Isolation and phylogenetic analysis of Bartonella species from Rusa deer (Rusa timorensis) in Thailand.

The aim of the present study is to investigate the prevalence of Bartonella infection in deer in Thailand and to characterize the isolates by biochemical, morphological and genetic analysis. A total of 247 blood samples were collected from Rusa deer (Rusa timorensis) in a livestock breeding facility in Thailand. Bartonella bacteria were isolated in 3.6% of the blood samples. Three out of 110 (2.7%) males and 6 of 137 (4.4%) females were positive for Bartonella. A higher prevalence of Bartonella was observed in young deer under 4 years of age compared to adults over 4 years of age, but no Bartonella was isolated from deer over 8 years of age. Phylogenetic analysis of concatenated sequences of seven loci of Bartonella indicated that all the isolates from Rusa deer in Thailand were identical and formed a distinct cluster from other known Bartonella species.

Bartonella dromedarii sp. nov. isolated from domesticated camels (Camelus dromedarius) in Israel.

Bartonella spp. are fastidious, Gram-negative bacilli that cause a wide spectrum of diseases in humans. Most Bartonella spp. have adapted to a specific host, generally a domestic or wild mammal. Dromedary camels (Camelus dromedarius) have become a focus of growing public-health interest because they have been identified as a reservoir host for the Middle East respiratory syndrome coronavirus. Nevertheless, data on camel zoonoses are limited. We aimed to study the occurrence of Bartonella bacteremia among dromedaries in Israel. Nine of 51 (17.6%) camels were found to be bacteremic with Bartonella spp.; bacteremia levels ranged from five to >1000 colony-forming units/mL. Phylogenetic reconstruction based on the concatenated sequences of gltA and rpoB genes demonstrated that the dromedary Bartonella isolates are closely related to other ruminant-derived Bartonella spp., with B. bovis being the nearest relative. Using electron microscopy, the novel isolates were shown to be flagellated, whereas B. bovis is nonflagellated. Sequence comparisons analysis of the housekeeping genes ftsZ, ribC, and groEL showed the highest homology to B. chomelii, B. capreoli, and B. birtlesii, respectively. Sequence analysis of the gltA and rpoB revealed ∼96% identity to B. bovis, a previously suggested cutoff value for sequence-based differentiation of Bartonella spp., suggesting that this approach does not have sufficient discriminatory power for differentiating ruminant-related Bartonella spp. A comprehensive multilocus sequence typing (MLST) analysis based on nine genetic loci (gltA, rpoB, ftsZ, internal transcribed spacer (ITS), 16S rRNA, ribC, groEL, nuoG, and SsrA) identified seven sequence types of the new dromedary isolates. This is the first description of a Bartonella sp. from camelids. On the basis of a distinct reservoir and ecological niche, sequence analyses, and expression of flagella, we designate these isolates as a novel Bartonella sp. named Bartonella dromedarii sp. nov. Further studies are required to explore its zoonotic potential.

Identification of Bartonellae in the soft tick species Ornithodoros sonrai in Senegal.

Ticks, belonging to the soft ticks species Ornithodorus sonrai, have been collected from six sites in Senegal and were tested for the presence of Bartonella spp. Initial screening by PCR revealed the presence of these bacteria in ticks from two villages, Soulkhou Thissé (5/8, 62.5%) and Maka Gouye (1/24, 4.2%). Three bacterial strains were isolated from live ticks, and the genetic characterization of these strains suggests that they belong to two previously unknown species. The pathogenicity of these two new species of Bartonella is not yet known. The new isolates described here are the first strains of Bartonella spp. from soft ticks and the first isolates from any arthropod species in Africa.

Identification of Bartonella Trw host-specific receptor on erythrocytes.

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

Isolation & characterization of Bartonella sp. from optic neuritis patients.

BACKGROUND & OBJECTIVES: Optic neuritis (ON) is characterized by sudden and rapid impairment of vision. Bartonella henselae is a known aetiological agent of cat scratch disease (CSD), which is a common cause of neuroretinitis, the least common type of optic neuritis. The present study was carried out to determine the microbiological aetiology of optic neuritis in patients attending a tertiary care eye hospital in north India, which was later confirmed with molecular characterization. METHODS: Of the 50 patients suffering from optic neuritis reported to the Ophthalmology OPD of a tertiary care eye hospital in New Delhi, India, 29 were included in the study. Blood culture from these patients were processed for aerobic and anerobic cultures to rule out infective aetiology. Subsequently, PCR was done on archive, glycerol-stocked cultures. RESULTS: Gram-negative pleomorphic coccobacilli grew in four of 29 patients tested. Characterization of these revealed Bartonella like organism as tested by the API 20E, API Staph, API Strept and RapID ANA systems. Electron microscopy revealed presence of polar flagella and bleb like projection all over the bacterial surface. PCR performed on preserved culture confirmed these as Bartonella sp. INTERPRETATION & CONCLUSIONS: Infections with Bartonella like organisms have not been demonstrated from India in cases of optic neuritis or in any of the other clinical syndromes in the past. The present study shows the isolation and characterization of Bartonella like organisms from optic neuritis patients. From clinical point of view it will be important to look for these organisms as aetiological agents in ON cases in order to treat with appropriate antibiotics.

Bartonella tamiae sp. nov., a newly recognized pathogen isolated from three human patients from Thailand.

Three strains of a novel Bartonella species (Bartonella tamiae) were isolated from human patients from Thailand. Sequence analysis of six chromosomal regions (16S rRNA, gltA, groEL, ftsZ, rpoB, and the intergenic spacer region) and phenotypical analysis supported the similarity of the three strains and placed them within the genus Bartonella separately from previously described species.

Bacillary angiomatosis in HIV-infected patients--an epidemiological and clinical study.

BACKGROUND: No data were available on the epidemiological and clinical characteristics of bacillary angiomatosis (BA) in Germany. OBJECTIVE: To determine epidemiological and clinical data on HIV-associated BA. METHODS: A chart review of all BA cases between 1990 and 1998 was performed in 23 German AIDS treatment units. RESULTS: A total of 21 cases of BA was diagnosed. During this period, the participating HIV centers treated about 17,000 HIV-infected patients. As a result, a BA prevalence of 1.2 cases/1,000 patients can be assumed. 19 BA were localized in the skin; in 5 cases bones and in 4 cases the liver were involved. Out of 20 patients who received antibiotic therapy, 13 had complete remission. The median time of duration up to complete remission was 32 days (9-82). During the follow-up of the 20 patients, 7 relapses were observed. CONCLUSION: BA is a rare HIV-associated disease with a prevalence of 1,2 cases/1,000 patients in the presented study.

Bartonellosis in Ecuador: serosurvey and current status of cutaneous verrucous disease.

Human bartonellosis is a classically biphasic disease caused by infection with the alpha-2 Proteobacteria Bartonella bacilliformis, which is phylogenetically related to the etiologic agents of cat scratch disease, bacillary angiomatosis, and trench fever. In Ecuador, typical bartonellosis has remained endemic for the past century in highland provinces near the Peruvian border. During the past six years, public health officials have noted an increasing number of atypical cases in which monophasic verrucous cutaneous disease is the only clinical manifestation. Epidemiologic, immunologic, histopathologic, and molecular biological studies have confirmed the presence of sporadic, atypical bartonellosis in residents of the lowland province of Manabi, where archeologic evidence exists of bartonellosis in pre-Colombian times. Between 1987 and 1995, 11 cases of cutaneous bartonellosis were investigated and serologic studies were done on 224 persons from five villages, two lowland and three highland. In the lowland village of Pajan in the province of Manabi, there was a 21% seropositivity proportion in contacts of index cases. These combined data suggest that bartonellosis is significantly under-reported due to the existence of mild clinical disease, possibly associated with less virulent bacterial strains, which are now disseminating or re-emerging in previously disease-free areas.

Bartonellosis and human immunodeficiency disease (AIDS): L-forms as persisters, activating factors, and mechanism of disease.

Bartonella, genus Proteus, can cause immunodepressive disease. The organisms, in parasitized red blood cells, may invade the brain and every other system and space in the human body. Bartonella henselae is proposed to have a role in the pathogenesis of acquired immunodeficiency syndrome (AIDS) encephalopathy. Bartonella bacilliformis produces two known toxins that can induce spasm and angiomatosis, respectively, and manifest as diseases associated with symptomatic AIDS. The skin lesions of bartonellosis may be mistaken clinically and histologically for Kaposi's sarcoma. Bacteria of the genus Proteus produce L-forms: their elementary bodies may be mistaken for what are called the 'human immunodeficiency viruses' (HIV). Antibiotics, especially penicillin, induce bacteria to produce L-forms. Air pollution and high sugar, salt and fat diets are factors that may increase the lipid content of microbes that produce toxins and L-forms that may persist or revert to bacterial form.

Bartonella spp. as emerging human pathogens.

Members of the genus Bartonella (formerly Rochalimaea) were virtually unknown to modern-day clinicians and microbiologists until they were associated with opportunistic infections in AIDS patients about 6 years ago. Since that time, Bartonella species have been associated with cat scratch disease, bacillary angiomatosis, and a variety of other disease syndromes. Clinical presentation of infection with Bartonella ranges from a relatively mild lymphadenopathy with few other symptoms, seen in cat scratch disease, to life-threatening systemic disease in the immunocompromised patient. In some individuals, infection manifests as lesions that exhibit proliferation of endothelial cells and neovascularization, a pathogenic process unique to this genus of bacteria. As the spectrum of disease attributed to Bartonella is further defined, the need for reliable laboratory methods to diagnose infections caused by these unique organisms also increases. A brief summary of the clinical presentations associated with Bartonella infections is presented, and the current status of laboratory diagnosis and identification of these organisms is reviewed.

Cell entry and the pathogenesis of Bartonella infections.

Erythrocyte parasitism, or hemotrophy, is not a common strategy for bacteria. However, Bartonella species are elegantly adapted to parasitize a variety of cell types including red blood cells. Bartonella bacilliformis, a useful model organism for the genus, has been used to study virulence determinants involved in colonization, attachment and invasion of host cells.

Endocarditis in a dog due to infection with a novel Bartonella subspecies.

Vegetative valvular endocarditis involving the aortic and, to a lesser extent, mitral valves was diagnosed echocardiographically in a 3-year-old spayed female Labrador retriever. Historically, the dog had been treated with tetracycline hydrochloride and prednisolone for positive seroreactivity to Ehrlichia canis and antinuclear antigens. Although three aerobic and anaerobic blood cultures failed to grow bacteria, blood cultured simultaneously by the lysis centrifugation technique grew a fastidious, gram-negative organism. Despite an initial therapeutic response, the owner elected euthanasia 17 days later. Necropsy confirmed aortic and mitral valvular endocarditis. Bacteria phenotypically similar to Bartonella species were visualized in the heart valve by light and electron microscopy, and Bartonella DNA from a frozen heart valve was amplified by PCR. Subsequent phenotypic and genotypic characterization of the isolate, including biochemical testing, cellular fatty acid analysis, DNA hybridization, and sequencing of the 16S rRNA gene indicated that this organism, which can induce endocarditis in dogs, is a novel Bartonella subspecies containing an insertion sequence unique among currently recognized Bartonella species. The name Bartonella vinsonii subsp. berkoffii subsp. nov. will be proposed for this organism.

Histological and ultrastructural study of one case of oral bacillary angiomatosis in HIV disease and review of the literature.

Bacillary angiomatosis (BA) is a new clinicopathological entity defined as a pseudo-neoplastic capillary proliferation secondary to an opportunistic infection by one of two Rochalimaea sp.: R. quintana or R. henselae. Although BA is a recently recognised entity, numerous cases have been reported. Most of the patients affected are reported to have low absolute CD4 lymphocyte counts associated with AIDS. Yet, very few oral cases associated or not with cutaneous lesions have been reported or simply identified. Histopathological and ultrastructural features of one case of oral BA with gingival and palatal lesions are presented. Clinical aspects of oral BA do not hold pathognomonic features and the lesions may resemble either a reactive lesion of the gingiva, pyogenic granuloma or Kaposi's sarcoma. The lesion is characteristically composed of circumscribed lobular capillary proliferations and the presence of granular amphophilic material on haematoxylin and eosin sections surrounded by neutrophils and neutrophilic debris is a clue to diagnosis. Demonstration of bacilli in the interstitium by the Warthin-Starry silver method or, better, by electron microscopy is diagnostic. BA may contribute to the death of the patient but erythromycin has proved to be very effective treatment.

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.

Interactions between live Bartonella bacilliformis and endothelial cells.

Bartonella bacilliformis, a gram-negative, flagellated, motile bacterium, is the etiologic agent of verruca peruana. It is found within the verruca, where it can form large cytoplasmic (Rocha-Lima) inclusions in endothelial cells. Previously, an activity has been described in homogenates of B. bacilliformis that in vitro increases the proliferation of endothelial cells and their production of tissue-type plasminogen activator (t-PA) and in vivo is angiogenic. The aim of the present study was to determine if live B. bacilliformis similarly stimulated endothelial cells and produced the Rocha-Lima inclusion. By measuring proliferation of cells and the production of t-PA in vitro, it was found that the live bacteria increased both parameters in a fashion similar to the homogenates of B. bacilliformis. Interaction between the bacteria and endothelial cells appeared to be necessary for proliferation. On electron microscopy, bacteria penetrated the endothelial cell within 1 h, forming a small membrane-bound inclusion. By 12 h, a large membrane-bound inclusion, similar to the Rocha-Lima inclusion, containing numerous bacteria was present. These data provide further evidence that B. bacilliformis has an angiogenic activity and that the bacteria are at least in part responsible for the vascular proliferation of the verruca.

A phage in Bartonella bacilliformis.

Bacteriophage-like particles were found in Bartonella bacilliformis culture. The particles consisted of head (icosahedral), 40 nm in diameter, and tail, 16 nm in length.

Bb65, a major immunoreactive protein of Bartonella bacilliformis.

A 65 kDa protein (Bb65) has been identified as one of the major specific antigens of Bartonella bacilliformis, the causative agent of bartonellosis which is a bacterial infectious disease of inhabitants of the Andes. The gene encoding this antigen (7B2) was isolated from an expression library made directly from randomly generated fragments of B. bacilliformis genomic DNA using Bartonella antibodies raised in rabbits and sera of bartonellosis patients. The Bartonella 7B2 gene was expressed in Escherichia coli and the recombinant Bb65 protein was purified by column chromatography. Using polyclonal antibodies raised in rabbits, the antigen was shown to be present in all of 13 B. bacilliformis isolates from different Peruvian regions. Immune electron microscopy demonstrated the probable cytoplasmatic localization of Bb65. When applied to enzyme immunoassays, Bb65 sensitively and specifically bound to IgG antibody of sera of bartonellosis patients, convalescents, and immunes from various Peruvian regions. IgM antibody was not recognized by Bb65, neither was IgG antibody circulating during the first 2 weeks of illness. The amino-terminal amino acid sequence of Bb65 was 53% homologous to the 65 kDa heat shock protein of Mycobacterium tuberculosis.

Common surface epitope of Bartonella bacilliformis and Chlamydia psittaci.

A serosurvey revealed intense cross-reactivity between Bartonella bacilliformis and Chlamydia psittaci. One of the cross-reacting Bartonella antigens was identified as lipopolysaccharide which reacted with Bartonella as well as with Chlamydia serum antibodies. A monoclonal Bartonella antibody bound to Bartonella lipopolysaccharide as well as to the surfaces of Bartonella bacilliformis and Chlamydia psittaci. It was thus demonstrated that Chlamydia psittaci carries a surface epitope identical to an epitope of Bartonella lipopolysaccharide. The lipopolysaccharide was preliminarily characterized by polyacrylamide gel electrophoresis and by a lectin-binding assay. The lipopolysaccharides of Bartonella bacilliformis and Chlamydia psittaci are not identical.

Verruga peruana mimicking malignant neoplasms.

We have shown that in some cases fully developed florid verruga peruana nodules, as well as late-resolving, deeply situated lesions, can histologically suggest a variety of tumors to experienced pathologists. The compact proliferation of endothelial cells characteristic of florid verruga lesions can give rise to two pseudoneoplastic histologic patterns. One consists of sheets or islands of cells arranged in an epithelioid or pseudoepithelioid pattern (cases 1 and 2) in which the following histologic diagnoses were considered: squamous carcinoma, sweat gland carcinoma, epithelioid hemangioendothelioma, epithelioid sarcoma, melanoma and metastatic carcinoma. The other pattern of the florid lesion is characterized by a predominantly spindle cell arrangement (case 3) and in it the following diagnoses were made: Kaposi's sarcoma, fibrosarcoma, melanoma and leiomyosarcoma. The dense lymphoplasmacytic and histiocytic infiltrates of the late-resolving, deeply situated nodules (case 4) suggested the following histologic conditions: malignant lymphoma, nodular Hodgkin's disease, reticuloendotheliosis, and reticulosarcomatosis. Some histologic features thought to be of value to help in the differential diagnoses are discussed. It is emphasized that only the finding of Rocha-Lima's inclusions on light-microscopic studies and/or the demonstration of bartonella organisms in the lesions by electron-microscopic studies can objectively establish a diagnosis in a given lesion. However, knowledge of the epidemiologic data and particularly the presence of other lesions in the patient make it relatively easy to rule out neoplasia.