Application of mathematical models to mycotoxins children risk assessment: a case study of Portuguese children exposure to co-occurring mycotoxins in processed cereal-based foods

People, animals and the environment can be exposed to multiple chemicals at once from a variety of sources, but current risk assessment is usually carried out based on one chemical substance at a time. In human health risk assessment, ingestion of food is considered a major route of exposure to many contaminants, namely mycotoxins, a wide group of fungal secondary metabolites that are known to potentially cause toxicity and carcinogenic outcomes. Mycotoxins are commonly found in a variety of foods including those intended for consumption by infants and young children and have been found in processed cereal-based foods available in the Portuguese market. The use of mathematical models, including probabilistic approaches using Monte Carlo simulations, constitutes a prominent issue in human health risk assessment in general and in mycotoxins exposure assessment in particular. The present study aims to characterize, for the first time, the risk associated with the exposure of Portuguese children to single and multiple mycotoxins present in processed cereal-based foods (CBF). Portuguese children (0-3 years old) food consumption data (n=103) were collected using a 3 days food diary. Contamination data concerned the quantification of 12 mycotoxins (aflatoxins, ochratoxin A, fumonisins and trichothecenes) were evaluated in 20 CBF samples marketed in 2014 and 2015 in Lisbon; samples were analyzed by HPLC-FLD, LC-MS/MS and GC-MS. Daily exposure of children to mycotoxins was performed using deterministic and probabilistic approaches. Different strategies were used to treat the left censored data. For aflatoxins, as carcinogenic compounds, the margin of exposure (MoE) was calculated as a ratio of BMDL (benchmark dose lower confidence limit) to the aflatoxin exposure. The magnitude of the MoE gives an indication of the risk level. For the remaining mycotoxins, the output of exposure was compared to the dose reference values (TDI) in order to calculate the hazard quotients (ratio between exposure and a reference dose, HQ). For the cumulative risk assessment of multiple mycotoxins, the concentration addition (CA) concept was used. The combined margin of exposure (MoET) and the hazard index (HI) were calculated for aflatoxins and the remaining mycotoxins, respectively. 71% of CBF analyzed samples were contaminated with mycotoxins (with values below the legal limits) and approximately 56% of the studied children consumed CBF at least once in these 3 days. Preliminary results showed that children exposure to single mycotoxins present in CBF were below the TDI. Aflatoxins MoE and MoET revealed a reduced potential risk by exposure through consumption of CBF (with values around 10000 or more). HQ and HI values for the remaining mycotoxins were below 1. Children are a particularly vulnerable population group to food contaminants and the present results point out an urgent need to establish legal limits and control strategies regarding the presence of multiple mycotoxins in children foods in order to protect their health. The development of packaging materials with antifungal properties is a possible solution to control the growth of moulds and consequently to reduce mycotoxin production, contributing to guarantee the quality and safety of foods intended for children consumption.

Contribute of the MYCOMIX project and future perspectives for Human Biomonitoring and health risk assessment in Portugal

There is a growing concern within public health about mycotoxin involvement in human diseases, namely those related to children. The MycoMix project (2012-2015), funded by the Portuguese Foundation for Science and Technology, gathered a multidisciplinary team aiming at answering several questions: 1) Are Portuguese children exposed daily to one or several mycotoxins through food? 2) Can this co-exposure affect children´s health? and 3) Are there interaction effect between mycotoxins? Mycomix results revealed that Portuguese children (< 3 years old, n=103) are exposed to multiple mycotoxins through food consumption. Cumulative risk assessment results revealed a potential health concern for the high percentiles of intake, specially for aflatoxins which are carcinogenic compounds. This fact assumes particular importance considering the interactive effects found in in vitro bioassays. These results highlight the need for a more accurate approach to assess the human exposure to mycotoxins6. Within the Mycomix project the assessment of mycotoxin exposure was based on calculations combining mycotoxin data in food with population data on food consumption. This approach does not consider some aspects as the inter-individual metabolism variation, the exposure through sources other than food and the heterogeneous distribution of mycotoxins in food. Exposure assessment of mycotoxins in Portuguese population through biomarkers is still missing and further studies are urgent to be developed. The European Human Biomonitoring Initiative (EHBMI), a proposal within the European Joint Programme, aims to advance the understanding of the extent of exposure to environmental chemicals across Europe and the impact on human health, by gathering national expertise in human biomonitoring domain. At national level Mycomix project uncovered the potential health risk of exposure of Portuguese children to multiple mycotoxins. The risk assessment expertise acquired within Mycomix, namely in analysis and toxicology of chemical mixtures, will be brought together as a contribute to EHBMI objectives.

Portuguese children exposure to multiple mycotoxins through food consumption: toward a holistic approach for multiple mycotoxins risk assessment

Humans can be exposed to multiple chemicals at once from a variety of sources, and human risk assessment of multiple chemicals poses several challenges to scientists, risk assessors and risk managers. Ingestion of food is considered a major route of exposure to many contaminants, namely mycotoxins, especially for vulnerable population groups, as children. A lack of sufficient data regarding mycotoxins children risk assessment, could contribute to an inaccuracy of the estimated risk. Efforts must be undertaken to develop initiatives that promote a broad overview of multiple mycotoxins risk assessment. The present work, developed within the MYCOMIX project, aims to assess the risk associated to the exposure of Portuguese children (< 3 years old) to multiple mycotoxins through consumption of foods primarily marketed for this age group. A holistic approach was developed applying deterministic and probabilistic tools to the calculation of mycotoxin daily intake values, integrating children food consumption (3-days food diary), mycotoxins occurrence (HPLC-UV, HPLC-FD, LC-MS/MS and GC-MS), bioaccessibility (standardized in vitro digestion model) and toxicological data (in vitro evaluation of cytotoxicity, genotoxicity and intestinal impact). A case study concerning Portuguese children exposure to patulin (PAT) and ochratoxin A (OTA), two mycotoxins co-occurring in processed cereal-based foods (PCBF) marketed in Portugal, was developed. Main results showed that there is low concern from a public health point of view relatively to PAT and OTA Portuguese children exposure through consumption of PCBF, considering the estimated daily intakes of these two mycotoxins (worst case scenarios, 22.930 ng/kg bw/day and 0.402 ng/kg bw/day, for PAT and OTA, respectively), their bioaccessibility and toxicology results. However, the present case study only concerns the risk associated with the consumption of PCBF and child diet include several other foods. The present work underlines the need to adopt a holistic approach for multiple mycotoxins risk assessment integrating data from exposure, bioacessibility and toxicity domains in order to contribute to a more accurate risk assessment.

Using percentiles to diagnose familial hypercholesterolemia in Portugal

Aims: Familial hypercholesterolemia (FH) is a genetic disorder of lipid metabolism, clinically characterised by high levels of low-density lipoprotein cholesterol (LDL-C) that leads to cholesterol accumulation in tendons and arteries, premature atherosclerosis and increased risk of premature coronary heart disease. In 1999, the Portuguese FH Study was established at the National Institute of Health to identify the genetic cause of hypercholesterolemia in individuals with a clinical diagnosis of FH and to perform an epidemiologic study to determine the prevalence and distribution of FH in Portugal. In the last 16 years, a genetic defect was identified in 749 patients, representing 3. 7 % of the cases estimated to exist in Portugal. Index patients were included in this study using the Simon Broome (SB) criteria. However, there are different FH clinical criteria to diagnose index cases. Since there are no clinical criteria to identify relatives with FH, the aim of this work was to investigate if a diagnostic tool based on population specific 95 th percentile improves the clinical identification of Portuguese FH patients comparing with SB criteria.

Caracterização genotípica de doentes portugueses com patologias associadas ao complexo multienzimático lisossomal: sialidose e galactosialidose

Dissertação de mestrado em Biologi apresentada à Faculdade de Ciências da Universidade do Porto, 2008

Excess Mortality associated to Influenza epidemic in Portugal, from 2007/2008 to 2014/2015 seasons

Mortality in the north hemisphere is higher in winter than in summer seasons, due to the influenza epidemics as well as cold temperatures. Portuguese influenza surveillance comprises clinical and laboratorial notifications of Influenza-like Illness (ILI) attended in the primary health care units and emergency rooms. Without information on specific cause of deaths in real time, estimation of influenza impact has been accessed using Portuguese Daily Mortality Monitoring System (VDM), that covers all cause mortality of Portuguese population. The aim of this study was to provide excess mortality, potentially associated to Influenza each season (between 2007/08 and 2014/15).