Mesosphaerum suaveolens (Lamiaceae) is a medicinal plant commonly used in Brazil for the treatment of diseases related to the digestive tract and respiratory diseases, so we hypothesized that the essential oil of this species may have antibacterial activity. Thus, we aimed to evaluate the in vitro antibacterial and modulatory activity of the essential oil of M. suaveolens as well as to characterize its chemical composition. The identification of the constituents was performed by gas chromatography-flame ionization detector (GC-FID) and the antibacterial and modulating activity by the plate microdilution method. We found the oil had sesquiterpene β-caryophyllene as the major component. This compound may account for the antibacterial activity against Staphylococcus aureus strains, since the essential oil had a MIC of 64 μg/mL for the standard strain and 256 μg/mL for the multiresistant strain, demonstrated that the oil does not exhibit drug modulating activity. Thus, M. suaveolens oil has bioactive compounds which can be used in the preparation of drugs.