Handheld Nanopore DNA Sequencer Rapidly Detects Viral Infections in Blood Samples

A handheld nanopore DNA sequencer was used to accurately identify and differentiate viruses in patient blood samples with an unprecedented sample-to-answer turnaround time of less than six hours.

The Oxford Nanopore Technologies (Oxford, United Kingdom) MinION nanopore sequencer rapidly determines the sequence of subject DNA through the application of protein nanopore technology. The method is based on a protein channel—only a few nanometers in diameter—through which a single strand of DNA can pass. As the DNA strand passes through the nanopore, it generates a series of characteristic electrical signatures, from which nucleotide bases can be identified, and the sequence of the strand determined. The instrument is powered and operated by a laptop computer via a USB connection.

Investigators at the University of California, San Francisco (USA; www.ucsf.edu) used a MinION instrument to analyze blood samples from four patients for Chikungunya virus (CHIKV), Ebolavirus (EBOV), and Hepatitis C virus (HCV).

They reported that at high titers ranging from 107 to 108 copies per milliliter, reads to EBOV from two patients with acute hemorrhagic fever and CHIKV from an asymptomatic blood donor were detected within 4 to 10 minutes of data acquisition, while lower titer HCV virus (1x105 copies per milliliter) was detected within 40 minutes. Confirmation of results was obtained by sequencing with an Illumina Inc. (San Diego, CA, USA) MiSeq instrument.

Nanopore sequencing is a third-generation sequencing technology that has two key advantages over second-generation technologies—longer reads and the ability to perform real-time sequence analysis. As of mid-2015, the MinION nanopore sequencer was capable of producing at least 100,000 sequences with an average read length of five kilobases, in total producing up to one gigabase of sequence in 24 hours on one flow cell.

In the current study, the investigators presented nanopore sequencing for metagenomic detection of viral pathogens from clinical samples with a sample-to-answer turnaround time of less than six hours. They also introduced MetaPORE, a real-time web-based sequence analysis and visualization tool for pathogen identification from nanopore data.

“To our knowledge, this is the first time that nanopore sequencing has been used for real-time metagenomic detection of pathogens in complex clinical samples in the setting of human infections,” said senior author Dr. Charles Y Chiu, associate professor of laboratory medicine at the University of California, San Francisco. “Unbiased point-of-care testing for pathogens by rapid metagenomic sequencing has the potential to radically transform infectious disease diagnosis in both clinical and public health settings. This point-of-care genomic technology will be particularly attractive in the developing world, where critical resources, including reliable electric power, laboratory space, and computational server capacity, are often severely limited.

The study was published in the September 29, 2015, online edition of the journal Genome Medicine.

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Oxford Nanopore Technologies
University of California, San Francisco
Illumina Inc.