Surveillance for and Discovery of Borrelia Species in US Patients Suspected of Tickborne Illness

Abstract

Background: Tick-transmitted Borrelia fall into 2 heterogeneous bacterial complexes comprised of multiple species, the relapsing fever (RF) group and the Borrelia burgdorferi sensu lato group, which are the causative agents of Lyme borreliosis (LB), the most common tickborne disease in the Northern Hemisphere. Geographic expansion of LB in the United States and discovery of emerging Borrelia pathogens underscores the importance of surveillance for disease-causing Borrelia.

Methods: De-identified clinical specimens, submitted by providers throughout the United States, for patients suspected of LB, anaplasmosis, ehrlichiosis, or babesiosis were screened using a Borrelia genus-level TaqMan polymerase chain reaction (PCR). Borrelia species and sequence types (STs) were characterized by multilocus sequence typing (MLST) utilizing next-generation sequencing.

Results: Among 7292 specimens tested, 5 Borrelia species were identified: 2 causing LB, B. burgdorferi (n = 25) and B. mayonii (n = 9), and 3 RF borreliae, B. hermsii (n = 1), B. miyamotoi (n = 8), and Candidatus B. johnsonii (n = 1), a species previously detected only in the bat tick, Carios kelleyi. ST diversity was greatest for B. burgdorferi-positive specimens, with new STs identified primarily among synovial fluids.

Conclusions: These results demonstrate that broad PCR screening followed by MLST is a powerful surveillance tool for uncovering the spectrum of disease-causing Borrelia species, understanding their geographic distribution, and investigating the correlation between B. burgdorferi STs and joint involvement. Detection of Candidatus B. johnsonii in a patient with suspected tickborne disease suggests this species may be a previously undetected cause of illness in humans exposed to bat ticks.

Figure 1. Origin of clinical specimens from patients suspected of tickborne illness

Origin of 7,292 clinical specimens tested in this study. Differential shading indicates the number of patient specimens originating from each state.

Figure 2. Phylogenetic relationships of identified Borrelia species

The phylogenetic tree is based on alignment of in frame concatenated DNA sequence fragments from 8 housekeeping genes (clpA, clpX, nifS, pepX, pyrG, recG, rplB, and uvrA) (n = 4,791 bp). Strains sequenced in this study are highlighted as follows: blue, B. burgdorferi; purple, B. mayonii; red, B. miyamotoi; pink, B. hermsii, and orange, Candidatus B. johnsonii. Publicly available sequences for 18 different Bbsl genospecies and 10 different RF borreliae (gray) were included for comparison. The scale bar corresponds to 0.1 substitutions per nucleotide position.

Figure 3. Identification of Candidatus Borrelia johnsonii in patient blood

Phylogenetic relationship of Borrelia spp. 15-3581 (red) based on analysis of concatenated glpQ, flaB, and 16S rDNA sequences (n = 3048 bp). Publicly available sequences for 6 different RF borreliae (gray) and B. hermsii 15-5636 from this study were included for comparison. Bootstrap support values greater than 50% are shown. The scale bar corresponds to 0.01 substitutions per nucleotide position.