Maria Holzmann

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

The 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.

CBOL Protist Working Group: Barcoding Eukaryotic Richness beyond the Animal, Plant, and Fungal Kingdoms

A group of protist experts proposes a two-step DNA barcoding approach, comprising a universal eukaryotic pre-barcode followed by group-specific barcodes, to unveil the hidden biodiversity of microbial eukaryotes.

Morphological distinction of molecular types in Ammonia – towards a taxonomic revision of the world’s most commonly misidentified foraminifera

Abstract In this study, morphometric analysis has been performed on 178 Ammonia specimens belonging to 12 different molecular types, plus non-sequenced type specimens of Ammonia beccarii and A. tepida. Molecular type distinction is based on phylogenetic analysis of 267 partial LSU rDNA sequences, obtained from 202 living Ammonia specimens, sampled in 30 localities from 17 countries bordering the Pacific Ocean, Atlantic Ocean, Mediterranean Sea, Caribbean Sea and North Sea. Restriction fragment length polymorphism (analysis was carried out for another seven specimens. Morphometric analysis was based on measurements or assessments of 37 external test characters in spiral, umbilical, profile and close-up Scanning Electron Microscopic views. Cluster analysis, canonical variates analysis, and detrended correspondence analysis, performed on the morphological data set, suggest that each molecular type can be distinguished morphologically and can be regarded as a separate species. Primary types of A. tepida and topotypes of A. beccarii are shown to be morphologically separate from any of the molecular types so far recognised. We are aware of at least 9 more distinctive morphotypes that have not yet been sequenced, and thus we infer that the total number of genetically distinct and morphologically separable living species of Ammonia worldwide is likely to exceed 25–30. At this stage not all molecular types can be unequivocally assigned to formally described species. Several genetically-based species can be distinguished by the presence of one distinct character, but most are discriminated on the basis of a combination of many different characters. Morphological characters (e.g. test shape, chamber shape, porosity, prolocular diameter, folium shape, radial furrow length, umbilical diameter) are shown to be slightly more valuable in separating the molecular types than surficial ornament (beads, pustules, bosses, secondary calcite). One highly distinctive group (2–3 species – beccarii, batava, ?inflata) is readily discriminated on the basis of its large test size, strongly beaded and grooved ornament, and the presence of fissures along the sutures on the spiral side. The results of this study imply that the widespread practice of recognising only one, two or three species of Recent Ammonia worldwide should be abandoned. The most commonly used name, Ammonia beccarii, should be restricted to a large, compressed, highly ornamented species, so far not recognised beyond its type locality in the Adriatic Sea. Other commonly used names, such as A. parkinsoniana and A. tepida, apply to species with far more restricted distributions than the literature would suggest.

The evolution of early Foraminifera

Fossil Foraminifera appear in the Early Cambrian, at about the same time as the first skeletonized metazoans. However, due to the inadequate preservation of early unilocular (single-chambered) foraminiferal tests and difficulties in their identification, the evolution of early foraminifers is poorly understood. By using molecular data from a wide range of extant naked and testate unilocular species, we demonstrate that a large radiation of nonfossilized unilocular Foraminifera preceded the diversification of multilocular lineages during the Carboniferous. Within this radiation, similar test morphologies and wall types developed several times independently. Our findings indicate that the early Foraminifera were an important component of Neoproterozoic protistan community, whose ecological complexity was probably much higher than has been generally accepted.

PHYLOGENY OF ALLOGROMIID FORAMINIFERA INFERRED FROM SSU rRNA GENE SEQUENCES

Allogromiids are classically defined as a group of monothalamous, soft-walled foraminiferans. Recent morphological, cytological, and molecular studies, however, challenge this view, showing that the soft-walled allogromiids are closely related to naked athalamids and unilocular agglutinated foraminiferans. To establish the phylogenetic relationships among these three groups we obtained partial small-subunit ribosomal DNA sequences of 50 species and undetermined morphotypes, and compared them to other foraminiferal taxa. Phylogenetic analyses of our data show that allogromiids, athalamids and astrorhizids comprise an assemblage of 13 lineages branching together at the base of the foraminiferal tree. Among these lineages, two are represented by a single species and four comprise similar genera, while the remaining seven are heterogeneous groups composed of several species having different types of wall structure and different test morphologies. All lineages are relatively well supported, yet the relationships among them are not resolved. In view of our data, we propose to revise the definition of allogromiids to include all naked and testate unilocular granuloreticuloseans that diverged early in the evolution of Foraminifera.

Molecular Identification of Algal Endosymbionts in Large Miliolid Foraminifera: 2. Dinoflagellates

Abstract Large miliolid foraminifers of the subfamily Soritinae bear symbiotic dinoflagellates morphologically similar to the species of the “Symbiodinium” complex, commonly found in corals and other marine invertebrates. Soritid foraminifers are abundant in coral reefs and it has been proposed that they share their symbionts with other dinoflagellate-bearing reef dwellers. In order to test this hypothesis, we have analysed partial large subunit ribosomal DNA sequences from dinoflagellates symbionts obtained from 28 foraminiferal specimens, and compared them to the corresponding sequences of Symbiodinium-like endosymbionts from various groups of invertebrates. Phylogenetic analysis of our data shows that all soritid symbionts belong to the “Symbiodinium” species complex, within which they form seven different molecular types (Fr1–Fr7). Only one of these types (Fr1) branches within a group of invertebrate symbionts, previously described as type C. The remaining six types form sister groups to coral symbionts previously designed as types B, C, and D. Our data indicate a high genetic diversity and specificity of Symbiodinium-like symbionts in soritids. Except for type C, we have found no evidence for the transmission of symbionts between foraminifers and other symbiont-bearing invertebrates from the same localities. However, exchanges must have occurred frequently between the different species of Soritinae, as suggested by the lack of host specificity and some biogeographical patterns observed in symbiont distribution. Our data suggest that members of the subfamily Soritinae acquired their symbionts at least three times during their history, each acquisition being followed by a rapid diversification and independent radiation of symbionts within the foraminiferal hosts.

Freshwater Foraminiferans Revealed by Analysis of Environmental DNA Samples

Abstract Sediment-dwelling protists are among the most abundant meiobenthic organisms, ubiquitous in all types of aquatic ecosystems. Yet, because their isolation and identification are difficult, their diversity remains largely unknown. In the present work, we applied molecular methods to examine the diversity of freshwater Foraminifera, a group of granuloreticulosan protists largely neglected until now. By using specific PCR primers, we detected the presence of Foraminifera in all sediment samples examined. Phylogenetic analysis of amplified SSU rDNA sequences revealed two distinct groups of freshwater foraminiferans. All obtained sequences branched within monothalamous (single-chambered), marine Foraminifera, suggesting a repeated colonization of freshwater environments. The results of our study challenge the traditional view of Foraminifera as essentially marine organisms, and provide a conceptual framework for charting the molecular diversity of freshwater granuloreticulosan protists.

Molecular phylogeny of Foraminifera a review

Foraminifera are traditionally defined as marine granuloreticuloseans characterized by the presence of a membraneous, agglutinated or calcareous test. This definition has been recently challenged by molecular phylogenetic studies which showed that Foraminifera include both testate and naked species and that they occur in marine, freshwater, and terrestrial environments. Molecular data also revealed high taxonomic diversity of monothalamous (single-chambered) foraminiferans that developed different types of organic and agglutinated tests. First analyses of ribosomal DNA sequences suggested an early divergence of Foraminifera in the evolutionary history of Eukaryotes, but this result was not confirmed by later protein sequence data. Furthermore, analysis of variable regions in ribosomal DNA revealed the presence of several cryptic species, whose geographic distribution seems to be related to oceanic water mass circulation and productivity.

Molecular phylogeny of large miliolid foraminifera (Soritacea Ehrenberg 1839)

Abstract The foraminiferal superfamily Soritacea belongs to the suborder Miliolina and is divided in two families, Peneroplidae and Soritidae, the latter one comprising two subfamilies, Archaiasinae and Soritinae. Phylogenetic relationships of 11 genera of soritid foraminifera were investigated by sequencing the complete SSU rDNA gene for 25 specimens. Additionally, partial SSU rDNA sequences were obtained from another 15 specimens of Soritinae. DNA sequence analysis confirms the monophyly of each family. Caribbean Archaiasinae form a monophyletic clade with Pacific Laevipeneroplis at the base. The genus Parasorites apppears as a sister taxa to Soritinae. Complex morphological features that characterize the genus Marginopora seem to have evolved independently at least twice, as the examined representatives cluster within two other soritine genera. Molecular analysis further shows that Sorites orbiculus and Sorites marginalis represent two different morphotypes of one species. Our data indicate that morphological changes and acquisition of new endosymbiont types in each group played an important role in the adaptation and radiation of Soritacea.

Small subunit ribosomal DNA suggests that the Xenophyophorean Syringammina corbicula is a foraminiferan

Abstract Xenophyophorea are giant deep-sea rhizopodial protists of enigmatic origins. Although species were described as Foraminifera or sponges in the early literature, the xenophyophoreans are currently classified either as a class of Rhizopoda or an independent phylum. To establish the phylogenetic position of Xenophyophorea, we analysed the small subunit (SSU) rRNA gene sequence of Syringammina corbicula Richardson, a newly described xenophyophorean species from the Cape Verde Plateau. The SSUrDNA analyses showed that S. corbicula is closely related to Rhizammina algaeformis, a tubular deep-sea foraminiferan. Both species branch within a group of monothalamous (single-chambered) Foraminifera, which include also such agglutinated genera as Toxisarcon, Rhabdammina, and Saccammina, and the organic-walled genera Gloiogullmia and Cylindrogullmia. Our results are congruent with observations of similar cytoplasmic organisation in Rhizammina and Syringammina. Thus, the Xenophyophorea appear to be a highly specialised group of deep-sea Foraminifera.