Eleventh external quality assessment scheme for Listeria monocytogenes typing in EU/EEA and EU enlargement countries, 2024

Human listeriosis is a relatively rare but serious food-borne disease. In 2023, the European Union (EU) notification rate was 0.66 cases per 100 000 population [1]. The number of cases in 2023 was even higher than before the COVID-19 pandemic (2019, 2 621 cases) [1], with an increase of 5.8% from 2022 to 2023 (2 738 and 2 952 cases, respectively). 

Since 2007, ECDC has been responsible for EU-wide surveillance of listeriosis, including facilitating the detection and investigation of food-borne outbreaks. Surveillance data, including certain basic typing parameters, are reported by European Union/European Economic Area (EU/EEA) countries to The European Surveillance System (TESSy). Since 2012, the EQA scheme has covered molecular typing methods used for EU-wide surveillance.

EQA-11, conducted between May and November 2024, involved serotyping/serogrouping and molecular typing-based cluster analyses. The objective of this EQA was to assess the quality and comparability of typing data reported by NPHRLs participating in FWD-Net. Seven test strains were selected for serotyping/grouping and molecular typing-based cluster analysis for the EQA to cover strains that are currently pertinent for public health in Europe, and to represent a broad range of clinically relevant types of invasive listeriosis. An additional ten sequences, some with modifications to simulate poor data quality or contamination, were included for the molecular typing-based cluster analysis. Twenty-three of the 24 laboratories that signed up completed the exercise. This was an increase of two laboratories compared with EQA-10; however, the composition was different, as one laboratory from EQA-10 did not participate in EQA-11. Most participating laboratories (18/23; 78%) completed the full EQA scheme. 

In total, 21 laboratories (91%) participated in the serotyping part of the exercise; all of them conducted PCR-based/WGS molecular serogrouping, and four laboratories (17%, 4/23) also conducted conventional antigen-based serotyping. Molecular serogrouping was performed well, with all (100%, 23/23) participants correctly identifying the serogroups. For the conventional method, 100% of the participants correctly serotyped all seven test strains. Since the first EQA in 2012, the trend has been towards replacing conventional serotyping with molecular serogrouping, showing strong performance. 

Of the 23 laboratories participating in the EQA-11, 20 (87%) performed molecular typing-based cluster analysis using a method of their choice. The intent of the cluster analysis component of the EQA was to evaluate the NPHRLs’ capability to identify a genetically closely related cluster and to accurately categorise the cluster test strains – regardless of the method used – rather than strictly being able to follow a specific procedure. 

The cluster of six closely related strains (three test strains and three strain sequences) was predefined by the EQA provider using data derived from whole genome sequencing (WGS). For this cluster, a correct cluster delineation was not expected to be possible to obtain using less discriminatory methods (e.g. pulsed-field gel electrophoresis (PFGE)). However, this year, none of the participating laboratories used PFGE for cluster identification. Twenty laboratories performed cluster analysis using WGS-derived data; only two used single-nucleotide polymorphism (SNP) as the main analysis. The submitted allelic differences (AD) clearly showed coherence despite the different approaches and schemes that the participants used. The most widely used core genome multilocus sequence type (cgMLST) scheme was Ruppitsch (13/20 laboratories), while the Pasteur scheme was less common (4/20). All laboratories reported 0–7 AD for the strains in the predefined cluster. When analysing the predefined cluster of the seven closely related strains, 75% (15/20) of the participants reported the same list of strains as the EQA provider. All of the participating laboratories were able to identify the pre-defined cluster of strain12, strain13 and strain16. However, a few participating laboratories wrongly included an additional one to three other strains.

In general, most of the participants were able to identify the different characteristics and modifications of the EQA-provided sequences. As part of their quality control (QC), 12 of the 20 participating laboratories (60%) excluded a modified strain based on correct identification of contamination. The issues with the sequence that had a mix of sequence types were detected and excluded by 90% (18/20) of the laboratories and finally, the two strains modified for reduced coverage, were identified and excluded by 95% (19/20) and 90% (18/20) of participants, respectively. 

A feedback survey was sent to the participants to assess their experience of EQA-11; 43% (10/23) of the participants responded. Overall, responses were positive, with minimal critical feedback. Few comments were provided, and no major concerns were raised, indicating that the process met expectations effectively. Decreased feedback compared to EQA-10 suggests a well-received experience with no immediate areas requiring improvement.