Agnes Weiner

The cryptic and the apparent reversed: lack of genetic differentiation within the morphologically diverse plexus of the planktonic foraminifer Globigerinoides sacculifer

Abstract Previous genetic studies of extant planktonic foraminifera have provided evidence that the traditional, strictly morphological definition of species in these organisms underestimates their biodiversity. Here, we report the first case where this pattern is reversed. The modern (sub)tropical species plexus Globigerinoides sacculifer is characterized by large morphological variability, which has led to the proliferation of taxonomic names attributed to morphological end-members within the plexus. In order to clarify the taxonomic status of its morphotypes and to investigate the genetic connectivity among its currently partly disjunct (sub)tropical populations, we carried out a global survey of two ribosomal RNA regions (SSU and ITS-1) in all recent morphotypes of the plexus collected throughout (sub)tropical surface waters of the global ocean. Unexpectedly, we find an extremely reduced genetic variation within the plexus and no correlation between genetic and morphological divergence, suggesting taxonomical overinterpretation. The genetic homogeneity within the morphospecies is unexpected, considering its partly disjunct range in the (sub)tropical Atlantic and Indo-Pacific and its old age (early Miocene). A sequence variant in the rapidly evolving ITS-1 region indicates the existence of an exclusively Atlantic haplotype, which suggests an episode of relatively recent (last glacial) isolation, followed by subsequent resumption of unidirectional gene flow from the Indo-Pacific into the Atlantic. This is the first example in planktonic foraminifera where the morphological variability in a morphospecies exceeds its rDNA genetic variability. Such evidence for inconsistent scaling of morphological and genetic diversity in planktonic foraminifera could complicate the interpretation of evolutionary patterns in their fossil record.

Vertical niche partitioning between cryptic sibling species of a cosmopolitan marine planktonic protist.

A large portion of the surface‐ocean biomass is represented by microscopic unicellular plankton. These organisms are functionally and morphologically diverse, but it remains unclear how their diversity is generated. Species of marine microplankton are widely distributed because of passive transport and lack of barriers in the ocean. How does speciation occur in a system with a seemingly unlimited dispersal potential? Recent studies using planktonic foraminifera as a model showed that even among the cryptic genetic diversity within morphological species, many genetic types are cosmopolitan, lending limited support for speciation by geographical isolation. Here we show that the current two‐dimensional view on the biogeography and potential speciation mechanisms in the microplankton may be misleading. By depth‐stratified sampling, we present evidence that sibling genetic types in a cosmopolitan species of marine microplankton, the planktonic foraminifer Hastigerina pelagica, are consistently separated by depth throughout their global range. Such strong separation between genetically closely related and morphologically inseparable genetic types indicates that niche partitioning in marine heterotrophic microplankton can be maintained in the vertical dimension on a global scale. These observations indicate that speciation along depth (depth‐parapatric speciation) can occur in vertically structured microplankton populations, facilitating diversification without the need for spatial isolation.

Phylogeography of the tropical planktonic foraminifera lineage Globigerinella reveals isolation inconsistent with passive dispersal by ocean currents

Morphologically defined species of marine plankton often harbor a considerable level of cryptic diversity. Since many morphospecies show cosmopolitan distribution, an understanding of biogeographic and evolutionary processes at the level of genetic diversity requires global sampling. We use a database of 387 single-specimen sequences of the SSU rDNA of the planktonic foraminifera Globigerinella as a model to assess the biogeographic and phylogenetic distributions of cryptic diversity in marine microplankton on a global scale. Our data confirm the existence of multiple, well isolated genetic lineages. An analysis of their abundance and distribution indicates that our sampling is likely to approximate the actual total diversity. Unexpectedly, we observe an uneven allocation of cryptic diversity among the phylogenetic lineages. We show that this pattern is neither an artifact of sampling intensity nor a function of lineage age. Instead, we argue that it reflects an ongoing speciation process in one of the three major lineages. Surprisingly, four of the six genetic types in the hyperdiverse lineage are biogeographically restricted to the Indopacific. Their mutual co-occurrence and their hierarchical phylogenetic structure provide no evidence for an origin through sudden habitat fragmentation and their limitation to the Indopacific challenges the view of a global gene flow within the warm-water provinces. This phenomenon shows that passive dispersal is not sufficient to describe the distribution of plankton diversity. Rather, these organisms show differentiated distribution patterns shaped by species interactions and reflecting phylogenetic contingency with unique histories of diversification rates.

PFR2: a curated database of planktonic foraminifera 18S ribosomal DNA as a resource for studies of plankton ecology, biogeography and evolution

Planktonic foraminifera (Rhizaria) are ubiquitous marine pelagic protists producing calcareous shells with conspicuous morphology. They play an important role in the marine carbon cycle, and their exceptional fossil record serves as the basis for biochronostratigraphy and past climate reconstructions. A major worldwide sampling effort over the last two decades has resulted in the establishment of multiple large collections of cryopreserved individual planktonic foraminifera samples. Thousands of 18S rDNA partial sequences have been generated, representing all major known morphological taxa across their worldwide oceanic range. This comprehensive data coverage provides an opportunity to assess patterns of molecular ecology and evolution in a holistic way for an entire group of planktonic protists. We combined all available published and unpublished genetic data to build PFR2, the Planktonic foraminifera Ribosomal Reference database. The first version of the database includes 3322 reference 18S rDNA sequences belonging to 32 of the 47 known morphospecies of extant planktonic foraminifera, collected from 460 oceanic stations. All sequences have been rigorously taxonomically curated using a six‐rank annotation system fully resolved to the morphological species level and linked to a series of metadata. The PFR2 website, available at http://pfr2.sb-roscoff.fr, allows downloading the entire database or specific sections, as well as the identification of new planktonic foraminiferal sequences. Its novel, fully documented curation process integrates advances in morphological and molecular taxonomy. It allows for an increase in its taxonomic resolution and assures that integrity is maintained by including a complete contingency tracking of annotations and assuring that the annotations remain internally consistent.

Methodology for single-cell genetic analysis of planktonic foraminifera for studies of protist diversity and evolution

Single-cell genetic analysis is an essential method to investigate the biodiversity and evolutionary ecology of marine protists. In protist groups that do not reproduce under laboratory conditions, this approach provides the only means to directly associate molecular sequences with cell morphology. The resulting unambiguous taxonomic identification of the DNA sequences is a prerequisite for barcoding and analyses of environmental metagenomic data. Extensive single-cell genetic studies have been carried out on planktonic foraminifera over the past 20 years to elucidate their phylogeny, cryptic diversity, biogeography and the relationship between genetic and morphological variability. In the course of these investigations, it has become evident that genetic analysis at the individual specimen level is confronted by innumerable challenges ranging from the negligible amount of DNA present in the single cell to the substantial amount of DNA contamination introduced by endosymbionts or food particles. Consequently, a range of methods has been developed and applied throughout the years for the genetic analysis of planktonic foraminifera in order to enhance DNA amplification success rates. Yet, the description of these methods in the literature rarely occurred with equivalent levels of detail and the different approaches have never been compared in terms of their efficiency and reproducibility. Here, aiming at a standardization of methods, we provide a comprehensive review of all methods that have been employed for the single-cell genetic analysis of planktonic foraminifera. We compile data on success rates of DNA amplification and use these to evaluate the effects of key parameters associated with the methods of sample collection, storage and extraction of single-cell DNA. We show that the chosen methods influence the success rates of single-cell genetic studies, but the differences between them are not sufficient to hinder comparisons between studies carried out by different methods. The review thus not only provides a comprehensive reference with guidelines for future genetic studies on foraminifera, but it also establishes an important benchmark for investigations using existing single-cell datasets. The methods are widely applicable and the review may help to establish similar standard principles for their utilization in other protist groups.

Caught in the act: anatomy of an ongoing benthic–planktonic transition in a marine protist

The transition from benthos to plankton requires multiple adaptations, yet so far it remains unclear how these are acquired in the course of the transition. To investigate this process, we analyzed the genetic diversity and distribution patterns of a group of foraminifera of the genus Bolivina with a tychopelagic mode of life (same species occurring both in benthos and plankton). We assembled a global sequence data set for this group from single-cell DNA extractions and occurrences in metabarcodes from pelagic environmental samples. The pelagic sequences all cluster within a single monophyletic clade within Bolivina. This clade harbors three distinct genetic lineages, which are associated with incipient morphological differentiation. All lineages occur in the plankton and benthos, but only one lineage exhibits no limit to offshore dispersal and has been shown to grow in the plankton. These observations indicate that the emergence of buoyancy regulation within the clade preceded the evolution of pelagic feeding and that the evolution of both traits was not channeled into a full transition into the plankton. We infer that in foraminifera, colonization of the planktonic niche may occur by sequential cooptation of independently acquired traits, with holoplanktonic species being recruited from tychopelagic ancestors