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Nanoplastics are considered contaminants of emerging concern at the global scale. The recent evidence of their occurrence in seawater from the Mediterranean Sea calls for a thorough evaluation of their impact on marine life and in particular on vulnerable life stages such as planktonic embryos. Here, we investigated the impact of increasing nominal concentrations of 50 nm amino-modified (PS-NH2) and 60 nm carboxy-modified (PS-COOH) polystyrene nanoparticles (PS NPs) on the embryonic development of the ascidian Ciona robusta (phylum Chordata), a common benthic invertebrate living in Mediterranean coastal areas with the peculiarity of being an early chordate developmental model. A strong agglomeration of PS-COOH (approx. 1 µm) was observed in natural sea water (NSW) already at time 0, while PS-NH2 resulted still monodispersed (approx. 130 nm) but largely aggregated after 22 h with a microscale dimension similar to those negatively charged. However, their effect on C. robusta embryos development largely differed at 22 h: PS-COOH did not affect larvae phenotypes nor their development, while PS-NH2 caused a dose-dependent effect (EC50 (22 h) of 7.52 μg mL−1) with various degrees of phenotype malformations (from mild to severe) and impairment of larval swimming. Embryos (up to 30%) exposed to 15 µg mL−1 PS-NH2 resulted not developed and the majority was unable to hatch. Calculated PS-NH2 EC50 resulted higher than those available for other marine invertebrate species, suggesting a protective role of the egg envelopes surrounding C. robusta embryos toward nanoplastics exposure.

Bacteria are an important component of the ecosystem, and microbial community alterations can have a significant effect on biogeochemical cycling and food webs. Toxicity testing based on microorganisms are widely used because they are relatively quick, reproducible, cheap, and are not associated with ethical issues. Here, we describe an ecotoxicological method to evaluate the biological response of the marine bacterium Vibrio anguillarum. This method assesses the acute toxicity of chemical compounds, including new contaminants such as nanoparticles, as well as environmental samples. The endpoint is the reduction of bacterial culturability (i.e., the capability to replicate and form colonies) due to exposure to a toxicant. This reduction can be generally referred to as mortality. The test allows for the determination of the LC50, the concentration that causes a 50% decrease of bacteria actively replicating and forming colonies, after a 6 h exposure. The culturable bacteria are counted in terms of colony forming units (CFU), and the "mortality" is evaluated and compared to the control. In this work, the toxicity of copper sulphate (CuSO4) was evaluated. A clear dose-response relationship was observed, with a mean LC50 of 1.13 mg/L, after three independent tests. This protocol, compared to existing methods with microorganisms, is applicable in a wider range of salinity and has no limitations for colored/turbid samples. It uses saline solution as the exposure medium, avoiding any possible interferences of growth medium with the investigated contaminants. The LC50 calculation facilitates comparisons with other bioassays commonly applied to ecotoxicological assessments of the marine environment.

Our research activities target the use of biological methods for the evaluation of environmental quality, with particular reference to saltwater/brackish water and sediment. The choice of biological indicators must be based on reliable scientific knowledge and, possibly, on the availability of standardized procedures. In this article, we present a standardized protocol that used the marine crustacean Artemia to evaluate the toxicity of chemicals and/or of marine environmental matrices. Scientists propose that the brine shrimp (Artemia) is a suitable candidate for the development of a standard bioassay for worldwide utilization. A number of papers have been published on the toxic effects of various chemicals and toxicants on brine shrimp (Artemia). The major advantage of this crustacean for toxicity studies is the overall availability of the dry cysts; these can be immediately used in testing and difficult cultivation is not demanded1,2. Cyst-based toxicity assays are cheap, continuously available, simple and reliable and are thus an important answer to routine needs of toxicity screening, for industrial monitoring requirements or for regulatory purposes3. The proposed method involves the mortality as an endpoint. The numbers of survivors were counted and percentage of deaths were calculated. Larvae were considered dead if they did not exhibit any internal or external movement during several seconds of observation4. This procedure was standardized testing a reference substance (Sodium Dodecyl Sulfate); some results are reported in this work. This article accompanies a video that describes the performance of procedural toxicity testing, showing all the steps related to the protocol.

Diatom-derived polyunsaturated aldehydes (PUAs), decadienal, heptadienal and octadienal, derive from the oxidation of fatty acids and have cytotoxic and anticancer effects. PUAs, tested separately, induce malformations in sea urchin Paracentrotus lividus embryos. Decadienal induces the worst malformations and lowest survival rates. Interestingly, decadienal, heptadienal and octadienal place in motion several genes to counteract their negative effects. To date, no studies are available reporting on the effects of PUA mixtures on marine invertebrates. Here we test binary and ternary mixtures on embryonic development of P. lividus. Our findings demonstrate that mixtures of PUAs act (i) at morphological level in synergistic way, being much more severe compared to individual PUAs; (ii) at molecular level also reveal an additive effect, affecting almost all fifty genes, previously tested using individual PUAs. This study is relevant from an ecological point of view since diatoms are a major food source for both pelagic and benthic organisms. This work opens new perspectives for understanding the molecular mechanisms that marine organisms use in reacting to environmental natural toxin mixtures such as diatom PUAs.