Tigers continue to face unprecedented threats to their existence due to poaching, habitat loss, habitat fragmentation and anthropogenic disturbances. The present study examines the physiological stress response of tigers due to anthropogenic activities including wildlife tourism in Bandhavgarh Tiger Reserve and Kanha Tiger Reserve using faecal glucocorticoid metabolite (fGCM) measurement. We collected a total of 341 faecal samples from both reserves during tourism and non-tourism periods. Data on various anthropogenic disturbances including tourism activities like number of vehicles and visitors were also collected. We ascertained the species identity and sex of all the samples collected using genetic markers. fGCMs were extracted using a previously reported procedure, and fGCM concentrations were subsequently determined using an established enzyme immunoassay. There was no significant difference in overall mean fGCM concentrations between the two tiger reserves, but within each reserve, concentrations were significantly higher in tigers during the tourism period as compared to the non-tourism period. We also found that the number of tourist vehicles and disturbance level significantly correlated with fGCM concentrations. This study further supports the assumption that unbridled tourism associated with high anthropogenic disturbance can be related to perceived stress and consequently may have an impact on the reproductive fitness of tigers and long-term survival of isolated populations.