<p>Karst areas typically exhibit a dual geological structure characterized by the coexistence of highly developed surface pore‑fissure systems and underground conduit flow systems, resulting in rapid connectivity and high vulnerability between surface water and groundwater. The Outstanding Universal Value (OUV) of the South China Karst World Natural Heritage sites is increasingly threatened by anthropogenic nitrate pollution. This study focuses on the Huanghou underground river basin, where water samples were collected during the rainy and dry seasons for analysis. By integrating hydrochemistry, nitrogen and oxygen isotope techniques, and a Bayesian mixing model, nitrate sources were identified and their relative contributions were quantified. The results showed that: (1) nitrate concentrations at all sites met China’s drinking-water standards and were higher in the rainy season owing to rainfall-driven flushing and percolation; (2) nitrate mainly originated from soil organic nitrogen, chemical fertilizers, and manure and sewage, and its transformation was dominated by nitrification; (3) in the rainy season, the contributions of these three sources were 34.4%, 45.7%, and 12.4%, respectively, whereas in the dry season they were 41.6%, 28.1%, and 22%, respectively; and (4) the key to controlling nitrate pollution in the Huanghou underground river basin lies in limiting excessive fertilizer application in upstream farmland and reducing domestic sewage discharge from residential areas. This study provides a scientific basis for groundwater management and nitrate pollution control in karst World Natural Heritage sites, and offers theoretical support for protecting the OUV and promoting the sustainable development of such heritage sites.</p>