The small subunit ribosomal RNA (SSU rRNA) gene is a widely used molecular marker to study the diversity of life. Sequencing of SSU rRNA gene amplicons has become a standard approach for the investigation of the ecology and diversity of microbes. However, a well-curated database is necessary for correct classification of these data. While available for many groups of Bacteria and Archaea, such reference databases are absent for most eukaryotes. The primary goal of the EukRef project (eukref.org) is to close this gap and generate well-curated reference databases for major groups of eukaryotes, especially protists. Here we present a set of EukRef-curated databases for the excavate protists-a large assemblage that includes numerous taxa with divergent SSU rRNA gene sequences, which are prone to misclassification. We identified 6121 sequences, 625 of which were obtained from cultures, 3053 from cell isolations or enrichments and 2419 from environmental samples. We have corrected the classification for the majority of these curated sequences. The resulting publicly available databases will provide phylogenetically based standards for the improved identification of excavates in ecological and microbiome studies, as well as resources to classify new discoveries in excavate diversity.

Biology Centre Institute of Parasitology Czech Academy of Sciences 370 05 České Budeějovice Czech Republic

Department of Biology and Centre of Comparative Genomics and Evolutionary Bioinformatics Dalhousie University Halifax NS B3H 4R2 Canada

Department of Biology Center for Genomics and Systems Biology New York University New York NY 10003 USA

Department of Biology University of Western Ontario London ON N6A 5B7 Canada

Department of Parasitology BIOCEV Faculty of Science Charles University 128 43 Vestec Czech Republic

Department of Zoology Charles University 128 00 Prague Czech Republic

Faculty of Science University of South Bohemia 370 05 České Budeějovice Czech Republic

Institute of Evolutionary Biology Faculty of Biology Biological and Chemical Research Centre University of Warsaw 02 089 Warsaw Poland

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de Vargas C., Audic S., Henry N., et al. (2015) Eukaryotic plankton diversity in the sunlit ocean. Science, 348, 1261605. PubMed

Del Campo J., Kolisko M., Boscaro V., et al. (2018) EukRef: phylogenetic curation of ribosomal RNA to enhance understanding of eukaryotic diversity and distribution. PLoS Biol., 16, 1–14. PubMed PMC

Simpson A.G.B. (2003) Cytoskeletal organization, phylogenetic affinities and systematics in the contentious taxon Excavata (Eukaryota). Int. J. Syst. Evol. Microbiol., 53, 1759–1777. PubMed

Hampl V., Hug L., Leigh J.W.  et al. (2009) Phylogenomic analyses support the monophyly of Excavata and resolve relationships among eukaryotic “supergroups”. Proc. Nat. Acad. Sci. USA, 106, 3859–3864. PubMed PMC

Adl S.M., Bass D., Lane C.E.  et al. (2019) Revisions to the classification, nomenclature, and diversity of eukaryotes. J. Eukaryot. Microbiol., 66, 4–119. PubMed PMC

Heiss A.A., Kolisko M., Ekelund F.  et al. (2018) Combined morphological and phylogenomic re-examination of malawimonads, a critical taxon for inferring the evolutionary history of eukaryotes. R. Soc. Open Sci., 5, 171707. PubMed PMC

Cavalier-Smith T. (2002) The phagotrophic origin of eukaryotes and phylogenetic classification of protozoa. Int. J. Syst. Evol. Microbiol., 52, 297–354. PubMed

Brown M.W., Heiss A.A., Kamikawa R.  et al. (2018) Phylogenomics places orphan protistan lineages in a novel eukaryotic super-group. Genome Biol. Evol., 10, 427–433. PubMed PMC

Adl S.M., Leander B.S., Simpson A.G.B.  et al. (2007) Diversity, nomenclature, and taxonomy of protists. Syst. Biol., 56, 684–689. PubMed

Kolisko M., Čepička I., Hampl V.  et al. (2008) Molecular phylogeny of diplomonads and enteromonads based on SSU rRNA, alpha-tubulin and HSP90 genes: implications for the evolutionary history of the double karyomastigont of diplomonads. BMC Evol. Biol., 8, 205. PubMed PMC

Kolisko M., Silberman J.D., Čepička I.  et al. (2010) A wide diversity of previously undetected free-living relatives of diplomonads isolated from marine/saline habitats. Environ. Microbiol., 12, 2700–2710. PubMed

Pánek T., Simpson A.G., Brown M.W.  et al. (2017) Heterolobosea In: Archibald JM, Simpson AGS, Slamovitz CH  et al. (ed). Handbook of the Protists. Springer, Cham, pp. 1–42.

Pánek T., Táborský P., Pachiadaki M.G.  et al. (2015) Combined culture-based and culture-independent approaches provide insights into diversity of jakobids, an extremely plesiomorphic eukaryotic lineage. Front. Microbiol., 6, 1288. PubMed PMC

Cavalier-Smith T. (2016) Higher classification and phylogeny of Euglenozoa. Eur. J. Protistol., 56, 250–276. PubMed

Maritz J.M., Land K.M., Carlton J.M.  et al. (2014) What is the importance of zoonotic trichomonads for human health?  Trends. Parasitol., 30, 333–341. PubMed PMC

Brune A. (2014) Symbiotic digestion of lignocellulose in termite guts. Nat. Rev. Microbiol., 12, 168–180. PubMed

Flegontova O., Flegontov P., Malviya S.  et al. (2016) Extreme diversity of diplonemid eukaryotes in the ocean. Curr. Biol., 26, 3060–3065. PubMed

Gawryluk R.M.R., Del Campo J., Okamoto N.  et al. (2016) Morphological identification and single-cell genomics of marine diplonemids. Curr. Biol., 26, 3053–3059. PubMed

Park J.S. and Simpson A.G.B. (2015) Diversity of heterotrophic protists from extremely hypersaline habitats. Protist, 166, 422–437. PubMed

Guillou L., Bachar D., Audic S.  et al. (2013) The protist ribosomal reference database (PR2): a catalog of unicellular eukaryote small sub-unit rRNA sequences with curated taxonomy. Nucleic Acids Res., 41, D597–D604. PubMed PMC

Quast C., Pruesse E., Yilmaz P.  et al. (2013) The SILVA ribosomal RNA gene database project: improved data processing and web-based tools. Nucleic Acids Res., 41, D590–D596. PubMed PMC

Moriya S., Dacks J.B., Takagi A.  et al. (2003) Molecular phylogeny of three oxymonad genera: pyrsonympha, dinenympha and oxymonas. J. Euk. Microbiol., 50, 190–197. PubMed

Heiss A.A. and Keeling P.J. (2006) The phylogenetic position of the oxymonad Saccinobaculus based on SSU rRNA. Protist, 157, 335–344. PubMed

Zhang Q., Táborský P., Silberman J.D.  et al. (2015) Marine isolates of Trimastix marina form a plesiomorphic deep-branching lineage within Preaxostyla, separate from other known trimastigids (Paratrimastix n. gen.). Protist, 166, 468–491. PubMed

Keeling P.J. and Leander B.S. (2003) Characterisation of a non-canonical genetic code in the oxymonad Streblomastix strix. J. Mol. Biol., 326, 1337–1349. PubMed

Silberman J.D., Simpson A.G.B., Kulda J.  et al. (2002) Retortamonad flagellates are closely related to diplomonads—Implications for the history of mitochondrial function in eukaryote evolution. Mol. Biol. Evol., 19, 777–786. PubMed

Simpson A.G.B., Roger A.J., Silberman J.D.  et al. (2002b) Evolutionary history of “early-diverging” eukaryotes: the excavate taxon Carpediemonas is a close relative of Giardia. Mol. Biol. Evol., 19, 1782–1791. PubMed

Čepička I., Kostka M., Uzlíková M.  et al. (2008) Non-monophyly of Retortamonadida and high genetic diversity of the genus Chilomastix suggested by analysis of SSU rDNA. Mol. Phylogenet. Evol., 48, 770–775. PubMed

Leger M.M., Kolisko M., Kamikawa R.  et al. (2017) Organelles that illuminate the origins of Trichomonas hydrogenosomes and Giardia mitosomes. Nat. Ecol. Evol., 1, 0092. PubMed PMC

Čepička I., Dolan M. and Gile G. (2017) Parabasalia In: Archibald JM, Simpson AGS, Slamovitz CH  et al. (eds). Handbook of the Protists. Springer, Cham, pp. 1–44.

Gile G.H. and Slamovits C.H. (2012) Phylogenetic position of Lophomonas striata Butschli (Parabasalia) from the hindgut of the cockroach Periplaneta americana. Protist, 163, 274–283. PubMed

Lara E., Chatzinotas A. and Simpson A.G.B. (2006) Andalucia (n. gen.) - the deepest branch within jakobids (Jakobida; Excavata), based on morphological and molecular study of a new flagellate from soil. J. Euk. Microbiol., 53, 112–120. PubMed

Kamikawa R., Kolisko M., Nishimura Y.  et al. (2014) Gene content evolution in discobid mitochondria deduced from the phylogenetic position and complete mitochondrial genome of Tsukubamonas globosa. Genome Biol. Evol., 6, 306–315. PubMed PMC

Yabuki A., Gyaltshen Y., Heiss A.A.  et al. (2018) Ophirina amphinema n. gen., n. sp., a new deeply branching discobid with phylogenetic affinity to Jakobids. Sci. Rep., 8, 16219. PubMed PMC

Yang J., Harding T., Kamikawa R.  et al. (2017) Mitochondrial genome evolution and a novel RNA editing system in deep-branching heteroloboseids. Genome Biol. Evol., 9, 1161–1174. PubMed PMC

Pánek T., Silberman J.D., Yubuki N.  et al. (2012) Diversity, evolution and molecular systematics of the Psalteriomonadidae, the main lineage of anaerobic/microaerophilic heteroloboseans (Excavata: discoba). Protist, 163, 807–831. PubMed

Hanousková P., Táborský P. and Čepička I. (2019) Dactylomonas gen. nov., a novel lineage of heterolobosean flagellates with unique ultrastructure, closely related to the amoeba Selenaion koniopes Park, De Jonckheere & Simpson, 2012. J. Euk. Microbiol., 66, 120–139. PubMed

Pánek T., Simpson A.G.B., Hampl V.  et al. (2014) Creneis carolina gen. et sp. nov. (Heterolobosea), a novel marine anaerobic protist with strikingly derived morphology and life cycle. Protist, 165, 542–567. PubMed

De Jonckheere J.F. (2004) Molecular definition and the ubiquity of species in the genus Naegleria. Protist, 155, 89–103. PubMed

Lax G. and Simpson A.G.B. (2013) Combining molecular data with classical morphology for uncultured phagotrophic euglenids (Excavata): a single-cell approach. J. Euk. Microbiol., 60, 615–625. PubMed

Lukomska-Kowalczyk M., Karnkowska A., Krupska M.  et al. (2016) DNA barcoding in autotrophic euglenids: evaluation of COI and 18S rDNA. J. Phycol., 52, 951–960. PubMed

Tashyreva D., Prokopchuk G., Yabuki A.  et al. (2018) Phylogeny and morphology of new diplonemids from Japan. Protist, 169, 158–179. PubMed

Yabuki A. and Tame A. (2015) Phylogeny and reclassification of Hemistasia phaeocysticola (Scherffel) Elbrachter & Schnepf, 1996. J. Euk. Microbiol., 62, 426–429. PubMed

Lara E., Moreira D., Vereshchaka A.  et al. (2009) Pan-oceanic distribution of new highly diverse clades of deep-sea diplonemids. Environ. Microbiol., 11, 47–55. PubMed

Okamoto N., Gawryluk R.M.R., Del Campo J.  et al. (2019) A revised taxonomy of diplonemids including the Eupelagonemidae n. fam. and a type species, Eupelagonema oceanica n. gen. & sp. J. Euk. Microbiol., 66, 519–524. PubMed

Moreira D., Lopez-Garcia P. and Vickerman K. (2004) An updated view of kinetoplastid phylogeny using environmental sequences and a closer outgroup: proposal for a new classification of the class Kinetoplastea. Int. J. Syst. Evol. Microbiol., 54, 1861–1875. PubMed

Simpson A.G.B., Lukeš J. and Roger A.J. (2002a) The evolutionary history of kinetoplastids and their kinetoplasts. Mol. Biol. Evol., 19, 2071–2083. PubMed

Deschamps P., Lara E., Marande W.  et al. (2011) Phylogenomic analysis of kinetoplastids supports that trypanosomatids arose from within bodonids. Mol. Biol. Evol., 28, 53–58. PubMed

Yazaki E., Ishikawa S.A., Kume K.  et al. (2017) Global Kinetoplastea phylogeny inferred from a large-scale multigene alignment including parasitic species for better understanding transitions from a free-living to a parasitic lifestyle. Genes Genet. Syst., 92, 35–42. PubMed

Flegontova O., Flegontov P., Malviya S.  et al. (2018) Neobodonids are dominant kinetoplastids in the global ocean. Environ. Microbiol., 20, 878–889. PubMed

Callahan H.A., Litaker R.W. and Noga E.J. (2002) Molecular taxonomy of the suborder Bodonina (order Kinetoplastida), including the important fish parasite. Ichthyobodo Necator. J. Euk. Microbiol., 49, 119–128. PubMed

Simpson A.G.B., Gill E.E., Callahan H.A.  et al. (2004) Early evolution within kinetoplastids (Euglenozoa), and the late emergence of trypanosomatids. Protist, 155, 407–422. PubMed

von der Heyden S., Chao E.E., Vickerman K.  et al. (2004) Ribosomal RNA phylogeny of bodonid and diplonemid flagellates and the evolution of euglenozoa. J. Euk. Microbiol., 51, 402–416. PubMed

von der Heyden S. and Cavalier-Smith T. (2005) Culturing and environmental DNA sequencing uncover hidden kinetoplastid biodiversity and a major marine clade within ancestrally freshwater Neobodo designis. Int. J. Syst. Evol. Microbiol., 55, 2605–2621. PubMed

Mukherjee I., Hodoki Y. and Nakano S. (2015) Kinetoplastid flagellates overlooked by universal primers dominate in the oxygenated hypolimnion of Lake Biwa, Japan. FEMS Microbiol. Ecol., 91, 1–11. PubMed

Goodwin J.D., Lee T.F., Kugrens P.  et al. (2018) Allobodo chlorophagus n. gen. n. sp., a kinetoplastid that infiltrates and feeds on the invasive alga Codium fragile. Protist, 169, 911–925. PubMed

Katoh K. and Toh H. (2008) Recent developments in the MAFFT multiple sequence alignment program. Brief. Bioinformatics, 9, 286–298. PubMed

Capella-Gutierrez S., Silla-Martinez J.M. and Gabaldon T. (2009) trimAl: a tool for automated alignment trimming in large-scale phylogenetic analyses. Bioinformatics, 25, 1972–1973. PubMed PMC

Katoh K. and Standley D.M. (2013) MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol. Biol. Evol., 30, 772–780. PubMed PMC

Stamatakis A. (2014) RAxML version 8: a tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics, 30, 1312–1313. PubMed PMC

Comparative Analysis of Protist Communities in Oilsands Tailings Using Amplicon Sequencing and Metagenomics

Water masses shape pico-nano eukaryotic communities of the Weddell Sea

Euglenozoa: taxonomy, diversity and ecology, symbioses and viruses