Carnivores, as focal species, play a significant role in ecosystems stability. In recent decades, increase in carnivore-human conflict has threatened the survival of these species. The assessment of species habitat requirements, landscape connectivity and the protection of biological corridors can ensure the long-term survival of focal and beneficiary species of an ecosystem. We investigated the level of spatial and trophic niche overlap between gray wolf and golden jackal in Isfahan province. Firstly, the landscape resistance surfaces were mapped as a function of 10 environmental and human variables using the ensemble distribution model. Due to the lack of data on dispersal movements of the studied species in Iran, we used different scenarios in relation to dispersal distances to evaluate how robust our predicted core patches are. We used resistant kernel and factorial least-cost path modelling to identify important core habitats and corridors between patches. We calculated the spatial niche overlap using the predicted distribution maps in different suitability thresholds. Also, the trophic niche overlap was assessed using scat analysis and Pianka’s niche overlap index. The results of predicted distribution maps showed that a large part of suitable habitats of ??the two species is located in central and western parts of the studied landscape. There was a high density of predicted corridors between Abbas-Abad Wildlife Refuge and Kahyaz Protected Area indicating significant role of these protected areas in maintaining landscape connectivity of carnivores. The results of trophic niche overlap showed that the abundance of domesticated ungulates, herbs and poultry was higher than other trophic items. This was attributed to the decrease in quality of natural habitats and the opportunistic nature of the studied species. The results of spatial and trophic niche overlap showed relatively high level of overlap between the two species (Schoener's D= 0.78; Pianka index=0.50). Our findings revealed that conserving these species requires integrated landscape-level management. As domestic animals constituted a considerable portion of the species diets, maintaining the quality of natural habitats should be considered in planning future conservation programs in order to mitigate human-wildlife conflict.