The natural occurrence of entomophthoralean fungi pathogenic towards aphids on cereal and potato crops was investigated in the years 2009, 2010, and 2011. Infected aphids were sampled in three bioclimatic zones in Tunisia (Beja, Cap bon, and Kairouan) and fungal species were determined based on morphological characters such as shape, size, and number of nuclei in the primary conidia. Polymerase Chain Reaction (PCR) on the internal transcribed spacer 1 region (ITS1) was used to verify morphological determination. Both methods gave consistent results and we documented for the first time the natural occurrence of two fungal species from the order Entomophthorales (phylum Entomophthoromycota), Pandora neoaphidis and Entomophthora planchoniana. Both fungi were recorded on the aphid species Sitobion avenae and Myzus persicae on barley ears and potato leaves, respectively. Moreover, natural mixed infections by both species (P. neoaphidis and E. planchoniana) were documented on the target aphids. This investigation provides basic information of entomopathogenic fungi infecting economically important aphids in Tunisia.