title: Whole Genome Sequence Analysis of Salmonella Enteritidis PT4 Outbreaks from a National Reference Laboratory's Viewpoint.
abstract: BACKGROUND
  In April and May 2014, two suspected egg-related outbreaks of Salmonella enterica subsp. enterica serovar Enteritidis (S. Enteritidis) were investigated by the Belgian National Reference Laboratory of Foodborne Outbreaks. Both the suspected food and human isolates being available, and this for both outbreaks, made these the ideal case study for a retrospective whole genome sequencing (WGS) analysis with the goal to investigate the feasibility of this technology for outbreak investigation by a National Reference Laboratory or National Reference Centre without thorough bioinformatics expertise.
  
  
  METHODS
  The two outbreaks were originally investigated epidemiologically with a standard questionnaire and with serotyping, phage typing, multiple-locus variable-number of tandem repeats analysis (MLVA) and antimicrobial susceptibility testing as classical microbiological methods. Retrospectively, WGS of six outbreak isolates was done on an Illumina HiSeq. Analysis of the WGS data was performed with currently available, user-friendly software and tools, namely CLC Genomics Workbench, the tools available on the server of the Center for Genomic Epidemiology and BLAST Ring Image Generator (BRIG).
  
  
  RESULTS
  To all collected human and food outbreak isolates, classical microbiological investigation assigned phage type PT4 (variant phage type PT4a for one human isolate) and MLVA profile 3-10-5-4-1, both of which are common for human isolates in Belgium. The WGS analysis confirmed the link between food and human isolates for each of the outbreaks and clearly discriminated between the two outbreaks occurring in a same time period, thereby suggesting a non-common source of contamination. Also, an additional plasmid carrying an antibiotic resistance gene was discovered in the human isolate with the variant phage type PT4a.
  
  
  CONCLUSIONS
  For the two investigated outbreaks occurring at geographically separated locations, the gold standard classical microbiological subtyping methods were not sufficiently discriminative to distinguish between or assign a common origin of contamination for the two outbreaks, while WGS analysis could do so. This case study demonstrated the added value of WGS for outbreak investigations by confirming and/or discriminating food and human isolates between and within outbreaks. It also proved the feasibility of WGS as complementary or even future replacing (sub)typing method for the average routine laboratory.
authors.affiliation: Department of Microbial and Molecular Systems, Centre of Microbial and Plant Genetics, KU Leuven, Leuven, Belgium; Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, Belgium; Platform Biotechnology and Molecular Biology, Scientific Institute of Public Health (WIV-ISP), Brussels, Belgium.
authors.forename: Véronique
authors.initials: V
authors.lastname: Wuyts
authors.affiliation: Scientific Institute of Public Health (WIV-ISP), Scientific Service of Foodborne Pathogens, NRL Foodborne Outbreaks, Brussels, Belgium.
authors.forename: Sarah
authors.initials: S
authors.lastname: Denayer
authors.affiliation: Scientific Institute of Public Health (WIV-ISP), Platform Biotechnology and Molecular Biology, Brussels, Belgium.
authors.forename: Nancy H C
authors.initials: NH
authors.lastname: Roosens
authors.affiliation: National Reference Centre for Salmonella and Shigella, Bacterial Diseases Division, Communicable and infectious Diseases, Scientific Institute of Public Health, Brussels, Belgium.
authors.forename: Wesley
authors.initials: W
authors.lastname: Mattheus
authors.affiliation: National Reference Centre for Salmonella and Shigella, Bacterial Diseases Division, Communicable and Infectious Diseases, Scientific Institute of Public Health (WIV-ISP), Brussels, Belgium.
authors.forename: Sophie
authors.initials: S
authors.lastname: Bertrand
authors.affiliation: Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, Belgium; Department of Information Technology, Ghent University, IMinds, Ghent, Belgium.
authors.forename: Kathleen
authors.initials: K
authors.lastname: Marchal
authors.affiliation: Scientific Institute of Public Health (WIV-ISP), Scientific Service of Foodborne Pathogens, NRL Foodborne Outbreaks, Brussels, Belgium.
authors.forename: Katelijne
authors.initials: K
authors.lastname: Dierick
authors.affiliation: Scientific Institute of Public Health (WIV-ISP).
authors.forename: Sigrid C J
authors.initials: SC
authors.lastname: De Keersmaecker
keywords: Salmonella; Whole genome sequence; disease outbreak; infectious disease