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Meiofaunal organisms are the predominant metazoans in benthic systems and important members of the benthic food web. They are defined by mesh size and specifically by their retention on a sieve with a 44-µm mesh size. In this study, we examined the accuracy of this standard collection method by counting the number of meiobenthic individuals, life stages and nematode species in a sample. A filter cascade consisting of five different mesh sizes (41 µm, 30 µm, 20 µm, 10 µm and 1 µm) was used to fractionate a natural freshwater meiobenthic collection, and the individuals in each fraction were then counted. In line with the current definition of meiofauna, all tardigrades, microcrustaceans, chironomids and oligochaetes were retained by the largest mesh size, whereas 9% of the rotifers were first retained on the 30-µm meshes. For nematodes, 23% were not retained on the 41-µm meshes and individuals were collected even from the 1-µm fraction. With declining mesh size, the yield of retained nematodes increased, the age structure shifted to juveniles, evenness declined, and the species composition changed. As all of these findings were significant, this study therefore shows that the current definition of meiofauna is not sufficient to encompass the entire spectrum of meiofauna present in a sample and may result in misleading assessments of the diversity and composition of these organisms. We therefore propose that, especially for nematodes, a definition based on a smaller mesh size (at least 20 µm) is more appropriate.