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- Analytical determination of nitrates in vegetablesPublication . Laia, Ana; Serra, Maria; Vasco, ElsaIntroduction: Nitrate compounds are common in the nature and widely found in food, water and soil. Generally, the nitrate compounds from vegetables are considered to be the main source of dietary nitrate intake. When consumed in excessive quantities, nitrate can be harmful to health because it could cause carcinogenesis and metahemoglobinemia. However, vegetables have an essential nutritional function and play an important role in health protection. In this way, to assess the dietary exposure of the population to these compounds, it is important to know the nitrate contents in vegetable products as they are consumed. As the nitrate concentration in vegetables varies according to season it is also important to study the effect of seasonality on the nitrate content.
- Influence of seasonality in carotenoids content of vegetables commercially available in PortugalPublication . Dias, Maria da Graça; Serra, Celeste; Almeida, AnaAim: To study the influence of seasonality in carotenoids content present in vegetable samples commercially available in Portuguese market.
- Enterovirus D68 diagnosed in severe respiratory and neurological illness in children during 2015-2016 season in PortugalPublication . Guiomar, Raquel; Costa, Inês; Pechirra, Pedro; Palminha, Paula; Ribeiro, Carlos; Roque, Carla; Peres, Maria João; Viseu, Regina; Balseiro, Maria Jesus; Brito, Maria João; Neves, João; Branquinho, Paula; Côrte-Real, RitaBackground: Enterovirus D68 (EV-D68) was first isolated in 1962, and since then associated with respiratory illness. The report of severe respiratory and neurological disease including deaths associated to EV-D68 in United States and Canada during August 2014 highlighted the need of epidemiological information regarding EV-D68 circulation. In Europe information was scarce, available only for few countries. In Portugal there was no data available and was critical to know the epidemiology of EV-D68, especially in children hospitalized with severe respiratory or neurological disease. This study aims to identify EV-D68 in Enterovirus positive respiratory samples in children under 18 with clinical diagnosis of severe respiratory infection or neurological illness. Methods: During 2015/16 winter season, between November/2015 and March/2016, 29 EV positive cases were reported to the National Influenza and Other Respiratory Virus Reference Laboratory (NIC) by two hospitals located in Lisbon and Setubal districts. EV diagnosis was performed in hospitals by biomolecular methods using commercial kits (real time multiplex-PCR, FTD Respiratory pathogens 21 and CLART Pneumovir, Genomica, respectively). EV-D68 was diagnosed by an in house real-time PCR [1]. Virus isolation in RD cell line and phylogenentic analysis of the VP1/VP3 genomic regions will enable the identification of genetic groups in circulation. All samples were irreversibly anonymized. Demographic and clinical data were collected. Results: EV-D68 was confirmed in 20 respiratory samples previously positive for EV (69%; 20/29). Samples were collected from children with age ranging from 2 months to 6 years old, both genders (9 female; 11 male) with diagnosis of severe respiratory or neurological illness. Eighteen cases were hospitalized (90%; 18/20). Bronchiolitis and pneumonia were the most frequently reported diagnosis, corresponding to 70% (14/20). Two cases have neurologic diagnosis. EV-D68 was identified throughout all study period with the higher number of positive cases detected during January 2016, in week 3. Virus isolation and genetic characterization are under way with expected results in virus phylogeny and evaluation on similarity with recent circulating strains in United States, Canada and European countries. Conclusions: EV-D68 was detected in a high positive rate (69%) among EV positive cases. This positive rate of EV-D68 was higher compared to the positivity rate of 10,2 %, calculated in a European study during 2014 [2]. This finding could be linked to the selection of severe and hospitalized patients in present study, highlighting the involvement of EV-D68 with severe respiratory disease in children. The identification of EV-D68 is also crucial in respiratory samples in children with clinical diagnosis of neurological illness. This study is the first attempt to describe the prevalence of EV-D68 in severe paediatric cases, in Portugal. The strength of EV-D68 surveillance in paediatric and adult population at the national level will be important to understand the epidemiology of EV-D68, age-related susceptibility and association with disease severity.
- Molecular characterization of respiratory syncycial virus during 2015-2016 season in PortugalPublication . Cristóvão, Paula; Pereira, Diogo; Pechirra, Pedro; Pereira-Vaz, João; Correia, Lurdes; Rodrigues, Fernando; Andrade, Graça; Corte-Real, Rita; Branquinho, Paula; Peres, Maria João; Viseu, Regina; Balseiro, Maria Jesus; Mota, Paula; Guiomar, RaquelBackground: Respiratory syncytial virus (RSV) is one of the most frequent and important respiratory viral agent that causes respiratory infection complications in younger children and elderly. RSV has an autumn / winter seasonality. Genetic diversity in both of RSV A and B subtypes increased in last years with the spread of new genotypes. This study aims to describe the genetic variability of RSV during 2015/2016 season in Portugal and correlate the circulating genotypes with detected ones in previous seasons. Will also be evaluated the association between genotype, clinical diagnosis and age. Methods: During 2015/16 winter season, between November/2015 and February/2016, 45 RSV were genetically characterized. RSV positive respiratory samples were collected in two settings: children under 5 years old diagnosed by hospital laboratories from the Portuguese Laboratory Network for the Diagnosis of Influenza Infection, and all age Influenza-like illness (ILI) patients reported by primary care health services and diagnosed by the National Influenza Reference Laboratory. All samples were irreversibly anonymized. Demographic and clinical data were collected. RSV detection was performed by real-time PCR and other biomolecular methods. RSV genotype was assigned by the nucleotide sequence of the hypervariable C-terminal region of the G protein gene and the phylogenetic analysis was performed in MEGA 6.0. Results: From 45 RSV genetically characterized, 31 (69%) were reported by hospitals, patients age ranged from newborn to 4 years old. From these, 25 (81%; 25/31) patients were hospitalized, being the bronchiolitis the most frequent diagnosis. While 14 (31%) RSV cases came from primary care health services, patients age ranged from 3 to 83 and all had a clinical diagnosis of ILI. Were included patients from both genders in equal proportions. RSV A and B co-circulated during 2015/2016 season. Were genetically characterized 21 (47%) RSV A and 24 (53%) RSV B. 90% (19/21) of RSV A clustered in ON1 genotype, the others 2 clustered with NA1 genotype. All RSV B present a BA-like genotype, 70% (17/24) were similar to BA9 and 30% (7/24) clustered with BA10 genotype. Conclusions: During 2015/2016 season was observed a co-circulation of RSVA and RSVB. In present study ON1genotype was predominant in circulation among RSVA, this was also detected as the major RSVA genotype at the global level. Only two RSVA belonged to NA1 genotype. In Portugal, NA1 was in circulation during 2010-2012 period. Undetected since 2012, it seems to reappear during 2015/16 season. All RSVB characterized belonged to BA genotypes, the majority clustered within BA9 genotype. BA10 genotype was also identified in circulation at low frequency. BA9 and BA10 were being found in co-circulation since 2011/12. No association was found between age, clinical diagnosis and RSVA and B genotypes. RSV has an important impact in children in high-risk groups highlighting the need off a continuous RSV surveillance each winter.
- Molecular characterization of human rhinovirus in Portugal: 2014-2015 seasonPublication . Cristóvão, Paula; Pechirra, Pedro; Conde, Patrícia; Costa, Inês; Guiomar, RaquelBackground: Human rhinoviruses (HRV) frequently cause mild upper respiratory tract infections and more severe disease manifestations such as bronchiolitis and pneumonia. Plays an important role in asthma and chronic lung disease exacerbations. Genetic characterization of HRVs detected by molecular methods has revealed much greater diversity, enabling the identification of three species (HRV-A, HRV-B and HRV-C) and a great number of types. The aim of this study is to characterize HRV detected in respiratory specimens from influenza-like illness cases (ILI), received in the scope of the Portuguese Influenza Surveillance Program, during 2014-2015 season in Portugal. Methods: During the 2014 - 2015 season, 411 nasopharyngeal swabs negative for influenza were tested for HRV in a multiplex PCR [1]. 18% (75/411) of the samples were positive for HRV, and from these 83% (62) were sequenced by a nested RT-PCR [2]. Nucleotide sequences of the VP4/VP2 region were used for genotyping and phylogenetic tree construction in Mega 6.0. Demographic and clinical data (according EU ILI case definition) were recorded in a questionnaire,. Results: HRV were detected throughout the study period, between week 40/2014 (October) and week 9/2015 (March) with a peak in January 2015. Phylogenetic analysis showed that 45% (28/62) strains belonged to species HRV-A, 15% (9/62) to species B and 40% (25/62) to species C. Overall were identified 35 different types. All species co-circulated in Portugal with the exception of the Algarve and Açores, being HRV-A predominat in north region and HRV-C predominant in Alentejo. HRV positives cases had a median age of 42,5. HRV-C were the most frequently detected in all age groups, apart from young adults aged 15 to 44. In this age group HRV-A were identified in 73% (16/22). HRV-B was detected sporadically in all age groups, except in children (5-14 years old). HRV was found in similar proportions in both genders (52% in female; 48% in male). Data on influenza vaccination was reported in 51 HRV positive cases, but only 9 (18%) had been previously vaccinated. Information on chronic diseases was reported in 54 cases, of these 15 (28%) had a chronic disease (mainly cardiovascular or diabetes). HRV was detected in 4 pregnant women, 14% (4/28). Cough, myalgias, weakness and fever were the most frequent symptoms reported by HRV confirmed cases. Conclusions: During 2014-2015 was observed a co-circulation of the three species of HRV (A, B and C) with a predominance of HRV-A followed by the recently identified specie C. A wide genetic diversity of 35 types was identified, with a higher diversity among HRV-A. HRV was most frequently diagnosed in adults. Our study included few children under 5, preventing conclusions about this group. Diabetes and cardiovascular disease were found has a possible risk for HRV infection, highlighting the relevance of respiratory disease prevention measures that should be undertaken. This was the first study to attempt the gentic diversity of rhinovirus circulating in Portugal during a winter season in ILI cases. Further studies in the general population and in high risk groups for severe respiratory disease will aid knowledge in HRV epidemiology and exarcebation of respiratory infections.
- Tumor-related splicing variant RAC1b in colorectal cancer: regulation and role in tumorigenesisPublication . Jordan, PeterPurpose: Alternative splicing of the small GTPase RAC1 generates RAC1b, a hyperactivated variant that is overexpressed in a subtype of colorectal tumors. The objective of our studies is to understand the molecular regulation of this alternative splicing event and how it contributes to tumorigenesis. Experimental description: The regulation of the RAC1b splicing event in human colon cell lines was dissected using a transfected RAC1 minigene and the role of upstream regulating protein kinases through an RNA interference approach. The functional properties of the RAC1b protein were characterized by experimental modulation of Rac1b levels in colon cell lines. Results: The RAC1b protein results from an in-frame inclusion of an additional alternative exon encoding 19 amino acids that change the regulation and signaling properties of the protein. RAC1b is a hyperactive variant that exists predominantly in the GTP-bound active conformation in vivo and promotes cell cycle progression and cell survival through activation of the transcription factor NF-κB. RAC1b overexpression functionally cooperates with the oncogenic mutation in BRAF-V600E to sustain colorectal tumor cell survival. The splicing factor SRSF1 was identified to bind an exonic splice enhancer element in the alternative exon and acts as a prime regulator of Rac1b alternative splicing in colorectal cells. SRSF1 is controlled by upstream protein kinase SRPK1, the inhibition or depletion of which led to reduced SRSF1 phosphorylation and nuclear translocation with a concomitant reduction in RAC1b levels. As further SRSF1-regulating pathways we discovered kinase GSK3 and a cyclooxygenase independent effect of the non-steroidal anti-inflammatory drug ibuprofen. Conclusions: Expression of tumor-related RAC1b in colorectal cancer depends critically on SRSF1 for the observed deregulation of alternative splicing during tumorigenesis and is controlled by upstream protein kinases that can be pharmacologically targeted.
- Diabetes: desigualdades socioeconómicos na população portuguesa em 2014Publication . Santos, Joana; Kislaya, Irina; Antunes, Liliana; Santos, Ana João; Rodrigues, Ana Paula; Neto, MarianaIntrodução: A Diabetes é considerada como um dos maiores problemas de saúde pública. Em Portugal, estima-se que a prevalência seja de 13.1% na população entre os 20-79 anos, dos quais cerca de 44% não tem conhecimento do diagnóstico. A par das diferenças observadas entre géneros (maior prevalência no sexo masculino) é necessário equacionar potenciais desigualdades socioeconómicas particularmente em grupos mais desfavorecidos. O objetivo deste estudo foi estimar a prevalência da diabetes auto-reportada por género, grupo etário, nível de escolaridade e determinar as desigualdades socioeconómicas na distribuição da diabetes na população adulta portuguesa. Métodos: Foram analisados dados de Inquérito Nacional de Saúde de 2014, cuja amostra é representativa da população residente e é constituída por 18204 indivíduos selecionados por amostragem probabilística, por grupos em três etapas, estratificada por região. Para este estudo a análise incidiu em indivíduos com 25 ou mais anos (n=16786). Calcularam-se estimativas da prevalência da diabetes total e estratificada por variáveis de caracterização socioeconómica. Para testar associações foi utilizada a estatística F-modificada variante do ajustamento de 2ª ordem do Qui-Quadrado de Rao-Scott. O grau de desigualdade socioeconómica foi estimado através do índice relativo de desigualdades e a curva de concentração. Todas as estimativas foram ponderadas para o desenho amostral. Resultados: Em 2014, a prevalência da diabetes em Portugal foi de 10,6% IC95%=[9,9%; 11,3%] sendo superior nos grupos etários de 55-64 anos 14,5% IC95%=[12,9%; 16,3%] e 65 e mais anos 23,5% IC95%=[21,7%; 25,1%], e na população com ensino básico 14,8% IC95%=[13,8%; 15,8%]. O índice relativo de desigualdade evidenciou desigualdades a favor dos grupos com um maior nível de escolaridade, sendo 0,33 IC95%=[0,19; 0,60] para os homens e 0,1 IC95%=[0,05; 0,21] para as mulheres. Conclusão: Os resultados evidenciam desigualdades de género e desigualdades educacionais. A desigualdade educacional na prevalência da diabetes é superior nas mulheres, o que sugere que a aposta na melhoria do nível de educação, em especial no sexo feminino poderá ter um efeito favorável na adoção de comportamentos mais saudáveis e consequente redução da carga da doença. A educação permanece um pilar central no desenvolvimento de intervenções para a promoção da saúde. O planeamento destas intervenções deve, por isso, prever as diferenças de género e o seu impacto na educação.
- Influenza surveillance during 2015/2016 season in PortugalPublication . Guiomar, Raquel; Pechirra, Pedro; Cristóvão, Paula; Costa, Inês; Conde, Patrícia; Pereira da Silva, Susana; Nunes, Baltazar; Rodrigues, Ana PaulaBackground: The National Influenza Surveillance Program ensures influenza epidemiological surveillance, integrating clinical and laboratory data. The clinical component describes the transmissibility over time, the virological component detects and characterizes influenza virus in circulation. Data is generated through 2 sentinel surveillance structures, the General Practitioner’s Sentinel Network (since 1990) and the Network of Emergency Units (since 1999). Here we provide the evaluation of 2015/2016 influenza season in Portugal. Methods: Influenza-Like illness (ILI) cases were reported to the National Influenza Reference Laboratory and to the Epidemiology Department of the National Institute of Health, in the context of the National Influenza Surveillance Program, from week 39/2015 through week 20/2016. The intensity and duration of the epidemic period were described based on the weekly ILI incidence rates. Nasopharyngeal swabs were collected for influenza and other respiratory viruses (RV: respiratory syncytial virus, adenovirus, rhinovirus, metapneumovirus, coronavirus, parainfluenza virus) for diagnosis and characterization. The detection of influenza and RV was performed by multiplex real-time RT-PCR. Influenza virus isolation, antigenic analysis (hemagglutination inhibition assay) and genetic characterization (HA1 gene segment) were performed. Results: During 2015/2016 season the influenza activity was low and the epidemic period occurred between week 1/2016 and 9/2016 with a maximum of 59.4 ILI cases per 105 inhabitants in week 3/2016. No impact on mortality from all causes was observed in any age group. 1097 nasopharyngeal swabs were tested for influenza and other respiratory viruses, influenza was detected in 444 (41%) and others respiratory viruses were found in 265 (24%). The influenza A(H1)pdm09 was predominant (91% of flu confirmed cases). Influenza B/ Victoria was identified sporadically (7% of flu cases) in late season. A(H1)pdm09 were predominantly detected in adults aged 15-64. Other respiratory viruses were detected in influenza negative cases, being rhinovirus (101; 38%) and coronavirus (78; 29%) found in higher frequencies. The antigenic and genetic analysis of circulating influenza A(H1)pdm09 showed similarity with vaccine strain. The majority of influenza B virus belonged to Victoria lineage and clade 1A, dissimilar from 2015/16 vaccine strain. Although few A(H3) viruses in circulation, almost all were similar to strain recommended for next season, 2016/2017, influenza vaccine. None of the 420 A(H1)pdm09 viruses analysed showed the H275Y substitution, correlated to high reduced susceptibility to oseltamivir. Conclusions: Influenza activity during 2015/2016 flu season was low, that can be linked with a higher influenza activity in last season and exceptional climatic conditions during the winter (higher temperature than usual). A(H1pdm)09 viruses were dominant, although in co-circulation with influenza B/Victoria. Situation that contrasts with European influenza picture, that showed a late peak of influenza B/ Victoria. Influenza A(H1)pdm09 detections reached the highest percentage in adults (15-64 years old). Most influenza detected viruses were similar to the 2015/2016 vaccine strains, although circulating influenza B/Victoria were from a different lineage comparing with vaccine strain. Observed mortality from all causes was within expected values during study period.
- Teor de iodo em alimentos: comparação entre os Açores e Portugal ContinentalPublication . Coelho, Inês; Delgado, Inês; Andrade, Pedro; Antunes, Carolina; Castanheira, Isabel; Calhau, Maria AntóniaNeste trabalho pretende-se contribuir com novos dados sobre o teor de iodo em alimentos disponíveis para consumo em Portugal. O iodo é um oligoelemento indispensável para a síntese das hormonas da tiroide. O seu papel assume particular importância em grávidas e crianças. Estudos clínicos em Portugal demonstraram a existência de uma carência generalizada das grávidas no país com particular severidade nos arquipélagos da Madeira e Açores [1]. Em relação às crianças em idade escolar, cerca de metade apresenta iodúrias abaixo do recomendado [2]. Assim, optou-se por comparar o teor de iodo de alimentos produzidos no arquipélago dos Açores com alimentos oriundos de Portugal Continental. Foram selecionadas amostras de lacticínios, frutas e produtos hortícolas de ambas as regiões em estudo. No total analisaram-se 110 amostras, em triplicado. Destas, 50 eram oriundas da região dos Açores e 60 de Portugal Continental. A metodologia escolhida para a determinação foi a espectrometria de massa com plasma indutivo acoplado (ICP-MS). O leite apresentou um maior teor de iodo em Portugal Continental (255 µg/l) do que nos Açores (133 µg/l). Para o queijo foi igualmente em Portugal Continental que se obteve teores de iodo mais elevados, 275 µg/kg, quando comparado com os Açores, 117 µg/kg. Todas as frutas analisadas apresentaram valores abaixo do limite de quantificação. Os resultados evidenciam que os lacticínios analisados são uma boa fonte de iodo e devem fazer parte das dietas de populações carenciadas. Os dados analíticos estão, também, alinhados com estudos recentes, evidenciando que os alimentos ricos em iodo representam um beneficio claro no aporte diário deste micronutriente, para a população em geral.
- Bioacessibilidade de Folatos em Quinoa: Efeito do ProcessamentoPublication . Motta, Carla; Assunção, Ricardo; Martins, Carla; Delgado, Inês; Coelho, Inês; Santos, Mariana; Torres, Duarte; Alvito, Paula; Castanheira, IsabelA quinoa é uma semente que pode ser usada e consumida como um cereal. Destaca-se pela elevada qualidade proteica, elevado teor em vitaminas e minerais e ausência de glúten 1. O folato ou vitamina B9 é um termo genérico que engloba as formas naturais e sintéticas, ácido fólico, tetrahidrofolato, 5-metil-tetrahidrofolato, 10-formil-tetrahidrofolato e 5-formil-tetrahidrofolato. O objetivo deste estudo foi avaliar a bioacessibilidade das diferentes formas de folatos em quinoa crua, cozida, cozida a vapor e maltada. Para a identificação e determinação dos folatos foi utilizado um cromatógrafo líquido acoplado a um espectrómetro de massas (UPLC-MS/MS). A digestão das amostras foi realizada, utilizando um protocolo de digestão estático, harmonizado "in vitro" 2. Os processos de cozedura aumentam a bioacessibilidade dos folatos, com valores de bioacessibilidade perto dos 100% para o 5-metil-tetrahidrofolato, verificando-se igualmente um aumento significativo da bioacessibilidade do 10-metil-tetrahidrofolato. Durante a maltagem a bioacessibilidade dos folatos também aumenta, com valores semelhantes aos da cozedura. O conhecimento da bioacessibilidade deste alimento, relacionada com o processamento culinário é uma indicação muito útil para cálculo da composição nutricional das dietas. A quinoa processada constitui uma fonte importante de folatos, e uma excelente alternativa nutricional, para a população em geral.