PCR-based assessment of shellfish traceability and sustainability in international Mediterranean seafood markets Asmaa Galal-Khallaf a,b , Alba Ardura a , Yaisel J. Borrell a , Eva Garcia-Vazquez a,⇑ a Departamento de Biologia Funcional, Universidad de Oviedo, C/Julian Claveria s/n., 33006 Oviedo, Spain b Genetic Engineering and Molecular Biology Division, Department of Zoology, Faculty of Science, Menoufia University, Egypt article info Article history: Received 6 July 2014 Received in revised form 24 January 2016 Accepted 29 January 2016 Keywords: DNA barcoding Ecological sustainability Egyptian markets Shellfish Spanish markets abstract Two mitochondrial markers (cytochrome oxidase COI and 16S rDNA) were employed for species identi- fication of commercial shellfish from two Mediterranean countries. New COI Barcodes were generated for six species: Pleoticus robustus, Metapenaeopsis barbata, Parapenaeus fissuroides, Hymenopenaeus debilis, Metapenaeus affinis and Sepia aculeata. Biodiversity of the seafood species analyzed was greater in Egypt, with nine crustacean and two cephalopod species found compared with only three crustaceans and three cephalopods in Spain. In total, 17.2% and 15.2% products were mislabeled in Egypt and Spain, respectively. Population decline is a problem for some of the substitute species. Others were exotic and/or invasive in exporters’ regions. This study offers the first comparable study of shellfish traceability in these Mediterranean markets. The PCR-based method used in this study proved to be reliable, effective and, therefore, could be employed for routine seafood analysis. Ó 2016 Elsevier Ltd. All rights reserved. 1. Introduction Food authentication is important in preventing commercial fraud as well as health problems, such as allergies or intoxications from undeclared ingredients (Lockley & Bardsley, 2000; Ortea, Cañas, Calo-Mata, Barros-Velázquez, & Gallardo, 2010). Mislabeling occurs in products that cannot be identified because they are pro- cessed or pre-cooked. For example, shellfish are often sold in pieces (cephalopods) or without shells (mollusks, crustaceans). Because they are consumed worldwide, controls are important. Shrimps (Decapoda) are amongst the most commercially valuable shellfish accounting for more than 30% of global consumption of crus- taceans (Pérez-Farfante & Kensley, 1997). Cephalopods are also a major product with squid and cuttlefish making up most of the catch. Theoretically, using shellfish as a protein source might be more sustainable than fish because the environmental impacts of shell- fish production are relatively low (Crawford, Macleod, & Mitchell, 2003). However, shellfish consumption does generate ecological concerns. On the one hand, over-exploitation occurs in some shell- fish fisheries, such as jinga shrimp Metapenaeus affinis, which rep- resents around half (52%) of the catch in Kuwaiti waters (Mohammed, 1995). On the other hand, shellfish farming can harm local ecosystems through accidental releases of non-native farmed species. For example, the banana prawn, Fenneropenaeus merguien- sis, collected in Bay Iskenderun (southeastern Turkey), is not native and is likely to have escaped from commercial farms (Özcan, Galil, Bakír & Katag ˘an, 2006). Visual identification of the species in shellfish products is diffi- cult and, thus, tools to identify species accurately are needed urgently. Protein-based methods, such as sodium dodecylsulphate polyacrylamide gel electrophoresis (SDS–PAGE), two dimensional gel electrophoresis (2-DE), enzyme-linked immunosorbent assay (ELISA), native isoelectric focusing (IEF) and urea isoelectric focus- ing (urea IEF) have been applied to identify fish and shellfish spe- cies of commercial interest (Asensio, González, García, & Martín, 2008; Berrini, Tepedino, Borromeo, & Secchi, 2006; Mackie et al., 2000; Ortea et al., 2010). Such methods depend mainly on polypeptide targets that are unstable in processed products. In contrast, DNA is more stable, and can resist processing including high temperatures. PCR-based methods are employed increasingly to authenticate shellfish products (Espiñeira, Vieites, & Santaclara, 2010; Pascoal, Barros-Velázquez, Cepeda, Gallardo, & Calo-Mata, 2008). Several DNA regions can be used as species-specific mark- ers. For example, the mitochondrial 5 ´ region of the cytochrome oxidase subunit I gene (COI) has been reported as an effective mar- ker for shellfish identification (e.g. Haye, Segovia, Vera, Gallardo, & Gallardo-Escárate, 2012). The variable 16S rDNA region has been http://dx.doi.org/10.1016/j.foodchem.2016.01.131 0308-8146/Ó 2016 Elsevier Ltd. All rights reserved. ⇑ Corresponding author. E-mail address: egv@uniovi.es (E. Garcia-Vazquez). Food Chemistry 202 (2016) 302–308 Contents lists available at ScienceDirect Food Chemistry journal homepage: www.elsevier.com/locate/foodchem