BBE02 disruption mutants of Borrelia burgdorferi B31 have a highly transformable, infectious phenotype.
We constructed highly transformable and infectious Borrelia burgdorferi B31 by inactivating BBE02, a putative restriction-modification gene on the linear plasmid lp25. The low-passage-number B31 clones 5A4 (containing all plasmids) and 5A18 (lp28-4(-) lp56(-)) were used for this study, and BBE02 was disrupted by homologous recombination. The transformation efficiency with the shuttle vector pBSV2C03::gntDeltakan was increased from <1 to approximately 10 colonies per mug of DNA for 5A4 and 5A4 BBE02::Kan(r) and from 14 to approximately 600 colonies per mug of DNA for 5A18 and 5A18 BBE02::Kan(r). lp25, which is required for infectivity in mice, was retained in BBE02 mutants transformed with pBSV2C03::gntDeltakan, but lp25 was not detected in transformants of the parental clones 5A4 and 5A18. BBE02 disruptants and pBSV2C03::gntDeltakan transformants of these clones remained infectious in C3H/HeN mice, and the 50% infective doses of the BBE02 disruptants were <10(2) organisms per mouse. The inactivation of BBE02 thus eliminates a transformation barrier for infectious B. burgdorferi B31 and will provide a valuable tool for studying the virulence factors of Lyme disease.
Kawabata
Hiroki
H
Department of Bacteriology, National Institute of Infectious Diseases, Toyama 1-23-1, Shinjuku-ku, Tokyo 162-8640, Japan. kbata@nih.go.jp
Norris
Steven J
SJ
Watanabe
Haruo
H
D000818:Animals; D001426:Bacterial Proteins / Q000502:physiology*; D025065:Borrelia burgdorferi / Q000235:genetics* / Q000472:pathogenicity; D005260:Female; D007596:Joints / Q000473:pathology; D008193:Lyme Disease / Q000209:etiology / Q000473:pathology; D051379:Mice; D008809:Mice, Inbred C3H; D009154:Mutation; D009535:Nicotinamidase / Q000235:genetics / Q000502:physiology*; D010641:Phenotype; D014169:Transformation, Bacterial; D037521:Virulence Factors / Q000502:physiology
D001426:Bacterial Proteins; D037521:Virulence Factors; D009535:Nicotinamidase