Roots can recruit beneficial microorganisms to suppress plant pathogens. However, conventional and organic practices differently shape the soil microbiome and consequently the root protection. Here, we investigated the suppressive activity of soil microbiome against the root-knot nematode (RKN) Meloidogyne incognita in horticultural areas under organic or conventional practices and the microbiome profiles in non-inoculated (RKN-absent) and inoculated (RKN-present) rhizospheres. Soils were collected from neighboring areas under long-term conventional or organic practices, but physicochemically similar. After a set of bio-tests in autoclaved and non-autoclaved soils, we concluded that the soil suppressiveness was of biological origin. However, plant growth, RKN suppression and defense induction were higher in organic soils. Also, RKN was highly suppressed when the organic soil microbiome was transferred to infested substrates. We used Illumina MiSeq platform to determine bacterial and fungal profiles in organic and conventional tomato rhizospheres, inoculated or not with M. incognita. Our data suggest that despite the higher bacterial abundance in the conventional RKN-absent rhizosphere, the organic RKN-present rhizosphere recruited more efficiently antagonistic bacteria and fungi. Microbiome α-diversity increased in the organic RKN-present rhizosphere. Antagonistic bacterial genera Pseudomonas, Serratia, Bradyrhizobium, Burkholderia and Azospirillum and fungal genera Beauveria, Clonostachys, Metarhizium, Purpureocillium and Arthrobotrys were highly abundant only in the organic RKN-present rhizosphere. This work suggests the organic and conventional practices interfere in the rhizosphere microbiome composition and consequently in the suppression of RKN, where organic practices intensified the assembly of bacterial and fungal antagonists in the presence of M. incognita.