Nontarget and suspect screening reveals the presence of multiple plastic-related compounds in polar bear, killer whale, narwhal and long-finned pilot whale blubber from East Greenland.

The monitoring of legacy contaminants in sentinel northern marine mammals has revealed some of the highest concentrations globally. However, investigations into the presence of chemicals of emerging Arctic concern (CEACs) and other lesser-known chemicals are rarely conducted, if at all. Here, we used a nontarget/suspect approach to screen for thousands of different chemicals, including many CEACs and plastic-related compounds (PRCs) in blubber/adipose from killer whales (Orcinus orca), narwhals (Monodon monoceros), long-finned pilot whales (Globicephala melas), and polar bears (Ursus maritimus) in East Greenland. 138 compounds were tentatively identified mostly as PRCs, and four were confirmed using authentic standards: di(2-ethylhexyl) phthalate (DEHP), diethyl phthalate (DEP), di(2-propylheptyl) phthalate (DPHP), and one antioxidant (Irganox 1010). Three other PRCs, a nonylphenol isomer, 2,6-di-tert-butylphenol, and dioctyl sebacate, exhibited fragmentation patterns matching those in library databases. While phthalates were only above detection limits in some polar bear and narwhal, Irganox 1010, nonylphenol, and 2,6-di-tert-butylphenol were detected in >50% of all samples. This study represents the first application of a nontarget/suspect screening approach in Arctic cetaceans, leading to the identification of multiple PRCs in their blubber. Further nontarget analyses are warranted to comprehensively characterize the extent of CEAC and PRC contamination within Arctic marine food webs.

Development and validation of a modified QuEChERS method for extracting polychlorinated biphenyls and organochlorine pesticides from marine mammal blubber.

The monitoring of legacy persistent organic pollutants (POPs) in blubber of key sentinel marine mammal species has been conducted using established techniques for decades. Although these methods for polychlorinated biphenyl (PCB) and organochlorine (OC) pesticide determination provide accurate and reproducible results, they possess some drawbacks in terms of cost, time, and a need for large volumes of toxic solvents. QuEChERS (quick, easy, cheap, effective, rugged, and safe) extractions may help address these issues, but have not been applied to marine mammal blubber/adipose. As such, our aim was to develop, validate, and apply a QuEChERS method for the extraction of PCB and OC contaminants in marine mammal blubber. First, we tested multiple solid-phase extraction and clean-up steps to find the approach that provided the cleanest extracts along with consistent and acceptable analyte recovery, accuracy, and precision. QuEChERS extractions followed by two enhanced matrix removal-lipid (EMR-lipid), one primary-secondary amine (PSA), and one silica gel clean-up showed the highest matrix removal and acceptable recoveries of spiked internal (62-97%) and external standards (61-94%). Solvent usage was reduced by ∼393% and extraction time was reduced by ∼25% (from 16 to 12 h). Next, the method was validated using standard reference material (SRM) NIST 1945. Recovery experiments on SRM (n = 5) showed acceptable recovery for 76% and 77% of PCBs and OC pesticides, respectively, and high precision for 73% and 69% of PCBs and OCs, respectively. Finally, the method was used on a set of southeast Greenland killer whales (n = 13), with previously published PCB and OC data. Bland-Altman plots indicated good agreement between QuEChERS and current-use methods for ΣPCBs and some OCs with no significant constant or proportional bias. These results demonstrate that this QuEChERS extraction method represents an effective, lower cost alternative to current-use extractions for PCBs and OCs in blubber, and likely other high-lipid samples.

Validation of quantitative fatty acid signature analysis for estimating the diet composition of free-ranging killer whales.

Accurate diet estimates are necessary to assess trophic interactions and food web dynamics in ecosystems, particularly for apex predators like cetaceans, which can regulate entire food webs. Quantitative fatty acid analysis (QFASA) has been used to estimate the diets of marine predators in the last decade but has yet to be implemented on free-ranging cetaceans, from which typically only biopsy samples containing outer blubber are available, due to a lack of empirically determined calibration coefficients (CCs) that account for fatty acid (FA) metabolism. Here, we develop and validate QFASA for killer whales using full blubber from managed-care and free-ranging individuals. First, we compute full, inner, and outer blubber CCs from the FA signatures across the blubber layers of managed-care killer whales and their long-term diet items. We then run cross-validating simulations on the managed-care individuals to evaluate the accuracy of diet estimates by comparing full-depth and depth-specific estimates to true diets. Finally, we apply these approaches to subsistence-harvested killer whales from Greenland to test the utility of the method for free-ranging killer whales, particularly for the outer blubber. Accurate diet estimates for the managed-care killer whales were only achieved using killer whale-specific and blubber-layer-specific CCs. Modeled diets for the Greenlandic killer whales largely consisted of seals (75.9 ± 4.7%) and/or fish (20.4 ± 2.4%), mainly mackerel, which was consistent with stomach content data and limited literature on this population. Given the remote habitats and below surface feeding of most cetaceans, this newly developed cetacean-specific QFASA method, which can be applied to outer-layer biopsies, offers promise to provide a significant new understanding of diet dynamics of free-ranging odontocetes and perhaps other cetacean species throughout the world's oceans.

Microplastic ingestion by quagga mussels, Dreissena bugensis, and its effects on physiological processes.

The impacts of microplastic particulates in benthic freshwater organisms have been largely unexplored despite abundant plastic accumulation in the sediments of these systems. We investigated the uptake of plastic particles by benthic filter feeding quagga mussels (Dreissena bugensis) and associated toxicity exhibited through impacts on mortality, filtration rate, reproduction and oxygen consumption. Matrix Assisted Laser Desorption/Ionization Imaging Mass Spectrometry (MALDI-IMS) technology was used to assess the microplastic inclusion. For this purpose, quagga mussels were exposed to four treatments ranging from 0.0 to 0.8 g/L of a high density fluorescent red polyethylene powder in the size range of 10-45 µm for 24-h, and the targeted endpoints were quantified. Identification of several micrograms of microplastics in the digestive tract suggests rapid clearance from the water column by filtering. At the higher concentrations, about 95% of the microplastics ingested remained in the mussels after 24-h. Microplastics were found in the gills which correlated with decreasing filtration rate at higher microplastic concentrations. Despite large-scale ingestion, plastic exposure did not affect survivorship, reproduction rates, or oxygen consumption in the period examined. MALDI-IMS identified unique mass spectra that correlated with microplastic inclusion. This research suggests that microplastics can impair feeding through decreased filtration rates of filter feeding organisms, potentially resulting in a reduction of overall fitness over time and that MALDI-IMS may have the potential to identify microplastics and changes in tissue at the borders of plastic inclusion.

Nanoplastics impact the zebrafish (Danio rerio) transcriptome: Associated developmental and neurobehavioral consequences.

Microplastics (MPs) are a ubiquitous pollutant detected not only in marine and freshwater bodies, but also in tap and bottled water worldwide. While MPs have been extensively studied, the toxicity of their smaller counterpart, nanoplastics (NPs), is not well documented. Despite likely large-scale human and animal exposure to NPs, the associated health risks remain unclear, especially during early developmental stages. To address this, we investigated the health impacts of exposures to both 50 and 200 nm polystyrene NPs in larval zebrafish. From 6 to 120 h post-fertilization (hpf), developing zebrafish were exposed to a range of fluorescent NPs (10-10,000 parts per billion). Dose-dependent increases in accumulation were identified in exposed larval fish, potentially coinciding with an altered behavioral response as evidenced through swimming hyperactivity. Notably, exposures did not impact mortality, hatching rate, or deformities; however, transcriptomic analysis suggests neurodegeneration and motor dysfunction at both high and low concentrations. Furthermore, results of this study suggest that NPs can accumulate in the tissues of larval zebrafish, alter their transcriptome, and affect behavior and physiology, potentially decreasing organismal fitness in contaminated ecosystems. The uniquely broad scale of this study during a critical window of development provides crucial multidimensional characterization of NP impacts on human and animal health.

Debugging diversity - a pan-continental exploration of the potential of terrestrial blood-feeding leeches as a vertebrate monitoring tool.

The use of environmental DNA (eDNA) has become an applicable noninvasive tool with which to obtain information about biodiversity. A subdiscipline of eDNA is iDNA (invertebrate-derived DNA), where genetic material ingested by invertebrates is used to characterize the biodiversity of the species that served as hosts. While promising, these techniques are still in their infancy, as they have only been explored on limited numbers of samples from only a single or a few different locations. In this study, we investigate the suitability of iDNA extracted from more than 3,000 haematophagous terrestrial leeches as a tool for detecting a wide range of terrestrial vertebrates across five different geographical regions on three different continents. These regions cover almost the full geographical range of haematophagous terrestrial leeches, thus representing all parts of the world where this method might apply. We identify host taxa through metabarcoding coupled with high-throughput sequencing on Illumina and IonTorrent sequencing platforms to decrease economic costs and workload and thereby make the approach attractive for practitioners in conservation management. We identified hosts in four different taxonomic vertebrate classes: mammals, birds, reptiles and amphibians, belonging to at least 42 different taxonomic families. We find that vertebrate blood ingested by haematophagous terrestrial leeches throughout their distribution is a viable source of DNA with which to examine a wide range of vertebrates. Thus, this study provides encouraging support for the potential of haematophagous terrestrial leeches as a tool for detecting and monitoring terrestrial vertebrate biodiversity.