Kleptoplastidic benthic foraminifera from aphotic habitats: Assimilation (or not) of inorganic C, N, and S studied at sub-cellular resolution

The assimilation of inorganic compounds and their role in the foraminiferal metabolism compared to predation or organic matter assimilation is unknown. Here we investigate these processes with a study of the species Nonionellina labradorica (Dawson, 1860), a common kleptoplastic benthic foraminifer from Arctic and North Atlantic sublittoral regions, including aphotic zones of the Gullmar Fjord (Sweden). The objectives of this work were to (1) identify the origin of N. labradorica kleptoplasts, (2) assess their photosynthetic functionality for both oxygen production and conversion of inorganic carbon into photosynthates, and (3) investigate inorganic nitrogen and sulfate assimilation by N. labradorica. We used DNA barcoding to identify the origin of N. labradorica kleptoplasts and transmission electron microscope (TEM) imaging, correlated with nanometer-scale secondary ion mass spectrometry (NanoSIMS) isotopic imaging, to study 13C-bicarbonate, 15N-ammonium, and 34S-sulfate uptake. In addition, respiration rate measurements were determined to assess N. labradorica response to light. The partial 18S and 16S rDNA sequencing established that the kleptoplasts belonged to two species of Thalassiosira, a cosmopolitan planktonic diatom genus. TEM-NanoSIMS imaging revealed degraded cytoplasm in specimens exposed to light for 20 h and a complete absence of 13C assimilation. Oxygen measurements showed higher respiration rates under light than dark conditions, and no O2 production was detected. These combined results indicate that the photosynthetic pathways in N. labradorica are not functional. Under dark conditions, N. labradorica assimilated both 15N-ammonium and 34S-sulfate into its cytoplasm. Light exposure negatively impacted foraminiferal metabolism, ultimately causing specimen death. Our results suggest that foraminifera might have several ammonium or sulfate assimilation pathways, involving either the kleptoplasts or bona fide foraminiferal pathway(s) not yet identified.

Kleptoplastidic benthic foraminifera from aphotic habitats: insights into assimilation of inorganic C, N and S studied with sub-cellular resolution

Emmanuelle Geslin, Charlotte Madeleine Nicole Lekieffre, Anders Meibom

The assimilation of inorganic compounds in foraminiferal metabolism compared to predation or organic matter assimilation is unknown. Here, we investigate possible inorganic-compound assimilation in Nonionellina labradorica, a common kleptoplastidic benthic foraminifer from Arctic and North Atlantic sublittoral regions. The objectives were to identify the source of the foraminiferal kleptoplasts, assess their photosynthetic functionality in light and darkness and investigate inorganic nitrogen and sulfate assimilation. We used DNA barcoding of a similar to 830 bp fragment from the SSU rDNA to identify the kleptoplasts and correlated transmission electron microscopy and nanometre-scale secondary ion mass spectrometry (TEM-NanoSIMS) isotopic imaging to study C-13-bicarbonate, N-15-ammonium and S-34-sulfate uptake. In addition, respiration rate measurements were determined to assess the response of N. labradorica to light. The DNA sequences established that over 80% of the kleptoplasts belonged to Thalassiosira (with 96%-99% identity), a cosmopolitan planktonic diatom. TEM-NanoSIMS imaging revealed degraded cytoplasm and an absence of C-13 assimilation in foraminifera exposed to light. Oxygen measurements showed higher respiration rates under light than dark conditions, and no O-2 production was detected. These results indicate that the photosynthetic pathways in N. labradorica are not functional. Furthermore, N. labradorica assimilated both N-15-ammonium and S-34-sulfate into its cytoplasm, which suggests that foraminifera might have several ammonium or sulfate assimilation pathways, involving either the kleptoplasts or bona fide foraminiferal pathway(s) not yet identified.

WILEY2019

Ammonium is the preferred source of nitrogen for planktonic foraminifer and their dinoflagellate symbionts

Emmanuelle Geslin, Charlotte Madeleine Nicole Lekieffre, Anders Meibom

The symbiotic planktonic foraminiferaOrbulina universainhabits open ocean oligotrophic ecosystems where dissolved nutrients are scarce and often limit biological productivity. It has previously been proposed thatO. universameets its nitrogen (N) requirements by preying on zooplankton, and that its symbiotic dinoflagellates recycle metabolic 'waste ammonium' for their N pool. However, these conclusions were derived from bulk(15)N-enrichment experiments and model calculations, and our understanding of N assimilation and exchange between the foraminifer host cell and its symbiotic dinoflagellates remains poorly constrained. Here, we present data from pulse-chase experiments with(13)C-enriched inorganic carbon,N-15-nitrate, and(15)N-ammonium, as well as a(13)C- and(15)N- enriched heterotrophic food source, followed by TEM (transmission electron microscopy) coupled to NanoSIMS (nanoscale secondary ion mass spectrometry) isotopic imaging to visualize and quantify C and N assimilation and translocation in the symbiotic system. High levels of(15)N-labelling were observed in the dinoflagellates and in foraminiferal organelles and cytoplasm after incubation with(15)N-ammonium, indicating efficient ammonium assimilation. Only weak(15)N-assimilation was observed after incubation with(15)N-nitrate. Feeding foraminifers with(13)C- and(15)N-labelled food resulted in dinoflagellates that were labelled with(15)N, thereby confirming the transfer of(15)N-compounds from the digestive vacuoles of the foraminifer to the symbiotic dinoflagellates, likely through recycling of ammonium. These observations are important for N isotope-based palaeoceanographic reconstructions, as they show that delta N-15 values recorded in the organic matrix in symbiotic species likely reflect ammonium recycling rather than alternative N sources, such as nitrates.

ROYAL SOC2020

Feeding behaviour of a benthic species (Ammonia tepida) under oxic and anoxic conditions: TEM-NanoSIMS correlation

Emmanuelle Geslin, Charlotte Madeleine Nicole Lekieffre, Anders Meibom

More and more marine areas are subjected to depleted-O2 concentration, mainly due to eutrophication induced by human activities. These phenomena affect benthic ecosystems, in particular continental shelves and coastal areas where the renewal of the bottom waters is low and/or the organic matter flux is high. Some species of benthic foraminifera are resistant to this hypoxic/anoxic events, surviving weeks to months without oxygen. One of the species known to survive to such extreme condition is the benthic one Ammonia tepida. However the metabolic process by which it succeed in it remains unknown. One hypothesis is that this species would be able to lower its metabolism under oxygen depleted conditions, until the return of better conditions. The aim of the present study is to understand the feeding behaviors of A. tepida under anoxia. For this purpose a laboratory experiment involving incubation with living A. tepida was carried out under controlled oxygen concentrations. The individuals were fed with a pulse of 13C-labeled diatoms (Navicula sp.) and then incubate in oxic or anoxic conditions during 28 days. Along the incubation time, observation of the cell ultrastructure with Transmission Electronic Microscope (TEM) and detection in the cell of the isotopic 13C signal using nanoSIMS, were made. The nanoSIMS (nanoscale Secondary-Ion Mass Spectrometry) is an analytical technique that allows to visualize the incorporation and transfer of isotopically labeled compounds in organisms; thus in our case to follow the ingestion of labeled-diatoms and further transfer of carbon in the cytoplasm of the foraminifera. According to the results acquired in this study, in both conditions the foraminifera directly integrate the diatoms in their cytoplasm during the first hours of the incubation. Then, in oxic conditions, these diatoms are quickly fully degraded, within 3 days, and the 13C-labeled compounds are transferred in the organelles of the cytoplasm. Whereas in anoxia, only a fraction of the diatoms are degraded and the transfer to the foraminifera cytoplasm is weak. These results are confirmed by the δ13C of the foraminifera, measured in bulk by mass spectrometry (GC-MS). In anoxia the δ13C is slightly increasing the first 24h, meaning that the 13C content of the cell increased, and then it remains stable until the end of the experiment. This leads us to assume that the foraminifera stop, or at least strongly lower their metabolic rate under anoxic condition.

2015

Ammonium is the preferred source of nitrogen for planktonic foraminifer and their dinoflagellate symbionts

Emmanuelle Geslin, Charlotte Madeleine Nicole Lekieffre, Anders Meibom

ROYAL SOC2020

Feeding behaviour of a benthic species (Ammonia tepida) under oxic and anoxic conditions: TEM-NanoSIMS correlation

Emmanuelle Geslin, Charlotte Madeleine Nicole Lekieffre, Anders Meibom

2015