The Ebola virus disease epidemic in West Africa is the largest on record, responsible for over 28,599 cases and more than 11,299 deaths1. Genome sequencing in viral outbreaks is desirable to characterize the infectious agent and determine its evolutionary rate. Genome sequencing also allows the identification of signatures of host adaptation, identification and monitoring of diagnostic targets, and characterization of responses to vaccines and treatments. The Ebola virus (EBOV) genome substitution rate in the Makona strain has been estimated at between 0.87 × 10−3 and 1.42 × 10−3 mutations per site per year. This is equivalent to 16–27 mutations in each genome, meaning that sequences diverge rapidly enough to identify distinct sub-lineages during a prolonged epidemic2,3,4,5,6,7. Genome sequencing provides a high-resolution view of pathogen evolution and is increasingly sought after for outbreak surveillance. Sequence data may be used to guide control measures, but only if the results are generated quickly enough to inform interventions8. Genomic surveillance during the epidemic has been sporadic owing to a lack of local sequencing capacity coupled with practical difficulties transporting samples to remote sequencing facilities9. To address this problem, here we devise a genomic surveillance system that utilizes a novel nanopore DNA sequencing instrument. In April 2015 this system was transported in standard airline luggage to Guinea and used for real-time genomic surveillance of the ongoing epidemic. We present sequence data and analysis of 142 EBOV samples collected during the period March to October 2015. We were able to generate results less than 24 h after receiving an Ebola-positive sample, with the sequencing process taking as little as 15–60 min. We show that real-time genomic surveillance is possible in resource-limited settings and can be established rapidly to monitor outbreaks.
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European Nucleotide Archive
The EMLab is a technical partner in the WHO Emerging and Dangerous Pathogens Laboratory Network (EDPLN), and the Global Outbreak Alert and Response Network (GOARN) and the deployments in West Africa have been coordinated and supported by the GOARN Operational Support Team at WHO/HQ and the African Union. This work was carried out in the context of the project EVIDENT (Ebola virus disease: correlates of protection, determinants of outcome, and clinical management) that received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement No 666100 and in the context of service contract IFS/2011/272-372 funded by Directorate-General for International Cooperation and Development. J.Q. is funded by the NIHR Surgical Reconstruction and Microbiology Research Centre (SRMRC). N.J.L. is funded by a Medical Research Council Special Training Fellowship in Biomedical Informatics (to September 2015) and a Medical Research Council Bioinformatics Fellowship. J.T.S. is supported by the Ontario Institute for Cancer Research through funding provided by the Government of Ontario. Dstl support was funded by the UK Ministry of Defence (MOD). Dstl authors thank S. Lonsdale, C. Lonsdale and C. Mayers for supply of RNA, previous assistance, and review of the manuscript. The views expressed in this paper are not necessarily endorsed by the UK MOD. A.R. was supported by EU Seventh Framework Programme [FP7/2007-2013] under Grant Agreement no. 278433-PREDEMICS and ERC Grant agreement no. 260864. We are grateful for the generous support of University of Birmingham alumni for donations in support of the pilot work. The MRC Cloud Infrastructure for Microbial Bioinformatics (CLIMB) cyberinfrastructure was used to conduct bioinformatics analysis. The authors would like to thank B. Oppenheim and C. Wardius for help with logistics and the staff of Alta Biosciences, University of Birmingham and Sigma-Aldrich for generating PCR primers especially rapidly for this project. The authors would like to thank scientists deployed from the Special Pathogens Program from the National Microbiology Laboratory, Public Health Agency of Canada, who worked on EBOV diagnostics in Guinea. We are grateful to I. Goodfellow, M. Cotten and P. Kellam for permission to include sequences from Sierra Leone in this analysis. We thank R. Vipond for assistance with validation experiments. We thank H. Eno and B. Myers for help with proofreading. We are thankful for the generous support of reagents and technical support from Oxford Nanopore. We thank the staff at Oxford Nanopore for technical and logistical support during this project with special thanks to S. Brooking, O. Hartwell, R. Pettett, C. Brown, G. Sanghera and R. Ronan. We thank T. Bedford and R. Neher for developing the Nextstrain website.
Extended data figures
This file contains a Field Guide to Nanopore Sequencing - a detailed discussion of logistical issues that arose during this project and Supplementary Tables 1-4.