Core Outcome Set for IgE-mediated food allergy clinical trials and observational studies of interventions: International Delphi consensus study 'COMFA'.

BACKGROUND: IgE-mediated food allergy (FA) is a global health concern with substantial individual and societal implications. While diverse intervention strategies have been researched, inconsistencies in reported outcomes limit evaluations of FA treatments. To streamline evaluations and promote consistent reporting, the Core Outcome Measures for Food Allergy (COMFA) initiative aimed to establish a Core Outcome Set (COS) for FA clinical trials and observational studies of interventions. METHODS: The project involved a review of published clinical trials, trial protocols and qualitative literature. Outcomes found as a result of review were categorized and classified, informing a two-round online-modified Delphi process followed by hybrid consensus meeting to finalize the COS. RESULTS: The literature review, taxonomy mapping and iterative discussions with diverse COMFA group yielded an initial list of 39 outcomes. The iterative online and in-person meetings reduced the list to 13 outcomes for voting in the formal Delphi process. One more outcome was added based on participant suggestions after the first Delphi round. A total of 778 participants from 52 countries participated, with 442 participating in both Delphi rounds. No outcome met a priori criteria for inclusion, and one was excluded as a result of the Delphi. Thirteen outcomes were brought to the hybrid consensus meeting as a result of Delphi and two outcomes, 'allergic symptoms' and 'quality of life' achieved consensus for inclusion as 'core' outcomes. CONCLUSION: In addition to the mandatory reporting of adverse events for FA clinical trials or observational studies of interventions, allergic symptoms and quality of life should be measured as core outcomes. Future work by COMFA will define how best to measure these core outcomes.

Correction: Exploiting Locusta migratoria as a source of bioactive peptides with anti-fibrosis properties using an in silico approach.

Correction for 'Exploiting Locusta migratoria as a source of bioactive peptides with anti-fibrosis properties using an in silico approach' by Carla S. S. Teixeira et al., Food Funct., 2024, 15, 493-502, https://doi.org/10.1039/D3FO04246D.

First nanoplate digital PCR method to trace allergenic foods: Improved sensitivity for the detection of sesame.

Sesame (Sesamum indicum L.) is an important allergenic food whose presence can be the cause of severe allergic reactions in sensitised individuals. In this work, nanoplate digital PCR (ndPCR) was used to develop two methods to detect trace amounts of sesame in processed foods and compared with previously proposed real-time PCR assays. Two independent ndPCR approaches were successfully advanced, achieving sensitivities of 5 and 0.1 mg/kg of sesame in dough/biscuits, targeting the CO6b-1 and ITS regions, respectively. The sensitivity using both targets was improved by one order of magnitude comparing with real-time PCR and was not affected by food processing. CO6b-1 system was not influenced by food matrix, exhibiting similar performance regardless the use of complex matrix extracts or serial diluted DNA. Herein, ndPCR was proposed for the first time for the detection of allergenic foods with the advantage of providing better performance than real-time PCR regarding sensitivity and robustness.

Electrochemical immunosensor for point-of-care detection of soybean Gly m TI allergen in foods.

Soybean is a legume with high technological functionality, commonly used by the food industry as an ingredient in different products. However, soybean is an allergenic food whose undeclared presence in processed foods may represent a public health risk. In this work, it was developed an efficient electrochemical immunosensor, targeting the soybean trypsin inhibitor (Gly m TI) allergen using commercial anti-Gly m TI IgG, aiming at detecting/quantifying minute amounts of soybean in different food formulations. For this purpose, model mixtures of different foods (sausages, cooked-hams, biscuits) were prepared to contain known amounts of soybean protein isolate (100,000-0.1 mg kg-1) and submitted to specific thermal treatments (autoclaving, oven-cooking, baking). The electrochemical immunosensor allowed quantifying down to 0.1 mg kg-1 of soybean in the three food matrices, raw and processed (0.0012 mg of Gly m TI/kg of matrix). Accordingly, the immunosensor is suitable for detecting traces of soybean in raw, processed, and complex foods, thus protecting 99 % of soybean-allergic patients.

Exploiting Locusta migratoria as a source of bioactive peptides with anti-fibrosis properties using an in silico approach.

Edible insects have been proposed as an environmentally and economically sustainable source of protein, and are considered as an alternative food, especially to meat. The migratory locust, Locusta migratoria, is an edible species authorised by the European Union as a novel food. In addition to their nutritional value, edible insects are also sources of bioactive compounds. This study used an in silico approach to simulate the gastrointestinal digestion of selected L. migratoria proteins and posteriorly identify peptides capable of selectively inhibiting the N-subunit of the somatic angiotensin-I converting enzyme (sACE). The application of the molecular docking protocol enabled the identification of three peptides, namely TCDSL, IDCSR and EAEEGQF, which were predicted to act as potential selective inhibitors of the sACE N-domain and, therefore, possess bioactivity against cardiac and pulmonary fibrosis.

Mass spectrometry-based approaches to assess the botanical authenticity of dietary supplements.

Dietary supplements are legally considered foods despite frequently including medicinal plants as ingredients. Currently, the consumption of herbal dietary supplements, also known as plant food supplements (PFS), is increasing worldwide and some raw botanicals, highly demanded due to their popularity, extensive use, and/or well-established pharmacological effects, have been attaining high prices in the international markets. Therefore, botanical adulteration for profit increase can occur along the whole PFS industry chain, from raw botanicals to plant extracts, until final PFS. Besides the substitution of high-value species, unintentional mislabeling can happen in morphologically similar species. Both cases represent a health risk for consumers, prompting the development of numerous works to access botanical adulterations in PFS. Among different approaches proposed for this purpose, mass spectrometry (MS)-based techniques have often been reported as the most promising, particularly when hyphenated with chromatographic techniques. Thus, this review aims at describing an overview of the developments in this field, focusing on the applications of MS-based techniques to targeted and untargeted analysis to detect botanical adulterations in plant materials, extracts, and PFS.

Quantitative Real-Time PCR for the Detection of Allergenic Species in Foods.

Food allergy is an increasing challenge to public health, with widespread global distribution. With no cure for this pathology, the food-allergic individuals are forced to adopt food eviction measurements, relying on label information to avoid consuming the offending foods. To safeguard these individuals, the analytical methods based on real-time PCR approaches are currently faced as excellent tools to verify labeling compliance, aiding industry and regulatory agencies to efficiently manage food allergen control programs. Therefore, this chapter intends to describe a protocol of real-time PCR to analyze allergenic food species. For method development, the main steps to be considered are (i) in silico sequence analysis and primer/hydrolysis probe design, (ii) preparation of calibrators (model foods containing the allergenic ingredient), (iii) efficient DNA extraction from complex food matrices, (iv) amplification by real-time PCR with hydrolysis probe (90-200 bp) targeting a highly specific DNA region (allergen-encoding gene), (v) sequencing PCR products for identity confirmation, and (vi) validation and application to commercial foods. Herein, a real-time PCR approach for the detection and quantification of cashew nut as an allergenic food is described as an example protocol, including all the steps for method development and validation.

CRISPR/Cas-mediated plant genome editing: outstanding challenges a decade after implementation.

The discovery of the CRISPR/Cas genome-editing system has revolutionized our understanding of the plant genome. CRISPR/Cas has been used for over a decade to modify plant genomes for the study of specific genes and biosynthetic pathways as well as to speed up breeding in many plant species, including both model and non-model crops. Although the CRISPR/Cas system is very efficient for genome editing, many bottlenecks and challenges slow down further improvement and applications. In this review we discuss the challenges that can occur during tissue culture, transformation, regeneration, and mutant detection. We also review the opportunities provided by new CRISPR platforms and specific applications related to gene regulation, abiotic and biotic stress response improvement, and de novo domestication of plants.

Edible Insects as a Novel Source of Bioactive Peptides: A Systematic Review.

The production of food and feed to meet the needs of the growing world's population will soon become a serious challenge. In search for sustainable solutions, entomophagy is being proposed as an alternative source of proteins, with economic and environmental advantages when compared to meat. Edible insects are not only a valuable source of important nutrients, but their gastrointestinal digestion also originates small peptides with important bioactive properties. The present work intends to provide an exhaustive systematic review on research articles reporting bioactive peptides identified from edible insects, as demonstrated by in silico, in vitro, and/or in vivo assays. A total of 36 studies were identified following the PRISMA methodology, gathering 211 potentially bioactive peptides with antioxidant, antihypertensive, antidiabetic, antiobesity, anti-inflammatory, hypocholesterolemia, antimicrobial, anti-severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), antithrombotic, and immunomodulatory properties, originated from the hydrolysates of 12 different insect species. From these candidates, the bioactive properties of 62 peptides were characterized in vitro and 3 peptides were validated in vivo. Data establishing the scientific basis of the health benefits associated with the consumption of edible insects can be a valuable contribution to overcoming the cultural issues that hinder the introduction of insects in the Western diet.

An in silico approach to unveil peptides from Acheta domesticus with potential bioactivity against hypertension, diabetes, cardiac and pulmonary fibrosis.

Entomophagy is a sustainable alternative source of proteins for human nutrition. Acheta domesticus is one of the three insect species that complies with the European Union Regulation on novel foods, but to date, there are no reports on their potential bioactive peptides. In this study, an in silico approach was applied to simulate the gastrointestinal (GI) digestion of six A. domesticus proteins and identify new peptides with potential anti-hypertensive and/or anti-diabetic properties, resulting from their capability to inhibit the somatic Angiotensin-I converting enzyme (sACE) and/or dipeptidyl peptidase 4 (DPP-4), respectively. A molecular docking protocol was applied to evaluate the binding interactions between the 43 peptides ranked with high probability of being bioactive and three drug targets: DPP-4 and two catalytic domains (N- and C-) of sACE. Five peptides (AVQPCF, CAIAW, IIIGW, DATW and QIVW) showed high docking scores for both enzymes, suggesting their potential to inhibit the DPP-4 and both catalytic domains of sACE, thus possessing multifunctional bioactive properties. Two peptides (PIVCF and DVW) showed higher docking scores for the N-domain of sACE, indicating a potential action as selective inhibitors and consequently with anti-cardiac and pulmonary fibrosis bioactivities. This is the first study identifying peptides originated from the simulated GI digestion of A. domesticus with potential activities against hypertension, diabetes, cardiac and pulmonary fibrosis.

Plasmonic genosensor for detecting hazelnut Cor a 14-encoding gene for food allergen monitoring.

A plasmonic nanostructure was constructed as a biorecognition element coupled to an optical sensing platform in sandwich format, targeting the hazelnut Cor a 14 allergen-encoding gene. The analytical performance of the genosensor presented a linear dynamic range between 100 amol L-1 and 1 nmol L-1, a limit of detection (LOD) < 19.9 amol L-1, and a sensitivity of 13.4 ± 0.6 m°. The genosensor was successfully hybridized with hazelnut PCR products, tested with model foods, and further validated by real-time PCR. It reached a LOD <0.001% (10 mg kg-1) of hazelnut in wheat material (corresponding to 1.6 mg kg-1 of protein) and a sensitivity of -17.2 ± 0.5 m° for a linear range of 0.001%-1%. Herein, a new genosensing approach is proposed as a highly sensitive and specific alternative tool with potential application in monitoring hazelnut as an allergenic food, protecting the health of sensitized/allergic individuals.

Techniques for Food Authentication: Trends and Emerging Approaches.

Food producers and retailers are obliged to provide correct food information to consumers; however, despite national and international legislation, food labels frequently contain false or misleading statements regarding food composition, quality, geographic origin, and/or processing [...].

Monitoring Yellow Mealworm (Tenebrio molitor) as a Potential Novel Allergenic Food: Effect of Food Processing and Matrix.

The consumption of insects has increased in western countries, raising concerns about their potential to induce food allergic reactions in sensitized/allergic individuals. This work intended to develop a real-time PCR approach for the detection/quantification of yellow mealworm (Tenebrio molitor) as a potential allergenic food in complex matrices. For this purpose, reference mixtures simulating the production of pork sausages and wheat biscuits containing known amounts of mealworm were used. Real-time PCR with TaqMan probe targeting the cytochrome b gene of T. molitor was able to detect up to 2 fg of insect DNA, and 1.0 and 0.1 mg/kg of mealworm flour in autoclaved sausages and baked biscuits, respectively. Generally, the method showed acceptable analytical performance parameters, confirming its suitability/applicability for a wide range of foods. However, real-time PCR data showed significant differences among food matrix and processing, highlighting the importance of using appropriate calibration models for quantitative analysis. Finally, the real-time PCR approach was successfully validated with blind mixtures and applied to commercial samples, demonstrating its efficacy and reliability in the quantification of mealworm in processed foodstuffs.

Authentication of incense (Pittosporum undulatum Vent.) honey from the Azores (Mel dos Açores) by a novel real-time PCR approach.

'Mel dos Açores' is a unique nectar honey produced from the exceptional and diverse flora of the Azores archipelago, categorised as incense honey ('mel de incenso') or multifloral honey ('mel multiflora'). Incense honey should contain over 30 % of pollen grains of Pittosporum undulatum Vent. In this work, a real-time PCR method targeting the ITS region was proposed for the first time to detect P. undulatum in the honey from the Azores. The approach exhibited high analytical performance, achieving a quantification limit of 0.01 pg of incense DNA. The method was successfully applied to 22 honey samples, from which incense was detected in all 9 monofloral incense honeys and in 5 out of 10 multifloral samples from the Azores. Generally, the quantitative results for incense DNA were in good agreement with the melissopalynological data. Therefore, a simple, cost-effective and reliable tool was herein proposed to authenticate and valorise the Azores honey.

Critical features of an in vitro intestinal absorption model to study the first key aspects underlying food allergen sensitization.

New types of protein sources will enter our diet in a near future, reinforcing the need for a straightforward in vitro (cell-based) screening model to test and predict the safety of these novel proteins, in particular their potential risk for de novo allergic sensitization. The Adverse Outcome Pathway (AOP) for allergen sensitization describes the current knowledge of key events underlying the complex cellular interactions that proceed allergic food sensitization. Currently, there is no consensus on the in vitro model to study the intestinal translocation of proteins as well as the epithelial activation, which comprise the first molecular initiation events (ME1-3) and the first key event of the AOP, respectively. As members of INFOGEST, we have highlighted several critical features that should be considered for any proposed in vitro model to study epithelial protein transport in the context of allergic sensitization. In addition, we defined which intestinal cell types are indispensable in a consensus model of the first steps of the AOP, and which cell types are optional or desired when there is the possibility to create a more complex cell model. A model of these first key aspects of the AOP can be used to study the gut epithelial translocation behavior of known hypo- and hyperallergens, juxtaposed to the transport behavior of novel proteins as a first screen for risk management of dietary proteins. Indeed, this disquisition forms a basis for the development of a future consensus model of the allergic sensitization cascade, comprising also the other key events (KE2-5).

Rosaceae food allergy: a review.

This review provides a global overview on Rosaceae allergy and details the particularities of each fruit allergy induced by ten Rosaceae species: almond/peach/cherry/apricot/plum (Amygdaleae), apple/pear (Maleae), and raspberry/blackberry/strawberry (Rosoideae). Data on clinical symptoms, prevalence, diagnosis, and immunotherapies for the treatment of Rosaceae allergy are herein stated. Allergen molecular characterization, cross-reactivity/co-sensitization phenomena, the impact of food processing and digestibility, and the methods currently available for the Rosaceae detection/quantification in foods are also described. Rosaceae allergy has a major impact in context to pollen-food allergy syndrome (PFAS) and lipid transfer protein (LTP) allergies, being greatly influenced by geography, environment, and presence of cofactors. Peach, apple, and almond allergies are probably the ones most affecting the quality of life of the allergic-patients, although allergies to other Rosaceae fruits cannot be overlooked. From patients' perspective, self-allergy management and an efficient avoidance of multiple fruits are often difficult to achieve, which might raise the risk for cross-reactivity and co-sensitization phenomena and increase the severity of the induced allergic responses with time. At this point, the absence of effective allergy diagnosis (lack of specific molecular markers) and studies advancing potential immunotherapies are some gaps that certainly will prompt the progress on novel strategies to manage Rosaceae food allergies.

Sesame as a source of food allergens: clinical relevance, molecular characterization, cross-reactivity, stability toward processing and detection strategies.

Sesame is an allergenic food with an increasing allergy prevalence among the European/USA population. Sesame allergy is generally life-persisting, being the cause of severe/systemic adverse immune responses in sesame-allergic individuals. Herein, clinical data about sesame allergy, including prevalence, diagnosis, relevance, and treatments are described, with focus on the molecular characterization of sesame allergens, their cross-reactivity and co-sensitization phenomena. The influence of food processing and digestibility on the stability/immunoreactivity of sesame allergens is critically discussed and the analytical approaches available for their detection in foodstuffs. Cross-reactivity between sesame and tree nuts or peanuts is frequent because of the high similarities among proteins of the same family. However, cross-reactivity phenomena are not always correlated with true clinical allergy in sensitized patients. Data suggest that sesame allergens are resistant to heat treatments and digestibility, with little effect on their immunoreactivity. Nevertheless, data are scarce, evidencing the need for more research to understand the effect of food processing on sesame allergenicity modulation. The demands for identifying trace amounts of sesame in foods have prompted the development of analytical methods, which have targeted both protein and DNA markers, providing reliable, specific, and sensitive tools, crucial for the effective management of sesame as an allergenic food.

Authentication of Argan (Argania spinosa L.) Oil Using Novel DNA-Based Approaches: Detection of Olive and Soybean Oils as Potential Adulterants.

Argan oil is a traditional product obtained from the fruits of the argan tree (Argania spinosa L.), which is endemic only to Morocco. It is commercialized worldwide as cosmetic and food-grade argan oil, attaining very high prices in the international market. Therefore, argan oil is very prone to adulteration with cheaper vegetable oils. The present work aims at developing novel real-time PCR approaches to detect olive and soybean oils as potential adulterants, as well as ascertain the presence of argan oil. The ITS region, matK and lectin genes were the targeted markers, allowing to detect argan, olive and soybean DNA down to 0.01 pg, 0.1 pg and 3.2 pg, respectively, with real-time PCR. Moreover, to propose practical quantitative methods, two calibrant models were developed using the normalized ΔCq method to estimate potential adulterations of argan oil with olive or soybean oils. The results allowed for the detection and quantification of olive and soybean oils within 50-1% and 25-1%, respectively, both in argan oil. Both approaches provided acceptable performance parameters and accurate determinations, as proven by their applicability to blind mixtures. Herein, new qualitative and quantitative PCR assays are proposed for the first time as reliable and high-throughput tools to authenticate and valorize argan oil.

Single-tube nested real-time PCR versus normalised real-time PCR for the quantification of allergenic cashew nut in foods: Impact of thermal processing and matrix.

In this work, three allergen-encoding genes (Ana o 1, Ana o 2, Ana o 3) were investigated for the detection of cashew nut as an allergenic food. Normalised and single-tube nested real-time PCR approaches targeting the Ana o 2 or Ana o 3 genes are proposed and compared. Normalised real-time PCR detected 10 pg, while single-tube nested real-time PCR achieved 1 pg of cashew nut DNA. Single-tube nested real-time PCR targeting Ana o 3 allowed the best relative sensitivities (10 mg/kg cashew nut in dough/biscuit), being successfully validated regarding precision/accuracy. The normalised real-time PCR did not show acceptable accuracy for both targets. Sensitivity of single-tube nested real-time PCR was affected by the matrix (pasta), but not by thermal processing (dough/biscuit). Herein, two highly sensitive and specific single-tube nested real-time PCR targeting allergen-encoding genes are proposed for the first time as quantitative/validated tools for cashew nut analysis as an allergenic food.

Animal Species Authentication in Dairy Products.

Milk is one of the most important nutritious foods, widely consumed worldwide, either in its natural form or via dairy products. Currently, several economic, health and ethical issues emphasize the need for a more frequent and rigorous quality control of dairy products and the importance of detecting adulterations in these products. For this reason, several conventional and advanced techniques have been proposed, aiming at detecting and quantifying eventual adulterations, preferentially in a rapid, cost-effective, easy to implement, sensitive and specific way. They have relied mostly on electrophoretic, chromatographic and immunoenzymatic techniques. More recently, mass spectrometry, spectroscopic methods (near infrared (NIR), mid infrared (MIR), nuclear magnetic resonance (NMR) and front face fluorescence coupled to chemometrics), DNA analysis (real-time PCR, high-resolution melting analysis, next generation sequencing and droplet digital PCR) and biosensors have been advanced as innovative tools for dairy product authentication. Milk substitution from high-valued species with lower-cost bovine milk is one of the most frequent adulteration practices. Therefore, this review intends to describe the most relevant developments regarding the current and advanced analytical methodologies applied to species authentication of milk and dairy products.