The aim of this study was to identify and characterise Bacillus cereus from a unique national collection of 564 strains associated with 140 strong-evidence food-borne outbreaks (FBOs) occurring in France during 2007 to 2014. Starchy food and vegetables were the most frequent food vehicles identified; 747 of 911 human cases occurred in institutional catering contexts. Incubation period was significantly shorter for emetic strains compared with diarrhoeal strains A sub-panel of 149 strains strictly associated to 74 FBOs and selected on Coliphage M13-PCR pattern, was studied for detection of the genes encoding cereulide, diarrhoeic toxins (Nhe, Hbl, CytK1 and CytK2) and haemolysin (HlyII), as well as panC phylogenetic classification. This clustered the strains into 12 genetic signatures (GSs) highlighting the virulence potential of each strain. GS1 (nhe genes only) and GS2 (nhe, hbl and cytK2), were the most prevalent GS and may have a large impact on human health as they were present in 28% and 31% of FBOs, respectively. Our study provides a convenient molecular scheme for characterisation of B. cereus strains responsible for FBOs in order to improve the monitoring and investigation of B. cereus-induced FBOs, assess emerging clusters and diversity of strains.

Background: Listeria monocytogenesClonal Complexes (CCs) have been epidemiologically associated with foods, especially ready-to-eat (RTE) products for which the most likely source of contamination depends on the occurrence of persisting clones in food-processing environments (FPEs). As the ability ofL. monocytogenesto adapt to environmental stressors met in the food chain challenges the efforts to its eradication from FPEs, the threat of persistent strains to the food industry and public health authorities continues to rise. In this study, 94 food and FPEsL. monocytogenesisolates, representing persistent subtypes contaminating three French seafood facilities over 2–6 years, were whole-genome sequenced to characterize their genetic diversity and determine the biomarkers associated with long-term survival in FPEs.Results: Food and FPEs isolates belonged to five CCs, comprising long-term intra- and inter-plant persisting clones. Mobile genetic elements (MGEs) such as plasmids, prophages and transposons were highly conserved within CCs, some of which harboured genes for resistance to chemical compounds and biocides used in the processing plants. Some of these genes were found in a 90.8 kbp plasmid, predicted to be” mobilizable”, identical in isolates from CC204 and CC155, and highly similar to an 81.6 kbp plasmid from isolates belonging to CC7. These similarities suggest horizontal transfer between isolates, accompanied by deletion and homologous recombination in isolates from CC7. Prophage profiles characterized persistent clonal strains and several prophage-loci were plant-associated. Notably, a persistent clone from CC101 harboured a novel 31.5 kbp genomic island that we namedListeriagenomic island 3 (LGI3), composed by plant-associated loci and chromosomally integrating cadmium-resistance determinantscadA1C.Conclusions: Genome-wide analysis indicated that inter- and intra-plant persisting clones harbour conserved MGEs, likely acquired in FPEs and maintained by selective pressures. The presence of closely related plasmids inL. monocytogenesCCs supports the hypothesis of horizontal gene transfer conferring enhanced survival to FPE-associated stressors, especially in hard-to-clean harbourage sites. Investigating the MGEs evolutionary and transmission dynamics provides additional resolution to trace-back potentially persistent clones. The biomarkers herein discovered provide new tools for better designing effective strategies for the removal or reduction of residentL. monocytogenesin FPEs to prevent contamination of RTE seafood.

Variant strains of Salmonella enterica serovar Typhimurium, lacking one or both flagellar phases have been widely reported. The monophasic S.1,4,[5],12:i:‐ variant has emerged worldwide in the past few years and has become one of the most frequently encountered in many countries. In contrast, monophasic S.1,4,[5],12:‐:1,2 and nonmotile S.1,4,[5],12:‐:‐ strains are rarely described. This study investigated seven molecular markers to identify and delineate monophasic S.1,4,[5],12:i:‐ (n = 90), S.1,4,[5],12:‐:1,2 (n = 25), nonmotile S.1,4,[5],12:‐:‐ (n = 17) strains, and some serovar Typhimurium strains (n = 124) collected through the French Salmonella network between 2001 and 2010. Three markers were commonly detected in serovar Typhimurium and in all variant strains: STM2757, mdh and fliA‐B. Monophasic S.1,4,[5],12:i:‐ were genotypically confirmed by the absence of the fljB, fljA, and hin genes. Nevertheless, 13 (14.5%) of them were positive for these last three genes, revealing monophasic strains named “inconsistent” as previously described. All nonmotile 1,4,[5],12:‐:‐ strains had the fliC, fljA, fljB, and hin genes and the fliC gene was detected in 88% of monophasic S.1,4,[5],12:‐:1,2 strains. The combination of the seven markers detection enables to recognize eight different genotypes within the S.1,4,[5],12:i:‐ collection, among which the Spanish and the U.S. clones previously described could be distinguished and assigned to a genotype. Based on this molecular approach, 71% of the French S.1,4,[5],12:i:‐ collection belonged to the Spanish clone, whereas only 2% were assigned to the U.S. clone. This study highlights the usefulness of these molecular markers and genotypes for identifying lineages, especially among the epidemiologically important monophasic S.1,4,[5],12:i:‐ variant.

Background: Typhimurium is the main serotype of Salmonella enterica subsp. enterica implicated in food-borne diseases worldwide. This study aimed to detect the prevalence of ten markers combined in a macro-array based on multiplex real-time PCR. We targeted characteristic determinants located on pathogenicity islands (SPI-2 to -5, virulence plasmid pSLT and Salmonella genomic island 1 (SGI1)) as well as a specific 16S-23S rRNA intergenic spacer sequence of definitive type 104 (DT104). To investigate antimicrobial resistance, the study also targeted the presence of genes involved in sulfonamide (sul1) and beta-lactam (bla _TEM) resistance. Finally, the intI1 determinant encoding integrase from class 1 integron was also investigated. Results: A total of 538 unrelated S. Typhimurium strains isolated between 1999 and 2009 from various sources, including food animals, food products, human and environmental samples were studied. Based on the combined presence or absence of these markers, we distinguished 34 different genotypes, including three major genotypes encountered in 75% of the studied strains, Although SPI determinants were almost always detected, SGI1, intI1, sul1 and bla _TEM determinants were found 47%, 52%, 54% and 12% of the time respectively, varying according to isolation source. Low-marker patterns were most often detected in poultry sources whereas full-marker patterns were observed in pig, cattle and human sources. Conclusion: The GeneDisc^® assay developed in this study madeit easier to explore variability within serotype Typhimurium by analyzing ten relevant gene determinants in a large collection of strains. This real-time multiplex method constitutes a valuable tool for strains characterization on epidemiological purposes.

BACKGROUND: Fish is a highly perishable food, so it is important to be able to estimate its freshness to ensure optimum quality for consumers. The present study describes the development of an SPME‐GC‐MS technique capable of quantifying both trimethylamine (TMA) and dimethylamine (DMA), components of what has been defined as partial volatile basic nitrogen (PVB‐N). This method was used, together with other reference methods, to monitor the storage of cod fillets (Gadus morhua) conserved under melting ice.RESULTS: Careful optimisation enabled definition of the best parameters for extracting and separating targeted amines and an internal standard. The study of cod spoilage by sensory analysis and TVB‐N assay led to the conclusion that the shelf‐life of cod fillet was between 6 and 7 days. Throughout the study, TMA and DMA were specifically quantified by SPME‐GC‐MS; the first was found to be highly correlated with the values returned by steam distillation assays. Neither TMA‐N nor DMA‐N were able to successfully characterise the decrease in early freshness, unlike dimethylamine/trimethylamine ratio (DTR), whose evolution is closely related to the results of sensory analysis until the stage where fillets need to be rejected.CONCLUSION: DTR was proposed as a reliable indicator for the early decrease of freshness until fish rejection. © 2015 Society of Chemical Industry

Listeriosis is a severe infection which mainly affects pregnant women, neonates and immuno-compromised adults. ANSES’s Laboratory for Food safety has been the European Union Reference Laboratory (EURL) for L. monocytogenes in the food chain since 2006. Pulsed Field Gel Electrophoresis (PFGE) is routinely used in the EURL for the surveillance of L. monocytogenes isolated from foods, animals and the environment. One of the main EURL activities is to evaluate alternative molecular subtyping methods to PFGE, and integrate their use within the National Reference Laboratories (NRL) network. Since 2008, the United Kingdom (UK)-NRL for L. monocytogenes at the Health Protection Agency (HPA), London, has used fluorescent Amplified Fragment Length Polymorphism (fAFLP) for the routine surveillance of L. monocytogenes isolated from human clinical cases, food and food processing environments in the UK. This study compares fAFLP with PFGE for subtyping L. monocytogenes.