Multi-laboratory validation study of a real-time PCR method for detection of Salmonella in baby spinach

Deng, Kaiping; Wang, Shizhen  Steven; Kiener, Shannon; Smith, Emily; Chen, Kai-Shun; Pamboukian, Ruiqing; Laasri, Anna; Pelaez, Catalina; Ulaszek, Jodie; Kmet, Matthew; Jesús, Antonio de; Hammack, Thomas S.; Reddy, Ravinder; Wang, Hua

doi:10.1016/j.fm.2023.104299

Cited by 4 publications

(1 citation statement)

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“…There was good concordance between detection via culturing and multiplex qPCR on enriched materials, as has been seen with various qPCR methods [45, 46, 47, 48]. Although culturing, qPCR, and sequencing of the 16S rRNA V3-V4 region had equivalent LODs when tested on spiked enriched feed samples, only qPCR was able to match culturing results when used on naturally contaminated samples.…”

Section: Discussionmentioning

confidence: 56%

Limit of detection ofSalmonellaser. Enteritidis using culture-based versus culture-independent diagnostic approaches

Bradford,

Yao,

Anastasiadis

et al. 2024

Preprint

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In order to prevent the spread of foodborne illnesses, the presence of pathogens in the food chain is monitored by government agencies and food producers. The culture-based methods currently employed are sensitive but time- and labour- intensive, leading to increasing interest in exploring culture-independent diagnostic tests (CIDTs) for pathogen detection. However, sensitivity and reliability of these CIDTs relative to current approaches has not been well established. To address this issue, we conducted a comparison of the limit of detection (LOD50) forSalmonellabetween a culture-based method and three CIDT methods: qPCR (targetinginvAandstn), metabarcode (16S) sequencing, and shotgun metagenomic sequencing. Samples of chicken feed and chicken caecal contents were spiked withSalmonellaserovar Enteritidis and subjected to culture- and DNA-based detection methods. To explore the impact of non-selective enrichment on LOD50, all samples underwent both immediate DNA extraction and an overnight enrichment prior to gDNA extraction. In addition to this spike-in experiment, feed and caecal samples acquired from the field were tested with culturing, qPCR, and metabarcoding. In general, LOD50was comparable between qPCR and shotgun sequencing methods. Overnight microbiological enrichment resulted in an improvement in LOD50with up to a three log decrease, comparable to culture-based detection. However,Salmonellareads were detected in some unspiked feed samples, suggesting false-positive detection ofSalmonella. Additionally, the LOD50in feeds was three logs lower than in caecal contents, underscoring the impact of background microbiota onSalmonelladetection using all methods.

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