The biodiversity crisis is one of the greatest challenges facing humanity, but our understanding of the drivers remains limited. Thus, after decades of studies and regulation efforts, it remains unknown whether to what degree and at what concentrations modern agricultural pesticides cause regional-scale species losses. We analyzed the effects of pesticides on the regional taxa richness of stream invertebrates in Europe (Germany and France) and Australia (southern Victoria). Pesticides caused statistically significant effects on both the species and family richness in both regions, with losses in taxa up to 42% of the recorded taxonomic pools. Furthermore, the effects in Europe were detected at concentrations that current legislation considers environmentally protective. Thus, the current ecological risk assessment of pesticides falls short of protecting biodiversity, and new approaches linking ecology and ecotoxicology are needed.