CHICAGO – Canadian startup BugSeq Bioinformatics has devised a method of detecting pathogens via metagenomic nanopore sequencing that the company says is more accurate and sensitive than PCR and microbial cultures. Notably, the technology does not need to target specific pathogens like PCR and microbial culture tests must.
The firm teamed with researchers from the Future of Humanity Institute, the University of British Columbia, Vancouver General Hospital, and the British Columbia Centre for Disease Control to describe the method in a manuscript posted on MedRxiv last week.
BugSeq cofounder and CEO Sam Chorlton, lead author of the study, said that metagenomic sequencing is “unbiased” when it comes to detecting pathogens because it is not just looking for a specific virus or a bacterial strain.
In a pilot study, the researchers examined samples collected from 2,714 people in and around Oxford, UK, from April to June 2020. The British Columbia team ran PCR tests, serology assays, and nanopore sequencing on this population to compare each method of determining SARS-CoV-2 positivity.
They reported 100 percent concordance in five samples for which they had both assembled SARS-CoV-2 genomes and PCR results for “variants of concern,” as designated by the World Health Organization. “Additionally, our assay was able to distinguish between the Alpha and Gamma variants, which was not possible with our VOC PCR technique,” they wrote.
The preprint also said that BugSeq demonstrated 95.2 percent sensitivity for samples with a RT-qPCR cycle threshold value of less than 30 in detecting SARS-CoV-2. Although performance drops when the Ct value is 30 or more, Chorlton said that BugSeq is “optimistic” about methods for increasing accuracy above Ct 30, including through host nucleic acid depletion and pathogen enrichment.
Vancouver, British Columbia-based BugSeq’s technology features a classification system that corrects for barcode cross-talk between specimens, according to the preprint.
The pilot study measured the performance of sequence-independent single primer amplification for detecting and characterizing SARS-CoV-2 through metagenomic next-generation sequencing. “[These] approaches provide the opportunity to examine the entire genomic material of a sample; allowing for detection of emerging and clinically relevant pathogens that may be missed in targeted assays,” the authors explained.
“This is a proof of principle that we can detect any pathogen, whether it’s known or novel, whether it’s emerging, prevalent, whether we have genome sequences online or not,” Chorlton said. “That’s really what we think is the future of diagnostic microbiology, this universal pathogen-agnostic diagnosis.”
Making the study possible, according to Chorlton, was the Oxford Nanopore Technologies handheld MinIon sequencer, which costs less than $1,000, putting long-read sequencing technology within reach of even the smallest lab. BugSeq has also been optimized for Pacific Biosciences Sequel II sequencers, he said.
“The portability and cost-effectiveness of MinIon sequencing makes Nanopore mNGS uniquely tailored for clinical applications,” the study authors wrote. “Despite these advances in long-read clinical sequencing applications, the field of nanopore clinical metagenomics has been largely unexplored.”
They described BugSeq as a bioinformatics platform featuring a graphical user interface and cloud-based data processing that enables clinical microbiology labs to conduct end-to-end analysis of nanopore sequencing data.
While the Oxford Nanopore MinIon produces high-throughput sequencing data in real time, it is slower than real-time qualitative reverse transcription PCR for producing a diagnostic result. “The use of liquid handling robots for automated sample extraction, nucleic acid amplification, and library preparation may aid in standardization,” the preprint said.
Founded in April 2020, BugSeq grew out of the needs for a bioinformatics platform to manage metagenomic testing from long-read sequencers. Although Chorlton is a resident medical microbiologist at UBC, BugSeq was independently developed.
In his capacity as a UBC faculty member, Chorlton consulted for microbiology diagnostic labs across Canada, including the National Microbiology Laboratory in Winnipeg, Manitoba.
“It was easier and easier [to perform] third-generation sequencing,” Chorlton said, “but basically none of them knew how to do any of the analysis.”
BugSeq has positioned itself as a platform for clinical defense, public health, and applied microbiology labs to analyze sequencing data, particularly sequences from long-read instruments from PacBio and Oxford Nanopore.
The firm has a two-tiered business model. The core software is open-source and freely available to individual users, but BugSeq also offers a paid package to help labs trying to adopt long-read sequencing.
“The value there is everything from taking their computational infrastructure off-site into the cloud that can be accelerated [to] doing all of the analysis automatically without needing a bioinformatician,” Chorlton said. “[We’re] giving them leading-accuracy data analysis that’s tailored for the clinical defense and public-health space.”
Already, BugSeq claims customers on six continents and even beyond, because the company processes data from the International Space Station. Users include the US Centers for Disease Control and Prevention, Canadian Health Networks, the Natural History Museum of London, and the British Columbia Centre for Disease Control, according to the firm.
The company has been self-funded so far, starting with bootstrapped investments from the founders and now earning revenue from paid users.
Since it has not been given regulatory clearance as a diagnostic aid in any jurisdiction yet, BugSeq is for research purposes only, but Chorlton said that this is on the company’s roadmap. He did not give a timeline for seeking regulatory review.
It has been used to identify SARS-CoV-2 in clinical specimens, though. “We believe that that will be an important part of the future where it’s one test that can detect all pathogens,” Chorlton said.
In the preprint study, the researchers were able to detect not only SARS-CoV-2, but other respiratory pathogens as well. They showed that none of the patients had influenza or respiratory syncytial virus, but did find Haemophilus influenzae and Streptococcus pneumoniae.
“Users are using us for every organism in every domain. We have people using BugSeq for bacterial infections, viral infections, fungal, and parasitic,” Chorlton said. This was described in a peer-reviewed paper about the technology that appeared in BMC Informatics in March.
BugSeq currently can only detect previously identified pathogens, but Chorlton said that the next iteration will be able to detect novel infections as well. He said that this update would be out “shortly,” but did not specify a time frame.
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