Distinct but Intertwined Evolutionary Histories of Multiple Salmonella enterica Subspecies

Abstract

Salmonella is responsible for many nontyphoidal foodborne infections and enteric (typhoid) fever in humans. Of the two Salmonella species, Salmonella enterica is highly diverse and includes 10 known subspecies and approximately 2,600 serotypes. Understanding the evolutionary processes that generate the tremendous diversity in Salmonella is important in reducing and controlling the incidence of disease outbreaks and the emergence of virulent strains. In this study, we aim to elucidate the impact of homologous recombination in the diversification of S. enterica subspecies. Using a data set of previously published 926 Salmonella genomes representing the 10 S. enterica subspecies and Salmonella bongori, we calculated a genus-wide pan-genome composed of 84,041 genes and the S. enterica pan-genome of 81,371 genes. The size of the accessory genomes varies between 12,429 genes in S. enterica subsp. arizonae (subsp. IIIa) to 33,257 genes in S. enterica subsp. enterica (subsp. I). A total of 12,136 genes in the Salmonella pan-genome show evidence of recombination, representing 14.44% of the pan-genome. We identified genomic hot spots of recombination that include genes associated with flagellin and the synthesis of methionine and thiamine pyrophosphate, which are known to influence host adaptation and virulence. Last, we uncovered within-species heterogeneity in rates of recombination and preferential genetic exchange between certain donor and recipient strains. Frequent but biased recombination within a bacterial species may suggest that lineages vary in their response to environmental selection pressure. Certain lineages, such as the more uncommon non-enterica subspecies (non-S. enterica subsp. enterica), may also act as a major reservoir of genetic diversity for the wider population.IMPORTANCE S. enterica is a major foodborne pathogen, which can be transmitted via several distinct routes from animals and environmental sources to human hosts. Multiple subspecies and serotypes of S. enterica exhibit considerable differences in virulence, host specificity, and colonization. This study provides detailed insights into the dynamics of recombination and its contributions to S. enterica subspecies evolution. Widespread recombination within the species means that new adaptations arising in one lineage can be rapidly transferred to another lineage. We therefore predict that recombination has been an important factor in the emergence of several major disease-causing strains from diverse genomic backgrounds and their ability to adapt to disparate environments.

Keywords: Salmonella; genome; pan-genome; recombination; subspecies.

FIG 1

Genomic differences among Salmonella enterica subspecies genomes. (a) Pairwise genome-wide ANI values. ANI calculates the average nucleotide identity of all orthologous genes shared between any two genomes. The phylogeny was reconstructed using the concatenated alignment of 1,596 genus-wide core genes. The scale bar represents nucleotide substitutions per site. (b) Frequency distribution of all pairwise ANI values. The 95% ANI cutoff is a frequently used standard for species demarcation. (c) Number of SNPs in the core genome alignment per subspecies. The box shows the median SNP count and the lower and upper quartiles. The whiskers represent the minimum and maximum SNP counts. (d) Number of accessory genes per genome for each subspecies. Subspecies classification is based on core genome variation calculated by Alikhan et al. (12).

FIG 2

Recombination parameters of the five largest S. enterica subspecies calculated using mcorr (34). Histograms show the frequency distribution of each recombination parameter for all pairs of genomes.

FIG 3

Variable patterns of recombination. (a) Size distribution of lengths of recombined core and accessory DNA fragments. (b) Genes that have undergone recent or ancestral recombination. The horizontal axis shows the estimated number of ancestral recombinations, and the vertical axis shows the estimated number of recent recombinations. For clarity, names of some of the most frequently recombined genes with known functions are shown. (c) The maximum likelihood phylogenetic tree was calculated using the concatenation of 1,596 core genes present in all 926 genomes and rooted using S. bongori. The scale bar represents nucleotide substitutions per site. The outer ring shows the different subspecies identified by Alikhan et al. (12). For visual clarity, only intersubspecies highways of recombination events identified by fastGEAR are shown (as gray arrowlines), and nonhighway recombination pairs are not shown. Inferred recipient genomes are indicated by the arrowheads.