Genetic diversity and delineation of Salmonella Agona outbreak strains by next generation sequencing, Bavaria, Germany, 1993 to 2018

Background In 2017, a food-borne Salmonella Agona outbreak caused by infant milk products from a French supplier occurred in Europe. Simultaneously, S. Agona was detected in animal feed samples in Bavaria. Aim Using next generation sequencing (NGS) and three data analysis methods, this study’s objectives were to verify clonality of the Bavarian feed strains, rule out their connection to the outbreak, explore the genetic diversity of Bavarian S. Agona isolates from 1993 to 2018 and compare the analysis approaches employed, for practicality and ability to delineate outbreaks caused by the genetically monomorphic Agona serovar. Methods In this observational retrospective study, three 2017 Bavarian feed isolates were compared to a French outbreak isolate and 48 S. Agona isolates from our strain collections. The later included human, food, feed, veterinary and environmental isolates, of which 28 were epidemiologically outbreak related. All isolates were subjected to NGS and analysed by: (i) a publicly available species-specific core genome multilocus sequence typing (cgMLST) scheme, (ii) single nucleotide polymorphism phylogeny and (iii) an in-house serovar-specific cgMLST scheme. Using additional international S. Agona outbreak NGS data, the cluster resolution capacity of the two cgMLST schemes was assessed. Results We could prove clonality of the feed isolates and exclude their relation to the French outbreak. All approaches confirmed former Bavarian epidemiological clusters. Conclusion Even for S. Agona, species-level cgMLST can produce reasonable resolution, being standardisable by public health laboratories. For single samples or homogeneous sample sets, higher resolution by serovar-specific cgMLST or SNP genotyping can facilitate outbreak investigations.