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ABSTRACT: The study of non-typhoid Salmonella in broiler integrations has been limited by the resolution of typing techniques. Although serotyping of Salmonella isolates is used as a traditional approach, it is not of enough resolution to clearly understand the dynamics of this pathogen within poultry companies. The aim of this research was to investigate the epidemiology and population dynamics of Salmonella serotypes in 2 poultry integrations using a whole genome sequencing approach. Two hundred and forty-three Salmonella isolates recovered from the broiler production chain of 2 integrated poultry companies were whole genome sequenced and analyzed with dedicated databases and bioinformatic software. The analyses of sequences revealed that S. Infantis was the most frequent serotype (82.3%). Most isolates showed a potential for resistance against medically important antibiotics and disinfectants. Furthermore, 97.5% of isolates harbored the pESI-like mega plasmid, that plays an important role in the global dissemination of AMR. SNP tree analysis showed that there were clones that are niche-specific while other ones were distributed throughout the broiler production chains. In this study, we demonstrated the potential of whole genome sequencing analysis for a comprehensive understanding of Salmonella distribution in integrated poultry companies. Data obtained with these techniques allow determination of the presence of genetic factors that play an important role in the environmental fitness and pathogenicity of Salmonella.

Listeria monocytogenes is a foodborne pathogen with a highly clonal population structure comprising multiple phylogenetic sub-groups that can persist within food processing environments and contaminate food. The epidemiology of L. monocytogenes is well-described in some developed countries; however, little is known about the prevalence and population structure of this pathogen in food and food processing environments located in less developed regions. The aim of this study was to determine the genetic characteristics and clonal relatedness of L. monocytogenes that were isolated from two Jamaican meat processing facilities. Of the 37 isolates collected between 2011 and 2015, only a single lineage II isolate was recovered (serotype 1/2c), and the remaining were lineage I isolates representing serotypes 4b, 1/2b, 3b, and two untypeable isolates. Pulsed-field gel electrophoresis (PFGE) delineated isolates into seven pulsotypes, and whole-genome sequencing (WGS) categorized most isolates within one of three clonal complexes (CC): CC2 (N = 12), CC5 (N = 11), and CC288 (N = 11). Isolates representing CC1 (N = 2) and CC9 (N = 1) were also recovered. Virulence-associated genes such as inlA and the LIPI-3 cluster were detected in multiple isolates, along with the stress survival islet cluster-1 (SSI-1), and benzalkonium (bcrABC) and cadmium (cad1, cad2, cad4) resistance cassettes. Multiple isolates that belong to the same CC and matching PFGE patterns were isolated from food and the environment from both facilities across multiple years, suggesting the presence of persistent strains of L. monocytogenes, and/or constant re-entry of the pathogens into the facilities from common sources. These findings highlight the ability of lineage I isolates of L. monocytogenes to colonize, persist, and predominate within two meat-producing environments, and underscores the need for robust surveillance strategies to monitor and mitigate against these important foodborne pathogens.

Salmonella Infantis, a common contaminant of poultry products, is known to harbor mobile genetic elements that confer multi-drug resistance (MDR) and have been detected in many continents. Here, we report four MDR S. Infantis strains recovered from poultry house environments in Santa Cruz Island of the Galapagos showing extended-spectrum β-lactamase (ESBL) resistance and reduced fluoroquinolone susceptibility. Whole-genome sequencing (WGS) revealed the presence of the ESBL-conferring blaCTX-M-65 gene in an IncFIB-like plasmid in three S. Infantis isolates. Multi-locus sequence typing (MLST) and single nucleotide variant/polymorphism (SNP) SNVPhyl analysis showed that the S. Infantis isolates belong to sequence type ST32, likely share a common ancestor, and are closely related (1–3 SNP difference) to blaCTX-M-65-containing clinical and veterinary S. Infantis isolates from the United States and Latin America. Furthermore, phylogenetic analysis of SNPs following core-genome alignment (i.e., ParSNP) inferred close relatedness between the S. Infantis isolates from Galapagos and the United States. Prophage typing confirmed the close relationship among the Galapagos S. Infantis and was useful in distinguishing them from the United States isolates. This is the first report of MDR blaCTX-M-65-containing S. Infantis in the Galapagos Islands and highlights the need for increased monitoring and surveillance programs to determine prevalence, sources, and reservoirs of MDR pathogens.

The emergence of multidrug-resistant bacterial strains worldwide has become a serious problem for public health over recent decades. The increase in antimicrobial resistance has been expanding via plasmids as mobile genetic elements encoding antimicrobial resistance (AMR) genes that are transferred vertically and horizontally. This study focuses on Salmonella enterica, one of the leading foodborne pathogens in industrialized countries. S. enterica is known to carry several plasmids involved not only in virulence but also in AMR. In the current paper, we present an integrated strategy to detect plasmid scaffolds in whole genome sequencing (WGS) assemblies. We developed a two-step procedure to predict plasmids based on i) the presence of essential elements for plasmid replication and mobility, as well as ii) sequence similarity to a reference plasmid. Next, to confirm the accuracy of the prediction in 1750 S. enterica short-read sequencing data, we combined Oxford Nanopore MinION long-read sequencing with Illumina MiSeq short-read sequencing in hybrid assemblies for 84 isolates to evaluate the proportion of plasmid that has been detected. At least one scaffold with an origin of replication (ORI) was predicted in 61.3% of the Salmonella isolates tested. The results indicated that IncFII and IncI1 ORIs were distributed in many S. enterica serotypes and were the most prevalent AMR genes carrier, whereas IncHI2A/IncHI2 and IncA/C2 were more serotype restricted but bore several AMR genes. Comparison between hybrid and short-read assemblies revealed that 81.1% of plasmids were found in the short-read sequencing using our pipeline. Through this process, we established that plasmids are prevalent in S. enterica and we also substantially expand the AMR genes in the resistome of this species.

Non-typhoidal Salmonella is a leading cause of foodborne illness worldwide. Prompt and accurate identification of the sources of Salmonella responsible for disease outbreaks is crucial to minimize infections and eliminate ongoing sources of contamination. Current subtyping tools including single nucleotide polymorphism (SNP) typing may be inadequate, in some instances, to provide the required discrimination among epidemiologically unrelated Salmonella strains. Prophage genes represent the majority of the accessory genes in bacteria genomes and have potential to be used as high discrimination markers in Salmonella. In this study, the prophage sequence diversity in different Salmonella serovars and genetically related strains was investigated. Using whole genome sequences of 1,760 isolates of S. enterica representing 151 Salmonella serovars and 66 closely related bacteria, prophage sequences were identified from assembled contigs using PHASTER. We detected 154 different prophages in S. enterica genomes. Prophage sequences were highly variable among S. enterica serovars with a median ± interquartile range (IQR) of 5 ± 3 prophage regions per genome. While some prophage sequences were highly conserved among the strains of specific serovars, few regions were lineage specific. Therefore, strains belonging to each serovar could be clustered separately based on their prophage content. Analysis of S. Enteritidis isolates from seven outbreaks generated distinct prophage profiles for each outbreak. Taken altogether, the diversity of the prophage sequences correlates with genome diversity. Prophage repertoires provide an additional marker for differentiating S. enterica subtypes during foodborne outbreaks.

The Salmonella Syst-OMICS consortium is sequencing 4,500 Salmonella genomes and building an analysis pipeline for the study of Salmonella genome evolution, antibiotic resistance and virulence genes. Metadata, including phenotypic as well as genomic data, for isolates of the collection are provided through the Salmonella Foodborne Syst-OMICS database (SalFoS), at https://salfos.ibis.ulaval.ca/. Here, we present our strategy and the analysis of the first 3,377 genomes. Our data will be used to draw potential links between strains found in fresh produce, humans, animals and the environment. The ultimate goals are to understand how Salmonella evolves over time, improve the accuracy of diagnostic methods, develop control methods in the field, and identify prognostic markers for evidence-based decisions in epidemiology and surveillance.