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Bartonella bacilliformis and Bartonella henselae, the respective agents of Oroya fever and cat-scratch disease in humans, are known to produce bacteriophage-like particles (BLPs) that package 14 kbp segments of the host chromosome. Data from this study suggest that other Bartonella species including Bartonella quintana, Bartonella doshiae and Bartonella grahamii also contain similar BLPs, as evidenced by the presence of a 14 kbp extrachromosomal DNA element in their genomes, whereas Bartonella elizabethae and Bartonella clarridgeiae do not. A purification scheme utilizing chloroform, DNase I and centrifugation was devised to isolate BLPs from B. bacilliformis. Intact BLPs were observed by transmission electron microscopy and were round to icosahedral in shape and approximately 80 nm in diameter. RFLP and Southern blot analysis of BLP DNA from B. bacilliformis suggest that packaging, while non-selective, is less than the near-random packaging previously reported for the B. henselae phage. Data also suggest that the linear, double-stranded BLP DNA molecules have blunt ends with noncovalently closed termini. Packaging of the BLP DNA molecules into a protein coat appears to be closely related to nucleic acid synthesis, as unpackaged phage DNA is not detectable within the host cell. SDS-PAGE analysis of purified BLPs from B. bacilliformis showed three major proteins with apparent molecular masses of 32, 34 and 36 kDa; values that closely correspond to proteins found in B. henselae BLPs. Western blot analysis performed with patient convalescent serum showed that BLP proteins are slightly immunogenic in humans. To determine if BLPs contribute to horizontal gene transfer, mutants of B. bacilliformis were generated by allelic exchange with an internal fragment of the 16S-23S rDNA intergenic spacer region and a suicide vector construct, termed pKB1. BLPs from one of the resultant strains were able to package the mutagenized region containing the kanamycin-resistance cassette; however, numerous approaches and attempts at intraspecies transduction using these BLPs were unsuccessful.
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PMID:A bacteriophage-like particle from Bartonella bacilliformis. 1074 63

Bartonella bacilliformis is the causative agent of the biphasic human disease, Oroya fever. During the primary disease phase, up to 100% of the circulating erythrocytes can be parasitized and 80% lysed. During the secondary phase of this disease, bacterial invasion shifts to endothelial cells lining the vasculature. B. bacilliformis is transferred between human hosts by the sandfly, Lutzomyia verrucarum. To investigate the regulation of ialB by environmental cues signaling vector-to-host transmission; nuclease protection assays were performed to compare the amount of ialB mRNA in bacteria subjected to temperature shift, pH change, oxidative stress, or hemin limitation. The amount of ialB mRNA increased by 223-310% in acid-treated samples and decreased by 28-39% in base-treated samples as compared to bacteria kept at pH 7.2. B. bacilliformis samples showed a 56-63% and 74-80% decrease in ialB mRNA when shifted to 37 degrees C from growth temperatures of 20 and 30 degrees C, respectively. Oxidative stress (1 mM H(2)O(2)) and hemin limitation had no significant effect on mRNA levels. Determination of IalB protein amounts using SDS-PAGE and immunoblotting showed the greatest amounts of IalB under acidic conditions or at 20 degrees C. The least amount of IalB was synthesized under basic conditions or at 37 degrees C. The viability of wild-type B. bacilliformis under the various experimental culture conditions was determined and found not to affect ialB mRNA amounts in these experiments. Finally, we compared the survival of wild-type and ialB mutant B. bacilliformis and found no difference in the viability of these two strains, demonstrating that IalB does not aid bacterial survival under these conditions.
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PMID:Differential expression of the invasion-associated locus B (ialB) gene of Bartonella bacilliformis in response to environmental cues. 1266 41