Essential oils of four different Eucalyptus species (Eucalyptus salubris, Eucalyptus salmonophloia, Eucalyptus oleosa, and Eucalyptus gracilis) grown in southern Tunisia were screened for their antioxidant and antimicrobial properties as well as their chemical compositions. According to gas chromatography-flame ionization detection and gas chromatography-mass spectrometry analysis, chemical compositions of the Eucalyptus species E. salubris (27 compounds; 99.2%), E. salmonophloia (31 compounds; 99.2%), E. oleosa (32 compounds; 97.6%), and E. gracilis (18 compounds; 97.7%) were identified. In the 1,1-diphenyl-2-picrylhydrazyl assay, the antioxidant activity was in the range of 12.0-52.8 mg/mL, whereas in the 2,2'-azinobis-3-ethylbenzothiazoline-6-sulfonate assay, E. oleosa (176.5 +/- 3.1 mg/L) gave the best inhibition result. To evaluate antimicrobial activity, all essential oils were tested against bacteria (two Gram-positive and two Gram-negative), two yeast, and two fungi. Essential oils exhibited an interesting antibacterial activity against all microorganisms tested (activity was better against Gram-positive bacteria) except for Staphylococcus aureus and Escherichia coli. Correlations between chemical composition and biological and antioxidant activities were studied.