The catfish Genidens barbus is a commercial species from South America. The aim of the present study was to examine the inter-annual variability in estuary-specific chemical signatures of otolith cores (Ba:Ca, Mg:Ca, Mn:Ca, Sr:Ca and Zn:Ca ratios) for three estuaries from Argentina and Brazil where adults were collected over multiple years. Secondarily, we evaluated whether the percentages of classification of individuals to their natal origin place are affected by the grouping of several cohorts. Most element:Ca ratios were not significantly different among year cohorts. Results from PERMANOVA revealed significant differences in the multi-element signatures of the otolith core between cohorts for the Plata River estuary (PR) (P = 0.006) and the Patos Lagoon (PL) (P = 0.03), while no significant differences (P = 0.9) were found for Paranaguá Bay (PB). The percentages of spatial classification (discriminant function analyses) decreased to between 15.5 and 25% for PR and PL when cohorts were grouped. This work makes it clear that the temporal variation in the chemical signature of the adult catfish otolith core can greatly affect the percentages of spatial classification.