Loss of plasmids of Borrelia burgdorferi sensu lato during prolonged in vitro cultivation.
ABSTRACT In the present study we analyzed stability of plasmid content in 34 Borrelia strains of three different species (13 Borrelia afzelii, 10 Borrelia garinii and 11 Borrelia burgodorferi sensu stricto) using pulse field gel electrophoresis (PFGE). During long-term in vitro cultivation consisting of 50 passages, plasmid loss was established in 46% of B. afzelii, 40% of B. garinii and 36% of B. burgdorferi sensu stricto strains. Loss of plasmids occurred as early as between the 5th and 10th passage, affected only plasmids in the range 9-41 kb but not plasmids in the range 50-68 kb and manifested with the loss of one to up to three plasmids.
Article: Detection of established virulence genes and plasmids to differentiate Borrelia burgdorferi strains.[show abstract] [hide abstract]
ABSTRACT: Borrelia burgdorferi sensu stricto is the major causative agent of Lyme disease in the United States, while B. garinii and B. afzelii are more prevalent in Europe. The highly complex genome of B. burgdorferi is 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 culture in vitro. Given that many of the B. burgdorferi virulence 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 sequenced B. burgdorferi strains, 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 gene spacer restriction fragment length polymorphism types (RSTs), ospC group determination, and sequencing of the variable dbpA and ospC genes. We show that two strains isolated from the same tick and both originally named N40 are in fact very distinct. Furthermore, we failed to detect bbk32, 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 their ospC and dbpA sequences. However, both belong to group RST3B.Infection and immunity 01/2012; 80(4):1519-29. · 4.21 Impact Factor
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ABSTRACT: With cases now documented in every province, Lyme borreliosis (LB) is emerging as a serious public health risk in Canada. Controversy over the contribution of LB to the burden of chronic disease is maintained by difficulty in capturing accurate Canadian statistics, especially early clinical cases of LB. The use of dogs as sentinel species demon-strates that potential contact with Borrelia burgdorferi spirochetes, as detected by C6 peptide, extends across the country. Dissemination of infected ticks by migratory birds and rapid establishment of significant levels of infection have been well described. Canadian public health response has focused on identification of established populations of the tick vectors, Ixodes scapularis and I. pacificus, on the assumption that these are the only important vectors of the disease across Canada. Strains of B. burgdorferi circulating in Canada and the full range of their reservoir species and coinfections remain to be explored. Ongoing surveys and historical records demonstrate that Borrelia-positive Ixodes species are regu-larly present in regions of Canada that have previously been considered to be outside of the ranges of these species in re-cent modeling efforts. We present data demonstrating that human cases of LB are found across the nation. Consequently, physician education and better early diagnoses are needed to prevent long term sequelae. An international perspective will be paramount for developing improved Canadian guidelines that recognize the complexity and diversity of Lyme borreliosis.The Open Neurology Journal 01/2012; 6:94-103.