Quantitative assessment and monitoring of microplastics and nanoplastics distributions and lipid metabolism in live zebrafish using hyperspectral stimulated Raman scattering microscopy.

Microplastics (MP) and nanoplastics (NP) pollutions pose a rising environmental threat to humans and other living species, given their escalating presence in essential resources that living subjects ingest and/or inhale. Herein, to elucidate the potential health implications of MP/NP, we report for the first time by using label-free hyperspectral stimulated Raman scattering (SRS) imaging technique developed to quantitatively monitor the bioaccumulation and metabolic toxicity of MP/NP within live zebrafish larvae during their early developmental stages. Zebrafish embryos are exposed to environmentally related concentrations (3-60 µg/ml) of polystyrene (PS) beads with two typical sizes (2 µm and 50 nm). Zebrafish are administered isotope-tagged fatty acids through microinjection and dietary intake for in vivo tracking of lipid metabolism dynamics. In vivo 3D quantitative vibrational imaging of PS beads and intrinsic biomolecules across key zebrafish organs reveals that gut and liver are the primary target organs of MP/NP, while only 50 nm PS beads readily aggregate and adhere to the brain and blood vessels. The 50 nm PS beads are also found to induce more pronounced hepatic inflammatory response compared to 2 µm counterparts, characterized by increased biogenesis of lipid droplets and upregulation of arachidonic acid detected in zebrafish liver. Furthermore, Raman-tagged SRS imaging of fatty acids uncovers that MP/NP exposure significantly reduces yolk lipid utilization and promotes dietary lipid storage in zebrafish, possibly associated with developmental delays and more pronounced food dilution effects in zebrafish larvae exposed to 2 µm PS beads. The hyperspectral SRS imaging in this work shows that MP/NP exposure perturbs the development and lipid metabolism in zebrafish larvae, furthering the understanding of MP/NP ingestions and consequent toxicity in different organs in living species.

Improving preservation state assessment of carbonate microfossils in paleontological research using label-free stimulated Raman imaging.

In micropaleontological and paleoclimatological studies based on microfossil morphology and geochemistry, assessing the preservation state of fossils is of the highest importance, as diagenetic alteration invalidates textural features and compromises the correct interpretation of stable isotope and trace elemental analysis. In this paper, we present a novel non-invasive and label-free tomographic approach to reconstruct the three-dimensional architecture of microfossils with submicron resolution based on stimulated Raman scattering (SRS). Furthermore, this technique allows deciphering the three-dimensional (3D) distribution of the minerals within these fossils in a chemically sensitive manner. Our method, therefore, allows to identify microfossils, to chemically map their internal structure and eventually to determine their preservation state. We demonstrate the effectiveness of this method by analyzing several benthic and planktonic foraminifera, obtaining full 3D distributions of carbonate, iron oxide and porosity for each specimen. Subsequently, the preservation state of each microfossil can be assessed using these 3D compositional maps. The technique is highly sensitive, non-destructive, time-efficient and avoids the need for sample pretreatment. Therefore, its predestined application is the final check of the state of microfossils before applying subsequent geochemical analyses.

Assessment tools for microplastics and natural fibres ingested by fish in an urbanised estuary.

Microplastics and fibres occur in high concentrations along urban coastlines, but the occurrence of microplastic ingestion by fishes in these areas requires further investigation. Herein, the ingestion of debris (i.e., synthetic and natural fibres and synthetic fragments of various polymer types) by three benthic-foraging fish species Acanthopagrus australis (yellowfin bream), Mugil cephalus (sea mullet) and Gerres subfasciatus (silverbiddy) in Sydney Harbour, Australia has been quantified and chemically speciated by vibrational spectroscopy to identify the polymer type. Ingested debris were quantified using gut content analysis, and identified using attenuated total reflectance Fourier transform infrared (ATR-FTIR) and Raman microspectroscopies in combination with principal component analysis (PCA). The occurrence of debris ingestion at the time of sampling ranged from 21 to 64% for the three species, and the debris number ranged from 0.2 to 4.6 items per fish for the different species, with ∼53% of debris being microplastic. There was a significant difference in the amount of debris ingested among species; however, there was no difference among species when debris counts were standardised to fish weight or gut content weight, indicating that these species ingest a similar concentration of debris relative to their ingestion rate of other material. ATR-FTIR microspectroscopy successfully identified 72% of debris. Raman spectroscopy contributed an additional 1% of successful identification. In addition, PCA was used to non-subjectively classify the ATR-FTIR spectra resulting in the identification of an additional 9% of the debris. The most common microplastics found were polyester (PET), acrylic-polyester blend, and rayon (semi-synthetic) fibres. The potential of using Raman microspectroscopy for debris identification was investigated and provided additional information about the nature of the debris as well as the presence of specific dyes (and hence potential toxicity).