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The developing resistance of plant pathogenic fungi to commercial fungicides has become a serious problem for efficient plant disease control. The use of antifungal preparations based on living microorganisms or their metabolites represents one of the possible environmentally friendly approaches. However, since a complete rejection of chemical fungicides is impossible, the combining of biopreparations and fungicides may be considered a promising biocontrol approach. Promising strains for the development of antifungal biopreparations include Penicillium fungi producing various biologically active compounds with antimicrobial and antiviral activities. A dry biomass of the P. chrysogenum F-24-28 strain (DMP) obtained from the P. chrysogenum VKPM F-1310 strain by induced mutagenesis possessed a high antifungal efficiency. According to in vitro experiments, supplementation of agarized medium with DMP (7.5–10 g/L) resulted in a significant growth inhibition in several plant pathogenic Fusarium fungi. The combination of DMP with a commercial azoxystrobin-based fungicide resulted in a prolonged growth inhibition in F. oxysporum, F. graminearum and F. culmorum even at fungicide concentrations significantly below the recommended level (0.5–2.5 mg/L or 2.5–12.5 g/ha vs. the recommended 100–275 g/ha). These results demonstrate a possibility to develop an efficient environmentally friendly biopreparation suitable to control crop diseases caused by a wide range of plant pathogens, and to prevent a possible selection and spreading of resistant pathogen strains.