U.S. flag An official website of the United States government

On Oct. 1, 2024, the FDA began implementing a reorganization impacting many parts of the agency. We are in the process of updating FDA.gov content to reflect these changes.

U.S. Food and Drug Administration
  1. Home
  2. Science & Research
  3. About Science & Research at FDA
  4. The FDA Science Forum
  5. The plasmid puzzle: challenges for Salmonella monitoring
  1. The FDA Science Forum

2023 FDA Science Forum

The plasmid puzzle: challenges for Salmonella monitoring

Download the Poster (PDF; 0.49 MB)
Authors:
Poster Author(s)
Kristina Feye, FDA/NCTR, Danielle Sopovski FDA/NCTR, Javier Revollo FDA/NCTR, Jaimie Miranda-Colon FDA/NCTR, Dereje Gudeta FDA/NCTR, Jing Han FDA/NCTR, Steven Foley FDA/NCTR
Center:
Contributing Office
National Center for Toxicological Research

Abstract

Poster Abstract

Salmonella is known to harbor small and large plasmids of different incompatibility groups. Fourth generation sequencing technologies are actively contributing to food safety monitoring. However, small and large plasmids may present a number of challenges to both sequencing platforms. In order to understand how different sequencing platforms impact Salmonella sequence data, we evaluated how PacBio Sequel II whole genome sequencing compares to traditional Illumina sequencing pipelines. Ten colony forming units from ten strains of Salmonella were sequenced. The plasmids from the strains were predicted to range from 6 KB to over 400,000 KB. This distribution of plasmids that were predicted within these strains should have both difficult and easy to identify plasmids. Putatively identified plasmids were harvested from the sequences and a BLAST analysis was performed to identify the plasmid and PCR was used to confirm plasmid replicon type. The plasmids were bioinformatically predicted via two independent plasmid programs, MOB-SUITE and PlasmidFinder. The resulting data varied profoundly. For example, an isolate had six plasmid replicons identified via the Illumina platform, whereas PacBio only identified four plasmids (PacBio: IncC, IncFIA, Inc FIB, IncX4 vs. Illumina: IncA/C, IncFIA, IncFIB, IncX4, ColpVC and IncFII (pCoo) using the same annotation pipeline (PlasmidFinder). When using MOB-SUITE (Typer), plasmid annotations shifted significantly in PacBio sequences, with plasmid replicons identified from the same strain via PacBio as: IncFIB, IncFII, IncFIA, ColRNAI_rep_cluster_1857, MOBP, IncA/C2, and IncX4. Two of the plasmids identified by their replicons appear to be re-arranged and merged (ColRNAI_rep_cluster_1857 and MOBP). The overall predicted sizes range for that particular isolate were from 3.176 to 126.356 kb with PacBio and 2 to 120 kb when sequenced via Illumina MiSeq for a single isolate. Each tested strain exhibited heterogeneity in the plasmids and the annotations. Additionally, mobility predictions and antibiotic resistance annotations on each plasmid (Abricate) varied significantly on the plasmids between the sequencing and bioinformatics platforms. Some of this is expected as different bioinformatics platforms can result in different data as programs rely on different means for annotation. Yet, basic plasmid identification and annotation challenges are identified in this study and are likely driven by sequencing platform limitations and that may require benchtop validations to overcome sequencing pitfalls.

Poster Image
The plasmid puzzle: challenges for Salmonella monitoring

Download the Poster (PDF; 0.49 MB)

Back to Top