Is the metabarcoding ripe enough to be applied to the authentication of foodstuff of animal origin? A systematic review.

Food authentication using molecular techniques is of great importance to fight food fraud. Metabarcoding, based on the next-generation sequencing (NGS) technologies, allowing large-scale taxonomic identification of complex samples via massive parallel sequencing of fragments (called DNA barcodes) simultaneously, has become increasingly popular in many scientific fields. A systematic review to answer the question "Is the metabarcoding ripe enough to be applied to the authentication of foodstuff of animal origin?" is presented. The inclusion criteria were focused on the selection of scientific papers (SPs) only applying metabarcoding to foodstuff of animal origin collected on the market. The 23 included SPs were first analyzed with respect to the metabarcoding phases: library preparation (target genes, primer pairs, and fragment length), sequencing (NGS platforms), and final data analysis (bioinformatic pipelines). Given the importance of primer selection, the taxonomic coverage of the used primers was also evaluated. In addition, the SPs were scored based on the use of quality control measures (procedural blanks, positive controls, replicates, curated databases, and thresholds to filter the data). A lack of standardized protocols, especially with respect to the target barcode/s and the universal primer/s, and the infrequent application of the quality control measures, leads to answer that metabarcoding is not ripe enough for authenticating foodstuff of animal origin. However, the observed trend of the SP quality improvement over the years is encouraging. Concluding, a proper protocol standardization would allow a wider use of metabarcoding by both official and private laboratories, enabling this method to become the primary for the authentication of foodstuffs of animal origin.