Eucalyptus grandis is a medicinal plant which has been indicated by Zulu traditional healer in the treatment of respiratory tract infections, bronchial infections, asthma and cough. The investigation of the essential oil of this plant could help to verify the rationale behind the use of the plant as a cure for these illnesses. Essential oil was hydro-distilled from the fresh leaves and characterised for the chemical constituents and bioactivity. The main constituents of the oil of the E. grandis are α-Pinene (29.69%), p-Cymene (19.89%), 1,8-cineole (12.80%), α-Terpineol (6.48%), Borneol (3.48%) and D-Limonene (3.14%). The essential oil of E. grandis showed high scavenging of DPPH and ABTS radicals, and was actively against 13 of the 16 organisms tested with the MIC ranging from 0.625 mg - 5.0 mg/ml; the MBC value ranged from 2.5 mg - 10 mg/ml. The essential oil also inhibited the growth of 7 of the 8 antibiotic resistant bacteria tested, with MIC ranging from 5 mg/ml - 10 mg/ml. The DNA extracted from the affected microorganisms did not show any damage however, there was an increase of released cytosolic LDH activity. We conclude that the antibacterial activity of the essential oil was exhibited through cell membrane damage rather than the damage of the DNA. It is apparent that the bioactivity of the essential oil of E. grandis plays an important role in the plants’ use in folk medicine for the treatment of respiratory tract illnesses.