Portable Nanopore Sequencing for Viral Surveillance.

Abstract

The recent Nature article (1) entitled, “Real-Time, Portable Genome Sequencing for Ebola Surveillance,” reports nanopore sequencing of the Ebola virus from 142 subjects near the end of the outbreak in West Africa from March to October 2015. The nanopore sequencer (MinION, Oxford Nanopore Technologies) has a mass of 87 g and plugs into a laptop computer by way of a USB 3.0 interface and cable. The consumable flow cell has up to 512 channels available for sequencing, each of which can simultaneously read single strands of DNA and their ligated complementary strands as “2D” reads, providing correlated data on both strands of DNA. The nanopores are typically made up of proteins like α-hemolysin that are pore-forming bacterial toxins embedded in an electrically insulating membrane. A voltage between the 2 sides of the membrane attracts DNA to the pore in an electrolyte solution. The DNA is captured by a polymerase near the pore that paces the rate of transport to 30–280 bases/s. The passage of single-stranded DNA affects the pore current, generating electrical “squiggle signals” that are sequence-specific. In simple terms, this is the Coulter principle applied to molecules rather than cells. In half of the cases, sufficient reads (25 2D reads for each amplicon) were obtained in <1 h of instrument time with a mean passing rate of 36.3%. The mean read rate was 138 amplicons/min, and in some cases, only 15 min of sequencing was required. Custom offline acquisition software was provided by the company so that Internet communication was not initially required. Nanopore sequencing is conceptually elegant and data acquisition is fast. However… Before sequencing, sample transport and preparation are required. To prevent delays of air transport of samples, a portable 50-kg laboratory was transported in airline luggage to Guinea. As samples became available locally, they were transported …