Submitted to: Journal of Applied Microbiology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 10/18/2004
Publication Date: 4/1/2005
Citation: Cyr, T.L., Jenkins, M.C., Gilborn, A., Masters, E.J., Mcdonald, G.A. 2005. Improving the specificity of 16s-rDNA-based polymerase chain reaction for detecting Borrelia burgdoferi sensu lato-causative agents of human Lyme disease. Journal of Applied Microbiology. 98:962-970. Interpretive Summary: Lyme disease, the most common tick-borne disease in North America, Europe and Asia, is a debilitating, multi-systemic illness caused by various genospecies of the spirochete, Borrelia burgdorferi. Prompt treatment with antibiotics during early stages of the disease affords complete recovery with less direct and indirect medical costs. However, later stages, which can occur months or years after the initial infection and may involve debilitating chronic arthritis or chronic neurological effects, are more difficult to treat, and frequently result in significantly higher medical costs. For this reason, a prompt and accurate diagnosis is critical. Although cell culture is a definitive method for identifying Borrelia, this technique is time-consuming and often results in false negatives. In this study, the sequences of B. burgdorferi coding for ribosomal RNA were aligned with similar sequences of other Lyme and non-Lyme disease causing Borrelia in order to identify PCR primers that might be used to specifically identify agents of human Lyme Disease. A primer pair termed Bbsl was developed and used to amplify strains of B. burgdorferi, B. afzelii, and B. garinii, but not species of Borrelia that cause relapsing fever. The Bbsl primer pair was used to amplify B. burgdorferi from skin biopsies of patients with Lyme disease symptoms as well as from field-collected deer ticks, Lonestar ticks and American dog ticks.
Technical Abstract: Differentiating Borrelia burgdorferi sensu lato from non-Lyme disease causing Borrelia species has relied on heterogeneity of various DNA sequences, including those coding for 16S-ribosomal RNA. In the present study, the 16S-rDNA sequences of B. burgdorferi sensu lato were aligned with the 16S-rDNA sequences of B. hermsii, B. turicatae, and B. lonestari in order to identify primers that might be used to specifically identify agents of human Lyme disease. An oligonucleotide sequence, designated TEC1, was shown, in combination with a previously developed primer (LD2), to amplify strains of B. burgdorferi sensu stricto, B. afzelii, and B. garinii, but not B. hermsii or B. turicatae. This primer pair, designated Bbsl, was used to amplify B. burgdorferi sensu lato from skin biopsies of patients with Lyme disease symptoms as well as from Ixodes scapularis, Amblyomma americanum and Dermacentor variabilis ticks.