Genetic structure of the protist Physarum albescens (Amoebozoa) revealed by multiple markers and genotyping by sequencing

Myxomycetes are terrestrial protists with many presumably cosmopolitan species dispersing via airborne spores. A truly cosmopolitan species would suffer from outbreeding depression hampering local adaptation, while locally adapted species with limited distribution would be at a higher risk of extinction in changing environments. Here, we investigate intraspecific genetic diversity and phylogeography of Physarum albescens over the entire Northern Hemisphere. We sequenced 324 field collections of fruit bodies for 1-3 genetic markers (SSU, EF1A, COI) and analysed 98 specimens with genotyping by sequencing. The structure of the three-gene phylogeny, SNP-based phylogeny, phylogenetic networks, and the observed recombination pattern of three independently inherited gene markers can be best explained by the presence of at least 18 reproductively isolated groups, which can be seen as cryptic species. In all intensively sampled regions and in many localities, members of several phylogroups coexisted. Some phylogroups were found to be abundant in only one region and completely absent in other well-studied regions, and thus may represent regional endemics. Our results demonstrate that the widely distributed myxomycete species Ph. albescens represents a complex of at least 18 cryptic species, and some of these seem to have a limited geographical distribution. In addition, the presence of groups of presumably clonal specimens suggests that sexual and asexual reproduction coexist in natural populations of myxomycetes

Unexplored diversity of microscopic myxomycetes: evidence from environmental DNA

Background and aims – Recent studies showed the position of two slime mould species with microscopic sporocarps, Echinosteliopsis oligospora and Echinostelium bisporum, within the class Myxomycetes. These minute species are seldom seen in studies based on detection of sporocarps and can easily be confused with protosteloid amoebozoans.Methods – We searched all published ePCR data sets that targeted myxomycete 18S rDNA for the presence of environmental sequences similar to E. oligospora and Echinosteliales in traditional circumscription, and performed phylogenetic analyses that included short environmental sequences and full-length 18S rDNA sequences representing all the major groups of myxomycetes.Key results – We report 19 unique sequences which are closely related to E. bisporum or E. oligospora based on sequence similarity (73.1–95.2% similarity) and which form well-supported monophyletic clades with these species in phylogenetic analyses. They may represent new species that are not yet described. Our phylogeny based on full-length 18S rDNA sequences further confirms the position of E. bisporum and E. oligospora within myxomycetes and the paraphyly of the order Echinosteliales in its traditional circumscription.Conclusions – Our results show that ePCR-based studies can reveal myxomycete taxa that often escape detection by traditional approaches, including potentially new species, and thus provide valuable new data on diversity and ecology of myxomycetes. As such, strategies for studying myxomycetes biodiversity should be revised, focusing also on molecular detection techniques in addition to the sporocarp-based ones

<b>New genus </b><b><i>Valtocarpus</i></b><b> (Myxomycetes = Eumycetozoa): molecular phylogeny and morphological analysis of aethalioid species in the order Stemonitidales</b>

The dataset includes:Fig. 1 with edited three-gene phylogenetic tree in PDFThe Table S1 with the list of specimens used for DNA extraction and sequencing and the list of sequences retrieved from GenBankThe File S1 with concatenated alignmentPartition file File S2 in RAxML formatUnedited three-gene phylogenetic tree File S3</p

Two independent genetic markers support separation of the myxomycete <i>Lycogala epidendrum</i> into numerous biological species

Lycogala epidendrum is one of the most widely known myxomycete species and the first-ever discovered representative of this group. Using 687 original DNA sequences from 330 herbarium specimens from Europe, Asia, North and Central America, and Australia, we constructed the first detailed phylogenies of the genus Lycogala, based on two independently inherited genetic markers, the ribosome small subunit 18S rRNA nuclear gene (18S rDNA) and the mitochondrial cytochrome oxidase subunit I gene (COI). In both phylogenies, L. epidendrum appeared to be a polyphyletic group, represented by numerous clades. The four other recognized species of the genus (L. confusum, L. conicum, L. exiguum, and L. flavofuscum) are scattered between branches corresponding to L. epidendrum. A barcode gap analysis revealed 60 18S rDNA phylogroups of L. epidendrum, which are distant from each other not less than from other species of the genus Lycogala. For 18 of these phylogroups with both 18S rDNA and COI sequences available, recombination patterns were analyzed to test for reproductive isolation. In contrast to the results of a simulation assuming panmixis, no crossing between ribosomal and mitochondrial phylogroups was found, thus allowing the conclusion that all tested phylogroups represent biospecies. More than one third (39.6%) of the studied specimens share a single 18S rDNA phylogroup, which we consider to be L. epidendrum s. str. This group displays the broadest geographic distribution and the highest intraspecific genetic variability. Nearly all (93.3%) of the remaining non-singleton 18S rDNA phylogroups are restricted to certain continents or even regions. At the same time, various reproductively isolated phylogroups occur sympatric at a given location.</p

<b><i>Diachea racemosa</i></b><b> (Myxomycetes = Myxogastrea): a new species with cespitose sporocarps from southern Vietnam and its position within the phylogenetic clade </b><b><i>Diachea</i></b><b> sensu lato (Physarales)</b>

The dataset includes:Fig. 1 and Fig. 2 with edited three-gene phylogenetic trees in PDFThe Table S1 with the list of specimens used for DNA extraction and sequencing and the list of sequences retrieved from GenBankThe File S1 with concatenated alignmentPartition file File S2 in RAxML formatUnedited three-gene phylogenetic tree File S3</p

Genetic structure of the protist Physarum albescens (Amoebozoa) revealed by multiple markers and genotyping by sequencing

Abstract Myxomycetes are terrestrial protists with many presumably cosmopolitan species dispersing via airborne spores. A truly cosmopolitan species would suffer from outbreeding depression hampering local adaptation, while locally adapted species with limited distribution would be at a higher risk of extinction in changing environments. Here, we investigate intraspecific genetic diversity and phylogeography of Physarum albescens over the entire Northern Hemisphere. We sequenced 324 field collections of fruit bodies for 1–3 genetic markers (SSU, EF1A, COI) and analysed 98 specimens with genotyping by sequencing. The structure of the three‐gene phylogeny, SNP‐based phylogeny, phylogenetic networks, and the observed recombination pattern of three independently inherited gene markers can be best explained by the presence of at least 18 reproductively isolated groups, which can be seen as cryptic species. In all intensively sampled regions and in many localities, members of several phylogroups coexisted. Some phylogroups were found to be abundant in only one region and completely absent in other well‐studied regions, and thus may represent regional endemics. Our results demonstrate that the widely distributed myxomycete species Ph. albescens represents a complex of at least 18 cryptic species, and some of these seem to have a limited geographical distribution. In addition, the presence of groups of presumably clonal specimens suggests that sexual and asexual reproduction coexist in natural populations of myxomycetes

Strategies for Intraspecific Genotyping of Duckweed: Comparison of Five Orthogonal Methods Applied to the Giant Duckweed Spirodela polyrhiza

The predominantly vegetative propagating duckweeds are of growing commercial interest. Since clonal accessions within a respective species can vary considerably with respect to their physiological as well as biochemical traits, it is critical to be able to track the clones of species of interest after their characterization. Here, we compared the efficacy of five different genotyping methods for Spirodela polyrhiza, a species with very low intraspecific sequence variations, including polymorphic NB-ARC-related loci, tubulin-gene-based polymorphism (TBP), simple sequence repeat variations (SSR), multiplexed ISSR genotyping by sequencing (MIG-seq), and low-coverage, reduced-representation genome sequencing (GBS). Four of the five approaches could distinguish 20 to 22 genotypes out of the 23 investigated clones, while TBP resolved just seven genotypes. The choice for a particular method for intraspecific genotyping can depend on the research question and the project budget, while the combination of orthogonal methods may increase the confidence and resolution for the results obtained

Parahydrogen-induced polarization of 14N nuclei

Hyperpolarization techniques provide a dramatic increase in sensitivity of nuclear magnetic resonance spectroscopy and imaging. In spite of the outstanding progress in solution-state hyperpolarization of spin-1/2 nuclei, hyperpolarization of quadrupolar nuclei remains challenging. Here, hyperpolarization of quadrupolar 14N nuclei with natural isotopic abundance of >99% is demonstrated. This is achieved via pairwise addition of parahydrogen to tetraalkylammonium salts with vinyl or allyl unsaturated moieties followed by a subsequent polarization transfer from 1H to 14N nuclei at high magnetic field using PH-INEPT or PH-INEPT+ radiofrequency pulse sequence. Catalyst screening identified water-soluble rhodium complex [Rh(P(m-C6H4SO3Na)3)3Cl] as the most efficient catalyst for hyperpolarization of the substrates under study, providing up to 1.3% and up to 6.6% 1H polarization in the cases of vinyl and allyl precursors, respectively. The performance of PH-INEPT and PH-INEPT+ pulse sequences was optimized with respect to interpulse delays, and the resultant experimental dependences were in good agreement with simulations. As a result, 14N NMR signal enhancement of up to 760-fold at 7.05 T (corresponding to 0.15% 14N polarization) was obtained