Listeria monocytogenes Transmission from Donated Blood to Platelet Transfusion Recipient, Italy.

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.

Klebsiella pneumoniae carrying multiple alleles of antigen 43-encoding gene of Escherichia coli associated with biofilm formation.

A clinical strain of Klebsiella pneumoniae typed as sequence type 307 carrying three different alleles of the flu gene encoding the Escherichia coli virulence factor antigen 43 associated with biofilm formation was detected and characterized. The flu alleles are located in the chromosome inside putative integrative conjugative elements. The strain displays the phenotypes associated with Ag43, i.e. bi-phasic colony morphology and enhanced biofilm production. Furthermore, the strain produces low amount of capsule known to affect Ag43 function. Analysis of 1431 worldwide deposited genomes revealed that 3.7% Klebsiella pneumoniae carry one or two flu alleles.

Reduction trend of mcr-1 circulation in Emilia-Romagna Region, Italy.

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.

On the optimal control of SIR model with Erlang-distributed infectious period: isolation strategies.

Mathematical models are formal and simplified representations of the knowledge related to a phenomenon. In classical epidemic models, a major simplification consists in assuming that the infectious period is exponentially distributed, then implying that the chance of recovery is independent on the time since infection. Here, we first attempt to investigate the consequences of relaxing this assumption on the performances of time-variant disease control strategies by using optimal control theory. In the framework of a basic susceptible-infected-removed (SIR) model, an Erlang distribution of the infectious period is considered and optimal isolation strategies are searched for. The objective functional to be minimized takes into account the cost of the isolation efforts per time unit and the sanitary costs due to the incidence of the epidemic outbreak. Applying the Pontryagin's minimum principle, we prove that the optimal control problem admits only bang-bang solutions with at most two switches. In particular, the optimal strategy could be postponing the starting intervention time with respect to the beginning of the outbreak. Finally, by means of numerical simulations, we show how the shape of the optimal solutions is affected by the different distributions of the infectious period, by the relative weight of the two cost components, and by the initial conditions.

The Potential Role of Direct and Indirect Contacts on Infection Spread in Dairy Farm Networks.

Animals' exchanges are considered the most effective route of between-farm infectious disease transmission. However, despite being often overlooked, the infection spread due to contaminated equipment, vehicles, or personnel proved to be important for several livestock epidemics. This study investigated the role of indirect contacts in a potential infection spread in the dairy farm network of the Province of Parma (Northern Italy). We built between-farm contact networks using data on cattle exchange (direct contacts), and on-farm visits by veterinarians (indirect contacts). We compared the features of the contact structures by using measures on static and temporal networks. We assessed the disease spreading potential of the direct and indirect network structures in the farm system by using data on the infection state of farms by paratuberculosis. Direct and indirect networks showed non-trivial differences with respect to connectivity, contact distribution, and super-spreaders identification. Furthermore, our analyses on paratuberculosis data suggested that the contributions of direct and indirect contacts on diseases spread are apparent at different spatial scales. Our results highlighted the potential role of indirect contacts in between-farm disease spread and underlined the need for a deeper understanding of these contacts to develop better strategies for prevention of livestock epidemics.

A Model for Temporal Dynamics of Brown Rot Spreading in Fruit Orchards.

Brown rot, caused by Monilinia spp., is a major disease of stone fruit and, in favorable environmental conditions and in the absence of fungicide treatments, it causes important economic losses. In the present work, we propose a modification of classical susceptible, exposed, infectious and removed compartmental models to grasp the peculiarities of the progression of brown rot epidemics in stone fruit orchards in the last stage of the fruit growth (i.e., from the end of the pit hardening to harvest time). Namely, we took into account (i) the lifespan of airborne spores; (ii) the dependence of the latent period on the cuticle crack surface area, which itself varies in time with fruit growth; (iii) the impossibility of recovery in infectious fruit; and (iv) the abrupt interruption of disease development by the elimination of the host fruit at harvest time. We parametrized the model by using field data from a peach Prunus persica orchard infected by Monilinia laxa and M. fructicola in Avignon (southern France). The basic reproduction number indicates that the environmental conditions met in the field were extremely favorable to disease development and the model closely fitted the temporal evolution of the fruit abundance in the different epidemiological compartments. The model permits us to highlight crucial mechanisms undergoing brown rot build up and to evaluate the consequences of different agricultural practices on the quantity and quality of the yield. We found that winter sanitation practices (which decrease the initial infection incidence) and the control of the fruit load (which affects the host fruit density and the single fruit growth trajectory) can be effective in controlling brown rot in conjunction with or in place of fungicide treatments.

Rise and fall of outbreak-specific clone inside endemic pulsotype of Salmonella 4,[5],12:i:-; insights from high-resolution molecular surveillance in Emilia-Romagna, Italy, 2012 to 2015.

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.

Salmonella Detection and Counting on Pig Carcasses and Cutting Lines in Italian Slaughterhouses.

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 cm2 each (from both the exterior and interior side) were swabbed with a single sponge. In abattoir C, four 100 cm2 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]/cm2) in 46.6% of the carcasses and as high as 2.7 MPN/cm2 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/cm2.

Processing-Dependent and Clonal Contamination Patterns of Listeria monocytogenes in the Cured Ham Food Chain Revealed by Genetic Analysis.

The quantitative and qualitative patterns of environmental contamination by Listeria monocytogenes were investigated in the production chain of dry-cured Parma ham. Standard arrays of surfaces were sampled in processing facilities during a single visit per plant in the three compartments of the food chain, i.e., ham production (19 plants) and postproduction, which was divided into deboning (43 plants) and slicing (25 plants) steps. The numbers of sampled surfaces were 384 in ham production, with 25 positive for L. monocytogenes, and 1,084 in postproduction, with 83 positives. Statistical analysis of the prevalence of contaminated surfaces showed that in ham production, contamination was higher at the beginning of processing and declined significantly toward the end, while in postproduction, prevalence rose toward the end of processing. Prevalence was higher in the deboning facilities than in slicing facilities and was dependent on the type of surface (floor/drainage > clothing > equipment). The qualitative pattern of contamination was investigated through an analysis of the survey isolates and a set of isolates derived from routine monitoring, including longitudinal isolations. Pulsed-field gel electrophoresis (PFGE) and whole-genome single-nucleotide polymorphism (SNP) analysis revealed a remarkable clonality of L. monocytogenes within plants, with the detection of 16 plant-specific clones out of 17 establishments with multiple isolates. Repeated detections of clonal isolates >6 months apart were also observed. Six was the maximum number of between-isolate differences in core SNPs observed within these clones. Based on the same six-SNP threshold, three clusters of clonal isolates, shared by six establishments, were also identified. The spread of L. monocytogenes within and between plants, as indicated by its clonal behavior, is a matter of concern for the hygienic management of establishments.

Differential single nucleotide polymorphism-based analysis of an outbreak caused by Salmonella enterica serovar Manhattan reveals epidemiological details missed by standard pulsed-field gel electrophoresis.

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_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.

Pattern of tick aggregation on mice: larger than expected distribution tail enhances the spread of tick-borne pathogens.

The spread of tick-borne pathogens represents an important threat to human and animal health in many parts of Eurasia. Here, we analysed a 9-year time series of Ixodes ricinus ticks feeding on Apodemus flavicollis mice (main reservoir-competent host for tick-borne encephalitis, TBE) sampled in Trentino (Northern Italy). The tail of the distribution of the number of ticks per host was fitted by three theoretical distributions: Negative Binomial (NB), Poisson-LogNormal (PoiLN), and Power-Law (PL). The fit with theoretical distributions indicated that the tail of the tick infestation pattern on mice is better described by the PL distribution. Moreover, we found that the tail of the distribution significantly changes with seasonal variations in host abundance. In order to investigate the effect of different tails of tick distribution on the invasion of a non-systemically transmitted pathogen, we simulated the transmission of a TBE-like virus between susceptible and infective ticks using a stochastic model. Model simulations indicated different outcomes of disease spreading when considering different distribution laws of ticks among hosts. Specifically, we found that the epidemic threshold and the prevalence equilibria obtained in epidemiological simulations with PL distribution are a good approximation of those observed in simulations feed by the empirical distribution. Moreover, we also found that the epidemic threshold for disease invasion was lower when considering the seasonal variation of tick aggregation.

React or wait: which optimal culling strategy to control infectious diseases in wildlife.

We applied optimal control theory to an SI epidemic model to identify optimal culling strategies for diseases management in wildlife. We focused on different forms of the objective function, including linear control, quadratic control, and control with limited amount of resources. Moreover, we identified optimal solutions under different assumptions on disease-free host dynamics, namely: self-regulating logistic growth, Malthusian growth, and the case of negligible demography. We showed that the correct characterization of the disease-free host growth is crucial for defining optimal disease control strategies. By analytical investigations of the model with negligible demography, we demonstrated that the optimal strategy for the linear control can be either to cull at the maximum rate at the very beginning of the epidemic (reactive culling) when the culling cost is low, or never to cull, when culling cost is high. On the other hand, in the cases of quadratic control or limited resources, we demonstrated that the optimal strategy is always reactive. Numerical analyses for hosts with logistic growth showed that, in the case of linear control, the optimal strategy is always reactive when culling cost is low. In contrast, if the culling cost is high, the optimal strategy is to delay control, i.e. not to cull at the onset of the epidemic. Finally, we showed that for diseases with the same basic reproduction number delayed control can be optimal for acute infections, i.e. characterized by high disease-induced mortality and fast dynamics, while reactive control can be optimal for chronic ones.

Long-term gait analysis in patients after total knee arthroplasty: A systematic review and meta-analysis.

BACKGROUND: Gait abnormalities have been described in patients after total knee arthroplasty (TKA), leading to the development of inter-joint coordination abnormalities and increased risk of falling. Such impairments have been reported to persist in the long-term, although the majority of studies assessed gait pattern especially in the first months after TKA. RESEARCH QUESTION: What are the long-term gait impairments in patients after TKA compared to healthy age-matched subjects? METHODS: A systematic search was conducted on MEDLINE/PubMed, EMBASE, CENTRAL and Scopus databases. Observational studies or randomized controlled trials investigating gait spatial-temporal, kinematic and kinetics parameters in a time-window longer than 6 months in patients with TKA compared to healthy age-matched subjects were included. Methodological quality was assessed using the modified Downs and Black (D&B) checklist and participants' characteristics, surgical procedures details and outcome measures were extracted. Pooled or un-pooled findings were categorized into "6 months - 1 year" and "more than 1 year" timepoint categories. RESULTS: Twenty-eight studies (976 patients) were included. Overall quality was fair with a mean modified D&B score of 63.5 %. Reduced speed, stride length, cadence and longer stance phase were found in patients when compared to healthy individuals at "6 months - 1 year" follow-up. Spatial-temporal parameters deficits were also found at more than 1 year after TKA, where lower single-limb support and longer double-limb support durations were detected. These impairments occurred in concomitance with decreased knee range of motion along the sagittal and frontal planes and altered kinetic parameters. Hip kinematic and kinetic long-term impairments were also detected after TKA. SIGNIFICANCE: These findings highlighted long-term gait pattern alterations in patients with TKA compared to age-matched healthy subjects. Future studies should identify interventions able to reduce long-term gait pattern alterations and improve function in patients after TKA.

Unexpected consequences of culling on the eradication of wildlife diseases: the role of virulence evolution.

The removal of individuals from an infected population (culling) is a common strategy used to eradicate wildlife diseases. The manipulation of host density can impose strong selective pressures on pathogen virulence by changing the ecological conditions, thus affecting the effectiveness of eradication programs. We present an analysis of the effect of virulence evolution on culling by extending a susceptible-infected model to the case of competing strains with superinfection. To assess both short- and long-term effects, we first carried out the analysis on an ecological timescale, with a two-strain competition model; then we explore the dynamics of a continuum of pathogenic strains on evolutionary timescales using a quantitative genetics approach (when infection and evolutionary processes occur on comparable timescales) and a game-theoretic approach (when evolutionary processes occur on a slower scale). We demonstrate that the competition among pathogenic variants in the presence of superinfection affects outcome of culling campaigns, since increased host mortality may select for less virulent strains able to establish in sparser populations. This can lead to the counterintuitive result that disease abundance and prevalence may even increase with culling, thus making the eradication of infections considerably less likely. This is particularly relevant in the case of zoonoses where higher prevalence and abundance of pathogens in wild reservoirs may increase the risk of spillover in livestock and humans.

Exclusion and spatial segregation in the apparent competition between two hosts sharing macroparasites.

In this paper we investigate the spatial dynamics of a deterministic model describing two host species that partially share a common spatial domain, experiencing apparent competition mediated by macroparasites. The aim of this work is to understand the mechanisms underlying apparent competition processes in a spatially structured environment, which have been generally overlooked up to now. First, we analyse the behaviour of a single-host macroparasite partial differential equation (PDE) model, both in the cases of uniform or spatially-dependent vital rates of the host, focusing on the role of spatial diffusion on parasite persistence and host abundance. We obtained the threshold condition for parasite persistence, and (in contrast to what occurs in reaction-diffusion models for an isolated population) we found that, in the case of spatially-dependent vital rates, increasing the host diffusion coefficient results in an increase of the overall host population. Then, a PDE model featuring spatial diffusion and apparent competition mediated via shared macroparasites between two species is analysed in order to understand the role of spatial heterogeneity in host coexistence. We assumed a partial overlap among the habitats of the two species and found that the shared parasites could cause, depending on the values of the diffusion coefficients and differences in induced mortality among host species, a decrease of the realized habitat and, eventually, the extinction of the species less tolerant to parasite infection. This shows that the presence of regulating parasites complicates the effect of dispersal on population dynamics and that the dynamics of apparent competition cannot be adequately understood from spatially-independent models.

Different Roles of Wild Boars and Livestock in Salmonella Transmission to Humans in Italy.

Wild boar (Sus scrofa) is the most widely distributed large wildlife mammal worldwide. To investigate the transmission of Salmonella enterica amongst wild boars (Sus scrofa), humans, and livestock, we compared via pulsed-field gel electrophoresis and whole genome sequences the isolates of S. enterica serovar Typhimurium (biphasic and monophasic variants) and Enteritidis collected from wild boars, food-producing animals, and human patients in Emilia-Romagna region (Northern Italy) between 2017 and 2020. Specifically, we analysed 2175 isolates originated from human (1832), swine (117), bovine (128), poultry (76), and wild boar (22). The genomic analyses showed that wild boars shared most of their lineages of biphasic Typhimurium with bovines and most of Enteritidis with poultry, whilst we did not find any lineage shared with swine. Moreover, almost 17% of human biphasic Typhimurium and Enteritidis belonged to genomic clusters including wild boar isolates, but the inclusion of bovine and poultry isolates in the same clusters and the peculiar spatial distribution of the isolates suggested that human cases (and wild boar infections) likely originated from bovines and poultry. Consequently, wild boars appear not to play a significant role in infecting humans with these serovars, but seem to get infected themselves from livestock, probably through the environment.

Using Expert Elicitation for ranking hazards, promoters and animal-based measures for on-farm welfare assessment of indoor reared beef cattle: an Italian experience.

On-farm welfare assessment gives the opportunity to monitor and improve the quality of the animal life on the farm. In order to build the first Italian public standard for the welfare of indoor reared beef cattle, a list of 25 potential hazards and 22 potential promoters of beef cattle welfare was judged by a group of national experts by taking into account their negative or positive impacts on the welfare-state of the target population. In addition, the experts were asked to rank a list of 11 animal-based measures for identifying the most appropriate and important for measuring negative welfare outcomes in beef cattle. Based on experts' ratings, an "impact score" (ISoverall) was calculated for the proposed measures. Management hazards and promoters were ranked to have a greater impact on beef cattle welfare than housing factors. Keeping cattle in large (≥ 40 animals) and heterogeneous groups obtained the highest ISoverall among the proposed hazards (ISoverall = 5.54), followed by the presence of animals without free access to drinking water (ISoverall = 4.39) and the use of high-concentrate corn silage diets (concentrate > 80% and fiber < 6%) (ISoverall = 4.39). On the other hand, housing animals in small (≤ 20 animals) and homogeneous groups (ISoverall = 5.41), checking them at least twice a day (ISoverall = 4.62) and rearing cattle in loose housing systems with access to an outdoor area/pasture (ISoverall = 4.27) were ranked among the top 3 promoters. Concerning animal-based measures, experts scored lameness, severe respiratory diseases, body condition scoring and mortality rate to be measures most important for assessing serious welfare impairment.

Salmonella Derby adaptation to swine and simultaneous attenuation for humans go through decay of Salmonella Pathogenicity Island I.

IMPORTANCE: This study integrated population data with in vitro assessment of virulence phenotypes to unveil that a considerable part of the global population of Salmonella Derby is evolving to enhance its host adaptation to the swine host and that this evolution is simultaneously increasing its attenuation for humans. The study shows that the fixation of deleterious mutations in SPI-1 has a role in this process. This evidence indicates that SPI-1 has a key role for S. Derby virulence in humans but not for its circulation in swine. The results show that genes generally considered essential for Salmonella pathogenesis do not play the same key role for all Salmonella serovars or lineages and/or all hosts. The study helps in understanding the molecular mechanisms underlying the ecology and host adaptation of Salmonella showing that the adaptation process can vary for different types of Salmonella and hosts.

Population Structure of Listeria monocytogenes in Emilia-Romagna (Italy) and Implications on Whole Genome Sequencing Surveillance of Listeriosis.

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.