DNA and Mini-DNA barcoding for the identification of Porgies species (family Sparidae) of commercial interest on the international market

Abstract

a b s t r a c tThe morphological similarity among Sparidae species, which are characterized by a different market price, represents a serious problem for their trade and for stock management, since it encourages frauds for substitution. The most accredited morphological method for their identification is based on the dental-plate, but this approach is not simple and cannot be used for prepared products. When molecular methods are used the DNA degradation induced by cooking is the main drawback. In this work, we collected 314 reference tissues belonging to 75 Sparidae species and we produced a dataset of full (FDB) and mini-barcode (MDB) reference sequences starting from DNA extracted from fresh and ethanolpreserved tissues using universal primes. Moreover, some fresh samples were cooked. The FDB was successfully amplified in 91% (fresh), 50% (cooked) and 81% (ethanol-preserved) samples, while the amplification rates of the MDB were considerably higher in case of cooked (100%) and ethanol-preserved (94%) samples. The same primers were used for the amplification of the DNA obtained from 58 market samples (MS). All the DNA barcodes were compared with BOLD and GenBank using IDs and BLAST analysis. FDB was able to provide unambiguous species-level identifications for 53 (78%) and 44 (64.7%) reference samples analyzed on BOLD and GenBank, respectively. The Mini-DNA barcode (MDB) showed a lower discriminating power with 32 (45.7%) and 29 (41.4%) sequences unambiguously matched to a species on BOLD and GenBank. However, the MDB allowed to identify all the reference sequences as belonging to the Sparidae family. FDB and MDB showed a similar performance in analyzing the MS, allowing to highlight 21 (38%) mislabeled MS. Our study, while confirming the FDB as a reliable tool for fish authentication, proposes the MDB as a promising tool to recover molecular information in case of cooked products.

Fish species identification in canned pet food by BLAST and Forensically Informative Nucleotide Sequencing (FINS) analysis of short fragments of the mitochondrial 16s ribosomal RNA gene (16S rRNA)

“…All the fish samples were stored at À20 C until total DNA extraction, which was performed according to the protocol proposed by Armani, Guardone, et al (2014); Armani, Tinacci, et al (2014), starting from a whole specimen in case of M and from 100 mg of tissue in case of F or RS. The DNA concentration and purity were assessed by evaluating the absorbance at 260 nm and the ratios A260/280 and A260/230 nm using a NanoDrop ND-1000 spectrophotometer (NanoDrop Technologies, Wilmington, DE, US).…”

“…Unfortunately, all these primers target a fragment of 700 bp, and are not suitable for the analysis of processed fish products because of the marked DNA degradation (Armani et al, 2013;Armani, Guardone, et al, 2014;Armani, Tinacci, et al, 2014). On the other hand, the high level of sequence variation of the cytb gene makes difficult to locate conserved areas to design universal primers for the amplification of short gene fragments (Zhang & Hanner, 2012).…”

DNA barcoding reveals commercial and health issues in ethnic seafood sold on the Italian market

“…Mislabeling is very frequent in the fishery supply chain, where it occurs in different forms and at any stage (Armani et al, 2015;Carvalho, Palhares, Drummond, & Frigo, 2015;Cawthorn, Steinman, & Witthuhn, 2012). A part of mislabeling is probably unintentional due to the fact that different species can be referred to by the same names in different regions and due to the lack of specific denominations, especially for new exotic species Lamendin, Miller, & Ward, 2015).…”

“…This DNA region usually shows a greater interspecific than intraspecific variation, allowing efficient discrimination among species (Hebert, Ratnasingham & de Waard, 2003). In addition, mini DNA barcoding (139 bp) has been successfully used as an alternative approach for species identification in case of processed products (Armani et al, 2015). In fact, the amplification of a shorter region could represent the only chance to obtain molecular information from products containing degraded DNA.…”

Overview on Untargeted Methods to Combat Food Frauds: A Focus on Fishery Products

“…More recently, a study based on the use of pyrosequencing as a rapid fish tool for species identification has been published [101]. DNA barcoding has also been widely used but especially for species identification [17][18][19]. A recent study based on the DNA barcoding approach was applied to reveal the incorrect labeling of imported fish products in China, amplifying the mitochondrial cytochrome C oxidase 1 (COI) gene of the target fish to demonstrate the correspondence to the morphological identification [20].…”