Biological control is an environmentally safe alternative technique for management of plant pathogenic fungi. This work aims to identify and characterize potential bioagents prevalent in soybean soil. A total of 27 fungal isolates were recovered from soybean plants exhibiting typical wilt and root rot symptoms. Pathogenicity of each isolate was examined on soybean seedlings, and the most pathogenic isolates, were selected. Those isolates were identified based on their microscopic features as Fusarium oxysporum Schlecht, F. solani Mart and Rhizoctonia solani Kuhn. The identity of each isolate was confirmed by molecular phylogeny based on internal transcribed spacer (ITS) sequence. Additional 31 microorganisms of the population within soybean rhizosphere were isolated and their potential antagonistic ability was characterized in vitro and in vivo. A collection of three fungi, four bacteria, one actinomycete and one yeast isolate showed significant reduction of mycelial growth of the tested fungal pathogens. The same potential bioagents allocate significant decrease in disease severity percentage of wilt-root rot complex disease under greenhouse and open field conditions compared with mycorrhizae and Rhizolex-T. Potential mechanisms by which the bioagents suppressed soybean wilt-root rot complex disease, were addressed. Application with bioagents induced expression of peroxidase, polyphenol oxidase, chitinase, β-1,3-gluctronase, total carbohydrates and total phenols in treated soybeans, which significantly contribute to plant disease resistance. Quality parameters of soybean seeds from bioagent-treated plants showed favorable elevation in total proteins and oils within the seeds.