The Cayo archaeological deposits documenting the contact period between European and Amerindian populations before the colonization of the Lesser Antilles are rare. Among the few sites dated to this period, the site of Roseau, excavated by Gérard Richard in the early 2000s, remains poorly investigated, especially from a stratigraphic and zooarchaeological point of view. In this study, we performed 20 new radiocarbon datings on the collagen content of rodent dentin along with new studies of the ceramic and faunal assemblages of the site. The results indicate that the archaeological assemblage is mainly associated to the Late Troumassoid, with some admixture of Early Troumassoid and Cayo elements represented in various proportions in the whole stratigraphy. Despite these elements, a chronological trend appears preserved in the stratigraphy allowing for a research of behavioral change in subsistence pattern through time. Vertebrate faunal data indicate that the two Troumassoid layers we recognized document different subsistence strategies. Indeed, the earliest inhabitants of the site were more focused on the exploitation of aquatic resources than the latest ones who more significantly exploited terrestrial fauna, especially rodents and iguanas. Carbon and oxygen stable isotope analyses performed on fossil tooth enamel of rodents (Antillomys rayi) suggest that Amerindians hunted them in both dry and wet environments. Paleontological approaches also led to the discovery of several now-extinct taxa that were part of the Amerindian diet. Our investigations point to a strong chronological variability of subsistence behaviors and improve our understanding of the accumulation history of the site.