Arbuscular mycorrhizal fungi (AMF) have been scarcely documented in relation to semiarid agroforestry systems. The "milpa-chichipera" ethno-agroforestry system (EAFS) in the semiarid Tehuac & aacute;n-Cuicatl & aacute;n Valley, Mexico, results from the conversion of xeric scrub dominated by Polaskia chichipe (Cactaceae). This EAFS promotes conservation of native plants linked to crops as an alternative to monocropping agriculture. We assessed how this EAFS impacts AMF communities, compared to preserved scrub and agricultural fields, considering soil features and rainfall seasonality. Soil samples were collected in the dry and rainy seasons at a depth of 15 cm within five plots per land-use type: i) preserved scrub, ii) vegetation patches and open areas within the EAFS, and iii) agricultural fields with monocropping. A total of 58 AMF species were recovered belonging to 17 genera and 10 families. AMF spore abundance, species richness, and diversity (alpha and beta) were enhanced by the preserved scrub and vegetation patches within the EAFS as compared to open areas and agricultural fields. Intensive agriculture modified the assembly of AMF communities at the genera and species levels, substantially depleting the spore abundance, species richness, and diversity of these fungi. In addition, all land-use types exhibited a particular AMF composition varying seasonally according to their native host plants and edaphic features. Our findings indicate that the "milpa-chichipera" EAFS, similarly to the preserved scrub, represents a mycorrhizal hotspot that may be buffering the impacts of land-use change to intensive agriculture in AMF communities.