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Comparison of conventional molecular and whole-genome sequencing methods for subtyping Salmonella enterica serovar Enteritidis strains from Tunisia

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Abstract

We sought to determine the relative value of conventional molecular methods and whole-genome sequencing (WGS) for subtyping Salmonella enterica serovar Enteritidis recovered from 2000 to 2015 in Tunisia and to investigate the genetic diversity of this serotype. A total of 175 Salmonella Enteritidis isolates were recovered from human, animal, and foodborne outbreak samples. Pulsed-field gel electrophoresis (PFGE), multiple locus variable-number tandem repeat analysis (MLVA), and whole-genome sequencing were performed. Eight pulsotypes were detected for all isolates with PFGE (DI = 0.518). Forty-five Salmonella Enteritidis isolates were selected for the MLVA and WGS techniques. Eighteen MLVA profiles were identified and classified into two major clusters (DI = 0.889). Core genome multilocus typing (cgMLST) analysis revealed 16 profiles (DI = 0.785). Whole-genome analysis indicated 660 single-nucleotide polymorphism (SNP) divergences dividing these isolates into 43 haplotypes (DI = 0.997). The phylogenetic tree supported the classification of Salmonella Enteritidis isolates into two distinct lineages subdivided into five clades and seven subclades. Pairwise SNP differences between the isolates ranged between 302 and 350. We observed about 311 SNP differences between the two foodborne outbreaks, while only less or equal to 4 SNP differences within each outbreak. SNP-based WGS typing showed an excellent discriminatory power comparing with the conventional methods such as PFGE and MLVA. Besides, we demonstrate the added value of WGS as a complementary subtyping method to discriminate outbreak from non-outbreak isolates belonging to common subtypes. It is important to continue the survey of Salmonella Enteritidis lineages in Tunisia using WGS.

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Acknowledgments

We thank the Veterinary Research team of Sfax-Tunisia and especially Dr. Sonia Zormati for providing animal Salmonella Enteritidis isolates.

Funding

This work was supported financially by the Research Laboratory Microorganisms and Human Disease “MPH LR03SP03” - Higher Education and Scientific Research in Tunisia.

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Authors and Affiliations

  1. Habib Bourguiba University Hospital, Research Laboratory Microorganisms and Human Disease, University of Sfax, Sfax, Tunisia

    Boutheina Ksibi, Sonia Ktari, Sonda Maalej, Basma Mnif, Faouzia Rhimi & Adnene Hammami

  2. Faculty of Medicine Sfax, Laboratory of Microbiology, Avenue Majida Boulila, 3027, Sfax, Tunisia

    Sonia Ktari

  3. Sydney Brenner Institute for Molecular Bioscience, Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, South Africa

    Houcemeddine Othman

  4. Laboratory of Bioinformatics, Biomathematics and Biostatistics, Institut Pasteur de Tunis, University of Tunis El Manar (UTM), Tunis, Tunisia

    Kais Ghedira

  5. Veterinary Research, Tunis, Tunisia

    Mohamed salah Abbassi

  6. Centre National de Référence des Escherichia coli, Shigella and Salmonella, Unité des Bactéries Pathogènes Entériques, Institut Pasteur, 75015, Paris, France

    Laetitia Fabre & Simon Le Hello

  7. Groupe de Recherche sur l’Adaptation Microbienne (GRAM 2.0), Normandie Univ, UNICAEN, UNIROUEN, GRAM 2.0, 14000, Caen, France

    Simon Le Hello

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  1. Boutheina Ksibi
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Correspondence to Sonia Ktari.

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Fig. S1

Phylogenetic results of Salmonella Enteritidis clustering isolates (lineage A), NCBI Pathogen detection cluster PDS000004725.23. The scale bar indicates the number of substitutions per site, where the branch length is proportional to the number of SNPs present. Detailed SNP differences also are available at the NCBI Pathogen Detection site by searching for either strain IDs or SNP cluster IDs. (PDF 1774 kb)

Fig. S2

Phylogenetic results of Salmonella Enteritidis clustering isolates (lineage B), NCBI Pathogen detection cluster PDS000056272.14. The scale bar indicates the number of substitutions per site where the branch length is proportional to the number of SNPs present. Detailed SNP differences also are available at the NCBI Pathogen Detection site by searching for either strain IDs or SNP cluster IDs. (PDF 2048 kb)

Fig. S3

Population structure of Salmonella Enteritidis isolates. (A) Maximum-likelihood tree of chromosomal sequences (B) Maximum-likelihood tree of plasmid sequences (PDF 638 kb)

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Ksibi, B., Ktari, S., Othman, H. et al. Comparison of conventional molecular and whole-genome sequencing methods for subtyping Salmonella enterica serovar Enteritidis strains from Tunisia. Eur J Clin Microbiol Infect Dis 40, 597–606 (2021). https://doi.org/10.1007/s10096-020-04055-8

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