Dataset Information

Quasimetagenomics-Based and Real-Time-Sequencing-Aided Detection and Subtyping of Salmonella enterica from Food Samples.

ABSTRACT: Metagenomics analysis of food samples promises isolation-independent detection and subtyping of foodborne bacterial pathogens in a single workflow. The selective concentration of Salmonella genomic DNA by immunomagnetic separation (IMS) and multiple displacement amplification (MDA) shortened the time for culture enrichment of Salmonella-spiked raw chicken breast samples by over 12 h while permitting serotyping and high-fidelity single nucleotide polymorphism (SNP) typing of the pathogen using short shotgun sequencing reads. The herein-termed quasimetagenomics approach was evaluated on Salmonella-spiked lettuce and black peppercorn samples as well as retail chicken parts naturally contaminated with different serotypes of Salmonella Culture enrichment of between 8 and 24 h was required for detecting and subtyping naturally occurring Salmonella from unspiked chicken parts compared with 4- to 12-h culture enrichment when Salmonella-spiked food samples were analyzed, indicating the likely need for longer culture enrichment to revive low levels of stressed or injured Salmonella cells in food. A further acceleration of the workflow was achieved by real-time nanopore sequencing. After 1.5 h of analysis on a potable sequencer, sufficient data were generated from sequencing the IMS-MDA products of a cultured-enriched lettuce sample to enable serotyping and robust phylogenetic placement of the inoculated isolate.IMPORTANCE Both culture enrichment and next-generation sequencing remain time-consuming processes for food testing, whereas rapid methods for pathogen detection are widely available. Our study demonstrated a substantial acceleration of these processes by the use of immunomagnetic separation (IMS) with multiple displacement amplification (MDA) and real-time nanopore sequencing. In one example, the combined use of the two methods delivered a less than 24-h turnaround time from the collection of a Salmonella-contaminated lettuce sample to the phylogenetic identification of the pathogen. An improved efficiency such as this is important for further expanding the use of whole-genome and metagenomics sequencing in the microbial analysis of food. Our results suggest the potential of the quasimetagenomics approach in areas where rapid detection and subtyping of foodborne pathogens are important, such as for foodborne outbreak response and the precision tracking and monitoring of foodborne pathogens in production environments and supply chains.

SUBMITTER: Hyeon JY

PROVIDER: S-EPMC5795075 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Quasimetagenomics-Based and Real-Time-Sequencing-Aided Detection and Subtyping of Salmonella enterica from Food Samples.

Hyeon Ji-Yeon JY Li Shaoting S Mann David A DA Zhang Shaokang S Li Zhen Z Chen Yi Y Deng Xiangyu X

Applied and environmental microbiology 20180131 4

Metagenomics analysis of food samples promises isolation-independent detection and subtyping of foodborne bacterial pathogens in a single workflow. The selective concentration of <i>Salmonella</i> genomic DNA by immunomagnetic separation (IMS) and multiple displacement amplification (MDA) shortened the time for culture enrichment of <i>Salmonella</i>-spiked raw chicken breast samples by over 12 h while permitting serotyping and high-fidelity single nucleotide polymorphism (SNP) typing of the pat ...[more]

PMID: 29196295

Similar Datasets

Project description:Food safety is a new area for novel applications of metagenomics analysis, which not only can detect and subtype foodborne pathogens in a single workflow but may also produce additional information with in-depth analysis capabilities. In this study, we applied a quasimetagenomic approach by combining short-term enrichment, immunomagnetic separation (IMS), multiple-displacement amplification (MDA), and nanopore sequencing real-time analysis for simultaneous detection of Salmonella and Escherichia coli in wheat flour. Tryptic soy broth was selected for the 12-h enrichment of samples at 42°C. Enrichments were subjected to IMS using beads capable of capturing both Salmonella and E. coli MDA was performed on harvested beads, and amplified DNA fragments were subjected to DNA library preparation for sequencing. Sequencing was performed on a portable device with real-time basecalling adaptability, and resulting sequences were subjected to two parallel pipelines for further analysis. After 1 h of sequencing, the quasimetagenomic approach could detect all targets inoculated at approximately 1 CFU/g flour to the species level. Discriminatory power was determined by simultaneous detection of dual inoculums of Salmonella and E. coli, absence of detection in control samples, and consistency in microbial flora composition of the same flour samples over several rounds of experiments. The total turnaround time for detection was approximately 20 h. Longer sequencing for up to 15 h enabled serotyping for many of the samples with more than 99% genome coverage, which could be subjected to other appropriate genetic analysis pipelines in less than a total of 36 h.IMPORTANCE Enterohemorrhagic Escherichia coli (EHEC) and Salmonella are of serious concern in low-moisture foods, including wheat flour and its related products, causing illnesses, outbreaks, and recalls. The development of advanced detection methods based on molecular principles of analysis is essential to incorporate into interventions intended to reduce the risk from these pathogens. In this work, a quasimetagenomic method based on real-time sequencing analysis and assisted by magnetic capture and DNA amplification was developed. This protocol is capable of detecting multiple Salmonella and/or E. coli organisms in the sample within less than a day, and it can also generate sufficient whole-genome sequences of the target organisms suitable for subsequent bioinformatics analysis. Multiplex detection and identification were accomplished in less than 20 h and additional whole-genome analyses of different nature were attained within 36 h, in contrast to the several days required in previous sequencing pipelines.

Dataset Information

Quasimetagenomics-Based and Real-Time-Sequencing-Aided Detection and Subtyping of Salmonella enterica from Food Samples.

Publications

Quasimetagenomics-Based and Real-Time-Sequencing-Aided Detection and Subtyping of Salmonella enterica from Food Samples.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets