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Background Biological drugs have improved the management of immune-mediated inflammatory diseases (IMIDs) despite being associated with important safety issues such as immunogenicity, infections, and malignancies in real-world settings. Objective The aim of this study was to explore the potential of a large Italian multi-database distributed network for use in the postmarketing surveillance of biological drugs, including biosimilars, in patients with IMID. Methods A retrospective cohort study was conducted using 13 Italian regional claims databases during 2010–2019. A tailor-made R-based tool developed for distributed analysis of claims data using a study-specific common data model was customized for this study. We measured the yearly prevalence of biological drug users and the frequency of switches between originator and biosimilars for infliximab, etanercept, and adalimumab separately and stratified them by calendar year and region. We then calculated the cumulative number of users and person-years (PYs) of exposure to individual biological drugs approved for IMIDs. For a number of safety outcomes (e.g., severe acute respiratory syndrome coronavirus 2 [SARS-COV-2] infection), we conducted a sample power calculation to estimate the PYs of exposure required to investigate their association with individual biological drugs approved for IMIDs, considering different strengths of association. Results From a total underlying population of almost 50 million inhabitants from 13 Italian regions, we identified 143,602 (0.3%) biological drug users, with a cumulative exposure of 507,745 PYs during the entire follow-up. The mean age ± standard deviation of biological drug users was 49.3 ± 16.3, with a female-to-male ratio of 1.2. The age-adjusted yearly prevalence of biological drug users increased threefold from 0.7 per 1000 in 2010 to 2.1 per 1000 in 2019. Overall, we identified 40,996 users of biosimilars of tumor necrosis factor (TNF)-α inhibitors (i.e., etanercept, adalimumab, and infliximab) in the years 2015–2019. Of these, 46% (N = 18,845) switched at any time between originator and biosimilars or vice versa. To investigate a moderate association (incidence rate ratio 2) between biological drugs approved for IMIDs and safety events of interest, such as optic neuritis (lowest background incidence rate 10.4/100,000 PYs) or severe infection (highest background incidence rate 4312/100,000 PYs), a total of 43,311 PYs and 104 PYs of exposure to individual biological drugs, respectively, would be required. As such, using this network, of 15 individual biological drugs approved for IMIDs, the association with those adverse events could be investigated for four (27%) and 14 (93%), respectively. Conclusion The VALORE project multi-database network has access to data on more than 140,000 biological drug users (and > 0.5 million PYs) from 13 Italian regions during the years 2010–2019, which will be further expanded with the inclusion of data from other regions and more recent calendar years. Overall, the cumulated amount of person-time of exposure to biological drugs approved for IMIDs provides enough statistical power to investigate weak/moderate associations of almost all individual compounds and the most relevant safety outcomes. Moreover, this network may offer the opportunity to investigate the interchangeability of originator and biosimilars of several TNFα inhibitors in different therapeutic areas in real-world settings. Supplementary Information The online version contains supplementary material available at 10.1007/s40259-021-00498-3.

Background: Magna Graecia is the ancient name for the modern geopolitical region of South Italy extensively populated by Greek colonizers, shown by archeological and historical evidence to be the oldest wine growing region of Italy, crucial for the spread of specialized viticulture around Mediterranean shores. Here, the genetic diversity of Magna Graecia grape germplasm was assessed and its role in grapevine propagation around the Mediterranean basin was underlined. Results: A large collection of grapevines from Magna Graecia was compared with germplasm from Georgia to the Iberian Peninsula using the 18 K SNP array. A high level of genetic diversity of the analyzed germplasm was determined; clustering, structure analysis and DAPC (Discriminant Analysis of Principal Components) highlighted the genetic relationships among genotypes from South Italy and the Eastern Mediterranean (Greece). Gene flow from east (Georgia) to west (Iberian Peninsula) was identified throughout the large number of detected admixed samples. Pedigree analysis showed a complex and well-structured network of first degree relationships, where the cultivars from Magna Graecia were mainly involved. Conclusions: This study provided evidence that Magna Graecia germplasm was shaped by historical events that occurred in the area due to the robust link between South Italian and Greek genotypes, as well as, by the availability of different thermal resources for cultivars growing in such different winegrowing areas. The uniqueness of this ampelographic platform was mainly an outcome of complex natural or human-driven crosses involving elite cultivars.

BACKGROUND Thyroid nodules diagnosed as Thy3B at fine-needle aspiration biopsy have a relevant risk of malignancy (15-30%) and are usually addressed to surgery. However surgery will result unnecessary in most cases. The present study aims at evaluating the possible increase of diagnostic accuracy for predicting malignancy using novel sonographic and elastographic parameters. METHODS In fifty patients undergoing thyroidectomy because of a Thy3B thyroid nodule, sonographic and elastosonographic evaluation was carried out by single operator before surgery. Five sonographic parameters (echogenicity, irregular margins, microcalcifications, intra-nodule blood flow and its irregularity) and two elastosonographic parameters (intra-nodule stiffness and its extension to adjacent tissue) were considered. After obtaining histological diagnosis, diagnostic accuracy was calculated. RESULTS When the two procedures were analyzed separately, sensitivity, specificity, positive (PPV) and negative (NPV) predictive values were 100%, 85%, 63% and 100% for ultrasonography and 60%, 92.5%, 67%, 90% for elastrosonography, respectively. The newly introduced evaluation procedures increased sensitivity. When a combined sonographic and elastosonographic evaluation was introduced, diagnostic accuracy was significantly improved: when ≥4 out of the seven parameters indicated were present, the risk of malignancy was very high (sensitivity 100%, specificity 92.55%, PPV 77%, NPV 100%). CONCLUSIONS A novel combined sonographic and elastosonographic parameter evaluation improved diagnostic accuracy for identifying thyroid nodules suspicious of malignancy.

Pyrethrum (Chrysanthemum cinerariifolium (Trev.) Vis. = Tanacetum cineariaefolium (Trev.) Schultz-Bip.) is a perennial herbaceous plant belonging to the family Asteraceae, native to Albania and the area of former Yugoslavia. It is the only species in the genus Tanacetum having an agronomic importance, although the genus consists of several species producing similar types of bioactive metabolites. The species is grown in order to obtain the insecticidal compounds collectively termed pyrethrins, which are found primarily in the flower heads. In this work we discuss the results found from a worldwide literature review about the micropropagation techniques followed on Pyrethrum, the in vitro culture conditions, and the ex vitro establishment trials under Mediterranean environmental conditions. Many technical problems concerning the propagation of the species seem to have been solved, and detailed protocols are available for an easy and fast propagation by seeds, vegetative splits, stem cuttings (rooted under mist or not), and tissue culture. The first attempts to introduce its cultivation into the semi-arid Mediterranean environments have brought to satisfactory results, and the species may be suggested as a valuable opportunity for the development strategies in new Mediterranean farming systems.

In Italy, grapevines are extensively cultivated, with Sicily representing one of the most significant wine regions. The high number of autochthonous grapevine varieties represents an important source of genetic diversity, and the many Sicilian varieties have anatomical and physiological traits that allow them to resist to different levels of drought stress. We investigated the water relation parameters of four cultivars of Vitis vinifera L. (Catarratto, Corinto, Nero d'Avola and Zibibbo) and characterized their leaf hydraulics. Measurements were conducted during summer on plants growing in the experimental field of the IBBR-CNR near Palermo. Daily patterns of leaf water potential (Yleaf) and stomatal conductance (gs) were measured in the field. Pressure-volume curves were constructed by the bench dehydration method to obtain leaf water potential at turgor loss point (Ytlp), osmotic potential at full rehydration (p0) and bulk modulus of elasticity (εmax). Leaf samples were collected to determine vein density using ImageJ. Major vein density was measured on digitally scanned leaves, while minor vein density was measured on photomicrographs of cleared and stained leaf portions.

Potassium availability affects drought responses in plants through several metabolic roles, among which stomatal regulation, cell growth and xylem hydraulics. Vitis vinifera L. is a highly valuable crop and several genotypes have been selected during its millennial cultivation. Varieties show differences in their adaptability to stress conditions, making them more or less suitable to certain climatic and edaphic conditions. The varieties cultivated in Sicily are characterized by high variability. We investigated the response of two Sicilian cultivars (Nero d'Avola and Catarratto) to potassium deficiency and drought stress. Two-year-old grafted plants were grown in agriperlite, with or without potassium in the fertigation solution, and subjected to moderate drought stress by suspending irrigation for 6-8 days. Potassium content of xylem sap, leaf and root tissues were measured with an ion-selective electrode. Changes in stomatal conductance, plant transpiration and hydraulic conductance were compared between genotypes and treatments, in order to gain information for the development of optimal fertigation practices and the selection of the most drought tolerant varieties.

L'obiettivo è quello di identificare più tipologie di pectina aventi caratteristiche chimico-fisiche, di purezza e trasparenza tali da poter essere applicate, da sole o in miscela con altre sostanze, in forma di gel. Tale studio permetterà di identificare quelle miscele che meglio si prestano ad essere utilizzate come film edibili da applicare sui prodotti ittici, con lo scopo di garantirne il mantenimento della shelf life.

The interest in sustainable agriculture has increased the demand of plant-derived compounds which can be less toxic both to mammals and to the environment than the synthetic agrochemicals. Chrysanthemum cinerariaefolium L. (Asteraceae), commonly termed pyrethrum, is an economically important crop from highlands of tropical and subtropical regions of the world. It is grown for the extraction of pyrethrins, natural insect repellents of plant origin. Pyrethrins are a mixture of six compounds produced by esterification of two acids (chrysanthemic and pyrethric acid) with three mono-terpene-alcohols (pyrethrolone-5, jasmolone-3 and cinerolone-4). The principale source of pyrethrins are the dried flowers of Chrysanthemum cinerariaefolium. Thanks to the low toxicity to mammals and other warm blooded animals, pyrethrum is the only plant species whose metabolites are currently commercially exploited in insecticides, and its worldwide demand exceeds the supply. Asteraceae species are considered as recalcitrant to successful growth in in vitro condition. Recalcitrance in root or shoot formation or in regeneration are associated to endogenous bacterial contamination, hyperhydricity, and tissue browning, hence studies on in vitro culture of Chrysanthemum cinerariaefolium have been rather limited. The aim of this study was to establish highly reproducible in vitro regeneration systems for an efficient multiplication of Chrysanthemum cinerariaefolium, since the pointing out of an efficient regeneration system could play an important role for the industrial exploitation of this plant. Petiole explants and leaf segments were used for micropropagation, callus induction and protoplast isolation. Sterile seeds were used as plant material source. The ability of petiole cuttings to produce direct shoot buds varied depending upon the different media composition tested for the experiments. Growth rate of shoots as well as root induction from shoots have been periodically analyzed. The in vitro raised plantlets were acclimatized and transferred to greenhouse with 60% success. For protoplast isolation and culture, young leaves of in vitro grown plants were used as initial plant material. Different enzymatic combinations were tested to achieve the highest protoplast release. We were able to recover a reasonable number of viable protoplast for further manipulation at the ploidy level with the aim to enhance the biological activity of Chrysanthemum cinerariaefolium and for the regeneration of novel insecticidal plant germplasm.

Pyrethrum (Chrisanthemum cinerariaefolium L. = Tanacetum cineariaefolium (Trev.) Schultz-Bip.) is a perennial herbaceous plant belonging to the family Asteraceae, native to Albania and the area of former Yugoslavia. Pyrethrum is the only species in the genus Tanacetum having an agronomic importance, although the genus consists of several species producing similar types of bioactive metabolites. The species is grown in order to obtain the insecticidal compounds collectively termed pyrethrins, which are found primarily in the flower head. Pyrethrum may be easily propagated by seeds, vegetative splits, stem cuttings (rooted or not under mist), and tissue culture. The first attempts to introduce its cultivation into the semi-arid Mediterranean environments have brought to satisfactory results, and the species has shown a good response in terms of biomass and flowers yield, even when technical inputs were applied in a reduced amount. Much work must still be done, however, in order to set a properly detailed management protocol for the genetic improvement of the species by means of biotechnology. In this work we discuss the first results of a specific experimental activity aimed to point out a micropropagation protocol for Pyrethrum, with the purpose to optimize the in vitro culture conditions, using explants sources as tissue donors for protoplast isolation and setting preliminary experiments in protoplast fusion methods to improve the flowers yield per plant and to increase the pyrethrins level.

To report contemporary estimates of the prevalence of hip-related osteoarthritis (OA) outcomes in African-Americans and Caucasians aged ≥ 45 years.

GENETIC CHARACTERISATION THROUGH MICROSATELLITES ANALYSIS OF SICILIAN WILD OLIVE FOR CONSERVATION AND VALORISATION OF NATURAL GENETIC RESOURCES Within the issues related to biodiversity conservation and valorisation of local plant germplasm a genetic analysis, through molecular markers, was carried out on 10 wild olive populations, from different areas of Sicily. Moreover, wild material was compared with five Sicilian cultivars, selected among the most widespread at regional scale. Results pointed out the absence of genetic affinities between wild populations and domestic varieties, supporting the assumption that domestication processes took place outside the regional territory. Although current oleaster communities are usually small relics of fragmented and degraded forests, this species still exhibits a certain intra- and inter-population genetic diversity. These results demonstrate that the gene pool of Sicilian wild olive represents a remarkable, but so far not sufficiently explored resource, of potential great concern both in agronomy and forestry.

The current rates of biodiversity decrease are estimated from 100 to 1,000 times the natural background levels. Among the initiatives to stop this trend, IUCN launched the campaign "Countdown 2010" to encourage and enable decision makers and stakeholders to uphold their commitments and achieve the goal to "halt the loss of biodiversity by 2010". Accordingly, the Italian Ministry of Environment funded in 2006 a project to save from the extinction the tree species Zelkova sicula, a very rare endemism from Sicily. Zelkova sicula is a relic of the Tertiary flora and belongs to a genus which became extinct in the whole continental Europe during the Quaternary glacial age. Currently the entire population consists of about 230 individuals and the species is included in the IUCN Red List as "Critically Endangered" of extinction. Several investigations showed that its current habitat is somewhat far away from a bioclimatic optimum and the most recent trends in climate change certainly give rise to discouraging scenarios. Summer water stress, exclusive vegetative regeneration, heavy anthropic disturbances and lack of active protection measures are the main points of criticism in the conservation of this species. Furthermore, the long geographic isolation and the limited population size might have caused a rapid loss of intra-specific genetic variability. The project to save Zelkova sicula consists of specific actions to promote in-situ and ex-situ conservation. It entails research activities, on one side, focusing on the multiplication and the genetic structure of the residual population, and, on the other side, education and awareness campaigning. CNR-IGV and Legambiente, respectively, are the subjects involved. Several steps have been performed. Numbering, labelling and positioning of each single tree through a GPS device were first carried out. Leaf material and current year twigs were then sampled from all trees in order to perform genetic analyses and vegetative multiplication. Different protocols for the vegetative propagation were developed. Cuttings were collected at three periods of the year (February, May and October) from three types of plant stocks (herbaceous, semi-woody and woody) and then treated with several concentrations of indole-3-butyric acid (IBA) and naphtalene acetic acid (NAA). Introduction in vitro and micropropagation were performed using newly flushed nodal explants as starting material. Explants were firstly sterilized under several conditions and then introduced in vitro on WPM medium supplemented with 6-benzylaminopurine (BAP) at different concentrations. The genetic diversity was investigated by molecular markers. Preliminary screening of genetic variation was performed using three ISSR primers. Then, codominant and potentially highly polymorphic nuclear SSRs were screened using primers designed in Zelkova serrata. Eleven SSR primers produced amplification products of the expected sizes in Zelkova sicula. Near absence of variation was observed in this species.

The polyphenol profile of an interspecific allotetraploid somatic hybrid, achieved by protoplast fusion, the 'Valencia' sweet orange (Citrus sinensis L. Osbeck) + 'Femminello' lemon (Citrus limon L. Burm), and three sexual hybrids obtained by backcrosses between Femminello lemon and the allotetraploid somatic hybrid ('Valencia' + 'Femminello') was studied by liquid chromatography-ultraviolet-diode array detector-mass spectrometry (LC-UV-DAD-MS). The aim of the work was to evaluate whether superior traits and improved performance can be observed in these new genotypes. Ten flavonoids (TF), comprising seven flavanones and three flavones, and four hydroxycinnamic acids (HCA) have been characterized and quantified in all hybrids and compared with those of the respective parents. The 'Valencia' + 'Femminello' somatic hybrid shows an intermediate polyphenol composition with respect to those of the parents, with a slight prevalence of lemon influence. The three sexual hybrids show, instead, different and more complex chromatographic profiles.

One of the most important goal in Citrus genetic improvement is the obtainment of seedless cultivars. Among the available strategies, we used the accomplishment of suitable interploid crosses (2X x 4X). To this purpose, three allotetraploid somatic hybrids have been used as pollen parent in sexual crosses with diploid 'Femminello' lemon, and autotetraploid 'Dancy' mandarin as pollen parent in crosses with diploid grapefruit and mandarin.

Five clones of sour orange (Citrus aurantium L.) showing significant morphological differences were selected from our germplasm collection and characterized both by genetic and leaf volatiles analysis. The genetic studies were undertaken by the use of molecular markers developed by PCR-based techniques (ISSR and RAPD), while the leaf essential oil patterns were obtained by chromatographic and mass spectrometric determination. Data obtained suggest that reasonably similar information can be achieved from the two techniques, supporting each other in characterization studies.

In order to obtain new somatic hybrids for genetic improvement of mandarin, several protoplast fusion experiments have been carried out. Isozyme banding pattern analysis and flow cytometry were used for early nuclear characterization. DNA of the hybrids was then extracted and subjected to ISSR-PCR using different primers anchored at 5’ and 3’ends. Successively, for cytoplasmic characterization of the hybrids, CAPs using universal mtDNA and cpDNA primers were performed. Our results showed that these new genotypes are definitely cybrids resulting asymmetric hybrids, in which the nucleus is inherited from the leaf parent and mitochondria from callus parent.

One of the most important goals in genetic improvement of lemon in the Mediterranean area is certainly the obtainment of new genotypes tolerant or resistant to mal secco disease. Two lemon cybrids (one diploid and one autotetraploid) with an intermediate degree of resistance to mal secco disease, spontaneously obtained by symmetric protoplast fusion between ‘Valencia’ orange and ‘Femminello’ lemon, were used as mother plants in backcrosses with a diploid clone of ‘Femminello’ Lemon Tolerant to the Mal Secco disease (LTMS) to improve tolerance and fruit quality. Since ‘Femminello’ lemon normally reproduces apomictically by nucellar embryony, an embryo-rescue technique was applied 105 days after pollination to recover zygotic embryos. A total of 66 plantlets were regenerated as follows: 20 from the 4nX2n cross, and 46 from the 2nX2n cross. In order to distinguish zygotic embryos from nucellars, flow cytometry was applied on all the 66 plantlets; and a modified AFLP (Amplified Fragment-Length Polymorphism) technique and microsatellite analysis were carried out on a sample of 20 seedlings (6 triploids and 14 diploids). Here we report the usefulness and reliability of the tested techniques and discuss the results obtained. Flow cytometry (FCM) was useful only in inter-ploid crosses, identifying 6 triploids, 4 of which were also detected by SSR (Simple Sequence Repeat) analysis and only 2 by modified AFLP. Among the 14 tested diploids, 5 zygotic genotypes were identified by SSR and 4 by modified AFLP. In our hands, the SSR technique appeared to be more suitable than modified AFLP.