Metagenomics-Based Proficiency Test of Smoked Salmon Spiked with a Mock Community.

Claudia Sala,Alessandra De Cesare,Clara Amid,Gastone Castellani,Sünje Johanna Pamp,Andreas Nitsche,Richard J Ellis,Soizick Le Guyader,Fiona Crispie,Paul D Cotter,Hanne Mordhorst,Julien Schaeffer,Gerardo Manfreda,Catherine Ragimbeau,Annika Brinkmann,Mikhayil Hakhverdyan,Josephine Grützke,Moon Y F Tay,Joël Mossong ,Casper Sahl Poulsen ,Jørgen Schlundt ,René S Hendriksen ,Thomas Nordahl Petersen ,Frank Møller Aarestrup

doi:10.3390/microorganisms8121861

Abstract

An inter-laboratory proficiency test was organized to assess the ability of participants to perform shotgun metagenomic sequencing of cold smoked salmon, experimentally spiked with a mock community composed of six bacteria, one parasite, one yeast, one DNA, and two RNA viruses. Each participant applied its in-house wet-lab workflow(s) to obtain the metagenomic dataset(s), which were then collected and analyzed using MG-RAST. A total of 27 datasets were analyzed. Sample pre-processing, DNA extraction protocol, library preparation kit, and sequencing platform, influenced the abundance of specific microorganisms of the mock community. Our results highlight that despite differences in wet-lab protocols, the reads corresponding to the mock community members spiked in the cold smoked salmon, were both detected and quantified in terms of relative abundance, in the metagenomic datasets, proving the suitability of shotgun metagenomic sequencing as a genomic tool to detect microorganisms belonging to different domains in the same food matrix. The implementation of standardized wet-lab protocols would highly facilitate the comparability of shotgun metagenomic sequencing dataset across laboratories and sectors. Moreover, there is a need for clearly defining a sequencing reads threshold, to consider pathogens as detected or undetected in a food sample.

Highlights

Foodborne illnesses associated with pathogenic microorganisms are a global public health and economic challenge
An inter-laboratory proficiency test was organized to assess the ability of participants to perform shotgun metagenomic sequencing of cold smoked salmon, experimentally spiked with a mock community composed of six bacteria, one parasite, one yeast, one DNA, and two RNA viruses
Our results highlight that despite differences in wet-lab protocols, the reads corresponding to the mock community members spiked in the cold smoked salmon, were both detected and quantified in terms of relative abundance, in the metagenomic datasets, proving the suitability of shotgun metagenomic sequencing as a genomic tool to detect microorganisms belonging to different domains in the same food matrix

Summary

Introduction

Foodborne illnesses associated with pathogenic microorganisms are a global public health and economic challenge. The techniques used for the detection and characterization of foodborne pathogens in food products evolved tremendously over the past several decades, but they generally focused on the detection of a single pathogen or just a few pathogens at a time [2]. This analytical restriction hampers the mapping of shifting microbial communities, which potentially affect the persistence of foodborne pathogens in the food production chain [2,3], and can result in pathogens being overlooked by virtue of being novel or not being traditionally associated with a particular environment. At present, deconvoluting the metagenomic data to definitively associate those markers with the same genome is still challenging [5,8]

Methods

Results

Conclusion