COI Barcoding of Nebelid Testate Amoebae (Amoebozoa: Arcellinida): Extensive Cryptic Diversity and Redefinition of the Hyalospheniidae Schultze

We used Cytochrome Oxidase Subunit 1 (COI) to assess the phylogenetic relationships and taxonomy of Nebela sensu stricto and similar taxa (Nebela group, Arcellinida) in order to clarify the taxonomic validity of morphological characters. The COI data not only successfully separated all studied morphospecies but also revealed the existence of several potential cryptic species. The taxonomic implications of the results are: (1) Genus Nebela is paraphyletic and will need to be split into at least two monophyletic assemblages when taxon sampling is further expanded. (2) Genus Quadrulella, one of the few arcellinid genera building its shell from self-secreted siliceous elements, and the mixotrophic Hyalosphenia papilio branch within the Nebela group in agreement with the general morphology of their shell and the presence of an organic rim around the aperture (synapomorphy for Hyalospheniidae). We thus synonymise Hyalospheniidae and Nebelidae. Hyalospheniidae takes precedence and now includes Hyalosphenia, Quadrulella (previously in the Lesquereusiidae) and all Nebelidae with the exception of Argynnia and Physochila. Leptochlamys is Arcellinida incertae sedis. We describe a new genus Padaungiella Lara et Todorov and a new species Nebela meisterfeldi n. sp. Heger et Mitchell and revise the taxonomic position (and rank) of several taxa. These results show that the traditional morphology-based taxonomy underestimates the diversity within the Nebela group, and that phylogenetic relationships are best inferred from shell shape rather than from the material used to build the shell. (C) 2011 Elsevier GmbH. All rights reserved.

Contrasting processes drive alpha and beta taxonomic, functional and phylogenetic diversity of orthopteran communities in grasslands

Bertrand Fournier, François Gillet

Taxonomic, functional and phylogenetic diversities can respond differently to biotic and abiotic filters. However, biodiversity management tends to focus on a single index, generally taxonomic diversity, assuming a strong positive correlation among biodiversity components across scales. This can result in a mismanagement of functional and phylogenetic diversities with negative consequences for ecosystem functioning and long-term maintenance of services. Understanding the relationships among biodiversity components, how they change across scales and which their main drivers are can lead to more sustainable management of biodiversity and its associated ecosystem services and functions. We used an integrative approach of biodiversity where we investigated alpha and beta taxonomic (TD), functional (FD) and phylogenetic (PD) diversity of orthopteran communities as well as species and functional traits composition and their associated drivers at the local and landscape scales in permanent mesic grasslands of the French Jura Mountains. We assessed whether orthopteran TD, FD and PD were positively correlated. We also determined the drivers of TD, FD and PD and their changes across scales and among indices using data related to soil, agricultural practices, elevation, and biotic interactions with plants. Our results showed that (i) elevation was a strong driver of orthopteran community trait composition, (ii) orthopteran alpha TD, FD and PD were correlated among themselves and increased with plant species richness, and (iii) local beta diversity was not correlated with alpha diversity. Beta diversity had different drivers at local and landscape scale: it was influenced by soil chemistry and texture at the local scale and increased with the difference in elevation among plots at the landscape scale. This study evidences distinct processes driving alpha and beta diversity of orthopteran communities at both the local and landscape scales. It supports the hypothesis that less intensive agricultural practices enhance orthopteran diversity and highlights the importance to consider beta diversity at both local and landscape scale when designing and assessing the management regimes of grassland ecosystems. Prioritizing the importance of the different biodiversity components and spatial scales constitutes an important challenge for sustainable grassland management.

Elsevier Science Bv2017

Comparing COI and SSU rDNA barcodes for assessing the diversity and phylogenetic relationships of cyphoderiid testate amoebae (Rhizaria: Euglyphida)

Thierry Jean Heger, Enrique Lara, Edward Mitchell

The mitochondrial Cytochrome Oxidase Subunit 1 gene (COI) has been promoted as an ideal “DNA barcode” for animal species and other groups of eukaryotes. However, the utility of the COI marker for species level discrimination and for phylogenetic analyses has yet to be tested within the Rhizaria. Accordingly, we analysed mitochondrial COI gene sequences and nuclear small subunit rDNA (SSU) sequences from several morphospecies of euglyphid testate amoebae (Cercozoa, Rhizaria) in order to evaluate the utility of these DNA markers for species discrimination and phylogenetic reconstructions. Sequences were obtained from eleven populations belonging to six Cyphoderia morphospecies that were isolated from field samples in North America and Europe. Mean inter-population COI sequence dissimilarities were on average 2.9 times greater than in the SSU, while the intra-population sequence dissimilarities were higher in the SSU (0-0.95 %) than in the COI (0%); this suggests that the COI fragment is valuable for discriminating Cyphoderiidae isolates. Our study also demonstrated that COI sequences are useful for inferring phylogenetic relationships among Cyphoderiidae isolates. COI and SSU tree topologies were very similar even though the COI fragment used in these analyses (500 bp) was much shorter than the SSU sequences (1600 bp). Altogether, these results demonstrate the utility of the COI as a potential taxonomic DNA barcode for assessing cyphoderiid species diversity and for inferring phylogenetic relationships within the group.

2011

Biogeography, Evolution and Diversity of Free-Living Protists

Thierry Jean Heger

Although widely recognized as essential actors of ecosystem processes and representing a significant part of the Earth's biodiversity, free-living protists are poorly known. Major questions concerning their diversity, ecology, and evolution remain completely unresolved, partly because our knowledge of their taxonomy, at each different levels, is still extremely limited. In this thesis, I used testate amoebae as model organisms to get insights into the diversity and biogeography of protists. Testate amoebae are a polyphyletic group of free-living protists that are characterized by a shell composed of proteinaceous, calcareous, or siliceous material. The shells are differentiated by their shapes and ultrastructures, allowing for the identification of morphospecies more easily than in the vast majority of other protists. However, the taxonomic validity of the characters used to define morphospecies and the true diversity of testate amoebae are largely unknown. We studied the taxonomy of cyphoderiid testate amoebae (Rhizaria: Euglyphida), by combining morphological and molecular genetic approaches (chapters 2 and 3). We used light- and scanning electron microscopy to analyze the shell ultrastructure and biometry of several Cyphoderia morphospecies and sequenced gene portions of the Cytochrome Oxidase Subunit 1 (COI) and nuclear small subunit rDNA (SSU). These analyses revealed that while several described species are morphologically distinct and well-supported by the genetic data, other species, such as Cyphoderia ampulla, represent a complex of cryptic or pseudo-cryptic species. The presence of cryptic or pseudo-cryptic Cyphoderia species illustrates the limits of a morphology-based approach for assessing protist diversity using only characters visible by light microscopy. DNA sequence analyses may allow for a better resolution for both species identification and diversity estimates, provided that appropriate genetic markers can be developed. Genetic marker development in protists is particularly challenging because their representation in the international genomic databases is still very limited. The SSU marker also allowed us to build an extensive phylogeny of eulgyhid testate amoebae. We then used this phylogeny as a framework to analyze macro-evolutionary processes in testate amoebae, in particular, how easily a species can cross an ecological barrier and colonize new habitats. The taxa included in our phylogeny occurred either in marine, marine supralittoral or freshwater habitats, and species of each habitat segregated into distinct sub-clades. This indicates that transitions between these environments occur only rarely in the course of evolution (chapter 4). A reliable taxonomy for microorganisms is not only an essential prerequisite for estimating species diversities, it would also be a key component to resolve the long-lasting debate of whether free-living microorganisms are strictly cosmopolitan, or whether at least some species have limited geographic distributions (chapter 1). We analyzed Arcellinida and Euglyphida testate amoeba communities from Amsterdam Island, one of the most remote islands in the world located in the Indian Ocean, to search for endemic species with limited geographic distributions (chapter 5). In macroorganisms, the degree of endemism is generally high in such remote islands. Our extensive inventory of testate amoeba morphospecies provided no clear evidence for endemism. However, given that the morphology-based species identification gives only a limited taxonomic resolution, we cannot exclude the presence of morphologically non-distinctive endemic species (cryptic species). Thus, such taxonomic uncertainties undermine biogeographical studies of testate amoebae and other protists. To circumvent taxonomical uncertainties in biogeographical studies, we analyzed the distribution of a morphologically highly distinctive testate amoeba, Nebela ansata (chapter 6). We report the presence of this species in Nova Scotia, Canada and at sites in New Jersey where it was initially described. An extensive literature survey further confirmed the absence of this species from both, the remaining North American continent as well as other continents. Together, these data provide an unusually convincing case of a free-living microorganism with a very limited distribution range along the temperate East American coast. Overall, my thesis illustrates how, in most cases, the current morphology-based species identification greatly underestimates testate amoeba diversity and calls for an improvement of the taxonomic resolution and reliability. The extension of the current taxonomy using molecular-genetic tools should greatly contribute to this task and also shed light on the debate on the distribution of protists and other microorganisms. Together, such studies will provide a first step towards a better understanding of the diversity and evolution of microorganisms and improve their utility as model organisms in both fundamental and applied questions.