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60 • "PROTIST—2016
functionalized with several types of ligands, which will be the core of the immunochromatographic RDT with potential for point-of-care diagnostics. Acknowledgments: Partially supported by Gilead GENESE-PGG/001/2014.
DIVERSITY OF PROTISTS IN SALINE AND BRACKISH CONTINENTAL WATER BODIES REVEALED BY HIGH-THROUGHPUT SEQUENCING
Plotnikov Andrey12, Gerasimova Elena1, Poshvina Daria1, Gogoleva Natalya3, Khlopko Yuri1
1 - Institute for Cellular and Intracellular Symbiosis UB RAS, Orenburg, Russia
2 - Orenburg State Medical University, Orenburg, Russia
3 - Kazan Institute ofBiochemistry and Biophysics KSC RAS, Kazan, Russia
protoz.@mail.ru
Modern methods of high-throughput sequencing (NGS) are widely used for characterization of protists biodiversity in fresh and marine waters and often result in new data changing our knowledge about natural microbial communities. At present only protistian communities from marine biotops have been studied with NGS, whereas the data on continental saline water bodies are rare (Heidelberg et al., 2013; Triado-Margarit and Casamayor, 2013). The aim ofthis investigation was characterization of structure and biodiversity ofprotistian communities in saline and brackish water bodies of the South Urals (Russia) with 18S metagenomic sequencing. For this purpose water samples from saline and brackish lakes and a brackish river were filtered through membranes with diameter ofpores 0.45 ^m. Total DNA was isolated from the filters and DNAlibraries were made by PCR with universal primers for V4 region of the gene 18S. High-throughput sequencing was conducted with MiSeq (Illumina). The obtained reads were treated with complex of bioinformatic tools. In the report the first data on the biodiversity of eukaryotes in the deeply continental saline and brackish water bodies of the South Urals (Russia) will be presented.
The research was performed in the Center of Shared Scientific Equipment «Persistence of microorganisms» of ICIS UB RAS and was supported by RFBR (16-44-560234, 15-29-02749, 15-2902518).
AN EARLY-BRANCHING CYANOBACTE-RIUM AT THE ORIGIN OF PRIMARY PHO-TOSYNTHETIC EUKARYOTES Ponce-Toledo R.I.1, Deschamps P.1, Lopez-Garcia P.1, Zivanovic Y.2, Benzerara K.3, Moreira D.1
1 - Unité d'Ecologie, Systématique et Evolution, Centre National de la Recherche Scientifique UMR 8079, Université Paris-Sud, 91405 Orsay, France
2 - Institut de Génétique et Microbiologie, Centre National de la Recherche Scientifique UMR 8621, Université Paris-Sud, 91405 Orsay, France
3 - IMPMC, Sorbonne Universités, Paris 75005, France
rafael.ponce@u-psud.fr
Primary plastid-bearing eukaryotes evolved by the endosymbiosis of a cyanobacterium within a heterotrophic host. This gave rise to the supergroup called Archaeplastida, comprising Viridiplantae (green algae and land plants), Rhodophyta (red algae) and Glaucophyta. Although the monophyly of primary plastids has been extensively recovered, the present-day closest cyanobacterial lineage to the chloroplast ancestor is still debated. We performed phylogenetic analyses using two concatenated datasets containing 97 plastid-encoded proteins and the plastid 16S+23S rRNA cluster, and found in both phylogenetic reconstructions that the ancestor of primary plastids was an early-branching cyanobacterium related to Gloeomargarita lithopho-ra, the first cultured member of a recently discovered freshwater cyanobacterial lineage widely present in stromatolites and thermophilic microbial mats. This discovery has implications for the environmental conditions in which the endosymbiosis took place.
THE SPECIAL CASE OF HOLOSPORA CARYO-PHILA, BACTERIAL SYMBIONT OF CILIATES PARAMECIUM
Potekhin A.1, Schrallhammer M.2, Schweikert M.3, Nekrasova I.1, Lebedeva N.4, Kaltz O.5, Petroni G.6
1 - Faculty of Biology, St Petersburg State University, Saint Petersburg, Russia
2 - Institute ofBiology, University ofFreiburg, Freiburg, Germany
3 - Institute of Biology, University of Stuttgart, Stuttgart, Germany
4 - Centre of Core Facilities "Culture Collections of Microorganisms", St Petersburg State University, Saint Petersburg, Russia
5 - Institut of Evolutionary Science, Montpellier University 2, Montpellier, France
6 - Department of Biology, University of Pisa, Pisa, Italy
alexey.potekhin@spbu.ru
Infectious bacterium Holospora caryophila, described as symbiont of the macronucleus of Paramecium biaurelia, is an unconventional Holospora. While other Holospora species are highly selective for the host, H. caryophila has been isolated from nature in several species of the P. aurelia complex, and in
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P. caudatum. Infection by H. caryophila sometimes kills paramecia, but stable association may last for many years. These premises led to detailed investigation of 6 isolates of H. caryophila, 4 ofthem inhabiting the species of the P. aurelia complex, and 2 found in P. caudatum. Screening of more than 70 potential hosts — strains belonging to 10 morphological species of Paramecium — showed that H. caryophila can infect majority of species of the P. aurelia complex, P. jenningsi, P. caudatum, P. multimicronucleatum, and P. putrinum. Symbiosis could be formed only in certain combinations ofthe host and the symbiont, though some strains appeared to be universal recipients for all H. caryophila isolates studied. However, most of the checked strains were never infected. P. caudatum strains often died during infection by H. caryophila isolated from P. aurelia strains. Thus, the chance that symbiont can not develop in host or kills it is higher than possibility of successful infection, explaining why ciliates harboring symbiotic bacteria are rather rare in nature. Molecular phylogenetic analysis of 16S rDNA sequences proved that phylogenetic relationships of H. caryophila with other Holospora species are quite distant. Physiological and phylogenetic features support transfer of H. caryophila to the new genus Preeria.
Supported by RFBR 16-04-01195.
CHASING ION CHANNELS OF DINOFLA-GELLATES
Pozdnyakov I., Matantseva O., Skarlato S. Institute ofCytology RAS, St. Petersburg, Russia pozdnyakov@cytspb.rssi.ru
Ion channels are transmembrane protein complexes permeable for ions and playing a crucial role in cell physiology. Ion channels of animals, plants and fungi have been intensively studied for many decades; however, noticeably less information is available concerning ion channels of other eukaryotes. This lack of knowledge hampers our understanding of both evolution of ion transport and physiology of protists. Dinoflagellates are the group of aquatic unicellular eukaryotes of high ecological relevance, but rather little is known about their physiology. At the same time, dinoflagellates are phylogenetically distant from animals, plants and fungi and thus are attractive objects to study evolution of ion channels. However, investigation of dinoflagellate ion channels is complicated by the lack of sufficient genomic data and obstacles in applying electrophysiological techniques to dinoflagellates due to their complex cell coverings. We analyzed publicly available transcriptomes of ten dinoflagellate species and found 12 ion channel
families, including four-domain voltage-gated ion channels (FDVGIC) that played a crucial role in evolution of exciting membranes of eukaryotes and nervous system in metazoans. We revealed a high degree of phylogenetic, structural and functional diversity in FDVGIC of dinoflagellates. We developed a new method to produce spheroplasts of armored dinoflagellates and for the first time obtained single-channel recordings oftheir ion channels. Our method allowed us to detect considerable diversity of cation channels in Prorocentrum minimum at the electrophysiological level: potassium-selective channels, inwardly rectifying cation channels, "fast" and "slow" cation channels, and nonselective cation channels. Funded by the Russian Science Foundation, project 16-14-10116.
PHYLOGENY OF PROTISTAN FOUR-DOMAIN VOLTAGE-GATED ION CHANNELS Pozdnyakov I., Skarlato S. Institute ofCytology RAS, St. Petersburg, Russia pozdnyakov@cytspb.rssi.ru
Four-domain voltage-gated ion channels (FDV GIC) drive the initial phase of the action potential propagation in metazoans and many protists. Therefore, these channels are considered as major players in evolution of eukaryotic excitability and metazoan nervous system. In addition to cellular excitability, they are important for cellular motility, intracellular signaling and regulation of rhythmical activity. There are five well studied subfamilies of FDVGIC, and all of them are associated with the opisthokont lineage: voltage-gated and voltage-insensitive sodium channels (Nav and NALCN, respectively), as well as voltage-gated calcium channels (LVA-Cav and HVA-Cav) and voltage-insensitive calcium channels of fungi (Cch). Using publicly available genomic, transcriptomic and protein databases and blast search, we identified 277 members of FDVGIC family from different eukaryotic groups to reconstruct phylogeny of this ion channel family employing the maximal likelihood (ML) method and Bayesian analysis (BA). In this work, we demonstrated that most ofthe considered protist groups have their own subfamilies of FDVGIC that do not form clades with any known subfamily ofFDVGIC (i.e. Nav, NALCN, LVA-Cav, HVA-Cav, and Cch). Moreover, both ML and BA approaches showed that similar to metazoans some protist groups, such as alveolates and stramenopiles, possess high phylogenetic diversity of FDVGIC. Although obtained phylogenies are not fully resolved due to the limited data on ion channel sequences, the present study advances our understanding of the diversity and evolution of FDVGIC family. Funded
