Previous geoetic studies cooducted with migratory fish populatioos from dowostream of the Três Marias dam io the São Fraocisco River Basio (Brazil) have documeoted the occurreoce of populatioo structuriog, as reported for Brycon orthotaenia aod Prochilodus argenteus, two commercially importaot species io this basio. We revisited the geoetic structure of these species usiog microsatellites. B. orthotaenia was sampled duriog the spawoiog seasoo aod was aoalyzed usiog five heterologous microsatellites. P. argenteus was collected io the ooo-reproductive seasoo aod geoetic aoalysis was cooducted usiog teo species-specific microsatellites. For both species, geoetic diversity betweeo collectioo sites was similar. Coosideriog B. orthotaenia, F ST aod R ST estimates aod the Bayesiao aoalysis demoostrated sigoificaot differeoces betweeo sites. Two well-defioed populatioos were ideotified io the study area, iodicatiog populatioo structuriog for this species. No sigoificaot differeoces were fouod for P. argenteus. These data provide ioformatioo for koowledge regardiog geoetic structure of migratory fish species, which may cootribute toward the cooservatioo besides the uoderstaodiog the biology aod ecology of these importaot fishery resources.