Triclopyr-based herbicides are broadly used worldwide for site preparation and forest vegetation management. Thus, following application, these agrochemicals can inadvertently reach the aquatic ecosystems. Garlon® is one of the most popular commercial denominations of this group of herbicides, considered as highly toxic to fish, even by its manufacturer. Although DNA is frequently regarded as a target of pesticide toxicity, the genotoxic potential of Garlon® to fish remains completely unknown. Hence, the main goal of this study was to evaluate the genotoxicity of Garlon® and its active ingredient (triclopyr), clarifying the underlying mechanisms. Therefore, the comet assay, implemented as the standard procedure, with an extra step involving DNA lesion-specific repair enzymes (formamidopyrimidine DNA glycosylase and endonuclease III), was used to identify DNA damage in blood cells of Anguilla anguilla L. Short-term exposures (1 and 3 days) to Garlon® and triclopyr were carried out, adopting environmentally realistic concentrations (67.6 and 270.5 µg L−1 Garlon® and 30 and 120 µg L−1 triclopyr). The results concerning the nonspecific DNA damage proved the risk of the herbicide Garlon® and its active ingredient triclopyr in both tested concentrations and exposure lengths. In addition, the higher genotoxic potential of the formulation, in comparison with the active ingredient, was demonstrated. When the additional breaks corresponding to net enzyme-sensitive sites were considered, none of the conditions revealed significant levels of oxidative damage. This identification of the genotoxic properties of triclopyr-based herbicides to fish highlights the need to develop less hazardous formulations, as well as the adoption of mitigation measures related to the application of these agrochemicals in the framework of forestry and agriculture sustainable management. © 2014 Wiley Periodicals, Inc. Environ Toxicol 30: 1073–1081, 2015.