The aim of this study was to examine the community structure of the order Ephemeroptera in the Eastern Part of the Black Sea Region of Turkey by using multivariate analyses such as Canonical Correspondence Analysis (CCA) and Two-Way Indicator Species Analysis (TWINSPAN). For this purpose, 12,117 individuals of Ephemeroptera belonging to seven families, 11 genera and 28 species were sampled at 24 collecting sites and identified. Environmental variables measured at each collecting site were dissolved oxygen (DO), water temperature, electrical conductivity (EC), pH, PO4-P, NO3-N, NO2 -N, NH4-N and SO4. The study area covered the eastern part of the Black Sea Region of Turkey, as it is a sub-ecoregion of the Caucasus Biodiversity Hotspots. With CCA analysis, the relationship between the mayfly species and the environmental variables was analysed for the first time for the region. In addition, TWINSPAN analysis determined the indicator species at the collecting sites. According to the results of CCA, Baetis milani, B. gemellus, Alainites muticus and Electrogena quadrilineata were positively related to PO4-P but negatively related to temperature. Baetis rhodani, B. lutheri, B. fuscatus, Electogena affinis, E. starmachi, Caenis macrura, C. luctuosa and Epeorus zaitzevi were positively related to SO4. NO2-N. EC and pH but negatively related to dissolved oxygen. Rhithrogena iridina, R. zelinkai, Ecdyonurus picteti, E. helveticus, Epeorus sylvicola, E. znojkoi, E. caucasicus and Habroleptoides confusa were positively correlated with DO but negatively correlated with SO4, EC, NO2-N and pH. Rhithrogena semicolorata, R. puytoraci, R. beskidensis, Seratella ignita and Potamanthus luteus were positively correlated with temperature but negatively correlated with PO4-P. In addition, according to the results of TWINSPAN, the collecting sites divided into two main group (Group A and B). Epeorus sylvicola, E. znojkoi and E. caucasicus were assigned as the indicator species for the sites in Group A. However, there were no indicator species assigned for Group B.