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68 • "PROTIST—2016
evolution as an organism gives up autotrophy in favor of moving onto its relatives' couch and raiding their refrigerator. We sequenced draft transcripto-mes for the parasitic red alga Choreocolax polysi-phoniae and its free-living host Vertebrata lanosa. These data, in combination with genomic DNA sequence data for these taxa was compared with other published red algal genomes and transcripto-mes to investigate the early consequences of tran-sitioning from autotrophy to parasitism. Choreocolax polysiphoniae appears to have lost genes involved in plastid maintenance and photosynthetic processes. Investigation ofthese data and their implications for the evolution of parasitism remains ongoing.
MOLECULAR DIVERSITY AND PHYLO-GENY OF OLIGOTRICHIA AND CHOREO-TRICHIA (CILIOPHORA, SPIROTRICHEA) Santoferrara L.F., McManus G.B. Department of Marine Sciences, University of Connecticut
[email protected] We revise and expand the molecular data on the subclasses Oligotrichia and Choreotrichia in terms of both genetic markers and taxonomic sampling. Two approached are used, one based on sequences from morphologically-characterized specimens, and another one including also the unidentified environmental sequences available in NCBI Gen Bank. First, concatenated sequences of ribosomal DNA markers (18S rDNA, 28S rDNA and ITS regions) from seventeen families and four clades of the paraphyletic genus Tintintinnopsis were analyzed, including two families newly studied by single cell sequencing and six genera newly sequenced for at least one marker. Second, all the 18S rDNA sequences available in GenBank for these subclasses were retrieved and curated as part of the EukRef initiative, resulting in almost 3,200 sequences and 1,000 OTUs (operational taxonomic units clustered at 99% similarity). The concatenated dataset allowed confirming or discovering novel phylogenetic relationships within Choreotrichia. The 18S rDNA dataset allowed rough comparisons of molecular diversity between the two subclasses. Because Choreotrichia has been easier to barcode (mainly for the loricate Tintinnida), analyses have not revealed novel diversity, but exposed known and new misidentifications and classification inconsistencies. In contrast, three clades within Oligotrichia included only environmental sequences, thus highlighting a high proportion of partially-characterized or novel diversity in this subclass. These analyses help solving classification conflicts and will provide a reference for studying the diversity and distribution
of these abundant and ecologically relevant ciliates in marine plankton.
TAXONOMIC COMPOSITION OF PROKA-RYOTES-ASSOCIATES OF HALOPHILIC PRO-TISTS
Selivanova Olga1, Gogoleva Natalya2, Khlopko Yuri1, Plotnikov Andrey1
1 - Institute for Cellular and Intracellular Symbiosis UB RAS, Orenburg, Russia
2 - Kazan Institute ofBiochemistry and Biophysics KSC RAS, Kazan, Russia [email protected]
As known, taxonomic structure of communities in hypersaline environments is simple. It includes prokaryotes, photosynthetic and heterotrophic protists, fungi, and crustaceans. Under extreme salinity the community is simplified up to only microorganisms, where phototrophic protists are the main producers of organic matter, and heterotrophic protists are the only consumers. Symbiotic associations with halophilic phototrophic protists are well studied for green alga Dunaliella and its prokaryotic associates, while heterotrophic protists and their role in forming of halophilic microbial community are studied poorly. Aim of the study was to estimate taxonomic composition of prokaryotes — associates of autotrophic and heterotrophic protists isolated from salt water bodies in Salt-Iletsk and Elton regions. The method of 16S metagenomic sequencing was used. 111 OTUs were found in association with heterotrophic flagellate Pleurostomum salinum. The genera of archaea Natronomonas sp., Haloarcula sp., Haloplanus sp., Halorubrum sp. and bacteria Halovibrio sp., uncultured bacterium (Bacteroide-tes) were predominant. Heterotrophic flagellate Tulamoeba bucina had very similar taxonomic composition of prokaryotes. There were 129 OTUs including uncultured bacterium (Bacteroidetes), Natronomonas sp., Halovibrio sp., Haloferax sp., Halorubrum sp. as predominant genera. There were 32 and 41 OTUs in associations with Pharyngomonas sp. and Heterolobosea aff. Euplaesiobystra respectively. Bacteria Idiomarina sp., Halomonas sp., Gracilimonas sp. were predominant in both cultures. Phototrophic flagellate Dunaliella parva had 19 OTUs of prokaryotes — associates. Bacteria Halovibrio sp. and archaea Halorubrum sp. were the most abundant. A crucial factor influencing the taxonomic composition of prokaryotes in the associations with halophilic protists was level of medium mineralization. Taxonomic diversity was higher in associations with heterotrophic protists than in associations with autotrophs. The highest
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species richness ofprokaryotes — associates has been registered in the culture Tulamoeba bucina. The research was performed in the Center of Shared Scientific Equipment «Persistence of microorganisms» of ICIS UB RAS and was supported by RFBR (16-44-560316, 14-04-01796).
CHARACTERIZATION OF MONOCLONAL ANTIBODIES FOR CATHEPSIN B AND CA-THEPSIN B-LIKE PROTEINS OF NAEGLERIA FOWLERI
Seong G.S.12, Sohn H.J.1-2, Kang H.K.12, Shin H.J.12
1 - DepartmentofMicrobiology, Ajou University School of Medicine
2 - Department of Biomedical Science, Graduate School of Ajou University, Suwon 443-721, Republic of Korea
Naegleria fowleri causes a fatal primary amoebic meningoencephalitis (PAM) in human and experimental animals. Cathepsin B (NfCPB) and cathepsin B-Like (NfCPBL) gene in N. fowleri, are consists of 1,038bp of DNA (345 amino acid) and 939bp of DNA (313 amino acid), and molecular weights of recombinant proteins are 38.4 and 34 kDa, respectively. In the previous study, rNfCPB and rNfCPB-L refolding protein may play important roles in host tissue invasion, immune evasion and nutrient uptake. In this study, we produced anti-NfCPB and anti-NfCPBL monoclonal antibody (rNfCPB-McAb and rNfCPBL-McAb) using the cell fusion technique and observed the immunological characteristics. Seven hybridoma cells secreting rNfCPB-McAb (2D6, 2D11, 2A7, 2B7, 2E2, 2E9, 2C9) and three hybridoma cells secreting rNfCPB-McAb (1E5, 1C8, 1D9) were produced. Among them, 2C9 (a cell line producing rNfCPB-McAb) and 1C8 (a cell line producing rNfCPBL-McAb) which showed high antibody titre, respectively, were selected. 2C9 monoclonal antibody was reacted with N. fowleri whole lysate or rNfCPB fusion protein by western blotting, as which showed about 28 kDa and 38.4 kDa ofband pattern, respectively. 1C9 monoclonal antibody was reacted with N. fowleri whole lysate or rNfCPBL fusion protein by western blotting, as which showed 24 kDa and 34 kDa ofband pattern. 2C9 and 1C8 monoclonal antibodies were not reacted with another amoebic lysates such as N. gruberi, Acanthamoeba castellanii, A. polyphagia in western blot analysis. In the results of the immunocytochemistry analysis, NfCPB and NfCPB-L protein were mainly detected in cytoplasm and cell membrane, especially pseudopodia, of N. fowleri trophozoites under a confocal microscope.
These results suggested that monoclonal antibodies against rNfCPB and rNfCPBL may be useful for the further immunological study.
BENTHIC PROTISTS (CILIOPHORA, GRO-MIIDA, FORAMINIFERA) IN THE BLACK SEA: THEIR CONTRIBUTION TO THE BOTTOM COMMUNITIES Sergeeva N.G.
Institute of Marine Biological Research of Russian Academy of Sciences, 2, Nakhimov ave, Sevastopol, Russia
Ciliophora, Gromiida and Foraminifera (hard- shell and soft-shell forms) are the significant components of the benthic communities of the Black Sea. These protozoans are of great role in the transfer of bacterial and algal production to the next trophic levels. They themselves are consumers of bottom bacteria, microalgae, pollen terrestrial vegetation and marine filamentous fungi; as predators, they might prey upon unicellular organisms. In addition, the representatives of above listed groups are an important food source for many bottom invertebrates in different habitats. The changes in taxa composition and abundance, and also the proportions of these protozoan representatives in the benthic communities along deep-water oxic/ anoxic interface (75-300m) in the Istanbul Strait's (Bosporus) outlet area of the Black Sea and NW shelf of Crimea Peninsula were studied. In the oxic zone of the Black Sea, benthic Protozoa and Metazoa were studied in area near the Kerch Strait's on NE shelf ofCrimean Peninsula. Our data allowed concluding about tolerance ofmany benthic protozoans to the hypoxic / anoxic conditions and sulfidic pollution of bottom sediments of the Black Sea and of their significant share in the deep-water benthic communities. Benthic Ciliophora, Gromiida and Foraminifera are numerous, specific and diverse in these conditions. The contribution of these protozoans in the meiobenthos communities had high values and extremely prone to variability. The spatial-bathymetrical distribution of these protists along with benthic multicellular organisms had uneven character in the Black Sea.
CHARACTERIZATION OF " CANDIDATUS GORTZIA SHAHRAZADIS", A NOVEL ENDO-SYMBIONT OF PARAMECIUM MULTIMI-CRONUCLEATUM FROM INDIA Serra V.1, Fokin S.I.1'2, Castelli M.1, Basuri C.K.3, Nitla V.M.3, Verni F.1, Sandeep B.V.34, Kalavathi C.3, Petroni G.1
1 - Department of Biology, University of Pisa, Pisa,
