Medicinal plants maintain structures and diversities of bacteria, fungi, and arbuscular mycorrhizal fungi (AMF) that can interact to promote growth and therapeutic properties. Therefore, the purpose of this research was to evaluate the microbiome of Lippia alba and Petiveria alliacea, species known for their high potential for medicinal benefits in Colombia. To achieve this, rhizosphere soils and roots were sampled from five departments in Colombia: Boyacá, Cundinamarca, Tolima, Putumayo, and Valle del Cauca. The results revealed that the dominant bacterial groups in both plants were primarily Proteobacteria, Acidobacteriota, and Actinobacteriota, with the first phylum showing the highest number of differentially abundant genera between the sampling points. In fungi, Ascomycota tended to dominate in most of the sampled locations, while Mortierellomycota was particularly abundant in roots of P. alliacea in Valle. Furthermore, the study of AMF indicated differentiation in the colonization for both plants, with the genera Glomus and Paraglomus being predominant. Differences in the Shannon diversity index were recorded between sampling types within these sampling points, possibly influenced by local and environmental factors. Our findings reveal that the microbiomes of both medicinal plants exhibit distinct community assemblies, which could be a significant factor for their future therapeutic use.