Benchmarking and quality control for nanopore sequencing and feasibility of rapid genomics in New Zealand: validation phase at a single quaternary hospital

Approximately 200 critically ill infants and children in New Zealand are in high-dependency neonatal/paediatric acute care at any given time, many with suspected genetic conditions, necessitating a scalable distributed solution for rapid genomic testing. We adopt the existing acute care genomics protocol of an accredited laboratory and established an expandable acute care clinical pipeline based around the Oxford Nanopore Technologies PromethION 2 solo system connected to a Bayesian AI-based clinical decision support tool (Fabric GEM software). In the establishment phase, we performed benchmarking using Global Alliance for Genomics and Health (GA4GH) benchmarking tools and Genome in a Bottle samples HG002-HG007. We evaluated single nucleotide variants (SNVs) and small insertions-deletions (indels) calls and achieved SNV precision and recall of 0.997 and 0.992, respectively. Small indel identification approached a precision of 0.922 and recall of 0.838. Rarefaction analyses demonstrated that SNV identification plateaus at ~20X coverage, while small indels plateaus at ~40X coverage. Large genomic variations from Coriell Copy Number Variation Reference Panel 1 (CNVPANEL01) were reliably detected with ~2M long reads. Finally, we present results obtained from ten trio samples that were processed through the pipeline validation phase, averaging a 5-day turnaround time, conducted in parallel with a clinically accredited short-read rapid genomic testing pipeline. ### Competing Interest Statement The authors have declared no competing interest. ### Funding Statement We gratefully acknowledge the generous donations from the Dines Family Trust, The Toutoku Trust, and The Kelliher Trust. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: Ethics approval was obtained from the Northern B Health and Disability Ethics Committee for the study entitled: Newborn Genomics Te Ira oo Te Arai (Ethics reference: 2023 FULL 15542). Locality approval was obtained from the Research Review Committee Te Toka Tumai Auckland for the project entitled Newborn Genomics Te Ira oo Te Arai (Reference A+9855 [FULL 15542]) I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes The genomic data (i.e. bam files) for the Genome in a Bottle (GIAB) samples is publicly available from NCBI Sequence Read Archive (SRA). The genomic and phenotypic data from families analysed in this study cannot be shared publicly due to privacy and ethical restrictions.