Evaluation of capture sequencing on the Oxford Nanopore MinION as a stand-alone molecular test for tick-borne diseases.

Molecular diagnostics of tick-borne diseases (TBD) require the capacity to target a broad range of microbial pathogens circulating in ticks, along with the high sensitivity needed for detection of low burden agents such as Borrelia burgdorferi. Next generation sequencing (NGS) can have broad applicability for TBD. However, insufficient sensitivity and large infrastructure requirements limit the utility of this method for diagnostics. The application of capture-based NGS onto the Oxford Nanopore instruments presents an alternative approach that can provide high sensitivity coupled with smaller laboratory footprint. We tested the utility of our capture probe panel targeting tick-borne agents, the TBDCapSeq, on the Oxford Nanopore MinION. In direct comparison to quantitative PCR, we examined 64 whole blood samples, including 41 from patients diagnosed with Lyme disease, 7 from patients with suspected Lyme disease without seroconversion, 12 from symptomatic individuals suspected of tick-borne disease and 4 healthy controls. TBDCapSeq was more sensitive than qPCR. All agents detected by qPCR were correctly identified by TBDCapSeq. However, in analyses of 41 samples from patients with Lyme disease, 7 were positive for B. burgdorferi DNA with TBDCapSeq versus 3 with qPCR. Furthermore, with TBDCapSeq we also recovered substantial portions of the genome essential for downstream phylogenetic analyses. Our study demonstrated the potential utility of capture sequencing for molecular TBD diagnostics.