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Abstract We investigated the impacts of resource access roads on soil enzyme activities in contrasting forested boreal peatlands (bog and fen). In August 2016, a total of 72 peat samples were collected from twelve 20 m long transects perpendicular to access roads, with a further six samples collected from undisturbed reference areas. Sampling locations represent a range in three variables associated with roads: 1) side of the road (upstream/downstream), 2) distance to a culvert (longitudinal; <2 and >20 m), and 3) distance from the road (lateral; 2, 6, and 20 m). Phenol oxidase and hydrolase (glucosidase, sulfatase, xylosidase, glucosaminidase, and phosphatase) enzyme activities were determined for each sample, in addition to water table depth, phenolic concentration, pH, and peat temperature. The average hydrolase activities in the fen were ~four times higher than in the bog. At the bog, the water table depth, phenolic concentration, pH and the activities of phenol oxidase, sulfatase, glucosidase, xylosidase and glucosaminidase were all significantly influenced by one or more road associated factors. The highest enzyme activities in the bog occurred on the downstream side of the road at plots located far from the culvert. In contrast, the flow of water in the fen was not perpendicular to the road. Consequently, no significant variations in water table depth, phenolic concentration, pH or enzyme activity were found with respect to road associated factors. Results indicate that road crossings in boreal peatlands can indirectly alter enzyme activities, likely as part of a causal chain following changes to hydrology and redox conditions. Two of six investigated enzymes had significantly higher activities in the road disturbed areas compared to undisturbed areas, suggesting ultimately that roads may enhance organic matter decomposition rates. However, adequate hydrologic connections through culverts and road construction parallel to the water flow can minimize the road-induced impacts. Graphical abstract Unlabelled Image Highlights • Roads alter local ecohydrology and hydrochemistry that may impact enzyme activity. • We observed changes in water table and enzyme activity in a road-fragmented bog. • Water flow parallel to the road in the fen limited road impact on enzyme activity. [ABSTRACT FROM AUTHOR]