The composition and antimicrobial activity of the essential oil from Verbesina negrensis Steyerm. leaves is reported. Analysis was performed by GC/MS. Major constituents were α-pinene (43.1%), α-humulene (13.8%), Δ-cadinene (8.1%), limonene (4.6%) and bicyclogermacrene (4.2%). The essential oil showed in vitro activity against the Gram-positive bacteria Staphylococcus aureus ATCC 25923 and Enterococcus faecalis ATCC 29212 (MIC 500 and 350 μL/mL respectively).