Oxford Nanopore MinION Sequencing Enables Rapid Whole Genome Assembly of Rickettsia typhi in a Resource Limited Setting
Abstract. The infrastructure challenges and costs of next generation sequencing have been largely overcome, for many sequencing applications, by Oxford Nanopore Technologies' portable MinION sequencer. However, the question remains open whether MinION based bacterial whole genome sequencing is by itself sufficient for the accurate assessment of phylogenetic and epidemiological relationships between isolates and whether such tasks can be undertaken in resource limited settings. To investigate this question, we sequenced the genome of an isolate of Rickettsia typhi, an important and neglected cause of fever across much of the tropics and subtropics, for which only three genomic sequences previously existed. We prepared and sequenced libraries on a MinION in Vientiane, Lao PDR, using v9.5 chemistry, and in parallel, we sequenced the same isolate on the Illumina platform in a genomics laboratory in the United Kingdom. The MinION sequence reads yielded a single contiguous assembly, in which the addition of Illumina data revealed 226 base substitution and 5,856 indel errors. The combined assembly represents the first complete genome sequence of a human R. typhi isolate collected in the last 50 years and differed from the genomes of existing strains collected over a 90 year time period at very few sites, with no rearrangements. Filtering based on the known error profile of MinION data improved the accuracy of the nanopore only assembly. However, the frequency of false positive errors remained greater than true sequence divergence from recorded sequences. Although nanopore only sequencing cannot yet recover phylogenetic signals in R. typhi, such an approach may be applicable for more diverse organisms.
The American journal of tropical medicine and hygiene