Taxonomic studies in cycad genera using multiple approaches have refined the delimitation of many species. In the case of Dioon Lindl., a Mesoamerican genus, a reliable classification has been achieved through taxonomic work carried out since the description of the genus and pioneering studies in Mexico during the 1980–90s. Here, we describe a new species from Guerrero based on evidence collected from populations encountered during fieldwork carried out in 2019. These populations had previously been considered morphologically similar to Dioon holmgrenii De Luca, Sabato & Vázq. Torres, which has a markedly disjunct distribution in Oaxaca. After studying herbarium specimens and making extensive observations on vegetative and reproductive structures from different populations in Oaxaca, we have concluded that the disjunct populations analysed in 2019 represent a distinct and new species of Dioon, separate from the two most phenotypically similar species—namely, D. stevensonii Nic.-Mor. & Vovides and D. holmgrenii. A key to geographically proximal and morphologically similar species as well as to the other species occurring in Guerrero and Oaxaca States is also presented. The proposed new species, Dioon nuusaviorum Mart.-Domínguez, Nic.-Mor. & D.W.Stev., is endemic to Guerrero and inhabits pine and pine-oak forest. Its conservation status, assessed on the based IUCN guidelines and criteria, qualifies as Endangered.