Incidence and Genomic Background of Antibiotic Resistance in Food-Borne and Clinical Isolates of Salmonella enterica Serovar Derby from Spain

Abstract

Salmonella enterica serovar Derby (S. Derby) ranks fifth among nontyphoidal Salmonella serovars causing human infections in the European Union. S. Derby isolates (36) collected between 2006 and 2018 in a Spanish region (Asturias) from human clinical samples (20) as well as from pig carcasses, pork- or pork and beef-derived products, or wild boar (16) were phenotypically characterized with regard to resistance, and 22 (12 derived from humans and 10 from food-related samples) were also subjected to whole genome sequence analysis. The sequenced isolates belonged to ST40, a common S. Derby sequence type, and were positive for SPI-23, a Salmonella pathogenicity island involved in adherence and invasion of the porcine jejune enterocytes. Isolates were either susceptible (30.6%), or resistant to one or more of the 19 antibiotics tested for (69.4%). Resistances to tetracycline [tet(A), tet(B) and tet(C)], streptomycin (aadA2), sulfonamides (sul1), nalidixic acid [gyrA (Asp87 to Asn)] and ampicillin (bla_TEM-1-like) were detected, with frequencies ranging from 8.3% to 66.7%, and were higher in clinical than in food-borne isolates. The fosA7.3 gene was present in all sequenced isolates. The most common phenotype was that conferred by the tet(A), aadA2 and sul1 genes, located within identical or closely related variants of Salmonella Genomic Island 1 (SGI1), where mercury resistance genes were also present. Diverse IncI1-I(α) plasmids belonging to distinct STs provided antibiotic [bla_TEM-1, tet(A) and/or tet(B)] and heavy metal resistance genes (copper and silver), while small pSC101-like plasmids carried tet(C). Regardless of their location, most resistance genes were associated with genetic elements involved in DNA mobility, including a class one integron, multiple insertion sequences and several intact or truncated transposons. By phylogenetic analysis, the isolates were distributed into two distinct clades, both including food-borne and clinical isolates. One of these clades included all SGI1-like positive isolates, which were found in both kinds of samples throughout the entire period of study. Although the frequency of S. Derby in Asturias was very low (0.5% and 3.1% of the total clinical and food isolates of S. enterica recovered along the period of study), it still represents a burden to human health linked to transmission across the food chain. The information generated in the present study can support further epidemiological surveillance aimed to control this zoonotic pathogen.

Keywords: IncI1-I(α); SGI1-like; SPI-23; ST40; Salmonella enterica serovar Derby; antimicrobial drug resistance; fosA7.3; pSC101-like; phylogenetic analysis; resistance plasmid.

Figure 1

Genetic environment of chromosomal resistance genes found in S. Derby isolated from human clinical samples and food samples in Spain. (A). Genetic organization of the SGI1-variant, using LSP 138/08 as a model. (B). Comparison of the genetic context of fosA7.3 in LSP 82/16 (used as a model) and LSP 25/16. The alignments were created with Easyfig BLASTn. The gray shading between regions reflects nucleotide sequence identities according to the scale shown at the right lower corner of the figure. Genes are represented by arrows pointing to the direction of transcription. Genes with similar functions are shown in the same color. Color code: yellow, DNA replication; brown, conjugal transfer; blue, genes involved in DNA mobility; red, resistance genes; grey, other functions; white, hypothetical proteins.

Figure 2

Genetic map of resistance genes carried by plasmids in S. Derby recovered from food and human clinical samples in Spain. Genes are represented by arrows pointing to the direction of transcription and color-coded according to function (see below). (A). Comparison of the resistance regions found in IncI1-I(α) plasmids of bla_TEM-1-positive isolates. The alignments were created with Easyfig BLASTn. The gray shading between regions shows nucleotide sequence identities according to the scale shown at the right lower corner of the figure. (B). Resistance region of the tet(B)-positive IncI1-I(α) plasmids found in LSP 218/06 and LSP 217/09. (C). pSC101-like plasmid of LSP 25/16 used as a model. Color code: yellow, plasmid replication and maintenance; brown, conjugational transfer; blue, DNA mobility; red, resistance; grey, other functions; white, hypothetical proteins.

Figure 3

Phylogenetic tree showing the relationships between S. Derby isolates collected from food and human clinical samples in Spain. The SNP (single nucleotide polymorphism)-based tree was constructed with CSI Phylogeny 1.4 (https://cge.cbs.dtu.dk/services/CSIPhylogeny/, accessed on 10 May 2023), using the genome of S. Derby strain LSP 138/08 as the reference. Numbers at the nodes represent bootstrap values based on 1000 replicates. The observed clades (A and B) and subclades (B1 and B2) are indicated. The SNP similarity matrix is shown in Table S2. Human isolates are highlighted in bold.