium. It has been difficult to isolate B. henselae directly from the blood of infected humans or animals or to grow the bacteria in liquid culture media under laboratory conditions. Therefore, we have developed a liquid growth medium that supports reproducible in vitro growth (3-h doubling time and a growth yield of approximately 5 x 10(8) CFU/ml) and permits the isolation of B. henselae from the blood of infected cats. During the development of this medium, we observed that B. henselae did not derive carbon and energy from the catabolism of glucose, which is consistent with genome nucleotide sequence data suggesting an incomplete glycolytic pathway. Of interest, B. henselae depleted amino acids from the culture medium and accumulated ammonia in the medium, an indicator of amino acid catabolism. Analysis of the culture medium throughout the growth cycle revealed that oxygen was consumed and carbon dioxide was generated, suggesting that amino acids were catabolized in a tricarboxylic acid (TCA) cycle-dependent mechanism. Additionally, phage particles were detected in the culture supernatants of stationary-phase B. henselae, but not in mid-logarithmic-phase culture supernatants. Enzymatic assays of whole-cell lysates revealed that B. henselae has a complete TCA cycle. Taken together, these data suggest B. henselae may catabolize amino acids but not glucose to derive carbon and energy from its host. Furthermore, the newly developed culture medium should improve isolation of B. henselae and basic research into the pathogenesis of the bacterium.

Four Bartonella species have been isolated from domestic cats, of which two serotypes/genotypes of Bartonella henselae and possibly B. clarridgeiae are human pathogens, causing cat scratch disease (CSD).Our objectives were to evaluate infection and potential cross-protection during re-infection in domestic cats with various Bartonella species or types.Thirty-six cats were primarily inoculated with B. henselae type I (n=16), B. henselae type II (n=10), B. clarridgeiae (n=6) or B. koehlerae (n=4). They were challenged with B. henselae type I (n=15), B. henselae type II (n=13) or B. clarridgeiae (n=8). All 36 cats became bacteremic (1.25x10(2)-1.44x10(6)CFU/ml) and bacteremia lasted from 37 to 582 days. Duration of bacteremia for cats inoculated with B. henselae type I was shorter than for cats inoculated with either B. henselae type II (P=0.025) or B. clarridgeiae (P=0.011). After challenge, 26 cats became bacteremic. Among the nine cats primarily inoculated with B. henselae type I and challenged with B. henselae type II, six cats stayed abacteremic. The three bacteremic cats had a transient low-level bacteremia. No bacteremia was observed in three cats primarily inoculated with B. henselae type I and challenged with another strain of B. henselae type I. Bacteremia levels in the 26 cats were significantly lower than for primary inoculation (P=0.022) and its duration was shorter (P=0.012). Among the eight cats challenged with B. clarridgeiae, duration of bacteremia in the four cats primarily inoculated with B. henselae type I was shorter than in the four cats primarily inoculated with B. henselae type II (P=0.01). Bartonella clarridgeiae inoculated cats were more likely to have relapses for both primary and secondary infections. This is the first demonstration of cross-protection, evidenced by absence of bacteremia, in cats primarily infected with B. henselae type I and challenged with B. henselae type II, whereas no cross-protection was previously shown for cats primarily infected with B. henselae type II and challenged with B. henselae type I. Such results are of major importance for future feline Bartonella vaccine development.

Bartonella henselae is a fastidious, Gram-negative bacterial pathogen of cats and humans. Previous workers have shown that serial passage in vitro leads to attenuation of virulence-associated attributes such as expression of pili, invasion of human epithelial cell lines and the stimulation of endothelial cell proliferation. In contrast to the published data, it was found that pilin expression is frequently preserved in organisms which have undergone phase variation in vitro. Transition from a slow-growing, dry agar-pitting (DAP) to a faster-growing, smooth non-agar-pitting (SNP) form appears to occur predictably and may reflect competition between two populations growing at different rates. Better survival of the slower-growing (DAP) form may explain its relatively easy retrieval from piliated SNP populations allowed to age on solid media. Pilin expression is associated with auto-agglutination in liquid suspension or broth cultures, and appears to be necessary but not sufficient for expression of the agar-pitting phenotype and for the formation of biofilms. Outer-membrane protein variation is seen in association with phase variation, but lipopolysaccharide expression is preserved in piliated as well as extensively passaged non-piliated isolates. The EagI/HhaI infrequent restriction site-PCR fingerprint, which has been previously used to discriminate between serotypes Marseille and Houston, is shown to alter with phase variation in vitro, and there is evidence that genetic change accompanies these events. The extent of genetic and phenotypic variability of phase-variant B. henselae has previously been underestimated. It may lead to new insights into the pathogenicity of this organism, and must be considered when interpreting data arising from such studies.

Two out of three pools of cat fleas, Ctenocephalides felis (Bouche), that were fed Bartonella henselae-positive cat blood for 3 d and then bovine blood for 3 d, were polymerase chain reaction (PCR) positive for B. henselae. In a second experiment, three cats were inoculated with a streptomycin-resistant strain of B. henselae. After the cats were inoculated, caged cat fleas were fed on the cats during three different periods, and then pooled and transferred to noninfected recipient cats. In the first trial, the bacteria in the flea feces were below level of detection when the fleas were transferred from the infected cats to the recipient cat. After the fleas had fed on the recipient cat for 6 d, a bacteria level of 4.00 x 10(3) CFU/ mg was detected in the flea feces. Subsequently, the bacteria level increased for 4 d and then declined. In another experiment, the bacteria level in the flea feces was 1.80 x 10(3) CFU/mg at 2 h after collection and 3.33 x 10(2) CFU/mg at 72 h after collection. These data indicated that this strain of B. henselae can persist in flea feces in the environment for at least 3 d, and that B. henselae can multiply in the cat flea.

Bacterial infection due to Bartonella henselae commonly develops in children and young adults following cat/dog contacts and/or cat/dog scratches. Regional lymphadenopathy is its most common clinical expression. However, encephalitis and Parinaud's syndrome (oculoglandular syndrome) have also been reported as has systemic illness. A review of the international literature in all languages revealed no fatal complications in immunocompetent hosts. A four-year-old white child with no underlying illness began to have seizure-like activity. She was taken to a local hospital and subsequently transferred to a medical center. The child was treated aggressively for seizures and fever of unknown origin. However, her condition rapidly declined and she died without a specific diagnosis. At autopsy there was marked cerebral edema with no gross evidence of acute meningitis. Microscopic exams revealed multiple granulomatous lesions as well as a meningitis and encephalitis. A variety of cultures and stains were negative for acid fast and fungal organisms. Warthin-Starry stains of involved tissue including brain and liver revealed pleomorphic rod shaped bacilli consistent with Barronella henselae. Analysis of brain tissue with polymerase chain reaction (PCR) and Southern blot for the deoxyribonucleic acid (DNA) was definitive for DNA of Bartonella henselae bacteria.

Bartonella henselae causes cat-scratch disease and bacillary angiomatosis peliosis. The bacteria reside in erythrocytes of asymptomatic cats, which represent the natural reservoir for this pathogen. B. henselae is usually grown on blood-enriched media. Growth experiments on Brucella medium without blood demonstrated that heme compounds are essential for the growth of B. henselae and can completely substitute the addition of blood components. The heme precursor protoporphyrin IX alone, or in combination with FeCl(2) or FeCl(3), as well as transferrin or lactoferrin did not support growth, indicating that B. henselae cannot synthesize heme itself. Hemin supported growth even when free iron was chelated, indicating that hemin is also used as an iron source. Binding assays showed that hemin starvation increased the binding capacity of B. henselae for hemin, providing evidence that the bacteria carry a specific hemin uptake system, which might be regulated by hemin.

Bartonella quintana and Bartonella henselae are fastidious gram-negative bacteria responsible for bacillary angiomatosis, trench fever, cat scratch disease, and endocarditis. During a 5-year period, we received 2,043 samples for culture of Bartonella sp. We found Bartonella sp. to be the etiologic agent in 38 cases of endocarditis, 78 cases of cat scratch disease, 16 cases of bacteremia in homeless people, and 7 cases of bacillary angiomatosis. We correlated the results of positive cultures with the clinical form of the disease, type of sample, culture procedure, PCR-based genomic detection, and antibody determination. Seventy-two isolates of B. quintana and nine isolates of B. henselae from 43 patients were obtained. Sixty-three of the B. quintana isolates and two of the B. henselae isolates, obtained from patients with no prior antibiotic therapy, were stably subcultured. The sensitivity of culture was low when compared with that of PCR-based detection methods in valves of patients with endocarditis (44 and 81%, respectively), skin biopsy samples of patients with bacillary angiomatosis (43 and 100%, respectively), and lymph nodes of cat scratch disease (13 and 30%, respectively). Serological diagnosis was also more sensitive in cases of endocarditis (97%) and cat scratch disease (90%). Among endocarditis patients, the sensitivity of the shell vial culture assay was 28% when inoculated with blood samples and 44% when inoculated with valvular biopsy samples, and the sensitivity of both was significantly higher than that of culture on agar (5% for blood [P = 0.045] and 4% for valve biopsy samples [P < 0.0005]). The most efficient culture procedure was the subculture of blood culture broth into shell vials (sensitivity, 71%). For patients with endocarditis, previous antibiotic therapy significantly affected results of blood culture; no patient who had been administered antibiotics yielded a positive blood culture, whereas 80% of patients with no previous antibiotic therapy yielded positive blood cultures (P = 0.0006). Previous antibiotic therapy did not, however, prevent isolation of Bartonella sp. from cardiac valves but did prevent the establishment of strains, as none of the 15 isolates from treated patients could be successfully subcultured. For the diagnosis of B. quintana bacteremia in homeless people, the efficiency of systematic subculture of blood culture broth onto agar was higher than that of direct blood plating (respective sensitivities, 98 and 10% [P < 10(-7)]). Nevertheless, both procedures are complementary, since when used together their sensitivity reached 100%. All homeless people with positive blood cultures had negative serology. The isolation rate of B. henselae from PCR-positive lymph nodes, in patients with cat scratch disease, was significantly lower than that from valves of endocarditis patients and skin biopsy samples from bacillary angiomatosis patients (13 and 33%, respectively [P = 0.084]). In cases of bacillary angiomatosis for which an agent was identified to species level, the isolation rate of B. henselae was lower than the isolation rate of B. quintana (28 and 64%, respectively [P = 0.003]). If culture is to be considered an efficient tool for the diagnosis of several Bartonella-related diseases, methodologies need to be improved, notably for the recovery of B. henselae from lymph nodes of patients with cat scratch disease.

Objectives: To report seven cases of bacillary angiomatosis; to evaluate the most useful diagnostic tools; to analyse the clinical and epidemiological features associated with Bartonella quintana or Bartonella henselae infections., Design: Clinical, diagnostic and epidemiological evaluation of 37 speciated bacillary angiomatosis cases in the literature, including the seven patients in our study., Methods: Pathological examination of tissue samples, including Warthin-Starry staining and immunohistology; titre of antibodies to Bartonella sp.; detection of Bartonella sp. in blood and biopsy materials by culture or PCR; and statistical analysis of clinical and epidemiological features associated with B. quintana or B. henselae bacillary angiomatosis cases., Results: Seven immunocompromised patients (six with AIDS and one patient with acute leukaemia) had bacillary angiomatosis confirmed by histology. B. quintana was cultured in three patients, whereas B. henselae DNA was amplified by PCR in the remaining four patients. Serum from only one patient reacted with Bartonella antigens. Amongst the 14 B. quintana and 23 B. henselae bacillary angiomatosis cases now reported in the literature, lymphadenopathies were significantly more frequent in B. henselae-infected patients, and neurological disorders of the central nervous system in B. quintana-infected patients. Risk factors were contact with cats, and homelessness or poor socioeconomic status in B. henselae and B. quintana bacillary angiomatosis cases, respectively., Conclusions: Although diagnosis of bacillary angiomatosis often remains solely based upon histology, culture or PCR-based methods are useful for the detection of Bartonella sp., and allow identification of the species involved, which is necessary to further characterize clinical and epidemiological features associated with B. quintana or B. henselae infections.

Members of the genus Bartonella are unique in that they are bacteria which cause proliferation of microvascular endothelial cells and neovascularization (angiogenesis). The mechanisms by which Bartonella henselae causes these processes are unknown. Given the importance of surface-exposed determinants in the pathogenesis of many organisms, outer membrane proteins (OMPs) of B. henselae were identified. Enrichment of the outer membrane fraction of B. henselae by sarkosyl treatment of total membranes, together with radioiodination and biotinylation of intact organisms, suggest that at least nine proteins, with molecular weights of 28, 30, 35, 43, 58, 61, 79, 92 and 171 kDa, are located in the outer membrane. Triton X-100-extracted biotinylated human umbilical vein endothelial cell (HUVEC) surface proteins bound to the 43 kDa B. henselae OMP after B. henselae whole-cell lysates and sarkosyl-fractionated OMPs were separated by SDS-PAGE and transferred onto nylon. Biotinylated B. henselae surface proteins of 28, 32, 43, 52 and 58 kDa were shown to bind intact HUVEC, with the 43 kDa protein being the major adhesin. Preincubation of HUVEC with an increasing concentration (20 microg/ml to 4 mg/ml) of sarkosyl-fractionated unlabelled B. henselae outer membrane proteins inhibited the attachment of all identified HUVEC binding proteins. The identification of B. henselae OMPs, as well as adhesins, should provide a basis for further investigation of the role of adherence in the pathogenesis of B. henselae., (Copyright 1998 Academic Press)

Rochalimaea (Bartonella) henselae is a fastidious, slowly growing, gram-negative bacillus that is an etiologic agent of bacillary angiomatosis, cat scratch disease, and related syndromes. Accumulation of direct microbiologic evidence of the relationship between the organism and the syndromes compatible with cat scratch disease has been hindered by the difficulties in the primary isolation of the organism from infected tissue specimens. A chemically defined liquid medium was developed to support the growth of Rochalimaea species to facilitate study of the organism. This medium was also used successfully to isolate R. henselae from clinical specimens from infected patients and a domestic cat. Recovery of R. henselae in this was more successful than when recovery was attempted on solid agar. This cell-free, extract-free, defined medium additionally supported the growth of Rochalimaea quintana and Afipia felis.

Several years ago, Rochalimaea henselae has emerged as an agent of bacillary angiomatosis, bacillary peliosis and recurrent septicaemia that generally occur in patients infected with human immunodeficiency virus. An aetiologic role in cat scratch disease is also suspected widely on the basis of a serologic survey. Its slow growth and its culture requirement explain that this pathogen, a gram-negative bacterium, could not be isolated until 1990. Moreover, blood and tissue samples request lysis and crushing for recovering by culture. The clinical, histological, microbiological and pathogenic aspects of these infections are described and discussed.