A New Whole Genome Culture-Independent Diagnostic Test (WG-CIDT) for Rapid Detection of Salmonella in Lettuce.

Dele Ogunremi,Roger C Lévesque,Hongsheng Huang,Andrée Ann Dupras,Nur A Hasan,Imelda Galván Márquez,Marc-Olivier Duceppe,Katayoun Omidi,Sohail Naushad,Lawrence Goodridge,Manoj Dadlani,Sebastian Magierowski,Luke Masson,Brent Dixon,Ruimin Gao

doi:10.3389/fmicb.2020.00602

Abstract

The rapid detection of foodborne microbial pathogens contaminating fresh fruits and vegetables during the intervening period between harvest and consumption could revolutionize microbial quality assurance of food usually consumed raw and those with a limited shelf life. We have developed a sensitive, shotgun whole genome sequencing protocol capable of detecting as few as 1 colony forming unit (cfu) of Salmonella enterica serovar Typhimurium spiked on 25 g of lettuce. The Ion Torrent sequencing platform was used to generate reads of globally amplified DNA from microbes recovered from the surface of lettuce followed by bioinformatic analyses of the nucleotide sequences to detect the presence of Salmonella. The test is rapid and sensitive, and appropriate for testing perishable foods, and those consumed raw, for Salmonella contamination. The test has the potential to be universally applicable to any microbial contaminant on lettuce as long as a suitable bioinformatics pipeline is available and validated. A universal test is expected to pave the way for preventive and precision food safety and the re-shaping of the entire spectrum of food safety investigations from the current disease-limiting, reactive procedure to a proactive, disease prevention process.

Highlights

Consumption of fresh fruits and vegetables is increasingly linked with outbreaks of foodborne illnesses, and the bacterial pathogen Salmonella is one of the most commonly incriminated causes (Centers for Disease Control and Prevention, 2005; Lynch et al, 2009; Kozak et al, 2013)
The exact count of Salmonella delivered to the surface of lettuce was determined by plating a replicate aliquot of the bacterial suspension used for the spiking experiment on Brain Heart infusion (BHI) and Xylosine Lysine Tergitol-4 (XLT-4) agar plates followed by an overnight incubation at 37◦C and plate examination and colony enumeration the following day
Toward the goal of developing a rapid and robust WG-CIDT, we investigated and established conditions under which Salmonella DNA sequences present in a sample of lettuce (25 g) spiked with a low number of the bacterial cells can be reproducibly detected

Summary

Introduction

Consumption of fresh fruits and vegetables is increasingly linked with outbreaks of foodborne illnesses, and the bacterial pathogen Salmonella is one of the most commonly incriminated causes (Centers for Disease Control and Prevention, 2005; Lynch et al, 2009; Kozak et al, 2013). Over 2,600 serovars of Salmonella have been identified and, while only a restricted number of these serovars are commonly associated with foodborne illnesses (Guibourdenche et al, 2010; Jackson et al, 2013), the diversity within and among these serovars is considerable and can limit effective detection, isolation and recovery of Salmonella organisms from food. In food matrices contaminated with low numbers of Salmonella, recovery of organisms may be very difficult or unsuccessful and this could explain the discrepancy between reports of low prevalence of Salmonella in food surveys and high prevalence of clinical salmonellosis in humans. As part of the epidemiological investigation into clinical cases of foodborne illnesses, interviews are conducted on patients to identify food consumed in the days leading to the clinical episode. Apart from difficulties in remembering what was consumed, a second reason limiting the identification of contaminated food as a source of a foodborne illness is that a patient may not have been aware of all the constituents of a meal and may not be able to accurately or

Methods

Results

Conclusion