Biomonitoring of least disturbed areas is a crucial issue to accurately assess the ecological status of surface waters and ensure their sustainability. Diatom metrics are important tools for the assessment of environmental conditions of lotic ecosystems to achieve environmental sustainability. The present study was aimed to evaluate the ecological status of least disturbed areas in the Antalya River basin (Turkey) using diatom indices developed from different ecoregion. Diatom species such as Cymbella excisa, Achnanthidium minutissimum, Cocconeis euglypta, and Cocconeis placentula were the most contributing species to the dissimilarity of sampling stations between rainy and dry seasons. The first two axes of canonical correspondence analysis (CCA) elucidated a strong (86%) correlation between diatom taxa and stressors. Results of the Monte Carlo permutation test indicated that total phosphorus (TP), water temperature, Tin (Sn), total organic carbon, zinc, and pH are significantly influential ecological factors on the distribution of diatom taxa among sampling stations. The Spearman correlation rank test indicated that diatom indices had significant positive and negative correlations with TP gradient. Results revealed that diatom indices developed from different ecoregions have different scores for the eco-assessment of similar sampling stations. European diatom indices mostly showed similar behavior in the bioassessment of the ecological status of rivers in the Antalya region compared to the different ecoregion. Among European diatom indices, TIT was more competitive and could give better results in the bio-evaluation of rivers in the Antalya River basin. Using diatom indices developed from different ecoregions may lead to an erroneous assessment of water quality. Accordingly, ecoregional specific diatom metrics are needed to accurately determine the surface water quality. Eco-assessment of least disturbed areas of the Antalya River basin: application of diatom indices from different ecoregions: Abuzer Çelekli, Ömer Lekesiz, Tolga Çetin.