Recent changes in taxonomy of Rhizopus, which are now heavily relying on molecular approach, create significant problem in assigning species name to particular Rhizopus strains isolated from various sources, including tempeh. The present study aims to determine 36 strains of Rhizopus from tempeh originated from 26 locations in Indonesia, using combination of molecular phylogenetic analysis based on ITS rDNA sequence, physiology, and morphology to species level. The results showed that most of the strains belong to Rhizopus microsporus complex, and only one strain belongs to Rhizopus delemar. Morphological variation within R. microsporus were observed, but under current approach they were insufficient for infraspecific delimitation. The current report is an important contribution in validating Rhizopus identity from fresh tempeh in Indonesia.