Chitwan-Annapurna Landscape (CHAL) with an area of 32,057 km² is a major biodiversity hotspot in central Nepal. It acts as a biological corridor that links Chitwan National Park to Annapurna Conservation Area (ACA). Four potential north south corridors were identified along CHAL but the functionality of these corridors was not clear.  We study to assess the validity of two major corridors: i) Barandabhar Forest – Gaighat –Seti River Valley – Madi River Valley – ACA and ii) Nawalparasi Forest–Palpa – Syangja – Panchase – ACA. Mammals, birds, fishes, plants and major infrastructure development were studied using direct and indirect methods (field observation, transects and plots, camera trapping, sign surveys, mist-netting, and questionnaire survey) in summer and winter seasons. Besides, potential corridors, biodiversity of eight Biodiversity Important Areas (Madi River valley, Eastern Himalayan Broadleaf Forest, Daraundi River Valley, Eastern Himalayan Alpine Meadows, Bhimtang Forest, Resunga Forest, Ganesh Himal Base Camp, and Simbhanjhyang Forest) were also explored for their baseline information. The field survey identified 28 species of mammals (herbivores, carnivores, primates, rodents and bats), 283 species of birds, 44 species of fish, 125 species of trees, 129 species of shrubs, and 234 species of herbs from the study sites. Mini corridors in different study sites supporting north-south linkage for wildlife movement have been identified as i. Panchase-Putalibazar-Tikhedunga, Jhyankatta-Khudi, Raipur-Saranghat, SukauraAahaldanda with their interlinkages ii. Chakchake – Sildjure, Devghat- Sikles, Majthana- Raniban, Gajarkot- Rhinoblock with their interlinkages.