The rice blast pathogen Magnaporthe oryzae is a global threat to rice production. Here we characterized RHO2 gene (MGG_02457) that belongs to the Rho GTPase family, using a deletion mutant. This mutant ΔMorho2 exhibited no defects in conidiation and germination but developed only 6% of appressoria in response to a hydrophobic surface when compared to the wild-type progenitor. This result indicates that MoRHO2 plays a role in appressorium development. Furthermore, exogenous cAMP treatment on the mutant led to appressoria that exhibited abnormal morphology on both hydrophobic and hydrophilic surfaces. These outcomes suggested the involvement of MoRHO2 in cAMP-mediated appressorium development. ΔMorho2 mutation also delayed the development of appressorium-like structures (ALS) at hyphal tips on hydrophobic surface, which were also abnormally shaped. These results suggested that MoRHO2 is involved in morphological development of appressoria and ALS from conidia and hyphae, respectively. As expected, ΔMorho2 mutant was defective in plant penetration, but was still able to cause lesions, albeit at a reduced rate on wounded plants. These results implied that MoRHO2 plays a role in M. oryzae virulence as well.