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Agricultural development was the major contributor to South America’s designation as the continent with the highest rates of forest loss from 2000–2012. As the apex predator in the Neotropics, jaguars (Panthera onca) are dependent on forest cover but the species’ response to habitat fragmentation in heterogeneous agricultural landscapes has not been a subject of extensive research. We used occupancy as a measure of jaguar habitat use in Colombia’s middle Magdalena River valley which, as part of the intercontinental Tumbes-Chocó-Magdalena biodiversity hotspot, is exceedingly fragmented by expanding cattle pastures and oil palm plantations. We used single-season occupancy models to analyze 9 months of data (2015–2016) from 70 camera trap sites. Given the middle Magdalena’s status as a “jaguar corridor” and our possible violation of the occupancy models’ demographic closure assumption, we interpreted our results as “probability of habitat use (Ψ)” by jaguars. We measured the associations between jaguar presence and coverage of forest, oil palm, and wetlands in radii buffers of 1, 3, and 5 km around each camera trap. Our camera traps recorded 77 jaguar detections at 25 of the camera trap sites (36%) during 15,305 trap nights. The probability of detecting jaguars, given their presence at a site, was 0.28 (0.03 SE). In the top-ranked model, jaguar habitat use was positively influenced by wetland coverage (β = 7.16, 3.20 SE) and negatively influenced by cattle pastures (β = -1.40, 0.63 SE), both in the 3 km buffers. We conclude that wetlands may serve as keystone habitats for jaguars in landscapes fragmented by cattle ranches and oil palm plantations. Greater focus on wetland preservation could facilitate jaguar persistence in one of the most important yet vulnerable areas of their distribution.