Toward on-site food authentication using nanopore sequencing

Food fraud is a major concern as it has economic impact and affects consumer confidence (Grunert, 2002, Spink and Moyer, 2011). Deliberate adulteration of a product for financial profit may have far-reaching consequences with for the food industry, as observed with the horsemeat scandal in 2013 (O’Mahony, 2013). This scandal raised questions about the effectiveness of controls along the food chain by businesses and governmental authorities. In case of a food safety risk or indications of fraudulent raw materials food inspection services are required to act fast. Incidents like the horsemeat scandal call for methods that allow the rapid assessment of the authenticity and quality of raw materials and food commodities. A promising fast detection technology is the MinION DNA sequencer from Oxford Nanopore Technologies (ONT) which is smaller than a smartphone and able to produce data within minutes. The sequencer with low purchasing price is able to sequence individual DNA molecules as they drive through biological nanopores by an applied electrical field (Loman & Watson, 2015). Currently, MinION sequencing shows a higher error rate compared to traditional Next-Generations Sequencing (NGS) equipment such as Illumina MiSeq technology (Judge et al., 2015, Laver et al., 2015). Nonetheless, the technology is already used for fast medical applications, even in the field, for instance for the mobile real-time surveillance of Zika virus in Brazil (Faria et al., 2016). Recently, Menegon et al. (2017) showed that DNA barcoding and MinION sequencing can also be used in on-site biodiversity assessments, despite the reported limitations of the technology. So far, no food-related applications of the MinION technology have been reported. The technology has the potential to screen complex food products quickly and comprehensively for a multitude of DNA-based markers. Here, we assessed the potential of MinION-based DNA metabarcoding as a system that may lead to fast food authentication, without the need for a highly priced sequencing platform. Two experimental fish mixtures were prepared and metabarcoded, after which the performance, i.e. the number and level of correctly identified fish species, of MinION sequencing was compared to the Illumina MiSeq technology. Fish was selected as example of food products vulnerable to food fraud (Everstine et al., 2013, Ogden, 2008, Warner et al., 2013). The mitochondrial DNA barcodes cytochrome oxidase subunit I gene (COI) (Ward, Hanner, & Hebert, 2009) and cytochrome b (cytb) gene (Griffiths et al., 2014) were selected as they have proven to be effective in identifying fish products (Cawthorn et al., 2012, Hanner et al., 2011, Logan et al., 2008, Miller and Mariani, 2010, Wong and Hanner, 2008). In this paper, the applicability of the MinION device for effective metabarcoding has been assessed. Furthermore, in order to process the typical quality of MinION reads a dedicated bioinformatics workflow was developed and the degree of run-to-run contamination was determined.