Microhabitat use in the endangered cyprinid species spirlin Alburnoides bipunctatus and accompanying species was examined in three water courses of Slovakia to determine the species’ environmental requirements as a basis for informing conservation policy and management. In all three rivers, water velocity, water depth and substratum character were central features of spirlin microhabitat use, regardless of year or season of sampling, with only limited variation in microhabitat associations as a function of time of day. Clear differences in microhabitat use and intra-specific associations during development were observed in two of the rivers. In particular, a shift in velocity preference towards faster flowing waters appears characteristic of spirlin during their larval and juvenile development, and possibly also that of gudgeon Gobio gobio, European minnow Phoxinus phoxinus, and chub Leuciscus cephalus, which are all species that may be significantly associated with young spirlin. Disproportionate use of deeper waters tended to increase with age in spirlin, and in gudgeon and barbel Barbus barbus, but spirlin preference for substratum was less uniform, with affinities ranging from indifference to strong preference. This contrasts the clear preferences for sand in gudgeon and for cobbles in European bullhead Cottus gobio. Of particular importance to young-of-the-year (YOY) spirlin are lentic zones with some sort of ligneous debris – habitat also used by YOY gudgeon and minnow. Contrary to previous reports elsewhere, spirlin did not avoid in-stream vegetation where present, and in one river it was preferred by YOY and 1+ spirlin. To avoid declines in spirlin and accompanying stream-dwelling species, such as reported elsewhere in Europe, river management in water courses such as these should be limited to the rehabilitation of regulated sections to achieve a natural, heterogeneous channel character.