Landuse changes might alter N cycling in tropical aquatic ecosystems, but understanding of N cycling in tropical streams is limited. We measured actual and potential denitrification rates during the dry season in Río Las Marías, a 4^th-order Andean piedmont stream in Venezuela. Our objectives were to describe spatial and temporal variation in denitrification, quantify the effects of nutrient availability and substratum particle size on denitrification, and explore potential effects of anthropogenic sedimentation on denitrification. In 4 experiments, actual and potential denitrification rates ranged from 0 to 160 and from 0 to 740 μg N₂O-N m⁻² h⁻¹, respectively. Rates were distributed approximately log-normally because of spatial variation. During a 1-mo period, actual denitrification rates decreased exponentially from 37 ± 39 to 5 ± 7 μg N₂O-N m⁻² h⁻¹ (mean ± SD), probably because of a decline in water-column NO₃-N concentration from 41 ± 14 to 12 ± 3 μg NO₃-N/L. The texture (particle size) of stream substrata markedly affected denitrification rates. Actual rates were low in cobble, gravel, and fine sediments (<5 mm), but in fine sediments, rates increased in response to addition of excess NO₃-N and organic C. In a 3-km stream reach, actual (but not potential) denitrification rates increased with the proportion of fine sediments (<2 mm) in mixed substrata. This increase was nonlinear, and the threshold value occurred at 37% fine particles, above which actual denitrification rates were almost always high. An experiment simulating the effects of anthropogenic sedimentation showed that topsoil inputs resulted in denitrification rates ∼8× higher than rates in trials where excess NO₃-N and organic C were supplied. Denitrification is a small but potentially significant sink for available N in this N-limited system. Anthropogenic sedimentation associated with landuse change might significantly increase denitrification rates in streams.