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Infective endocarditis due to Bartonella spp. and Coxiella burnetii: experience at a cardiology hospital in Sao Paulo, Brazil.

Bartonella spp. and Coxiella burnetii are recognized as causative agents of blood culture-negative endocarditis (BCNE) in humans and there are no studies of their occurrences in Brazil. The purpose of this study is to investigate Bartonella spp. and C. burnetii as a causative agent of culture-negative endocarditis patients at a cardiology hospital in São Paulo, Brazil. From January 2004 to December 2004 patients with a diagnosis of endocarditis at our Institute were identified and recorded prospectively. They were considered to have possible or definite endocarditis according to the modified Duke criteria. Those with blood culture-negative were tested serologically using the indirect immunofluorescent assay (IFA) for Bartonella henselae, B. quintana, and C. burnetii. IFA-IgG titers >800 for Bartonella spp. and C. burnetii were considered positive. A total of 61 patients with endocarditis diagnosis were evaluated, 17 (27%) were culture-negative. Two have had IgG titer greater than 800 (>/=3,200) against Bartonella spp. and one against C. burnetii (phase I and II>/=6,400). Those with Bartonella-induced endocarditis had a fatal disease. Necropsy showed calcifications and extensive destruction of the valve tissue, which is diffusely infiltrated with mononuclear inflammatory cells predominantly by foamy macrophages. The patient with C. burnetii endocarditis received specific antibiotic therapy. Reports of infective endocartitis due to Bartonella spp. and C. burnetii in Brazil reveal the importance of investigating the infectious agents in culture-negative endocarditis.

Adult↗

Quantitative analysis of valvular lesions during Bartonella endocarditis.

Cardiac valve pathology was evaluated in 15 patients with confirmed diagnosis of Bartonella endocarditis. Ten were infected by Bartonella quintana and 5 by Bartonella henselae. Histologic features of these cases, including fibrosis, calcification, vegetation, pattern of inflammation, and vascularization, were compared with those of valves from 25 cases of non-Bartonella endocarditis as controls using a computerized quantitative image analysis. Pathologic and immunohistologic testing for localization of Bartonella species in resected valves included Warthin-Starry stain and polyclonal antibody-based immunodetection. Compared with other cases of infective endocarditis, cases of Bartonella endocarditis are more fibrotic and calcified, less vascularized, with less extensive vegetation and chronic inflammation. These pathologic changes are suggestive of a prolonged infection. Warthin-Starry stain and immunohistologic testing demonstrated the presence of the organism, respectively, in 11 and 10 of the 13 tested valves. Results of both staining methods showed microorganisms in extracellular locations and in regions unaccompanied by inflammation. Pathology and immunohistology may contribute to the etiologic diagnosis of Bartonella endocarditis when serology and molecular techniques are not available.

Adult↗

Seroprevalence of antibodies against Bartonella species and evaluation of risk factors and clinical signs associated with seropositivity in dogs.

OBJECTIVE: To determine the seroprevalence of antibodies against Bartonella spp in a population of sick dogs from northern California and identify potential risk factors and clinical signs associated with seropositivity. SAMPLE POPULATION: Sera from 3,417 dogs. PROCEDURE: Via an ELISA, sera were analyzed for antibodies against Bartonella vinsonii subsp berkhoffii, Bartonella clarridgeiae, and Bartonella henselae; test results were used to classify dogs as seropositive (mean optical density value > or = 0.350 for B henselae or > or = 0.300 for B clarridgeiae or B vinsonii subsp berkhoffi) or seronegative. Overall, 305 dogs (102 seropositive and 203 seronegative dogs) were included in a matched case-control study. RESULTS: 102 of 3,417 (2.99%) dogs were seropositive for > or = 1 species of Bartonella. Of these, 36 (35.3%) had antibodies against B henselae only, 34 (33.3%) had antibodies against B clarridgeiae only, 2 (2.0%) had antibodies against B vinsonii subsp berkhoffii only, and 30 (29.4%) had antibodies against a combination of those antigens. Compared with seronegative dogs, seropositive dogs were more likely to be herding dogs and to be female, whereas toy dogs were less likely to be seropositive. Seropositive dogs were also more likely to be lame or have arthritis-related lameness, nasal discharge or epistaxis, or splenomegaly. CONCLUSIONS AND CLINICAL RELEVANCE: Only a small percentage of dogs from which serum samples were obtained had antibodies against Bartonella spp. Breed appeared to be an important risk factor for seropositivity. Bartonella infection should be considered in dogs with clinical signs of lameness, arthritis-related lameness, nasal discharge or epistaxis, or splenomegaly.

Animals↗

[Study on the prevalence of Bartonella species in rodent hosts from different environmental areas in Yunnan].

OBJECTIVE: To investigate Bartonella infections in small mammalian reservoir hosts from different environments and types of climate in Yunnan. METHODS: Femoral blood samples were collected from the anesthetic captured animals from five counties including three types of climate. All isolates were grown on brain and heart infusion agar plates containing 5% defibrinated rabbit blood. The agar plates were incubated at 35 degrees C in a humidified with 5% CO2 environment for at least 4 weeks. Bartonella-like isolates were confirmed by the polymerase chain reaction and visualizing the target gene fragment by gel electrophoresis. RESULTS: Bartonella species were isolated from 69 of 176 small animals including 4 species of 3 genera from 4 counties and the total prevalence in rodents was 39.2%. The maximal prevalence was 42.0% of Rattus tanezumi flavipectus usually inhabiting indoors and courtyard and contacting closely to human. Moreover, Bartonella isolates were obtained from Rattus noruegicus, Eothenomys miletus and Mus pahari. Life environments of captured animals involved indoors, courtyard, brush and forest in mountain. CONCLUSION: The finding in this study suggested the characteristic of diversity of Bartonella infections in rodent hosts in southern China included Bartonella species parasiting in a wide range of animal hosts in different environments as well as climate types. Further investigations were needed in different areas in China to confirm more mammalian reservoir hosts with Bartonella infections.

Animals↗

[Study on Bartonella vinsonii berkhoffii isolated from blood of native dogs in China].

OBJECTIVE: To isolate and identify Bartonella strains from native dogs in Shandong province in China. METHODS: EDTA-anticoagulated blood samples were collected from 71 native dogs in Yanggu county of Shandong province in March 2005. All isolates were grown on brain heart infusion agar plates containing 5% defibrinated rabbit blood. The agar plates were incubated at 37 degrees C in a humidified with 5% CO2 environment for 4 weeks or longer. All Bartonella-like isolates were examined by routine Gram and Giménez staining and then followed by polymerase chain reaction (PCR) and PCR-RFLP analysis for identification and differentiation of the isolates. Sequencing 16S rRNA, citrate synthase (gltA) gene and 16S-23S rRNA ITS were carried out and sequential similarities were calculated using the DNASTAR5 software package. The phylogenetic tree was inferred from each bootstrap sample, using the neighbor-joining methods as executed in the MEGA 3.1 software. The translation from DNA to protein were determined by DNASIS 2.5. RESULTS: The two Bartonella-like organisms (strains Q52SHD and Q64SHD) were isolated from the blood of 71 dogs. Light microscopic examination of the Gram and Giménez-stained micro-organisms showed small, short and slightly curved pleomorphic gram-negative bacilli. Amplified products of the three pairs of Bartonella genus-specific primers carried the same size as the predicted of those Bartonella species. Data from PCR-RFLP analysis showed that the two strains that having the same profiles were all different from the B. henselae type strain-16S rRNA, gltA and 16S-23S rRNA ITS sequences from the two isolates were 100.0%, 99.7% and 97.2% homologous to B. vinsonii berkhoffii. CONCLUSIONS: Based on these findings, the two isolates Q52SHD and Q64SHD were demonstrated as B. vinsonii berkhoffii. To our knowledge, this was the first report on the presence of Bartonella infection in native dogs from China, which constituted a large reservoir of Bartonella species in this country.

Animals↗

In vitro Bartonella quintana infection modulates the programmed cell death and inflammatory reaction of endothelial cells.

Bartonella quintana is an epicellular bacterium, which in vivo as well as in vitro, invades endothelial cells and develops within them inducing proliferative effects that play a pivotal role in neovascular manifestation of this disease. We investigated the effect of live Bartonella quintana and its LPS on apoptosis and inflammatory response in HUVEC-C, an endothelial cell line. The kinetics of the programmed cell death of Bartonella quintana-infected HUVEC-C showed a peculiar course. Even if early during infection apoptosis reached a peak after 6 h, later on apoptosis was inhibited. Such apoptosis inhibition was not observed during Bartonella quintana lipopolysaccharide treatment because LPS-stimulated HUVEC-C did progress to cell death. Evaluation of multiple cell signal transduction pathways revealed an overexpression of Apaf 1 and caspase 8 in HUVEC-C after 2 h of infection, and of bcl-2 starting from 10 h post Bartonella quintana infection. Moreover, Bartonella quintana and its LPS showed a different effect on the activation of genes involved in inflammatory response as revealed by molecular analysis of host cells. Bartonella quintana appears to be able to inhibit programmed cell death, inducing intracellular signals leading to survival and proliferation through the bcl-2 gene, despite the early increase of inflammatory status induced in endothelial cells. This mechanism, together with a poor endotoxin ability to stimulate strong inflammatory response, could contribute to the capability of the bacteria to persist intracellularly, causing chronic disease and producing neovascular manifestations.

Apoptosis↗

Identification of Rickettsia spp. and Bartonella spp. in ffrom the Thai-Myanmar border.

During a survey for possible rickettsial vectors in villages of the central part of the Thai-Myanmar border from September 2001 to February 2002, four species of fleas were collected from common peridomestic animals. All fleas were tested by PCR to detect DNA of bacteria of the genera Rickettsia (gltA and ompB genes) and Bartonella (ITS and ftsZ genes). Sequencing of PCR-amplified products was done using gltA fragments for Rickettsia and ftsZ fragments for BARTONELLA: Two genotypes related to Rickettsia felis were identified in three Ctenocephalides canis and one C. felis specimen. Further, the following Bartonella spp. were detected: Bartonella henselae in two C. felis specimens; Bartonella clarridgeiae in three C. felis specimens; and a new Bartonella genotype in one Nosopsylla fasciatus specimen. Rickettsia and Bartonella may be frequently detected in fleas infesting peridomestic animals from the western border of Thailand.

Animals↗

Evaluation of human seroreactivity to Bartonella species in Sweden.

Among the species that compose the expanding genus Bartonella, thus far only B. henselae and B. quintana have reportedly been isolated from humans in Europe. To evaluate the prevalence of Bartonella infection in Sweden, we conducted a retrospective serological examination of 126 human serum samples. These samples were analyzed for antibodies to B. henselae, B. quintana, and B. elizabethae. Serum samples from 100 blood donors, who spanned the ages of 20 to 60 and had no apparent clinical signs of illness, were also studied as a control group. An immunoglobulin G indirect fluorescence antibody assay revealed 4 and 8.3% Bartonella positivity rates for the blood donor and patient group, respectively, when a cutoff titer of >/=64 was chosen. Among the blood donors, four were seropositive to B. elizabethae; one of these also had concordant positive titer to B. henselae. In the patient group, 14 serum samples were positive against Bartonella spp. These serum specimens represented nine patients. In three of these seropositive patients, paired serum samples displayed a fourfold increase in antibody titer to at least one of the three antigens. These three patients are discussed. In this report we also present a case study of a 60-year-old Swedish male with fatal myocarditis. Postmortem serological analysis revealed a high titer against B. elizabethae. PCR and nucleotide sequencing of the myocardial tissue from this patient, and of liver tissue from one of the other three patients, showed sequences similar to B. quintana. The age, geographical origin, animal contacts, and serological response pattern to the different Bartonella antigens differed among the four patients. This study substantiates the presence of Bartonella spp. in Sweden, documents the seroreactivity to three Bartonella antigens in Swedish patients, and reports the first two cases of B. quintana-like infections in Sweden.

Adolescent↗

Potential limitations of the 16S-23S rRNA intergenic region for molecular detection of Bartonella species.

PCR targeting the 16S-23S rRNA gene intergenic transcribed spacer (ITS) region has been proposed as a rapid and reliable method for the detection of Bartonella species DNA in clinical samples. Because of variation in ITS sequences among Bartonella species, a single PCR amplification can be used to detect different species within this genus. Therefore, by targeting the ITS region, multiple PCRs or additional sample-processing steps beyond the primary amplification can be avoided when attempting to achieve molecular diagnostic detection of Bartonella species. Although PCR amplification targeting this region is considered highly sensitive, amplification specificity obviously depends on primer design. We report evidence of nonspecific PCR amplification of Mesorhizobium species with previously published primers that were designed to amplify the Bartonella consensus ITS region. Use of these or other, less species-specific, primers could lead to a false-positive diagnostic test result when evaluating clinical samples. We also report the presence of Mesorhizobium species DNA as a contaminant in molecular-grade water, a series of homologous sequences in the ITS region that are common to Bartonella and Mesorhizobium species, the amplification of Mesorhizobium DNA with unpublished primers designed in our laboratory targeting the ITS region, and the subsequent design of unambiguous ITS primers that avoid nonspecific amplification of Mesorhizobium species. Our results define some potential limitations associated with the molecular detection of Bartonella species in patient samples and indicate that primer specificity is of critical importance if the ITS region is used as a diagnostic target for detection of Bartonella species.

Bartonella↗

Rodent-associated Bartonella febrile illness, Southwestern United States.

Serum specimens from 114 patients hospitalized with a febrile illness were tested with an indirect immunofluorescence assay (IFA) using Bartonella antigens prepared from 6 species of sigmodontine rodents and 3 known human Bartonella pathogens: B. henselae, B. quintana, and B. elizabethae. Acute- and convalescent-phase serum samples from 5 of these patients showed seroconversion with an IFA titer >512 to rodent-associated Bartonella antigens. The highest titer was against antigen derived from the white-throated woodrat (Neotoma albigula), although this rodent is not necessarily implicated as the source of infection. Three of the 5 who seroconverted showed no cross-reaction to the 3 Bartonella human pathogens. Common clinical characteristics were fever, chills, myalgias, leukopenia, thrombocytopenia, and transaminasemia. Although antibodies to Bartonella are cross-reactive, high-titer seroconversions to rodent-associated Bartonella antigens in adults with common clinical characteristics should stimulate the search for additional Bartonella human pathogens.

Adolescent↗

[Presence of Bartonella henselae in cats: natural reservoir quantification and human exposition risk of this zoonoses in Chile].

BACKGROUND: The availability of a serologic test for cat scratch disease in humans has allowed the diagnosis of an increasing number of cases of this disease in Chile. AIM: To perform a serological survey for Bartonella henselae among cats in Chile. MATERIAL AND METHODS: Blood samples from 187 cats living in three Chilean cities were obtained. IgG antibodies against Bartonella henselae were measured using indirect immunofluorescence. Blood cultures were done in 60 samples. The presence of Bartonella henselae in positive cultures was confirmed by restriction fragment length polymorphism polymerase chain reaction (RFLP-PCR). RESULTS: The general prevalence of IgG antibodies against Bartonella henselae was 85.6%. No differences in this prevalence were found among cats younger or older than 1 year, or those infested or not infested with fleas. However domestic cats had a lower prevalence when compared with stray cats (73 and 90% respectively, p <0.01). Bartonella henselae was isolated in 41% of blood cultures. All the isolated were confirmed as Bartonella henselae by RFLP-PCR. CONCLUSIONS: This study found an important reservoir of Bartonella henselae in Chilean cats and therefore a high risk of exposure in humans who have contact with them.

Animals↗

Analysis and preparation of Bartonella bacilliformis antigens.

Twenty-four antigens of Bartonella bacilliformis, a bacterium which causes bartonellosis in residents of high altitude valleys of the Andes, were identified by immunoblot and immunoprecipitation using rabbit anti-Bartonella sera as well as sera of patients. The antigens were designated according to their relative molecular mass which ranged from 16 to 160 kDa. Twelve antigens were detected by antibodies in sera of bartonellosis patients using immunoblot, of which six antigens were detected by immunoprecipitation. Antigens 25, 46, 65, 75, 99, and 160 were identified as probable cell wall antigens. Antigens 50, 65, and 75 detected long-persisting antibodies. Crude Bartonella antigen applied to ELISA reacted with anti-Chlamydia psittaci antibody as well as with antibody of unknown identity in human sera, whereas immunoblot and immunoprecipitation with Triton soluble antigens revealed Bartonella-specific results. Seven Bartonella antigens were prepared by high performance liquid chromatography of which one antigen (48 kDa) reacted Bartonella-specific when applied to ELISA. It was concluded that specificity of antibody determination with crude Bartonella antigen should be confirmed by either immunoblot or immunoprecipitation.

Animals↗

Use of Bartonella antigens for serologic diagnosis of cat-scratch disease at a national referral center.

BACKGROUND: Bartonella henselae (formerly the genus Rochalimaea) has recently been isolated from patients with cat-scratch disease and their cats, and since September 1992 the Centers for Disease Control and Prevention has offered an indirect fluorescent antibody assay for Bartonella-specific antibody. METHODS: Physicians submitted serum samples from patients suspected of having cat-scratch disease or other Bartonella-associated illness and completed a questionnaire that recorded clinical information. Indirect fluorescent antibody assay was performed with the use of antigen derived from three Bartonella species: B henselae, Bartonella quintana, and Bartonella elizabethae. RESULTS: During 16 months, 3088 serum samples were received. The largest numbers of specimens and the highest percentages positive (titer, > or = 64) were observed in the fall and winter. Clinical histories of the first 600 patients for whom serum samples and completed information forms were received were examined in detail; seropositivity was significantly associated with cat contact, cat age of less than 1 year, cat scratch, presence of an inoculation papule, and regional adenopathy. Of 91 patients whose illness met a strict clinical definition of cat-scratch disease, 86 (95%) had titers of 64 or greater to either B henselae or B quintana. A fourfold rise or fall in titer was observed in 87 of 132 patients with paired serum samples. CONCLUSIONS: The indirect fluorescent antibody assay for Bartonella-specific antibody is sensitive for the diagnosis of cat-scratch disease. Redefinition of cat-scratch disease on the basis of cause and use of this assay as a diagnostic criterion is recommended.

Adolescent↗

Seroprevalence of Bartonella henselae in cats in Germany.

Bartonella henselae and B. quintana infections in man are associated with various clinical manifestations including cat-scratch disease, bacillary angiomatosis and bacteraemia. While cats are the natural reservoir for B. henselae, the source of B. quintana is unclear. In this study, the sera of 713 cats from Germany were examined for the presence of antibodies against B. henselae, B. quintana or Afipia felis by an indirect immunofluorescence assay (IFA). Bartonella-specific antibody titres of > or =50 were found in 15.0% of the cats. There was substantial cross-reactivity among the various Bartonella antigens, although single sera showed high titres against B. henselae but not against B. quintana and vice versa. Antibodies against A. felis were not detected in any of these cats. Statistical analysis indicated that there is no correlation between Bartonella infections and the sex, age or breed of the cat or its hunting behavior. There was also no correlation between bartonella and toxoplasma infections in cats. However, whereas 16.8% of cats from northern Germany had B. quintana-specific antibodies, only 8.0% of cats from southern Germany were seropositive for B. quintana. No statistically significant difference was found for B. henselae. IFA-positive and IFA-negative sera were used for immunoblot analysis including B. henselae and B. quintana. Marked reactivity was observed with protein bands at 80, 76, 73, 65, 37, 33 and 15 kDa. The results of this study suggest that B. henselae, and possibly a B. quintana-related pathogen, but not A. felis, are common in cats in Germany, and that there are differences in the geographic distribution of bartonella infections in cats.

Animals↗

Use of the cell division protein FtsZ as a means of differentiating among Bartonella species.

Genes coding for homologs of the highly conserved cell division protein FtsZ were isolated from Bartonella henselae and Bartonella quintana, the causative agents of cat scratch disease and trench fever, respectively. DNA fragments coding for the ftsZ open reading frames (ORFs) were cloned into Escherichia coli following PCR amplification with primers based on the ftsZ sequence of the closely related species Bartonella bacilliformis. The amino acid sequences predicted from the cloned B. henselae and B. quintana ftsZ ORFs are 81 to 83% identical to the corresponding protein in B. bacilliformis. Like the FtsZ protein of B. bacilliformis, the B. henselae and B. quintana homologs are about twice as large as the FtsZ proteins reported in most other organisms. Localized sequence differences within the C-terminal coding regions of the Bartonella ftsZ genes were used as the basis for species-specific identification of these organisms at both the DNA and protein levels. Oligonucleotide primers which permit the amplification of an ftsZ fragment from each of the Bartonella species without amplifying DNA from the other two species were designed. Anti-FtsZ antisera raised in rabbits against synthetic peptides corresponding to the relatively divergent C-terminal regions were shown via Western blot analysis to react only with the FtsZ protein from the cognate Bartonella species. These observations raise the possibility that the differences in ftsZ sequences can be used as the basis for diagnostic tests to differentiate among these closely related pathogens.

Amino Acid Sequence↗

Bartonella koehlerae, a new cat-associated agent of culture-negative human endocarditis.

Bartonella koehlerae is reported for the first time to be a human pathogen that causes culture-negative endocarditis. It is also shown that this species, isolated twice before from domestic cats, can be recovered as well from a stray cat population in Israel. This work follows a recent report of the same case in which the causative agent was misidentified as B. henselae, based on serology and PCR-restriction fragment length polymorphism (RFLP) analysis (A. Schattner, O. Zimhony, B. Avidor, and M. Gilad, Lancet 361:1786, 2003). B. koehlerae was identified in the valvular tissue of an endocarditis patient by DNA sequencing of the PCR products of two Bartonella genes: the genes for citrate synthase (gltA) and riboflavin synthase (ribC). The commonly used PCR-RFLP analysis of the TaqI-digested gltA PCR product did not distinguish between B. koehlerae and B. quintana or between B. elizabethae and B. clarridgeiae. PmlI digestion of the gltA amplification product failed to differentiate between B. quintana, B. clarridgeiae, and B. elizabethae. RFLP analysis of the heat shock protein (htrA) gene by TaqI digestion misidentified B. koehlerae as B. henselae. However, RFLP analysis of the ribC PCR product, digested with TaqI, was able to distinguish between the human endocarditis-associated Bartonella species tested, B. henselae, B. quintana, B. elizabethae, and B. koehlerae, as well as between the cat-associated Bartonella species, B. henselae and B. clarridgeiae. Given the expanding number of Bartonella species emerging as human pathogens, it is suggested that PCR-RFLP analysis for the diagnosis of Bartonella infections target several genes and be coupled with DNA sequencing to avoid species identification.

Bartonella↗

Subtyping of uncultured bartonellae using sequence comparison of 16 S/23 S rRNA intergenic spacer regions amplified directly from infected blood.

This study aimed to assess the usefulness of a PCR-based approach to the detection and differentiation of Bartonella strains in infected blood. The conservation of potential genus-specific PCR primer hybridisation sites within the 16 S/23 S rRNA gene intragenic spacer regions of Bartonella species was confirmed following sequence analysis of the intragenic spacer regions of four previously untested species. The extent of intra-species variation within the specific amplicons was assessed by comparison of sequences obtained from 17 strains of four Bartonella species. Eight sequence variants were obtained. Each species for which multiple strains were tested possessed at least two intragenic spacer regions variants, but the differences between these strains were markedly less than those observed inter-species. Sequence analysis was performed on 60 amplicons obtained from blood pellets collected from woodland rodent communities in which bartonella infections were known to be highly prevalent. Twelve variants were encountered, only five of which had been found among the studied isolates. Partial intragenic spacer region amplification followed by product sequence analysis offers a potentially sensitive and totally transferable means of inter- and intra-species differentiation of Bartonella strains, and its use in this study has broadened our knowledge of the genotypic spectrum of bartonellae associated with natural infections among UK woodland rodents.

Animals↗

Laboratory diagnosis of Bartonella infections.

Bartonella species are pathogens of emerging and reemerging significance, causing a wide array of clinical syndromes. In North America and Europe, they are increasingly recognized as a cause of culture negative endocarditis, neuroretinitis, and disease among homeless, HIV-infected, and other immunosuppressed individuals. In South America, bartonellosis continues to plague those in endemic regions and poses a significant threat to travelers in these areas. As the clinician is increasingly faced with these illnesses, which may be difficult to diagnose, laboratory techniques to confirm or refute the diagnosis are becoming increasingly important. Culture methods have improved over the past decade demonstrating increased sensitivity, but still require prolonged periods before isolation of the organism. Specimen handling, media selection, and growth conditions all may affect results and must be optimized in order to provide the highest likelihood of recovering the organism. Pure culture of the bacteria not only provides morphologic information, but also provides material for further diagnostic testing. Work with liquid media, which may provide a more rapid means of cultivation has shown some promise and should continue to be pursued. Improved blood culture techniques were a primary factor in the discovery of Bartonella endocarditis and continued improvements will likely demonstrate further clinical insights. Serologic testing for B henselae infections has become the cornerstone of clinical diagnosis, replacing the skin test that was poorly standardized and posed a potential risk to the patient. Immunofluorescence assays have been well characterized and validated in clinical trials, however they are not universally available. Vero cell cocultivated antigens appear to provide higher sensitivity and specificity when compared with agar-derived antigens. IFA assays are inherently difficult to perform, requiring significant expertise to provide reproducible results. On the contrary, enzyme immunoassays offer ease of use and a high level of reproducibility, however ideal antigens for use in the diagnosis of Bartonella infections have not been clearly identified. Continued work to define antigenic targets of the human response to infection and incorporation of these into a widely available EIA will provide a cost-effective tool for the clinician and epidemiologist alike. Due to the close phylogenetic relationship of B henselae and B quintana, differentiation between these species by serologic means may prove difficult. Molecular techniques including PCR offer high sensitivity and specificity, rapid availability of information, and the ability to differentiate Bartonella organisms at the highest level. Results of studies to date are promising and as methods are refined it will be important to conduct clinical studies to define the role of these assays. In disseminated Bartonella infections such as bacillary angiomatosis, peliosis, endocarditis, and urban trench fever, PCR currently offers the ability to establish the diagnosis when other tests may be unrevealing. For CSD, this technique should be used as a confirmatory technique when the diagnosis is unclear by other means. PCR analysis of blood specimens offers a minimally invasive approach to diagnosis, but clinical data are scarce and further studies are needed. As DNA microarrays move into the clinical arena, specific hybridization probes may allow improved identification and differentiation of Bartonellae at the molecular level.

Antibodies, Bacterial↗