The yield, the chemical composition, the antibacterial and the antifungal properties of the essential oil extracted from Rosa canina (leaves) and their extracts from two regions in Tunisia (Ain draham and El Feija) were studied. Remarkable differences in the chemical composition of the essential oil and the antibacterial activity according to the province. The yield is situated between 0.45% a 0.6%. Fifty four compounds were identified by GC and GC/MS. The major components of essential oil of Rosa canina resulting from Ain Draham are the Vitispirane (22.5%), the hexdecanoique acid (15.5%), the linoleic acid (13.5%). Whereas, the major constituents of oil obtained from El Feija plant were hexadecanoic acid (23.2%), vitispirane (9.1%), linoleic acid (7.9%), dodecanoic acid (6.4%). The essential oil of R. canina and its extracts showed a significant antimicrobial activity against all four tested microorganisms. To the best of our knowledge, this work is the first of its kind.