In July 2017, symptoms of root rot, leaf yellowing, and death were observed on Mexican lime (Citrus aurantiifolia) plants in a nursery located in Roudan, Hormozgan province. The rotted roots were dark brown with numerous black microsclerotia present in the vascular and on cortical tissues. Symptomatic root fragments were surface-disinfested with 2% sodium hypochlorite solution for 3 min, rinsed three times in sterile distilled water, dried on sterilised paper, and plated onto potato dextrose agar (PDA) supplemented with 150 μg ml−1 of streptomycin sulfate. Plates were incubated at 30 °C for 6 days with a 12-h photoperiod. In culture, the mycelium was initially hyaline, later becoming grey and after 4-5 days of incubation, numerous black, spherical, oblong or irregularly shaped microsclerotia, 60-118 x 52-98 µm, developed in the colonies. Pycnidia were not produced by any isolate. The morphology of the isolates was identical to that described for M. phaseolina. To confirm the identification, genomic DNA from a representative isolate was extracted and the internal transcribed spacer region (ITS1-5.8S-ITS2) was amplified by PCR and sequenced using the universal primers ITS1 and ITS4. BLASTn search of the sequence (Accession No. MG636998) showed 100% identity with sequences of M. phaseolina (KR012878.1, GU046902.1 and GU046898.1) obtained from National Center for Biotechnology Information (NCBI). To establish pathogenicity, microsclerotia of six hyphal-tipped isolates of M. phaseolina, including the representative isolate, were used as inoculum. Two-month-old seedlings were transplanted into pots containing autoclaved peat/soil mix infested with the microsclerotia (100 viable microsclerotia g-1 soil) and maintained in a greenhouse at 30+/-2°C. Control seedlings were transferred to non-infested soil. After three weeks, all inoculated seedlings began to show symptoms on leaves and roots, similar to the symptoms of the nursery seedlings. Five weeks after inoculation, disease severity ranged from 75 to 100% depending on isolate. Microsclerotia were produced after seven weeks on roots of the seedlings. No symptoms were observed on control seedlings. M. phaseolina was reisolated only from inoculated seedlings. Based on the incidence and severity of symptoms, charcoal root rot of citrus, can be considered as a potential threat to citrus production in southern Iran. This is the first report of charcoal rot caused by M. phaseolina on citrus in Iran.