Genetic diversity of Listeria monocytogenes strains contaminating food and food producing environment as single based sample in Italy (retrospective study).

Human listeriosis outbreaks are often associated with food products, which could be contaminated, at the same time, also by different clones of Listeria monocytogenes. This emphasize the need to type more than one L.monocytogenes isolate found in a single food or environmental sample. The purpose of the present study was to evaluate the presence of different L.monocytogenes strains in food and food production environment in order to understand if there is need to type more isolates from the same sample in case of presence of L.monocytogenes. Between 2011 and 2015, at the Italian National Reference Laboratory for L.monocytogenes, for each positive sample, from two to twenty-three isolates of L.monocytogenes were collected. All the isolates were characterized by conventional serotyping and pulsed field gel electrophoresis (PFGE). Moreover, isolates from the same sample, having indistinguishable PFGE profile, were subjected to whole genome sequencing in order to perform core genome Multi Locus Sequence Typing (cgMLST). Within each sample, more than one serotype and one pulsotype were found in 11.9% and 27.5%, respectively. For indistinguishable PFGE patterns the cgMLST analysis showed 96.2% of concordance demonstrating the added value of new sequencing technologies. This study has demonstrated the need to select and type more than one L.monocytogenes colony in one food or food environmental sample to detect the diversity of L.monocytogenes strains and facilitate downstream investigations and effective source attribution in foodborne outbreak inquiry.

Genetic relationships and biofilm formation of Listeria monocytogenes isolated from the smoked salmon industry.

Among pathogens, L. monocytogenes has the capability to persist on Food Processing Environment (FPE), first of all posing safety issues, then economic impact on productivity. The aim of this work was to determine the influence of biofilm forming-ability and molecular features on the persistence of 19 Listeria monocytogenes isolates obtained from FPE, raw and processed products of a cold-smoked salmon processing plant. To verify the phenotypic and genomic correlations among the isolates, different analyses were employed: serotyping, Clonal Complex (CC), core genome Multi-Locus Sequence Typing (cgMLST) and Single Nucleotide Polymorphisms (SNPs) clustering, and evaluation of the presence of virulence- and persistence-associated genes. From our results, the biofilm formation was significantly higher (*P < 0.05) at 37 °C, compared to 30 and 12 °C, suggesting a temperature-dependent behaviour. Moreover, the biofilm-forming ability showed a strain-specific trend, not correlated with CC or with strains persistence. Instead, the presence of internalin (inL), Stress Survival Islet (SSI) and resistance to erythromycin (ermC) genes was correlated with the ability to produce biofilms. Our data demonstrate that the genetic profile influences the adhesion capacity and persistence of L. monocytogenes in food processing plants and could be the result of environmental adaptation in response to the external selective pressure.

A Real-Time PCR Screening Assay for Rapid Detection of Listeria Monocytogenes Outbreak Strains.

From January 2015 to March 2016, an outbreak of 23 human cases of listeriosis in the Marche region and one human case in the Umbria region of Italy was caused by Listeria monocytogenes strains showing a new pulsotype never described before in Italy. A total of 37 clinical strains isolated from patients exhibiting listeriosis symptoms and 1374 strains correlated to the outbreak were received by the Italian National Reference Laboratory for L. monocytogenes (It NRL Lm) of Istituto Zooprofilattico Sperimentale dell'Abruzzo e del Molise (IZSAM) for outbreak investigation. A real-time PCR assay was purposely designed for a rapid screening of the strains related to the outbreak. PCR-positive strains were successively typed through molecular serogrouping, pulsed field gel electrophoresis (PFGE), and Next Generation Sequencing (NGS). Applying the described strategy, based on real-time PCR screening, we were able to considerably reduce time and costs during the outbreak investigation activities.

A severe outbreak of listeriosis in central Italy with a rare pulsotype associated with processed pork products.

PURPOSE: From May 2015 to March 2016, an outbreak due to Listeria monocytogenes serotype 1/2a and clinical pulsotype never previously isolated in Europe occurred in central Italy, involving 24 confirmed clinical cases. The article provides a description of the outbreak and the investigation carried out by a multidisciplinary network. METHODOLOGY: Epidemiological and microbiological surveillance was conducted to confirm the outbreak and to detect the food vehicle of infection. The origin and destination of the implicated food and its ingredients were investigated by tracing-back and -forward investigation. RESULTS: Next-generation sequencing confirmed the unique outbreak strain. On 4 January 2016, a L. monocytogenes strain with pulsotype indistinguishable from that isolated from clinical cases in the outbreak was detected in a sample of hog head cheese purchased from a retail supermarket by one of the patients. The hog head cheese was produced by a small meat processing plant in the Marche region, where microbiological investigation confirmed environmental and food contamination by the outbreak strain. Plant production was suspended and all contaminated batches of the hog head cheese were withdrawn from the market by 19 February by local health authority. We subsequently observed a sharp decline in clinical cases, the last being reported on 11 March 2016. CONCLUSION: The key factor in the timely conclusion of this investigation was intersectoral collaboration among epidemiologists, microbiologists, veterinarians, statisticians and health and food safety authorities at national, regional and local levels.

Whole-Genome Sequences of Two Listeria monocytogenes Serovar 1/2a Strains Responsible for a Severe Listeriosis Outbreak in Central Italy.

We report the whole-genome sequences of two Listeria monocytogenes strains responsible for a severe invasive listeriosis outbreak in central Italy that occurred in 2015 and 2016. These two strains differ by a single band in their pulsed-field gel electrophoresis (PFGE) profiles.