Triazole resistance in Aspergillus fumigatus develops in patients with chronic lung diseases receiving long-term azole therapy or by environmental selection of resistant A. fumigatus. Here we report for the first time the isolation of triazole-resistant A. fumigatus (TRAF) harbouring the G54E mutation from environmental samples in India, Romania and Tanzania. This mutation in the cyp51A azole target gene of A. fumigatus is so far considered as de novo occurring in patients due to prolonged exposure to azoles. A total of 81 soil and woody debris samples from India, Romania and Tanzania were processed for detection of TRAF and determination of their susceptibility to medical triazoles and fungicides. cyp51A sequencing and real-time PCR were performed for detection of mutations. The isolates were genotyped by microsatellite typing. Overall, 25% of samples (20/81) from India, Romania and Tanzania harboured TRAF. Of the 20 samples harbouring TRAF, a single resistance mechanism, the G54E mutation, was found in 16 samples from three countries. This mechanism was responsible for 46.4% of resistant isolates from Tanzania, 30.4% from Romania and 20.0% from India. The G54E isolates revealed high MICs of itraconazole and posaconazole and were cross-resistant to agricultural fungicides. The majority of the Romanian and Tanzanian G54E isolates had an identical genotype. The present report describes the genetic heterogeneity of TRAF strains harbouring the G54E mutation in the environment of India, Romania and Tanzania. It may be anticipated that long-term exposure of A. fumigatus to fungicides may induce selection of G54 mutants in the environment.