Genomic survey of multidrug resistant Salmonella enterica serovar Minnesota clones in chicken products

Abstract

Salmonella enterica serovar Minnesota (S. Minnesota) is an emerging serovar that persists within poultry supply chains, potentially causing outbreaks in humans. Understanding its population genomics is crucial for designing preventive measures. We performed a genomic surveillance study of S. Minnesota by analyzing 259 isolates from poultry in Saudi Arabia. Whole-genome sequencing data for these isolates were analyzed to characterize emerging clones and the genetic factors underlying antimicrobial resistance and virulence. We compared the isolates to all available global genomes of S. Minnesota. Our results revealed the emergence of four clones, three of which were mixed with global strains. These clones exhibited higher levels of antimicrobial resistance and virulence due to the acquisition of multiple plasmids, particularly IncC plasmids, carrying resistance and virulence genes. IncC plasmids underwent genomic rearrangements, presenting diverse configurations of resistance genes. Our findings demonstrate the emergence and persistence of pathogenic and multidrug-resistant S. Minnesota clones.

Fig. 1. The midpoint-rooted minimum spanning tree reconstructed from the mean SNP distances between genome pairs in different BAPS clones, shown in Fig. S1A.

The boxplots for age show the distribution of the years of collection for the strains in each BAPS clone. The boxplots for diversity correspond to pairwise SNP distances between genomes within each clone. The boxplots for the count (#) of virulence and resistance genes show genes identified by the genomic pipelines. Internal genomes are those sequenced as part of this study, while external genomes are from public databases (EnteroBase/PathogenDetection). The * and *** signs on the each panel denote significance levels of <0.05 and <0.01, respectively, based on a one-sided Wilcoxon ranked test. These levels indicate the significance of differences in age, diversity, virulence factor gene count, and resistance gene count for isolates in the BAPS1–6 clones (which included isolates from the current study) compared to the other BAPS clones (BAPS7–11).

Fig. 2. Phylodynamic analysis of the clones circulating in Saudi Arabia.

A Estimated clone age for the four clones containing Saudi isolates. Error bars represent the 95% highest posterior density (HPD). B Skyline growth plot depicting changes in population size over time. The shaded region indicates the 95% confidence interval. C Phylodynamic trees for BAPS2 and BAPS4 clones, including genomes from the global collection. Internal branch colors denote the inferred status of the country of origin, with horizontal red bars indicating the 95% HPD.

Fig. 3. Transmission network for four BAPS clones.

Each node represents one genome, and edges indicate genetic relatedness, with a distance cutoff of 10 SNPs for defining the networks. External (global) isolates are colored gray. All external isolates in the BAPS4 clone originated from South Africa. White nodes within BAPS4 represent Brazilian isolates from the current study. Colored squares denote different brand codes/suppliers for isolates from the same city. Each color represents a distinct brand code/supplier.

Fig. 4. The distribution of.

A Antimicrobial resistance genes, B virulence factor genes that are variably present in the clones, and C plasmids across the BAPS clones. Bubble sizes correspond to relative frequency, with numbers indicating relative frequencies within each clone. The tree is the same as in Fig. 1. For the detection of virulence factor genes and antimicrobial resistance genes, SRST2 pipeline with a threshold of 90% for identity and coverage was used. The shaded squares in (A) indicate the genes that were significantly overrepresented in each clone compared to other clones, as determined by a one-sided proportion test with a significance level of 0.01. For plasmids in (C), short reads for each strain were mapped against one of the plasmid backbones from each cluster presented in Fig. S3A, using a cutoff of 90% coverage to determine the presence of the plasmid. Genome IDs on the x-axis in (C) correspond to the isolate from which the reference plasmid genome was obtained. For IncC plasmid, the shortest IncC plasmid backbone in the population was used as reference. The labels “chr” and “Cl.” denote “chromosomal” and “cluster” indicating the genomic context of the gene on chromosomal or plasmid, respectively. “Unk” denotes “unknown,” indicating cases where the genomic context could not be resolved due to either the absence of the genome in the collection or fragmented contigs.

Fig. 5. The structure and population genomics of IncC plasmids.

A Genomic map of the largest IncC plasmid (from isolate K_153) identified in the long-read sequencing data. B Linearized IncC plasmid with alignment of all IncC plasmids against the plasmid in (A). The bands for other plasmids are ordered from top to bottom and from left to right as indicated in the caption. Horizontal bars represent matches of 90% identity. Each color for the horizontal bars corresponds to the BAPS clones from which the plasmid was isolated. C Pangenome graph of IncC plasmids from 16 aligned IncC plasmids. Nodes and edges represent genes and their colocation within the same genomic context. Mobile genetic elements (MGE)s linked with resistance genes are colored blue.