Background:Plant extracts are important products in the world and have been widely used for isolation of important biologically active products. Because of their significant environmental impact, extensive research has been explored to determine the antimicrobial activity of plant extracts.Methods:Acetone extracts of the bark and leaf of Cupressus sempervirens and Juniperus phoenicea, collected from three different altitudes (125, 391, and 851 m high of sea level) at Al- Jabel Al-Akhdar area, Libya were obtained and analyzed by GC/MS. The antimicrobial activity of the extracts was further evaluated against plant bacteria Rhizobium radiobacter, Erwinia carotovora, Rhodococcus fascians and Ralstonia solanacearum and fungus Botrytis cinerea.Results:The impact of the altitude from the sea level on the quantity and chemical constituents of the extracts was investigated. The yield was largely dependent on tree species and the highest yield (6.50%) was obtained with C. sempervirens L bark of altitude III (851 m of the sea level), while the lowest (1.17%) was obtained with the leaf extract of C. sempervirens L from altitude I (125 m). The chemical composition analyzed by GC/MS confirmed that the leaf extracts of C. sempervirens and J. phoenicea contained a complex mixture of monoterpene hydrocarbons, sesquiterpenes, diterpenes, diterpenoids, terpenophenolic, steroids and phthalates. However, the bark extracts of both trees contained a mixture of sesquiterpenes, diterpenes, diterpenoids, terpenophenolics, phthalates, retinol and steroids. These constituents revealed some variability among the extracts displaying the highest interesting chemotype of totarol (terpenophenolic) in all extracts (14.63-78.19% of the total extract). The extracts displayed a noteworthy antifungal potency with varying degrees of inhibition of growth with EC50 values ranged from 78.50 to 206.90 mg/L. The extracts obtained from the leaves of C. sempervirens showed that the highest inhibitory activity was obtained with the extract of altitude II (391 m) with MIC 565, 510, 380 and 710 mg/L against E. carotovora, R. fascians, and R. radiobacter and R. solanacearum, respectively.Conclusion:Based on antimicrobial activity, raw plant extracts can be a cost-effective way to protect crops from microbial pathogens. Because plant extracts contain several antimicrobial compounds, the development of resistant pathogens can be delayed.