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Testate amoebae are unicellular organisms that occur worldwide in lacustrine habitats and have been widely used as proxies for palaeoreconstruction and bioindicators of contemporary ecosystems. Our knowledge of the environmental factors determining the spatial distribution of testate amoebae in freshwater stratified lakes is still limited. The aim of this study was to assess the influence of environmental variables on testate amoebae assemblages inhabiting both relatively close and distant habitats within one lake basin and to determine the factors influencing the species structure and spatial distribution of testate amoebae communities along a depth gradient and within one limnological zone In our study we used multystation approach within one waterbody and collected samples at five transects in four limnological zones, total 69 samples from 24 sites. Redundancy and k-means analysis distinguished four groups of samples; two represent shallow water, one sublittoral and one deep depth habitats. The analysis of the dominance species structure of groups allows to select variables influencing tetstae amoebae communities along a depth gradient and within limnological zones. o We found that the number of groups of samples identified in the limnological zone reflects the level of variability of environmental conditions within the limnological zone. Our results highlight three mechanisms determining species structure along a depth gradient, in shallow water habitats and in deep depth habitats in stratified freshwater lake. Changes of species structure along a depth gradient were determined by changes in TEMP, COND and BGA-PE, in shallow water by amount of sand content in sediments, presence or absence of water plants and ORP, BGA-PC, pH, COND and PPT and in deep depth habitats by DOC and BGA-PE.