Inulin-Coated ZnO Nanoparticles: A Correlation between Preparation and Properties for Biostimulation Purposes.

Within the framework of plant biostimulation, a pivotal role is played by the achievement of low-cost, easily prepared nanoparticles for priming purposes. Therefore, in this report, two different synthetic strategies are described to engineer zinc oxide nanoparticles with an inulin coating. In both protocols, i.e., two-step and gel-like one-pot protocols, nanoparticles with a highly pure ZnO kernel are obtained when the reaction is carried out at T ≥ 40 °C, as ascertained by XRD and ATR/FTIR studies. However, a uniformly dispersed, highly homogeneous coating is achieved primarily when different temperatures, i.e., 60 °C and 40 °C, are employed in the two phases of the step-wise synthesis. In addition, a different binding mechanism, i.e., complexation, occurs in this case. When the gel-like process is employed, a high degree of coverage by the fructan is attained, leading to micrometric coated aggregates of nanometric particles, as revealed by SEM investigations. All NPs from the two-step synthesis feature electronic bandgaps in the 3.25-3.30 eV range in line with previous studies, whereas the extensive coating causes a remarkable 0.4 eV decrease in the bandgap. Overall, the global analysis of the investigations indicates that the samples synthesized at 60 °C and 40 °C are the best suited for biostimulation. Proof-of-principle assays upon Vicia faba seed priming with Zn5 and Zn5@inu indicated an effective growth stimulation of seedlings at doses of 100 mgKg-1, with concomitant Zn accumulation in the leaves.

Seed Priming with Zinc Oxide Nanoparticles to Enhance Crop Tolerance to Environmental Stresses.

Drastic climate changes over the years have triggered environmental challenges for wild plants and crops due to fluctuating weather patterns worldwide. This has caused different types of stressors, responsible for a decrease in plant life and biological productivity, with consequent food shortages, especially in areas under threat of desertification. Nanotechnology-based approaches have great potential in mitigating environmental stressors, thus fostering a sustainable agriculture. Zinc oxide nanoparticles (ZnO NPs) have demonstrated to be biostimulants as well as remedies to both environmental and biotic stresses. Their administration in the early sowing stages, i.e., seed priming, proved to be effective in improving germination rate, seedling and plant growth and in ameliorating the indicators of plants' well-being. Seed nano-priming acts through several mechanisms such as enhanced nutrients uptake, improved antioxidant properties, ROS accumulation and lipid peroxidation. The target for seed priming by ZnO NPs is mostly crops of large consumption or staple food, in order to meet the increased needs of a growing population and the net drop of global crop frequency, due to climate changes and soil contaminations. The current review focuses on the most recent low-cost, low-sized ZnO NPs employed for seed nano-priming, to alleviate abiotic and biotic stresses, mitigate the negative effects of improper storage and biostimulate plants' growth and well-being. Taking into account that there is large variability among ZnO NPs and that their chemico-physical properties may play a role in determining the efficacy of nano-priming, for all examined cases, it is reported whether the ZnO NPs are commercial or lab prepared. In the latter cases, the preparation conditions are described, along with structural and morphological characterizations. Under these premises, future perspectives and challenges are discussed in relation to structural properties and the possibility of ZnO NPs engineering.

Recent Advances in the Synthesis of Inorganic Materials Using Environmentally Friendly Media.

Deep Eutectic Solvents have gained a lot of attention in the last few years because of their vast applicability in a large number of technological processes, the simplicity of their preparation and their high biocompatibility and harmlessness. One of the fields where DES prove to be particularly valuable is the synthesis and modification of inorganic materials-in particular, nanoparticles. In this field, the inherent structural inhomogeneity of DES results in a marked templating effect, which has led to an increasing number of studies focusing on exploiting these new reaction media to prepare nanomaterials. This review aims to provide a summary of the numerous and most recent achievements made in this area, reporting several examples of the newest mixtures obtained by mixing molecules originating from natural feedstocks, as well as linking them to the more consolidated methods that use "classical" DES, such as reline.

An ELIME assay for hepatitis A virus detection.

An Enzyme Linked ImmunoMagnetic Electrochemical assay (ELIME) was developed for the detection of the hepatitis A virus (HAV). This system is based on the use of new polydopamine-modified magnetic nanobeads as solid support for the immunochemical chain, and an array of 8 screen-printed electrodes as a sensing platform. Enzymatic-by-product is quickly measured by differential pulse voltammetry. For this purpose, all analytical parameters were optimized; in particular, different blocking reagents were evaluated in order to minimize the nonspecific interaction of bioreagents. Using the ELIME assays, a quantitative determination of HAV can be achieved with a detection limit of 1·10-11 IU mL-1 and a working range between 10-10 - 5 × 10-7 IU mL-1. The cross-reactivity of the commercial monoclonal antibodies against HAV used in ELIME assays was tested for Coxsackie B4, resulting very low. The sensitivity was also investigated and compared with spectrophotometric sandwich ELISA. The average relative standard deviation (RSD) of the ELIME method was less than 5% for the assays performed on the same day, and 7% for the measurements made on different days. The proposed system was applied to the cell culture of HAV, which title was quantified by Real-Time Quantitative Reverse Transcription PCR (RT¬qPCR). To compare the results, a correlation between the units used in ELIME (IU mL-1) and those used in RT¬qPCR (genome mL-1) was established using a HAV-positive sample, resulting in 1 IU mL-1-10-4 gen mL-1 (R2 = 0.978). The ELIME tool exhibits good stability and high biological selectivity for HAV antigen detection and was successfully applied for the determination of HAV in tap water.

Molecular characterization of norovirus GII strains identified in Albania.

Noroviruses (NoVs) are considered as the leading cause of diarrheal diseases in all groups of age. In the last decade the number of NoV outbreaks worldwide is increasing. Data published by the systems of NoV surveillance show the GII.4 strain as the human predominant genotype circulating worldwide and new genetic variants of GII.4 were associated with epidemic events. In Albania the economy transformation has damaged significantly the environment and a large circulation of enteric viruses was reported in the past with the presence of NoV among the genotyped strains. This study aimed to characterize, by molecular analysis, the NoV GII strains detected in Albania during two time periods: in 2010 from the outbreak occurred in Ballsh and in 2002-2003 from sporadic cases of diarrhoea. A total of 21 Nov GII strains were characterized. The NoV GII.4 was genotyped more frequently and it was related closely to the pandemic variants recorded in GenBank. During 2002-2003, six NoV GII recombinant strains have been characterized.

Presence of hepatitis E RNA in mussels used as bio-monitors of viral marine pollution.

Mussels (Mytilus galloprovincialis), collected from a harvesting area approved by European Community Regulation, were transplanted to four polluted sites located in the Northwestern Mediterranean area (Tuscany). They were used as bio-monitors to test the quality of the marine water pollution. At different times after the transplantation, mussels were withdrawn and tested for presence of phages and enteric viruses by molecular tests. 52.4% of the transplanted mussel samples were positive for at least one enteric virus. Hepatitis A virus (HAV) was identified in each site (17/37; 45.9%). Three samples were positive for hepatitis E virus (HEV) (8.1%) and two (5.4%) for norovirus (NoV) genogroup I. Coliphages and RYC 2056 phages were detected in all sites, while HSP 40 phages were detected in three sites. Results demonstrate the ability of transplanted mussels in accumulating and retaining different species of enteric microorganisms. Their utility as bio-monitor organisms enables testing for viral marine pollution.

Analytical solution for the modeling of the natural time-dependent reduction of waterborne viruses injected into fractured aquifers.

We propose an analytical solution in order to explain the processes that determine the fate and behavior of the viruses during transport in a fractured aquifer at Salento (Italy). The calculations yield the efficiency of filtration in fractures at a site near Nardò (Southern Italy) in reducing the numbers of enteric viruses (i.e., Enteroviruses and Norovirus) in secondary municipal effluents that have been injected in the aquifer over the period 2006-2007. The model predicted, by a theoretical expression, the time-dependent rate of virus reduction, which was in good agreement with field data. The analytical solution yields the achievable "Log reduction credits" for virus reduction in wells located at the setback distances that are usually adopted in local drinking water regulations. The resulting new analytical formula for the time-dependent reduction of viruses during subsurface transport can easily be applied in health risk-based models used to forecast the spread of waterborne diseases and provides appropriate criteria (i.e., distances) needed to meet standards for the quality of drinking water derived from undisinfected groundwater.

A strategy to increase the specificity of Syber Green I qRT-PCR in hepatitis A detection.

Various methods for the recovery and detection of HAV have been suggested, and molecular tests have recently provided an effective replacement for the traditional methods. Real-time RT-PCR technology offers many advantages over conventional RT-PCR in terms of rapidity and specificity. Most procedures are based on the TaqMan chemistry, but some researchers have used the SYBR Green I approach, which is less expensive and simpler to carry out. However the formation of primer-dimers needs to be distinguished from specific products through a melting curve analysis. This study focused on a strategy to increase the specificity of Syber Green I chemistry, thus nullifying the primer-dimers interference. To this end, forward and reverse primers were specially designed for hairpin loop formation, a strategy widely used to improve the specificity and the efficiency of PCR. Two different concentrations of primers were assayed (200 nM and 400 nM) in a one-step, real-time RT-PCR procedure, evaluating the specificity of the amplicons and the optimization of the real-time protocol. We demonstrated that this approach can increase the specificity of the Syber Green I qRT-PCR performance with a good reproducibility of the method. Because of the simplicity of the assay and the lower costs involved, this procedure could be a valid alternative to HAV monitoring from environmental matrices.

Interaction between single wall carbon nanotubes and a human enteric virus.

Activated single wall carbon nanotubes have been used for biomedical purposes as carriers for drugs, peptides, proteins and nucleic acids. A large volume of data speaks to their suitability to act as a carrier. The ability of two differently activated SWNTs (with carboxyl groups or with carboxyl groups and polyethylenimine (PEI)) to form a complex with the hepatitis A virus was evaluated. Both types of activations permitted the formation of a virus-SWNT complex. However, their patterns were different. The carboxyl-activated nanotubes had a somewhat low adsorptive capacity that was related inversely to the concentrations of the SWNTs and viruses. Statistical analysis, using the chi(2)-test, showed no significant differences between the SWNT-PEI ratios of 1:2.5, 1:1 and 1:0.5. The addiction of PEI improved the adsorption, probably because of the electropositive charge of the molecule. Adsorption was optimal between 100 microg and 10 ng with a SWNTs-PEI weight ratio of 1:0.2 up to an inoculum of 10(5) genome equivalents of hepatitis A virus. Reducing or increasing this weight ratio reduced the adsorptive capacity of the PEI, and this adsorption activity was time and contact-dependent. Thus, SWNTs coated with PEI are able to complex with viruses, and they might be used in the future to transfect non-permissive cell lines.

Detection and molecular characterization of human rotaviruses isolated in Italy and Albania.

Rotaviruses are one of the most important causes of gastroenteritis in children under 5 years old. Analysis of G and P rotavirus genotypes in circulation is crucial in evaluating the appropriacy of mass vaccination of children worldwide. Overall, 592 stool samples were collected in Tirana (Albania), the Salento peninsula (South Italy), and three different hospitals in Rome (Central Italy). Of the total samples, 31.3% were rotavirus positive in Albania, 78.3% in the Salento, and 40.3% in Rome. The samples collected in Tirana and Rome were G-P typed, whereas the samples collected in the Salento were only G typed. Overall, in Italy the most frequent combinations were G4 P[8] (54.5%), G1 P[8] (27.3%), and G2 P[4] (18.2%); in Albania they were G9 P[8] (72.1%), G4 P[8] (8.8%), G1 P[8] (5.9%), and G2 P[4] (2.9%). The prevalence in Albania of atypical combinations was 7.4% for G4 P[4] and 2.9% for G9 P[4]. Phylogenetic analysis was also performed to assess the genetic relatedness of the strains. J. Med. Virol. 82:510-518, 2010. (c) 2010 Wiley-Liss, Inc.

[The birth of nanobiotechnologies: new nanomaterials, potential uses, toxic effects and implications for public health]. / La nascita delle nanobiotecnologie: I nuovi nanomateriali, i potenziali applicativi, gli effetti tossici e le implicazioni per la Sanità Pubblica.

Nanotechnologies hold considerable promise of advances in many sectors especially the biomedical field, since the materials used are of the appropriate dimensions to interact with important biological matter such as proteins, DNA and viruses. In this field the use of nanotechnologies will probably be second in importance only to biotechnologies. However many characteristics of nanomaterials that make them so promising from a technological point of view may also lead to negative effects on the environment and human health. It is important therefore that the environmental and work-related exposure effects to these materials be evaluated. In this article the potential uses, toxic effects and public health implications of nanobiotechnologies are discussed.

[Virus removal in conventional wastewater treatment process]. / Rimozione dei virus nei processi convenzionali di trattamento delle acque reflue.

Little has been published in the scientific literature on the distribution of pathogenic viruses and on their removal in the different treatment phases in wastewater treatment plants in Italy. Recent advances in molecular biology have reduced analytic costs and allowed the detection of virus directly in the environment. This study evaluates the presence of enteric viruses and their removal in three treatment plants of different sizes and which use different treatment methods. Overall 92 samples were collected during three seasonal campaigns (winter, spring-summer, and autumn). Samples were collected at different phases of the treatment process. Chemical and microbiological analyses were performed. Samples were concentrated by ultrafiltration and tested for the presence of hepatitis A virus, adenovirus, rotavirus, astrovirus, norovirus and enteroviruses. All viruses, excluding the Enteroviruses were detected through biomolecular testing. Enteroviruses, were isolated on BGM cells and subsequently genotyped at the VP1 region. Sixty-two of the 92 samples tested positive for viruses; of these, 21 tested positive for HAV, 4 for adenovirus, 9 for rotavirus, 21 for astrovirus, and 7 for norovirus. Enteroviruses were present on average in 44% of samples collected during the three sampling campaigns. Hepatitis A virus was isolated during the spring-summer period while Astroviruses were identified in all three sampling campaigns. Different enteroviruses were isolated, including four types of Coxsackie B virus variably isolated in all three campaigns, and type 1 Echovirus isolated in the second and third campaigns. All detected polioviruses were of the vaccine-type and were identified as types 1 and 3. Both viral genotypes of Norovirus were identified. Adenoviruses were only sporadically isolated and rotaviruses were identified as type A. As expected, results show a greater prevalence of Enteroviruses and this reflects viral circulation in the population. Regarding the efficacy of the different treatment stages, primary treatment was found to remove only small percentages of viruses, while activated sludge e subsequent sedimentation were found to be more effective.

Influence of enteric viruses on gastroenteritis in Albania: epidemiological and molecular analysis.

Gastroenteritis is one of the most important diseases in developing country and viral infections are well documented. To understand better the epidemiological aspect of gastroenteritis in Albania and especially viral gastroenteritis, one-year study was carried out with the cooperation of physicians working in the Paediatric Hospital in University Hospital Center "Mother Thereza" in Tirana. Three hundred thirteen stool samples were collected from children with diarrhoea and a questionnaire was filled by the health personnel for each child. Analysis of the questionnaires revealed that overcrowding families and the limited availability of drinking water at home were risk factors for gastroenteritis. All the tests for enteroviruses were carried out using the molecular methods. One hundred and forty-seven out of three hundred thirteen stool samples showed a specific amplification band for one of the enteric viruses: astrovirus, adenovirus, rotavirus, and norovirus with an overall positive specimen rate of 46.9%. Rotavirus was the most frequent virus identified in 105 out of 147 samples (71.4%), astrovirus in 5 (3.4%), norovirus in 19 (12.9%), and enteric adenovirus in 18 (12.3%) samples. Double infection was present only in 14 samples (9.5%). The data suggest an evident circulation of viruses involved in gastroenteritis with a higher prevalence of rotavirus.

Analysis of the shift of the transmission pattern for hepatitis C in a community in Central Italy.

Anti-HCV and HCV-RNA prevalence among adults from an isolated Central Italian community were 16.3% and 9.0%; 3.0%, 2.3% among 25-to-49-year-olds, 27.6%, 14.7% among older subjects. 1b genotype prevailed (88.5%). 1a, la/1b, 4 were also isolated. Seropositivity was associated with age > 50 years, public dental health care, glass syringe use, surgical interventions. The transmission pattern seems to be shifting from that typical of Southern Italy (high prevalence, particularly in the elderly, 1b genotype predominant, various routes of transmission involved, cohort effect), to that typical of Europe (lower prevalence, particularly among younger adults, other genotypes involved, association with intravenous drug use and immigrants).

Effect of extremely low frequency electromagnetic fields (ELF-EMF) on Kaposi's sarcoma-associated herpes virus in BCBL-1 cells.

Association between extremely low frequency electromagnetic fields (ELF-EMF) and human cancers is controversial, and few studies have been conducted on their influence on oncogenic viruses. We studied the effects of 1 mT, 50 Hz sine waves, applied for 24-72 h, on Kaposi's sarcoma (KS)-associated herpesvirus (KSHV or HHV-8) in BCBL-1, a latently infected primary effusion lymphoma (PEL) cell line. ELF-EMF exposure did not affect the growth and viability of BCBL-1 cells, either stimulated or not with TPA. The total amount of KSHV DNA detected in ELF-EMF exposed cultures not stimulated with TPA did not differ from that of the unexposed controls (P = ns). However, in the presence of TPA stimulation, total KSHV DNA content was found higher in ELF-EMF exposed than in control BCBL-1 cultures (P = .024) at 72 h exposure, but not earlier. Viral DNA increase significantly correlated with increased mean fluorescence intensity/cell for the lytic antigen gp K8.1A/B (P < .01), but not with percentage of gp K8.1A/B-positive cells or of cells containing virions. Viral progeny produced under ELF-EMF exposure consisted mainly of defective viral particles.

[Problems related to the use of real-time RT-PCR in environmental analysis]. / Problematiche connesse con l'applicazione della RT-PCR real-time nelle analisi ambientali.

Molecular biology techniques allow high sensitivity and specificity in the detection of enteric viruses in various environmental samples, and are considerably less costly and more rapid than traditional analytical methods. Real time RT-PCR technology allows accurate, efficient, and reproducible quantification of viral genes, by amplifying enteroviral RNA directly from an adequately treated environmental sample. It uses different chemical systems, including TaqMan and Syber Green probes, for detection of the amplificon. Both systems allow quantification of the initial number of copies in each cycle by comparing values with those of an external calibration curve (standard curve), generated by serial dilutions of a reference RNA sample with a known concentration. Difficulties in generating a standard curve for each enteric virus however, make standardization of the system time consuming. In an attempt to overcome this obstacle, we used an internal standard with a known concentration, to obtain a valid calibration curve for the quantification of environmental enteroviruses. A comparative analysis was performed with various commercially available extraction and amplification systems to evaluate the method's efficiency and reproducibility.

Detection and genotypic characterization of Giardia cysts in water samples.

Water samples were collected from a wastewater treatment plant and from two lakes in the vicinity of Rome (Italy) for detection and genotypic characterization of Giardia. All wastewater samples and all but two of the lake samples were scored by immunofluorescence as positive for Giardia. In addition, Giardia cysts were detected, by nested PCR, in all of the examined samples. Analysis of the SSU-rRNA gene sequences showed that all Giardia isolates belonged to Assemblage A. The presence of Assemblage A genotype in wastewater and surface bathing water represents an important public health concern. In fact, isolates showing this genotype are considered to be potential human pathogens and also have zoonotic implications.

[Compulsory and additional parameters in water control]. / Applicazione dei parametri obbligatori ed accessori nel controllo di acque di diversa origine.

The authors have analyzed 22 water samples from the Arno River (both river and drinking samples), as well as 11 effluent samples from an urban plant. Chemical and microbiological features of river samples were in line with the A3 class, needing treatment for drinking use (Executive Order n. 152/99); drinkable water met requirements (Presidents Decree n degrees 236/1988 and Executive Order n degrees 31/2001). As regards chemical parameters, effluents complied with the law (Executive Order n. 152/1999) but their bacteriological figures exceeded the standards set by the Authorities due to a lack in chlorination. Enteric viruses reacted to the RT-PCR test in 5 of the samples, including a Coxsackievirus Type B2 and a Poliovirus. No association was shown between bacteriophage parameters and virus.