Abstract: The fungal flora with special attention to Trichoderma in 23 soil samples collected from Al-Jabal Al-Akhdar Region, Libya was studied using different culture media. Trichoderma Selective Medium (TSM), Martin's medium (MT) and Potato Dextrose Agar (PDA) were the superior media for isolating Trichoderma. Martin's medium (MT) and Malt Extract medium (ME) were the most valuable for isolating the greatest number of total fungal count. Trichoderma Selective Medium (TSM) supplemented with 100 μg mL-1 PCNB was the most effective medium for counting Trichoderma and recorded 120-140% efficacy of re-isolation. Trichoderma occurred in moderate frequency in the tested soil and was isolated from 5 soil samples. Trichoderma counted 0.5-1x103 CFU g-1 dry soil and five Trichoderma isolates were identified as Trichoderma harzianum. Aspergillus and Penicillium sp. were the most frequent fungi isolated from the tested soil and were averaged 8.3-5.5 CFU mg-1 soil, respectively. Further studies are needed to clarify the distribution of fungal flora especially Trichoderma sp. in the Libyan soil.