In Bhutan, protected areas constitute 51.4% of the total geographical area, out of which 7.7% is designated biological corridors (BCs) that serve as connectors between protected areas. The biological corridor (BC-03) constitutes a total area of 407.7 km² and connects Phibsoo Wildlife Sanctuary in the west, Jigme Singye Wangchuck National Park and Royal Manas National Park in the north and southeast through Sarpang–Tsirang District landscapes. However, most BCs, including BC-03, lack comprehensive inventories of mammal species. We conducted joint rapid biodiversity assessment (RBA) using camera trap surveys from November 2019 to February 2020. Our findings from BC-03 include records of 26 mammal species belonging to 11 families in five orders. Nearly half of these species are listed as Endangered, Vulnerable, or Near Threatened. Overall, the BC-03 landscape was found to support 77.8% of the mammal species of Sarpang District and 21.7% of Bhutan (as per Biodiversity Statistics of Bhutan, 2017). Therefore, BC-03 is of conservation interest. Comprehensive conservation plans, periodic monitoring of keystone species, and restrictions on the expansion of cardamom plantations in BCs are suggested for securing wildlife habitats and ensuring long-term persistence of keystone species, including within BC-03 in southcentral Bhutan.