The Neotropical otter, Lontra longicaudis (Mustelidae) is a semi-aquatic mustelid that exploits a variety of freshwater habitats. To understand the relative influence of human activities and environmental factors affecting its distribution and habitat use, we conducted systematic, seasonal surveys of otter signs along the middle Guamá River, in Pará state in the eastern Brazilian Amazon. We applied generalized additive models to compare distribution of otters along the river with data collected on environmental factors (landcover type derived from satellite imagery, and in situ measurements of physicochemical water characteristics) and anthropogenic factors (fishing gear in the river and human habitation along the river). Most otter signs (indicators of otter habitat use) occurred along the shoreline of the main river channel during the dry season; we observed fewer signs during peak flow, probably because the shoreline and floodplain are flooded, which hid signs and made access to the floodplain difficult. The best-fit model included variables for proportion of forest, presence of fishing gear and boats, bank steepness, and presence of rock formations and sand banks. Otter occurrence was negatively related to forested area and positively related to the presence of fishing gear and boats. Otters are likely attracted to fish trapped in fishing gear because they can easily predate on the trapped fish.