Dataset Information

Implementation of Nationwide Real-time Whole-genome Sequencing to Enhance Listeriosis Outbreak Detection and Investigation.

ABSTRACT: Listeria monocytogenes (Lm) causes severe foodborne illness (listeriosis). Previous molecular subtyping methods, such as pulsed-field gel electrophoresis (PFGE), were critical in detecting outbreaks that led to food safety improvements and declining incidence, but PFGE provides limited genetic resolution. A multiagency collaboration began performing real-time, whole-genome sequencing (WGS) on all US Lm isolates from patients, food, and the environment in September 2013, posting sequencing data into a public repository. Compared with the year before the project began, WGS, combined with epidemiologic and product trace-back data, detected more listeriosis clusters and solved more outbreaks (2 outbreaks in pre-WGS year, 5 in WGS year 1, and 9 in year 2). Whole-genome multilocus sequence typing and single nucleotide polymorphism analyses provided equivalent phylogenetic relationships relevant to investigations; results were most useful when interpreted in context of epidemiological data. WGS has transformed listeriosis outbreak surveillance and is being implemented for other foodborne pathogens.

SUBMITTER: Jackson BR

PROVIDER: S-EPMC4946012 | biostudies-other | 2016 Aug

REPOSITORIES: biostudies-other

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Implementation of Nationwide Real-time Whole-genome Sequencing to Enhance Listeriosis Outbreak Detection and Investigation.

Jackson Brendan R BR Tarr Cheryl C Strain Errol E Jackson Kelly A KA Conrad Amanda A Carleton Heather H Katz Lee S LS Stroika Steven S Gould L Hannah LH Mody Rajal K RK Silk Benjamin J BJ Beal Jennifer J Chen Yi Y Timme Ruth R Doyle Matthew M Fields Angela A Wise Matthew M Tillman Glenn G Defibaugh-Chavez Stephanie S Kucerova Zuzana Z Sabol Ashley A Roache Katie K Trees Eija E Simmons Mustafa M Wasilenko Jamie J Kubota Kristy K Pouseele Hannes H Klimke William W Besser John J Brown Eric E Allard Marc M Gerner-Smidt Peter P

Clinical infectious diseases : an official publication of the Infectious Diseases Society of America 20160418 3

Listeria monocytogenes (Lm) causes severe foodborne illness (listeriosis). Previous molecular subtyping methods, such as pulsed-field gel electrophoresis (PFGE), were critical in detecting outbreaks that led to food safety improvements and declining incidence, but PFGE provides limited genetic resolution. A multiagency collaboration began performing real-time, whole-genome sequencing (WGS) on all US Lm isolates from patients, food, and the environment in September 2013, posting sequencing data i ...[more]

PMID: 27090985

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Project description:Whole genome sequencing (WGS) has become the new gold standard for bacterial outbreak investigation, due to the high resolution available for typing. While sequencing is currently predominantly performed on Illumina devices, the preceding library preparation can be performed using various protocols. Enzymatic fragmentation library preparation protocols are fast, have minimal hands-on time, and work with small quantities of DNA. The aim of our study was to compare three library preparation protocols for molecular typing: Nextera XT (Illumina); Nextera Flex (Illumina); and QIAseq FX (Qiagen). We selected 12 ATCC strains from human Gram-positive and Gram-negative pathogens with %G+C-content ranging from 27% (Fusobacterium nucleatum) to 73% (Micrococcus luteus), each having a high quality complete genome assembly available, to allow in-depth analysis of the resulting Illumina sequence data quality. Additionally, we selected isolates from previously analyzed cases of vancomycin-resistant Enterococcus faecium (VRE) (n = 7) and a local outbreak of Klebsiella aerogenes (n = 5). The number of protocol steps and time required were compared, in order to test the suitability for routine laboratory work. Data analyses were performed with standard tools commonly used in outbreak situations: Ridom SeqSphere+ for cgMLST; CLC genomics workbench for SNP analysis; and open source programs. Nextera Flex and QIAseq FX were found to be less sensitive than Nextera XT to variable %G+C-content, resulting in an almost uniform distribution of read-depth. Therefore, low coverage regions are reduced to a minimum resulting in a more complete representation of the genome. Thus, with these two protocols, more alleles were detected in the cgMLST analysis, producing a higher resolution of closely related isolates. Furthermore, they result in a more complete representation of accessory genes. In particular, the high data quality and relative simplicity of the workflow of Nextera Flex stood out in this comparison. This thorough comparison within an ISO/IEC 17025 accredited environment will be of interest to those aiming to optimize their clinical microbiological genome sequencing.

Dataset Information

Implementation of Nationwide Real-time Whole-genome Sequencing to Enhance Listeriosis Outbreak Detection and Investigation.

Publications

Implementation of Nationwide Real-time Whole-genome Sequencing to Enhance Listeriosis Outbreak Detection and Investigation.

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