Detection of Established Virulence Genes and Plasmids To Differentiate Borrelia burgdorferi Strains

Abstract

ABSTRACTBorrelia burgdorferisensu stricto is the major causative agent of Lyme disease in the United States, whileB. gariniiandB. afzeliiare more prevalent in Europe. The highly complex genome ofB. burgdorferiis comprised of a linear chromosome and a large number of variably sized linear and circular plasmids. Many plasmids of this spirochete are unstable during its culturein vitro. Given that many of theB. burgdorferivirulence factors identified to date are plasmid encoded, spirochetal plasmid content determination is essential for genetic analysis of Lyme pathogenesis. Although PCR-based assays facilitate plasmid profiling of sequencedB. burgdorferistrains, a rapid genetic content determination strategy for nonsequenced strains has not yet been described. In this study, we combined pulsed-field gel electrophoresis (PFGE) and Southern hybridization for detection of genes encoding known virulence factors,ribosomal RNA genespacer restriction fragment length polymorphismtypes (RSTs),ospCgroup determination, and sequencing of the variabledbpAandospCgenes. We show that two strains isolated from the same tick and both originally named N40 are in fact very distinct. Furthermore, we failed to detectbbk32, which encodes a fibronectin-binding adhesin, in one “N40” strain. Thus, two distinct strains that show different plasmid profiles, as determined by PFGE and PCR, were isolated from the same tick and vary in theirospCanddbpAsequences. However, both belong to group RST3B.