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[Display omitted] • Detection of Yersinia enterocolitica in bivalve mollusks by a reference test (ISO 10273:2017). • Validation "in house" of the molecular method employed to detect pathogenic Yersinia enterocolitica in bivalve mollusks. • Detection of Yersinia enterocolitica in real samples of bivalve mollusks. Yersinia enterocolitica (Ye) is a foodborne pathogen isolated from humans, food, animals, and the environment. Yersiniosis is the third most frequently reported foodborne zoonosis in the European Union. Ye species are divided into six biotypes 1A, 1B, 2, 3, 4, and 5, based on biochemical reactions and about 70 serotypes. Biotype 1A is non-pathogenic, 1B is highly pathogenic, and biotypes 2–5 have moderate or low pathogenicity. The reference analysis method for detecting pathogenic Ye species underestimates the presence of the pathogen due to similarities between Yersinia enterocolitica -like species and other Yersiniaceae and/or Enterobacteriaceae , low concentrations of distribution pathogenic strains and the heterogeneity of Yersinia enterocolitica species. In this study, the real-time PCR method ISO/TS 18867 to identify pathogenic biovars of Ye in bivalve molluscs was validated. The sensitivity, specificity and accuracy of the molecular method were evaluated using molluscs experimentally contaminated. The results fully agree with those obtained with the ISO 10273 method. Finally, we evaluated the presence of Ye in seventy commercial samples of bivalve molluscs collected in the Gulf of Naples using ISO/TS 18867. Only one sample tested resulted positive for the ail gene, which is considered the target gene for detection of pathogenic Ye according to ISO/TS 18867. Additionally, the presence of the ystB gene, used as target for Ye biotype 1A, was assessed in all samples using a real-time PCR SYBR Green platform. The results showed amplification ystB gene aim two samples. [ABSTRACT FROM AUTHOR]

Verocytotoxin-producing Escherichia coli, also referred to as Shiga-toxin-producing Escherichia coli (STEC), can be transmitted to humans through person-to-person contact, consumption of contaminated food or water, or by direct contact with animals. Its clinical and economic consequences have prompted the development of alternative approaches to the official method of analysis "UNI CEN ISO/TS 13136: 2012", which describes the identification of STEC through the detection of its main virulence genes. Recently, droplet digital PCR (ddPCR) has been proposed as a technique for the sequence-specific detection and direct quantification of nucleic acids. The present study aimed to investigate if ddPCR could be able to detect STEC in less time than that required by the official method. This study consisted of the ddPCR of slices of beef contaminated with STEC and of the sponges used for beef official control at the slaughter stage. The results showed the ability of ddPCR to detect STEC in slices of beef already after sample incubation for 7 h at 37 °C while, in the case of sponges used for official controls, 9 h at 37 °C was needed. In this way, the ddPCR could represent an efficient method for detecting STEC and providing results in less time than the official method. [ABSTRACT FROM AUTHOR]