Open AccessBBE02 Disruption Mutants of Borrelia burgdorferi B31 Have a Highly Transformable, Infectious Phenotype
Author(s) -
Hiroki Kawabata,
Steven J. Norris,
Haruo Watanabe
Publication year - 2004
Publication title -
infection and immunity
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.508
H-Index - 220
eISSN - 1070-6313
pISSN - 0019-9567
DOI - 10.1128/iai.72.12.7147-7154.2004
Subject(s) - borrelia burgdorferi , plasmid , biology , shuttle vector , infectivity , virulence , transformation (genetics) , mutant , microbiology and biotechnology , virology , dna , spirochaetaceae , homologous recombination , gene , vector (molecular biology) , genetics , recombinant dna , antibody , virus
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.