ena-STUDY-OXFORD UNIVERSITY-22-03-2018-06:58:12:306-640
MinION sequencing enables rapid whole genome assembly of Rickettsia typhi in a resource-limited setting
The infrastructure and costs associated with performing next-generation sequencing have been barriers to its introduction into resource-limited settings. Oxford Nanopore Technologies' (ONT) portable MinION sequencer overcomes many of these challenges to allow small scale sequencing projects with minimal laboratory and computing infrastructure. We performed whole-genome sequencing (WGS) of Rickettsia typhi, an important and neglected cause of fever across much of the tropics and subtropics. Sequencing libraries were prepared from extracted DNA following standard ONT protocols and sequenced on a laptop in our laboratory in Vientiane, Laos. Basecalling was performed subsequently using ONT's Albacore software and assembled locally using a bioinformatics pipeline. This generated a single contig assembly of 1,078,916bp. Frequent SNP and indel error rates, commonly seen with ONT sequencing data, resulted in multiple gene annotation errors. As a result, DNA was also sequenced in the United Kingdom using the Illumina platform to generate a polished genome. Only 3 previous R. typhi whole genome sequences exist. Comparison with these shows evidence of reductive genome evolution with no rearrangements and few differences between strains collected over a 90-year time period. This is the first WGS to be performed in Laos and the first WGS of an R. typhi sample collected in the last 50 years. We demonstrate the utility of portable sequencing platforms such as the MinION for carrying out next-generation sequencing projects in resource-limited settings.
The infrastructure and costs associated with performing next-generation sequencing have been barriers to its introduction into resource-limited settings. Oxford Nanopore Technologies' (ONT) portable MinION sequencer overcomes many of these challenges to allow small scale sequencing projects with minimal laboratory and computing infrastructure. We performed whole-genome sequencing (WGS) of Rickettsia typhi, an important and neglected cause of fever across much of the tropics and subtropics. Sequencing libraries were prepared from extracted DNA following standard ONT protocols and sequenced on a laptop in our laboratory in Vientiane, Laos. Basecalling was performed subsequently using ONT's Albacore software and assembled locally using a bioinformatics pipeline. This generated a single contig assembly of 1,078,916bp. Frequent SNP and indel error rates, commonly seen with ONT sequencing data, resulted in multiple gene annotation errors. As a result, DNA was also sequenced in the United Kingdom using the Illumina platform to generate a polished genome. Only 3 previous R. typhi whole genome sequences exist. Comparison with these shows evidence of reductive genome evolution with no rearrangements and few differences between strains collected over a 90-year time period. This is the first WGS to be performed in Laos and the first WGS of an R. typhi sample collected in the last 50 years. We demonstrate the utility of portable sequencing platforms such as the MinION for carrying out next-generation sequencing projects in resource-limited settings.