Whole genome sequencing for typing and characterisation of Listeria monocytogenes isolated in a rabbit meat processing plant

Listeria monocytogenes is a food-borne pathogen able to survive and grow in different environments including food processing plants where it can persist for month or years. In the present study the discriminatory power of Whole Genome Sequencing (WGS)-based analysis (cgMLST) was compared to that of molecular typing methods on 34 L. monocytogenes isolates collected over one year in the same rabbit meat processing plant and belonging to three genotypes (ST14, ST121, ST224). Each genotype included isolates indistinguishable by standard molecular typing methods. The virulence potential of all isolates was assessed by Multi Virulence-Locus Sequence Typing (MVLST) and the investigation of a representative database of virulence determinant genes. The whole genome of each isolate was sequenced on a MiSeq platform. The cgMLST, MVLST, and in silico identification of virulence genes were performed using publicly available tools. Draft genomes included a number of contigs ranging from 13 to 28 and N50 ranging from 456298 to 580604. The coverage ranged from 41 to 187X. The cgMLST showed a significantly superior discriminatory power only in comparison to ribotyping, nevertheless it allows the detection of two singletons belonging to ST14 that were not observed by other molecular methods. All ST14 isolates belonged to VT107, which 7-loci concatenated sequence differs for only 4 nucleotides to VT1 (Epidemic clone III). Analysis of virulence genes showed the presence of a fulllength inlA version in all ST14 isolates and of a mutated version including a premature stop codon (PMSC) associated to attenuated virulence in all ST121 isolates

Evaluation of Real-Time PCR to complement ISO 6579:2004 method for the detection of Salmonella in pork cuts

According to Commission Regulation (EC) No 2073/2005 of 15 November 2005 on microbiological crite-ria for foodstuff , the analytical reference method for the detection of Salmonella in food is ISO 6579:2004. However this long and labor-intensive method is not in line with the short production times of the food industry. In the last years, Real-Time PCR is used more and more by scientists for the relia-ble, fast and specific detection of bacterial pathogens in food. The aim of the present study was to eval-uate the Salmonella detection capability of a validated Real-Time PCR assay on naturally contaminated pork cuts in comparison with the reference method ISO 6579:2004. Three sampling were performed and included 16 pork cut packaging. From each packaging, three aliquots of 10 g each were tested separate-ly by ISO 6579:2004 method and by Real-Time PCR. In particular this molecular method was applied on DNA samples extracted from pre-enrichment broth after 1 and 18 hours of incubation. Within the three sampling periods, Real-Time PCR detected Salmonella in 81%, 100% e 62,5% of pork cut samples respectively, whereas the corresponding percentages of detection of the reference method were 56%, 81% e 62,5% respectively. In conclusion the Real-Time PCR assay used in the present study might be a reliable tool for a fast detection of Salmonella on pork cuts, especially when large number of samples needs to be tested. The reference method might be applied only on positive samples for isolation purpos-es mandatory in epidemiological investigations

New technologies to enhance quality and safety of table eggs: ultra-violet treatment and modified atmosphere packaging

In the present study the effect of ultra-violet (UV) treatment alone and in combination with 100% CO2 modified atmosphere packaging (MAP) was evaluated both on the survival of naturally occurring bacteria, as well as on quality parameters of table eggs during 28 days of storage at 21\ub0C. Table eggs were collected from the conveyor belt after the UV module, and placed on carton trays. A representative number of carton trays were packed in a high barrier multilayer pouch filled with 100% CO2. All eggs were stored at 21\ub0C and analysed at 0, 1, 7, 14, 21 and 28 days of storage. Eggs not treated with UV and not packed were also included. On the eggshells total colony count, total coliforms and faecal coliforms counts, as well as the detection of Salmonella spp. were investigated. Moreover, chemical-functional parameters such as weight loss, albumen pH and Haugh Unit (HU) were evaluated. The total colony count on UV treated table eggs was approximately 1 log10 CFU/g lower than untreated eggs (2.27 vs 3.29 log10 CFU/g). During storage, CO2 packed eggs maintained the initial values of HU, whereas the albumen pH decreased up to 1.5-2 points in comparison to unpacked eggs. The UV treatment was effective in reducing the total colony count on the surface of table eggs. MAP showed a great potential in maintaining/enhance the technological properties of egg constituents (higher foam stability of the albumen for meringue preparation) without significantly impacting on the microbial load of table eggs

Salmonella detection and aerobic colony count in deep-frozen carcasses of house sparrow (Passer domesticus) and starling (Sturnus vulgaris) intended for human consumption

Wild birds are potential vehicles of zoonotic pathogen transmission to humans. The zoonotic concern increases for small wild birds like house sparrows (Passer domesticus) and starlings (Sturnus vulgaris) which are hunted in developing countries and commercialised in Italy for human consumption. From June to October 2011, 330 house sparrows and 140 starlings were hunted and slaughtered. Deepfrozen carcasses were transported to Italy and stored for 6-8 months at -18\ub0C. Aerobic colony count and Salmonella detection in carcasses were assessed following standard microbiological methods (ISO 4833:2003 and ISO 6579:2004, respectively). Carcasses of house sparrows showed higher levels of aerobic bacteria in comparison to starling carcasses (5.7 vs 3.2 log10 CFU/g). Moreover, 7 out of 11 lots of carcasses of house sparrows were positive for Salmonella. Among the 18 isolates of Salmonella, 14 were S. Typhimurium, 2 were S. Enteritidis, and 2 were not distinguishable. All of them were susceptible to antibiotics. All tested carcasses of starling were Salmonella negative. Deep-freezing was not efficient as a decontamination technique on carcasses of house sparrows