Phthalates and fatty acid markers in free-ranging cetaceans from an insular oceanic region: Ecological niches as drivers of contamination.

Plastic additives, such as phthalates, are ubiquitous contaminants that can have detrimental impacts on marine organisms and overall ecosystems' health. Valuable information about the status and resilience of marine ecosystems can be obtained through the monitoring of key indicator species, such as cetaceans. In this study, fatty acid profiles and phthalates were examined in blubber biopsies of free-ranging individuals from two delphinid species (short-finned pilot whale - Globicephala macrorhynchus, n = 45; common bottlenose dolphin - Tursiops truncatus, n = 39) off Madeira Island (NE Atlantic). This investigation aimed to explore the relations between trophic niches (epipelagic vs. mesopelagic), contamination levels, and the health status of individuals within different ecological and biological groups (defined by species, residency patterns and sex). Multivariate analysis of selected dietary fatty acids revealed a clear niche segregation between the two species. Di-n-butylphthalate (DBP), diethyl phthalate (DEP), and bis(2-ethylhexyl) phthalate (DEHP) were the most prevalent among the seven studied phthalates, with the highest concentration reached by DEHP in a bottlenose dolphin (4697.34 ± 113.45 ng/g). Phthalates esters (PAEs) concentration were higher in bottlenose dolphins (Mean ∑ PAEs: 947.56 ± 1558.34 ng/g) compared to pilot whales (Mean ∑ PAEs: 229.98 ± 158.86 ng/g). In bottlenose dolphins, DEHP was the predominant phthalate, whereas in pilot whales, DEP and DBP were more prevalent. Health markers suggested pilot whales might suffer from poorer physiological conditions than bottlenose dolphins, although high metabolic differences were seen between the two species. Phthalate levels showed no differences by ecological or biological groups, seasons, or years. This study is the first to assess the extent of plastic additive contamination in free-ranging cetaceans from a remote oceanic island system, underscoring the intricate relationship between ecological niches and contaminant exposure. Monitoring these chemicals and their potential impacts is vital to assess wild population health, inform conservation strategies, and protect critical species and habitats.

Optimization and validation of a micro-QuEChERS method for phthalates detection in small samples of cetacean blubber.

In this study, an innovative method was developed to detect and quantify phthalates in fresh cetacean blubber. An adaptation of the ammonium formate QuEChERS method was used and adapted as a micro-extraction for small quantities of samples. Significantly, this technique utilized minimal quantities of reagents and salts, with the additional implementation of rigorous Quality Assurance/Quality Control protocols to further reduce background contamination. To ensure the reliability of this method, comprehensive validation procedures were conducted, with a specific focus on two widely studied cetacean species: the common bottlenose dolphin (Tursiops truncatus) and the short-finned pilot whale (Globicephala macrorhynchus). Determination coefficients (R2) for matrix-matched calibration were >0.93 with limits of quantifications (LOQ) of the method in the range of 5-10 ng/g. Mean recovery values were between 40 and 100 %. This novel methodology holds particular relevance for environmental research studies, offering the capability to detect emerging contaminants with minimal sample requirements. This aspect is particularly valuable in investigations that involve free-ranging animals and rely on biopsy sampling. It allows for the assessment of contaminant levels in healthy individuals within wild populations, enhancing our understanding of ecological impacts and potential conservation measures.•A micro-extraction adaptation of the ammonium formate QuEChERS method was developed and applied to a small quantity of fresh cetacean blubber to detect phthalates.•Small quantities of reagents and salts were used, and additional Quality Assurance/ Quality Control procedures were taken to further minimize background contamination.•Method validation was carried out for two cosmopolitan and extensively studied cetacean species: the common bottlenose dolphin (Tursiops truncatus) and the short-finned pilot whale (Globicephala macrorhynchus).

Microplastic ingestion and plastic additive detection in pelagic squid and fish: Implications for bioindicators and plastic tracers in open oceanic food webs.

The ubiquitous presence of microplastics (MPs) in the ocean represents a potential threat to marine organisms, with poorly understood long-term adverse effects, including exposure to plastic additives. The present study investigated the ingestion of MPs in two epipelagic fish species (Trachurus picturatus and Scomber colias) and three pelagic squid species (Loligo vulgaris, Ommastrephes caroli and Sthenoteuthis pteropus) from an open oceanic region of the Northeast Atlantic. Seven phthalate esters (PAEs) were also analysed in the organisms' tissue, and the potential correlation between PAEs concentrations and ingested MPs was investigated. Seventy-two fish and 20 squid specimens were collected and analysed. MPs were found in the digestive tract of all species and in the squid species' gills and ink sacs. The highest occurrence of MPs was in the stomach of S. colias (85 %) and the lowest in the stomach and ink sac of O. caroli and L. vulgaris (12 %). Most of the particles identified (>90 %) were fibres. Among all the ecological and biological factors considered (dietary preferences, season, body size, total weight, liver weight, hepatosomatic index and gastrosomatic index), only gastrosomatic index (GSI) and season were significant predictors of MPs ingestion in fish species, with a greater likelihood of ingestion in the cold season and in specimens with higher GSI values (i.e. higher feeding intensity). Four PAEs (DEP, DIBP, BBP, DEHP) were detected in all the species analysed, with average ∑PAEs concentrations ranging between 10.31 and 30.86 ng/g (wet weight). DIBP was positively correlated with ingested MPs, suggesting this compound might represent a "plastic tracer". This study looks into the problem of MPs ingestion for pelagic species in an open oceanic region, highlighting the most suitable bioindicators and providing essential insights into the factors that may influence ingestion rates. Additionally, the detection of PAEs in all species indicates the need for further research on the contamination sources, the effects of these chemicals on marine organisms, and the potential risks to human health through seafood consumption.

Seasonal variation in microplastics and zooplankton abundances and characteristics: The ecological vulnerability of an oceanic island system.

The ingestion of microplastics (MPs - plastic particles <5 mm) by planktivorous organisms represents a significant threat to marine food webs. To investigate how seasonality might affect plastic intake in oceanic islands' ecosystems, relative abundances and composition of MPs and mesozooplankton samples collected off Madeira Island (NE Atlantic) between February 2019 and January 2020 were analysed. MPs were found in all samples, with fibres accounting for 89 % of the particles. MPs and zooplankton mean abundance was 0.262 items/m3 and 18.137 individuals/m3, respectively. Their monthly variations follow the seasonal fluctuation of environmental parameters, such as currents, chlorophyll-a concentration, sea surface temperature and precipitation intensity. A higher MPs/zooplankton ratio was recorded in the warm season (May-Oct), reaching 0.068 items/individual when considering large-sized particles (1000-5000 µm). This is the first study to assess the seasonal variability of MPs in an oceanic island system providing essential information respecting its ecological impact in pelagic environments.

Determination of phthalic acid esters and di(2-ethylhexyl) adipate in fish and squid using the ammonium formate version of the QuEChERS method combined with gas chromatography mass spectrometry.

In the present study, the ammonium formate version of the QuEChERS method, considered highly advantageous in relation to instrument maintenance and other issues, was applied for the first time to extract a group of twelve phthalic acid esters (PAEs, i.e. dipropyl phthalate, DPP; diisobutyl phthalate, DIBP; dibutyl phthalate, DBP; diisopentyl phthalate, DIPP; di-n-pentyl phthalate, DNPP; dihexyl phthalate, DHP; butyl benzyl phthalate, BBP; dicyclohexyl phthalate, DCHP; di(2-ethylhexyl) phthalate, DEHP; di-n-octyl phthalate, DNOP; diisononyl phthalate, DINP; and diisodecyl phthalate, DIDP) and one adipate (di(2-ethylhexyl) adipate, DEHA) from two species of fish (Scomber colias and Katsuwonus pelamis) and one of squid (Loligo gahi). The method was validated in terms of linearity, trueness and matrix effects. Determination coefficients (R2) for matrix-matched calibration curves were higher than 0.99 in all cases, being the lowest calibration levels in the range 0.5-10 ng/g. Mean recovery values were between 70 and 117% with relative standard deviation values ≤20%. Matrix effects were soft (between -20 and +20%) for most analytes and matrices, except in squid samples, which was mostly medium with a moderate ion suppression. The analysis of 10 samples of each type showed the presence of DIBP, DBP and DEHP at concentrations up to 44.2 ± 2.1 ng/g of wet weight in some of the samples and species, still not representing concerning values when considering the daily intake of such species of seafood in the human diet (tolerable daily intake -TDI- values were not exceeded). Results demonstrated that the ammonium formate version of the QuEChERS method can be applied with success for the extraction and determination of the selected PAEs and DEHA in fish and squid samples.

Large-scale movements of common bottlenose dolphins in the Atlantic: dolphins with an international courtyard.

Wide-ranging connectivity patterns of common bottlenose dolphins (Tursiops truncatus) are generally poorly known worldwide and more so within the oceanic archipelagos of Macaronesia in the North East (NE) Atlantic. This study aimed to identify long-range movements between the archipelagos of Macaronesia that lie between 500 and 1,500 km apart, and between Madeira archipelago and the Portuguese continental shelf, through the compilation and comparison of bottlenose dolphin's photo-identification catalogues from different regions: one from Madeira (n = 363 individuals), two from different areas in the Azores (n = 495 and 176), and four from different islands of the Canary Islands (n = 182, 110, 142 and 281), summing up 1791 photographs. An additional comparison was made between the Madeira catalogue and one catalogue from Sagres, on the southwest tip of the Iberian Peninsula (n = 359). Results showed 26 individual matches, mostly between Madeira and the Canary Islands (n = 23), and between Azores and Madeira (n = 3). No matches were found between the Canary Islands and the Azores, nor between Madeira and Sagres. There were no individuals identified in all three archipelagos. The minimum time recorded between sightings in two different archipelagos (≈ 460 km apart) was 62 days. Association patterns revealed that the individuals moving between archipelagos were connected to resident, migrant and transient individuals in Madeira. The higher number of individuals that were re-sighted between Madeira and the Canary Islands can be explained by the relative proximity of these two archipelagos. This study shows the first inter-archipelago movements of bottlenose dolphins in the Macaronesia region, emphasizing the high mobility of this species and supporting the high gene flow described for oceanic dolphins inhabiting the North Atlantic. The dynamics of these long-range movements strongly denotes the need to review marine protected areas established for this species in each archipelago, calling for joint resolutions from three autonomous regions belonging to two EU countries.