Microdochium spp. are the causal agents of seedling blight, crown rot, leaf blotch, and head blight on wheat and other small-grain cereals grown in many areas worldwide. Twenty isolates of Microdochium sp. obtained from symptomatic wheat crowns were collected from nine provinces in the center and to the east of northern Algeria. The assignment of isolates to the genus Microdochium was based on microscopic and macroscopic morphological criteria. Using species-specific primers, the two species M. nivale and M. majus were identified. Molecular identification revealed that 75% of isolates were M. nivale (n = 16), and 25% of isolates were M. majus (n = 4). The two species showed optimal conidiation and mycelial growth at 20 °C and 25 °C, respectively. Furthermore, potato sucrose-salt agar together with potato dextrose agar media appeared to be the most favorable for conidiation and mycelial growth, contrary to synthetic nutrient-poor agar, which was unfavorable for both conidiation and mycelial growth. The aggressiveness of Microdochium spp. isolates was assessed by in vitro and in planta bioassays on wheat leaves, crowns, and heads. The results obtained showed no significant difference between the crown rot severity of M. nivale and M. majus (P = 0.22), with mean disease indices of 1.7 and 1.38, respectively. In contrast, M. nivale was more aggressive on wheat leaves and heads than M. majus (P < 0.01 and P < 0.05, respectively). The data obtained in the current study revealed that M. nivale isolates were more aggressive than M. majus isolates on Algerian wheat.