Reassessment of the Putative Ciliate Fossils Eotintinnopsis, Wujiangella, and Yonyangella from the Neoproterozoic Doushantuo Formation in China

Three putative ciliate fossils were described from the Neoproterozoic Doushantuo Formation in China: Eotintinnopsis, Wujiangella, and Yonyangella. The identity of these fossils is important for our understanding of the origins and early morphological evolution within ciliate clades. Here we compare the homology of the fossil characteristics with those in their proposed ciliate relatives. Eotintinnopsis resembles a tintinnid, but its feathery tentacle-like apical structure is probably not homologous within any known ciliate. Wujiangella presents homology issues with the size and distribution of its putative somatic cilia. Yonyangella appears to be a suctorian with its tentaclelike structures, but the presence and size of its putative somatic cilia pose homology issues. We suggest that these three fossils are likely to be taphonomically and diagenetically distorted and altered acritarchs. These alterations include secondary mineral encrustations on the interiors of vesicles, the crushing, folding and other distortions of the vesicles, the bending and crushing of the acritarch spines, and the preservation of organic material in and outside of the cysts. The earliest known ciliate fossil remains a tintinnid that occurs in the Ordovician of Kazakhstan

Interpreting phylogenetic placements for taxonomic assignment of environmental DNA

Taxonomic assignment of operational taxonomic units (OTUs) is an important bioinformatics step in analyzing environmental sequencing data. Pairwise alignment and phylogenetic-placement methods represent two alternative approaches to taxonomic assignments, but their results can differ. Here we used available colpodean ciliate OTUs from forest soils to compare the taxonomic assignments of VSEARCH (which performs pairwise alignments) and EPA-ng (which performs phylogenetic placements). We showed that when there are differences in taxonomic assignments between pairwise alignments and phylogenetic placements at the subtaxon level, there is a low pairwise similarity of the OTUs to the reference database. We then showcase how the output of EPA-ng can be further evaluated using GAPPA to assess the taxonomic assignments when there exist multiple equally likely placements of an OTU, by taking into account the sum over the likelihood weights of the OTU placements within a subtaxon, and the branch distances between equally likely placement locations. We also inferred the evolutionary and ecological characteristics of the colpodean OTUs using their placements within subtaxa. This study demonstrates how to fully analyze the output of EPA-ng, by using GAPPA in conjunction with knowledge of the taxonomic diversity of the clade of interest

Impact de la recherche d'amorces mutées sur les résultats d'analyses métagénomiques

National audienceEn métagénomique ciblée, une problématique récurrente concerne la quantité de lectures réellement exploitables en sortie d'un séquenceur à haut débit. L'une des étapes influençant cette quantité est la détection dans chaque séquence des amorces utilisées pour amplifier le gène ciblé. Usuellement les séquences n’ayant pas les amorces parfaites sont rejetées. Notre étude interroge l’impact d’accepter également les séquences avec amorces mutées. Il s’agit de répondre aux questions suivantes : Accepter les séquences avec amorces mutées permet-il d'augmenter le nombre de séquences exploitables par échantillon ? Cette augmentation impacte-t-elle les résultats obtenus au terme de l'analyse métagénomique , c’est-à-dire permet-elle d’augmenter le nombre d’OTU (Operationnal taxonomic unit), et donc potentiellement d’espèces, détectés ? L’étude a été menée sur 9 échantillons, dans le cadre d’une étude de la biodiversité des sols tropicaux chez les eucaryotes unicellulaires. Les échantillons ont été séquencés à la fois en Roche/454 et en Illumina MiSeq, afin de pouvoir constater l'impact de la technologie sur les résultats. Le séquençage a ciblé la région V4 de la sous-unité 18S de l'ARN ribosomique. En résultat, on observe que la recherche des amorces V4F et V4R mutées permet d'améliorer de façon non-négligeable la sensibilité de l’analyse métagénomique. Cela augmente le rappel parmi les séquences d'un échantillon et cela permet également la détection de nouveaux OTU (+7 à +10 %, dans notre étude, en fonction de la technologie de séquençage), que ce soit en augmentant l'abondance de séquences détectées en nombre insuffisant pour être validées ou que ce soit en détectant des séquences totalement nouvelles qui n'étaient pas visibles auparavant. Une étude supplémentaire a par ailleurs montré que ces nouveaux OTU, et notamment les totalement nouveaux, étaient aussi crédibles que les OTU avec amorces exactes

Testing ecological theories with sequence similarity networks: marine ciliates exhibit similar geographic dispersal patterns as multicellular organisms

International audienceBackground : High-throughput sequencing technologies are lifting major limitations to molecular-based ecological studies of eukaryotic microbial diversity, but analyses of the resulting millions of short sequences remain a major bottleneck for these approaches. Here, we introduce the analytical and statistical framework of sequence similarity networks, increasingly used in evolutionary studies and graph theory, into the field of ecology to analyze novel pyrosequenced V4 small subunit rDNA (SSU-rDNA) sequence data sets in the context of previous studies, including SSU-rDNA Sanger sequence data from cultured ciliates and from previous environmental diversity inventories.Results : Our broadly applicable protocol quantified the progress in the description of genetic diversity of ciliates by environmental SSU-rDNA surveys, detected a fundamental historical bias in the tendency to recover already known groups in these surveys, and revealed substantial amounts of hidden microbial diversity. Moreover, network measures demonstrated that ciliates are not globally dispersed, but are structured by habitat and geographical location at intermediate geographical scale, as observed for bacteria, plants, and animals.Conclusions : Currently available ‘universal’ primers used for local in-depth sequencing surveys provide little hope to exhaust the significantly higher ciliate (and most likely microbial) diversity than previously thought. Network analyses such as presented in this study offer a promising way to guide the design of novel primers and to further explore this vast and structured microbial diversity

Broad Taxon Sampling of Ciliates Using Mitochondrial Small Subunit Ribosomal DNA

Mitochondrial SSU-rDNA has been used recently to infer phylogenetic relationships among a few ciliates. Here, this locus is compared with nuclear SSU-rDNA for uncovering the deepest nodes in the ciliate tree of life using broad taxon sampling. Nuclear and mitochondrial SSU-rDNA reveal the same relationships for nodes well-supported in previously-published nuclear SSU-rDNA studies, although support for many nodes in the mitochondrial SSU-rDNA tree are low. Mitochondrial SSU-rDNA infers a monophyletic Colpodea with high node support only from Bayesian inference, and in the concatenated tree (nuclear plus mitochondrial SSU-rDNA) monophyly of the Colpodea is supported with moderate to high node support from maximum likelihood and Bayesian inference. In the monophyletic Phyllopharyngea, the Suctoria is inferred to be sister to the Cyrtophora in the mitochondrial, nuclear, and concatenated SSU-rDNA trees with moderate to high node support from maximum likelihood and Bayesian inference. Together these data point to the power of adding mitochondrial SSU-rDNA as a standard locus for ciliate molecular phylogenetic inferences

Swarm v3: towards tera-scale amplicon clustering

Motivation: Previously we presented swarm, an open-source amplicon clustering programme that produces fine-scale molecular operational taxonomic units (OTUs) that are free of arbitrary global clustering thresholds. Here, we present swarm v3 to address issues of contemporary datasets that are growing towards tera-byte sizes. Results: When compared with previous swarm versions, swarm v3 has modernized C++ source code, reduced memory footprint by up to 50%, optimized CPU-usage and multithreading (more than 7 times faster with default parameters), and it has been extensively tested for its robustness and logic

Evaluating Support for the Current Classification of Eukaryotic Diversity

Perspectives on the classification of eukaryotic diversity have changed rapidly in recent years, as the four eukaryotic groups within the five-kingdom classification—plants, animals, fungi, and protists—have been transformed through numerous permutations into the current system of six “supergroups.” The intent of the supergroup classification system is to unite microbial and macroscopic eukaryotes based on phylogenetic inference. This supergroup approach is increasing in popularity in the literature and is appearing in introductory biology textbooks. We evaluate the stability and support for the current six-supergroup classification of eukaryotes based on molecular genealogies. We assess three aspects of each supergroup: (1) the stability of its taxonomy, (2) the support for monophyly (single evolutionary origin) in molecular analyses targeting a supergroup, and (3) the support for monophyly when a supergroup is included as an out-group in phylogenetic studies targeting other taxa. Our analysis demonstrates that supergroup taxonomies are unstable and that support for groups varies tremendously, indicating that the current classification scheme of eukaryotes is likely premature. We highlight several trends contributing to the instability and discuss the requirements for establishing robust clades within the eukaryotic tree of life