A DNA Metabarcoding Workflow to Identify Species in Spices and Herbs.

BACKGROUND: Spices and herbs are food categories regularly cited as highly susceptible to be adulterated. To detect potential adulteration with undeclared species, DNA-based methods are considered the most suitable tools. OBJECTIVE: In this study, the performance of the ready-to-use Thermo Scientific™ NGS Food Authenticity Workflow (Thermo Fisher Scientific)-a commercial DNA metabarcoding approach-is described. The tool was further applied to analyze 272 commercial samples of spices and herbs. METHOD: Pure samples of spices and herbs were analyzed with the Thermo Scientific NGS Food Authenticity Workflow to assess its specificity, and spikings down to 1% (w/w) allowed evaluation of its sensitivity. Commercial samples, 62 and 210, were collected in Asian and European markets, respectively. RESULTS: All tested species were correctly identified often down to the species level, while spikings at 1% (w/w) confirmed a limit of detection at this level, including in complex mixtures composed of five different spices and/or herbs. The analysis of 272 commercial samples showed that 78% were compliant with the declared content, whereas the rest were shown to contain undeclared species that were in a few cases allergenic or potentially toxic. CONCLUSIONS: The Thermo Scientific NGS Food Authenticity Workflow was found to be suitable to identify food plant species in herbs and spices, not only when tested on pure samples, but also in mixtures down to 1% (w/w). The overall workflow is user-friendly and straightforward, which makes it simple to use and facilitates data interpretation. HIGHLIGHTS: The Thermo Scientific NGS Food Authenticity Workflow was found to be suitable for species identification in herbs and spices, and it allowed the detection of undeclared species in commercial samples. Its ease of use facilitates its implementation in testing laboratories.

Development of a DNA Barcoding-Like Approach to Detect Mustard Allergens in Wheat Flours.

The spread of food allergens is a topic of global importance due to its impact on public health. National and International regulations ask food producers and manufacturers to declare product compositions on the label, especially in case of processed raw materials. Wheat flour (Triticum aestivum) can be contaminated by a wide range of species belonging to the Brassicaceae in the field or during grain harvests, storage, and processing. Among them, mustards (Brassica nigra, Brassica juncea and Sinapis alba) are well known allergenic species. Often, food quality laboratories adopt an ELISA approach to detect the presence of mustard species. However, this approach shows cross-reactivity with other non-allergenic species such as Brassica napus (rapeseed). In the last few years, DNA barcoding was proposed as a valid identification method, and it is now commonly used in the authentication of food products. This study aims to set up an easy and rapid DNA-based tool to detect mustard allergenic species. DNA barcoding (matK and ITS2) and chromosome markers (A6, B, C1 genome regions) were selected, and specific primers were validated on incurred reference food matrices. The developed test was proven to be able to distinguish mustard from rapeseed and wheat, overcoming cross-reactivity with Brassica napus.

Development and evaluation of two multi-antigen serological assays for the diagnosis of bovine tuberculosis in cattle.

There is currently an increased interest in the use of serological approaches in combination with traditional cell-mediated immunity-based techniques to improve the detection of tuberculosis (TB)-infected animals. In the present study, we developed and validated two different serological TB-detection assays using four antigens, MPB70, MPB83, ESAT6 and CFP10, and the tuberculin PPDb. A conventional multi-antigen TB-ELISA method and a novel TB multiplex test, based on Luminex technology, were developed to detect antibodies to multiple antigen targets. The performance levels of the two tests were evaluated and compared using selected panels of samples having known TB states. The TB-ELISA test (containing five antigens, including PPDb) had a sensitivity (Se) of 74.2% and a specificity (Sp) of 94.9%, while the TB-Luminex test had higher Se (79.0%) and Sp (99.1%) rates even when only one reactive antigen was used to classify the test as positive. If a more restrictive criterion, requiring two positive antigens to classify the test as positive, was used, then the TB-ELISA's Sp rate increased to 99.8% but the Se decreased to 61.3%, while the TB-Luminex test's Sp rate increased to 100% but the Se decreased to 51.2%. TB-ELISA and TB-Luminex were applied to a panel of 257 sera collected from bTB-positive herds, as determined by a post-mortem inspection. They showed good performance levels, identifying 49 (80.3%) and 48 (78.7%), respectively, of 61 samples that had tested positive by the intradermal tuberculin (IDT) test and/or interferon-gamma assay. In addition, TB-ELISA and TB-Luminex were able to identify 60 and 42 samples as positive, respectively, out of the 196 samples that tested negative to IDT and interferon-gamma at the time of serum collection. Subsequent IDT tests performed after 1-2 months, confirmed the positivity of 18 samples, indicating the strategic value of having two serological assays to detect TB-infected herds that were not reactive to initial IDT testing, thereby allowing for the rapid control of outbreaks and eradication of the disease.