A bacteriophage-like particle from Bartonella bacilliformis

Bartonella bacilliformis and Bartonella henselae, the respective agents of Oroya fever and cat-scratch disease in humans, are known to produce bacteri ophage-like particles (BLPs) that package 14 kbp segments of the host chrom osome. Data from this study suggest that other Bartonella species including Bartonella quintana, Bartonella doshiae and Bartonella grahamii also conta in 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 a nd centrifugation was devised to isolate BLPs from B. bacilliformis, intact BLPs were observed by transmission electron microscopy and were round to i cosahedral in shape and approximately 80 nm in diameter. RFLP and Southern blot analysis of BLP DNA from B. bacilliformis suggest that packaging, whil e non-selective, is less than the near-random packaging previously reported for the B. henselae phage. Data also suggest that the linear, double-stran ded BLP DNA molecules have blunt ends with noncovalently closed termini. Pa ckaging of the BLP DNA molecules into a protein coat appears to be closely related to nucleic acid synthesis, as unpackaged phage DNA is not detectabl e within the host cell. SDS-PAGE analysis of purified BLPs from B. bacillif ormis 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 show ed that BLP proteins are slightly immunogenic in humans. To determine if BL Ps contribute to horizontal gene transfer, mutants of B. bacilliformis were generated by allelic exchange with an internal fragment of the 16S-23S rDN A intergenic spacer region and a suicide vector construct, termed pKB1. BLP s from one of the resultant strains were able to package the mutagenized re gion containing the kanamycin-resistance cassette; however, numerous approa ches and attempts at intraspecies transduction using these BLPs were unsucc essful.