Early Lyme disease with spirochetemia - diagnosed by DNA sequencing

Sin Hang Lee,Jessie Williams,William Jones,Jay Walshon,Jessica S Vigliotti,Veronica S Vigliotti

doi:10.1186/1756-0500-3-273

Abstract

BackgroundA sensitive and analytically specific nucleic acid amplification test (NAAT) is valuable in confirming the diagnosis of early Lyme disease at the stage of spirochetemia.FindingsVenous blood drawn from patients with clinical presentations of Lyme disease was tested for the standard 2-tier screen and Western Blot serology assay for Lyme disease, and also by a nested polymerase chain reaction (PCR) for B. burgdorferi sensu lato 16S ribosomal DNA. The PCR amplicon was sequenced for B. burgdorferi genomic DNA validation. A total of 130 patients visiting emergency room (ER) or Walk-in clinic (WALKIN), and 333 patients referred through the private physicians' offices were studied. While 5.4% of the ER/WALKIN patients showed DNA evidence of spirochetemia, none (0%) of the patients referred from private physicians' offices were DNA-positive. In contrast, while 8.4% of the patients referred from private physicians' offices were positive for the 2-tier Lyme serology assay, only 1.5% of the ER/WALKIN patients were positive for this antibody test. The 2-tier serology assay missed 85.7% of the cases of early Lyme disease with spirochetemia. The latter diagnosis was confirmed by DNA sequencing.ConclusionNested PCR followed by automated DNA sequencing is a valuable supplement to the standard 2-tier antibody assay in the diagnosis of early Lyme disease with spirochetemia. The best time to test for Lyme spirochetemia is when the patients living in the Lyme disease endemic areas develop unexplained symptoms or clinical manifestations that are consistent with Lyme disease early in the course of their illness.

Highlights

A sensitive and analytically specific nucleic acid amplification test (NAAT) is valuable in confirming the diagnosis of early Lyme disease at the stage of spirochetemia
Nested polymerase chain reaction (PCR) followed by automated DNA sequencing is a valuable supplement to the standard 2-tier antibody assay in the diagnosis of early Lyme disease with spirochetemia
After confirming a 100% identities match with a unique specific DNA sequence for B. burgdorferi sensu lato 16S rDNA stored in the GenBank database using the online Basic Local Alignment Search Tool (BLAST), the molecular identification of the nested PCR product as a genomic DNA of B. burgdorferi was established beyond a reasonable doubt

Summary

Introduction

A sensitive and analytically specific nucleic acid amplification test (NAAT) is valuable in confirming the diagnosis of early Lyme disease at the stage of spirochetemia. The polymerase chain reaction (PCR) technologies for the study of the most conserved genospecies-specific Borrelia burgdorferi sensu lato16S ribosomal RNA gene, or 16S rDNA, have been used in epidemiology research [5,6]. We have recently refined this research tool with a nested PCR technology for DNA detection, followed by automated direct DNA sequencing for validation of the genospecies-specific B. burgdorferi sensu lato 16S rDNA in patient body fluids to further augment the sensitivity and specificity of the procedure as a clinical laboratory test [8]. This report summarizes our experience in using this routine clinical laboratory test for molecular diagnosis of B. burgdorferi spirochetemia in an endemic suburban town during a summer season

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