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Background: Safinamide modulates both dopaminergic and glutamatergic systems with positive effects on motor and non-motor symptoms of Parkinson's disease (PD). The drug utilization study SYNAPSES was designed to investigate the use of safinamide in routine clinical practice, as recommended by the European Medicines Agency. Objective: To describe the occurrence of adverse events in PD patients treated with safinamide in real-life conditions. Methods: The SYNAPSES trial is an observational, European, multicenter, retrospective-prospective cohort study. Patients were followed up to 12 months with analyses performed in the overall population and in patients aged >75 years, with relevant comorbidities and with psychiatric conditions. Results: Of the 1610 patients included, 82.4% were evaluable after 12 months with 25.1% of patients >75 years, 70.8% with relevant comorbidities and 42.4% with psychiatric conditions. During observation 45.8% patients experienced adverse events, 27.7% patients had adverse drug reactions and 9.2% patients had serious adverse events. The adverse events were those already described in the patients' information leaflet. The majority were mild or moderate and completely resolved and no differences were detected between the subgroup of patients. Clinically significant improvements were seen in the UPDRS motor score and in the UPDRS total score in >= 40% of patients, according to the criteria developed by Shulman et al. Conclusion: The SYNAPSES study confirms the good safety profile of safinamide even in special groups of patients. Motor complications and motor scores improved with clinically significant results in the UPDRS scale maintained in the long-term.

The spittlebug Philaenus spumarius L. is known to be the predominant vector of Xylella fastidiosa Wells et al. (Xf) to olive trees in the Apulia Region of Italy. However, factors influencing Xf transmission by P. spumarius have not been studied and, consequently, the epidemiology of Xf-associated emerging diseases cannot be properly understood. Here we investigated 1) the kinetics of the bacterial persistence and multiplication, along with transmission efficiency by the vector, and 2) the spread rate of Xf among olive plants in summer and autumn. For the first objective, olive-to-olive transmission efficiency, Xf retention and multiplication in P. spumarius adults were tested in four separate assays carried out in 2017-2018 in different seasons. For the second objective, experimental assays in microcosms were carried out in the same periods to assess influence of i) IAP duration, ii) climatic conditions (semi-field vs controlled), and iii) season on the spread of Xf infection within an olive seedling population. The results show that i) P. spumarius is a competent Xf vector to olive throughout the season and adult life ii) bacterial load in the vector foregut reaches a plateau after 2-3 weeks from acquisition iii) different climatic conditions and period of the year may result in significant differences in transmission rates and iv) differential survival of vectors -- influenced by insect age, season and climatic conditions -- may affect the spread of Xf in olive plants. In summary, this work describes the influence of seasonal and environmental factors on the transmission characteristics of Xf by P. spumarius. Such transmission dynamics may explain epidemic patterns in Apulian olive agroecosystem. The transmission parameters obtained in this study can be used in modelling the pathogen spread, by explicitly incorporating the effect of insect vectors, with the aim of designing effective control and prevention measures.

The meadow spittlebug Philaenus spumarius (Linnaeus) is a common and widespread xylem-sap feeder in Europe. The species has been recently discovered as the main vector of the gram-negative bacterium Xylella fastidiosa (Wells) known as the causal agent of the olive dieback in Apulia (southern Italy). Since then, the vector became one of the major concerns posed to European agriculture. Only the adult stage of P. spumarius is able to acquire the bacterium X. fastidiosa through feeding on the xylem sap of infected host plants. Once the adult acquired the bacterium, it is persistently infected and can immediately transmit the pathogen to healthy plants, as latency in the vector is not required. Therefore, the capacity to predict the population phenology and abundance of P. spumarius is a key element for understanding the epidemiology of X. fastidiosa as well as for the development of targeting control interventions for rational management strategies against both the vector and the bacterium. Here, we present a physiologically-based mechanistic model predicting the spatio-temporal variation of phenology and population abundance of P. spumarius. The model describes the life-history strategies of the different biological stages by means of stage-specific development, mortality and fertility rate functions. The rate functions describe the physiological responses of the species to local environmental forcing variables (e.g. air temperature) and resources (e.g., host plant availability). The rate function's parameters have been estimated through data obtained from laboratory and field experiments. The model has been calibrated and validated using independent population dynamics datasets from Apulia and Liguria regions (Italy). Model outputs can be used for assessing the risks linked to P. spumarius population dynamics and X. fastidiosa epidemiology. Model simulations can also support the design and implementation of monitoring and control activities for managing the X. fastidiosa-plant-vector pathosystem.

"Insects viruses as biocontrol tool in crop protection have received much attention in recent years due to their narrowed target and their environmentally-safe properties, which make them harmless both to human and non-target organisms. Indeed, baculoviruses have already been used for decades as biocontrol strategies against Lepidoptera. Although many formulates have been successfully developed against Lepidoptera, little attention has been payed to the selection of new suitable viruses able to control insects belonging to other orders (i.e. Hemiptera). In this perspective, the H2020-funded VIROPLANT project aims at identifying potential biocontrol agents from the virosphere of phytoplasma vectors. Phytoplasmas are wall-less bacteria which multiplies in the plant phloem. In particular, Flavescence dorée phytoplasma of grapevine was classified as quarantine pest by EFSA, since its fast spread causes severe damages to the vineyards. Although Scaphoideus titanus Ball (Hemiptera: Cicadellidae) is the principal vector of flavescence dorée (FD) of grapevine, Euscelidius variegatus Kirschbaum (Hemiptera: Cicadellidae) is used as an efficient vector of FD phytoplasma in laboratory condition, because, contrary to S. titanus, it is a polivoltine and polyphagous leafhopper. In this work we present for the first time four novel +ssRNA viruses putatively belonging to the family Iflaviridae found to be naturally present in both the above mentioned leafhoppers populations. Two iflaviruses, named Euscelidius variegatus virus 1 (EVV-1) and Euscelidus variegatus virus 2 (EVV-2), were serendipitously found in our in-house rearing of E. variegatus during an RNA seq analysis. Finally, other two iflaviruses were identified during an NGS analysis of S. titanus transcriptome. The latters, named Scaphoideus titanus iflavirus 1 (STiV-1) and Scaphoideus titanus iflavirus 2 (STiV-2), were discovered in a wild population sampled in Europe and in another sampled in the US, respectively. In particular, EVV-1 was found to be present in our lab rearing with a 100% prevalence. The discovery of a virus-free lab population coming from France allowed us to characterize the transmission routes of this iflavirus. The ability to transmit FD phytoplasma was also tested for both EVV-1 infected and non-infected individuals. Preliminary results showed that the virus was constantly detected both in phytoplasma-exposed and non-exposed vectors, but, interestingly, the virus load was significantly lower in FD-exposed insects, suggesting a potential cross-talk among insects, endogenous bacteria and viruses. Further experiments aiming at setting up an efficient inoculative strategy of EVV-1 in E. variegatus virus-free individuals are ongoing. An infectious clone derived from EVV1 was able to infect and replicate in virus-free insects. Even though the infection rates still need to be improved, the EVV-1 infectious clone might provide the unprecedented opportunity to manipulate the expression of endogenous insect genes by promoting virus-induced gene silencing by the iflavirus and interfere with insect ability to transmit phytoplasmas in both leafhopper target"

Flavescence dorée of grapevine (FD) is a phytoplasma-associated disease present in several European countries and transmitted by the nearctic leafhopper Scaphoideus titanus among plants of the genus Vitis. The leafhopper spreads FD through primary infections (from gone-wild vines surrounding the vineyard to cultivated grapevines) and secondary infections (from vine to vine within the vineyard). FD transmission experiments to the main Piedmontese grapevine cultivars (cvs) were carried out with the vector S. titanus, in order to screen for different levels of cv susceptibility to FD phytoplasmas. We made the hypotheses that tolerance could be due to plant response to phytoplasmas or could be mediated by vector-plant interaction. To test this latter hypothesis, based on the results of the transmission experiments, S. titanus fitness and feeding behavior were studied on three cvs with different susceptibility to FD: Barbera (susceptible), Brachetto and Moscato (tolerant). Survival tests revealed a higher mortality rate on Moscato compared to Barbera and Brachetto, while no major differences were highlighted in developmental time measured on the three cvs. Feeding behavior of the vector was investigated with the electropenetrography (EPG) technique. Hemipteran insects insert the stylets inside the leaf to feed in different plant tissues, and this technology allows studying feeding behavior as different electric waveforms and the association with different feeding phases. The EPG experiments on S. titanus allowed a precise description of feeding behavior on the above mentioned grapevine cvs, thus shedding light on the mechanisms of susceptibility/tolerance of grapevine to FD eventually mediated by the vector.

Colony Collapse Disorder (CCD) of honey bee has been widely studied in the past decade and it is nowadays well recognized as a syndrome with multifactorial origin that leads to sudden depopulation of the hives. Among factors leading to CCD, pathogens and parasites may play a major role. According to the literature, the chronic exposure of bee colonies to biotic stressors, such as viruses and parasites, acts in an indirect way by deregulating the immune pathways linked to the antimicrobial response (Toll and Imd pathways). Nevertheless, a better comprehension of the impact of every actor involved is necessary to elucidate the mechanisms that underlie these interactions. In the frame of the Interreg-Alcotra project "Innov'api", which was monitoring the health status of several honey bee colonies managed with two alternative methods for varroa mite control, we selected a subset of samples collected from Piedmontese apiaries in July and September 2019 (before and after the summer acaricide treatment). The selected colonies were characterized by a wide range of viral loads, previously assessed by qPCR analyses carried out on pooled adult bee samples. We then investigated the variability of infection level among individual bees belonging to the same colonies, by focusing on the three most widespread ssRNA+ viruses (Deformed Wing virus, Black Queen Cell virus, Sacbrood virus). The microsporidian Nosema ceranae, another major stressor involved in CCD, was also included in the analysis. On the same samples, we quantified the expression levels of five genes belonging to the Toll and Imd pathways (three antimicrobial peptides, a peptidoglycan receptor and a NF-kB factor) and of vitellogenin, a protein involved in the immunocompetence, in order to investigate their correlations with biotic stress level. Viral loads as well as gene expression values showed very different levels among individuals within the same colony, allowing us to study how these pathogens may affect the individual immune responses. Our multivariate analysis suggests that DWV and BQCV loads positively correlated with the production of antimicrobial peptides such as apidaecin and hymenoptaecin and with the expression level of the NF-kB factor dorsal-1A. Moreover, the type of varroa control treatment had an impact on viral loads and gene expression levels, whereas vitellogenin transcript levels were mostly affected by the sampling time. As the microsporidian N. ceranae was seldom detected, we were not able to evaluate the impact of this pathogen on the regulation of the investigated pathways.

Varroa destructor mite is regarded as a major parasite of Apis mellifera, as it is responsible for many colony losses both directly and indirectly, by virus transmission. The current control strategies are based on the use of synthetic acaricides, which can compromise the quality of the hive products due to the presence of chemical residues, and whose activity is limited by the selection of resistant strains over time. Through a collaboration with the INRAE-PACA researchers from Avignon and with the Italian and French national associations of beekeepers UNAAPI and ADA France (Interreg-Alcotra project "Innov'api"), we tested a sustainable control technique against varroa. To this aim the innovative technique of brood removal combined with oxalic acid treatment was compared to conventional control with a synthetic acaricide (amitraz). In Piedmont, more than 200 colonies from both migratory and stationary apiaries managed by local beekeepers were monitored: the project took into account several parameters, such as population of adult workers and brood amount, productivity and health status of the hives. The latter was described by the assessment of varroa infestation levels (obtained by counting the phoretic mites) and infection levels of five viruses (deformed wing virus, DWV; acute and chronic bee paralysis virus, ABPV and CBPV; black queen cell virus, BQCV; sacbrood virus, SBV). Quantification of the viral loads was carried out through Real-time qPCR analyses, after reverse transcription of the RNA extracted from two pools of 40 nurse bees per colony. The "Innov'api" experimental design, by including at least six sampling dates per year and having covered three beekeeping seasons (2017-2020), has allowed us to describe the seasonal trends of the main honey bee viruses in relation to the population dynamics of the parasitic mite and host colonies. In Piedmont, a high prevalence of DWV, BQCV and SBV was found, with virus-specific fluctuations, whereas ABPV and CBPV were detected sporadically. However, considering the viral loads of the infected samples, we highlighted positive correlations between ABPV and DWV, as well as between BQCV and SBV. Showing higher levels in spring and summer, BQCV and SBV shared similar dynamics to those of the hives and positively correlated with the number of adult bees and brood. By reaching maximum peaks in autumn, DWV followed the trend of varroa population. Moreover, DWV load was positively correlated with the infestation levels of the hives. The overall results show that the brood removal technique is as effective, and sometimes quicker, in lowering the varroa population, as well as the associated viral load, as the conventional technique.

The spread of insect vectors is a key point in understanding the epidemiology of insect-borne plant diseases and in assessing effective vector control strategies. The transmission of the bacterium Xylella fastidiosa Wells in Europe is mainly due to spittlebugs (Hemiptera: Aphrophoridae). The meadow spittlebug Philaenus spumarius L. plays the major role in the spread of X. fastidiosa ssp. pauca ST53 on olives in the Apulia region (Italy). Despite its importance, little information is available on the mobility of P. spumarius. Dispersal capabilities of the meadow spittlebug were hence investigated by mark-release-recapture experiments. Several trials were carried out from May to October of 2016-2017 in two agroecosystems: olive grove and grass meadow, in the Apulia and the Piedmont regions (Italy), respectively. Adults of P. spumarius of both sexes were collected in natural grassland habitats, marked with an aqueous solution of albumin (10%) and then released at a single point in the centre of the experimental area. A total of ?6500 and ?3000 spittlebugs were release during Apulia and Piedmont experiments, respectively. Spittlebugs were recaptured through sweep net in fixed sampling points located up to a maximum of 250 m from release point. Recaptures were conducted every two or three days up to 17 days after the release. Marked insects were identified via an indirect ELISA. The dispersal capacity of P. spumarius adults was described estimating the probability density function describing the distribution of the end locations of insects relative to the source point (i.e. the dispersal kernel). Diffusion rates in the two agroecosystems were estimated under the hypothesis of a random walk and applying a Gaussian kernel. Spittlebugs were recaptured up to 155 meters and 200 m in Apulia and Piedmont, with 50% of total recaptures within 30 m and 40 m from release point, respectively. Results showed a high variability in the estimated median distance from the release point, ranging from 25 m (Apulia) to 35 m (Piedmont) per day, and from 356 m (Apulia) to 507 m for the estimated median of adult life-long dispersion. Estimated spread parameters of P. spumarius are fundamental information to model X. fastidiosa spread in Apulia and other European foci. Further researches are needed to investigate on influence of both vector's physiology and environmental factors on movement tendency and directionality of P. spumarius.

In the context of the severe epidemic spread of Xylella fastidiosa subsp. pauca ST53 (Xfp) in the Apulia Region (southern Italy), there is a need to urgently rethink cropping system for restoring agricultural productions and landscapes. Efforts are devoted to identifying species immune to Xfp (as alternative crops), and to assess the impact of host species showing low susceptibility/resistance to the infections. To this end, we performed transmission experiments from olives and almond, to assess vector acquisition and transmission from host plants showing different susceptibility and low bacterial population size (

Phytoplasmas are wall-less bacteria associated with severe diseases in hundreds of plant species, producing in many cases major yield loss in several crops. Phytoplasmas inhabit the phloem elements of infected plants and are transmitted from plant to plant by hemipteran insect species, belonging to the suborders Auchenorrhyncha (Cicadellidae and Cixiidae are the families with the largest number of vector species) and Sternorrhynca (Psyllidae family). Control of phytoplasma diseases is so far mainly based on insecticide treatments against vector populations. Taking into account the impacts of insecticides on agro-economy, public health, and environment, the development of innovative and more sustainable management strategies is becoming urgent. The leafhoppers Scaphoideus titanus and Euscelidius variegatus are known phytoplasma vectors. The former species is the natural vector of the phytoplasma associated with the grapevine flavescence dorée (FD), 16SrV-C and -D; the latter is the natural vector of the 'Candidatus Phytoplasma asteris' Chrysanthemum yellows strain, 16SrI-B, and a laboratory vector of FD phytoplasma. Previous studies indicated a crucial role of the insect ATP synthase during phytoplasma infection process. In the present work, the silencing of ATP synthase ? was obtained by injection of specific dsRNAs (dsATP) in both S. titanus and E. variegatus. After the injection of dsRNAs in E. variegatus and successive phytoplasma acquisition on infected plants, silenced E. variegatus showed significant reduction of the target protein and lower phytoplasma loads compared to the not-silenced controls. An in vivo role for insect ATP synthase ? could be hypothesized during phytoplasma infection process. Complete lack of progeny was also observed in several repeated experiments for E. variegatus insects following silencing of ATP synthase ?. Consistently, altered morphology of ovaries as well as deregulation of genes involved in egg development were observed in ATP synthase ? silenced E. variegatus adult females in comparison with specimen injected with dsGFP, used as negative controls. Our results are in line with the known role of ATP synthase in promoting germ cell differentiation, as already described in the fruit fly Drosophila melanogaster. The strong sterilizing effect obtained by silencing ATP synthase ? could pave the way towards the development of innovative control approaches to limit phytoplasma vector population, although many issues still need to be addressed, such as specificity, to avoid undesired off-target effects, and efficient strategies to deliver dsRNAs to phloem-sucking insects.

The Panel on Plant Health of EFSA conducted a pest categorisation of 17 viruses of Rubus L. that were previously classified as either non-EU or of undetermined standing in a previous opinion. These infectious agents belong to different genera and are heterogeneous in their biology. Blackberry virus X, blackberry virus Z and wineberry latent virus were not categorised because of lack of information while grapevine red blotch virus was excluded because it does not infect Rubus. All 17 viruses are efficiently transmitted by vegetative propagation, with plants for planting representing the major pathway for entry and spread. For some viruses, additional pathway(s) are Rubus seeds, pollen and/or vector(s). Most of the viruses categorised here infect only one or few plant genera, but some of them have a wide host range, thus extending the possible entry pathways. Cherry rasp leaf virus, raspberry latent virus, raspberry leaf curl virus, strawberry necrotic shock virus, tobacco ringspot virus and tomato ringspot virus meet all the criteria to qualify as potential Union quarantine pests (QPs). With the exception of impact in the EU territory, on which the Panel was unable to conclude, blackberry chlorotic ringspot virus, blackberry leaf mottle-associated virus, blackberry vein banding-associated virus, blackberry virus E, blackberry virus F, blackberry virus S, blackberry virus Y and blackberry yellow vein-associated virus satisfy all the other criteria to be considered as potential QPs. Black raspberry cryptic virus, blackberry calico virus and Rubus canadensis virus 1 do not meet the criterion of having a potential negative impact in the EU. For several viruses, the categorisation is associated with high uncertainties, mainly because of the absence of data on biology, distribution and impact. Since the opinion addresses non-EU viruses, they do not meet the criteria to qualify as potential Union regulated non-quarantine pests.

For insect-borne plant pathogens, transmission biology is of major importance in outlining the disease epidemiology. The characteristics of acquisition, persistence and transmission of X. fastidiosa ST53 by the spittlebug Philaenus spumarius, the main vector in Apulia, are not yet described; similarly, transmission competence of the potential vector Cicadella viridis, the most common sharpshooter in Europe, is unknown. In this perspective, two sets of experiments were performed in 2017 and 2018 to study: i) the kinetics of bacterial multiplication and persistence in P. spumarius and C. viridis; ii) the influence of temperature, season and age of P. spumarius adults on simulated epidemic progression on olive plants under indoor and outdoor conditions. For the kinetics experiments, following the acquisition, insects were serially transferred in groups of five to olive or periwinkle test plants. In simulated epidemic progression experiments, after the acquisition, groups of insects were isolated in cages with 16 olive seedlings for different inoculation periods. Acquisition and transmission rates were assessed by testing individual insects after inoculation and by testing recipient plants 6and 10 months post-inoculation. Furthermore, acquisition and transmission of X. fastidiosa ST53 by C. viridis were tested through an in vitro acquisition system. Overall, about 900 insects and 170 plants were tested in kinetics experiments, while about 800 spittlebugs and 1,500 plants were studied in simulated epidemic progression experiments. Preliminary results for P. spumarius indicate: a) a higher acquisition efficiency in September than July; b) a lower acquisition efficiency from periwinkle compared with olive as source plants, but higher transmission efficiency to periwinkle compared with olive as recipient plants. Cicadella viridis was able to acquire and transmit X. fastidiosa following acquisition on artificial diet or periwinkle, although with low efficiency.