Single Core Genome Sequencing for Detection of both Borrelia burgdorferi Sensu Lato and Relapsing Fever Borrelia Species.

Sin Hang Lee,John Eoin Healy,John S Lambert

doi:10.3390/ijerph16101779

Abstract

Lyme disease, initially described as Lyme arthritis, was reported before nucleic-acid based detection technologies were available. The most widely used diagnostic tests for Lyme disease are based on the serologic detection of antibodies produced against antigens derived from a single strain of Borrelia burgdorferi. The poor diagnostic accuracy of serological tests early in the infection process has been noted most recently in the 2018 Report to Congress issued by the U.S. Department of Health and Human Services Tick-Borne Disease Working Group. Clinical Lyme disease may be caused by a diversity of borreliae, including those classified as relapsing fever species, in the United States and in Europe. It is widely accepted that antibiotic treatment of Lyme disease is most successful during this critical early stage of infection. While genomic sequencing is recognized as an irrefutable direct detection method for laboratory diagnosis of Lyme borreliosis, development of a molecular diagnostic tool for all clinical forms of borreliosis is challenging because a “core genome” shared by all pathogenic borreliae has not yet been identified. After a diligent search of the GenBank database, we identified two highly conserved segments of DNA sequence among the borrelial 16S rRNA genes. We further developed a pair of Borrelia genus-specific PCR primers for amplification of a segment of borrelial 16S rRNA gene as a “core genome” to be used as the template for routine Sanger sequencing-based metagenomic direct detection test. This study presented examples of base-calling DNA sequencing electropherograms routinely generated in a clinical diagnostic laboratory on DNA extracts of human blood specimens and ticks collected from human skin bites and from the environment. Since some of the tick samples tested were collected in Ireland, borrelial species or strains not known to exist in the United States were also detected by analysis of this 16S rRNA “core genome”. We recommend that hospital laboratories located in Lyme disease endemic areas begin to use a “core genome” sequencing test to routinely diagnose spirochetemia caused by various species of borreliae for timely management of patients at the early stage of infection.

Highlights

Lyme disease and related borreliosis transmitted by the bite of borrelia-infected ticks is the most common vector-borne infectious disease in the United States and in many European countries [1,2,3,4,5].It is generally believed that at the early stage of infection this disease can be treated successfully withInt
The goal of this study was to present our evidence-based data to demonstrate that a highly conserved Borrelia genus-specific segment of the 16S ribosomal RNA (rRNA) gene can serve as a “core genome” shared by all species and isolates of Borrelia, including members of the B. burgdorferi sensu lato complex and the more heterogeneous species of the relapsing fever borreliae which are known to be capable of infecting the ticks and humans in North America and in Europe [1,2,3,4,5] as well as in Africa [29]
After reviewing the alignments of the borrelial 16S rRNA gene sequences of the common species of Borrelia capable of causing human infections and after testing all the candidate primers synthesized, we identified a unique segment of Borrelia burgdorferi sensu lato 16S rRNA gene and the corresponding segment of 16S rRNA genes of various relapsing fever borreliae, including that of B. recurrentis, all defined by a pair of highly conserved 21-base DNA sequences

Summary

Introduction

Lyme disease and related borreliosis transmitted by the bite of borrelia-infected ticks is the most common vector-borne infectious disease in the United States and in many European countries [1,2,3,4,5].It is generally believed that at the early stage of infection this disease can be treated successfully withInt. Public Health 2019, 16, 1779; doi:10.3390/ijerph16101779 www.mdpi.com/journal/ijerph

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