Mitchell, Edward

2013, Heger, Thierry J., Mitchell, Edward, Leander, Brian S.

Although free-living protists play essential roles in aquatic and soil ecology, little is known about their diversity and phylogeography, especially in terrestrial ecosystems. We used mitochondrial cytochrome c oxidase subunit 1 (COI) gene sequences to investigate the genetic diversity and phylogeography of the testate amoeba morphospecies Hyalosphenia papilio in 42 Sphagnum (moss)-dominated peatlands in North America, Europe and Asia. Based on 1% sequence divergence threshold, our results from single-cell PCRs of 301 individuals revealed 12 different genetic lineages and both the general mixed Yule-coalescent (GMYC) model and the automatic barcode gap discovery (ABGD) methods largely support the hypothesis that these 12 H.papilio lineages correspond to evolutionary independent units (i.e. cryptic species). Our data also showed a high degree of genetic heterogeneity within different geographical regions. Furthermore, we used variation partitioning based on partial redundancy analyses (pRDA) to evaluate the contributions of climate and dispersal limitations on the distribution patterns of the different genetic lineages. The largest fraction of the variation in genetic lineage distribution was attributed to purely climatic factors (21%), followed by the joint effect of spatial and bioclimatic factors (13%), and a purely spatial effect (3%). Therefore, these data suggest that the distribution patterns of H.papilio genetic lineages in the Northern Hemisphere are more influenced by climatic conditions than by dispersal limitations.

2012, Gomaa, Fatma, Todorov, Milcho, Heger, Thierry J., Mitchell, Edward, Lara, Enrique

The systematics of lobose testate amoebae (Arcellinida), a diverse group of shelled free-living unicellular eukaryotes, is still mostly based on morphological criteria such as shell shape and composition. Few molecular phylogenetic studies have been performed on these organisms to date, and their phylogeny suffers from typical under-sampling artefacts, resulting in a still mostly unresolved tree. In order to clarify the phylogenetic relationships among arcellinid testate amoebae at the inter-generic and inter-specific level, and to evaluate the validity of the criteria used for taxonomy, we amplified and sequenced the SSU rRNA gene of nine taxa - Difflugia bacillariarum, D. hiraethogii, D. acuminata, D. lanceolata, D. achlora, Bullinularia gracilis, Netzelia oviformis, Physochila griseola and Cryptodifflugia oviformis. Our results, combined with existing data demonstrate the following: 1) Most arcellinids are divided into two major clades, 2) the genus Difflugia is not monophyletic, and the genera Netzelia and Arcella are closely related, and 3) Cryptodifflugia branches at the base of the Arcellinida clade. These results contradict the traditional taxonomy based on shell composition, and emphasize the importance of general shell shape in the taxonomy of arcellinid testate amoebae. (C) 2012 Elsevier GmbH. All rights reserved.

2012, Gomaa, Fatma, Todorov, Milcho, Heger, Thierry J., Mitchell, Edward, Lara, Enrique

The systematics of lobose testate amoebae (Arcellinida), a diverse group of shelled free-living unicellular eukaryotes, is still mostly based on morphological criteria such as shell shape and composition. Few molecular phylogenetic studies have been performed on these organisms to date, and their phylogeny suffers from typical under-sampling artefacts, resulting in a still mostly unresolved tree. In order to clarify the phylogenetic relationships among arcellinid testate amoebae at the inter-generic and inter-specific level, and to evaluate the validity of the criteria used for taxonomy, we amplified and sequenced the SSU rRNA gene of nine taxa - Difflugia bacillariarum, D. hiraethogii, D. acuminata, D. lanceolata, D. achlora, Bullinularia gracilis, Netzelia oviformis, Physochila griseola and Cryptodifflugia oviformis. Our results, combined with existing data demonstrate the following: 1) Most arcellinids are divided into two major clades, 2) the genus Difflugia is not monophyletic, and the genera Netzelia and Arcella are closely related, and 3) Cryptodifflugia branches at the base of the Arcellinida clade. These results contradict the traditional taxonomy based on shell composition, and emphasize the importance of general shell shape in the taxonomy of arcellinid testate amoebae.

2011, Heger, Thierry J., Pawlowski, Jan, Lara, Enrique, Leander, Brian S., Todorov, Milcho, Golemansky, Vassil, Mitchell, Edward

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.

2013, Kosakyan, Anush, Gomaa, Fatma, Mitchell, Edward, Heger, Thierry J., Lara, Enrique

Species identification by means of morphology is often problematic in protists. Nebela tincta-collaris-bohemica (Arcellinida) is a species complex of small to medium-sized (ca. 100 mu m) testate amoebae common in peat bogs and forest soils. The taxonomic validity of characters used to define species within this group is debated and causes confusion in studies of biogeography, and applications in palaeoecology. We examined the relationship between morphological and genetic diversity within this species complex by combined analyses of light microscopy imaging and Cytochrome Oxidase Subunit 1(COI) sequences obtained from the same individual amoeba cells. Our goals were (1) to clarify the taxonomy and the phylogenetic relationships within this group, and (2) to evaluate if individual genotypes corresponded to specific morphotypes and the extent of phenotypic plasticity. We show here that small variations in test morphology that have been often overlooked by traditional taxonomy correspond to distinct haplotypes. We therefore revise the taxonomy of the group. We redefine Nebela tincta (Leidy) Kosakyan et Lara and N. collaris (Ehrenberg 1848) Kosakyan et Gomaa, change N. tincta var. rotunda Penard to N. rotunda (Penard 1890), describe three new species: N. guttata n. sp. Kosakyan et Lara, N. pechorensis n. sp. Kosakyan et Mitchell, and N. aliciae n. sp. Mitchell et Lara. (C) 2012 Elsevier GmbH. All rights reserved.

2012, Heger, Thierry J., Straub, Francois, Mitchell, Edward

Testate amoebae (Arcellinida and Euglyphida) and diatoms (Bacillariophyta) respond to different ecological gradients. These protists are useful tools for biomonitoring and paleoecological studies in aquatic and terrestrial ecosystems. However, little is known about the responses of these micro-eukaryotes to soil management practices. We analyzed the testate amoeba and diatom communities from the DOK-trial (D: biodynamic, O: bio-organic, K: german "konventionell" integrated conventional) agricultural experiment at Therwil, Switzerland. Soil samples were collected from biodynamic and conventional plots and subsequently incubated for four months in a growth chamber. The diatom diversity tended to be higher in the biodynamic than in the two conventional systems. Redundancy analysis (RDA) suggested that diatom community structure differed between organic and the two conventional systems. Testate amoeba abundance was about five times higher in biodynamic than in conventional systems (P < 0.05) but no significant differences in diversity were reported between treatments. Altogether, these data suggest that diatoms and testate amoebae are sensitive to farming systems. As direct analyses of soil samples are time-consuming, molecular tools would be very useful for further development of the use of protists in bioindication. Crown Copyright (C) 2011 Published by Elsevier Masson SAS. All rights reserved.

2011, Lara, Enrique, Heger, Thierry J., Scheihing, Rodrigo, Mitchell, Edward

Aim Propagule size and ecological requirements are believed to be major factors influencing the passive dispersal of free-living terrestrial protists. We compared the colonization potential of three closely related testate amoeba species (Assulina muscorum, A. seminulum, A. scandinavica, ranging from 40 to 100 mu m in length). Location Europe. Methods We collected individual Assulina species cells from Sphagnum peatlands across Europe. We sequenced a 550-bp fragment of the mitochondrial cytochrome c oxidase subunit I gene (COI) to estimate the within-species variability, as a proxy for gene flow. We reviewed existing ecological and palaeoecological data to assess the ecological tolerance of Assulina species and how rapidly they colonized developing peatlands. Results We obtained COI sequences for 30 individuals of A. seminulum from eleven sites, for 39 of A. muscorum from six sites, and for six of A. scandinavica from two sites. We observed three haplotypes for A. seminulum and two for A. muscorum, often co-existing in the same sites. The sequences of A. scandinavica from the two sites were identical. Assulina muscorum and A. seminulum haplotypes varied by only 1-2 nucleotides, resulting in > 99.5% similarity. Genetic diversity within A. seminulum was higher than that within A. muscorum. Ecological and palaeoecological records showed that A. muscorum was much more frequent and abundant than A. seminulum, and had a somewhat broader ecological tolerance for pH, moisture and water-table depth. Assulina muscorum always appeared early during the developmental history of peatlands, either before or simultaneously with A. seminulum. Main conclusions The lack of genetic structure observed with a variable marker such as COI suggests high gene flow between the sites and thus rapid transport (at an evolutionary scale) over large distances, in agreement with the palaeoecological records. Thus, geographical distance alone does not seem to prevent the dispersal of testate amoebae, at least within Europe. Nevertheless, genetic diversity was significantly lower within A. muscorum than within A. seminulum, suggesting that its smaller size and abundance and/or broader ecological tolerance influence its effective dispersal capacity. These results are in agreement with the often earlier colonization of peatlands by A. muscorum and its broader ecological tolerance.

2012, Heger, Thierry J., Imfeld, Gwenael, Mitchell, Edward

2012, Kosakyan, Anush, Heger, Thierry J., Leander, Brian S., Todorov, Milcho, Mitchell, Edward, Lara, Enrique

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.

2011, Heger, Thierry J., Booth, Robert K., Sullivan, Maura E., Wilkinson, David M., Warner, Barry G., Asada, Taro, Mazei, Yuri, Meisterfeld, Ralf, Mitchell, Edward

Aim The question whether free-living protists are generally cosmopolitan is currently a matter of debate. In this study we investigate the geographical distribution of a distinctive testate amoeba species, Nebela ansata, and use our data to assess the potential for highly restricted distribution patterns in some protist species. Location Global. Methods We analysed (1) 3400 testate amoeba publications from North America and other continents, (2) unpublished slides of the Penard Collection of the Natural History Museum, London, UK, and (3) 104 Sphagnum samples from eastern North America. Non-metric multidimensional scaling (NMDS) was used to visualize the similarities in testate amoeba community composition among 1012 North American samples, including two communities that contained N. ansata. Results We rediscovered N. ansata at a site in New Jersey located close to its type locality, and in Nova Scotia. We also report the existence of an apparently unpublished museum specimen originally collected from New Jersey. Our extensive literature survey confirmed the presence of this species only in the temperate part of eastern North America. The NMDS revealed that communities with N. ansata were less similar to each other than to communities from other parts of North America, suggesting that favourable habitats for N. ansata occur in other Sphagnum-dominated peatlands, a habitat type that has been extensively sampled in North America and elsewhere. Main conclusions These data provide an unusually convincing case of a free-living microorganism with a very limited distribution range in the temperate part of eastern North America. The remarkably restricted distribution of N. ansata highlights the extent of our ignorance about the natural history of free-living microorganisms, and raises questions about the lack of attention to microbial diversity in conservation biology.