Evaluation of nanopore sequencing technology to identify Salmonella enterica Choleraesuis var. Kunzendorf and Orion var. 15, 34.

Our previous study demonstrated that whole genome sequencing (WGS) data generated by Oxford Nanopore Technologies (ONT) can be used for rapid and accurate prediction of selected Salmonella serotypes. However, one limitation is that established methods for WGS-based serotype prediction, utilizing data from either ONT or Illumina, cannot differentiate certain serotypes and serotype variants with the same or closely related antigenic formulae. This study aimed to evaluate nanopore sequencing and additional data analysis for identification of Salmonella enterica Choleraesuis var. Kunzendorf and S. enterica Orion var. 15, 34, thus overcoming this limitation. Five workflows that combined different flow cells, library construction methods and basecaller models were evaluated and compared. The workflow that consisted of the R9 flow cell, rapid sequencing library construction kit and guppy basecaller with base modified model performed best for Single Nucleotide Polymorphism (SNP) analysis. With this workflow, 99.98% of matching identity between assembled genomes from ONT and that from Illumina was achieved. Less than five high-quality SNPs differed when comparing sequencing data between ONT and Illumina. SNP typing successfully identified Choleraesuis var. Kunzendorf. While prophage prediction further differentiated Orion var. 15, 34 from the other two Orion variants. Our study improves the readiness of ONT as a Salmonella subtyping and source tracking tool for food industry applications.