Hidden aliens: Application of digital PCR to track an exotic foraminifer across the Skagerrak (North Sea) correlates well with traditional morphospecies analysis.

The problem of invasive species is a well-studied one, but knowledge of free-living unicellular eukaryotic invasive species is lacking. A potentially invasive foraminifer (Rhizaria), Nonionella sp. T1, was recently discovered in the Skagerrak and its fjords. Digital polymerase chain reaction (dPCR) was applied to track the spread of this non-indigenous species using a new dPCR assay (T1-1). The use of dPCR appears highly complementary to traditional hand picking of foraminiferal shells from the sediment, and is far less time-consuming. This study indicates that Nonionella sp. T1 has bypassed the outer Skagerrak strait, instead becoming established in Swedish west coast fjords, constituting up to half of the living foraminiferal community in fjord mouth areas. The ecology of Nonionella sp. T1 and its potential invasive impacts are still largely unknown, but it appears to be an opportunist using several energy sources such as nitrate respiration and kleptoplasty along with a possibly more efficient reproductive strategy to gain an advantage over the native foraminiferal species. Future ecological studies of Nonionella sp. T1 could be aided by dPCR and the novel Nonionella sp. T1-specific T1-1 assay.

Effect of micro-plastic particles on coral reef foraminifera.

Foraminifera are single-celled protists which are important mediators of the marine carbon cycle. In our study, we explored the potential impact of polystyrene (PS) microplastic particles on two symbiont-bearing large benthic foraminifera species-Heterostegina depressa and Amphistegina lobifera-over a period of three weeks, employing three different approaches: investigating (1) stable isotope (SI) incorporation-via 13C- and 15N-labelled substrates-of the foraminifera to assess their metabolic activity, (2) photosynthetic efficiency of the symbiotic diatoms using imaging PAM fluorometry, and (3) microscopic enumeration of accumulation of PS microplastic particles inside the foraminiferal test. The active feeder A. lobifera incorporated significantly more PS particles inside the cytoplasm than the non-feeding H. depressa, the latter accumulating the beads on the test surface. Photosynthetic area of the symbionts tended to decrease in the presence of microplastic particles in both species, suggesting that the foraminiferal host cells started to digest their diatom symbionts. Compared to the control, the presence of microplastic particles lead to reduced SI uptake in A. lobifera, which indicates inhibition of inorganic carbon and nitrogen assimilation. Competition for particulate food uptake was demonstrated between algae and microplastic particles of similar size. Based on our results, both species seem to be sensitive to microplastic pollution, with non-feeding H. depressa being more strongly affected.

An 800-year record of benthic foraminifer images and 2D morphometrics from the Santa Barbara Basin.

The Santa Barbara Basin is an extraordinary archive of environmental and ecological change, where varved sediments preserve microfossils that provide an annual to decadal record of the dynamics of surrounding ecosystems. Of the microfossils preserved in these sediments, benthic foraminifera are the most abundant seafloor-dwelling organisms. While they have been extensively utilized for geochemical and paleoceanographic work, studies of their morphology are lacking. Here we use a high-throughput imaging method (AutoMorph) designed to extract 2D data from photographic images of fossils to produce a large image and 2D shape dataset of recent benthic foraminifera from two core records sampled from the center of the Santa Barbara Basin that span an ~800-year-long interval during the Common Era (1249-2008 CE). Information on more than 36,000 objects is included, of which more than 22,000 are complete or partially-damaged benthic foraminifera. The dataset also includes other biogenic microfossils including ostracods, pteropods, diatoms, radiolarians, fish teeth, and shark dermal denticles. We describe our sample preparation, imaging, and identification techniques, and outline potential data uses.

Benthic foraminifera in Gulf of Mexico show temporal and spatial dynamics of microplastics.

Microplastics have accumulated in the environment since plastic production began, with present-day observations that range from marine trenches to mountains. However, research on microplastics has only recently begun so it is unclear how they have changed over time in many oceanic regions. Our study addressed this gap by quantifying the temporal and spatial dynamics of microplastics in two deep-water regions of the Gulf of Mexico (GOM). We isolated agglutinated foraminifera from sediment cores and assessed microplastics that were incorporated into their tests. Our results indicated that microplastics were incorporated by agglutinated foraminifera after plastic production began. Microplastics were higher at deep-water sites and closer to the Mississippi River. This study confirms the presence of microplastic incorporation into agglutinated foraminifera tests and investigates microplastics in deep-water sediments in the GOM. Additional work is needed to fully identify the distribution of microplastics across the GOM and other oceanic basins.

Bone regeneration of rat tibial defect by zinc-tricalcium phosphate (Zn-TCP) synthesized from porous Foraminifera carbonate macrospheres.

Foraminifera carbonate exoskeleton was hydrothermally converted to biocompatible and biodegradable zinc-tricalcium phosphate (Zn-TCP) as an alternative biomimetic material for bone fracture repair. Zn-TCP samples implanted in a rat tibial defect model for eight weeks were compared with unfilled defect and beta-tricalcium phosphate showing accelerated bone regeneration compared with the control groups, with statistically significant bone mineral density and bone mineral content growth. CT images of the defect showed restoration of cancellous bone in Zn-TCP and only minimal growth in control group. Histological slices reveal bone in-growth within the pores and porous chamber of the material detailing good bone-material integration with the presence of blood vessels. These results exhibit the future potential of biomimetic Zn-TCP as bone grafts for bone fracture repair.

Foraminifera and plastic pollution: Knowledge gaps and research opportunities.

Plastic has become one of the most ubiquitous and environmentally threatening sources of pollution in the Anthropocene. Beyond the conspicuous visual impact and physical damages, plastics both carry and release a cocktail of harmful chemicals, such as monomers, additives and persistent organic pollutants. Here we show through a review of the scientific literature dealing with both plastic pollution and benthic foraminifera (Rhizaria), that despite their critical roles in the structure and function of benthic ecosystems, only 0.4% of studies have investigated the effects of micro- and nano-plastics on this group. Consequently, we urge to consider benthic foraminifera in plastic pollution studies via a tentative roadmap that includes (i) the use of their biological, physiological and behavioral responses that may unveil the effects of microplastics and nanoplastics and (ii) the evaluation of the indicative value of foraminiferal species to serve as proxies for the degree of pollution. This appears particularly timely in the context of the development of management strategies to restore coastal ecosystems.

First record of microplastic in the environmental matrices of a Mediterranean marine cave (Bue Marino, Sardinia, Italy).

This study investigates for the first time the presence of microplastics in sediment, water, and benthic organisms (foraminifera) of a marine cave in the Gulf of Orosei (Sardinia, Italy). Microplastics were found in all water, and sediment samples with similar shapes, sizes, and compositions; identified items were mainly fragments and fibers constituted by PVC and polyethylene. Their provenance was supposed to be predominantly from the sea than from the seasonal freshwater supplies from the karst system. Foraminiferal assemblages were mainly constituted by calcareous hyaline taxa in the outer station, while in the inner ones, the agglutinated Eggerelloides advenus was dominant. FTIR analyses on agglutinated shells identified polyethylene. Microplastic items are collected by the foraminifers and sediment grains building the shell chambers. This is the first study providing evidence that marine caves may be collectors of microplastics and that, in these habitats, microplastics enter the biotic matrix at the protist's level.

Fractal analysis highlights analogies in arenaceous tubes of Sabellaria alveolata (Metazoa, Polychaeta) and agglutinated tests of foraminifera (Protista).

Bioconstructions of Sabellaria alveolata (Polychaeta Sabellariidae) from southern Sicily (Central Mediterranean) were sampled and analysed through a multidisciplinary approach in order to unravel the construction pattern of arenaceous tubes and explore possible analogies existing between the worm tubes and the agglutinated tests of benthic foraminifera (Protista). Scanning Electron Microscopy and Energy Dispersive Spectroscopy analyses were carried out on entire tubes as well as sectioned ones. Results show that arenaceous tubes are built following a rigorous architectural framework, based on selection and methodical arrangement of the agglutinated grains, and show surprising analogies with the test microstructure previously observed in agglutinated foraminifera. The grain distribution detected in both model species bioconstructions was analysed using a fractal numerical model (Hausdorff fractal dimension). Collected data show that in both organisms the grains were distributed according to a fractal model, indicating that the evolutionary process may have led to finding the same optimal constructive strategy across organisms with an independent evolutionary history, notwithstanding different geometrical scales. Furthermore, in sectioned tubes we observed microplastic fragments agglutinated within the arenaceous wall and in the inter-tube area. This unexpected finding shows that marine animals can be affected by microplastic pollution not only in soft tissues, but also engineered hard structures, and suggests the problem is more pervasive than estimated so far.

Nanoplastic incorporation into an organismal skeleton.

Studies on the effects of global marine plastic pollution have largely focused on physiological responses of few organism groups (e.g., corals, fishes). Here, we report the first observation of polymer nanoparticles being incorporated into the calcite skeleton of a large benthic foraminifera (LBF), a significant contributor to global carbonate production. While previous work on LBF has documented selectivity in feeding behaviour and a high degree of specialization regarding skeletal formation, in this study, abundant cases of nanoplastic encrustation into the calcite tests were observed. Nanoplastic incorporation was associated with formation of new chambers, in conjunction with rapid nanoplastic ingestion and subsequent incomplete egestion. Microalgae presence in nanoplastic treatments significantly increased the initial feeding response after 1 day, but regardless of microalgae presence, nanoplastic ingestion was similar after 6 weeks of chronic exposure. While ~ 40% of ingesting LBF expelled all nanoplastics from their cytoplasm, nanoplastics were still attached to the test surface and subsequently encrusted by calcite. These findings highlight the need for further investigation regarding plastic pollution impacts on calcifying organisms, e.g., the function of LBF as potential plastic sinks and alterations in structural integrity of LBF tests that will likely have larger ecosystem-level impacts on sediment production.

Marine biofouling organisms on beached, buoyant and benthic plastic debris in the Catalan Sea.

Plastic debris provides long-lasting substrates for benthic organisms, thus acting as a potential vector for their dispersion. Its interaction with these colonizers is, however, still poorly known. This study examines fouling communities on beached, buoyant and benthic plastic debris in the Catalan Sea (NW Mediterranean), and characterizes the plastic type. We found 14 specimens belonging to two phyla (Annelida and Foraminifera) on microplastics, and more than 400 specimens belonging to 26 species in 10 phyla (Annelida, Arthropoda, Brachiopoda, Bryozoa, Chordata, Cnidaria, Echinodermata, Mollusca, Porifera and Sipuncula) on macroplastics. With 15 species, bryozoans are the most diverse group on plastics. We also report 17 egg cases of the catshark Scyliorhinus sp., and highlight the implications for their dispersal. Our results suggest that plastic polymers may be relevant for distinct fouling communities, likely due to their chemical structure and/or surface properties. Our study provides evidence that biofouling may play a role in the sinking of plastic debris, as the most abundant fouled plastics had lower densities than seawater, and all bryozoan species were characteristic of shallower depths than those sampled. More studies at low taxonomic level are needed in order to detect new species introduction and potential invasive species associated with plastic debris.

Plastics, (bio)polymers and their apparent biogeochemical cycle: An infrared spectroscopy study on foraminifera.

To understand the fate of plastic in oceans and the interaction with marine organisms, we investigated the incorporation of (bio)polymers and microplastics in selected benthic foraminiferal species by applying FTIR (Fourier Transform Infrared) microscopy. This experimental methodology has been applied to cultured benthic foraminifera Rosalina globularis, and to in situ foraminifera collected in a plastic remain found buried into superficial sediment in the Mediterranean seafloor, Rosalina bradyi, Textularia bocki and Cibicidoides lobatulus. In vitro foraminifera were treated with bis-(2-ethylhexyl) phthalate (DEHP) molecule to explore its internalization in the cytoplasm. Benthic foraminifera are marine microbial eukaryotes, sediment-dwelling, commonly short-lived and with reproductive cycles which play a central role in global biogeochemical cycles of inorganic and organic compounds. Despite the recent advances and investigations into the occurrence, distribution, and abundance of plastics, including microplastics, in marine environments, there remain relevant knowledge gaps, particularly on their effects on the benthic protists. No study, to our knowledge, has documented the molecular scale effect of plastics on foraminifera. Our analyses revealed three possible ways through which plastic-related molecules and plastic debris can enter a biogeochemical cycle and may affect the ecosystems: 1) foraminifera in situ can grow on plastic remains, namely C. lobatulus, R. bradyi and T. bocki, showing signals of oxidative stress and protein aggregation in comparison with R. globularis cultured in negative control; 2) DEHP can be incorporated in the cytoplasm of calcareous foraminifera, as observed in R. globularis; 3) microplastic debris, identified as epoxy resin, can be found in the cytoplasm and the agglutinated shell of T. bocki. We hypothesize that plastic waste and their associated additives may produce modifications related to the biomineralization process in foraminifera. This effect would be added to those induced by ocean acidification with negative consequences on the foraminiferal biogenic carbon (C) storage capacity.

Nanoparticle-Biological Interactions in a Marine Benthic Foraminifer.

The adverse effects of engineered nanomaterials (ENM) in marine environments have recently attracted great attention although their effects on marine benthic organisms such as foraminifera are still largely overlooked. Here we document the effects of three negatively charged ENM, different in size and composition, titanium dioxide (TiO2), polystyrene (PS) and silicon dioxide (SiO2), on a microbial eukaryote (the benthic foraminifera Ammonia parkinsoniana) using multiple approaches. This research clearly shows the presence, within the foraminiferal cytoplasm, of metallic (Ti) and organic (PS) ENM that promote physiological stress. Specifically, marked increases in the accumulation of neutral lipids and enhanced reactive oxygen species production occurred in ENM-treated specimens regardless of ENM type. This study indicates that ENM represent ecotoxicological risks for this microbial eukaryote and presents a new model for the neglected marine benthos by which to assess natural exposure scenarios.

Environmental regeneration processes in the Anthropocene: The Bilbao estuary case (northern Spain).

This work tackles a multidisciplinary study on the recent sedimentary record of the Bilbao estuary (northern Spain), which is the backbone of a city that was primarily industrial and now is widely recognized as a successful example of urban transformation. Although hotspots of heavily polluted materials still remain at the mouth of the two main tributaries (Galindo and Gobelas), the data obtained confirm the ongoing formation of a new layer of sediments (here called "postindustrial zone") covering historically polluted and azoic deposits. It is characterized by largely variable levels of metals and magnetic susceptibility and moderate-to-high abundances of benthic foraminifera. Monitoring of the evolution of this layer appears a key factor to assess environmental improvement and decision-making in polluted estuaries.

Characterization of unusual MgCa particles involved in the formation of foraminifera shells using a novel quantitative cryo SEM/EDS protocol.

Quantifying ion concentrations and mapping their intracellular distributions at high resolution can provide much insight into the formation of biomaterials. The key to achieving this goal is cryo-fixation, where the biological materials, tissues and associated solutions are rapidly frozen and preserved in a vitreous state. We developed a correlative cryo-Scanning Electron Microscopy (SEM)/Energy Dispersive Spectroscopy (EDS) protocol that provides quantitative elemental analysis correlated with spatial imaging of cryo-immobilized specimens. We report the accuracy and sensitivity of the cryo-EDS method, as well as insights we derive on biomineralization pathways in a foraminifer. Foraminifera are marine protozoans that produce Mg-containing calcitic shells and are major calcifying organisms in the oceans. We use the cryo-SEM/EDS correlative method to characterize unusual Mg and Ca-rich particles in the cytoplasm of a benthic foraminifer. The Mg/Ca ratio of these particles is consistently lower than that of seawater, the source solution for these ions. We infer that these particles are involved in Ca ion supply to the shell. We document the internal structure of the MgCa particles, which in some cases include a separate Si rich core phase. This approach to mapping ion distribution in cryo-preserved specimens may have broad applications to other mineralized biomaterials. STATEMENT OF SIGNIFICANCE: Ions are an integral part of life, and some ions play fundamental roles in cell metabolism. Determining the concentrations of ions in cells and between cells, as well as their distributions at high resolution can provide valuable insights into ion uptake, storage, functions and the formation of biomaterials. Here we present a new cryo-SEM/EDS protocol that allows the mapping of different ion distributions in solutions and biological samples that have been cryo-preserved. We demonstrate the value of this novel approach by characterizing a novel biogenic mineral phase rich in Mg found in foraminifera, single celled marine organisms. This method has wide applicability in biology, and especially in understanding the formation and function of mineral-containing hard tissues.

Biodiversity and abundance patterns of rock encrusting fauna in a temperate fjord.

Fjords are semi-enclosed systems often with usually strong physical and chemical gradients. These gradients provide the opportunity to test the influence of various physical and chemical factors on biodiversity. However study area of this investigation, Trondheimsfjord, is a large water body where especially salinity gradient along the fjord is not well pronounced. The goal of this study was to establish within a temperate fjord a baseline identifying encrusting fauna on rocks and determine the factors driving changes along the length of the fjord and changing depths. There was no trend in species composition change and increase or decrease in number of species, diversity and number of individuals along the fjord. This was likely due to the relative homogeneity of both substrate (rocks) and environmental parameters. Nevertheless, the influence of fresh water inflow in the vicinity of the river mouth was apparent by the presence of characteristic brackish-water species at these locations. Multidimensional scaling analysis revealed three separate assemblages: intertidal, shallow and deep subtidal (below 50 m). Intertidal assemblages were species poor (one to 11 species) but relatively abundant (six to 2374 indiv./m(2) of rocks). Number of individuals and biomass was highest in the shallow subtidal (2059-13,587 indiv./m(2) of rocks). Overall the highest species number (45) was recorded at 50 m depth which is probably result of low competition pressure yet still relatively high nutrient concentration in comparison to shallower locations. Environmental parameters (i.e., tidal currents, wave action, salinity) change more drastically with depth than along the fjord and these changes are the major driving forces in shaping encrusting assemblages in Trondheimsfjord.

Assessment of the health quality of Ria de Aveiro (Portugal): heavy metals and benthic foraminifera.

This work analyses the distribution of heavy metals in the sediments of Ria de Aveiro (Portugal) assessed by total digestion and sequential chemical extraction of the sediments. The influence of environmental parameters on the living benthic foraminiferal assemblages was studied. The most polluted parts in the Ria de Aveiro are areas where the residence time is high and cohesive sediments are deposited. Organic matter, which is an excellent scavenger for a number of metals, is in general more abundant in the finer deposits of this lagoon, which act as sinks of anthropogenic pollutants. This condition is observed in Aveiro canals and Murtosa channel where sediments with the highest concentrations of Zn, Pb, Cu, and Cr are found. The sediments of Murtosa channel are also enriched in As, Co and Hg. In Aveiro canals the enrichment of heavy metals is mostly related to the past industrial production at their margins (ceramic and metallurgy), whereas in Murtosa channel with effluent discharges of the Chemical Complex of Estarreja. Foraminiferal density and diversity reach higher values near the lagoon mouth under higher marine influence and decline in general under very low-oxygen conditions. Some species seems to be indifferent to the increasing of TOC (e.g. Haynesina germanica and Ammonia tepida) and some have an opportunistic behaviour in areas with very depressed levels of oxygen (e.g. A. tepida and Quinqueloculina seminulum) whereas other species can better tolerate sulphide/reducing conditions (e.g. H. germanica, Bolivina ordinaria, Buliminella elegantissima, Bulimina elongata/gibba and Nonionella stella) a widespread condition in this lagoon. Foraminiferal density and some species are negatively correlated with concentrations of heavy metals. A most sensitive group of species to higher concentrations of heavy metals is identified (such as B. ordinaria, B. pseudoplicata and B. elongata/gibba) and another one of more tolerant species (such as H. germanica A. tepida and Q. seminulum). Foraminifera are more tolerate higher available concentrations (AC) of Zn in any phase than higher AC of Cu adsorbed do clay minerals (F1) and associated with Fe and Mn oxides (F2) and of Pb in F2; the phase F2, probably the most mobile phase, and even phase F1 seems to be more toxic than the increasing of metals in organic matter (F3).

Foraminifera as indicators of marine pollutant contamination on the inner continental shelf of southern Brazil.

Analyses of living foraminiferal and environmental parameters near an outfall at Mar Grosso Beach (Laguna, SC, Brazil) demonstrate its usefulness as indicators of domestic sewage pollution. The low species diversity may be due to sand accumulation in the central part. Higher diversity was noted closer to the mouth of Laguna estuarine system where reduced salinity and higher temperatures indicate freshwater influence, suggesting a relationship between increased diversity and greater availability of terrestrial food. On the basis of foraminiferal diversity and average coliform count the higher values are closer to the mouth of the estuarine system and under the influence of the outfall. Due to the effect of local hydrodynamics, the particulate organic waste derived from the outfall does not settle down locally, and thus, do not accumulate nearby. Our hypothesis is that the fine material derived from the outfall is accumulating on the southwestern and northwestern parts of the beach.