Both forest fragmentation and overhunting have profound effects on the structure of large-vertebrate assemblages in neotropical forests. However, the long-term value of habitat fragments for forest mammals remains poorly understood and few regional scale studies have replicated sampling across spatially independent landscapes. Here, we assess the species occupancy and abundance of midsized to large-bodied mammals within three neighbouring Amazonian forest landscapes varying widely in extent of forest cover. One of these consisted of forest fragments surrounded by semi-natural scrub savannahs that had been occupied by paleoindian populations for at least 7,000 years, whereas forest cover in the other two landscapes was either variegated or continuous. Data on species occurrence and abundance from diurnal and nocturnal line-transect surveys and local interviews in each landscape were used to examine the effects of forest cover and hunting pressure on mammal persistence within forest patches. The extent of forest cover was a key determinant of species persistence across the three landscapes, but populations of large-bodied species were either reduced or driven to local extinction by hunting even in the most forested and least fragmented landscape. Many game and non-game species persisted in forest isolates, even though, individually, these were likely too small to support viable populations. This study indicates that even small, long-term forest fragments may retain significant conservation value if they can be managed within the context of enhanced connectivity across wider fragmented landscapes.