Telonemia are one of the oldest identified marine protists that for most part of their history have been recognized as a distinct incertae sedis lineage. Today, their evolutionary proximity to the SAR supergroup (Stramenopiles, Alveolates, and Rhizaria) is firmly established. However, their ecological distribution and importance as a natural predatory flagellate, especially in freshwater food webs, still remain unclear. To unravel the distribution and diversity of the phylum Telonemia in freshwater habitats, we examined over a thousand freshwater metagenomes from all over the world. In addition, to directly quantify absolute abundances, we analyzed 407 samples from 97 lakes and reservoirs using Catalyzed Reporter Deposition-Fluorescence in situ Hybridization (CARD-FISH). We recovered Telonemia 18S rRNA gene sequences from hundreds of metagenomic samples from a wide variety of habitats, indicating a global distribution of this phylum. However, even after this extensive sampling, our phylogenetic analysis did not reveal any new major clades, suggesting current molecular surveys are near to capturing the full diversity within this group. We observed excellent concordance between CARD-FISH analyses and estimates of abundances from metagenomes. Both approaches suggest that Telonemia are largely absent from shallow lakes and prefer to inhabit the colder hypolimnion of lakes and reservoirs in the Northern Hemisphere, where they frequently bloom, reaching 10%-20% of the total heterotrophic flagellate population, making them important predatory flagellates in the freshwater food web.

Aquatic Botany and Microbial Ecology Research Group HUN REN Balaton Limnological Research Institute 8237 Tihany Hungary

Biological Station Lake Neusiedl Seevorgelände 1 7142 Illmitz Austria

Cavanilles Institute of Biodiversity and Evolutionary Biology University of Valencia E 46980 Paterna Valencia Spain

Center of Excellence for Science and Technology Integration of Mediterranean Region Zagreb Croatia

Centre for Limnology Estonian University of Life Sciences 6117 Vehendi Tartu County Estonia

Department of Aquatic Microbial Ecology Institute of Hydrobiology Biology Centre of the Czech Academy of Sciences 37005 České Budějovice Czech Republic

Department of Aquatic Sciences and Assessment Swedish University of Agricultural Sciences 750 07 Uppsala Sweden

Department of Botany and Plant Biology Microbiology Unit University of Geneva Sciences 3 CH 1211 Geneva Switzerland

Department of Ecology and Genetics Limnology Uppsala University SE 75236 Uppsala Sweden

Department of Fisheries Ministry of Natural Resources and Climate Change 593 Lilongwe Malawi

Department of Fisheries Oceanography and Marine Ecology National Marine Fisheries Research Institute 81 332 Gdynia Poland

Department of Freshwater and Marine Ecology Institute for Biodiversity and Ecosystem Dynamics University of Amsterdam Amsterdam 1098 XH The Netherlands

Department of Functional Ecology Institute of Botany of the Czech Academy of Sciences 252 43 Průhonice Czech Republic

Department of Hydrobiology Faculty of Biology Institute of Ecology Biological and Chemical Research Centre University of Warsaw Żwirki i Wigury 101 02 089 Warsaw Poland

Department of Ichthyology and Aquatic Environment School of Agricultural Sciences University of Thessaly 38446 Volos Greece

Department of Molecular Biology and Biotechnology Faculty of Biology and Geology Babeş Bolyai University 5 7 Clinicilor Street 400006 Cluj Napoca Romania

Department of Plankton and Microbial Ecology Leibniz Institute for Freshwater Ecology and Inland Fisheries Alte Fischerhuette 2 D 16775 Neuglobsow Germany

Division of Materials Chemistry Ruđer Bošković Institute Bijenička Cesta 54 10000 Zagreb Croatia

État de Vaud Direction de l'environnement industriel urbain et rural 1066 Epalinges Switzerland

Evolutionary Genomics Group Departamento de Producción Vegetal y Microbiología Universidad Miguel Hernández 03550 San Juan de Alicante Alicante Spain

Faculty of Science University of South Bohemia 37005 České Budějovice Czech Republic

Hydrobiological Station Faculty of Biology University of Warsaw Pilchy 5 12 200 Pisz Poland

Institute of Biochemistry and Biology Potsdam University Maulbeerallee 2 D 14469 Potsdam Germany

Institute of Earth Sciences University of Applied Sciences and Arts of Southern Switzerland Campus Mendrisio Via Flora Ruchat Roncati 15 CH 6850 Mendrisio Switzerland

Institute of Microbiology University of Applied Sciences and Arts of Southern Switzerland Campus Mendrisio Via Flora Ruchat Roncati 15 CH 6850 Mendrisio Switzerland

Laboratoire Microorganismes Génome et Environnement CNRS Université Clermont Auvergne 63000 Clermont Ferrand France

Laboratory of Marine Microbiology Institute of Oceanography and Fisheries Šetalište Ivana Meštrovića 63 21000 Split Croatia

Lake and Glacier Ecology Research Group Department of Ecology University of Innsbruck A 6020 Innsbruck Austria

Leibniz Institute for Baltic Sea Research Warnemünde Seestrasse 15 D 18119 Rostock Germany

Limnological Station Department of Plant and Microbial Biology University of Zurich 8802 Kilchberg Switzerland

NBFC National Biodiversity Future Center 90133 Palermo Italy

Research and Innovation Centre Fondazione Edmund Mach Via E Mach 1 38098 S Michele all'Adige Italy

Research Department for Limnology Mondsee University of Innsbruck A 5310 Mondsee Austria

School of Life Sciences University of Warwick CV4 7AL Coventry United Kingdom

Unité d'Ecologie Systématique et Evolution CNRS Université Paris Saclay AgroParisTech 91190 Gif sur Yvette France

Water Research Institute National Research Council Largo Tonolli 50 Verbania 28922 Italy

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