Key biodiversity areas (KBAs) are critical regions for preserving global biodiversity. KBAs are identified by their importance to biodiversity rather than their legal status. As such, KBAs are often under pressure from human activities. KBAs can encompass many different land-use types (e.g., cropland, pastures) and land-use intensities. Here, we combine a global economic model with spatial mapping to estimate the biodiversity impacts of human land use in KBAs. We find that global human land use within KBAs causes disproportionate biodiversity losses. While land use within KBAs accounts for only 7% of total land use, it causes 16% of the potential global plant loss and 12% of the potential global vertebrate loss. The consumption of animal products accounts for more than half of biodiversity loss within KBAs, with housing the second largest at around 10%. Bovine meat is the largest single contributor to this loss, at around 31% of total biodiversity loss. In terms of land use, lightly grazed pasture contributes the most, accounting for around half of all potential species loss. This loss is concentrated mainly in middle- and low-income regions with rich biodiversity. International trade is an important driver of loss, accounting for 22–29% of total potential plant and vertebrate loss. Our comprehensive global, trade-linked analysis provides insights into maintaining the integrity of KBAs and global biodiversity.