Macrophomina phaseolina (Tassi) Goid is a destructive fungus causing root rot disease in more than 500 plant species (Su et al. 2001). During March 2017, sudden wilting of lentil (Lens culinaris) plants grown at NIAB were observed. When the diseased plants were uprooted their taproot was dark with signs of rotting and lateral roots were also destroyed. Pieces from these roots were surface disinfected and plated aseptically onto 2% PDA medium. The fungus growing out of the plated root pieces was purified using the hyphal tip method and incubated at 26 °C. The colony of the purified fungus was appressed, dense and dark brown to black with abundant microsclerotia. Microsclerotia were black and round to oblong or irregular shaped (65–110 μm in diameter). Based on the host symptoms and cultural characters the isolated pathogen was identified as M. phaseolina (Sarr et al. 2014). For molecular confirmation, the internal transcribed spacer (ITS) region of one representative isolate was direct sequenced (Eurofins Genomics, USA), submitted to GenBank (accession No. MH191216) and revealed 99–100% identity in BLASTn with strains of M. phaseolina. Furthermore, 100% identity with an epitype of M. phaseolina CBS 205.47 (KF951622) supports that our isolate (MH191216) fell into a well-defined clade of M. phaseolina as described by Sarr et al. (2014). The characterized culture was deposited (FMB0126) with fungal molecular biology lab-culture collection (FMB-CC) at University of Agriculture, Faisalabad. Pathogenicity of the isolated pathogen was confirmed by pouring 60 ml of microsclerotia suspension (ca. 1 × 104 mL−1) at the base of 40day old 15 potted lentil plants and by placing surface disinfected seeds on the fungal culture (Akhtar et al. 2011; Pastrana et al. 2016). Complete wilting of all the inoculated plants as observed under natural conditions was recorded after 2 weeks while the non-inoculated control plants were symptomless. M. phaseolina was consistently re-isolated from the inoculated plants. Lentil seeds placed on the fungal culture were rotten, failed to germinate or died if germinated covered with microsclerotia. Based on morpho-cultural, ITS region sequence analysis and pathogenicity test, the isolated pathogen was identified as M. phaseolina. M. phaseolina has been reported from multiple hosts in Pakistan, but this is the first report of this fungus naturally infecting lentil in Pakistan.