MUTATIONS IN BARTONELLA-BACILLIFORMIS GYRB CONFER RESISTANCE TO COUMERMYCIN A(1)

This study describes the first isolation and characterization of spont aneous mutants conferring natural resistance to an antibiotic for any Bartonella species. The Bartonella bacilliformis gyrB gene, which enco des the B subunit of DNA gyrase, was cloned and sequenced. The gyrB op en reading frame (ORF) is 2,079 bp and encodes a deduced amino acid se quence of 692 residues, corresponding to a predicted protein of simila r to 77.5 kDa. Sequence alignment indicates that B. bacilliformis GyrB is most similar to the GyrB protein from Bacillus subtilis (40.1% ami no acid sequence identity) and that it contains the longest N-terminal tail (52 residues) of any GyrB characterized to date. The cloned B. b acilliformis gyrB was expressed in an Escherichia coil S30 cell extrac t and was able to functionally complement a temperature-sensitive E. c oli Cou(r) gyrB mutant (strain N4177). We isolated and characterized s pontaneous mutants of B. bacilliformis resistant to coumermycin A(1), an antibiotic that targets GyrB. Sequence analysis of gyrB from 12 Cou (r) mutants of B. bacilliformis identified single nucleotide transitio ns at three separate loci in the ORF. The predicted amino acid substit utions resulting from these transitions are Gly to Ser at position 124 (Gly124-->Ser), Arg184-->Gln, and Thr214-->Ala or Thr214-->Ile, which are analogous to mutated residues found in previously characterized r esistant gyrB genes from Borrelia burgdorferi, E. coil, Staphylococcus aureus, and Haloferax sp. The Cou(r) mutants are three to five times more resistant to coumermycin A(1) than the wild-type parental strain.