Key-words:HER, macroinvertebrate, monitoring, source distance, watercourse Increasing the river habitat safeguard level is one of the main actions proposed by the European Water Framework Directive in the field of biomonitoring. To do so, watercourses within the same hydroecoregions (that is, homogeneous areas based on climate, geology and topography) ought to be compared. In addition, the source distance was thought to play an important role in comparing rivers and then planning monitoring activities. The purpose of this study was to evaluate if both hydroecoregion and source distance affect the response (in terms of taxa assemblage) of one of the most used group in the river monitoring activities: the benthic macroinvertebrate. Here we proposed the comparative influence of hydroecoregion and source distance on the invertebrate assemblage in Mediterranean rivers of central Italy. Our statistical outputs highlighted how macroinvertebrate differed for both hydroecoregions and source distance ranks. In particular, no differences were found among sites of different (1) source distance ranks and (2) hydroecoregion (that is, when this two descriptors were analyzed separately), while the highest difference in the macroinvertebrate assemblage was observed between the same source distance ranks of different hydroecoregions. Our results showed how the use of both hydroecoregion and source distance should be considered for planning monitoring activities to properly manage rivers and water resources.