Inactivation mechanisms of human adenovirus by e-beam irradiation in water environments.

This study aims to study the kinetics and mechanisms of human adenovirus inactivation by electron beam. Human adenovirus type 5 (HAdV-5) was inoculated in two types of aqueous substrates (phosphate-buffered saline - PBS, domestic wastewater - WW) treated by electron beam at a dose range between 3 and 21 kGy. Samples were evaluated for virus infectivity, PCR amplification of fragments of HAdV-5 genome and abundance and antigenicity of the virion structural proteins. The maximum reduction in viral titre, in plaque-forming units (PFU) per millilitre, was about 7 and 5 log PFU/mL for e-beam irradiation at 20 kGy in PBS and 19 kGy in wastewater, respectively. Among the virion structural proteins detected, the hexon protein showed the higher radioresistance. Long (10.1 kbp) genomic DNA fragments were differently PCR amplified, denoting a substrate effect on HAdV-5 genome degradation by e-beam. The differences observed between the two substrates can be explained by the protective effect that the organic matter present in the substrate may have on viral irradiation. According to the obtained results, the decrease in viral viability/infectivity may be due to DNA damage and to protein alterations. In summary, electron beam irradiation at a dose of 13 kGy is capable of reducing HAdV-5 viral titres by more than 99.99% (4 log PFU/mL) in both substrates assayed, indicating that this type of technology is effective for viral wastewater disinfection and may be used as a tertiary treatment in water treatment plants. KEY POINTS: • The substrate in which the virus is suspended has an impact on its sensitivity to e-beam treatment. • E-beam irradiation at 13 kGy is capable of reducing by 4 Log PFU/mL the HAdV-5 viral titre. • The decrease in viral viability/infectivity may be due to DNA damage and to protein alterations.

Is Gamma Radiation Suitable to Preserve Phenolic Compounds and to Decontaminate Mycotoxins in Aromatic Plants? A Case-Study with Aloysia citrodora Paláu.

This study aimed to determine the effect of gamma radiation on the preservation of phenolic compounds and on decontamination of dry herbs in terms of ochratoxin A (OTA) and aflatoxin B1 (AFB1), using Aloysia citrodora Paláu as a case study. For this purpose, artificially contaminated dry leaves were submitted to gamma radiation at different doses (1, 5, and 10 kGy; at dose rate of 1.7 kGy/h). Phenolic compounds were analysed by HPLC-DAD-ESI/MS and mycotoxin levels were determined by HPLC-fluorescence. Eleven phenolic compounds were identified in the samples and despite the apparent degradation of some compounds (namely verbasoside), 1 and 10 kGy doses point to a preservation of the majority of the compounds. The mean mycotoxin reduction varied between 5.3% and 9.6% for OTA and from 4.9% to 5.2% for AFB1. It was not observed a significant effect of the irradiation treatments on mycotoxin levels, and a slight degradation of the phenolic compounds in the irradiated samples was observed.

Tracking Human Adenovirus Inactivation by Gamma Radiation under Different Environmental Conditions.

UNLABELLED: Adenovirus is the most prevalent enteric virus in waters worldwide due to its environmental stability, which leads to public health concerns. Mitigation strategies are therefore required. The aim of this study was to assess the inactivation of human adenovirus type 5 (HAdV-5) by gamma radiation in aqueous environments. Various substrates with different organic loads, including domestic wastewater, were inoculated with HAdV-5 either individually or in a viral pool (with murine norovirus type 1 [MNV-1]) and were irradiated in a Cobalt-60 irradiator at several gamma radiation doses (0.9 to 10.8 kGy). The infectivity of viral particles, before and after irradiation, was tested by plaque assay using A549 cells. D10 values (dose required to inactivate 90% of a population or the dose of irradiation needed to produce a 1 log10 reduction in the population) were estimated for each substrate based on virus infectivity inactivation exponential kinetics. The capability of two detection methods, nested PCR and enzyme-linked immunosorbent assay (ELISA), to track inactivated viral particles was also assessed. After irradiation at 3.5 kGy, a reduction of the HAdV-5 titer of 4 log PFU/ml on substrates with lower organic loads was obtained, but in highly organic matrixes, the virus titer reduction was only 1 log PFU/ml. The D10 values of HAdV-5 in high organic substrates were significantly higher than in water suspensions. The obtained results point out some discrepancies between nested PCR, ELISA, and plaque assay on the assessments of HAdV-5 inactivation. These results suggest that the inactivation of HAdV-5 by gamma radiation, in aqueous environments, is significantly affected by substrate composition. This study highlights the virucidal potential of gamma radiation that may be used as a disinfection treatment for sustainable water supplies. IMPORTANCE: Human adenovirus (HAdV) is the most prevalent of the enteric viruses in environmental waters worldwide. The purposes of this study are to provide new insights on the inactivation of enteric virus by gamma irradiation and to introduce new concepts and reinforce the benefits and utility of radiation technologies as disinfection processes. This may be an effective tool to guarantee the reduction of viral pathogens and to contribute to public health and sustainable water supplies.

Effects of gamma radiation on wastewater microbiota.

Wastewater treatment by gamma radiation is a promising technology, with the capacity to reduce the impact of chemical and biological pollution of effluents in the environment. The aim of this study was to find out the effect of gamma radiation on the inactivation response of wastewater microorganisms. Wastewater samples were irradiated at a Co-60 facility, at different dose rates and at sublethal doses. The D10-values of total coliforms and mesophilic microbiota were determined for each sample and dose rate. Radio-resistant microorganisms in wastewater samples were isolated and their growth and inactivation kinetics in different composition substrates were determined, to find out the capacity of these bacteria to biodegrade the organic content of the wastewater. The results obtained suggest that irradiation substrate and dose rate influence the response of microorganisms to gamma radiation and could be also important factors for bioremediation.

Ga complexes of 8-hydroxyquinoline-2-carboxylic acid: Chemical speciation and biological activity.

The binding ability of 8-hydroxyquinoline-2-carboxylic acid (8-HQA) towards Ga3+ has been investigated by ISEH+ (Ion Selective Electrode, glass electrode) potentiometric and UV/Vis spectrophotometric titrations in KCl(aq) at I = 0.2 mol dm-3 and at T = 298.15 K. Further experiments were also performed adopting both the metal (with Fe3+ as competing cation) and ligand-competition approaches (with EDTA as competing ligand). Results gave evidence of the formation of the [Ga(8-HQA)]+, [Ga(8-HQA)(OH)], [Ga(8-HQA)(OH)2]- and [Ga(8-HQA)2]- species, the latter being so far the most stable, as also confirmed by ESI-MS analysis. Experiments were also designed to determine the stability constants of the [Ga(EDTA)]- and [Ga(EDTA)(OH)]2- in the above conditions. Due to the relevance of Ga3+ hydrolysis in aqueous systems, literature data on this topic were collected and critically analyzed, providing equations for the calculation of mononuclear Ga3+ hydrolysis constants at T = 298.15 K, in different ionic media, in the ionic strength range 0 < I / mol dm-3 ≤ 1.0. The synthesis and characterization (by ElectroSpray Ionization - Mass Spectrometry (ESI-MS), Attenuated Total Reflectance - Fourier-Transform Infrared Spectroscopy (ATR-FTIR) and ThermoGravimetric Analysis (TGA)) of Ga3+/8-HQA complexes were also performed, identifying [Ga(8-HQA)2]- as the main isolated species, even in the solid state. Finally, the potential effects of 8-HQA and Ga3+/8-HQA complex towards human microbiota exposed to ionizing radiation were evaluated (namely Actinomyces viscosus, Streptococcus mutans, Streptococcus sobrinus, Pseudomonas putida, Pseudomonas fluorescens and Escherichia coli), as well as their anti-proliferative and anti-inflammatory properties. A radioprotective effect of Ga3+/8-HQA complex was observed on Actinomyces viscosus, while showing a potential radiosensitizing effect against Streptococcus mutans and Streptococcus sobrinus. No cytotoxicity on RAW264.7 murine macrophage cells was observed, neither for the free ligand or Ga3+/8-HQA complex. Nevertheless, Ga3+/8-HQA complex highlighted potential anti-inflammatory properties.

Effects of Electron Beam Radiation on the Phenolic Composition and Bioactive Properties of Olive Pomace Extracts.

Olive pomace is an agro-industrial waste product generated from the olive oil industry and constituted by bioactive compounds with potential applications in several industrial sectors. The purpose of this work was to evaluate the effects of electron beam (e-beam) radiation on olive pomace, specifically on phenolic compounds (by HPLC-DAD-ESI/MS) and the bioactive properties (antioxidant, antiproliferative, and antimicrobial activities) of crude olive pomace (COP) and extracted olive pomace (EOP) extracts. The amount of total flavonoid content and the reducing power of COP extracts were higher than those obtained for EOP extracts. The results suggested that e-beam radiation at 6 kGy increased both total phenolic and total flavonoid contents as well as the reducing power of COP extracts, due to the higher extractability (>2.5-fold) of phenolic compounds from these samples, while decreasing the scavenging activity of extracts. The extracts of both olive pomaces showed antibacterial potential, and COP extracts at 400 µg/mL also presented antiproliferative activity against A549, Caco-2, 293T, and RAW264.7 cell lines, with both properties preserved with the e-beam treatment. All in all, e-beam radiation at 6 kGy appears to be a promising technology to valorize the pollutant wastes of the olive oil industry through enhancing phenolic extractability and bioactive properties, and, furthermore, to contribute to the environmental and economical sustainability of the olive oil industry.

Effect of Olive Pomace Extract Application and Packaging Material on the Preservation of Fresh-Cut Royal Gala Apples.

The efficiency of natural olive pomace extracts for enhancing the quality of fresh-cut apples was compared with commercial ascorbic acid and two different packaging films (biodegradable polylactic acid (PLA) and oriented polypropylene (OPP)) were tested. The composition of atmosphere inside the packages, the physicochemical parameters (firmness, weight loss and color), the microbial load, total phenolic content and antioxidant activity of fresh-cut apples were evaluated throughout 12 days of storage at 4 °C. After 12 days of refrigerated storage, a significant decrease in O2 was promoted in PLA films, and the weight loss of the whole packaging was higher in PLA films (5.4%) than in OPP films (0.2%). Natural olive pomace extracts reduced the load of mesophilic bacteria (3.4 ± 0.1 log CFU/g and 2.4 ± 0.1 log CFU/g for OPP and PLA films, respectively) and filamentous fungi (3.3 ± 0.1 log CFU/g and 2.44 ± 0.05 log CFU/g for OPP and PLA films, respectively) growth in fresh-cut apples after five days of storage at 4 °C, and no detection of coliforms was verified throughout the 12 days of storage. In general, the olive pomace extract preserved or improved the total phenolic index and antioxidant potential of the fruit, without significant changes in their firmness. Moreover, this extract seemed to be more effective when combined with the biodegradable PLA film packaging. This work can contribute to the availability of effective natural food additives, the sustainability of the olive oil industries and the reduction of environmental impact. It can also be useful in meeting the food industries requirements to develop new functional food products.

Microbiota Assessment of Fresh-Cut Apples Packaged in Two Different Films.

The aim of this work was to assess the natural microbiota of packed fresh-cut apples during refrigerated storage. Two different films were tested for the package, a biodegradable (PLA) film and a conventional and commercial one (OPP). Two antioxidant additives were applied, a natural olive pomace extract and the commercial ascorbic acid used by the industries. The results revealed lower bacteria counts in samples with olive pomace extract and PLA films than in those with ascorbic acid and OPP films after 5 and 12 days of storage. These findings suggest that the use of such natural extracts as additives in fruits could delay the growth of mesophilic bacteria. The characterization and identification of the bacterial isolates from fresh-cut apple samples showed that the most prevalent species were Citrobacter freundii, Staphylococcus warneri, Pseudomonas oryzihabitans, Alcalinogenes faecalis, Corynebacterium jeikeium, Micrococcus spp., Pantoea aglomerans and Bacillus spp. Furthermore, an increase in the microbial diversity during the storage time at refrigerated temperatures was observed, except for the sample treated with olive pomace extract and packaged in OPP film. The highest microbial diversity was found for samples with ascorbic acid as an additive. This could indicate a negative effect of ascorbic acid on the microbial inhibition of apple slices. The natural olive pomace extract demonstrated potential as an antimicrobial additive for fresh-cut apples.

Ultrasound-assisted extraction of hydroxytyrosol and tyrosol from olive pomace treated by gamma radiation: process optimization and bioactivity assessment.

Ultrasound-assisted extraction (UAE) was used to recover hydroxytyrosol and tyrosol from olive pomace, a residue generated by the olive oil industry. The extraction process was optimized using response surface methodology (RSM), with processing time, ethanol concentration and ultrasonic power as the combined independent variables. The highest amounts of hydroxytyrosol (36 ± 2 mg g-1 of extract) and tyrosol (14 ± 1 mg g-1 of extract) were obtained after 28 min of sonication at 490 W using 7.3% ethanol as the solvent. Under these global conditions, an extraction yield of 30 ± 2% was achieved. The bioactivity of the extract obtained under optimized UAE was evaluated and compared with that of an extract obtained under optimal heat-assisted extraction (HAE) conditions in a previous work of the authors. Compared to HAE, UAE reduced the extraction time and the solvent consumption, and also led to higher extraction yields (HAE yield was 13.7%). Despite this, HAE extract presented higher antioxidant, antidiabetic, anti-inflammatory and antibacterial activities and no antifungal potential against C. albicans. Furthermore, HAE extract also showed higher cytotoxic effects against the breast adenocarcinoma (MCF-7) cell line. These findings provide useful information for the food and pharmaceutical industries in developing new bioactive ingredients, which may represent a sustainable alternative to synthetic preservatives and/or additives.

Construction of HER2-Specific HIV-1-Based VLPs.

Virus-like particles (VLPs) are nanoplatforms comprised of one or more viral proteins with the capacity to self-assemble without viral genetic material. VLPs arise as promising nanoparticles (NPs) that can be exploited as vaccines, as drug delivery vehicles or as carriers of imaging agents. Engineered antibody constructs, namely single-chain variable fragments (scFv), have been explored as relevant molecules to direct NPs to their target. A vector containing the scFv of an antibody, aimed at the human epidermal growth factor receptor 2 (HER2) and fused to the human immunodeficiency virus (HIV) protein gp41, was previously constructed. The work herein describes the early results concerning the production and the characterization of HIV-1-based VLPs expressing this protein, which could function as potential non-toxic tools for transporting drugs and/or imaging agents.

How promising are HIV-1-based virus-like particles for medical applications.

New approaches aimed at identifying patient-specific drug targets and addressing unmet clinical needs in the framework of precision medicine are a strong motivation for researchers worldwide. As scientists learn more about proteins that drive known diseases, they are better able to design promising therapeutic approaches to target those proteins. The field of nanotechnology has been extensively explored in the past years, and nanoparticles (NPs) have emerged as promising systems for target-specific delivery of drugs. Virus-like particles (VLPs) arise as auspicious NPs due to their intrinsic properties. The lack of viral genetic material and the inability to replicate, together with tropism conservation and antigenicity characteristic of the native virus prompted extensive interest in their use as vaccines or as delivery systems for therapeutic and/or imaging agents. Owing to its simplicity and non-complex structure, one of the viruses currently under study for the construction of VLPs is the human immunodeficiency virus type 1 (HIV-1). Typically, HIV-1-based VLPs are used for antibody discovery, vaccines, diagnostic reagent development and protein-based assays. This review will be centered on the use of HIV-1-based VLPs and their potential biomedical applications.

Target specific post-harvest treatment by gamma radiation for the microbial safety of dried Melissa officinalis and Aloysia citrodora.

This study aimed to assess a specific gamma radiation dose to be applied as a post-harvest process to guarantee the microbial safety of two medicinal plants, Melissa officinalis and Aloysia citrodora. Dried plants treated with gamma radiation indicated that a dose of 5 kGy could be applied as a post-harvest treatment process of M. officinalis and A. citrodora, assuring the microbial safety of dried medicinal plants and lowering the potentiality of deleterious effects on plants' quality attributes. This will enhance the safety and quality of the dried plants to be used as raw materials in industrial applications.

Combined effects of irradiation and storage time on the nutritional and chemical parameters of dried Agaricus bisporus Portobello mushroom flour.

Portobello variety of Agaricus bisporus mushrooms, appreciated for its taste, makes it desirable to be eaten fresh and also as flour in soups and gravies. Gamma and electron-beam radiation at four doses (1, 2, 5, and 10 kGy) were used to analyze its preservation effect on Portobello mushroom flour. A proximate analysis, as well as the impact on fatty acids, tocopherols, soluble sugars, organic acids, and ergosterol profiles, were performed every 3 months, during a storage period of 1 year. Gamma rays preserved mannitol (most abundant soluble sugar) over the 12 months, while electron beam radiation preserved organic acids. No significant changes were sought for any radiation type, and the slight changes extracted from the estimated marginal means reveal a tendency for irradiation as having preserving effects of nutrients and other important molecules. Thus, both irradiation types, up to 10 kGy are suitable for preservation of A. bisporus Portobello flour.

Ionizing Radiation Technologies to Increase the Extraction of Bioactive Compounds from Agro-Industrial Residues: A Review.

Due to the growing demand in society for healthier foods, scientific communities are searching and developing new ingredients. In this context, agro-industrial residues, which can have a negative impact on the environment, represent a natural source for bioactive compounds and their recovery can contribute to economic and environmental sustainability. Ionizing radiation is a clean and eco-friendly technology that can be used to improve the extraction of bioactive compounds. The aim of this review, after presenting general aspects about bioactive compounds in agro-industrial residues and radiation technologies, is to focus on the effects of ionizing radiation on the extraction of bioactive compounds from these residues and related bioactive properties. Irradiated residues were demonstrated to have enhanced bioactive characteristics that turn the prepared extracts suitable for applications in food industry, resulting in high-added-value products as well as reducing adverse impacts on the environment.

Effect of Gamma-Radiation on Zearalenone-Degradation, Cytotoxicity and Estrogenicity.

Zearalenone (ZEA) is produced in cereals by different species of Fusarium, being a non-steroidal estrogenic mycotoxin. Despite having a low acute toxicity, ZEA strongly interferes with estrogen receptors. Gamma-radiation has been investigated to eliminate mycotoxins from food and feed, showing promising results. The present study aims to investigate the gamma-radiation effect on ZEA at different moisture conditions and to evaluate the cytotoxicity and estrogenicity of the irradiated ZEA. Different concentrations of dehydrated ZEA and aqueous solutions of ZEA were exposed to gamma-radiation doses ranging from 0.4 to 8.6 kGy and the mycotoxin concentration determined after exposure by high performance liquid chromatography (HPLC) with fluorescence detection. Following this, the cytotoxicity of irradiated samples was assessed in HepG2 cells, by measuring alterations of metabolic activity, plasma membrane integrity and lysosomal function, and their estrogenicity by measuring luciferase activity in HeLa 9903 cells. Gamma-radiation was found to be effective in reducing ZEA, with significant increases in degradation with increased moisture content. Furthermore, a reduction of cytotoxicity with irradiation was observed. ZEA estrogenicity was also increasingly reduced with increasing radiation doses, but mainly in aqueous solutions. These results suggest reduction of ZEA levels and of its toxicity in food and feed commodities may be achieved by irradiation.

The use of gamma radiation for extractability improvement of bioactive compounds in olive oil wastes.

Olive pomace is an environmentally detrimental waste from olive oil industry, containing large amounts of bioactive compounds that might be used by the food industry. In this work, the effects of gamma radiation on phenolic compounds and bioactive properties (antioxidant, antimicrobial activities and hepatotoxicity) of Crude Olive Pomace (COP) and Extracted Olive Pomace (EOP) extracts were evaluated. Hydroxytyrosol was the main phenolic compound identified in both olive pomace extracts (24-25 mg/g). The gamma radiation treatment of olive pomace improved at least 2-fold the extractability of phenolic compounds. Moreover, results suggested that gamma radiation at 5 kGy increased the antioxidant activity in EOP, while keeping the ability to protect erythrocytes against oxidation-induced haemolysis. Gamma radiation at 5 kGy could be a suitable technology for olive oil pomaces waste valorization, contributing to enhance extraction of phenolic compounds and bioactive properties, especially when applied on extracted material.

Compliance of indoor air quality during sleep with legislation and guidelines - A case study of Lisbon dwellings.

This study aimed to provide a comprehensive characterisation of the indoor air quality during the sleeping period of 10 couples at Lisbon dwellings, using a multi-pollutant approach, and to understand how the compliance with legislation and guidelines was to assure a good indoor air quality. The assessment of indoor air quality was conducted in the cold season using real time monitors during the sleeping period for comfort parameters (temperature and relative humidity) and air pollutants (carbon dioxide - CO2, carbon monoxide - CO, formaldehyde - CH2O, total volatile organic compounds - VOCs, and particulate matter - PM2.5 and PM10), together with active sampling of bioaerosols (fungi and bacteria) before and after the sleeping period. Lower compliance (less than 50% of the cases) with the Portuguese legislation was found for temperature, CO2 (3440 ± 1610 mg m-3), VOCs (1.79 ± 0.99 mg m-3) and both bioaerosol types. In 70% of the cases, PM2.5 (15.3 ± 9.1 µg m-3) exceeded the WHO guideline of 10 µg m-3. All bedrooms presented air change rates above the recommended minimum value of 0.7 h-1, highlighting that a good indoor air quality during sleep is not guaranteed.

Virucidal activity of gamma radiation on strawberries and raspberries.

The environmental stability of enteric viruses and resistance to conventional treatments and common disinfectants, leads to their persistence in waters and food, causing serious implications on public health. Among non-thermal treatment methods, ionizing radiation is recognized as a useful and effective mean of disinfection. The objective of this study was to estimate the inactivation of enteric virus by gamma radiation in raw berry fruits, in order to evaluate the potential of this technology to be applied as a disinfection treatment. Fresh strawberries and raspberries were inoculated either individually with murine norovirus type 1 (MuNoV; as a human norovirus surrogate) and human adenovirus type 5 (HAdV) or with a viral pool of both viruses, and irradiated in a Co-60 equipment at doses of 1 kGy up to 11 kGy. The infectivity of viral particles of MuNoV and HAdV was assessed by plaque assay using Raw 264.7 and A549 cells, respectively. A 2 log PFU/g reduction on MuNoV and HAdV titers was obtained after treatment with a dose of 4 kGy for both fruits. However, non-linear inactivation survival curves were obtained for MuNoV and HAdV in fresh fruits, leading to the detection of infective viral particles at a dose of 11 kGy. The irradiation process indicated virucidal potential, although the estimated gamma radiation dose to attain food safety (> 7 kGy) would compromise the preservation of food quality. Nevertheless, the irradiation technology could be an effective virus mitigation tool to treat polluted waters, which are a major vehicle of contamination for fresh produce.

Stability of total folates/vitamin B9 in irradiated watercress and buckler sorrel during refrigerated storage.

The suitability of post-packaging gamma radiation treatment for preserving total folates or vitamin B9 in watercress (Nasturtium officinale R. Br.) and buckler sorrel (Rumex induratus Boiss. & Reut.) during storage at 4 °C was evaluated. Comparable amounts of total folates were found in fresh, non-stored samples of both species. In watercress, the irradiation treatment of up to 5 kGy reduced the loss of total folates caused by 7 days of storage. In turn, the 12-day storage period did not affect the total folate content of buckler sorrel (while the 2 kGy dose decreased the initial levels), evidencing that packaging and refrigeration are enough for preservation. These results suggest that the suitability of post-packaging irradiation for preserving total folates may depend not only on the applied dose but also on the plant matrix under analysis. In addition, new data useful to complete food composition tables or databases is provided.

Gamma irradiation effects on ochratoxin A: Degradation, cytotoxicity and application in food.

Ochratoxin A (OTA) is one of the main mycotoxins that can be found in food. The use of gamma radiation is a technique for preserving food that may exert some effects on mycotoxins. OTA was irradiated in its dry form, in aqueous and in methanolic solutions, and in wheat flour, grape juice and wine. Additionally, the toxicity of OTA irradiated in water was tested. In aqueous solutions, more than 90% of the OTA was degraded by γ-radiation doses ≥2.5kGy, and a 2-fold reduction in OTA cytotoxicity was observed. In food matrices, the elimination of OTA by γ-radiation was found more difficult, as radiation doses of 30kGy eliminate at most 24% of the OTA. Higher moisture content of food matrices did not substantially increase OTA elimination. It is concluded that OTA is very sensitive to irradiation in water solutions but resistant in its dry form and in food matrices.