* The biodiversity of aquatic ecosystems is under threat and there is an urgent need to quantify the various facets of biodiversity to assess the conservation value of freshwater ecosystems. The effects of taxonomic relatedness have so far not been taken into account in biodiversity assessments of lake macrophytes. * We therefore tested the response of species richness and average taxonomic distinctness (AvTD) of aquatic macrophytes along environmental gradients using linear regression models and Bayesian Information Criterion variable selection method. We selected data from four regions, each with 50–60 lakes, situated in northern Europe (Finland and Sweden) and northern America (Minnesota and Wisconsin). We separately studied all macrophyte species, hydrophytes and helophytes. * Species richness and AvTD of aquatic macrophytes were generally negatively related in all regions, although it was not statistically significant. Both biodiversity measures responded to environmental gradients to various degrees among the studied macrophyte groups and regions. Species richness was best explained by alkalinity and lake area in Finland, by elevation, annual mean temperature and total phosphorus in Minnesota, and by alkalinity in Wisconsin. AvTD was best explained by alkalinity, annual mean temperature and total phosphorus in Finland and by alkalinity in Wisconsin. Very weak relationships were found in Sweden. * Our findings strongly suggest that complementary indices are needed to indicate more comprehensively the effects of environmental conditions on freshwater biodiversity. Species richness was found to be a better measure than AvTD to account for conservation value in freshwaters. However, further research is required to evaluate the usefulness of AvTD to indicate conservation value (e.g. randomisation tests), because alternative measures are clearly needed for those freshwater taxa lacking complete information on true phylogenetic diversity.