Maize rough dwarf disease (MRDD) is one of the most important viral diseases around the world. Rice black-streaked dwarf virus (RBSDV) and maize rough dwarf virus (MRDV) cause MRDD in East Asia and Mediterranean regions, respectively. RBSDV and MRDV are similar in many features such as host range, disease symptoms, vector, and serological relationships, which make it hard to differentiate them based on serological and biological properties. However, molecular analyses as powerful tools could be used for differentiating species of fijiviruses. In this study, the complete genome sequence, genetic variation of 13 isolates using the nucleotide sequence of segment 10 ORF, and host range of MRDD causal agent in Iran were determined. Whole-genome comparisons and phylogenetic analysis showed that Iranian isolate shared high nucleotide sequence identities ranging from 86.5–91.3% and closest evolutionary relationship to the RBSDV Chinese isolates. The Iranian isolates showed over 99% homology based on amino acid sequence of ORF10 and were grouped in one clade through phylogenetic analysis. The results revealed that ORF10 of MRDD causal agent was under negative selection. Iranian isolate of MRDD was experimentally transmitted to maize, rice, barley, wheat, millet, oat, rye, and some grasses by Loadelphax striatellus with about 32% efficiency. The results of this study revealed that the MRDD causal agent in Iran had the closest genetic similarity to RBSDV demonstrating it is closer to RBSDV than MRDV.