Phenotypic diversity of a clonal Ralstonia solanacearum pathogen lineage is explained by accessory genome variation

Ralstonia solanacearumis aplant pathogenic gram-negative bacterium capable of infecting several economically important crops such as potato and tomato. It can also persist in environmental reservoirs including soils, rivers and in asymptomatic wild hosts, causing disease outbreaks during pathogen spillover events when crossing agroecological interface. In the UK, R. solanacearum outbreaks originate from Solanum dulcamarawild hosts (woody nightshade) and river networks. To what extent selection in these natural environments drive R. solanacearumsurvival and life history evolution including virulence is unknown. To study this, we focused on a largely clonal R. solanacearum lineage inhabiting river networks across the UK consisting of a collection of 182 isolates spanning 30 years since the first outbreak in 1992. We first characterised strains phenotypically regarding 32 traits including resource catabolism, virulence and abiotic stress tolerance and then used microbial GWAS techniques to identify links between phenotypic traits and the presence of specific accessory genes. We found that isolates can be clustered into three phenotypic groups, which differed clearly regarding their resource specialism and stress tolerance. No effect of isolation location was found. However, isolates became more variable phenotypically along with time. While only few SNPs were found to vary among all isolates, the presence and absence of certain accessory genes, such asS-layer family protein,could be associated with phenotypic variation in terms ofsurvival in stressful environments. Together, our findings suggest that a low number of accessory genes can cause high phenotypic variability within highly clonal bacterial lineage.