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 At different times after the transplantation, mussels were withdrawn and tested for presence of phages and enteric viruses by molecular tests. 52.4% of the transplanted mussel samples were positive for at least one enteric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.

[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.

Enteric viruses in molluscan shellfish.

One hundred and thirty-seven bivalves were collected for environmental monitoring and the market; all the samples were analysed by RT-PCR test. Bacteriological counts meeting the European Union shellfish criteria were reached by 69.5% of all the samples, whereas the overall positive values for enteric virus presence were: 25.5%, 18.2%, 8.0% and 2.1% for Rotavirus, Astrovirus, Enteroviruses, Norovirus, respectively. Mussels appear to be the most contaminated bivalves, with 64.8% of positive samples, 55.7% and 22.7% respectively for clams and oysters, whereas in the bivalves collected for human consumption 50.7% were enteric virus positive, as compared to 56.4% of the samples collected for growing-area classification. The overall positive sample was 54.0%.

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.