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The study assessed the role of equids at slaughter as faecal carriers of Salmonella enterica and the occurrence of contaminated equid carcasses during the slaughter process in Northern Italy (Emilia-Romagna Region). From June to November 2021, 152 equids (146 horses, 5 donkeys and 1 mule) were tested for Salmonella both in caecal contents and through carcass swabs. Antimicrobial resistance (AMR) of recovered strains was tested against 15 antimicrobials. Salmonella was detected in 3/152 of the caecal contents (2.0 %), while all carcass samples were negative. S. enterica serovars Enteriditis, Typhimurium and Stanleyville were identified. The only AMR isolate was S. Typhimurium with AMR profile AmCStxT. Considering the consumption of raw horse meat (i.e., minced raw meat named "pesto di cavallo" and dried and smoked strips named "sfilacci di cavallo") in different areas of Northern Italy, we also investigated the possible link between horse meat eating and salmonellosis cases in the human population in the same area. Specifically, we compared the Salmonella strains collected during the study with those routinely processed in the laboratory surveillance system for human salmonellosis in Emilia-Romagna (a region with about 4.5 million inhabitants). The comparison was based on whole genome sequencing data through core genome multi-locus sequence typing (cgMLST) used in routine surveillance. A genomic match in cgMLST was found between the strain of S. enterica serovar Enteritidis isolated from a horse caecal content and an enduring outbreak of 17 human cases in Emilia-Romagna during the study period. The consequent epidemiological investigation highlighted that a number of cases with known food history reported the consumption of horse meat and traced different batches of the consumed meat, released weeks apart from each other, to the slaughter investigated in the study. The results of the epidemiological investigation suggested the role of horses in the S. enterica serovar Enteritidis outbreak affecting raw horse meat consumers. This study shows that, despite the low prevalence on equid carcasses, S. enterica in horse meat can represent a risk to consumers. From the perspective of the slaughter activities, this highlights the need to maintain a high level of hygiene during the entire process, starting from the hygiene at lairage up to the slaughtering phase and dressing of carcasses., Competing Interests: Declaration of competing interest None., (Copyright © 2023 The Authors. Published by Elsevier B.V. All rights reserved.)

We report Listeria monocytogenes infection in a patient in Italy who was transfused with pooled platelet concentrate. Genomic analysis revealed that L. monocytogenes isolates from the donor blood unit, the transfused platelets, and the patient's blood culture were genetically closely related, confirming transfusion transmission. Additional surveillance and secondary bacterial screening could improve transfusion safety.

This study aims to describe trends of mcr-positive Enterobacterales in humans based on laboratory surveillance with a defined catchment population. The data source is the Micro-RER surveillance system, established in Emilia-Romagna region (Italy), to monitor the trend of mcr resistance. Enterobacterales isolates from human clinical samples with minimum inhibitory concentration (MIC) ≥ 2 mg/L for colistin were sent to the study reference laboratory for the detection of mcr genes. Isolates prospectively collected in the period 2018-2020 were considered for the assessment of population rates and trends; further analyses were carried out for the evaluation of clonality and horizontal mcr gene transfer. Previous isolates from local laboratory collection were also described. In the period 2018-2020, 1164 isolates were sent to the reference laboratory, and 51 (4.4%) were confirmed as mcr-positive: 50 mcr-1 (42 Escherichia coli, 6 Klebsiella pneumoniae, 2 Salmonella enterica) and 1 mcr-4 (Enterobacter cloacae). The number of mcr-positive isolates dropped from 24 in the first half of 2018 to 3 in the whole of 2020 (trend p value < 0.001). Genomic analyses showed the predominant role of the horizontal transfer of mcr genes through plasmids or dissemination of transposable elements compared to clonal dissemination of mcr-positive microorganisms. The study results demonstrate a substantial decrease in the circulation of mcr-1 plasmid genes in Emilia-Romagna Region., (© 2021. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The population structure of human isolates of Listeria monocytogenes in Emilia-Romagna, Italy, from 2012 to 2018 was investigated with the aim of evaluating the presence of genomic clusters indicative of possible outbreaks, the proportion of cluster-associated vs. sporadic isolates and different methods and metrics of genomic analysis for use in routine surveillance. In the 2012-2018 period the notification rate of confirmed invasive cases in Emilia-Romagna was 0.91 per 100,000 population per year, more than twice the average rate of EU countries. Out of the total 283 cases, 268 (about 95%) isolates were typed through whole genome sequencing (WGS) for cluster detection with methods based on core-genome multi-locus sequence typing and single nucleotide polymorphisms. Between 66 and 72% of listeriosis cases belonged to genomic clusters which included up to 27 cases and lasted up to 5 years. This proportion of cluster-associated cases is higher than previously estimated in other European studies. Rarefaction analysis, performed by reducing both the number of consecutive years of surveillance considered and the proportion of isolates included in the analysis, suggested that the observed high proportion of cluster-associated cases can be ascribed to the long surveillance duration (7 years) and the high notification and typing rates of this study. Our findings show that a long temporal perspective and high surveillance intensity, intended as both exhaustiveness of the system to report cases and high WGS-typing rate, are critical for sensitive detection of possible outbreaks within a WGS-based surveillance of listeriosis. Furthermore, the power and complexity of WGS interpretation emerged from the integration of genomic and epidemiological information in the investigation of few past outbreaks within the study, indicating that the use of multiple approaches, including the analysis of the accessory genome, is needed to accurately elucidate the population dynamics of Listeria monocytogenes ., (Copyright © 2020 Scaltriti, Bolzoni, Vocale, Morganti, Menozzi, Re and Pongolini.)

The study assessed Salmonella carriage in wild boars (Sus scrofa) and compared their isolates with those recovered from the domestic swine population of the same area of northern Italy (Emilia-Romagna), characterized by intensive pig farming and rather high density of wild boars. A total of 189 wild boars hunted during twelve months (2017-2018) were tested for Salmonella in mesenteric lymph nodes (MLN) and faecal samples. Antimicrobial resistance of recovered strains was tested against 14 antimicrobials. Salmonella was detected in 33/189 wild boars (17.5%), specifically from 30/189 MLN (15.9%) and 6/189 faecal samples (3.2%). Three animals were positive in both samples. Thirteen Salmonella serovars were identified, i.e. Typhimurium (the most common), Bovismorbificans, Coeln, Derby, Enteritidis, Gaminara, Hessarek, Houtenae IV, Kottbus, Napoli, Stanleyville, Thompson and Veneziana. Salmonella carriage was higher in warm than in cold months (P = 0.0013). Pregnancy status was never associated with Salmonella carriage, with significant difference in the recovery of the pathogen between non-pregnant and pregnant females (P = 0.003). Only one resistance pattern to streptomycin and tetracycline was found in 15 isolates (41.7%) belonging to Typhimurium (14/14; 100%) and Kottbus (1/3; 33.3%) serovars. Overlap with isolates from farmed pigs was limited at serotype level (Typhimurium, Derby, Enteritis, Bovismorbificans, Kottbus) and absent at PFGE level, and also antimicrobial resistance patterns were substantially different. This evidence indicates a substantial segregation of the two animal populations with regard to infectious contacts, possibly suggesting that biosecurity measures in place at industrial farm level limit the exchange of Salmonella.

Salmonella enterica serovar Napoli (serovar Napoli) is an emerging cause of human salmonellosis in Northern Italy. No specific reservoirs of serovar Napoli have been identified in Italy, so far. However, the environment, especially surface waters, has been hypothesized as an important source of infection based on the observation that genotypically different clusters of serovar Napoli are detected in different geographical macro-areas. To further support the hypothesis of a spatially-restricted pattern of serovar Napoli diffusion, a spatial segregation of serovar Napoli lineages should be observed also at smaller geographical scale. However, classical genotyping techniques used for Salmonella, such as pulsed-field gel electrophoresis (PFGE), did not possess enough discriminatory power to highlight spatial clustering of serovar Napoli within the macro-areas. To this purpose, we performed phylogeographical analyses based on genome-wide single nucleotide polymorphisms to test whether spatio-temporal evolution patterns of serovar Napoli in Northern Italy could be recognized with high geographical resolution, i.e. at local level. Specifically, we analyzed the local spread of the main PFGE clonal group, responsible for more than 60% of human infections in the study area, that did not show any geographical differentiation by PFGE within Northern Italy, i.e. the macro-area considered in the study. Both discrete and continuous phylogeography highlighted the existence of two main geographically-restricted clades: a Southern clade corresponding to the Po Valley and a Northern clade corresponding to the Pre-Alps area. Furthermore, the phylogeographical analyses suggested that the most probable site of origin of the clone was in an area of the Po Valley at the confluence of the Po and Ticino rivers, one of the most important Italian wetlands. These findings provide further support to the hypothesis that environmental transmission may play an important role in the ecology of serovar Napoli., Competing Interests: The authors have declared that no competing interests exist.

During 2014-2015, 300 pig carcasses before chilling and 85 food contact surfaces (FCSs) at cutting lines were tested for Salmonella in three slaughterhouses (namely A, B, and C) of northern Italy. In slaughterhouses A and B, four carcass sites of 100 cm 2 each (from both the exterior and interior side) were swabbed with a single sponge. In abattoir C, four 100 cm 2 sites of the exterior and the interior sides were swabbed with two independent sponges. The population average prevalence of Salmonella-positive carcasses (which takes into account the structure of the study design, with multiple samples collected in a single day) in slaughterhouses A and B was 12.3%, while in slaughterhouse C it was 11.2%. Presence of Salmonella on exterior and interior sides of carcasses showed a low level of concordance (only 3/12 of the contaminated carcasses were positive on both sides). No significant difference was found for FCSs contamination in the three slaughterhouses, with a population average prevalence of Salmonella-positive FCSs of 19.9%. In addition, we found that the clustering due to the day of sampling account for more than 36% and 60% of the overall prevalence variation on carcasses and FCSs, respectively. Eight serovars were identified, with Salmonella Derby as the most common type. The counting of Salmonella on carcasses showed large variability. It was low (<0.0075 most probable number [MPN]/cm 2 ) in 46.6% of the carcasses and as high as 2.7 MPN/cm 2 in 4.7%. Specifically, we found that counts on carcasses fit with "heavy tailed" distributions (lognormal and Weibull with a small shape parameter), suggesting not negligible probability of episodes of high Salmonella contamination. The mean values of contamination obtained with the two distributions ranged from 0.235 to 0.435 MPN/cm 2 .

Background and aimEpidemiology of human non-typhoid salmonellosis is characterised by recurrent emergence of new clones of the pathogen over time. Some clonal lines of Salmonella have shaped epidemiology of the disease at global level, as happened for serotype Enteritidis or, more recently, for Salmonella 4,[5],12:i:-, a monophasic variant of serotype Typhimurium. The same clonal behaviour is recognisable at sub-serotype level where single outbreaks or more generalised epidemics are attributable to defined clones. The aim of this study was to understand the dynamics of a clone of Salmonella 4,[5],12:i:- over a 3-year period (2012-15) in a province of Northern Italy where the clone caused a large outbreak in 2013. Furthermore, the role of candidate outbreak sources was investigated and the accuracy of multilocus variable-number tandem repeat analysis (MLVA) was evaluated. Methods: we retrospectively investigated the outbreak through whole genome sequencing (WGS) and further monitored the outbreak clone for 2 years after its conclusion. Results: The study showed the transient nature of the clone in the population, possibly as a consequence of its occasional expansion in a food-processing facility. We demonstrated that important weaknesses characterise conventional typing methods applied to clonal pathogens such as Salmonella 4,[5],12:i:-, namely lack of accuracy for MLVA and inadequate resolution power for PFGE to be reliably used for clone tracking. Conclusions : The study provided evidence for the remarkable prevention potential of whole genome sequencing used as a routine tool in systems that integrate human, food and animal surveillance.

In this study we report the detection of the recently described mcr-4 gene in two human isolates of Salmonella enterica serovar Typhimurium. The strains were isolated from faecal samples of two Italian patients with gastroenteritis, collected in 2016. The identified mcr-4 genes (variant mcr-4.2 ) differed from the mcr-4 gene originally described in a Salmonella strain of swine origin from Italy. Salmonella species could represent a hidden reservoir for mcr genes.

Here we report the complete nucleotide sequence of a 49.257-bp IncL/M conjugative plasmid (pRAY) carrying the blaOXA-48 gene collected from a Klebsiella pneumoniae clinical strain isolated in Italy. The genetic environment of pRAY plasmid revealed that the blaOXA-48 gene was located within a Tn1999.2 transposon. The pRAY plasmid differed from blaOXA-48-harboring IncL/M plasmids by genetic context and size. Comparative analysis demonstrated that pRAY plasmid lacked a region of ~15 kb carrying genes encoding proteins involved in pilus assembly and plasmid conjugative apparatus.

Usutu virus is an arbovirus closely related to West Nile virus (genus Flavivirus), which circulates between mosquitoes and wild birds. This virus has been increasingly reported in Europe, raising concerns for its possible pathogenic potential for wild birds and humans. This study reports the whole genome sequences of 15 strains of USUV, isolated between 2010 and 2014 from mosquitoes and wild birds in the course of West Nile virus surveillance in the Emilia-Romagna and Lombardy regions of Northern Italy. Both whole and 656 partial genome sequences, obtained from isolated viruses and field samples (mosquitoes and wild birds), were analyzed to describe the temporal and geographical spread of USUV in the surveyed area. The detected sequences belonged to two groups, with one circulating primarily in the northwestern part of the area and the other in the southeastern part. This pattern is likely the result of different routes of introduction from the North (over the Alps) and from the East, respectively. The phylogenetic analysis of obtained sequences and other European sequences demonstrated that the majority of European strains belonged to one main clade, while less common strains, mainly from Western Europe, fell in other two clades. This analysis strongly suggested an autochthonous evolution process of strains of the main clade from a common ancestor with an estimated time of arrival in Europe at the beginning of the 1990s. In addition to causing mass mortality in wild birds, Usutu virus can infect humans and can sporadically cause disease. These factors and the endemization of the Usutu virus in a large area of Europe, sustained by the obtained data, strongly support the need to adequately survey Usutu virus in areas in which its circulation is detected., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Infectious diseases in livestock can be transmitted through fomites: objects able to convey infectious agents. Between-farm spread of infections through fomites is mostly due to indirect contacts generated by on-farm visits of personnel that can carry pathogens on their clothes, equipment, or vehicles. However, data on farm visitors are often difficult to obtain because of the heterogeneity of their nature and privacy issues. Thus, models simulating disease spread between farms usually rely on strong assumptions about the contribution of indirect contacts on infection spread. By using data on veterinarian on-farm visits in a dairy farm system, we built a simple simulation model to assess the role of indirect contacts on epidemic dynamics compared to cattle movements (i.e. direct contacts). We showed that including in the simulation model only specific subsets of the information available on indirect contacts could lead to outputs widely different from those obtained with the full-information model. Then, we provided a simple preferential attachment algorithm based on the probability to observe consecutive on-farm visits from the same operator that allows overcoming the information gaps. Our results suggest the importance of detailed data and a deeper understanding of visit dynamics for the prevention and control of livestock diseases.

Multidrug-resistant (MDR) Klebsiella pneumoniae is one of the most important causes of nosocomial infections worldwide. After the spread of strains resistant to beta-lactams at the end of the previous century, the diffusion of isolates resistant to carbapenems and colistin is now reducing treatment options and the containment of infections. Carbapenem-resistant K. pneumoniae strains have spread rapidly among Italian hospitals, with four subclades of pandemic clonal group 258 (CG258). Here we show that a single Italian hospital has been invaded by three of these subclades within 27 months, thus replicating on a small scale the "Italian scenario." We identified a single clone responsible for an epidemic outbreak involving seven patients, and we reconstructed its star-like pattern of diffusion within the intensive care unit. This epidemiological picture was obtained through phylogenomic analysis of 16 carbapenem-resistant K. pneumoniae isolates collected in the hospital during a 27-month period, which were added to a database of 319 genomes representing the available global diversity of K. pneumoniae strains. Phenotypic and molecular assays did not reveal virulence or resistance determinants specific for the outbreak isolates. Other factors, rather than selective advantages, might have caused the outbreak. Finally, analyses allowed us to identify a major subclade of CG258 composed of strains bearing the yersiniabactin virulence factor. Our work demonstrates how the use of combined phenotypic, molecular, and whole-genome sequencing techniques can help to identify quickly and to characterize accurately the spread of MDR pathogens., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Assessing the performance of a surveillance system for infectious diseases of domestic animals is a challenging task for health authorities. Therefore, it is important to assess what strategy is the most effective in identifying the onset of an epidemic and in minimizing the number of infected farms. The aim of the present work was to evaluate the performance of the bovine tuberculosis (bTB) surveillance system in the network of dairy farms in the Emilia-Romagna (ER) Region, Italy. A bTB-free Region since 2007, ER implements an integrated surveillance strategy based on three components, namely routine on-farm tuberculin skin-testing performed every 3 years, tuberculin skin-testing of cattle exchanged between farms, and post-mortem inspection at slaughterhouses. We assessed the effectiveness of surveillance by means of a stochastic network model of both within-farm and between-farm bTB dynamics calibrated on data available for ER dairy farms. Epidemic dynamics were simulated for five scenarios: the current ER surveillance system, a no surveillance scenario that we used as the benchmark to characterize epidemic dynamics, three additional scenarios in which one of the surveillance components was removed at a time so as to outline its significance in detecting the infection. For each scenario we ran Monte Carlo simulations of bTB epidemics following the random introduction of an infected individual in the network. System performances were assessed through the comparative analysis of a number of statistics, including the time required for epidemic detection and the total number of infected farms during the epidemic. Our analysis showed that slaughterhouse inspection is the most effective surveillance component in reducing the time for disease detection, while routine surveillance in reducing the number of multi-farms epidemics. On the other hand, testing exchanged cattle improved the performance of the surveillance system only marginally., (Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.)

Acute rheumatic fever (ARF) is a postsuppurative sequela caused by Streptococcus pyogenes infections affecting school-age children. We describe here the occurrence of an ARF outbreak that occurred in Bologna province, northeastern Italy, between November 2012 and May 2013. Molecular analysis revealed that ARF-related group A Streptococcus (GAS) strains belonged to the M-18 serotype, including subtypes emm18.29 and emm18.32. All M-18 GAS strains shared the same antigenic profile, including SpeA, SpeB, SpeC, SpeL, SpeM, and SmeZ. Matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) analysis revealed that M-18 GAS strains grouped separately from other serotypes, suggesting a different S. pyogenes lineage. Single nucleotide polymorphisms and phylogenetic analysis based on whole-genome sequencing showed that emm18.29 and emm18.32 GAS strains clustered in two distinct groups, highlighting genetic variations between these subtypes. Comparative analysis revealed a similar genome architecture between emm18.29 and emm18.32 strains that differed from noninvasive emm18.0 strains. The major sources of differences between M-18 genomes were attributable to the prophage elements. Prophage regions contained several virulence factors that could have contributed to the pathogenic potential of emm18.29 and emm18.32 strains. Notably, phage ΦSPBO.1 carried erythrogenic toxin A gene (speA1) in six ARF-related M-18 GAS strains but not in emm18.0 strains. In addition, a phage-encoded hyaluronidase gene (hylP.2) presented different variants among M-18 GAS strains by showing internal deletions located in the α-helical and TSβH regions. In conclusion, our study yielded insights into the genome structure of M-18 GAS strains responsible for the ARF outbreak in Italy, thus expanding our knowledge of this serotype., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

We retrospectively analyzed a rare Salmonella enterica serovar Manhattan outbreak that occurred in Italy in 2009 to evaluate the potential of new genomic tools based on differential single nucleotide polymorphism (SNP) analysis in comparison with the gold standard genotyping method, pulsed-field gel electrophoresis. A total of 39 isolates were analyzed from patients (n=15) and food, feed, animal, and environmental sources (n=24), resulting in five different pulsed-field gel electrophoresis (PFGE) profiles. Isolates epidemiologically related to the outbreak clustered within the same pulsotype, SXB&#95;BS.0003, without any further differentiation. Thirty-three isolates were considered for genomic analysis based on different sets of SNPs, core, synonymous, nonsynonymous, as well as SNPs in different codon positions, by Bayesian and maximum likelihood algorithms. Trees generated from core and nonsynonymous SNPs, as well as SNPs at the second and first plus second codon positions detailed four distinct groups of isolates within the outbreak pulsotype, discriminating outbreak-related isolates of human and food origins. Conversely, the trees derived from synonymous and third-codon-position SNPs clustered food and human isolates together, indicating that all outbreak-related isolates constituted a single clone, which was in line with the epidemiological evidence. Further experiments are in place to extend this approach within our regional enteropathogen surveillance system., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Klebsiella pneumoniae is at the forefront of antimicrobial resistance for Gram-negative pathogenic bacteria, as strains resistant to third-generation cephalosporins and carbapenems are widely reported. The worldwide diffusion of these strains is of great concern due to the high morbidity and mortality often associated with K. pneumoniae infections in nosocomial environments. We sequenced the genomes of 89 K. pneumoniae strains isolated in six Italian hospitals. Strains were selected based on antibiotypes, regardless of multilocus sequence type, to obtain a picture of the epidemiology of K. pneumoniae in Italy. Thirty-one strains were carbapenem-resistant K. pneumoniae carbapenemase producers, 29 were resistant to third-generation cephalosporins, and 29 were susceptible to the aforementioned antibiotics. The genomes were compared to all of the sequences available in the databases, obtaining a data set of 319 genomes spanning the known diversity of K. pneumoniae worldwide. Bioinformatic analyses of this global data set allowed us to construct a whole-species phylogeny, to detect patterns of antibiotic resistance distribution, and to date the differentiation between specific clades of interest. Finally, we detected an ∼ 1.3-Mb recombination that characterizes all of the isolates of clonal complex 258, the most widespread carbapenem-resistant group of K. pneumoniae. The evolution of this complex was modeled, dating the newly detected and the previously reported recombination events. The present study contributes to the understanding of K. pneumoniae evolution, providing novel insights into its global genomic characteristics and drawing a dated epidemiological scenario for this pathogen in Italy., (Copyright © 2015, American Society for Microbiology. All Rights Reserved.)

Hepatitis A virus (HAV) was detected in two samples of mixed frozen berries linked to Italian hepatitis A outbreak in April and September 2013. Both viruses were fully sequenced by next-generation sequencing and the genomes clustered with HAV complete genomes of sub-genotype IA with nucleotide identities of 95-97%.

Tonsils from 150 pigs slaughtered at 270 days or older were tested for Yersinia enterocolitica with different cultural methods. Samples were collected in three different abattoirs of Northern Italy between April and November 2012 and were analysed by direct plating on cefsulodin-irgasan-novobiocin (CIN) agar and by enrichment procedures following the ISO 10273:2003 reference method. Twenty-three (15.3%) samples were positive: 22 tonsils (14.7%) were positive for human pathogenic Y. enterocolitica bio-serotype 4/O:3 and one tonsil (0.7%) for Y. enterocolitica bio-serotype 1A/7,8-8,8,19. Seventeen samples out of 23 (73.9%) were positive by direct plating method. Among the enrichment procedures, the best recovery rate (8 positives out of 23; 34.8%) was obtained by the two-day enrichment in peptone-sorbitol-bile (PSB) broth followed by plating on CIN agar plates. The two-day enrichment in PSB followed by potassium hydroxide (KOH) treatment before plating onto CIN agar gave 7 positives out of 23 (30.4%), decreasing to 3 positives (13.0%) without KOH treatment. The worst results were obtained by prolonged (five days) enrichment in PSB, with or without KOH treatment, followed by plating on CIN agar: 4.3% (1 out of 23) and 0.0% recovery rates, respectively. The mean concentration was 1.9 × 10(4)CFU/g, with a minimum of 1.0 × 10(2)CFU/g and a maximum of 5.8 × 10(4)CFU/g, thus demonstrating that tonsils may play an important role in contamination of pluck sets, carcasses, and slaughterhouse environment. Prevalence of virulence genes among the Y. enterocolitica 4/O:3 isolates was as follows: 12/22 (54.5%) for yadA, 21/22 (95.5%) for ail, 21/22 (95.5%) for inv and 22/22 (100%) for ystA. All Y. enterocolitica 4/O:3 isolates were sensitive to amoxicillin/clavulanic acid, ciprofloxacin and ceftazidime and resistant to ampicillin and cephalotin. High proportions of 4/O:3 isolates (95%) were sensitive to cefotaxime, gentamicin, kanamicin and nalidixic acid. High levels of resistance were observed to sulphonamide compounds (91%), streptomycin (64%) and chloramphenicol (55%). Multi-resistant isolates were very common; resistance to three or more antimicrobials was observed in 91% (20/22) of 4/O:3 isolates. High level of resistance to chloramphenicol was possibly due to coresistance to tiamphenicol, which was detected in 100% of the isolates. XbaI-PFGE detected four clusters among the 22 Y. enterocolitica 4/O:3 isolates. The most represented accounted for 77% (17/22) of the isolates, the second most common was found in 14% (3/22) of the isolates and the two other profiles were observed in single isolates. The comparison with a selection of human isolates supported the role of the pig as reservoir of 4/O:3 Y. enterocolitica., (Copyright © 2014 Elsevier B.V. All rights reserved.)

In 2005-2008, 1152 samples (451 faecal samples, 451 carcass swabs and 250 tonsils) were collected from 451 finishing pigs slaughtered in three abattoirs of northern Italy. In two abattoirs, 34 scalding water samples were collected. The aim of this study was to investigate the faecal and palatine tonsil carriage rate of Salmonella enterica and Yersinia enterocolitica in pigs at slaughter and the degree of carcass contamination by these bacteria. Typing of the isolates, virulence characterization and antimicrobial testing were also performed. S. enterica was isolated from 21.5% of the faecal samples, 10.9% of the carcasses and 10.4% of the tonsils, but not from scalding water. Nineteen different serovars were identified among 172 S. enterica isolates. The prevalent serovars were Derby (41.3%), Rissen (12.2%), Typhimurium (11%), 4,[5],12:i:- (8.7%) and Give (4.1%). S. enterica ser. Typhimurium and S. enterica ser. 4,[5],12:i:- isolates were phage-typed and PT DT120 was the most common (23.5%). Y. enterocolitica was detected in 17.1% of the faecal samples, 2.4% of the carcasses, 10.8% of the tonsils and 11.8% of the scalding water samples. A total of 119 isolates were found, four of them in water. Of the 115 Y. enterocolitica isolates of pig origin, 24 (20.9%) were 4/O:3 and 4 (3.5%) were 2/O:9. Y. enterocolitica 4/O:3 represented 85.7% of the pathogenic isolates found in all types of samples and 100% of those found in tonsils. In 4/O:3 isolates the most common virulence-associated genes were ystA (100%), inv (95.8%), ail (87.5%) and yadA (54.2%). In 2/O:9 isolates the prevalent genes were ail (100%), inv (100%) and ystA (100%), followed by ystB (25.0%). The majority (75.7%) of Y. enterocolitica isolates was biotype 1A, belonging to 13 serotypes (O:3; O:5; O:4,32-4,33; O:6,30-6,31; O:7,8-8; O:7,8-8-8,19; O:7,13; O:8; O:9; O:13; O:16-16,29; O:41,42-41,43; O:52). The most common virulence genes in 1A isolates were inv (95.4%) and ystB (72.4%). The antimicrobial resistance test showed that all Salmonella isolates were susceptible to cefotaxime, ciprofloxacin, cefalothin, gentamicin and enrofloxacin. Resistances to tetracycline (56%), sulphonamide compounds (42%) and streptomycin (34%) were the most common. All Y. enterocolitica isolates were susceptible to ciprofloxacin, ceftazidime, cefotaxime, chloramphenicol, enrofloxacin, gentamicin, kanamicin and neomycin. Most isolates were resistant to cefalothin (92%) and ampicillin (89%). Apparently, carcass contamination by S. enterica and Y. enterocolitica was more likely attributable to cross-contamination than to self-contamination, suggesting that good hygienic measures and slaughtering procedures can control transmission of these pathogens to pork meat., (Copyright © 2013 Elsevier B.V. All rights reserved.)

In 2001, a compulsory active surveillance system was started in the European Union to assess the prevalence of bovine spongiform encephalopathy (BSE) in the cattle population. The aim of the current study was to report on the field performances of 3 rapid tests: a Western blot (WB), a chemiluminescence enzyme-linked immunosorbent assay (ELISA), and an immunochromatographic assay, routinely used at 3 laboratories of the Istituto Zooprofilattico Sperimentale of Lombardia and Emilia Romagna, over 8 years of BSE monitoring activity. A total of 2,802,866 samples from slaughtered animals and 202,453 samples from fallen stock were tested by 1 of 3 tests. Positive results of the rapid tests were confirmed by histopathological examination, immunohistochemistry, and confirmatory WB. The field performances (i.e., initial reactive and false-positive rates) and practical aspects regarding resources and applicability of the tests to high-throughput routine testing laboratories were evaluated. The 3 tests proved to be reliable tools when applied to slaughtered samples, showing no or very low false-positive rates (<1 per 100,000 negative samples tested) and low retesting frequencies (0.02-0.26%). When samples from fallen stock were analyzed, performances of the immunochromatographic assay, and especially the chemiluminescence ELISA, were negatively affected, resulting in higher false-positive and retesting rates. On the other hand, both tests are less expensive, much easier to use, provide more rapid results, and adapt well to application in routine laboratories as compared with WB. In the authors' experience, the immunochromatographic assay was a good compromise between performance and convenience.

Amino acid polymorphisms of the prion protein gene influence sheep susceptibility to classical and atypical scrapie. Substitutions at codons 136, 154 and 171 play an important role in classical scrapie. Codon 141 leucine to phenylalanine mutation (AFRQ) has been recognized as an increased risk factor for atypical scrapie. In addition a rare allele with lysine at codon 171 (ARK) has been detected in Mediterranean sheep breeds. The presence of ARK poses two problems: the determination of its frequency and its possible interference with genotyping output of routine methods lacking specific detection capacity for ARK. The aim of our work was the development of a routine genotyping method with the capacity to identify ARK and AFRQ in addition to the normally detected alleles and to determine the frequencies of all these alleles in 5 main Italian breeds: Sarda (n=2494), Bergamasca (n=2686), Appenninica (n=297), Comisana (n=361) and Massese (n=402). A multiplex primer extension assay targeting the six single nucleotide polymorphisms of interest was developed. Allele frequencies revealed a very low level of ARR in Bergamasca (6.91%) as opposed to the other breeds, very diverse levels of AFRQ ranging from absence in Comisana to 10.70% in Massese and a restricted presence of ARK. This allele has only been detected in Bargamasca with a significant 3.67% and marginally in Appenninica (0.34%). These results underline the need for adequate routine methods for genotyping of breeds with alleles that can interfere with typing of important codons such as the case of ARK for codon 171.