Anaerobic metabolism of Foraminifera thriving below the seafloor

Foraminifera are single-celled eukaryotes (protists) of large ecological importance, as well as environmental and paleoenvironmental indicators and biostratigraphic tools. In addition, they are capable of surviving in anoxic marine environments where they represent a major component of the benthic community. However, the cellular adaptations of Foraminifera to the anoxic environment remain poorly constrained. We sampled an oxic-anoxic transition zone in marine sediments from the Namibian shelf, where the genera Bolivina and Stainforthia dominated the Foraminifera community, and use metatranscriptomics to characterize Foraminifera metabolism across the different geochemical conditions. Relative Foraminifera gene expression in anoxic sediment increased an order of magnitude, which was confirmed in a 10-day incubation experiment where the development of anoxia coincided with a 20-40-fold increase in the relative abundance of Foraminifera protein encoding transcripts, attributed primarily to those involved in protein synthesis, intracellular protein trafficking, and modification of the cytoskeleton. This indicated that many Foraminifera were not only surviving but thriving, under the anoxic conditions. The anaerobic energy metabolism of these active Foraminifera was characterized by fermentation of sugars and amino acids, fumarate reduction, and potentially dissimilatory nitrate reduction. Moreover, the gene expression data indicate that under anoxia Foraminifera use the phosphogen creatine phosphate as an ATP store, allowing reserves of high-energy phosphate pool to be maintained for sudden demands of increased energy during anaerobic metabolism. This was co-expressed alongside genes involved in phagocytosis and clathrin-mediated endocytosis (CME). Foraminifera may use CME to utilize dissolved organic matter as a carbon and energy source, in addition to ingestion of prey cells via phagocytosis. These anaerobic metabolic mechanisms help to explain the ecological success of Foraminifera documented in the fossil record since the Cambrian period more than 500 million years ago

Worldwide Genotyping in the Planktonic Foraminifer Globoconella inflata: Implications for Life History and Paleoceanography

The planktonic foraminiferal morpho-species Globoconella inflata is widely used as a stratigraphic and paleoceanographic index. While G. inflata was until now regarded as a single species, we show that it rather constitutes a complex of two pseudo-cryptic species. Our study is based on SSU and ITS rDNA sequence analyses and genotyping of 497 individuals collected at 49 oceanic stations covering the worldwide range of the morpho-species. Phylogenetic analyses unveil the presence of two divergent genotypes. Type I inhabits transitional and subtropical waters of both hemispheres, while Type II is restricted to the Antarctic subpolar waters. The two genetic species exhibit a strictly allopatric distribution on each side of the Antarctic Subpolar Front. On the other hand, sediment data show that G. inflata was restricted to transitional and subtropical environments since the early Pliocene, and expanded its geographic range to southern subpolar waters ∼700 kyrs ago, during marine isotopic stage 17. This datum may correspond to a peripatric speciation event that led to the partition of an ancestral genotype into two distinct evolutionary units. Biometric measurements performed on individual G. inflata from plankton tows north and south of the Antarctic Subpolar Front indicate that Types I and II display slight but significant differences in shell morphology. These morphological differences may allow recognition of the G. inflata pseudo-cryptic species back into the fossil record, which in turn may contribute to monitor past movements of the Antarctic Subpolar Front during the middle and late Pleistocene

Fossil and genetic evidence for the polyphyletic nature of the planktonic foraminifera ‘Globigerinoides’, and description of the new genus Trilobatus

Planktonic foraminifera are one of the most abundant and diverse protists in the oceans. Their utility as paleo proxies requires rigorous taxonomy and comparison with living and genetically related counterparts. We merge genetic and fossil evidence of “Globigerinoides”, characterized by supplementary apertures on spiral side, in a new approach to trace their “total evidence phylogeny” since their first appearance in the latest Paleogene. Combined fossil and molecular genetic data indicate that this genus, as traditionally understood, is polyphyletic. Both datasets indicate the existence of two distinct lineages that evolved independently. One group includes “Globigerinoides” trilobus and its descendants, the extant “Globigerinoides” sacculifer, Orbulina universa and Sphaeroidinella dehiscens. The second group includes the Globigerinoides ruber clade with the extant G. conglobatus and G. elongatus and ancestors. In molecular phylogenies, the trilobus group is not the sister taxon of the ruber group. The ruber group clusters consistently together with the modern Globoturborotalita rubescens as a sister taxon. The re-analysis of the fossil record indicates that the first “Globigerinoides” in the late Oligocene are ancestral to the trilobus group, whereas the ruber group first appeared at the base of the Miocene with representatives distinct from the trilobus group. Therefore, polyphyly of the genus "Globigerinoides" as currently defined can only be avoided either by broadening the genus concept to include G. rubescens and a large number of fossil species without supplementary apertures, or if the trilobus group is assigned to a separate genus. Since the former is not feasible due to the lack of a clear diagnosis for such a broad genus, we erect a new genus Trilobatus for the trilobus group (type species Globigerina triloba Reuss) and amend Globoturborotalita and Globigerinoides to clarify morphology and wall textures of these genera. In the new concept, Trilobatus n. gen. is paraphyletic and gave rise to the Praeorbulina / Orbulina and Sphaeroidinellopsis / Sphaeroidinella lineages

Eukaryotic biodiversity and spatial patterns in the Clarion-Clipperton Zone and other Abyssal regions: Insights from sediment DNA and RNA metabarcoding

The abyssal seafloor is a mosaic of highly diverse habitats that represent the least known marine ecosystems on Earth. Some regions enriched in natural resources, such as polymetallic nodules in the Clarion-Clipperton Zone (CCZ), attract much interest because of their huge commercial potential. Since nodule mining will be destructive, baseline data are necessary to measure its impact on benthic communities. Hence, we conducted an environmental DNA and RNA metabarcoding survey of CCZ biodiversity targeting microbial and meiofaunal eukaryotes that are the least known component of the deep-sea benthos. We analyzed two 18S rRNA gene regions targeting eukaryotes with a focus on Foraminifera (37F) and metazoans (V1V2), sequenced from 310 surface-sediment samples from the CCZ and other abyssal regions. Our results confirm huge unknown deep-sea biodiversity. Over 60% of benthic foraminiferal and almost a third of eukaryotic operational taxonomic units (OTUs) could not be assigned to a known taxon. Benthic Foraminifera are more common in CCZ samples than metazoans and dominated by clades that are only known from environmental surveys. The most striking results are the uniqueness of CCZ areas, both datasets being characterized by a high number of OTUs exclusive to the CCZ, as well as greater beta diversity compared to other abyssal regions. The alpha diversity in the CCZ is high and correlated with water depth and terrain complexity. Topography was important at a local scale, with communities at CCZ stations located in depressions more diverse and heterogeneous than those located on slopes. This could result from eDNA accumulation, justifying the interim use of eRNA for more accurate biomonitoring surveys. Our descriptions not only support previous findings and consolidate our general understanding of deep-sea ecosystems, but also provide a data resource inviting further taxon-specific and large-scale modeling studies. We foresee that metabarcoding will be useful for deep-sea biomonitoring efforts to consider the diversity of small taxa, but it must be validated based on ground truthing data or experimental studies

The Protist Ribosomal Reference database (PR2): a catalog of unicellular eukaryote Small Sub-Unit rRNA sequences with curated taxonomy

International audienceThe interrogation of genetic markers in environmental meta-barcoding studies is currently seriously hindered by the lack of taxonomically curated reference data sets for the targeted genes. The Protist Ribosomal Reference database (PR2, http://ssu-rrna.org/) provides a unique access to eukaryotic small sub-unit (SSU) ribosomal RNA and DNA sequences, with curated taxonomy. The database mainly consists of nuclear-encoded protistan sequences. However, metazoans, land plants, macrosporic fungi and eukaryotic organelles (mitochondrion, plastid and others) are also included because they are useful for the analysis of high-troughput sequencing data sets. Introns and putative chimeric sequences have been also carefully checked. Taxonomic assignation of sequences consists of eight unique taxonomic fields. In total, 136 866 sequences are nuclear encoded, 45 708 (36 501 mitochondrial and 9657 chloroplastic) are from organelles, the remaining being putative chimeric sequences. The website allows the users to download sequences from the entire and partial databases (including representative sequences after clustering at a given level of similarity). Different web tools also allow searches by sequence similarity. The presence of both rRNA and rDNA sequences, taking into account introns (crucial for eukaryotic sequences), a normalized eight terms ranked-taxonomy and updates of new GenBank releases were made possible by a long-term collaboration between experts in taxonomy and computer scientists

Pour une meilleure caractérisation du registre fossile pélagique : diversité morphologique, biogéographie et écologie des espèces cryptiques de foraminifères planctoniques

The usefulness of calcareous shells of planktonic foraminifera as a paleoceanographic proxy relies on the key hypothesis that each morphospecies corresponds to a biological species with a specific habitat. This empirical relationship has been challenged since molecular analyses have revealed a significant level of cryptic genetic diversity among modern morphospecies of planktonic foraminifera. Previous workers have suggested that the cryptic species or genotypes (1) display narrower biogeographic and ecological ranges than their related morphospecies, and (2) exhibit shell morphological differences. In this work, we have characterized the genetic, morphological and ecological diversity among four planktonic foraminiferal morphospecies of significance in paleoceanography, i.e. Orbulina universa, Truncorotalia truncatulinoides, Globoconella inflata and Globigerina bulloides. Our study relies on the development of a new single-cell DNA extraction protocol that retains the shell, allowing direct morpho-genetic comparisons. Shape or porosity variations within each genotype have been quantified. It appears that the high degree of morphological plasticity widely documented in planktonic foraminifera and classically seen as ecophenotypy, is at least partly the spurious consequence of lumping several genotypes that display morphological and environmental preferences. Based on these observations, we developed several population-scale models, which allow recognition of the cryptic species based on their shell morphology. Finally, in order to quantify the impact of integrating cryptic diversity in assemblage-based aleoceanographic reconstructions, we have re-calibrated transfer functions based on the ecological ranges of the genotypes of O. universa, T. truncatulinoides, G. inflata and G. bulloides in the southern oceans. Such recalibrations led to a great, previously never reached improvement in the accuracy of the assemblage-based paleoceanographic reconstructions.L’utilisation des coquilles carbonatées de foraminifères planctoniques comme marqueurs paléocéanographiques repose sur l’hypothèse fondamentale que chaque espèce morphologique correspond à une espèce biologique caractéristique d’un habitat spécifique. Cette relation empirique a été récemment remise en cause par des analyses moléculaires qui ont révélé la présence systématique de plusieurs espèces génétiques (espèces cryptiques) au sein des morpho-espèces actuelles. Il a été suggéré que ces espèces cryptiques ou génotypes, présentaient (1) des préférences biogéographiques et écologiques restreintes par rapport à leur morpho-espèce respective et (2) des différences morphologiques. Dans ce travail, nous avons caractérisé la diversité génétique, morphologique et écologique de 4 morpho-espèces clefs de la paléocéanographie, i.e. Orbulina universa, Truncorotalia truncatulinoides, Globoconella inflata et Globigerina bulloides. Cette étude repose sur le développement d’un protocole d’extraction ADN non destructif pour la coquille calcaire, permettant l’analyse conjointe de la variabilité génétique et morphologique d’un même individu. Les variations de forme ou de porosité de chacun des génotypes des morphoespèces ont été quantifiées. Il apparaît que le fort degré de plasticité morphologique largement documenté chez les foraminifères planctonique et jusqu’alors interprété comme écophénotypique, est au moins en partie la conséquence du regroupement de plusieurs génotypes présentant des morphologies et préférences écologique particulières. Sur la base de ces observations, des modèles de reconnaissance morphologique permettant d’identifier les génotypes à partir de la morphologie de la coquille, utilisables à l’échelle populationnelle, ont été développés. Afin de quantifier l’impact de l’intégration de la diversité cryptique dans les reconstitutions paléocéanographiques basées sur les assemblages de foraminifères planctoniques, les fonctions de transfert couramment utilisées ont été re-calibrées en intégrant les distributions restreintes des génotypes d’O. universa, T. truncatulinoides, G. inflata et G. bulloides. Ces re-calibrations conduisent à un degré de précision jusqu’alors jamais atteint dans les reconstructions paléocéanographiques basées sur les assemblages de foraminifères planctoniques

For a better characterization of the fossil pelagic record : molecular, biogeographical and ecological diversity of planctonic foraminifers cryptic species

L’utilisation des coquilles carbonatées de foraminifères planctoniques comme marqueurs paléocéanographiques repose sur l’hypothèse fondamentale que chaque espèce morphologique correspond à une espèce biologique caractéristique d’un habitat spécifique. Cette relation empirique a été récemment remise en cause par des analyses moléculaires qui ont révélé la présence systématique de plusieurs espèces génétiques (espèces cryptiques) au sein des morpho-espèces actuelles. Il a été suggéré que ces espèces cryptiques ou génotypes, présentaient (1) des préférences biogéographiques et écologiques restreintes par rapport à leur morpho-espèce respective et (2) des différences morphologiques. Dans ce travail, nous avons caractérisé la diversité génétique, morphologique et écologique de 4 morpho-espèces clefs de la paléocéanographie, i.e. Orbulina universa, Truncorotalia truncatulinoides, Globoconella inflata et Globigerina bulloides. Cette étude repose sur le développement d’un protocole d’extraction ADN non destructif pour la coquille calcaire, permettant l’analyse conjointe de la variabilité génétique et morphologique d’un même individu. Les variations de forme ou de porosité de chacun des génotypes des morphoespèces ont été quantifiées. Il apparaît que le fort degré de plasticité morphologique largement documenté chez les foraminifères planctonique et jusqu’alors interprété comme écophénotypique, est au moins en partie la conséquence du regroupement de plusieurs génotypes présentant des morphologies et préférences écologique particulières. Sur la base de ces observations, des modèles de reconnaissance morphologique permettant d’identifier les génotypes à partir de la morphologie de la coquille, utilisables à l’échelle populationnelle, ont été développés. Afin de quantifier l’impact de l’intégration de la diversité cryptique dans les reconstitutions paléocéanographiques basées sur les assemblages de foraminifères planctoniques, les fonctions de transfert couramment utilisées ont été re-calibrées en intégrant les distributions restreintes des génotypes d’O. universa, T. truncatulinoides, G. inflata et G. bulloides. Ces re-calibrations conduisent à un degré de précision jusqu’alors jamais atteint dans les reconstructions paléocéanographiques basées sur les assemblages de foraminifères planctoniques.The usefulness of calcareous shells of planktonic foraminifera as a paleoceanographic proxy relies on the key hypothesis that each morphospecies corresponds to a biological species with a specific habitat. This empirical relationship has been challenged since molecular analyses have revealed a significant level of cryptic genetic diversity among modern morphospecies of planktonic foraminifera. Previous workers have suggested that the cryptic species or genotypes (1) display narrower biogeographic and ecological ranges than their related morphospecies, and (2) exhibit shell morphological differences. In this work, we have characterized the genetic, morphological and ecological diversity among four planktonic foraminiferal morphospecies of significance in paleoceanography, i.e. Orbulina universa, Truncorotalia truncatulinoides, Globoconella inflata and Globigerina bulloides. Our study relies on the development of a new single-cell DNA extraction protocol that retains the shell, allowing direct morpho-genetic comparisons. Shape or porosity variations within each genotype have been quantified. It appears that the high degree of morphological plasticity widely documented in planktonic foraminifera and classically seen as ecophenotypy, is at least partly the spurious consequence of lumping several genotypes that display morphological and environmental preferences. Based on these observations, we developed several population-scale models, which allow recognition of the cryptic species based on their shell morphology. Finally, in order to quantify the impact of integrating cryptic diversity in assemblage-based aleoceanographic reconstructions, we have re-calibrated transfer functions based on the ecological ranges of the genotypes of O. universa, T. truncatulinoides, G. inflata and G. bulloides in the southern oceans. Such recalibrations led to a great, previously never reached improvement in the accuracy of the assemblage-based paleoceanographic reconstructions

For a better characterization of the fossil pelagic record : molecular, biogeographical and ecological diversity of planctonic foraminifers cryptic species

L’utilisation des coquilles carbonatées de foraminifères planctoniques comme marqueurs paléocéanographiques repose sur l’hypothèse fondamentale que chaque espèce morphologique correspond à une espèce biologique caractéristique d’un habitat spécifique. Cette relation empirique a été récemment remise en cause par des analyses moléculaires qui ont révélé la présence systématique de plusieurs espèces génétiques (espèces cryptiques) au sein des morpho-espèces actuelles. Il a été suggéré que ces espèces cryptiques ou génotypes, présentaient (1) des préférences biogéographiques et écologiques restreintes par rapport à leur morpho-espèce respective et (2) des différences morphologiques. Dans ce travail, nous avons caractérisé la diversité génétique, morphologique et écologique de 4 morpho-espèces clefs de la paléocéanographie, i.e. Orbulina universa, Truncorotalia truncatulinoides, Globoconella inflata et Globigerina bulloides. Cette étude repose sur le développement d’un protocole d’extraction ADN non destructif pour la coquille calcaire, permettant l’analyse conjointe de la variabilité génétique et morphologique d’un même individu. Les variations de forme ou de porosité de chacun des génotypes des morphoespèces ont été quantifiées. Il apparaît que le fort degré de plasticité morphologique largement documenté chez les foraminifères planctonique et jusqu’alors interprété comme écophénotypique, est au moins en partie la conséquence du regroupement de plusieurs génotypes présentant des morphologies et préférences écologique particulières. Sur la base de ces observations, des modèles de reconnaissance morphologique permettant d’identifier les génotypes à partir de la morphologie de la coquille, utilisables à l’échelle populationnelle, ont été développés. Afin de quantifier l’impact de l’intégration de la diversité cryptique dans les reconstitutions paléocéanographiques basées sur les assemblages de foraminifères planctoniques, les fonctions de transfert couramment utilisées ont été re-calibrées en intégrant les distributions restreintes des génotypes d’O. universa, T. truncatulinoides, G. inflata et G. bulloides. Ces re-calibrations conduisent à un degré de précision jusqu’alors jamais atteint dans les reconstructions paléocéanographiques basées sur les assemblages de foraminifères planctoniques.The usefulness of calcareous shells of planktonic foraminifera as a paleoceanographic proxy relies on the key hypothesis that each morphospecies corresponds to a biological species with a specific habitat. This empirical relationship has been challenged since molecular analyses have revealed a significant level of cryptic genetic diversity among modern morphospecies of planktonic foraminifera. Previous workers have suggested that the cryptic species or genotypes (1) display narrower biogeographic and ecological ranges than their related morphospecies, and (2) exhibit shell morphological differences. In this work, we have characterized the genetic, morphological and ecological diversity among four planktonic foraminiferal morphospecies of significance in paleoceanography, i.e. Orbulina universa, Truncorotalia truncatulinoides, Globoconella inflata and Globigerina bulloides. Our study relies on the development of a new single-cell DNA extraction protocol that retains the shell, allowing direct morpho-genetic comparisons. Shape or porosity variations within each genotype have been quantified. It appears that the high degree of morphological plasticity widely documented in planktonic foraminifera and classically seen as ecophenotypy, is at least partly the spurious consequence of lumping several genotypes that display morphological and environmental preferences. Based on these observations, we developed several population-scale models, which allow recognition of the cryptic species based on their shell morphology. Finally, in order to quantify the impact of integrating cryptic diversity in assemblage-based aleoceanographic reconstructions, we have re-calibrated transfer functions based on the ecological ranges of the genotypes of O. universa, T. truncatulinoides, G. inflata and G. bulloides in the southern oceans. Such recalibrations led to a great, previously never reached improvement in the accuracy of the assemblage-based paleoceanographic reconstructions

Macroevolutionary patterns in intragenomic rDNA variability among planktonic foraminifera

Ribosomal intragenomic variability in prokaryotes and eukaryotes is a genomic feature commonly studied for its inflationary impact on molecular diversity assessments. However, the evolutionary mechanisms and distribution of this phenomenon within a microbial group are rarely explored. Here, we investigate the intragenomic variability in 33 species of planktonic foraminifera, calcifying marine protists, by inspecting 2,403 partial SSU sequences obtained from single-cell clone libraries. Our analyses show that polymorphisms are common among planktonic foraminifera species, but the number of polymorphic sites significantly differs among clades. With our molecular simulations, we could assess that most of these mutations are located in paired regions that do not affect the secondary structure of the SSU fragment. Finally, by mapping the number of polymorphic sites on the phylogeny of the clades, we were able to discuss the evolution and potential sources of intragenomic variability in planktonic foraminifera, linking this trait to the distinctive nuclear and genomic dynamics of this microbial group