Listeria monocytogenes serotypes in human infections (Italy, 2000-2010)

BACKGROUND: In developed countries invasive listeriosis is an infection of great concern to public health to due its clinical severity and high fatality rate, despite its low incidence. In Europe, statistically significant increasing trends in listeriosis notification rates from 2005 to 2009 were noted in Austria, Denmark, Hungary, Italy, Spain and Sweden. MATERIALS AND METHODS: The standardized techniques based on phenotype to typing Listeria monocytogenes is the serotyping. In Europe, as elsewhere in the world, about 95% of L. monocytogenes strains isolated from clinical and food samples belongs to serovars 1/2a, 1/2b, 1/2c and 4b. RESULTS: The target of this work is to draw attention to this important and atypical foodborne disease, reporting epidemiological data and serotypes distribution of 251 human L. monocytogenes isolates reported during 2000-2010 to Veterinary Public Health and Food Safety Department of Istituto Superiore di Sanità, focusing on epidemiological trend of invasive listeriosis in Lombardia, a North Italian Region. The serotypes most frequently identified are 1/2a, 4b, 1/2b (in total 92%), but the detection of uncommon serotypes is not missing (1/2c, 3a, 3b, 4d). CONCLUSIONS: In Italy the surveillance laboratory network, as well as the foodborne disease network (ENTER-NET), has revealed in the last 11 years an increase trend of listeriosis cases reported likewise with results of Notificable National Infectious Disease surveillance System. This is probably due to a real increase of listeriosis, even if there is a greater sensitivity of the network in some regions

Detection and Virulence Characterization of Listeria monocytogenes Strains in Ready-to-Eat Products

The public health risk posed by Listeria monocytogenes in ready-to-eat (RTE) foods depends on the effectiveness of its control at every stage of the production process and the strain involved. Analytical methods currently in use are limited to the identification/quantification of L. monocytogenes at the species level, without distinguishing virulent from hypovirulent strains. In these products, according to EU Regulation 2073/2005, L. monocytogenes is a mandatory criterion irrespective of strain virulence level. Indeed, this species encompasses a diversity of strains with various pathogenic potential, reflecting genetic heterogeneity of the species itself. Thus, the detection of specific L. monocytogenes virulence genes can be considered an important target in laboratory food analysis to assign different risk levels to foods contaminated by strains carrying different genes. In 2015–2016, a severe invasive listeriosis outbreak occurred in central Italy, leading to the intensification of routine surveillance and strain characterization for virulence genetic markers. A new multiplex real-time polymerase chain reaction targeting main virulence genes has been developed and validated against the enzyme-linked fluorescent assay (ELFA) culture-based method. Results of the improved surveillance program are now reported in this study

European validation of a real-time PCR-based method for detection of Listeria monocytogenes in soft cheese

The classical microbiological method for detection of Listeria monocytogenes requires around 7days for final confirmation, and due to perishable nature of RTE food products, there is a clear need for an alternative methodology for detection of this pathogen. This study presents an international (at European level) ISO 16140-based validation trial of a non-proprietary real-time PCR-based methodology that can generate final results in the following day of the analysis. This methodology is based on an ISO compatible enrichment coupled to a bacterial DNA extraction and a consolidated real-time PCR assay. Twelve laboratories from six European countries participated in this trial, and soft cheese was selected as food model since it can represent a difficult matrix for the bacterial DNA extraction and real-time PCR amplification. The limit of detection observed was down to 10CFU per 25 of sample, showing excellent concordance and accordance values between samples and laboratories (>75%). In addition, excellent values were obtained for relative accuracy, specificity and sensitivity (82.75%, 96.70% and 97.62%, respectively) when the results obtained for the real-time PCR-based methods were compared to those of the ISO 11290-1 standard method. An interesting observation was that the L. monocytogenes detection by the real-time PCR method was less affected in the presence of Listeria innocua in the contaminated samples, proving therefore to be more reliable than the reference method. The results of this international trial demonstrate that the evaluated real-time PCR-based method represents an excellent alterative to the ISO standard since it shows a higher performance as well as reduce the extent of the analytical process, and can be easily implemented routinely by the competent authorities and food industry laboratories

European validation of Real-Time PCR method for detection of Salmonella spp. in pork meat

The classical microbiological method for detection of Salmonella spp. requires more than five days for final confirmation, and consequently there is a need for an alternative methodology for detection of this pathogen particularly in those food categories with a short shelf-life. This study presents an international (at European level) ISO 16140-based validation study of a non-proprietary Real-Time PCR-based method that can generate final results the day following sample analysis. It is based on an ISO compatible enrichment coupled to an easy and inexpensive DNA extraction and a consolidated Real-Time PCR assay. Thirteen laboratories from seven European Countries participated to this trial, and pork meat was selected as food model. The limit of detection observed was down to 10CFU per 25g of sample, showing excellent concordance and accordance values between samples and laboratories (100%). In addition, excellent values were obtained for relative accuracy, specificity and sensitivity (100%) when the results obtained for the Real-Time PCR-based methods were compared to those of the ISO 6579:2002 standard method. The results of this international trial demonstrate that the evaluated Real-Time PCR-based method represents an excellent alternative to the ISO standard. In fact, it shows an equal and solid performance as well as it reduces dramatically the extent of the analytical process, and can be easily implemented routinely by the Competent Authorities and Food Industry laboratories