<p>The intensive use of soil and the increased consumption of chemical fertilizers have risen over the years, causing soil degradation and impacting human health. The application of the fungus <i>Trichoderma</i> spp. has expanded exponentially and has been used to enhance soil fortification and detoxification of both soil and wastewater. Additionally, it has been applied in agricultural systems as a biocontrol agent against pathogens, to stimulate plant growth, and as a biological fertilizer to improve soil fertility. In this study, three <i>Trichoderma</i> strains (<i>T. harzianum</i> MMBF 58/09, <i>T. asperellum</i> URM 6997/160821, and <i>T. harzianum</i> IB 19/17) were cultivated using a culture medium designed through Plackett-Burman statistical design, followed by the characterization of the microorganisms to increase enzyme production, siderophore synthesis, and indole-3-acetic acid production. Significant enzymatic activity was observed, including chitinase (0.022 and 0.004 U·mL^− 1), β-1,3-glucanase (0.199 and 0.007 U·mL^− 1), and protease (1.45 and 1.40 U mL^− 1), as well as propagules (9.67 × 10⁷ conidia·mL^− 1, 2.56 × 10⁷ microsclerotia·mL^− 1, and 1.79 × 10⁷ chlamydospores·mL^− 1). Siderophore production reached 33.26%, and biomass was measured at 1.32&#xa0;g·100 mL^− 1. Some assays of the strains MMBF 58/09, URM 6997/160,821, and IB 19/17 demonstrated that these strains have promising potential to contribute to food safety and agricultural quality. Therefore, the observed responses emphasize the need for oriented microbial selection, supporting the development of targeted bioinputs within the framework of applied microbiology.</p>