Horsegram (Macrotyloma uniflorum) (Lam.) is an economically important medicinal plant. The plant has been used for its antiurolithiatic property although it has other medicinal uses. Plant can establish mycorrhizal connections and rhizobial nodules because it belongs to the Leguminosae family. The main objective of the present research was to investigate the biodiversity of Arbuscular Mycorrhizal Fungi that are associated with the Horsegram plant grown in midhill region of Mandi district, Himachal Pradesh. Roots and soil samples were collected from Horsegram plants at the seedling, flowering, and maturity stages for AMF analysis and spore evaluation. The objective of this study was to extract, characterise, and classified vesicular arbuscular mycorrhizal fungal spores from root-adhering soil samples of conventional horsegram crop plants. The research was carried out to study the association of arbuscular mycorrhizal fungal spores with the roots and rhizosphere of the Horsegram plant.Total 33 species of VAM fungal spores were isolated belonging to 6 genera (Acaulospora, Glomus, Gigaspora, Scutellospora, Septoglomus and Pacisphora ) from the plant. The dominant genus was Acaulospora and Glomus. Analysis of root samples under a microscope showed varying levels of arbuscular mycorrhizal fungal colonisation, during this investigation, the qualitative characteristics of the horsegram plant were also examined.