Hygienic assessment of digestate from a high solids anaerobic co-digestion of sewage sludge with biowaste by testing Salmonella Typhimurium, Escherichia coli and SARS-CoV-2.

Anaerobic digestion is a consolidated technology to convert sewage sludge and other organic wastes into biogas and a nutrient-rich fertilizer (i.e. digestate). The origin of sewage sludge does not exclude the potential presence of pathogens (e.g. Salmonella spp. and SARS-CoV-2) in mature digestate that hence could represent a source of sanitary concerns when it is spread on soil for agriculture purpose. Therefore, an experimental study aimed at proving the sanitizing effect of a full scale thermophilic high solids anaerobic digestion process was conducted by monitoring the hygienic characteristics of mature digestate. Although Salmonella spp. was detected in the sewage sludge fed to the full scale plant, the anaerobic digestion treatment demonstrated sanitization capacity since the monitored pathogens were never found in the mature digestate over the entire duration of the monitoring survey. Furthermore, tests on the regrowth of Salmonella Typhimurium and Escherichia coli, artificially inoculated on mature digestate, were also conducted under both anaerobic and aerobic conditions with the aim to assess the effectiveness of mature digestate as microbial growth medium. Concentrations of Salmonella Typhimurium and Escherichia coli were drastically reduced after a short time of incubation under anaerobic process and the two microorganisms already resulted undetectable after 24-48 h, whereas, under aerobic conditions, two microorganisms' concentrations were stably high for longer than 10 days. The combination of no free oxygen, high temperature, anaerobic metabolites (e.g. total ammonium nitrogen, and volatile fatty acids) production, bacteria competition and lack of nutritional elements in mature digestate considerably reduced in 24-48 h the sanitary risks associated to accidently contaminated digestate. Furthermore, a SARS-CoV-2 monitoring survey on mature digestate during 13 months, resulted in the absence of the virus RNA in the analyzed digestate.

Metabolomic profiling of food matrices: Preliminary identification of potential markers of microbial contamination.

The research aimed to generate an early warning system highlighting in real-time bacterial contamination of meat matrices and providing information which could support companies in accepting or rejecting batches. Current microorganisms' detection methods rely on techniques (plate counting), which provide retrospective values for microbial contamination. The purpose of this research was to evaluate the ability of the headspace solid-phase microextraction (HS-SPME) and gas chromatography-mass spectrometry (GC/MS) methodologies to detect volatile organic carbons (VOCs), which may be associated to a peculiar microbiological contamination of food. The disposal of fast headspace gas chromatography-mass spectrometry (HS-SPME-GC/MS) able to accurately and rapidly (30 min per sample) detect pathogens in raw meat could replace the traditional and time-consuming (3 to 4 days) standardized microbiological analysis required by regulations. Experiments focused on qualitative and quantitative evaluations of VOCs produced by Salmonella Typhimurium, Campylobacter jejuni, and Staphylococcus aureus in different types of raw meat (beef, pork, chicken). HS-SPME-GC/MS allowed to use smaller sample volumes compared to traditional methods with no sample processing and the potentiality for its application on various food matrices for the detection of a wide variety of pathogens. Data analysis showed the identification of unique VOCs' profiles being possible markers of meat contamination due to their association to specific pathogens. The identification of VOCs markers in association to selected bacterial pathogens and their metabolites could support the rapid determination of specific meat samples contamination. Further research is required to outline-specific metabolic profiles for each microorganism responsible of meat contamination and prevent false positives.

Integrated characterization and risk management of marine sediments: The case study of the industrialized Bagnoli area (Naples, Italy).

The aim of the present work is to demonstrate the practical importance of a multidisciplinary approach and weighted criteria to synthesize and integrate different typologies of data (or lines of evidence, LOEs), including chemical levels in marine sediments, their bioavailability to specific indicator species, ecotoxicological effects measured through subcellular biomarkers and batteries of bioassays, and potential impacts of pollution on local benthic communities. The area of Bagnoli (Gulf of Naples, Southern Italy) was selected as a model case-study, as it is a coastal area chronically impacted by massive industrial contamination (trace metals and hydrocarbons), and dismissed decades ago without any subsequent remediation or habitat restoration. The results of each LOE were elaborated to provide specific hazard indices before their overall integration in a weight of evidence (WOE) evaluation. Levels of some trace metals and PAHs revealed a severe contamination in the entire study area. Bioavailability of hydrocarbons was evident particularly for high molecular weight PAHs, which also caused significant variations of cellular biomarkers, such as cytochrome P450 metabolization in fish, lysosomal membrane destabilization in mussels, genotoxic effects both in fish and molluscs. The results of a battery of bioassays indicated less marked responses compared to those obtained from chemical and biomarkers analyses, with acute toxicity still present in sediments close to the source of contamination. The analysis of benthic assemblages showed limited evidence of impact in the whole area, indicating a good functioning of local ecosystems at chronic contamination. Overall, the results of this study confirm the need of combining chemical and biological data, the quantitative characterization of various typologies of hazard and the importance of assessing an integrated environmental WOE risk, to orientate specific and scientifically-supported management options in industrialized areas.

A comparative assessment of metals and phthalates in commercial tea infusions: A starting point to evaluate their tolerance limits.

Tea is one of the most consumed beverages in the word. Here we report the concentrations of metals and phthalates in 32 commercial tea packages. The data were used to estimate the average daily intake of metals and phthalates, and associated Hazard Quotients (HQ) were calculated in order to determine risk of non-cancerous health effects for adults consuming tea on a daily basis. Tea samples were chosen based on the sales network, the price, the marketing quality and the presence of filters in the packages. Relatively high median concentrations of Al (5240 µg/L), Ni (44 µg/L), and Mn (2919 µg/L) were detected. No metals or phthalates quantified in the tea infusions and soluble tea showed an HQ greater than 1, indicating no risk of non-cancerous health effects. The data presented herein may serve as a starting point to evaluate tolerance limits of metals and phthalate in the tea infusion.

Protamine-like proteins have bactericidal activity. The first evidence in Mytilus galloprovincialis.

The major acid-soluble protein components of the mussel Mytilus galloprovincialis sperm chromatin consist of the protamine-like proteins PL-II, PL-III and PL-IV, an intermediate group of sperm nuclear basic proteins between histones and protamines. The aim of this study was to investigate the bactericidal activity of these proteins since, to date, there are reports on bactericidal activity of protamines and histones, but not on protamine-like proteins. We tested the bactericidal activity of these proteins against Gram-positive bacteria: Enterococcus faecalis and two different strains of Staphylococcus aureus, as well as Gram-negative bacteria: Proteus mirabilis, Proteus vulgaris, Pseudomonas aeruginosa, Salmonella typhmurium, Enterobacter aerogenes, Enterobacter cloacae, and Escherichia coli. Clinical isolates of the same bacterial species were also used to compare their sensitivity to these proteins. The results show that Mytilus galloprovincialis protamine-like proteins exhibited bactericidal activity against all bacterial strains tested with different minimum bactericidal concentration values, ranging from 15.7 to 250 µg/mL. Furthermore, these proteins were active against some bacterial strains tested that are resistant to conventional antibiotics. These proteins showed very low toxicity as judged by red blood cell lysis and viability MTT assays and seem to act both at the membrane level and within the bacterial cell. We also tested the bactericidal activity of the product obtained from an in vitro model of gastrointestinal digestion of protamine-like proteins on a Gram-positive and a Gram-negative strain, and obtained the same results with respect to undigested protamine-like proteins on the Gram-positive bacterium. These results provide the first evidence of bactericidal activity of protamine-like-proteins.

Comparative assessment of the quality of commercial black and green tea using microbiology analyses.

BACKGROUND: Drinking tea constitutes a tradition which is deeply rooted in the culture of several countries. Moreover, in recent years, tea consumption is growing all over the world. Improper herbal tea storage (long periods, humid environments) represents a relevant health hazard for consumers because of the growth of bacteria and molds. RESULTS: This study analyzed 32 samples of commercially available black and green teas - purchased from southern Italy markets and online-shops - and the monitoring of microbiological quality of the tea bag content was performed. Evaluations were conducted with the aim of characterizing pathogens indicated by the European and American guidelines (total bacterial count, fungi and Escherichia coli) and on the research of Pseudomonas spp. and Clostridium perfringens. The presence of ochratoxin A in tea matrix-leaves and infusions was further assessed, using a validated and accredited HPLC-FLD method. Microbial loads, for over 80% samples, ranged from 1.0 × 102 to 2.8 × 105 CFU/g tea: most of identified microorganisms were classified as Bacillaceae. The utilization of rapid detection and identification methods (PCR and sequencing), allowed the characterization of strains of Pseudomonas psychrotolerans, Staphylococcus warneri, Pantoea gaviniae and the isolation of one strain of Clostridium perfringens, whose ability to produce toxins can result in harmful outcomes for consumers. Fungi were isolated from 70% samples: the most prevalent molds were Aspergillus niger strains, followed by Aspergillus tubingensis. Ochratoxin A was detected in 22 of 32 tea solid samples investigated: concentrations resulted over the indicated limits for food products for 50% samples. CONCLUSIONS: Results obtained demonstrated the need to develop targeted regulations for the safety of herbal teas.

Correction: What is in your cup of tea? DNA Verity Test to characterize black and green commercial teas.

[This corrects the article DOI: 10.1371/journal.pone.0178262.].

Toxicity assessment within the application of in situ contaminated sediment remediation technologies: A review.

Polluted sediment represents a great problem for aquantic environments with potential direct acute and chronic effects for the biota and can be tackled with both in situ and ex situ treatments. Once dredging activities are not compulsory, sediment can be kept in place and managed with techniques involving the use of amendment and/or capping. Before their application, the assessment of their potential impact to the target environment cannot ignore the safe-by-design approach. The role of toxicity in in situ sediment remediation was reviewed discussing about how it can be used for the selection of amendments and the monitoring of treatment technologies. Results evidenced that capping technology coupled to activated carbon (AC) is the most frequently applied approach with effects varying according to the rate of contamination in treated sediment, the amount of AC used (% v/v), and target biological models considered. Little data are available for zerovalent iron as well as other minor amending agents such as hematite, natural zeolite, biopolymers and organoclays. Current (eco-)toxicological information for in situ sediment remediation technologies is fragmentary and incomplete or entirely missing, making also the interpretation of existing data quite challenging. In situ sediment remediation represents an interesting potentially effective approach for polluted sediment recovering. As its application in some lab-based and field studies reported to induce negative effects for target organisms, amendments and capping agents must be attentively evaluated for short- and long-term environmental effects, also in the perspective of the remediated site monitoring and maintenance.

What is in your cup of tea? DNA Verity Test to characterize black and green commercial teas.

In this study, we used several molecular techniques to develop a fast and reliable protocol (DNA Verity Test, DVT) for the characterization and confirmation of the species or taxa present in herbal infusions. As a model plant for this protocol, Camellia sinensis, a traditional tea plant, was selected due to the following reasons: its historical popularity as a (healthy) beverage, its high selling value, the importation of barely recognizable raw product (i.e., crushed), and the scarcity of studies concerning adulterants or contamination. The DNA Verity Test includes both the sequencing of DNA barcoding markers and genotyping of labeled-PCR DNA barcoding fragments for each sample analyzed. This protocol (DVT) was successively applied to verify the authenticity of 32 commercial teas (simple or admixture), and the main results can be summarized as follows: (1) the DVT protocol is suitable to detect adulteration in tea matrices (contaminations or absence of certified ingredients), and the method can be exported for the study of other similar systems; (2) based on the BLAST analysis of the sequences of rbcL+matK±rps7-trnV(GAC) chloroplast markers, C. sinensis can be taxonomically characterized; (3) rps7-trnV(GAC) can be employed to discriminate C. sinensis from C. pubicosta; (4) ITS2 is not an ideal DNA barcode for tea samples, reflecting potential incomplete lineage sorting and hybridization/introgression phenomena in C. sinensis taxa; (5) the genotyping approach is an easy, inexpensive and rapid pre-screening method to detect anomalies in the tea templates using the trnH(GUG)-psbA barcoding marker; (6) two herbal companies provided no authentic products with a contaminant or without some of the listed ingredients; and (7) the leaf matrices present in some teabags could be constituted using an admixture of different C. sinensis haplotypes and/or allied species (C. pubicosta).

Prevalence, Distribution, and Diversity of Salmonella spp. in Meat Samples Collected from Italian Slaughterhouses.

Recently worldwide food safety authorities indicated the rise of foodborne outbreaks linked to Salmonella: this highlighted the need to intensify monitoring and apply more targeted controls to help manage the spread of the disease. The aim of this study was to assess the prevalence and distribution of Salmonella serotypes in 7 slaughterhouses, located in different areas of Naples province (Regione Campania, Italy). Meat samples collected from the slaughterhouses were submitted for standardized microbiological analysis in 2015. Results of routine testing for Salmonella spp. were analyzed and then compared to biochemical and molecular evaluations. Salmonella spp. were detected in 12% of 320 samples examined (39/320) and the isolation rates ranged from 87% (32 samples) for raw poultry meat to 13% (7 samples) for pork meat. Biochemical serotyping showed that approximately 50% of the isolates belonged to Salmonella enterica serotype Choleraesuis. Rapid detection methods, such as molecular analysis (polymerase chain reaction and gel electrophoresis), able to confirm food matrices contamination, represent a valid support to the fast identification of Salmonella species. A further aspect of the study consisted, indeed, on analyzing isolated strains through molecular evaluations. By amplifying bacterial DNA-using invA primers, selective for Salmonella-it was possible, in less than 3 h, to classify the isolates as Salmonella spp., confirming the results of microbiological outcomes. Results of distribution analysis, supported by rapid molecular approaches, showed the difficulty of reducing Salmonella risk on food chain. This emphasized the importance of periodic surveillance to prevent outbreaks.

Rare earth elements in human and animal health: State of art and research priorities.

BACKGROUND: A number of applications have been developed using rare earth elements (REE), implying several human exposures and raising unsolved questions as to REE-associated health effects. METHODS: A MedLine survey was retrieved from early reports (1980s) up to June 2015, focused on human and animal exposures to REE. Literature from animal models was selected focusing on REE-associated health effects. RESULTS: Some REE occupational exposures, in jobs such as glass polishers, photoengravers and movie projectionists showed a few case reports on health effects affecting the respiratory system. No case-control or cohort studies of occupational REE exposures were retrieved. Environmental exposures have been biomonitored in populations residing in REE mining areas, showing REE accumulation. The case for a iatrogenic REE exposure was raised by the use of gadolinium-based contrast agents for nuclear magnetic resonance. Animal toxicity studies have shown REE toxicity, affecting a number of endpoints in liver, lungs and blood. On the other hand, the use of REE as feed additives in livestock is referred as a safe and promising device in zootechnical activities, possibly suggesting a hormetic effect both known for REE and for other xenobiotics. Thus, investigations on long-term exposures and observations are warranted. CONCLUSION: The state of art provides a limited definition of the health effects in occupationally or environmentally REE-exposed human populations. Research priorities should be addressed to case-control or cohort studies of REE-exposed humans and to life-long animal experiments.

Assessment of the concentrations of polycyclic aromatic hydrocarbons and organochlorine pesticides in soils from the Sarno River basin, Italy, and ecotoxicological survey by Daphnia magna.

We studied the contamination level of the soils of the Sarno River basin in southwestern Italy by combined acute toxicity test with Dapnia magna and chemical extraction of polycyclic aromatic hydrocarbons (PAHs) and organochlorine pesticides (OCPs). For the ecotoxicological assessment, 188 samples were taken and coincided with those of a previous study (2013) where heavy metals were surveyed. For the organics assessment, 21 samples were selected nearby representative areas of elevated anthropic pressure. About 10.1 % of the samples showed noticeable sign of D. magna mortality, 61-100 %, and fall along the potentially floatable areas of Sarno and Solofrana basins with high degree of contamination by Cr, As, Zn, and Hg. High levels of ecotoxicity, 61-100 %, were determined in the lower Sarno River basin in areas of moderate or low degree of contamination by Cd, Cu Hg, Pb, Sb, Sn, and Zn. Benzo(a)pyrene, indenopyrene, and benzo(g,h,i)perylene were present at concentrations of 0.32, 0.23, and 0.18 mg kg(-1), respectively, 2- to 3-fold the law limits with most of the samples falling nearby the points where the ecotoxicity output was close to 100 %. Among OCPs, pp'-DDT had a mean of 0.225 mg kg(-1) and hence about more than 200- and 2-fold the residential, 0.01 mg kg(-1), and commercial/industrial limits, 0.1 mg kg(-1) and determined mainly in the central Sarno valley in an area where elevated concentrations of benzopyrene and D. magna mortality were also observed. The study evidenced the high rate of contamination by PAHs and OCPs of the soils and the need of urgent remediation actions.

Antimicrobial activity of various cationic molecules on foodborne pathogens.

Antibacterial effects of various arginine- and lysine-rich polycationic proteins and polymers were evaluated by broth and solid dilution assay on a range of foodborne pathogens, Gram-positive and Gram-negative bacteria. The Minimum Inhibitory Concentration (MIC) and the Minimum Bactericidal Concentration (MBC) of α-poly-L-lysine (poly-lys), α-poly-L-arginine (poly-arg) and protamines from herring sperm (clupeine sulphate) and salmon sperm (salmine sulphate) were determined on Bacillus subtilis, Bacillus cereus, Staphylococcus aureus, Listeria monocytogenes, Salmonella typhimurium, Shigella sonnei, Escherichia coli O157:H7 and Pseudomonas aeruginosa. All these molecules showed antibacterial activity on all strains with different MIC and MBC values. The molecular mechanisms underlying the effect of α-poly-L-arginine might be related to the entrance of the molecule into the cell. In fact α-poly-L-arginine labelled with 7-Diethylamino coumarin-3-carboxylic acid, succinimidyl ester (DEAC,SE) showed ability to permeate the cell membrane of B. cereus and E. coli O157:H7.