1887

Abstract

There is a growing need for public health and veterinary laboratories to perform whole genome sequencing (WGS) for monitoring antimicrobial resistance (AMR) and protecting the safety of people and animals. With the availability of smaller and more affordable sequencing platforms coupled with well-defined bioinformatic protocols, the technological capability to incorporate this technique for real-time surveillance and genomic epidemiology has greatly expanded. There is a need, however, to ensure that data are of high quality. The goal of this study was to assess the utility of a small benchtop sequencing platform using a multi-laboratory verification approach. Thirteen laboratories were provided the same equipment, reagents, protocols and bacterial reference strains. The Illumina DNA Prep and Nextera XT library preparation kits were compared, and 2×150 bp iSeq i100 chemistry was used for sequencing. Analyses comparing the sequences produced from this study with closed genomes from the provided strains were performed using open-source programs. A detailed, step-by-step protocol is publicly available via protocols.io (https://www.protocols.io/view/iseq-bacterial-wgs-protocol-bij8kcrw). The throughput for this method is approximately 4–6 bacterial isolates per sequencing run (20–26 Mb total load). The Illumina DNA Prep library preparation kit produced high-quality assemblies and nearly complete AMR gene annotations. The Prep method produced more consistent coverage compared to XT, and when coverage benchmarks were met, nearly all AMR, virulence and subtyping gene targets were correctly identified. Because it reduces the technical and financial barriers to generating WGS data, the iSeq platform is a viable option for small laboratories interested in genomic surveillance of microbial pathogens.

Funding
This study was supported by the:
  • u.s. food and drug administration (Award 1U18 FD006444-01)
    • Principle Award Recipient: NotApplicable
  • u.s. food and drug administration (Award 5U18FD006160-02)
    • Principle Award Recipient: NotApplicable
  • u.s. food and drug administration (Award 1U18FD006866-01)
    • Principle Award Recipient: LeyiWang
  • u.s. food and drug administration (Award 1U18FD006714-01)
    • Principle Award Recipient: LauraB. Goodman
  • u.s. food and drug administration (Award 1U18 FD006859-01)
    • Principle Award Recipient: NotApplicable
  • u.s. food and drug administration (Award 1U18 FD006445-01)
    • Principle Award Recipient: NotApplicable
  • u.s. food and drug administration (Award 1U18 FD006719-01)
    • Principle Award Recipient: NotApplicable
  • u.s. food and drug administration (Award 1U18FD006671-01)
    • Principle Award Recipient: NotApplicable
  • u.s. food and drug administration (Award 1U18 FD006555-01)
    • Principle Award Recipient: NotApplicable
  • u.s. food and drug administration (Award 1U18 FD006714-01)
    • Principle Award Recipient: NotApplicable
  • u.s. food and drug administration (Award 1U18 FD006866-01)
    • Principle Award Recipient: NotApplicable
  • u.s. food and drug administration (Award 1U18 FD006460-01)
    • Principle Award Recipient: NotApplicable
  • u.s. food and drug administration (Award 1U18 FD006568-01)
    • Principle Award Recipient: NotApplicable
  • u.s. food and drug administration (Award 1U18 FD006593-01)
    • Principle Award Recipient: NotApplicable
  • This is an open-access article distributed under the terms of the Creative Commons Attribution NonCommercial License. This article was made open access via a Publish and Read agreement between the Microbiology Society and the corresponding author’s institution.
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2022-02-03
2024-03-28
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