Biocontrol products based on microorganisms and natural substances are promising alternatives to chemical pesticides that could contribute to develop a more sustainable agriculture. Here, we investigated the potential of cell-free culture filtrates (CFCFs) from two strains of the Bacillus subtilis group to inhibit Zymoseptoria tritici, a major fungal pathogen of wheat. Foliar application of CFCFs from Bacillus velezensis GA1 and Bacillus sp. III1 on wheat seedlings in a greenhouse strongly reduced Z. tritici disease severity (>90%). In vitro bioassays showed that CFCFs completely inhibited the spore germination and fungal growth (100%). In planta cytological investigations revealed a significant impact of the treatments on both spore germination (∼40% inhibition) and fungal growth of Z. tritici (>80% inhibition). High Performance Liquid Chromatography (HPLC) analysis showed that the Bacillus strains displayed different lipopeptide profiles. The CFCF obtained from Bacillus GA1 contained 90mg l-1 of iturin A+surfactins+fengycins and the CFCF obtained from Bacillus sp. III1 contained 25mg l-1 of mojavensin A (iturin family) + surfactins+fengycins. Strains of the B. subtilis group producing different iturins could provide several CFCF-based solutions for the biocontrol of Z. tritici.