Whole genome sequencing as a tool to investigate a cluster of seven cases of listeriosis in Austria and Germany, 2011-2013

Abstract

A cluster of seven human cases of listeriosis occurred in Austria and in Germany between April 2011 and July 2013. The Listeria monocytogenes serovar (SV) 1/2b isolates shared pulsed-field gel electrophoresis (PFGE) and fluorescent amplified fragment length polymorphism (fAFLP) patterns indistinguishable from those from five food producers. The seven human isolates, a control strain with a different PFGE/fAFLP profile and ten food isolates were subjected to whole genome sequencing (WGS) in a blinded fashion. A gene-by-gene comparison (multilocus sequence typing (MLST)+) was performed, and the resulting whole genome allelic profiles were compared using SeqSphere(+) software version 1.0. On analysis of 2298 genes, the four human outbreak isolates from 2012 to 2013 had different alleles at ≤6 genes, i.e. differed by ≤6 genes from each other; the dendrogram placed these isolates in between five Austrian unaged soft cheese isolates from producer A (≤19-gene difference from the human cluster) and two Austrian ready-to-eat meat isolates from producer B (≤8-gene difference from the human cluster). Both food products appeared on grocery bills prospectively collected by these outbreak cases after hospital discharge. Epidemiological results on food consumption and MLST+ clearly separated the three cases in 2011 from the four 2012-2013 outbreak cases (≥48 different genes). We showed that WGS is capable of discriminating L. monocytogenes SV1/2b clones not distinguishable by PFGE and fAFLP. The listeriosis outbreak described clearly underlines the potential of sequence-based typing methods to offer enhanced resolution and comparability of typing systems for public health applications.

Keywords: Foodborne infections; Listeria; listeriosis; outbreaks; typing; whole genome sequencing; zoonoses.

Fig 1

Cases of invasive listeriosis caused by Listeria monocytogenes (L.m.)serovar 1/2b by calendar week of diagnosis (Austria, 2010 to February 2014).

Fig 2

Phylogenetic relationships of 18 Listeria monocytogenes serovar 1/2b isolates from different sources based on whole genome sequencing. The minimum spanning tree is based on allelic profiles of 2998 genes present in all strains investigated. Each circle represents a given allelic profile, and contains the isolate's name as given in Materials and Methods. The different sample origin (human, cheese, and meat) is distinguished by colours of the circles. The numbers on the connecting lines illustrate the number of differing alleles in a pairwise comparison.

Fig 3

Occurrence of Listeria monocytogenes serovar 1/2b in food samples tested in Austria 2013: cluster strain in comparison with non-cluster strains. Outbreak investigation started on 22 January 2013. The producer of the unaged soft cheese, producer A, was first contacted on 26 February, and the producer of the ready-to-eat meat, producer B, on 4 March. Raised awareness and resulting intensification of in-factory sanitation caused the cluster strain to disappear.