While fish species mislabelling has emerged as a global problem, the tracking of improvements or deteriorations in seafood trading practices is challenging without a consistent basis for monitoring. The aim of this study was to develop a robust, repeatable species authentication protocol that could be used to benchmark the current and future incidences of fish mislabelling in South Africa. Using this approach, 149 fish samples collected from restaurants and retailers in three provinces (KwaZulu-Natal, Western Cape and Gauteng) were identified using DNA barcoding, supplemented in certain cases with mitochondrial control region sequencing. Overall, 18% of samples were incorrectly described in terms of species, with similar misrepresentation rates in restaurants (18%) and retail outlets (19%). While there appears to be some improvement in the transparency of local seafood marketing compared to previous studies, the results remain of concern and signal the need for enhanced seafood labelling regulations, monitoring and law enforcement.