Hydrogen production pathways in Clostridia and their improvement by metabolic engineering.

Biological production of hydrogen has a tremendous potential as an environmentally sustainable technology to generate a clean fuel. Among the different available methods to produce biohydrogen, dark fermentation features the highest productivity and can be used as a means to dispose of organic waste biomass. Within this approach, Clostridia have the highest theoretical H2 production yield. Nonetheless, most strains show actual yields far lower than the theoretical maximum: improving their efficiency becomes necessary for achieving cost-effective fermentation processes. This review aims at providing a survey of the metabolic network involved in H2 generation in Clostridia and strategies used to improve it through metabolic engineering. Together with current achievements, a number of future perspectives to implement these results will be illustrated.

Estrogenic activity in wastewater treatment plants through in vitro effect-based assays: Insights into extraction phase.

Effluents of wastewater treatment plants can abundantly spread endocrine disrupting chemicals in the environment. To improve water quality monitoring, the use of effect-based tools that measure estrogenic activity has been suggested, however their results could be influenced by different factors. This study compared the estrogenic activity of wastewater samples extracted with two stationary phases and tested with two in vitro effect-based assays to investigate whether and how stationary phases and assays could influence biomonitoring data. During four seasonal periods, the effluents of six WWTPs located in northern Italy were sampled. After the extraction using two different stationary phases (HLB, C18), the samples (n = 72) were tested using two effect-based assays: a gene reporter luciferase assay on mammalian cells (MELN) and yeast estrogen screen assay (YES). The results showed that estrogenic activity of HLB extracts was significantly different from the activity of C18 extracts, suggesting that extraction phase can influence biomonitoring data. Moreover, the estrogenic activity was overall higher using gene reporter MELN assay than using YES assay, suggesting that, due to difference in cell membrane permeability and metabolic activation, the applied cell model can affect the biomonitoring results. Finally, from the comparison between the activity of the final effluent and the environmentally safe estrogenic levels in surface waters, MELN data suggested that the activity of this effluent may pose an environmental risk, while YES data showed that it should not be considered a threat to the receiving surface waters. This study pointed out that a standardized approach is needed to assess the estrogenic activity of waters; it reported important data to select the most suitable stationary phase for samples extraction (samples extracted with C18 sorbent showed higher estradiol equivalent concentration values) and the most appropriate bioassay (gene reporter luciferase MELN assay was more sensitive than YES assay) to assess the environmental risk, thus protecting human health.

Atmospheric endocrine disruptors: A systematic review on oestrogenic and androgenic activity of particulate matter.

The alarming human health effects induced by endocrine disruptors (ED) have raised the attention of public opinion and policy makers leading worldwide to regulations that are continuously improved to reduce exposure to them. However, decreasing the exposure levels is challenging because EDs are ubiquitous and exposure occurs through multiple routes. The main exposure route is considered ingestion, but, recently, the inhalation has been hypothesized as an important additional route. To explore this scenario, some authors applied bioassays to assess the endocrine activity of air. This review summarizes for the first time the applied methods and the obtained evidences about the in vitro endocrine activity of airborne particulate matter (PM) collected outdoor. Among the bioassay endpoints, (anti)oestrogenic and (anti)androgenic activities were selected because are the most studied endocrine activities. A total of 24 articles were ultimately included in this review. Despite evidences are still scarce, the results showed that PM can induce oestrogenic, antioestrogenic, androgenic and antiandrogenic effects, suggesting that PM has an endocrine disrupting potential that should be considered because it could represent a further source of exposure to EDs. Although it is difficult to estimate how much inhalation can contribute to the total burden of EDs, endocrine activity of PM may increase the human health risk. Finally, the results pointed out that the overall endocrine activity is difficult to predict from the concentrations of individual pollutants, so the assessment using bioassays could be a valuable additional tool to quantify the health risk posed by EDs in air.

Occurrence of human pathogenic viruses in drinking water and in its sources: A review.

Since many waterborne diseases are caused by human pathogenic viruses, virus monitoring of drinking water (DW) and DW sources is crucial for public health. Therefore, the aim of this review was to describe the occurrence of human pathogenic viruses in DW and DW sources; the occurrence of two viruses proposed as novel indicators of human faecal contamination (Pepper mild mottle virus and Tobacco mosaic virus) was also reported. This research was focused on articles that assessed viral occurrence using molecular methods in the surface water used for DW production (SW-D), groundwater used for DW production (GW-D), DW and bottled-DW (BW). A total of 1544 studies published in the last 10 years were analysed, and 79 were ultimately included. In considering the detection methods, filtration is the most common concentration technique, while quantitative polymerase chain reaction is the most common quantification technique. Regarding virus occurrence in SW-D, GW-D, and DW, high percentages of positive samples were reported for adenovirus, polyomavirus and Pepper mild mottle virus. Viral genomes were frequently detected in SW-D and rarely in GW-D, suggesting that GW-D may be a safe DW source. Viral genomes were also detected in DW, posing a possible threat to human health. The lowest percentages of positive samples were found in Europe, while the highest were found in Asia and South America. Only three articles assessed viral occurrence in BW. This review highlights the lack of method standardization and the need for legislation updates.

Effectiveness of Workplace Health Promotion (WHP) interventions in university employees: a scoping review.

Workplace Health Promotion (WHP) interventions proved to be effective in several workplace contexts. Currently, the effectiveness of such interventions in the academic workplace is lacking, albeit evidence suggests similar patterns to those occurring in other workplace sectors. The aim of this study was to review WHP interventions in the university workplace that led to improve health- and work-related outcomes. Articles were selected using Pubmed, Web of Science, Scopus, Embase, PsycInfo, Cinhal and FSTA, using search strings focused on health- and/or work-related outcomes and involving University WHP interventions published between January 2010 and July 2021. The majority of the 12 studies selected reported positive results in their individuality, especially regarding health-related outcomes [biological such as weight loss, physical activity, mental health and lifestyle habits] and work-related outcomes concerning improvements either for the employee or for the working system. Studies on economic advantage and Return on Investment were limited and reported contrasting results. In conclusion, we have highlighted how the studies on effectiveness of WHP interventions in the university context are few and heterogeneous and need to be encouraged further research in order to build specific guidelines that are effective.

Interventions concerning prevention promotion in the workplace, involving activities targeting lifestyle habits, nutrition and/or physical activity as well as general wellbeing, are growing and becoming more common due to their benefits on health and work outcomes. The aim of this review was to investigate the effectiveness of such interventions in the university workplace context to help create directed guidelines for their correct implementation in this setting. Although it was difficult to draw peculiar conclusions due to the heterogeneity of the programs described in the studies, the overall positivity of the results and the perspective university environment, outlook that the implementation of multicomponent and multilevel interventions would improve employees' wellness and favor universities' welfare.

Direct Impact of the Air on Mutant Cells for Mutagenicity Assessments in Urban Environments.

BACKGROUND: Urban air pollution is recognized as a critical problem for public health and is classified as a carcinogen for humans. A great number of studies have focused on the monitoring of urban air mutagenicity. One of the best-known and applied methods for assessing mutagenicity is the Ames test, a bacterial reverse mutation test. The classic protocol for assessing air mutagenicity involves the concentration of particulate matter (PM) on filters and subsequent extraction using organic solvents. This work aimed to develop a method for the evaluation of air mutagenicity directly impacted by air on microbial plates already containing an Ames' microbial sensor. METHODS: A specific six-month sampling campaign was carried out in Turin in a period with high air pollution. Samples were tested for mutagenicity on Salmonella typhimurium strains TA98, TA100, and YG1024 with the traditional method and with the new direct method. RESULTS: The new protocol is able to evaluate the mutagenicity of the sampled air and obtain repeatable results. The final sensitivity is similar to the traditional method (≈10 net revertants/m3); however, the mutagenic response is due to the complete air pollution mixture, including volatile and semivolatile pollutants avoiding the concentration of filters and the following laborious extraction procedures. CONCLUSIONS: Despite some critical issues in contamination control, the method is easier, faster, and less expensive than traditional methods.

Use of online and paper-and-pencil questionnaires to assess the distribution of orthorexia nervosa, muscle dysmorphia and eating disorders among university students: can different approaches lead to different results?

PURPOSE: Administration of questionnaires to assess the diffusion of disordered eating behaviours via the web is becoming common today. The aim of this study is to assess whether two different approaches of administering a test to assess traits of eating disorders (EDs), orthorexia nervosa (ON) and muscle dysmorphia (MD) by email recruitment and online completion (web-based survey-WBS) and by in person recruitment and paper-and-pencil completion (paper-based survey-PBS), gives different results. METHODS: During 2 consecutive academic years, a self-reported questionnaire consisting of questions about personal characteristics and three tests for the evaluation of ON (ORTO-15), MD (MDDI-ITA), and EDs (EAT-26) were administered to two groups of undergraduates, respectively, as a WBS and a PBS. RESULTS: The WBS response rate was 6.7% (N = 137), and the PBS response rate was 86.5% (N = 372). The WBS group showed a statistically significant higher prevalence of students with eating disordered behaviours (21.2% vs 5.4%) and registered a higher mean score on the EAT-26 test (13.5 ± 11.1 vs 6.0 ± 8.0); no differences between the two groups emerged for ON and MD prevalence and test scores. Moreover, in the WBS group, the number of students with one or more tests with test scores above the cut-off values was significantly higher (46.0% vs 32.3%). CONCLUSION: The choice of the approach to administer a questionnaire to assess the diffusion of EDs and related issues must take into account all the factors that can result in selection bias and that can affect the reliability of the results. LEVEL OF EVIDENCE: Level V, descriptive cross-sectional survey.

In Vitro Effects of Particulate Matter Associated with a Wildland Fire in the North-West of Italy.

Wildland fires, increasing in recent decades in the Mediterranean region due to climate change, can contribute to PM levels and composition. This study aimed to investigate biological effects of PM2.5 (Ø < 2.5 µm) and PM10 (Ø < 10 µm) collected near a fire occurred in the North-West of Italy in 2017 and in three other areas (urban and rural areas). Organic extracts were assessed for mutagenicity using Ames test (TA98 and TA100 strains), cell viability (WST-1 and LDH assays) and genotoxicity (Comet assay) with human bronchial cells (BEAS-2B) and estrogenic activity using a gene reporter assay (MELN cells). In all sites, high levels of PM10 and PM2.5 were measured during the fire suggesting that near and distant sites were influenced by fire pollutants. The PM10 and PM2.5 extracts induced a significant mutagenicity in all sites and the mutagenic effect was increased with respect to historical data. All extracts induced a slight increase of the estrogenic activity but a possible antagonistic activity of PM samples collected near fire was observed. No cytotoxicity or DNA damage was detected. Results confirm that fires could be relevant for human health, since they can worsen the air quality increasing PM concentrations, mutagenic and estrogenic effects.

Comparison of UV, Peracetic Acid and Sodium Hypochlorite Treatment in the Disinfection of Urban Wastewater.

One source of water contamination is the release of wastewater that has not undergone efficient treatment. The aim of this study was to evaluate the reduction obtained with sodium hypochlorite (NaClO), UV and peracetic acid disinfection treatment of Salmonella spp., pathogenic Campylobacter, STEC and bacterial indicators in three full-scale municipal wastewater plants. A general reduction in Salmonella was observed after disinfection, but these bacteria were detected in one UV-treated sample (culture method) and in 33%, 50% and 17% of samples collected after NaClO, UV and PAA disinfection treatments, respectively (PCR method). A better reduction was also observed under NaClO disinfection for the microbial indicators. Independent of the disinfection treatment, E. coli O157:H7 was not detected in the disinfected samples, whereas some samples treated with UV and PAA showed the presence of the stx1 gene. No reduction in the presence of stx2 genes was verified for any of the disinfection treatments. Campylobacter was not detected in any of the analysed samples. The overall results highlight a better reduction in microbiological parameters with a NaClO disinfection treatment in a full-scale municipal wastewater plant compared with UV and PAA. However, the results indicate that a complete and specific monitoring program is necessary to prevent a possible risk to public health.

Traits of orthorexia nervosa and muscle dysmorphia in Italian university students: a multicentre study.

PURPOSE: This study aimed to evaluate the prevalence of traits of orthorexia nervosa (ON) and muscle dysmorphia (MD) in a group of undergraduates, investigate the associations between the risk of these conditions and the type of university course attended, the individual characteristics (gender, BMI, amount of physical activity, supplements and medicines use, dieting) and the risk of eating disorders (EDs). METHODS: A self-reported questionnaire consisting of a socio-demographic section and three tests validated for the evaluation of a risk of ON (ORTO-15), MD (MDDI-ITA) and EDs (EAT-26) was completed by 918 students from three Italian universities. RESULTS: 29.0% of participants demonstrated traits of ON and 5.0% of MD, without differences in prevalence in the three areas of study investigated (health-scientific, economic-humanistic, sport sciences); students of sport sciences exhibited a significantly higher score for MDDI-ITA (F = 6.493, p = 0.002). Participants with ON and MD traits were more on a diet (OR = 0.47, p ≤ 0.001 and OR = 0.428, p = 0.020, respectively) and showed a higher prevalence of EDs risk (OR = 3.55, p < 0.001 and OR = 10.23, p ≤ 0.001, respectively). The simultaneous presence of ON, MD, and EDs traits was seen in 5.4% of the students and the three test scores were correlated. CONCLUSIONS: The prevalence of ON and MD traits was found similar to that reported in the literature on undergraduates. Some associations observed improvement in the knowledge about these conditions, especially the association of participants with ON and MD traits with dieting and EDs traits and the correlation of the three test scores suggests a connection among these potential conditions. LEVEL OF EVIDENCE: Level V, descriptive cross-sectional survey.

Monitoring of Air Microbial Contaminations in Different Bioenergy Facilities Using Cultural and Biomolecular Methods.

Bioaerosol exposure linked to the bioenergy production from waste and its effects on human health in occupational and residential environments has rising interest nowadays. The health risk associated with the exposure includes mainly infective diseases, allergies, chronic bronchitis, and obstructive pulmonary disease. A risk assessment's critical point is the bioaerosol quality and quantity characterization. The aim of this study is to evaluate the application of different methods for the analysis of bioaerosol sampled in bioenergy plants. This study involved six Italian plants for the treatment of biomasses and energy production. Bioaerosol cultural evaluation was performed, by Surface Air System (SAS) sampler, and DNA was extracted from PM0.49 samples and Low Melting Agar plates. qRT-PCR followed by Denaturing Gradient Gel Electrophoresis (DGGE) and band sequencings were performed. The cultural method is able to detect less than 15% of what is evaluable with bio-molecular methods. A low sample alfa-diversity and a high beta-biodiversity in relation to feedstock and process were observed. Sequencing showed microorganisms with a hygienic-sanitary relevance such as Arcobacter, Pseudomonas, Enterobacter, Klebsiella, Enterococcus and Bacillus. Integrated cultural and biomolecular methods can be more exhaustive to evaluate bioaerosol's exposure in the occupational environment.

Cryptosporidium Oocyst Contamination in Drinking Water: A Case Study in Italy.

The aim of this study was to evaluate the occurrence of Cryptosporidium oocysts in a drinking water treatment plant (DWTP) located in a rural area of northern Italy. Influent and effluent samples were collected at the DWTP over three years (2013-2016). In parallel, tap water samples from a public drinking fountain were collected as well. All samples were analyzed for the presence of Cryptosporidium spp. oocysts by a common method based on an immunomagnetic separation (IMS)/immunofluorescence assay (IFA), complemented by 4,6-diamidino-2-phenylindole (DAPI) staining. A reverse transcriptase-PCR (RT-PCR) protocol was added to evaluate oocyst viability. The results highlighted a high variability of oocyst concentrations across all samples (mean 4.3 ± 5.8/100 L) and a high variability in the percentage of DAPI-positive specimens (mean 48.2% ± 40.3%). Conversely, RT-PCR did not reveal the presence of viable C. parvum and C. hominis oocysts. A nested PCR targeting Cryptosporidium 18S ribosomal DNA, carried out in two water samples, confirmed the presence of a Cryptosporidium genotype associated with wild animals in the river and in tap water. The results obtained underline the vulnerability of the investigated surface water to Cryptosporidium spp. contamination. Although the recovered Cryptosporidium genotype is not a human pathogen, its presence demonstrates the existence of a potential pathogen Cryptosporidium spp. contamination risk. Moreover, these results underline the importance of also considering unconventional (not bacterial) biological contaminations (protozoa) in water resources in rural areas, including those of developed countries.

Anodic microbial community analysis of microbial fuel cells based on enriched inoculum from freshwater sediment.

The characterization of anodic microbial communities is of great importance in the study of microbial fuel cells (MFCs). These kinds of devices mainly require a high abundance of anode respiring bacteria (ARB) in the anode chamber for optimal performance. This study evaluated the effect of different enrichments of environmental freshwater sediment samples used as inocula on microbial community structures in MFCs. Two enrichment media were compared: ferric citrate (FeC) enrichment, with the purpose of increasing the ARB percentage, and general enrichment (Gen). The microbial community dynamics were evaluated by polymerase chain reaction followed by denaturing gradient gel electrophoresis (PCR-DGGE) and real time polymerase chain reaction (qPCR). The enrichment effect was visible on the microbial community composition both during precultures and in anode MFCs. Both enrichment approaches affected microbial communities. Shannon diversity as well as ß-Proteobacteria and γ-Proteobacteria percentages decreased during the enrichment steps, especially for FeC (p < 0.01). Our data suggest that FeC enrichment excessively reduced the diversity of the anode community, rather than promoting the proliferation of ARB, causing a condition that did not produce advantages in terms of system performance.

Cytotoxicity and genotoxicity of light emitted by incandescent, halogen, and LED bulbs on ARPE-19 and BEAS-2B cell lines.

LED technology has the extraordinary ability to reduce energy consumption, constituting an economic and ecological advantage, so it is planned to replace incandescent, halogen and other inefficient bulbs for public and domestic lighting with LEDs. LEDs present specific spectral and energetic characteristics compared with those of other domestic light sources, so the potential risks for human health of these bulbs need to be explored. The aim of this study was to assess cytotoxicity and genotoxicity of light emitted by different commercial light bulbs: incandescent, halogen, and two LED bulbs with different correlated color temperatures. The evaluation was done on ARPE-19 as a specific cell model for eye toxicity and on BEAS-2B as a good cell model for toxicology tests. Light induced mainly cytotoxic effects on ARPE-19 and DNA damage on BEAS-2B, so different cell lines showed different biological responses. Moreover, our findings indicate that among the four bulbs, cold LED caused the highest cytotoxic effect on ARPE-19 and the highest genotoxic and oxidative effect on BEAS-2B. Cold LED is probably able to cause more cellular damage because it contains more high-energy radiations (blue). These results suggest that LED technology could be a safe alternative to older technologies, but the use of warm LED should be preferred to cold LED, which can potentially cause adverse effects on retinal cells.

Effectiveness of a neutral electrolysed oxidising water (NEOW) device in reducing Legionella pneumophila in a water distribution system: A comparison between culture, qPCR and PMA-qPCR detection methods.

Disinfection of hot water systems is critical for reducing Legionnaires' disease in high-risk buildings. The use of neutral electrolysed oxidising water (NEOW) is a promising method for the control of microorganisms in hot water systems. However, full-scale evaluations of the efficacy of NEOW devices to control Legionella pneumophila are currently lacking. The aim of this study was to assess the effectiveness of a NEOW device in reducing L. pneumophila in a hotel water network. Water samples (n = 67) were collected from different sites of a hotel distribution system before and after the installation of the NEOW device at the 1st, 4th, 8th and 12th week. Detection of L. pneumophila was performed comparing culture, qPCR and PMA-qPCR methods. Total bacterial counts (22 °C and 37 °C), Pseudomonas spp. and physico-chemical parameters were also monitored. The NEOW treatment resulted in a reduction of the amount of L. pneumophila positive samples (-32%) and of the number of heavily contaminated points (>104 CFU/L and >103 CFU/L) (-100% and -96%, respectively). Treatment maintained L. pneumophila at low levels (<102 CFU/L), which do not require specific intervention measures. The effectiveness of the disinfection system was also confirmed by PMA-qPCR (p < 0.001). The use of PMA resulted in a signal decrease in almost all samples upon the disinfection treatment. The NEOW disinfection device appears to be a promising approach to reduce the colonisation of hot water systems by L. pneumophila; however, further investigations are needed to ascertain its efficiency over longer time periods.

Physical Activity, Lifestyle Factors and Oxidative Stress in Middle Age Healthy Subjects.

Oxidative stress (OS) has been recognized to play a primary role in many acute and chronic diseases. Environmental and lifestyle factors, such as physical activity and dietary intake are involved in the oxidative balance, but their specific influence remains unclear. In order to contribute to a greater characterization of the oxidative status in relation to exercise training and to environmental and lifestyle factors, different biomarkers-pro-oxidant capacity (d-ROMs), anti-oxidant capacity (BAP), radical scavenging activity (DPPH) and DNA damage (8-OHdGuo)-were measured in biological samples of a group of healthy middle aged subjects. The evaluation of the investigated biomarkers highlighted a significant effect of exercise training on OS, measured as d-ROMs and 8OhdGuo, in subjects playing regular physical activity. An association of the OS status measured by DPPH and 8-OhdGuo with the condition of living in urban high traffic areas was also found. Otherwise dietary habits did not reveal any significant effect on OS levels by the investigated biomarkers. As a whole the results obtained in this investigation suggested that a correct lifestyle, with regular physical activity practice, contributes to control the OS status in middle age subjects.

Aerosol exposure and risk assessment for green jobs involved in biomethanization.

Anaerobic digestion is a consolidated biotechnology able to produce renewable energy from biomasses. In the European countries, quick growth of biogas production from different organic matrices including wastes has been observed. In relation to the characteristics and quantity of the anaerobic digestion of feedstock, there are different technologies, advantages and criticisms. An accurate occupational risk assessment and development of management tools for green jobs involved in the anaerobic digestion plants are due. The aim of this work is to assess the aerosol exposure for such workers, focusing on the bioaerosol risk. Full scale plants for the treatment of organic municipal waste, waste water treatment sludge, agro zootechnical and food producing byproducts were involved for this purpose. The bioaerosol levels were monitored during activities through culturing and biomolecular methods; moreover, the sub-fractionated PM10 and carried endotoxins were measured in different plant areas. Global microbial contamination is higher (>5000 UFC/m3) in the area where organic wastes are handled and pretreated, both for organic municipal waste plants - with a bacterial prevalence - and agro zootechnical plants - with a fungi prevalence. Moreover, the microbial contamination is higher where organic municipal waste is present in respect to other biomasses (ANOVA p < 0.01). Numerous pathogens are carried by the aerosol. HAdV-4 presence is lower than LOQ (50 gene copies/m3) in all the samples. Environmental PM10 reached the 280 µg/m3 level including PM3 for 78%. Endotoxin pollution overtakes the 90 EU/m3 limit sporadically. Personal PM4.5 reached 10 mg/m3 only for maintenance technicians in the pretreatment area for organic municipal waste. The risk can be evaluated under a quantitative and qualitative point of view highlighting risk management improvement for anaerobic digestion plants.

Viability of Legionella pneumophila in Water Samples: A Comparison of Propidium Monoazide (PMA) Treatment on Membrane Filters and in Liquid.

Legionella pneumophila is a ubiquitous microorganism widely distributed in aquatic environments and can cause Legionellosis in humans. A promising approach to detect viable cells in water samples involves the use of quantitative polymerase chain reaction (qPCR) in combination with photoactivatable DNA intercalator propidium monoazide (PMA). However, the PMA efficiency could be different depending on the experimental conditions used. The aim of this study was to compare two PMA exposure protocols: (A) directly on the membrane filter or (B) in liquid after filter washing. The overall PMA-induced qPCR means reductions in heat-killed L. pneumophila cells were 2.42 and 1.91 log units for exposure protocols A and B, respectively. A comparison between the results obtained reveals that filter exposure allows a higher PMA-qPCR signal reduction to be reached, mainly at low concentrations (p < 0.05). This confirms the potential use of this method to quantify L. pneumophila in water with low contamination.

Assessing Methanobrevibacter smithii and Clostridium difficile as not conventional faecal indicators in effluents of a wastewater treatment plant integrated with sludge anaerobic digestion.

Wastewater treatment plants (WWTP) are an important source of surface water contamination by enteric pathogens, affecting the role of environmental water as a microbial reservoir. We describe the release to the environment of certain anaerobes of human and environmental concern. The work was focused on emerging microbial targets. They are tracing, by RT-qPCR, on WWTP effluents, both liquid and solid, when an anaerobic digestion step is included. The focus is placed on Clostridium spp. with the specific quantification of Clostridium perfringens, as typical bioindicator, and Clostridium difficile, as emerging pathogen not only confined into nosocomial infection. Moreover methanogens were quantified for their involvement in the anaerobic digestion, and in particular on Methanobrevibacter smithii as major methanogenic component of the human gut microbiome and as not conventional faecal indicator. In the water samples, a reduction, statistically significant, in all microbial targets was observed (p < 0.01), 2 log for the total bacteria, 1.4 log for the Clostridium spp. and M. smithii, 1 log for total methanogens, C. perfringens and C. difficile. The AD process contribute to a significant change in microbial levels into the sludge for total bacteria and total methanogens (p < 0.01), both when the input sludge are primary and secondary, while for the presence of Clostridium spp. and C. difficile there was not a significant change. The produced data are innovative showing which is the diffusion of such anaerobic microorganisms throughout the WWTP and opening a discussion on the implementation of possible techniques for a more efficient microbial removal from effluents, particularly bio-solids, to reduce the potential release of pathogens into the environment.

The importance of indicators in monitoring water quality according to European directives.

Directive 2000/60/EC and subsequent legislation provide a list of priority substances to be measured and monitored in EU water bodies and require the adoption of analytical methods that ensure comparability of the data collected in all Member States. These regulations and standards have gradually improved water quality in the EU. However, new drugs, whose effects on ecosystems and health are still to be determined, are detected with growing frequency. The Member States are now called upon to characterize and monitor these pollutants in view of their possible inclusion in the priority substance list.