The relationship between humans and nature is defined by culture. Accordingly, the use, conceptions, and perceptions of resources differ among cultural groups, even among those inhabiting the same region or those who come into contact with the same biota. In particular, mushrooms evoke a wide range of sentiments. During ethnobiological tours in Mexico, semi-structured interviews were carried out with 37 individuals of each community, from ten Wixarika and mestizo communities, living in the same locality and sharing similar resources, in the municipality of Villa Guerrero in northern Jalisco, Mexico. Furthermore, informal interviews with four Wixarika and five mestizo key informants were conducted. The topics treated were regarding the traditional nomenclature and classification, uses, and knowledge of mushrooms and related practices. Wixarika names of 37 mushroom species with edible, medicinal, and recreational uses were recorded. In addition, the Wixaritari were found to associate toxic mushrooms with the divine, as evidenced by one case of the use of mushrooms as a hierophanic agent. Each culture’s knowledge of the phenology and ecology of mushrooms was recorded in addition to data highlighting the cultural exchange between the Wixaritari and mestizos. However, a loss in the knowledge and practices concerning mushrooms was observed as a result of social changes. Even so, both cultures prefer mushrooms to other foods, including meat, especially Volvariella bombycina and Pleurotus djamor.