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- Exposure to mycotoxins in the Portuguese adult populationPublication . Maris, Elias; Namorado, Sónia; Chen, A.; Pero-Gason, Roger; De Boevre, Marthe; De Saeger, Sarah; Silva, Maria João; Alvito, PaulaMycotoxins are toxic fungal metabolites commonly found in food, posing health risks such as immunosuppression, carcinogenicity, and endocrine disruption. Despite regulatory limits, chronic low-level exposure remains a concern. Understanding real-life exposure in populations is essential for effective risk assessment. This study aims to investigate mycotoxin exposure among young adults in Portugal, contributing to evidence-based public health interventions. This study leveraged data and biospecimens from the INSEF-ExpoQuim survey, a cross-sectional study nested within thPortuguese National Health Examination Survey (INSEF). Data was collected via REDCap-assisted telephone interviews, covering sociodemographic and exposure-relevant variables. A subset of 295 first morning urine samples was collected from adults aged 28–39 years between May 2019 and March 2020. Urine samples were analyzed by a newly optimized and validated LC-MS/MS method targeting 40 mycotoxins and/or their corresponding metabolites in urine. Urinary creatinine was measured using a validated colorimetric method to allow adjustment and standardization of mycotoxin concentrations, ensuring accurate exposure assessment and comparability. This methodological approach enabled a robust characterization of mycotoxin exposure in a representative Portuguese population cohort.The study included 58% females and 42% males. Most participants had medium to high education, and urbanization was nearly evenly split between towns/suburbs (36.9%) and rural areas (35.9%), with fewer living in cities (27.1%). The majority were employed, and sampling was primarily conducted in summer and autumn. The number of mycotoxin co-exposures in the Portuguese population ranged from 0 to 5, with two simultaneous exposures being most common (n = 160). Among the 40 mycotoxins analysed, deoxynivalenol and tenuazonic acid were most frequently detected, with frequency of detection of 85% and 96%, respectively. This study offers robust biomonitoring data on mycotoxin exposure in Portuguese young adults using a validated LC-MS/MS method. The high prevalence of deoxynivalenol and tenuazonic acid suggests low level dietary contamination. These findings support the need for continued monitoring and the integration ofhuman biomonitoring into national food safety strategies. Detailed sociodemographic analyses are planned to further clarify exposure patterns and enable targeted public health interventions.
- Genomic diversity of Enterobacteriaceae on food-processing surfaces: an overlooked non-clinical setting with public health implicationsPublication . Peixoto, Andreia; Nova, Lúcia; Mourão, Joana; Matos, Cátia; Santos, Susana; Rodrigues, João; Saraiva, Margarida; Correia, Cristina Belo; Batista, Rita; Peixe, Luísa; Novais, Carla; Antunes, PatríciaBacterial hazards on contaminated surfaces in food-processing environments pose significant food safety risks. Hygiene monitoring of surfaces in direct or indirect contact with food typically relies on a limited set of bacterial indicators, such as Enterobacteriaceae counts. However, their genomic diversity remains poorly characterised. The aim of this study was to provide a comprehensive genomic characterisation of Enterobacteriaceae recovered from food-contact surfaces after cleaning and disinfection with biocides in foodservice units. Diverse strains were identified, with some STs shared across samples and/or foodservices. - A diverse Enterobacteriaceae population persists on food-contact surfaces even after cleaning and disinfection with biocides, with evidence of cross-contamination. - These surfaces represent critical points of contact for strains that are genetically similar to those circulating in food, environment, and human clinical sources, including strains carrying clinically relevant AMR genes. - Further research is needed to understand the factors driving their persistence and dissemination in foodservice environments, aiming to enhance food safety risk management protocols and protect public health.
- Intestinal Permeability Studies using a more realistic barrier: performance of co-cultures of Caco-2/HT29-MTX cellsPublication . Figueira, Carolina; Gravato, Carlos; Alvito, PaulaThe intestinal barrier, essential for overall health, can have its permeability affected by certain food compounds and additives. Among various models, in vitro cellular monolayers are the most commonly used to study this process. Among these, Caco-2 cells—representing enterocytes—are commonly used, though they lack complexity to mimic some properties of the intestinal barrier. This limitation can be overwhelmed by co-culturing it with HT29-MTX cells, which allows the secretion of mucus and mimics goblet cell functioning. This study aimed to evaluate the intestinal permeability by assessing the paracellular and transcellular transport of lucifer yellow (LY) and propranolol (PR), two intestinal permeability markers, respectively, using a 9:1 co-culture of Caco-2/HT29-MTX cells. Cells were cultivated separately in complete medium. Functional monolayer formation was monitored over 28 days using transepithelial electrical resistance (TEER) measurements in triplicate plates, with values ranging from 450.6 to 1287.3 Ω·cm². Transport assays were conducted on day 21 by applying LY and PR to the apical compartment and measuring their passage to the basolateral side. Apparent permeability coefficients (Papp) and basal recovery values were estimated by fluorescence quantification. Statistical analyses were conducted to evaluate variability in TEER, Papp, and basal recovery results. The Shapiro-Wilk test was used to assess normality, and comparisons used one-way ANOVA followed by Tukey’s test or the Kruskal-Wallis test followed by Dunn’s test when requisites of ANOVA were not met. Significant differences in TEER values were observed between days 8–20 (p = 5.8e-11), 22–28 (p = 2.3e-09), and before vs. after transport on day 21 (p < 2.2e-16). For day 21 transport assays, a significant difference was found in LY basal recovery for two out of three plates (p = 0.03). As for the PR Papp values, there is a significant difference between duplicate plates (p = 0.005), as one plate was excluded due to a poor calibration curve fit. These findings will inform improvements to the protocol for assessing intestinal permeability using co-culture models. Thus, Caco-2/HT29-MTX co-culture appears to be a promising model for evaluating the impact of food components and additives on the intestinal barrier.
- Surface hygiene in food environments: challenges with biocide efficacy and bioindicatorsPublication . Peixoto, Andreia; Matos, Cátia; Santos, Susana; Rodrigues, João; Saraiva, Margarida; Belo Correia, Cristina; Batista, Rita; Peixe, Luísa; Novais, Carla; Antunes, PatríciaThe presence of bacterial hazards on contaminated surfaces in food-processing environments poses significant food safety risks. Monitoring and hygiene verification of surfaces in direct or indirect contact with food remains a top priority for food business operators. Sanitation uses antimicrobial compounds, biocides (disinfectants/antiseptics), whose effectiveness on surface microbiota depends on microorganisms’ susceptibility and factors such as concentration and contact time. The aim of this stydy was to assess the post-disinfection occurrence and diversity of hygiene indicator bacteria on kitchen surfaces of Portuguese foodservice facilities and correlate their presence with biocide usage. Despite recommended biocide use, poor hygiene of surfaces in direct or indirect contact with food was observed. The presence of diverse, potentially pathogenic and recurrent bacterial species suggests persistence, reintroduction, and/or cross-contamination. Identifying phenotypic and genetic markers of persistence, including biocide susceptibility, is crucial for developing real-time hygiene bioindicators and ensuring food safety.
- Towards safer edible insects: assessing contaminant bioaccumulation and depuration capacity in Tenebrio molitorPublication . Cardoso, Diogo N.; Alvito, Paula; de Boevre, Marthe; Silva, Ana Rita; Pinto, José; Rodrigues, Ana; Prodana, Marija; Silva, Patricia; Mostafaie, Amid; Loureiro, SusanaThe integration of insects into sustainable food and feed chains demands a robust evaluation of their capacity to accumulate and eliminate contaminants. Understanding these dynamics is fundamental to ensuring consumer safety and supporting the development of regulatory standards for insect-derived products. This study assessed the bioaccumulation and depuration kinetics of metal(loid)s (As, Cd, Pb) and mycotoxins (Aflatoxin B1, Ochratoxin A, Deoxynivalenol, and Zearalenone) in the yellow mealworm Tenebrio molitor, a species already authorized for human consumption in the EU. Larvae were reared on feed substrates artificially spiked at the maximum levels allowed for animal feed under EU legislation. The experimental design included 21 days of exposure followed by a 21-day elimination phase on uncontaminated substrate, to evaluate both uptake and depuration efficiency under realistic production conditions. Distinct toxicokinetic patterns were observed among metals. Arsenic displayed the highest accumulation potential, reaching internal concentrations of approximately 7 mg/kg dry weight (DW) at the end of exposure, with a progressive but incomplete decline during depuration. Cadmium accumulated steadily to ~2.5 mg/kg DW, exceeding the current feed limit, but was rapidly eliminated below regulatory thresholds within one day of clean feeding. Lead, in contrast, showed low assimilation and rapid clearance, remaining consistently below EU maximum levels. Mycotoxins and their metabolites were not detected above quantification limits in any treatment, confirming the species’ limited bioaccumulation potential for these compounds. Overall, the results highlight Tenebrio molitor’s high depuration capacity and resilience to transient contaminant exposure. These findings highlight the need to apply toxicokinetic principles in insect risk assessment to define effective depuration periods and ensure safe, circular production systems.