Direct molecular detection and genotyping of Borrelia burgdorferi from whole blood of patients with early Lyme disease.

Direct molecular tests in blood for early Lyme disease can be insensitive due to low amount of circulating Borrelia burgdorferi DNA. To address this challenge, we have developed a sensitive strategy to both detect and genotype B. burgdorferi directly from whole blood collected during the initial patient visit. This strategy improved sensitivity by employing 1.25 mL of whole blood, a novel pre-enrichment of the entire specimen extract for Borrelia DNA prior to a multi-locus PCR and electrospray ionization mass spectrometry detection assay. We evaluated the assay on blood collected at the initial presentation from 21 endemic area patients who had both physician-diagnosed erythema migrans (EM) and positive two-tiered serology either at the initial visit or at a follow-up visit after three weeks of antibiotic therapy. Results of this DNA analysis showed detection of B. burgdorferi in 13 of 21 patients (62%). In most cases the new assay also provided the B. burgdorferi genotype. The combined results of our direct detection assay with initial physician visit serology resulted in the detection of early Lyme disease in 19 of 21 (90%) of patients at the initial visit. In 5 of 21 cases we demonstrate the ability to detect B. burgdorferi in early Lyme disease directly from whole blood specimens prior to seroconversion.

Prospective study on the incidence of infection by Borrelia burgdorferi sensu lato after a tick bite in a highly endemic area of Switzerland.

The periurban forest of Neuchâtel (Switzerland) is a high-risk area for Lyme Borreliosis, due to a high density of infected Ixodes ricinus ticks. In this study, we evaluated the risk of subclinical (seroconversion) and clinical infection after a tick bite in Neuchâtel inhabitants from 2003 to 2005. Inhabitants have been invited, through media, to visit a physician after a tick bite. A questionnaire was filled out and two blood samples were taken at 8-week interval. EIA screening tests for IgM and IgG (IMX system, Abbott) were applied for paired sera. In case of a change in antibody titres between both samples, a homemade Western-blot using Borrelia afzelii, B. burgdorferi sensu stricto and B. garinii as antigens was performed. Participants were included into two groups. Group one included asymptomatic participants (n=255). Among them, nine (3.5%) seroconverted with seroconversion rates varying between 6.8% in 2003, 2.1% in 2004 and 2.3% in 2005. Participants who developed clinical symptoms of LB were included into group two (n=14). Erythema migrans (EM) was reported in 5.2% of participants (5.2%), varying between 7.5% in 2003, 5% in 2004 and 3.4% in 2005. Ticks obtained from 186 participants were examined for B. burgdorferi infection by PCR/Reverse Line Blotting, and by Real Time PCR and tick attachment duration was estimated. Among I. ricinus ticks collected from participants, 32.8% were infected by B. burgdorferi sensu lato. B. afzelii predominated among these ticks. Globally, 65.9% of nymphs remained attached for more than 24h whereas only 38.3% of female ticks remained attached for more than 24h. We observed that 6.6% and 2.4% of participants bitten by infected and uninfected ticks, respectively, developed EM.