Toxicity assessment of animal manure composts containing environmental microplastics by using earthworms Eisenia andrei.

Nowadays, animal manure composting constitutes a sustainable alternative for farmers to enhance the level of nutrients within soils and achieve a good productivity. However, pollutants may be present in manures. This study focuses on the detection of environmental microplastics (EMPs) into composts, as well as on the assessment of their potential toxicity on the earthworm Eisenia andrei. To these aims, animals were exposed to two types of compost, namely bovine (cow) and ovine (sheep) manure, besides to their mixture, for 7 and 14 days. The presence and characterization of EMPs was evaluated in all the tested composts, as well as in tissues of the exposed earthworms. The impact of the tested composts was assessed by a multi-biomarker approach including cytotoxic (lysosomal membrane stability, LMS), genotoxic (micronuclei frequency, MNi), biochemical (activity of catalase, CAT, and glutathione-S-transferase, GST; content of malondialdehyde, MDA), and neurotoxic (activity of acetylcholinesterase, AChE) responses in earthworms. Results indicated the presence of high levels of EMPs in all the tested composts, especially in the sheep manure (2273.14 ± 200.89 items/kg) in comparison to the cow manure (1628.82 ± 175.23 items/kg), with the size <1.22 µm as the most abundant EMPs. A time-dependent decrease in LMS and AChE was noted in exposed earthworms, as well as a concomitant increase in DNA damages (MNi) after 7 and 14 days of exposure. Also, a severe oxidative stress was recorded in animals treated with the different types of compost through an increase in CAT and GST activities, and LPO levels, especially after 14 days of exposure. Therefore, it is necessary to carefully consider these findings for agricultural good practices in terms of plastic mitigation in compost usage, in order to prevent any risk for environment health.

Microplastics in fillets of Mediterranean seafood. A risk assessment study.

Microplastics (MPs) are considered as emergent threat to human health. No complete data still exists on MPs presence in fish tissue and their transmission to humans. The present study aims to detect and quantify the presence of MPs (<3 µm) in several edible seafood (Sardina pilchardus, wild and farmed Sparus aurata, Mullus surmuletus, Solea solea and musselMytilus galloprovincialis) from the south coast of Mediterranean Sea. MPs were detected through an innovative extraction method coupled to the SEM-EDX technology. The Estimated Daily Intakes (EDIs) for adults and children for each species were calculated. The higher median level (IQR) of MPs (9.09E+04) was found inM. surmuletus. Conversely, the lower median (IQR) level was observed in S. pilchardus (7.04E+04). The smallest and biggest median (IQR)diameter of MPs (1.8 and 2.5 µm) were identified in M. galloprovincialisand S. solea, respectively. The highest EDIs (25.50E+03; 48.09E+03) arefor ingestion of farmedS. auratarespectively for adults and children. Instead, the lowest EDIs (2.37E+02; 4.48E+02) are due to M. galloprovincialisingestion for adults and children, respectively. Our data should be carefully considered in view of the direct exposure of humans to plastic particles under 3 µm through seafood consumption to better manage the related risks.

Assessment of heavy metal pollution transfer and human exposure risks from the consumption of chicken grown in mining-surrounding areas.

The purpose of this study was to assess heavy metal contamination in soil, plants, earthworms, and chicken in farmlands adjacent to an old mining site and to evaluate the potential exposure risks to humans through the consumption of chicken. For this purpose, soil, earthworms, plant, chickens, and eggs were sampled from 5 sites following a gradient of contamination. All samples were analyzed for heavy metals (Pb, Cd, Cu, and Zn). A food chain model was used in order to characterize heavy metal transfer between soil-plant-earthworm and chicken organs. Furthermore, target hazard quotient (THQ), estimated daily intake (EDI), and hazard index (HI) were employed to assess human health risks posed by heavy metal contamination. Despite the higher level of Pb, our data related to the calculation of EDI and THQ suggested that local consumers are more at risk of Cd contamination. The calculated HI showed values ranging from 2.58 to 4.74 for adults, and up to 12.34 for children, indicating a considerable risk to the health of local inhabitants, especially children. This study highlighted the crucial role of diets based on chickens grown in contaminated areas, on health risks especially for children.

Micro- and nano-plastics in edible fruit and vegetables. The first diet risks assessment for the general population.

Microplastics (MPs) represent a current public health concern since toxicity has not yet fully investigated. They were found in several foods, but to the best of our knowledge, at this time no data was reported for the edible vegetables and fruits. We focused on diet exposure aiming to evaluate the number and the size (<10 µm) of MPs in the most commonly consumed vegetables and fruits, in relation to their recommended daily intake too. MPs extraction and analysis were carried out using an innovative Italian methodology and SEM-EDX, respectively. Finally, we calculated the Estimated Daily Intakes (EDIs) for adults and children for each type of vegetal and fruit. The higher median (IQR) level of MPs in fruit and vegetable samples was 223,000 (52,600-307,750) and 97,800 (72,175-130,500), respectively. In particular, apples were the most contaminated fruit samples, while carrot was the most contaminated vegetable. Conversely, the lower median (IQR) level was observed in lettuce samples 52,050 (26,375-75,425). Both vegetable and fruit samples MPs levels were characterized by wide variability. The smallest size of MPs was found in the carrot samples (1.51 µm), while the biggest ones were found in the lettuce (2.52 µm). Both vegetable and fruit samples had size of the MPs characterized by low variability. We found the highest median level of MPs in samples purchased from the "fruiter 3" (124,900 p/g) and the lowest in those purchased in "supermarket" (87,600 p/g). The median size of the MPs had overlapping dimensions in all the purchase sites, with the exception of the samples purchased at the "shop at km zero 2″ which had slightly smaller size (1.81 µm). The highest adults' (4.62 E+05) and children's (1.41 E+06) EDIs are due the ingestion of apples, instead the lowest are due to the ingestion of carrots (adults: 2.96 E+04; children: 1.15 E+05). We hypothesized that the mechanism of uptake and translocation of MPs can be the same described and reported for carbon-nanomaterials. This may be a possible translocation route of MPs by environment to vegetables permitting, so, the translocation or uptake inside of their biological systems. Based on the results obtained it is urgent important to perform toxicological and epidemiological studies to investigate for the possible effects of MPs on human health.

Interactions of a pesticide/heavy metal mixture in marine bivalves: a transcriptomic assessment.

BACKGROUND: Mixtures of chemicals present in aquatic environments may elicit toxicity due to additive or synergistic effects among the constituents or, vice versa, the adverse outcome may be reduced by antagonistic interactions. Deviations from additivity should be explained either by the perturbations of toxicokinetic parameters and/or chemical toxicodynamics. We addressed this important question in marine mussels exposed subchronically to a binary mixture made of two wide-spread pollutants: the heavy metal nickel and the organic phosphorus pesticide Chlorpyrifos. To this aim, we carried out in tissues of Mytius galloprovincialis (Lam) a systems approach based on the evaluation and integration of different disciplines, i.e. high throughput gene expression profiling, functional genomics, stress biomakers and toxicokinetics. RESULTS: Cellular and tissue biomarkers, viz. digestive gland lysosomal membrane stability, lysosomal/cytosol volume ratio, neutral lipid content and gill acetylcholinesterase activity were, in general, altered by either the exposure to nickel and Chlorpyrifos. However, their joint action rendered (i) an overall decrease of the stress syndrome level, as evaluated through an expert system integrating biomarkers and (ii) statistically significant antagonistic deviations from the reference model systems to predict mixture toxicity. While toxicokinetic modeling did not explain mixture interactions, gene expression profiling and further Gene Ontology-based functional genomics analysis provided clues that the decrement of toxicity may arise from the development of specific toxicodynamics. Multivariate statistics of microarray data (238 genes in total, representing about 14% of the whole microarray catalogue) showed two separate patterns for the single chemicals: the one belonging to the heavy metal -135 differentially expressed genes (DEGs) was characterized by the modulation of transcript levels involved in nucleic acid metabolism, cell proliferation and lipid metabolic processes. Chlorpyrifos exposure (43 DEGs) yielded a molecular signature which was biased towards carbohydrate catabolism (indeed, chitin metabolism) and developmental processes. The exposure to the mixture (103 DEGs) elicited a composite complex profile which encompassed the core properties of the pesticide but also a relevant set of unique features. Finally, the relative mRNA abundance of twelve genes was followed by Q-PCR to either confirm or complement microarray data. These results, in general, were compatible with those from arrays and indeed confirmed the association of the relative abundance of two GM-2 ganglioside activator genes in the development of the hyperlipidosis syndrome observed in digestive gland lysosomes of single chemical exposed mussels. CONCLUSION: The transcriptomic assessment fitted with biological data to indicate the occurrence of different toxicodynamic events and, in general, a decrease of toxicity, driven by the mitigation or even abolition of lysosomal responses. Furthermore, our results emphasized the importance of the application of mechanistic approaches and the power of systems assessment to study toxicological responses in ecologically relevant organisms.

Mixture toxicity assessment of nickel and chlorpyrifos in the sea bass Dicentrarchus labrax.

The present research work was designed to study Dicentrarchus labrax biotransformation and detoxification responses to acute exposure to nickel (Ni) and chlorpyrifos (CHP). Sexually immature sea bass were treated by intraperitoneal injection of nickel chloride (500 µg kg⁻¹), chlorpyrifos (10 mg kg⁻¹), and their binary mixture for 1, 3, and 7 days. Ni and CHP accumulation was quantified in liver after the exposure periods. The following biological responses were measured: (1) NADPH cytochrome P450 reductase (NCR) activity, as phase I biotransformation parameter; (2) gluthathione S-transferase (GST) activity as a phase II conjugation enzyme, acetylcholinesterase activity, and metallothionein (MT) content. Ni bioaccumulation in the liver resulted in an increasing uptake up to 15.48 µg g⁻¹ wet weight (Ni-treated animals) and 16.73 µg g⁻¹ wet weight (mixture-treated animals) after 7 days of exposure. CHP accumulation showed a distinct pattern in animals exposed to the mixture of chemicals in comparison with CHP-treated animals. NCR activity exhibited a marked activation in CHP and mixture-treated animals. GST activity was significantly increased starting from 1 day exposure in CHP-treated animals and after 3 days in Ni-treated animals. MT accumulation increased in all conditions, with a marked synergetic effect after 7 days of exposure. These data should be carefully considered in view of the biological effects of mixture pollutants, particularly in fish farming conditions.

Metallothionein induction by Cu, Cd and Hg in Dicentrarchus labrax liver: assessment by RP-HPLC with fluorescence detection and spectrophotometry.

Metallothionein was quantified in sea bass Dicentrarchus labrax intraperitoneally (i.p.) injected with different Cu, Cd and Hg doses (50-250 microg kg(-1) wet wt) after 48 h exposure. A distinct peak with 16.8 min retention time was obtained by reversed-phase high performance liquid chromatography coupled to fluorescence detection (RP-HPLC-FD) with the three metals. Total metallothionein levels assayed in unheated liver extracts by RP-HPLC-FD were significantly higher (1.3-1.95-fold) than those obtained by the well-established spectrophotometric method. In the RP-HPLC-FD method, metallothionein increased linearly with Cu and Hg doses, being saturated beyond 100 mug kg(-1) Cd. Maximum induction was obtained at 100 microg kg(-1) Cd (5.3-fold), and 250 microg kg(-1) Cu or Hg (8- and 5.1-fold, respectively). At low doses no metallothionein induction was shown by the less sensitive spectrophotometric assay.