Programma della conferenza

Pigments used in various industries such as the production of paints, dyes and cosmetics are heterogeneous materials sized between 0.2-0.3 micrometers and can be classified as emerging pollutants. Unlike dyes, they do not penetrate the material but form a colored layer on the surface of the objects from which they are easily released and dispersed in the environment. Pigments are divided into four main categories: natural inorganic substances (from metals or rare earths extracted from mineral deposits), synthetic inorganic substances (industrially produced from raw minerals), natural organic substances (from plant or animal sources), and synthetic organic substances (from petroleum derivatives).

Their dispersion in the environment poses a significant threat to terrestrial and aquatic ecosystems, as they cause pollution both through the release of the chemical substances that compose them and through their nanometric nature, which has the property of interacting with organisms, thus threatened the conservation of the ecosystem biodiversity.

In this study, six pigments (three synthetic and three natural, similar colors) with the widest range of commodity applications were selected. Size, morphological properties, chemical composition (metals, metalloids, rare earths), and behavior in the aquatic environment, both freshwater and marine were determined. In addition, their ecotoxicity was assessed using two target species: the bacterium Aliivibrio fischeri (aquatic environment) and the monocotyledonous plant Lepidium sativum (terrestrial environment). In addition, in plants, the absorption of pigments at the root level was also evaluated. The results show a significant difference of behavior of the pigments in aquatic environments depending on the salinity of the matrix; content of metals, metalloids and rare earths also depending on their color; a significant absorption at the root system level by plants. Our results suggest that further research is needed to evaluate the risk for the environment related to the massive use and dispersion of pigments.

Per- and polyfluoroalkyl (PFAS) compounds constitute a class of chemicals that possess unique water and oil-repellent properties. For this reason, these synthetically produced fluorinated compounds are implicated in various applicative fields and are used to produce many items, such as food packaging, non-stick cookware and cosmetics. In this context, although PFAS improved many aspects of the everyday life, on the other hand their global distribution led to an abundant release in the environment, with an elevated consequent risk of bioaccumulation in the living organisms as already observed for other pollutants. Indeed, given their properties, PFAS resist to any kind of degrading process and this property, combined with a massive use and release of PFAS-containing products, make their waste a crucial environmental worldwide problem. In this context, while numerous data are available regarding the toxicity of the first generation “legacy” PFAS, scant information exists on newly synthesized emerging molecules, with limited details on the cellular-level effects. Given these premises, the medicinal leech Hirudo verbana has been proposed in the current study to assess the possible side effects of four different fluorinated PFAS compounds (HFPO-DA, PFMoBa, PFOA and PFMOPrA) during freshwater dispersion. In particular, two concentrations (0.6 and 229 μM) have been tested, evaluating PFAS ability to induce leech inflammatory response and oxidative stress. Results have been analyzed by means of morphological, immunohistochemical and molecular assays, revealing how these molecules were differently able to modulate both the cellular and the molecular response. As recently demonstrated for plastics particles, this work can lead new information also about PFAS, deepening the current knowledge on their potential harmful effects, deriving also from a possible accumulation in living organisms.

Fire training sites and airports are major PFAS contamination sources due
to the widespread use of Aqueous Film-Forming Foam (AFFF). Although recent
policies restrict PFAS congeners higher than C8, legacy contaminants such as
PFOS and PFOA persist, impacting ecosystems. Modern AFFF formulations
still include short-chain PFAS and their oxidizable precursors. In terrestrial
ecosystems, earthworms play a key role in soil processes. This study assesses
the impact of PFAS-contaminated soil on earthworms, highlighting ecological
consequences and the need for sustainable policies. Using a multi-tier
approach, the study investigated apical toxicity and sublethal responses of
earthworms across a PFAS contamination gradient at a drill site in Trelleborg,
Sweden. PFAS contamination was confirmed with 22 compounds measured,
mainly PFOS, PFHxS, PFOSA, 6:2 FTS, PFHpS, PFOA, and PFPeA, ranging
from 960 ppb to 8.7 ppb. OECD No. 207 (acute toxicity) and No. 222
(reproduction) tests, plus 30-day sublethal assessments, were conducted.
Biomarkers included mRNA relative abundances of immune-related genes
(lysenin and ccf-1) and oxidative burst in hemocytes; enzymatic responses in
tissues, including catalase, phenol oxidase, and acetylcholinesterase. A
behavioral test based on escape time was administered to each earthworm
after 30-day exposure. Pristine soil with similar granulometric size and organic
matter served as an external reference control. Acute toxicity (mortality) at 14
days was below the threshold of OECD test No. 207, but reproduction was
impacted at all sites except B7 according to OECD No. 222. In general,
sublethal responses across the PFAS gradient showed a significant impact
compared to the reference control, but a linear response was barely observed
with the PFAS concentration found in soil. Acetylcholinesterase activity and
escape time showed interesting correlations, warranting further investigation of
critical neurotransmitters for locomotion using LC-MS metabolomics. These
findings contribute to understanding PFAS-contaminated site ecology and aid
in constructing an environmental database for risk assessment.

After the spreading of COVID-19, the amount of plastics in the ocean was severely enhanced due to the increased production of PPE, in particular surgical masks. Once entered into the environment, these disposable items can both release potential toxic additives and undergo fragmentation leading to the formation of microplastics and nanoplastics. Moreover, microplastics and nanoplastics are considered the most harmful for corals and marine organisms since can easily be ingested. Current information regarding the effects of micro and nanoplastics on coral reefs is limited; especially the toxicity of nanoplastics and nanoplastics leachate from fibers degradation of synthetic fabrics. The alcyonacean Pinnigorgia flava was exposed for 72 h to different concentrations of polypropylene nanofibers (0.1 and 1 mg/L) and nanofibers leachate (0.1 and 1 mg/L) under controlled aquaria conditions. The cellular response was assessed through antioxidant enzymatic assays, namely Superoxide Dismutase (SOD), Catalase (CAT), Glutathione Reductase (GR), and Glutathione-S-Transferase (GST). Results showed that for all treatments, oxidative stress was experienced in all samples. However, different patterns of enzymatic activity were observed between nanofibers and leachates, with a general higher toxicity generated by leachates exposition that produced an inhibition and/or impairment of the antioxidant defense mechanisms at cellular level. This study represents a new area of investigation and is one of the first to focus on such stressors on soft coral species. In addition, the results highlight that nanofibers and leachates are indeed a major threat for coral reefs, which are already exposed to multiple climate change stressors and relentless anthropogenic pressures.

This study explores the potential of reduced graphene oxide (rGO) to mitigate metal stress in Lemna minor 5500 through its adsorption capacity. We investigated the co-exposure effects of nickel (Ni) and copper (Cu) on the duckweed plant.

Experimental set-up included different treatments (in triplicate): control plants (CP) in mineral water, plants with rGO (1 mg/L) (PG), plants exposed to either Cu (1 mg/L) or Ni (1.3 mg/L) (PM) and plants exposed to a combination of rGO (1 mg/L) and either Cu (1 mg/L) or Ni (1.3 mg/L) (PGM).

Cu exposure significantly reduced biomass (20%) by T14 in PM compared to CP. This effect was not statistically significant at T7, although a similar trend of RGR was observed. Interestingly, rGO treatment appeared to mitigate Cu stress. After 14 days, PGM plants showed a statistically significant increase in carotenoid content (43%) compared to PM, while total chlorophyll content also increased by 65% (though not statistically significant). These findings suggest rGO may alleviate Cu-induced stress on L. minor, potentially through enhanced chlorophyll and carotenoid production.

Copper accumulation in the PGM treatment was 25% lower than in the PM treatment after only seven days, suggesting its greater effectiveness in reducing copper accumulation. Our findings from the Cu co-exposure experiment support the hypothesis that GBMs can reduce metal bioavailability for plants. Ni uptake wasn't affected by rGO, suggesting a need for further study on this metal-rGO interaction. These results suggest a possible relationship between the observed effects and the different affinities that Cu and Ni may have for rGO and, consequently, their relative bioavailability for L. minor. The different physiological performances of the two treatments seem to confirm these hypotheses. Further investigations will be necessary to verify their applicability in the field of phytoremediation, such as large-scale experiments with different GBM and selected Lemna clones.

Antibiotic (ABs) use is surging globally, particularly in intensive livestock production. ABs are often incompletely metabolized by animals, leading to significant excretion through feces and urine. Consequently, aquatic environments are becoming major repositories of ABs, with significant negative impacts on ecosystems, as these emerging pollutants also affect non-target organisms. This study examines the effects of a mixture of sulfamethoxazole (SMX) and chlorotetracycline (CTC), along with their potential interaction with copper sulphate (CuSO₄ 5H₂O, Cu), released in aquatic environment from anthropogenic sources, on the aquatic plant Lemna sp., a common aquatic plant worldwide distributed and used as a model organism in ecotoxicology tests due to its rapid growth and sensitivity to environmental pollutants. Initially, following OECD guidelines¹, the toxicity Cu, SMX and CTC was investigated on Lemna mediterranea 9425a plants. The concentration-response relationship was used to estimate EC₅₀ values: Cu, EC₅₀= 7.27 mg/L; SMX, EC₅₀= 173.00 mg/L and CTC, EC₅₀= 6.99 mg/L.

To simulate environmental conditions, L. mediterranea was exposed to SMX (2.5 mg/L) and CTC (0.1 mg/L) mixture alone and in combination with different concentrations of copper sulphate, 1.5 mg/L and 4 mg/L, for 14 days. The inhibitory effects induced by contaminants on duckweed plant growth and physiology have been evaluated by measuring fronds biomass, relative growth rate (RGR), pigment content, chlorophyll fluorescence, tolerance index and bioconcentration of Cu. Interestingly, the presence of the antibiotic mixture in combination with Cu appeared to mitigate the harmful effects of copper alone respect to duckweed's biomass and chlorophyll content. This was observed at both exposure times (7 and 14 days). Antibiotics seemed to lessen how much copper the plants absorbed at the highest copper dose, but the plants still accumulated copper over time, just more slowly.

¹OECD. Lemna sp. growth inhibition test. Guideline 221. Organization for Economic Co-operation and Development: Paris, France 2006.

The INSIGHT project (https://insight-project.org/) showcases four case studies integrating ecotoxicology and chemo-nano informatics models, demonstrating the intersection of ecotoxicology, chemical and material science safety, sustainability, and advanced in silico approaches. These integrated models, both data-driven and physics-based, provide comprehensive assessments of material safety, sustainability, and functionality, forming a Knowledge Graph that offers new insights and linkages between disparate concepts. The case studies encompass different chemicals or materials, industrial applications, model sets, regulatory landscapes, and scientific questions. They highlight the innovative integration of models and the resultant technical and societal insights.

The case studies focus on the following 4 topics. Graphene Oxide (GO) for Batteries and Construction: GO enhances battery performance and durability of cement. Challenges include scalable production and toxicity assessment. The study addresses GO's impact on material durability and recyclability within regulatory frameworks like REACH and RoHS. Per and Poly Fluoroalkylated Substances (PFAS) for the aerospace industry: PFAS are valued for thermal stability and low friction but pose environmental and health risks. The study explores PFAS replacements and models regulatory compliance, focusing on REACH and the Water Framework Directive. Bio-based Synthetic Amorphous Silica (SAS) for Tyres: Derived from renewable resources, bio-based SAS offers a sustainable alternative to conventional silica. The study evaluates life-cycle and social impacts, aligning with regulations like the EU's End-of-Life Vehicle Directive. Antimicrobial Coatings: Used in healthcare, food packaging, and consumer products, these coatings face challenges like microbial resistance and environmental impact. The study compares nano-enabled coatings with conventional disinfectants, under the EU Biocidal Products Regulation.

The informatics-driven approach integrates diverse modelling methodologies, including social life cycle assessment, to provide a holistic view of material functionality, safety, and sustainability. Each case study is documented through nanopublications, forming a Knowledge Graph consistent with the INSIGHT Data Management Plan and demonstrating leadership in model FAIRification.

Microplastic (MP) pollution is a major environmental concern and a significant threat to aquatic life. This study presents a preliminary assessment of MP pollution in abiotic (water and sediment) and biotic (macrophyton, periphyton, the edible part of the gastropod Sinotaia quadrata, the bivalve Corbicula fluminea, and the crustacean Atyaephyra desmarestii) compartments along different rivers in the Tuscany region. Five sampling sites were identified along the Arno, Bisenzio, and Ombrone rivers, which are known to be exposed to anthropogenic pressures (e.g., presence of a textile factories, effluents from various urban centers). S. quadrata (N=325) was found at all five sampling sites, while C. fluminea (N=19) was only found at site five and A. desmarestii (N=50) at site two. Samples were sorted by stereomicroscopy at 10-80X, and potential targets were then chemically analyzed using microscopy coupled with Fourier transform infrared spectroscopy (μFT-IR). The analysis led to the identification of 101 items. The main MP colors were blue, white, and black, and the primary chemical types identified were polypropylene, polyethylene terephthalate, and polyethylene. A total of 56 MP items were found in S. quadrata, with a statistically significant difference in the average size of microplastics across different size classes. For C. fluminea and A. desmarestii, 9 and 5 MP items were found, respectively. Regarding the abiotic compartments and macrophyton, 31 MPs were identified with a mean size of 318.55 μm for water, 235.70 μm for sediment, and 291.77 μm for macrophyton samples. No MPs were found in periphyton. This study underscores the pervasive presence of microplastics in both abiotic and biotic components of river ecosystems in Tuscany, highlighting the need for further research and mitigation strategies to address this environmental threat.

The environmental challenge of plastic waste has been compounded by the global COVID-19 pandemic. During this period, countries have introduced the obligation to wear surgical face masks in public places to control the spread of the virus.

Since face masks are predominantly made of polystyrene (PP) and tend to release micro/nano fibres (MNFs), they pose a potential hazard to ecosystems.

The present study provides new information regarding the impacts of MNFs in aquatic organisms by evaluating the effects of secondary PP-MNFs derived from the non-woven PP fabrics of surgical face masks on D. rerio individuals.

The impact of MNFs on embryonic and larval zebrafish developmental stages has been evaluated by respectively short-term (up to 6 days) and median-term (up to 15 days) bioassays. The effects of low (0.2 mg/L), medium (1 mg/L), and high (5 mg/L) environmental-relevant contamination levels of MNFs were evaluated. Alterations in several apical endpoints (embryonic development, survival, growth, morphology, behaviour) and transcriptomic analysis were investigated.

After six days of exposure, a significant reduction in the eye area was observed in both the 0.2 mg/L and 5 mg/L treatments. It is noteworthy that the upregulation of genes related to the negative regulation of developmental processes could explain the observed morphological alterations. Moreover, the downregulation of genes involved in energy-related metabolic processes suggests an impairment in the correct development of organisms exposed to PP-MNFs. Furthermore, an increased mortality in MNF treatments occurred between 9 and 12 days, period when larval fish make the transition from endogenous feeding to exogenous feeding. This suggests an impairment in foraging behaviour occurred due the exposure to secondary PP-MNFs.

The findings of this study demonstrate that environmental levels of PP-MNFs may pose a hazard to aquatic organisms, suggesting the potential for an ecotoxicological risk associated with the improper disposal of surgical face masks.

Microplastics (MPs) and invasive species pose significant threats to aquatic conservation, representing critical global eco-environmental challenges. MPs affect aquatic habitats, including physical and ecotoxicological effects on biota, whereas invading species are a key cause of global biodiversity decrease. In Europe, crayfish are among the most common freshwater invaders, leading to local ecological and economic impacts. Lake Maggiore, characterized by growing urbanization, industrial and touristic activities is an important survey area for evaluating the combined effects of these environmental pressures due to its vast hydrographic basin, able of conveying large quantities of MPs towards the lake. This study aimed to assess the presence of invasive crayfish in Lake Maggiore, their biometry and their MPs degree of bioaccumulation. Three invasive crayfish species (Faxonius limosus, Pacifastacus leniusculus, and Procambarus clarkii) were captured in the Swiss sector of the lake along the littoral of Locarno area through baited traps. They were divided by sex, weighed and sized and their belonging to the species were also confirmed through molecular approach. A pool of 30 individuals per species and sex divided into small, medium and large sizes was considered for the analyzes of the presence of MPs in the intestine. The results revealed MPs accumulation of 0.07 ± 0.06 items/specimen in P. clarkii, 0.20 ± 0.10 items/specimen in P. leniusculus, and 0.37 ± 0.31 items/specimen in F. limosus, the latter showing higher concentrations. Polyester and polyacrylate were the most common MPs in the three species. Thus, intestinal content may reflect the bioavailability of MPs in Lake Maggiore, giving an alternative to monitoring abiotic matrices while additionally supporting invasive species containment. More research is needed to discover the best crayfish species for detecting MPs pollution and to assess how much MPs move from the intestine to other organs, particularly muscle edible tissues used in the food market.

Contaminants of emerging concern (CECs) include a vast array of currently unregulated xenobiotics, including chemicals, their by-products. These contaminants are not necessarily newly synthetized, but their presence in natural ecosystems has only recently started to be investigated. Some CECs are currently under scrutiny for regulation because they have potential adverse effects on wild organisms and human health, including pharmaceuticals and personal care products (PPCPs), flame retardants (FRs), certain pesticides, plastic additives, per- and polyfluoroalkyl substances (PFASs), nanoparticles, and micro and nanoplastics. During the last years, the occurrence and distribution of certain CECs have been detected in the Mediterranean basin, one of most polluted marine regions, and their bioaccumulation and biomagnification have been documented. However, the studies on the trophic transfer of CECs in marine ecosystems have mainly focused on species belonging to low trophic levels. We propose the Scopoli's shearwaters (Calonectris diomedea), a top-predator in the Mediterranean marine trophic chain, as an excellent sentinel organism for biomonitoring spatial and temporal pattern of CECs in this area. We will focus on obtaining biological samples from shearwaters breeding at several colonies across most of the Mediterranean Sea, from Greece to Spain. The accumulation of different CECs, including PPCPs, FRs, plastic additives and PFASs will be assessed in the blood from fledglings exposed to local levels of contamination. These analyses will shed light on the spatial distribution of CECs and will act as a baseline for biomonitoring the fate of CECs in Mediterranean ecosystems.

This study aims to enhance our understanding of emerging pollutants and their effects on energy transfer within trophic networks. The main goal is to investigate how exposure to substances can impact the ecological mechanisms regulating biomass distribution across trophic levels, to predict their effects and manage their thresholds. Specifically, the study concentrates on the impact of Gadolinium (Gd) on autotrophic and heterotrophic organisms.

The selected taxa as models for both marine (Aliivibrio fischeri, Phaeodactylum tricornutum, Paracentrotus lividus) and freshwater ecosystems (Aliivibrio fischeri, Raphidocelis subcapitata, Daphia magna) represent the foundational elements of total productivity, reflecting the carbon content within the natural capital. Gadolinium is an external element primarily used in medical settings as a contrast agent for enhancing magnetic resonance imaging (MRI) and magnetic resonance angiography (MRA) efficacy. However, the increased Gd content in recent years, exacerbated by the Covid-19 pandemic, has prompted a need for further investigation.

This research aims to perform reliable predictions applicable to empirical models and meeting legal requirements for Gd distribution and permissible levels in marine and freshwater environments. Through assessing ecotoxicological responses and essential biometric characteristics under relevant environmental concentrations, organisms in mesocosms were exposed to known pollutant levels (ranging from 0.1 to 100 μg/L). Significant ecotoxicological effects were recorded at 100 μg/L, prompting additional concentration tests at higher levels (250-500-800 μg/L). Trophodynamic analysis revealed shifts in carbon content per unit and their implications for overall productivity. The outcomes of this study provide insights into how Gd influences biomass loss, subsequently impacting energy transfer between trophic levels.

The Mediterranean Sea is facing rising anthropogenic impacts. In addition to marine traffic, tourism, and industrial activities, the basin is also subject to significant riverine runoff that transports contaminants from cities and rural areas. The combination of these pressures and the basin's unique characteristics makes the Mediterranean an accumulation sink for numerous pollutants, including the new generation of compounds and molecules classified as emerging contaminants.

Among these, pharmaceutical and personal care products (PPCPs) and phenolic endocrine‐disrupting compounds (PEDCs) have gained attention for their designed bioactivity, and mutagenic and/or carcinogenic properties for organisms, posing a risk to marine ecosystems and human health. Despite this, studies reporting the occurrence and concentration of these pollutants in the Mediterranean marine environment are still lacking.

This study aimed to evaluate the presence of selected PPCPs and PEDCs in two Italian coastal areas with different anthropogenic pressures and exposure to pollutant sources: the Marine Protected Area of Rome Municipality ‘Secche di Tor Paterno‘, located in the Central Tyrrhenian Sea, few miles southern from the River Tiber mouth, and the popular summer tourist destination of Giglio Island (Tuscany Archipelago). The Mediterranean endemic seagrass Posidonia oceanica meadows develop on seabed shallower than 40 m in both areas. Surface seawater, sediment, and P. oceanica (rhizomes and leaves) were collected to investigate the occurrence of pollutants and evaluate the seagrass's potential application as a bioindicator of PEDCs and PPCPs contamination. Additionally, an ecological risk evaluation was performed based on the measured concentrations of these bioactive pollutants in the seawater. Results showed that PPCPs were present at higher concentrations than PEDCs in the study areas and bioaccumulated in P. oceanica, suggesting that this seagrass can be a suitable bioindicator of organic contamination. The risk analysis performed also indicated that the selected contaminants may pose a high risk to the marine ecosystem.

Worm movement depends on alternating waves of inhibitory and excitatory neurotransmitters, namely GABA and acetylcholine, at the neuromuscular junction. We have previously shown that short-term exposure to per- and polyfluoroalkyl substances, such as PFOA and the short chain perfluoropropylene oxide dimer acid (HFPO-DA), also known as GenX, can affect neurotransmitter levels and impair locomotion in the red worm E. fetida. In this work, we performed molecular docking analysis of a battery of 34 different PFAS congeners with the 3D structure of the human GABA-a​ receptor, showing that several structurally different PFAS have varying affinities for different binding sites and that the majority of PFAS congeners prefer the more spacious flumazenil site, while at the intracellular side, all molecules showed affinity towards the picrotoxin site. We selected a subset of mostly potentially active compounds and assessed their toxicological properties in terms of survival and GABA chloride channel modulation in a modified neuronal-like neuroblastoma cell line. Despite no acute toxicity and hormetic effects, several PFAS compounds could affect GABA chloride currents. PFOS was the most effective in reversibly suppressing GABA current, followed by PFMOBA, PFOA, CH3-PFO3-3-6-9-TriDoA, branched ADONA, and PFMOPrA. These findings highlight the complex interactions and potential neurotoxicity of various PFAS compounds, emphasizing the need for comprehensive risk assessment strategies. The One Health approach, considering the interconnected health of humans, animals, and ecosystems, is crucial for addressing the multifaceted impacts of PFAS contamination. Integrating environmental, biological, and chemical data is essential to develop effective policies and interventions to protect public health and the environment.