Background: Although allergic reactions to honey are rare, there have been documented systemic allergic reactions following ingestion of honey. Moreover, there have been reports of supplementary bee feeding, honey adulteration and mould contamination in beehives that identify a potential for gluten and allergenic presence in honey. As such, this study aimed to investigate whether honey samples contained detectable levels of gluten, food and mould allergens that could pose a risk to consumers. Methods: Honey samples (n=40) of UK, EU and Non-EU origin were extracted and analysed for gluten using the R5 immunoassay and for major allergen content using quantitative multiplex arrays for allergens from milk, egg, peanut, soy, hazelnut, cashew and moulds. Results: Gluten was identified in 28% of samples analysed at concentrations ranging 2.5 to 13.8ppm. Milk allergens Bos d 5 and Bos d 11 were detected in 7% of samples, in concentrations ranging 0.37mg/kg-0.57mg/kg and 0.03mg/kg-0.18mg/kg respectively. Samples positive for milk allergen also contained detectable levels of gluten. No detectable amounts of egg, peanut, soy, hazelnut, cashew or mould allergen was identified in the samples. Conclusion: This novel study is the first of its kind in the identification of gluten and milk allergens within honey. Consumers may take reassurance in that levels of gluten detected in all samples fell below the current 20ppm ‘gluten free’ threshold. Similarly, none of the samples analysed for milk allergen content exceeded VITAL 3.0 reference doses. However, caution may be required for repeated consumption, leading to accumulating doses of gluten and allergen.

Background: T cell responses to natural SARS-CoV-2 infection may be more robust and longer lived than antibody responses, thus preventing re-infection. Accurate assessment of vaccine-induced T cell responses is critical for understanding the magnitude and longevity of vaccine-induced immunity across patient cohorts. Aims: To establish a simple, accurate and rapid whole blood test to determine natural and vaccine-induced SARS-CoV-2 immunity via a cytokine release assay. Methods: Cytokine release in whole blood stimulated with peptides specific for SARS-CoV-2 was measured in donors with PCR-confirmed previous infection (n=29), suspected infection (n=30) or with no history of exposure (n=69); and in donors pre- and post-vaccination (n=32). Cytokines were measured by enzyme immunoassay and multiplex array. Results: Cytokines interleukin-2 (IL-2) and interferon-gamma (IFN-γ) were highly elevated in PCR-confirmed or suspected SARS-CoV-2 infected donors at 20->2000pg/ml and 20-1000pg/ml, respectively, compared to history negative controls (<20-90pg/ml). Receiver operating curves showed IL-2 as the superior biomarker with AUC of 0.99 compared to IFN-γ (0.94). Following vaccination, 100% of PCR-confirmed donors and 94% of unexposed individuals demonstrated a positive IL-2 response. Mean IL-2 levels increased ~18-fold from 12pg/ml pre-vaccination to 202pg/ml and 216pg/ml after the 1 st and 2 nd vaccine doses, respectively. No other cytokines were suitable biomarkers for distinguishing SARS-CoV-2 infection or vaccination responses. Conclusion: This rapid, whole blood-based T cell test can be utilised to make accurate and comparable assessments of vaccine-induced T cell immunity across multiple population cohorts, and aid decision making on public health policies and vaccine efficacy.

Title :Identification of gluten and major milk allergens Bos d 5 and Bos d 11 within commercially available honey.To the Editor,Although allergy to honey is rare, there have been documented systemic allergic reactions following ingestion of honey[1]. Interestingly, supplementary bee feeding with mixtures of soybean flour, dried brewer’s yeast (containing high levels of residual gluten from brewing processes), and dry skimmed milk with sugar and water, can be implemented by beekeepers to maintain healthy honeybee colonies[2]. Additionally, there have been reports of mould contamination within beehives[3]. Both factors suggest a potential for gluten and allergenic protein presence in honey, which could account for some of the reported reactions following honey consumption. As such, the aim of this study was to determine if commercially available honey contained undeclared gluten and/or food or mould allergens, and at levels which could present a risk to individuals with hypersensitivities.To investigate this, honey samples (n=40) of UK, EU, Non-EU and blended Non-EU/EU origin were extracted and analysed for gluten using the R5 immunoassay and for major allergen content using quantitative multiplex arrays for allergens from cow’s milk, egg, peanut, soy, hazelnut, cashew and moulds[4] (see online supporting material for methods) .Of the forty samples analysed, it was observed that 11 of the 40 (27.5%) samples contained gluten in the range of 2.5ppm to 13.8 ppm. Positive honey samples categorised by origin are detailed in Table 1 and Figure 1. Out of the 21 non-EU honey samples analysed, 7 were positive for gluten; forming 63.6% of the positive samples. From the 9 UK and 9 EU/non-EU blend honey samples, both had 2 samples positive for gluten, each accounting for 18.2% of the positive samples.Milk allergens Bos d 5 and Bos d 11 were detected in 7% of samples. The positive results ranged from 0.368mg/kg, up to 0.567mg/kg for Bos d 5, and from 0.030mg/kg up to 0.182mg/kg for Bos d 11; with one sample being a blend of EU/Non-EU honey, and two being of UK origin. Samples found to contain Bos d 5 and Bos d 11 were also positive for gluten. Results of the 3 positive samples are shown in Table 1 and Figure 1. No detectable amounts of egg, peanut, soy, hazelnut, cashew or mould allergen was identified in the samples. To our knowledge, this novel study is the first of its kind to identify the presence of milk allergen and gluten within honey. Consumers may take some reassurance in that levels of gluten detected in all samples fell below the current 20ppm ‘gluten free’ threshold[5]. Similarly, in the three samples positive for cow’s milk allergens Bos d 5 and Bos d 11, the levels of milk allergen detected did not exceed the VITAL 3.0 reference dose of 0.2mg of milk protein per serving; a level that should prevent 99% of the allergic population from having a reaction[6] (see online supporting information for VITAL 3.0 calculations). However, for both milk allergen and gluten, caution may be required when considering repeated consumption, leading to accumulating doses. The fact that no allergen contaminants other than milk were identified in the honey samples can offer consumers further reassurances. Current literature does not give reason to suspect contamination from sources other than milk and soy, so it is encouraging the data shows agreement.The findings herein could indicate honey suppliers may wish to undertake allergen testing of their products to offer greater safety to consumers. Moreover, it may be useful for future research to investigate the ability of allergen contaminants to be transferred from bee supplementary feeds to the final product to prevent possible accidental contamination of undeclared allergens.WORD COUNT : 587