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ABC: microAlgae, Bivalves and Crustaceans as model organisms to assess transfer and effects of emerging contaminants through a marine food chain
No full textThousands of different natural and synthetic chemical compounds continuously enter aquatic ecosystems. Some of these compounds are emerging contaminants that include for instance agrochemicals, pharmaceuticals and personal care products, nanoparticles, per- and polyfluorinated compounds and plasticizers.
The present PhD thesis evaluated the effects of BPA analogues - a group of emerging contaminants - currently used to replace bisphenol A (BPA) which is largely used in polycarbonate plastic production, as well as in the production of thermal papers and food packaging. These new compounds are considered safer than BPA, even if they share a similar chemical structure. In this regard, the first chapter of the thesis (Chapter One) is a comprehensive review concerning BPA analogues, their presence in aquatic ecosystems and their biological effects on marine species.
The first aim of this research project was to assess the effects of BPA analogues on three different marine model species with a relevant ecological and economic value, and representative of three trophic levels, namely the microalgae Phaeodactylum tricornutum as a primary producer, the clam Ruditapes philippinarum as a primary consumer and the crab Carcinus aestuarii as a secondary consumer. In this project, we tested three different BPA analogues: bisphenol AF, bisphenol F and bisphenol S, which are the currently main substitutes for BPA. The effects of a mixture of these three BPA analogues were also evaluated as a more realistic scenario. The selected concentration was 300 ng/L for each single compound, while the mixture included 100 ng/L of each of the three components (300 ng/L in total).
The second aim of this research project was to improve knowledge concerning the potential bioaccumulation and transfer of BPA analogues through the marine food chain. For this reason, a brief marine food chain was simulated in laboratory conditions using the previously mentioned species. This project has been divided into two parts. In the first part (Chapter Two), each of the previously described marine species has been individually exposed to the three BPA analogues and their mixture. In the second part (Chapter Three), two trophic experiments were conducted, exposing clams or crabs to BPA analogue-contaminated diets. In detail, the lower trophic level was subject to waterborne exposure to BPA analogues and then provided as food for the upper trophic level. Several biomarkers regarding the antioxidant system, oxidative damage, immune system, epigenetic mechanisms and physiology were measured in each experiment. In addition, we evaluated the bioaccumulation of BPA analogues in the three species after exposure to contaminated seawater or contaminated food. Indeed, it is known that aquatic organisms can bioaccumulate contaminants from water or sediments and may transfer these contaminants to predators that forage on them.
Overall, the results obtained (Chapter Four) demonstrate that BPA analogues can cause toxic effects on all three tested species. In particular, BPA analogues were able to cause ROS-mediated toxicity increasing oxidative stress and oxidative damage in all the species, following both waterborne and foodborne exposures. Moreover, we observed alterations in the immune system and some physiological parameters. Such results agree with those of initial studies and demonstrate that these compounds can cause alterations similar to those already reported for BPA. As for bioaccumulation, it was observed that BPA analogues were bioaccumulated mainly following waterborne exposure. Lastly, the mixture of the three BPA analogues seems to be the worst experimental condition tested, in which synergic interactions between the three components probably occurred. In conclusion, based on the results reported in this thesis, it seems that BPA analogues are not harmless substitutes for BPA, being able to affect several biological parameters in the three marine species tested
DNA strand breaks in the crab Carcinus aestuarii exposed to three BPA analogues and their mixture
Full text linkIn this study, the genotoxic effects of three different bisphenols (BPAF, BPF and BPS) and their mixture were assessed in the crab Carcinus aestuarii. Crabs were exposed for 7 and 14 days to 300 ng/L of BPA analogues, alone or as a mixture (100 ng/L for each compound). After 7- and 14-day exposure, gills and hepatopancreas were sampled from crabs to evaluate damage to DNA by quantifying the levels of DNA single- and double-strand breaks. Results indicated that BPA analogues increased the DNA damage in both gills and hepatopancreas. In detail, BPAF, the most hydrophobic compound tested, exerted a marked effect on DNA. Overall, results suggest that such relatively new compounds can cause damage to DNA in crabs, raising doubts about the toxicological profile of BPA analogues, at least in the species studied and under the conditions tested in this study
Bisphenol Analogs in Aquatic Environments and Their Effects on Marine Species—A Review
Full text linkBisphenol A analogs are currently used in manufacturing and as plasticizers as a substitute
for bisphenol A. This replacement is taking place because bisphenol A is recognized as an endocrine
disruptor chemical (EDC) that can also cause oxidative stress and genotoxic effects in
aquatic species. Bisphenol A analogs have a similar chemical structure to BPA, raising doubts about
their use as safer substitutes. This review intends to summarize the concentrations of BPA analogs
found in aquatic environments and the effects of these emerging compounds on marine species.
Generally, studies indicate that BPA analogs have similar effects to their precursor, altering the neuroendocrine
system in several marine species. Furthermore, BPA analogs can cause oxidative stress
and developmental alterations. The available information on the biological effects of BPA analogs
suggests that more effort should be performed to assess the effects of these compounds in marine
organisms
Effects of acute exposure of Mytilus galloprovincialis to two neonicotinoid substances: Thiacloprid and Calypso
No full textNew insights into morpho-functional features of haemocytes from the blue crab Callinectes sapidus
No full textIn this study, we provided a comprehensive morpho-functional characterization of haemocytes in the blue crab Callinectes sapidus. Three haemocyte types were identified in the haemolymph: hyalinocytes (50 ± 4.7 %), lacking evident cytoplasmic granules; semigranulocytes (22.8 ± 2.02 %), containing a variable number of refractile granules; and granulocytes (27.2 ± 2 %), distinguished by their abundance of refractile granules. Haemocytes were predominantly oval or round. No significant size differences were observed among cell types, with granulocytes and semigranulocytes ranging from 7 to 22 μm, and hyalinocytes from 8 to 20 μm. Additionally, haemocytes were categorized into three cytochemical subpopulations: acidophils (38 %), basophils (36 %), and neutrophils (26 %). Notably, Neutral Red staining failed to reveal lysosomes in vivo, suggesting low membrane permeability under these conditions. Transmission electron microscopy corroborated the presence of the three haemocyte types. Both granulocytes and hyalinocytes exhibited phagocytic activity against yeast cells, although the phagocytic index remained low (∼4 %), implying that phagocytosis may not be the primary immune mechanism in C. sapidus. All haemocyte types generated superoxide anion and tested positive for several hydrolytic enzymes and phenoloxidase activity. Overall, these findings confirm the presence of three distinct haemocyte types in C. sapidus haemolymph and suggest that alternative immune pathways, beyond phagocytosis, may play a central role. Further research is needed to investigate additional immune functions, such as degranulation and inflammatory responses in C. sapidus
Effects of the Fragrance Galaxolide on the Biomarker Responses of the Clam Ruditapes philippinarum
Full text linkThe musk fragrance Galaxolide® (HHCB) is widely used in personal care and household products. Its large use leads to a continuous release of the compound into aquatic environments. Although some studies on the presence of HHCB in ecosystems and biota have been conducted, limited data about its effects on organism biomarkers are available. This study aimed at investigating both cellular and biochemical effects of HHCB in the clam Ruditapes philippinarum. Mussels were exposed for 7, 14 and 21 days to 100 ng/L and 500 ng/L of HHCB in seawater, and the effects on haemocyte parameters and antioxidant enzyme activities in the gills and digestive gland were evaluated. In addition, the neurotoxic potential of HHCB and its capacity to cause oxidative damage to proteins were assessed. Overall, our results demonstrated that exposure to HHCB was able to induce changes in biomarker responses of mussels, mainly at the cellular level
Effects of Three Widely Used Antibiotics and Their Mixture on the Haemocytes of the Clam Ruditapes philippinarum
Full text linkAlthough the presence of pharmaceutical and personal care products in aquatic
ecosystems is well documented, little information is available about their sublethal
effects, on aquatic invertebrates. From an ecotoxicological point of view, the use of
in vitro approaches has been recommended as a tool to assess adverse effects and to
understand the mechanisms of action of chemicals at the cellular level. In the present
in vitro study, the effects of Amoxicillin (AMX), Trimethoprim (TMP) and Ciprofloxacin (CIP)
(1 μg/L, each) were tested alone and—for the first time—as a mixture (MIX) on haemocytes
of the clam Ruditapes philippinarum. After the exposure, a battery of cellular parameters
was evaluated, such as haemocyte viability, lysosomal membrane stability, superoxide
anion production, acid phosphatase activity, the frequency of micronuclei and
chromosomal aberrations. The results demonstrated that AMX, TMP, CIP and MIX
affected lysosomal membrane stability, as well as superoxide anion and acid
phosphatase production, and promoted chromosomal aberrations. This study
highlighted that Manila clam haemocytes are a sensitive cell model to assess the
effects of exposure to pharmaceutical products on non-target species. Our study
demonstrated that the effects of pharmaceutical mixtures on marine species should be
experimentally evaluated because they are not predictable from single exposures as the
compounds can interact in different ways on the various biological endpoints considered
From a validated targeted method to a retrospective UHPLC-HRMS non-targeted analysis unveiling COVID-19-related contaminants in clams. Have we bias in marine model organisms for ecotoxicological studies?
Full text link: This study explores a retrospective non-targeted analysis (NTA), based on Ultra High-Performance Liquid Chromatography coupled to High-Resolution Mass Spectrometry (UHPLC-HRMS), to assess hidden chemicals of emerging concern (CECs) in marine model organisms. Conventional ecotoxicological studies do not include evaluating the natural habitats of the collected organisms, missing the possibility of highlighting unexpected pollutants, and thus compromising the correctness and reliability of the experimental results. In this paper we reprocessed samples previously collected from the Venice Lagoon for ecotoxicological studies and used for targeted analysis of three bisphenols-related compounds (i.e. BPS, BPF and BPAF) on seawater and specimens of the clam Ruditapes philippinarum. Results from the validation were the following: accuracy, expressed as percentage recoveries (R%), in the range 80%<120% for all the considered compounds and matrices, and precision, expressed as relative standard deviation of the absolute areas of IS, was <20% for clams (N=24) and <15% for seawater (N=30). LODs ranged from 5 to 50 ng/L for seawater and 5 to 12 ng/g for clam tissues. After validation, a retrospective NTA was carried out on control samples, showing the presence of some CECs, i.e. Lopinavir, Ritonavir, DEHS, and DEHA, two antiviral drugs used during the COVID-19 pandemic and two plasticizers, respectively, reported here for the first time in the Mediterranean Sea. CEC identifications were confirmed by matching MS/MS spectra with libraries. The present study emphasizes the importance of retrospective investigations to describe the contamination scenario of environmental matrices and the related effect in biota, to correctly address studies on model animals, also including possible "cocktail effects"
Effects of a mixture of glyphosate, 17α-ethynylestradiol and amyl salicylate on cellular and biochemical parameters of the mussel Mytilus galloprovincialis
No full textIn this study the effects of a mixture of glyphosate (herbicide), 17a-ethinylestradiol (synthetic estrogen) and amyl salicylate (fragrance) to the mussel Mytilus galloprovincialis were evaluated. Mussels were exposed for 7 days to two realistic concentrations of the mixture (10 and 100 ng/L) and the effects on total haemocyte counts, haemocyte diameter and volume, haemocyte proliferation, haemolymph lactate dehydrogenase activity and haemocyte lysate lysozyme activity were measured. In addition, superoxide dismutase, catalase, acetylcholinesterase, glutathione-S-transferase and glutathione reductase activities were measured in gills and digestive gland. The survival-in-air test was also performed. Results demonstrated that the mixture affected both cellular and biochemical biomarkers, but not tolerance to aerial exposure of M. galloprovincialis. The negative effects recorded in this study suggested that more efforts should be done to assess the ecotoxicological risks posed by contaminant mixture to aquatic invertebrates
Effects of Bisphenol A analogues and their mixture on the crab Carcinus aestuarii: Cytotoxicity, oxidative stress and damage, neurotoxicity, physiological responses, and bioaccumulation
Full text linkBisphenol A (BPA) analogues are emerging contaminants, whose ecotoxicological profile for aquatic species, particularly marine ones, is little known. In this study, the effects of an environmentally realistic concentration (300 ng/L) of three BPA analogues (BPAF, BPF, and BPS) - alone or as a mixture (MIX) – were evaluated for the first time on the crab Carcinus aestuarii. A multibiomarker approach was adopted to assess the effects of 7 and 14 days of exposure on haemolymph parameters, gill and hepatopancreas biochemical parameters, and physiological responses of crabs. Bioaccumulation of the three bisphenols was also investigated in crabs by UHPLC-HRMS. A significant reduction in total haemocyte counts was recorded in crabs exposed for 7 days to BPAF and MIX and for 14 days to the MIX, whereas an increase was found in crabs treated for 14 days with BPAF. Cell proliferation increased significantly in crabs exposed for 14 days to BPS and MIX. An imbalance of the antioxidant system, as well as oxidative damage, was recorded in gills and hepatopancreas. No neurotoxic effects were observed in crabs. At the physiological level, exposure to MIX increased the respiration rate of crabs. As for bioaccumulation, only bisphenol AF was detected in crabs. Overall, the present study demonstrated that BPA analogues can affect some important cellular parameters, induce oxidative stress and alter physiological responses in crabs
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