Understanding the factors and mechanisms shaping differences in species composition across space and time (ß-diversity) in human-modified landscapes has key ecological and applied implications. This topic is, however, challenging because landscape disturbance can promote either decreases (biotic homogenization) or increases (biotic differentiation) in ß-diversity. We assessed temporal differences in intersite ß-diversity of medium-bodied and large-bodied mammals in the fragmented Lacandona rainforest, Mexico. We hypothesized that, given the relatively short history of land-use changes in the region, and the gain and loss of some species caused by landscape spatial changes, ß-diversity would increase through time, especially its nestedness component. We estimated ß-diversity between 24 forest sites (22 forest patches and two continuous forest sites) in 2011 and 2017 to assess whether ß-diversity is decreasing or increasing in the region, and calculated its turnover and nestedness components to understand the mechanisms responsible for changes in ß-diversity, separately assessing mammal groups with different body mass, feeding guild, and habitat specialization. We then related such temporal changes in ß-diversity to temporal changes in five landscape variables (forest cover, matrix openness, number of patches, edge density and interpatch distance) to identify the landscape drivers of ß-diversity. In contrast with our expectations, ß-diversity decreased over time, suggesting an ongoing biotic homogenization process. This pattern was mostly driven by a decrease in species turnover in all mammal groups, especially in landscapes with decreasing forest cover and increasing forested matrices. Although the nestedness component showed a three-fold increase through time, species turnover was 22 and six times higher than nestedness in 2011 and 2017, respectively. The decreased turnover appears to be driven by an increase in dispersal (i.e., spillover) of native species among patches. The prevalence of species turnover over nestedness indicates that different forest sites have a fairly distinct subset of species (i.e., high complementarity in species composition). Therefore, conserving all remaining forest patches and increasing forest cover is of utmost importance to effectively maintain ß-diversity and conserve the total diversity (γ) of mammal assemblages in this Mesoamerican biodiversity hotspot.