Written by Charlotte Harrison, science writer
Whole-genome sequencing can identify genetic variants that cause disease, but the time needed for sequencing and analysis hinders its use in patients who are acutely ill.
Now, a study in Nature Biotechnology shows that a method using nanopore sequencing can identify genetic variants and provide a genome-based diagnosis much quicker than previous efforts. In the study, high-depth, human whole-genome data was generated in under 2 hours and diagnostic genetic variants were identified in under 8 hours.
All systems go
The team of scientists from Stanford University, UC Santa Cruz Genomics Institute, Stanford Health Care, Oxford Nanopore Technologies, NVIDIA Corporation and Baylor College of Medicine based their approach on Oxford Nanopore’s high-throughput sequencing device, PromethION 48, which uses multiple flow cells to sequence one sample.
They combined the use of this sequencing device with an optimized protocol for sample preparation, near real-time base calling and alignment using a cloud-based module, accelerated variant calling (that is, identification of genetic variants) and fast filtration of a reduced number of variants to enable efficient manual curation.
The feasibility and characteristics of this rapid sequencing pipeline were highlighted using a reference genome, showing that the platform provides accurate variant calls and efficient prioritization, and accelerates diagnostic genome sequencing twofold compared with previous approaches.
The need for clinical speed
To test the clinical application of the platform, the researchers sequenced samples from two critically ill patients.
The first was a middle-aged man with a severe SARS-CoV-2 infection and comorbidities, who needed a lung transplant. His clinicians requested rapid molecular testing to help clarify a wide differential diagnosis. In under 7.5 hours from the start of sample preparation, a diagnostic variant was identified in the TNNT2 gene (encoding cardiac troponin T). The genetic diagnosis reduced the need for multiple follow-up imaging studies and a cardiac biopsy.
The second case was an infant with a suspected but unconfirmed genetic disorder who was admitted to intensive care. In around 7 h after a blood sample was taken, 31 small variants and 21 structural variants had been prioritized for manual review. Less than an hour later, the researchers identified the LZTR1 gene — which likely stabilizes the Golgi complex — as a candidate variant gene, although the pathogenic contribution of this variant was unclear.
In a companion study published in the New England Journal of Medicine, the pipeline was used to sequence samples from 12 critically ill patients with hard to diagnose symptoms. Pathogenic genetic variations were identified for five of the 12 patients, and for all patients, the sequencing pipeline was completed within one day.
Issues regarding the high cost of this type of sequencing need to be addressed, and faster access to genetic information is only relevant in an acute care setting if a treatment or clinical decision can be made based on that information. Nevertheless, these studies show that rapid nanopore sequencing is moving towards clinical use.
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