Peru is a major fishing country and has a high consumption of fish per capita of approximately 25.4 kg/person. The gastronomic boom of the Peruvian seafood business has been increasing steadily, mostly led by seafood restaurants called cevicherias for the prevalence of their main dish “ceviche”. We investigated the incidence of seafood fraud (i.e. mislabeling and substitution) in restaurants, supermarkets, and fish retail markets from September 2017 and October 2018, in Lima, the capital of Peru. We conducted DNA barcode sequencing of 364 samples collected from fresh and frozen fish fillets and ready-to-eat dishes (i.e. ceviche, sushi, and sashimi) and compared the expected species (from sale name) to observed species (from genetic identification). We found incidences of seafood fraud in 43% of samples collected. Restaurants exhibited a significantly higher incidence (61%) compared to supermarkets. Within restaurants, 71% of samples collected in cevicherias and 47% from sushi-bars exhibited mislabeling/substitution. For cevicherias, mislabeling/substitution was inversely associated to the cost of the dish. A small number of species were commonly used as substitutes, including species without fishery management plans. Also found in the samples were threatened species, notably: Anguilla anguilla and Thunnus thynnus. Our results revealed that seafood fraud, whether as mislabeling or as substitution, occurs in Lima and increases along the supply chain, it is influenced by product price, and includes threatened species. But this is a first snapshot. We recommend the implementation of an official and regulated list of commercial fish products and continuous monitoring at points of sale to understand the prevalence of seafood fraud over large periods of time and in other cities with high consumption of seafood products.