Detection of <i>Borrelia burgdorferi </i>Cell-Free DNA in Human Plasma Samples for Improved Diagnosis of Early Lyme Borreliosis

Abstract

Background: Erythema migrans (EM), the skin rash of early Lyme borreliosis (LB), is an insensitive and non-specific physical finding. Confirmation of suspected cases is challenging, as serology and blood PCR are insensitive in early LB. Here, we demonstrate that detection of Borrelia burgdorferi (B.b.) cell-free DNA (cfDNA) in plasma can improve diagnosis of early LB. Methods: We compared B.b.cfDNA detection in plasma samples with serology and blood PCR in 40 patients with physician-diagnosed EM, 28 of whom were confirmed to have LB by skin biopsy culture (N=18), seroconversion (N=2) or both (N=8). Findings: B.b.cfDNA was detected in 18/28 patients (64%) with laboratory-confirmed EM. In comparison, sensitivity of acute-phase serology using modified two-tiered testing (MTTT) was 50% (P=0.45); sensitivity of blood PCR was 7% (P=0.0002). Combining B.b.cfDNA detection and MTTT increased diagnostic sensitivity to 86%, significantly higher than either approach alone (P < or = 0.04). B.b.cfDNA sequences matched precisely with strain-specific sequence generated from the same individual’s cultured B.b. isolate. B.b.cfDNA was not detected in plasma from 684 asymptomatic ambulatory individuals. Among 3000 hospitalized patients tested as part of clinical care, B.b.cfDNA was detected in only two individuals, both of whom had clinical presentations consistent with LB. Interpretation: This is the first report of B.b.cfDNA detection in early LB and a demonstration of potential diagnostic utility. The combination of B.b.cfDNA and acute-phase MTTT improves clinical sensitivity for diagnosis of early LB. Funding Statement: This work represents an unfunded collaboration between the academically-affiliated authors and Karius Inc. Declaration of Interests: JAB has received research support from Zeus Scientific, bioMerieux, Immunetics, Alere, DiaSorin, the Bay Area Lyme Foundation (BALF), and the National Institute of Allergy and Infectious Diseases (NIAID; Award 1R21AI119457-01) for LB-related research projects. NRP has received funding from BALF and NIAID (Award 1R21AI119457-01) for LB-related research. JEL has received payments from Sherlock Biosciences for consulting services. LB, AAA, DKH, SB, TB, DH and CH are employed by Karius, Inc. All other authors declare no competing interests. Ethics Approval Statement: The parent and current studies were approved by the Partners Healthcare Human Research Committee.