Kirill V. Mikhailov

The origin of Metazoa: a transition from temporal to spatial cell differentiation†

For over a century, Haeckels Gastraea theory remained a dominant theory to explain the origin of multicellular animals. According to this theory, the animal ancestor was a blastula‐like colony of uniform cells that gradually evolved cell differentiation. Today, however, genes that typically control metazoan development, cell differentiation, cell‐to‐cell adhesion, and cell‐to‐matrix adhesion are found in various unicellular relatives of the Metazoa, which suggests the origin of the genetic programs of cell differentiation and adhesion in the root of the Opisthokonta. Multicellular stages occurring in the complex life cycles of opisthokont protists (mesomycetozoeans and choanoflagellates) never resemble a blastula. Here, we discuss a more realistic scenario of transition to multicellularity through integration of pre‐existing transient cell types into the body of an early metazoon, which possessed a complex life cycle with a differentiated sedentary filter‐feeding trophic stage and a non‐feeding blastula‐like larva, the synzoospore. Choanoflagellates are considered as forms with secondarily simplified life cycles.

Obligately phagotrophic aphelids turned out to branch with the earliest-diverging fungi.

Reconstructing the early evolution of fungi and metazoans, two of the kingdoms of multicellular eukaryotes thriving on earth, is a challenging task for biologists. Among extant organisms having characters intermediate between fungi and hypothetical protistan ancestors, from which both fungi and metazoans are believed to have evolved, aphelids are unfairly neglected. The phylogenetic position of these microalgal endoparasites remained uncertain, since no nucleotide sequence data have been reported to date. Aphelids resemble some primitive zoosporic fungi in life cycle, but, unlike fungi, they live by phagotrophy. Here we present a phylogeny, in which a cultured aphelid species, Amoeboaphelidium protococcarum, forms a monophyletic group with Rozella and microsporidia as a sister group to Fungi. We also report a non-canonical nuclear genetic code in A. protococcarum.

Description of Colponema vietnamica sp.n. and Acavomonas peruviana n. gen. n. sp., two new alveolate phyla (Colponemidia nom. nov. and Acavomonidia nom. nov.) and their contributions to reconstructing the ancestral state of alveolates and eukaryotes.

The evolutionary and ecological importance of predatory flagellates are too often overlooked. This is not only a gap in our understanding of microbial diversity, but also impacts how we interpret their better-studied relatives. A prime example of these problems is found in the alveolates. All well-studied species belong to three large clades (apicomplexans, dinoflagellates, and ciliates), but the predatory colponemid flagellates are also alveolates that are rare in nature and seldom cultured, but potentially important to our understanding of alveolate evolution. Recently we reported the first cultivation and molecular analysis of several colponemid-like organisms representing two novel clades in molecular trees. Here we provide ultrastructural analysis and formal species descriptions for both new species, Colponema vietnamica n. sp. and Acavomonas peruviana n. gen. n. sp. Morphological and feeding characteristics concur with molecular data that both species are distinct members of alveolates, with Acavomonas lacking the longitudinal phagocytotic groove, a defining feature of Colponema. Based on ultrastructure and molecular phylogenies, which both provide concrete rationale for a taxonomic reclassification of Alveolata, we establish the new phyla Colponemidia nom. nov. for the genus Colponema and its close relatives, and Acavomonidia nom. nov. for the genus Acavomonas and its close relatives. The morphological data presented here suggests that colponemids are central to our understanding of early alveolate evolution, and suggest they also retain features of the common ancestor of all eukaryotes.

On the phylogenetic position of insects in the Pancrustacea clade

The current views on the phylogeny of arthropods are at odds with the traditional system, which recognizes four independent arthropod classes: Chelicerata, Crustacea, Myriapoda, and Insecta. There is compelling evidence that insects comprise a monophyletic lineage with Crustacea within a larger clade named Pancrustacea, or Tetraconata. However, which crustacean group is the closest living relative of insects is still an open question. In recent phylogenetic trees constructed on the basis of large gene sequence data insects are placed together with primitive crustaceans, the Branchiopoda. This topology is often suspected to be a result of the long branch attraction artifact. We analyzed concatenated data on 77 ribosomal proteins, elongation factor 1A (EF1A), initiation factor 5A (eIF5A), and several other nuclear and mitochondrial proteins. Analyses of nuclear genes confirm the monophyly of Hexapoda, the clade uniting entognath and ectognath insects. The hypothesis of the monophyly of Hexapoda and Branchiopoda is supported in the majority of analyses. The Maxillopoda, another clade of Entomostraca, occupies a sister position to the Hexapoda + Branchiopoda group. Higher crustaceans, the Malacostraca, in most analyses appear a more basal lineage within the Pancrustacea. We report molecular synapomorphies in low homoplastic regions, which support the clade Hexapoda + Branchiopoda + Maxillopoda and the monophyletic Malacostraca including Phyllocarida. Thus, the common origin of Hexapoda and Branchiopoda and their position within Entomostraca are suggested to represent bona fide phylogenetic relationships rather than computational artifacts.

Genomic survey of a hyperparasitic microsporidian Amphiamblys sp. (Metchnikovellidae)

Metchnikovellidae are a group of unusual microsporidians that lack some of the defining ultrastructural features characteristic of derived Microsporidia and are thought to be one of their earliest-branching lineages. The basal position of metchnikovellids was never confirmed by molecular phylogeny in published research, and thus far no genomic data for this group were available. In this work, we obtain a partial genome of metchnikovellid Amphiamblys sp. using multiple displacement amplification, next-generation sequencing, and metagenomic binning approaches. The partial genome, which we estimate to be close to 90% complete, displays genome compaction on par with gene-dense microsporidian genomes, but contains an unusual repertoire of unique repeat elements. Phylogenetic analyses of multigene datasets place Amphiamblys sp. as the first branch of the microsporidian lineage following the divergence of a mitochondriate microsporidian Mitosporidium. We find evidence for a mitochondrial remnant presumably functionally equivalent to a mitosome in Amphiamblys sp. and the common enzymatic complement for microsporidian anaerobic metabolism. Comparative genomic analyses identify the conservation of components for clathrin vesicle formation as one of the key features distinguishing the metchnikovellid from its highly derived relatives. The presented data confirm the notion of Metchnikovellidae as a less derived microsporidian group, and provide an additional stepping stone for reconstruction of an evolutionary transition from the early diverging parasitic fungi to derived Microsporidia.

Heterokont Predator Develorapax marinus gen. et sp. nov. - A Model of the Ochrophyte Ancestor.

Heterotrophic lineages of Heterokonta (or stramenopiles), in contrast to a single monophyletic group of autotrophs, Ochrophyta, form several clades that independently branch off the heterokont stem lineage. The nearest neighbors of Ochrophyta in the phylogenetic tree appear to be almost exclusively bacterivorous, whereas the hypothesis of plastid acquisition by the ancestors of the ochrophyte lineage suggests an ability to engulf eukaryotic alga. In line with this hypothesis, the heterotrophic predator at the base of the ochrophyte lineage may be regarded as a model for the ochrophyte ancestor. Here, we present a new genus and species of marine free-living heterotrophic heterokont Develorapax marinus, which falls into an isolated heterokont cluster, along with the marine flagellate Developayella elegans, and is able to engulf eukaryotic cells. Together with environmental sequences D. marinus and D. elegans form a class-level clade Developea nom. nov. represented by species adapted to different environmental conditions and with a wide geographical distribution. The position of Developea among Heterokonta in large-scale phylogenetic tree is discussed. We propose that members of the Developea clade represent a model for transition from bacterivory to a predatory feeding mode by selection for larger prey. Presumably, such transition in the grazing strategy is possible in the presence of bacterial biofilms or aggregates expected in eutrophic environment, and has likely occurred in the ochrophyte ancestor.

Do we need many genes for phylogenetic inference

Fifty-six nuclear protein coding genes from Taxonomically Broad EST Database and other databases were selected for phylogenomic-based examination of alternative phylogenetic hypotheses concerning intergroup relationship between multicellular animals (Metazoa) and other representatives of Opisthokonta. The results of this work support sister group relationship between Metazoa and Choanoflagellata. Both of these groups form the taxon Holozoa along with the monophyletic Ichthyosporea or Mesomycetozoea (a group that includes Amoebidium parasiticum, Sphaeroforma arctica, and Capsaspora owczarzaki). These phylogenetic hypotheses receive high statistical support both when utilizing whole alignment and when only 5000 randomly selected alignment positions are used. The presented results suggest subdivision of Fungi into Eumycota and lower fungi, Chytridiomycota. The latter form a monophyletic group that comprises Chytridiales + Spizellomycetales + Blastocladiales (Batrachochytrium, Spizellomyces, Allomyces, Blastocladiella), contrary to the earlier reports based on the analysis of 18S rRNA and a limited set of protein coding genes. The phylogenetic distribution of genes coding for a ubiquitin-fused ribosomal protein S30 implies at least three independent cases of gene fusion: in the ancestors of Holozoa, in heterotrophic Heterokonta (Oomycetes and Blastocystis), and in the ancestors of Cryptophyta and Glaucophyta. Ubiquitin-like sequences fused with ribosomal protein S30 outside of Holozoa are not FUBI orthologs. Two independent events of FUBI replacement by the ubiquitin sequence were detected in the lineage of C. owczarzaki and in the monophyletic group of nematode worms Tylenchomorpha + Cephalobidae. Bursaphelenchus xylophilus (Aphelenchoidoidea) retains a state typical of the rest of the Metazoa. The data emphasize the fact that the reliability of phylogenetic reconstructions depends on the number of analyzed genes to a lesser extent than on our ability to recognize reconstruction artifacts.

A new view on the morphology and phylogeny of eugregarines suggested by the evidence from the gregarine Ancora sagittata (Leuckart, 1860) Labbé, 1899 (Apicomplexa: Eugregarinida)

Background Gregarines are a group of early branching Apicomplexa parasitizing invertebrate animals. Despite their wide distribution and relevance to the understanding the phylogenesis of apicomplexans, gregarines remain understudied: light microscopy data are insufficient for classification, and electron microscopy and molecular data are fragmentary and overlap only partially. Methods Scanning and transmission electron microscopy, PCR, DNA cloning and sequencing (Sanger and NGS), molecular phylogenetic analyses using ribosomal RNA genes (18S (SSU), 5.8S, and 28S (LSU) ribosomal DNAs (rDNAs)). Results and Discussion We present the results of an ultrastructural and molecular phylogenetic study on the marine gregarine Ancora sagittata from the polychaete Capitella capitata followed by evolutionary and taxonomic synthesis of the morphological and molecular phylogenetic evidence on eugregarines. The ultrastructure of Ancora sagittata generally corresponds to that of other eugregarines, but reveals some differences in epicytic folds (crests) and attachment apparatus to gregarines in the family Lecudinidae, where Ancora sagittata has been classified. Molecular phylogenetic trees based on SSU (18S) rDNA reveal several robust clades (superfamilies) of eugregarines, including Ancoroidea superfam. nov., which comprises two families (Ancoridae fam. nov. and Polyplicariidae) and branches separately from the Lecudinidae; thus, all representatives of Ancoroidea are here officially removed from the Lecudinidae. Analysis of sequence data also points to possible cryptic species within Ancora sagittata and the inclusion of numerous environmental sequences from anoxic habitats within the Ancoroidea. LSU (28S) rDNA phylogenies, unlike the analysis of SSU rDNA alone, recover a well-supported monophyly of the gregarines involved (eugregarines), although this conclusion is currently limited by sparse taxon sampling and the presence of fast-evolving sequences in some species. Comparative morphological analyses of gregarine teguments and attachment organelles lead us to revise their terminology. The terms “longitudinal folds” and “mucron” are restricted to archigregarines, whereas the terms “epicystic crests” and “epimerite” are proposed to describe the candidate synapomorphies of eugregarines, which, consequently, are considered as a monophyletic group. Abolishing the suborders Aseptata and Septata, incorporating neogregarines into the Eugregarinida, and treating the major molecular phylogenetic lineages of eugregarines as superfamilies appear as the best way of reconciling recent morphological and molecular evidence. Accordingly, the diagnosis of the order Eugregarinida Léger, 1900 is updated.

Evolution of mitochondrial genomes in Baikalian amphipods

BackgroundAmphipods (Crustacea) of Lake Baikal are a very numerous and diverse group of invertebrates generally believed to have originated by adaptive radiation. The evolutionary history and phylogenetic relationships in Baikalian amphipods still remain poorly understood. Sequencing of mitochondrial genomes is a relatively feasible way for obtaining a set of gene sequences suitable for robust phylogenetic inferences. The architecture of mitochondrial genomes also may provide additional information on the mechanisms of evolution of amphipods in Lake Baikal.ResultsThree complete and four nearly complete mitochondrial genomes of Baikalian amphipods were obtained by high-throughput sequencing using the Illumina platform. A phylogenetic inference based on the nucleotide sequences of all mitochondrial protein coding genes revealed the Baikalian species to be a monophyletic group relative to the nearest non-Baikalian species with a completely sequenced mitochondrial genome - Gammarus duebeni. The phylogeny of Baikalian amphipods also suggests that the shallow-water species Eulimnogammarus has likely evolved from a deep-water ancestor, however many other species have to be added to the analysis to test this hypothesis.The gene order in all mitochondrial genomes of studied Baikalian amphipods differs from the pancrustacean ground pattern. Mitochondrial genomes of four species possess 23 tRNA genes, and in three genomes the extra tRNA gene copies have likely undergone remolding. Widely varying lengths of putative control regions and other intergenic spacers are typical for the mitochondrial genomes of Baikalian amphipods.ConclusionsThe mitochondrial genomes of Baikalian amphipods display varying organization suggesting an intense rearrangement process during their evolution. Comparison of complete mitochondrial genomes is a potent approach for studying the amphipod evolution in Lake Baikal.

The complete mitochondrial genome of a deep-water Baikalian amphipoda Brachyuropus grewingkii (Dybowsky, 1874)

Abstract In this study, we present a complete mitochondrial genome of a deep-water amphipoda Brachyuropus grewingkii (Dybowsky, 1874) from Lake Baikal. A circular mitochondrial DNA has 17,118 bp in length and contains 13 protein-coding genes, two ribosomal RNA genes, 22 transfer RNA genes, a putative control region, and five intergenic spacers. An extended control region and altered positions of some tRNA genes distinguish mitochondrial genome of B. grewingkii from the mitochondrial genomes described for other Baikalian amphipoda species.