Accepted_test

Data on naked lobose amoebae species distribution in natural ecosystems remain limited since their morphological identification and molecular barcoding using 18S rRNA and Cox 1 genes still require cultivation and long-term observation. However metabarcoding allows to asses amoebae diversity in environmental samples without labor-consuming maintenance of clonal cultures. In this study V4 marker region of 18S rRNA gene was amplified from total DNA extracted from 9 freshwater bottom sediments and 2 soil samples from geographically distant sampling sites and sequenced using llumina MiSeq platform. Sequencing data were analyzed using Mothur software package and MiSeq SOP pipeline. Representatives of all major phylogenetic groups of naked lobose amoebae (Variosea. Cutosea, Tubulinea, Flabellinia and Longamoebia) were identified. Surprisingly, Variosea (Filamoeba, Arboramoeba, Heliamoeba, Flamella, Telaepolella, Ischnamoeba, Dictyamoeba, Angulamoeba) and Tubulinea (Flabellula, Leptomyxa, Copromyxa, Ptolemeba, Hartmannella, Echinamoeba, Vermamoeba) visibly dominated within obtained Amoebozoa reads. In addition to species that are commonly mentioned in ecological studies, metabarcoding provided an access to data on amoebae that are rarely recovered in cultures, like representatives of Filamoeba, Angulamoeba, Echinamoeba, Vermamoeba and Mycamoeba genera. Newly obtained data also illustrated a predicted patchy pattern of amoebae occurrence in the environment, resulted from discreteness of freshwater and soil habitats in terms of ecological conditions on a microscopic level.