Multi-laboratory evaluation of the Illumina iSeq platform for whole genome sequencing of Salmonella, Escherichia coli and Listeria

Patrick K. Mitchell, Leyi Wang, Bryce J. Stanhope, Brittany D. Cronk, Renee Anderson, Shipra Mohan, Lijuan Zhou, Susan Sanchez, Paula Bartlett, Carol Maddox, Vanessa Deshambo, Rinosh Mani, Lindsy M. Hengesbach, Sarah C Gresch, Katie Wright, Sunil Mor, Shuping Zhang, Zhenyu Shen, Lifang Yan, Rebecca MackeyRebecca Franklin-Guild, Yan Zhang, Melanie Prarat, Katherine Shiplett, Akhilesh Ramachandran, Sai Narayanan, Justin Sanders, Andree A. Hunkapiller, Kevin Lahmers, Amanda A. Carbonello, Nicole Aulik, Ailam Lim, Jennifer Cooper, Angelica Jones, Jake Guag, Sarah M. Nemser, Gregory H. Tyson, Ruth Timme, Errol Strain, Renate Reimschuessel, Olgica Ceric, Laura B. Goodman

Research output: Contribution to journalArticlepeer-review

Abstract

There is a growing need for public health and veterinary laboratories to perform whole genome sequencing (WGS) for monitor-ing 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 cor-rectly 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.

Original languageEnglish (US)
Article number000717
JournalMicrobial genomics
Volume8
Issue number2
DOIs
StatePublished - 2022

Bibliographical note

Funding Information:
This study was funded (PAR-18-604) and performed in collaboration with the Food and Drug Administration?s Veterinary Laboratory Investigation and Response Network (FDA Vet-LIRN) under cooperative agreements 1U18FD006714-01 to L.B.G. and 1U18FD006866-01 to L.W. Additional funding for participating laboratories was as follows: 5U18FD006160-02, Florida Department of Agriculture and Consumer Services; 1U18 FD006444-01, Athens Veterinary Diagnostic Laboratory, The University of Georgia; 1U18 FD006593-01, Michigan State University, Veterinary Diagnostic Labora-tory; 1U18 FD006568-01, University of Minnesota, Veterinary Diagnostic Laboratory; 1U18 FD006460-01, University of Missouri, Veterinary Medical Diagnostic Laboratory; 1U18 FD006866-01, 1U18 FD006714-01, Mississippi State University, Veterinary Diagnostic Laboratory; 1U18 FD006555-01, Ohio Animal Disease Diagnostic Laboratory; 1U18FD006671-01, Oklahoma Animal Disease Diagnostic Laboratory, Oklahoma State University; 1U18 FD006719-01, Oregon Veterinary Diagnostic Laboratory, Oregon State University; 1U18 FD006445-01, Virginia-Maryland College of Veterinary Medicine; 1U18 FD006859-01, University of Wisconsin-Madison Veterinary Diagnostic Laboratory.

Funding Information:
This study was funded (PAR-18-604) and performed in collaboration with the Food and Drug Administration’s Veterinary Laboratory Investigation and Response Network (FDA Vet-LIRN) under cooperative agreements 1U18FD006714-01 to L.B.G. and 1U18FD006866-01 to L.W. Additional funding for participating laboratories was as follows: 5U18FD006160-02, Florida Department of Agriculture and Consumer Services; 1U18 FD006444-01, Athens Veterinary Diagnostic Laboratory, The University of Georgia; 1U18 FD006593-01, Michigan State University, Veterinary Diagnostic Laboratory; 1U18 FD006568-01, University of Minnesota, Veterinary Diagnostic Laboratory; 1U18 FD006460-01, University of Missouri, Veterinary Medical Diagnostic Laboratory; 1U18 FD006866-01, 1U18 FD006714-01, Mississippi State University, Veterinary Diagnostic Laboratory; 1U18 FD006555-01, Ohio Animal Disease Diagnostic Laboratory; 1U18FD006671-01, Oklahoma Animal Disease Diagnostic Laboratory, Oklahoma State University; 1U18 FD006719-01, Oregon Veterinary Diagnostic Laboratory, Oregon State University; 1U18 FD006445-01, Virginia-Maryland College of Veterinary Medicine; 1U18 FD006859-01, University of Wisconsin-Madison Veterinary Diagnostic Laboratory.

Publisher Copyright:
© 2022, Microbiology Society. All rights reserved.

Keywords

  • One Health
  • foodborne illness
  • sequencing accessibility
  • sequencing quality

PubMed: MeSH publication types

  • Journal Article
  • Research Support, Non-U.S. Gov't

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