Epidemic multiple drug resistant Salmonella Typhimurium causing invasive disease in sub-Saharan Africa have a distinct genotype

Abstract

Whereas most nontyphoidal Salmonella (NTS) are associated with gastroenteritis, there has been a dramatic increase in reports of NTS-associated invasive disease in sub-Saharan Africa. Salmonella enterica serovar Typhimurium isolates are responsible for a significant proportion of the reported invasive NTS in this region. Multilocus sequence analysis of invasive S. Typhimurium from Malawi and Kenya identified a dominant type, designated ST313, which currently is rarely reported outside of Africa. Whole-genome sequencing of a multiple drug resistant (MDR) ST313 NTS isolate, D23580, identified a distinct prophage repertoire and a composite genetic element encoding MDR genes located on a virulence-associated plasmid. Further, there was evidence of genome degradation, including pseudogene formation and chromosomal deletions, when compared with other S. Typhimurium genome sequences. Some of this genome degradation involved genes previously implicated in virulence of S. Typhimurium or genes for which the orthologs in S. Typhi are either pseudogenes or are absent. Genome analysis of other epidemic ST313 isolates from Malawi and Kenya provided evidence for microevolution and clonal replacement in the field.

Figure 1.

Radial phylogram showing the phylogenetic relationship of S. enterica serovar Typhimurium sequence types (STs). The S. Typhimurium STs (red and yellow circles) are rooted to S. Typhi (blue circle). The length of the connecting line is proportional to the degree of divergence. S. Typhimurium STs from various worldwide locations outside sub-Saharan Africa SSA (red) and NTS STs predominantly from SSA (yellow) are indicated.

Figure 2.

Whole-genome comparison of S. Typhimurium LT2, D23580, and DT104 NCTC13348. ACT comparison (http://www.sanger.ac.uk/Software/ACT/) of amino acid matches between the complete six-frame translations (computed using BLASTN) of the whole genome sequences of S. Typhimurium LT2 (LT2), D23580, and DT104 NCTC13348. The red bars between the DNA lines represent individual TBLASTX matches, with inverted matches colored blue. Large chromosomal features are indicated with a horizontal line labeled with the feature identity. These analyses indicate the overall colinearity and synteny of the chromosome of sequenced S. Typhimurium isolates. Large insertions or deletions (mainly prophage elements) are labeled. Smaller indels described in the text are not visible at this resolution.

Figure 3.

Radial phylogram based on chromosomal SNPs showing the phylogenetic relationship of sequenced S. Typhimurium isolates. Branch lengths indicate the number of SNPs, scale as indicated, separating the sequenced isolates indicated by circles. ST19 isolates (red circles) and ST313 isolates (yellow circles) are shown. The black square indicates the ancestral root.

Figure 4.

Genes and homologous regions of pSLT-BT and pSKT-A130. A circular map of pSLT-BT shows predicted genes on the forward and reverse strands on the outer circle (yellow). Genes with putative antibiotic resistance functions (red), integron- or transposon-related functions (blue), plasmid-associated functions (green), antiseptic and mercury resistance functions (purple), miscellaneous functions (gray), and pseudogenes (brown) are shown. Arrows below the pSLT-BT composite element gene map indicate the annealing site of oligonucleotide primers used to probe clinical isolates. Regions of each locus with similarity to Tn21 are indicated.