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Anthropic areas play a pivot role for main wolf conservation challenges. Wolf presence in the higher Pisan hills has been well documented while wolf settlement in the lower Pisan hills is still uncertain. In this study, long-term information on wolf presence in a highly anthropic area of the lower Pisan hills was collected by using non-invasive monitoring techniques. Furthermore, both the relationship of this predator with human activity and the impact of hunting on wolf presence have been investigated. The results obtained indicate the presence of a stable and reproductive wolf pack composed by both Italian wolf and hybrids individuals in the municipalities of Crespina Lorenzana and Casciana Terme Lari. A high impact of wolf on livestock was recorded in this area since no prevention systems were adopted by farmers. Wolf appears not to have a negative impact on wild boar population. Similarly, wild boar drive hunting does not appear to affect the wolf pack presence in the area. Thereby wolf may play a key role as controller of wild population. Prevention strategies improvement becomes instrumental to promote wolf-human coexistence. Further investigation to monitor pack hybridization level and turnover and to assess the impact of packs on wild population and livestock in anthropic areas is desirable.

This study shows the results of an interdisciplinary research aimed at devising artificial intelligence methods to support data understanding in Cultural Heritage. The objective is to create automated methods for automatically classifying and successively retrieving photographic data leveraging the current breed of AI methods based on Deep Learning paradigm. In this work, the lack of images to be used for the training of the AI system is addressed by testing an approach based on Generative Adversarial Networks (GANs) for automatically synthesizing unrealistic photos, which, in turn, can be used for training image classification and retrieval systems. Specifically, we propose a method to drive the generation of realistic classical order images using GAN approaches anchored to semantic ontology domain representation. More specifically, the proposed AI leverages the advantages of knowledge-based and data-driven methods and foresees a level image generation: the first one that synthesizes isolated objects corresponding to each ontology concept and the second one that, instead, combines the generated objects according to the spatial information provided by the ontology in order to generate realistic scenes. The resulting images can then be employed to train automated classifiers and more reliable retrieval methods by minimizing the efforts usually required by human operators to manually annotate data. In addition, the proposed generative strategy allows for a deeper understanding of cultural heritage visual data enabling the possibility to enhance design capabilities in simulation contexts.

Free-ranging corvids&mdash, 678 magpies (Pica pica) and 120 hooded crows (Corvus cornix) from nine protected areas of the Pisa province (central Italy)&mdash, were examined for Toxoplasma gondii and Trichinella pseudospiralis. The intracardiac blood clots from 651 magpies and 120 hooded crows were serologically examined for T. gondii. The DNA extracted from the hearts of seropositive birds was then used to perform a nested PCR for the amplification of the T. gondii B1 gene and for genotyping for SAG genetic markers. Breast muscle samples from 678 magpies and 91 hooded crows were tested by an artificial digestion method for Trichinella. Data were statistically analyzed. Forty-five (5.8%&mdash, 41 magpies and four hooded crows) out of the 771 examined animals scored seropositive for T. gondii, with titers ranging from 1:25 to 1:100. T. gondii DNA was detected in 15 of the 45 positive birds and T. gondii genotypes II and III were identified. No positivity for T. pseudospiralis was found. No significant differences between the two species of corvids and among the different areas of origin were observed for seropositivity to T. gondii. This is the first extensive study on both T. gondii and T. pseudospiralis in magpies and hooded crows, as well as the first detection of T. gondii SAG genotypes in magpies.

Integrating human-provided location priors into video object segmentation has been shown to be an effective strategy to enhance performance, but their application at large scale is unfeasible. Gamification can help reduce the annotation burden, but it still requires user involvement. We propose a video object segmentation framework that leverages the combined advantages of user feedback for segmentation and gamification strategy by simulating multiple game players through a reinforcement learning (RL) model that reproduces human ability to pinpoint moving objects and using the simulated feedback to drive the decisions of a fully convolutional deep segmentation network. Experimental results on the DAVIS-17 benchmark show that: 1) including user-provided prior, even if not precise, yields high performance; 2) our RL agent replicates satisfactorily the same variability of humans in identifying spatiotemporal salient objects; and 3) employing artificially generated priors in an unsupervised video object segmentation model reaches state-of-the-art performance.

The vegetarian diet of many herbivorous mammals is supplemented with proteins of animal origin, especially in young individuals and in breeding females, to provide key proteins necessary for both growth and breeding. Among porcupine species, only the Cape porcupine (Hystrix africaeaustralis) has been observed to consume carrion flesh. From June to August 2019, a pigeon carcass was placed together with corn in 7 study settlements and near 2 monitored capture-traps, in order to assess the carrion flesh feeding habits of the crested porcupine (Hystrix cristata). Scavenging behaviour was recorded on four occasions. All the recorded individuals were adults and at least one was female. This demonstrates that the crested porcupine occasionally does eat flesh. Such evidence raises important questions concerning the relationship between feeding habits and the physiological needs of this herbivorous rodent.

The crested porcupine is a mainly nocturnal mammal that shows both moonlight avoidance and some diurnal motor activity, the latter as an extension of its night-time foraging behaviour. Furthermore, a peculiar daytime behaviour, described as “sunbathing”, was reported as episodic in H. africaeaustralis. Between 2016 and 2019 a camera-trapping monitoring was performed within 10 porcupine settlements in order to detect the diurnal motor activity and to verify and describe the sunbathing behaviour in crested porcupine. Out of 1,003 trap days, a total of 148 events of diurnal motor activity were recorded. The diurnal motor activity occurred throughout the year mainly between December and June from 15:00 to 16:00, with no statistical difference between cubs, youngsters and adults. The sunbathing behaviour was detected for a total of 36 episodes recorded. Sunbathing was performed mainly by cubs. The sunbathing behaviour occurred only between April and June during the hottest hours of the day (11:00 to 12:00). Diurnal motor activity and sunbathing behaviour of porcupine are discussed in relation to food availability and porcupine physiology.

The continuous increasing of engineered nanomaterials (ENMs) in our environment, their combinatorial diversity, and the associated genotoxic risks, highlight the urgent need to better define the possible toxicological effects of ENMs. In this context, we present a new high-throughput screening (HTS) platform based on the cytokinesis-block micronucleus (CBMN) assay, lab-on-chip cell sorting, and automated image analysis. This HTS platform has been successfully applied to the evaluation of the cytotoxic and genotoxic effects of silver nanoparticles (AgNPs) and silica nanoparticles (SiO 2 NPs). In particular, our results demonstrate the high cyto- and genotoxicity induced by AgNPs and the biocompatibility of SiO 2 NPs, in primary human lymphocytes. Moreover, our data reveal that the toxic effects are also dependent on size, surface coating, and surface charge. Most importantly, our HTS platform shows that AgNP-induced genotoxicity is lymphocyte sub-type dependent and is particularly pronounced in CD2+ and CD4+ cells. Refereed/Peer-reviewed

We show that water soluble InP/ZnS core/shell QDs are a safer alternative to CdSe/ZnS QDs for biological applications, by comparing their toxicity in vitro (cell culture) and in vivo (animal model Drosophila). By choosing QDs with comparable physical and chemical properties, we find that cellular uptake and localization are practically identical for these two nanomaterials. Toxicity of CdSe/ZnS QDs appears to be related to the release of poisonous Cd(2+) ions and indeed we show that there is leaching of Cd(2+) ions from the particle core despite the two-layer ZnS shell. Since an almost identical amount of In(III) ions is observed to leach from the core of InP/ZnS QDs, their very low toxicity as revealed in this study hints at a much lower intrinsic toxicity of indium compared to cadmium.

Despite the extensive use of silica nanoparticles (SiO(2)NPs) in many fields, the results about their potential toxicity are still controversial. In this work, we have performed a systematic in vitro study to assess the biological impact of SiO(2)NPs, by investigating 3 different sizes (25, 60 and 115 nm) and 2 surface charges (positive and negative) of the nanoparticles in 5 cell lines (3 in adherence and 2 in suspension). We analyzed the cellular uptake and distribution of the NPs along with their possible effects on cell viability, membrane integrity and generation of reactive oxygen species (ROS). Experimental results show that all the investigated SiO(2)NPs do not induce detectable cytotoxic effects (up to 2.5 nM concentration) in all cell lines, and that cellular uptake is mediated by an endocytic process strongly dependent on the particle size and independent of its original surface charge, due to protein corona effects. Once having assessed the biocompatibility of SiO(2)NPs, we have evaluated their potential in gene delivery, showing their ability to silence specific protein expression. The results of this work indicate that monodisperse and stable SiO(2)NPs are not toxic, revealing their promising potential in various biomedical applications.

In this study, we show that 15 nm citrate-capped AuNPs exert a remarkable toxicity in living systems. The assessment was performed by using well-characterized AuNPs, the combination of in vitro and in vivo models (namely two different cell lines and Drosophila melanogaster), exposure to low dosages of nanoparticles (in the sub-nanomolar concentration range), along with the application of several biological assays to monitor different aspects of the toxic effects, such as viability, genotoxicity, and molecular biomarkers.

In this work we show the development and assessment of disposable, biocompatible, fully plastic microreactors, which are demonstrated to be highly efficient for performing on-chip direct PCR analyses of very diverse crude biological samples, such as human cells, whole blood, buccal scrape and hair bulbs, without requiring any purification steps before PCR analyses.

The growing use of nanomaterials in commercial goods and novel technologies is generating increasing questions about possible risks for human health and environment, due to the lack of an in-depth assessment of their potential toxicity. In this context, we investigated the effects of citrate-capped gold nanoparticles (AuNPs) on the model system Drosophila melanogaster upon ingestion. We observed a significant in vivo toxicity of AuNPs, which elicited clear adverse effects in treated organisms, such as a strong reduction of their life span and fertility, presence of DNA fragmentation, as well as a significant overexpression of the stress proteins. Transmission electron microscopy demonstrated the localization of the nanoparticles in tissues of Drosophila. The experimental evidence of high in vivo toxicity of a nanoscale material, which is widely considered to be safe and biocompatible in its bulk form, opens up important questions in many fields, including nanomedicine, material science, health, drug delivery and risk assessment.

In this article, we report the design and development of a plastic modular chip suitable for one-shot human papillomavirus (HPV) diagnostics, namely detection of the viral presence and relative genotyping, by two sequential steps performed directly on the same device. The device is composed of two modular and disposable plastic units that can be assembled or used separately. The first module is represented by a polydimethylsiloxane (PDMS) microreactor that is exploited for real-time polymerase chain reaction (PCR) and, thus, is suitable for detecting the presence of virus. The second unit is a PDMS microwell alpha-ray that allows virus genotyping by a colorimetric assay, based on DNA hybridization technology developed on plastic, requiring simple inspection by the naked eye. The two modules can be easily coupled to reusable hardware, enabling the heating/cooling processes and the real-time detection of HPV. By coupling real-time assay and colorimetric genotyping on the same chip, the assembled device may provide a low-cost tool for HPV diagnostics, thereby favoring the prediction of cancer risk in patients. (C) 2009 Elsevier Inc. All rights reserved.

The crested porcupine ( Hystrix cristata L., 1758) is the largest rodent among the Italian fauna. It is nocturnal, monogamous, herbivorous and lives in burrows that could be self made as well as made by other mammals such as fox ( Vulpes vulpes ) and badger ( Meles meles ). It exhibits moonlight avoidance and male performs parental care. In Europe, distribution of the species is limited to Italy, although the presence of porcupine in Balkans is still debated. Outside Europe crested porcupine distribution includes parts of northwestern Africa from Morocco to Libya and Sub-Saharan Africa from Gambia to Tanzania.

We show the possibility to fabricate highly controlled metal micropatterns on a variety of substrates, such as semiconducting or metallic materials, exploiting a combination of spontaneous galvanic displacement reactions with microfluidics. The process is reliable and quite versatile and allows the fabrication of complex patterns of different metals on a number of substrates in few minutes on a conventional laboratory bench.

We investigated the spatial structures of soil properties and snow-bed vegetation, and their relationships, in southern Italy. We analyzed data on 26 plant species and 10 soil traits from adjacent 1 × 1 m plots in two snow-bed patches. Measures of spatial autocorrelation revealed striking spatial structures for plant cover and soil properties at both sites. Bivariate statistics and Mantel tests highlighted a significant correlation between spatial patterns of plants and soil in the study sites. Canonical correspondence analysis related such relationships to an ecological gradient connecting soil properties and plant assemblages in this unusual ecological context. Among the variables significantly related to plant patterns is the soil organic matter, which is recognized as being sensitive to global warming. Our analyses suggest that soil dynamics due to increasing temperature may promote the replacement of species typical of southern snow-bed ecosystems by more mesophilous plants.

To obtain conformational data on the monomeric form of insulin, which is believed to be the physiologically active form of the hormone, insulin in sodium dodecyl sulfate solution was studied by Fourier-transform infrared (FTIR) spectroscopy and circular dichroism, and results were compared with those obtained with des(B23-30) octapeptide insulin (DOI) and dimeric insulin in buffer. The FTIR amide I band (1600-1700 cm-1) was examined, and a quantitative evaluation of the secondary structure fractions of the various conformations showed less of a beta-sheet component for both insulin in SDS and DOI in buffer than for insulin in buffer, corresponding to a lack of monomers binding to form dimers. At the concentrations used for FTIR (> or = 2 mg/mL), the CD spectra of insulin in SDS and DOI in buffer were qualitatively identical but different from that of insulin in buffer, which is associated at these concentrations. The CD spectrum pattern of insulin in very dilute solution (80 nM), where it is prevalently monomeric, is very similar to that of monomeric insulin in SDS, which suggests that the conformation of the hormone in the two cases is very similar.

The study assesses the influence of soil aspect on differences in soil chemical and biochemical properties. We examined soils on contiguous south- and north-facing slopes of the Aspromonte Mountains (Calabria, Southern Italy), influenced by the same climate, vegetation and parent material. In each of the two topographic aspects, six study sites were described. The investigated variables were air temperature, soil temperature, soil water content, photosynthetically active radiation, soil microbial biomass C, organic matter content, total nitrogen, total water-soluble phenols, humic and fulvic acids. Fluorescein diacetate hydrolytic activity, dehydrogenase, protease urease, alkaline and acid phosphatases, enzymes related to soil microbiological activity and hydrolysing coefficient, an empiric indicator of soil quality, were analyzed and interpreted. Except in few cases, all considered soil properties and microclimate variables showed significant differences between topographic aspects. In the soil on the north-facing slope, a lower content of organic matter and microorganisms and a lower activity of the enzymes related to soil microbiological activity were observed. The differences may be attributed to topographic aspect-induced microclimatic differences, which causing differences in the biotic soil component and organic matter trend, affect soil fertility.

Native glycosylated and enzymically deglycosylated conglutin γ (a lupin seed oligomeric protein) both showed an unusual resistance to tryptic degradation. The result of this treatment was that a single 40–residue peptide was cleaved from the N-terminus of conglutin γ light subunit. Acid treatment of the two protein forms led to their substantial unfolding, as indicated by CD spectra. After this treatment, both polypeptides were completely degraded by trypsin after a few minutes of incubation. Conversely, trypsin pulse experiments run under renaturing conditions demonstrated a different refolding behaviour of the two proteins: the glycosylated form became resistant to trypsin after a 7-h renaturation, while the deglycosylated form required 42 h renaturation. These results were confirmed by CD spectra and reverse-phase HPLC analyses of the glycosylated and deglycosylated conglutin γ forms. Therefore, it was concluded that the saccharide chain of conglutin γ increased the rate of formation of a trypsin-resistant conformation upon refolding of the acid-treated protein, without playing any direct role in the protection of the native protein from proteolysis.

Polydimethylsiloxane doped with colloidal CdSe/CdS nanocrystals is proposed as high-luminescent and biocompatible nanocomposite. A physical mixing procedure has been adopted to realize the optically active material and spectral and time-resolved optical analysis has been carried out in order to proof that nanocrystals quantum properties are preserved into the polymeric host matrix. Biocompatibility tests have shown no cadmium release in water ambient and no differences have been observed in cell viability cultured on pure PDMS substrates and on nanocomposite films. The obtained results let us envision possible applications of the proposed material as luminescent substrate for label-free cell imaging compatible with the most common optical microscopy techniques. (C) 2013 Elsevier B.V. All rights reserved.

We developed a new versatile strategy that allows the detection of several classes of RNases (i.e., targeting ss- or ds-RNA, DNA/RNA hetero-hybrid or junctions) with higher sensitivity than existing assays. Our two-step approach consists of a DNA-RNA-DNA chimeric Hairpin Probe (cHP) conjugated to magnetic microparticles and containing a DNAzyme sequence in its terminal region and molecular beacons for fluorescence signal generation. In the first step, the digestion of the RNA portion of the cHP sequences in presence of RNases leads to the release of multiple copies of the DNAzyme in solution. Then, after magnetic washing, each DNAzyme molecule elicits the catalytic cleavage of numerous molecular beacons, providing a strong amplification of the overall sensitivity of the assay. We successfully applied our approach to detect very low concentrations of RNase A, E. coli RNase I and RNase H. Furthermore, we analyzed the effect of two antibiotics (penicillin and streptomycin) on RNase H activity, demonstrating the applicability of our strategy for the screening of inhibitors. Finally, we exploited our system to detect RNase activity directly in crude biological samples (i.e., blood and saliva) and in cell culture medium, highlighting its suitability as cheap and sensitive tool for the detection of RNase levels.

Il Regolamento dell’Unione europea del 22 ottobre 2014, relativo allo statuto e al finanziamento dei partiti politici europei e delle fondazioni politiche europee, pone all’attenzione il problema della natura giuridica dei partiti politici. Al riguardo si possono individuare due ambiti distinti ma connessi di indagine. In primo luogo, quello del confronto fra il regime nazionale, alla luce della normativa italiana in materia, e il regolamento europeo. In secondo luogo, quello della comparazione fra i regimi nazionali di diritto privato in materia di partecipazione politica. Il saggio costituisce un primo tentativo di analisi nelle due direzioni indicate, e propone una lettura della normativa nazionale alla luce dei principi dell’Unione e delle esperienze di altri paesi membri dell’Unione

We show that Silicon Nitride (Si 3 N 4 ) photonic crystal (PhC) resonators are powerful building blocks to realize biocompatible photonic devices based on spontaneous emission engineering of nanoemitters in the visible spectral range. The versatility of this technological platform is demonstrated also in biological applications, where nanocavity modes are coupled to DNA strains marked with Cyanine 3 (Cy3) organic dyes and antibodies bounded to fluorescent proteins (TRITC).

We propose a technological approach aimed at improving biochips performances, based on an efficient spectral modeling and enhancement of markers fluorescence through the insertion of photonic crystal nanocavities (PhC-NCs) in the readout area of biochips. This strategy univocally associates a specific emission wavelength to a specific bioprobe immobilized on a nanocavity, therefore guaranteeing parallel detection of multiple elements and faster analysis time. Moreover, PhC-NCs significantly enhance the markers fluorescence, thus improving the detection sensitivity.

GM2 activator protein (GM2AP) is a cofactor for stimulating the enzymatic hydrolysis of the glycolipid GM2 by β-hexosaminidase A to produce GM3. We have examined the conformation of GM2AP before and after its interaction with GM2, GM3, and GA2 using circular dichroism and fluorescence spectroscopy techniques. In the presence of GM2, a blue shift of the fluorescence emission maximum and a strong decrease of molar ellipticity values in circular dichroism spectra were observed only at pH 4.5 and at GM2/GM2AP molar ratio higher than 10:1 (up to 50:1). These results suggest that GM2AP assumed a more organized α-helical conformation with the tryptophan residues moving from the polar medium toward the hydrophobic environment of the protein. The conformation of GM2AP in the presence of the downstream reaction product, GM3, or a less favorable substrate, GA2, clearly differed from that in the presence of GM2. The relationships between spectroscopic changes and enzymatic activity, herein discussed, strongly suggest that the specific conformation exhibited by GM2AP in the presence of GM2 is functional to serve as an activator for the enzymatic hydrolysis of GM2.

We demonstrate a general mechanism for the toxicity induced by metal-containing NPs, named “lysosome-enhanced Trojan horse effect”, which provides design rules to engineer safer NPs., The assessment of the risks exerted by nanoparticles is a key challenge for academic, industrial, and regulatory communities worldwide. Experimental evidence points towards significant toxicity for a range of nanoparticles both in vitro and in vivo. Worldwide efforts aim at uncovering the underlying mechanisms for this toxicity. Here, we show that the intracellular ion release elicited by the acidic conditions of the lysosomal cellular compartment – where particles are abundantly internalized – is responsible for the cascading events associated with nanoparticles-induced intracellular toxicity. We call this mechanism a “lysosome-enhanced Trojan horse effect” since, in the case of nanoparticles, the protective cellular machinery designed to degrade foreign objects is actually responsible for their toxicity. To test our hypothesis, we compare the toxicity of similar gold particles whose main difference is in the internalization pathways. We show that particles known to pass directly through cell membranes become more toxic when modified so as to be mostly internalized by endocytosis. Furthermore, using experiments with chelating and lysosomotropic agents, we found that the toxicity mechanism for different metal containing NPs (such as metallic, metal oxide, and semiconductor NPs) is mainly associated with the release of the corresponding toxic ions. Finally, we show that particles unable to release toxic ions (such as stably coated NPs, or diamond and silica NPs) are not harmful to intracellular environments.

Drosophila was the most important model organism used in the fields of medicine and biology over the last century. Recently, Drosophila was successfully used in several studies in the field of nanotoxicology. However, only a part of its potential has been exploited in this field until now. In fact, apart from macroscopic observations of the effect due to the interaction between nanomaterials and living organism (i.e. lifespan, fertility, phenotypic aberrations, etc.), Drosophila has the potential to be a very useful tool to deeply analyze the molecular pathways involved in response to the interactions at nano-bio level. The aim of this editorial is to encourage the use of Drosophila by the different research groups working in the fields of nanotoxicology and nanomedicine, in order to define the effects induced by nanomaterials at molecular level for their subsequent exploitation in the field of nanomedicine.

Chemical reconstitution of recombinant bovine adrenal mitochondrial apoadrenodoxin was carried out in the presence of the nonhomologous chaperone protein GroEL and of the cochaperone GroES, both in the presence and in the absence of ATP. The approach used here was different from the one characterizing studies on chaperone activity, as we used an adrenodoxin apoprotein, devoid of the cluster iron and sulfide, rather than a denaturant-unfolded form of the protein, and catalytic amounts of the chaperone proteins. A possible scaffolding role for two bacterial sulfurtransferases, namely, rhodanese from Azotobacter vinelandii and a rhodanese-like sulfurtransferase from Escherichia coli, was also investigated in the absence of the enzyme substrates. The extent and the rate of adrenodoxin refolding following cluster insertion was measured by spectroscopy and by monitoring the activity recovery in a NADPH–cytochrome c reduction assay. These measurements were carried out on the unresolved reaction mixture and on the adrenodoxin-containing fraction obtained by HPLC fractionation of the reconstitution mixture at different reaction times. The rate and extent of cluster insertion and activity recovery were substantially improved by addition of GroEL and increased with increasing the GroEL/apoadrenodoxin ratio. GroES and ATP had no effect by themselves, and did not enhance the effect of GroEL. A. vinelandii rhodanese, the E. coli sulfurtransferase, and bovine serum albumin had no effect on the rate and yield of chemical reconstitution. The accelerated chemical reconstitution of apoadrenoxin in the presence of GroEL is therefore attributable to a scaffolding effect of this protein.

Lipase from Pseudomonas cepacia was made soluble in 1,4-dioxane by lyophilization of the enzyme from aqueous solutions containing methoxypoly(ethylene glycol) (PEG). The solubility of the enzyme–PEG complex depended both on protein concentration and PEG protein ratio. Intrinsic protein fluorescence and far- and near-UV circular dichroism revealed that not only did the enzyme not unfold in the organic solvent, but rather became more compact. This was seen by the slight quenching of fluorescence intensity and by the enhancement of the near-UV circular dichroism negative signals, which are indicative of stronger interactions of tryptophanyl and/or tyrosyl residues among themselves or with other parts of the enzyme molecule. The specific activity of the lipase–PEG complex in the organic solvent was at least 2 orders of magnitude higher than that of the enzyme powder. This can be attributed both to the maintenance of native conformation and to enzyme dissolution in the reaction medium which should minimize possible limitations to enzyme–substrate interactions. © 1999 John Wiley & Sons, Inc., Biotechnol Bioeng 64: 624–629, 1999.