ABSTRACT The greatest diversity of culturally relevant fungi in Mexico has been recorded in temperate forests. Likewise, it has been proven that people who live in such environments possess greater ethnomycological knowledge, compared with people in the lowlands. In this study, we investigated whether the uses and perceptions of fungi were different between people living near forests in highlands and those living in lowlands near grasslands and subtropical scrublands. We selected seven communities from the municipality of Tlaltenango, Zacatecas, Mexico, where we conducted guided tours, 35 semistructured interviews, and free lists to 420 people. Since highlands surrounded by temperate forests give rise to high species diversity, we expected that the inhabitants, in comparison with those of the lowlands where there is less diversity, would recognize a larger number of fungi. To test this hypothesis, we employed discriminant function analysis, principal coordinate analysis, Mann-Whitney U test, and linear regression. The cultural importance of each species was calculated based on the frequency with which it was mentioned and the first principal coordinate. Although the highlanders have a deeper knowledge of mushrooms, lowlanders and highlanders had similar fungal backgrounds and preferred the same species, regardless of the fungi surrounding their territory. The lack of differences among communities was due in part to the fact that the most culturally important species were those that grew in grasslands and subtropical scrub areas. Agaricus campestris was the most culturally prevalent and only commercialized species both in the highlands and lowlands, followed by Pleurotus djamor and Volvariella bombycina. None of the mushrooms growing in the pine-oak forest had a high cultural importance value, even for the communities living in the vicinity of this forest, forcing them to travel long distances to collect Agaricus campestris. Further investigations are needed in order to assess the relevance of sociocultural factors, and their potential influence in the preference for particular mushrooms in this region.