The seasonal colonization by benthic macrofauna was investigated using implanted substrate trays in a second order Croatian stream (Veliki Potok) during 1998 and 1999. During each season, the composition of substrates was slightly modified by the accumulation of sand and silt. The trays also had high retention, especially during spring when CPOM dominated the accumulated POM. FPOM was dominant during the other seasons. The MacArthur-Wilson equilibrium model described the colonization during each season by revealing the differences in both the colonization rate and the number of colonizing taxa, suggesting there were seasonal equilibrium values in examined macrozoobenthos parameters. Colonization was fastest during autumn and summer and slowest during spring, probably resulting from important physical parameters, such as temperature, but also influenced by macrofauna life histories. Colonization in spring is suggested to be complex due to insect emergence patterns and high fluctuations in POM. Different seasonal colonization potentials of some abundant taxa were evident from their colonization curves. Overall, shredders dominated functional organization during spring and summer, while scrapers dominated assemblages in autumn and winter, suggesting a seasonal shift in the relative importance among trophic groups. Shredders and collector-gatherers were initial colonizers, followed by collector-filterers, and then predators and scrapers. Accumulated organic matter had a negative effect on the colonization by collector-filterers during winter and summer, while collector-gatherers responded positively to UPOM quantity during autumn. Predators seemed to respond to prey diversity (scrapers in particular) in most seasons.