title: Rapid detection and specific differentiation of Salmonella enterica subsp. enterica Enteritidis, Typhimurium and its monophasic variant 4,[5],12:i:- by real-time multiplex PCR.
abstract: Salmonella enterica is one of the most common zoonotic pathogens worldwide causing clinical diseases in human and animal hosts. Targeting a reduction of Salmonella prevalence in poultry, the EU set up a microbiological criterion that demands the absence of S. enterica subsp. enterica serovars Enteritidis and Typhimurium including its monophasic variant with seroformula 4,[5],12:i:- in 25 g of poultry neck skin samples and fresh meat according to regulation (EU) no 1086/2011. We developed and in-house validated a method that detects and differentiates these Salmonella serovars based on a 5-plex real-time PCR assay within 24 h after sampling. The inclusivity and exclusivity were between 98 and 99% analysing 456 bacterial strains. Validation according to ISO 16140:2003 against the traditional cultural reference method ISO 6579:2002 was performed using 60 artificially contaminated and 31 presumably naturally contaminated chicken neck skin samples resulting in a relative accuracy of 100%. The detection probability reached 100% between 3 and 5 CFU/25 g sample. We were also able to assign rough and non-motile strains to S. enterica subsp. enterica serovars Enteritidis and Typhimurium. In conclusion, we provide diagnostic laboratories a fast and accurate method to monitor these Salmonella serovars in chicken neck skin samples. Other matrices could be easily adapted.
authors.affiliation: Federal Institute for Risk Assessment, Unit Molecular Microbiology and Genome Analysis, National Salmonella Reference Laboratory, Diedersdorfer Weg 1, D-12277 Berlin, Germany.
authors.forename: Sven
authors.initials: S
authors.lastname: Maurischat
authors.affiliation: Federal Institute for Risk Assessment, Unit Molecular Microbiology and Genome Analysis, National Salmonella Reference Laboratory, Diedersdorfer Weg 1, D-12277 Berlin, Germany.
authors.forename: Beatrice
authors.initials: B
authors.lastname: Baumann
authors.affiliation: Federal Institute for Risk Assessment, Unit Epidemiology, Statistics and Mathematical Modelling, Max-Dohrn-Str. 8-10, D-10589 Berlin, Germany.
authors.forename: Annett
authors.initials: A
authors.lastname: Martin
authors.affiliation: Federal Institute for Risk Assessment, Unit Molecular Microbiology and Genome Analysis, National Salmonella Reference Laboratory, Diedersdorfer Weg 1, D-12277 Berlin, Germany. Electronic address: burkhard.malorny@bfr.bund.de.
authors.forename: Burkhard
authors.initials: B
authors.lastname: Malorny
mesh_terms: D000818:Animals; D002645:Chickens; D057230:Limit of Detection; D008460:Meat / Q000382:microbiology; D060885:Multiplex Polymerase Chain Reaction; D060888:Real-Time Polymerase Chain Reaction; D015203:Reproducibility of Results; D012477:Salmonella enteritidis / Q000235:genetics / Q000302:isolation & purification; D012486:Salmonella typhimurium / Q000235:genetics / Q000302:isolation & purification
keywords: Detection; Enteritidis; Real-time PCR; Salmonella; Typhimurium; Validation