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- Programa Nacional de Vigilância da Gripe: relatório da época 2023/2024Publication . Guiomar, Raquel; Verdasca, Nuno; Gomes, Licínia; Henriques, Camila; Dias, Daniela; Lança, Miguel; Rodrigues, Ana Paula; Silva, Susana Pereira da; Direcção-Geral da Saúde (colaboração); Rede Portuguesa de Laboratórios para o Diagnóstico da Gripe e Outros Vírus Respiratórios (colaboração)Relatório anual do Programa Nacional de Vigilância da Gripe e de Outros Vírus Respiratórios (PNVG), que descreve a caraterização clínica e laboratorial da atividade gripal na época 2023/2024. O PNVG assegura a vigilância epidemiológica da gripe em Portugal, integrando as componentes de vigilância clínica e laboratorial. A componente clínica possibilita descrever a intensidade e evolução da epidemia no tempo. A componente virológica tem por base o diagnóstico laboratorial do vírus da gripe, SARS-CoV-2 e vírus sincicial respiratório (RSV) o que permite detetar e caraterizar os vírus respiratórios em circulação em cada inverno. As atividades do PNVG são desenvolvidas pelo Laboratório Nacional de Referência para o Vírus da Gripe e Outros Vírus Respiratórios do Departamento de Doenças Infeciosas e pelo Departamento de Epidemiologia do INSA, em colaboração com a Direção-Geral da Saúde.
- Effectiveness of the autumn 2023 COVID-19 vaccine dose in hospital-based healthcare workers: results of the VEBIS healthcare worker vaccine effectiveness cohort study, seven European countries, season 2023/24Publication . Savulescu, Camelia; Prats-Uribe, Albert; Brolin, Kim; Uusküla, Anneli; Bergin, Colm; Fleming, Catherine; Murri, Rita; Zvirbulis, Viesturs; Zavadska, Dace; Gaio, Vania; Popescu, Corneliu P.; Hrisca, Raluca; Cisneros, Maria; Latorre-Millán, Miriam; Lohur, Liis; McGrath, Jonathan; Ferguson, Lauren; De Gaetano Donati, Katleen; Abolina, Ilze; Gravele, Dagne; Machado, Ausenda; Florescu, Simin-Aysel; Lazar, Mihaela; Subirats, Pilar; Clusa Cuesta, Laura; Sui, Jacklyn; Kenny, Claire; Santangelo, Rosaria; Krievins, Dainis; Barzdina, Elza Anna; Valadas Henriques, Camila; Kosa, Alma Gabriela; Pohrib, Saftica-Mariana; Muñoz-Almagro, Carmen; Milagro, Ana; Bacci, Sabrina; Nardone, Anthony; VEBIS HCW VE study group; Collaborators in VEBIS HCW study groupCOVID-19 vaccination recommendations prioritise healthcare workers (HCWs), considering their exposure to severe acute respiratory coronavirus 2 (SARS-CoV-2) and their key role in the functioning of healthcare systems. In the European Union/European Economic Area (EU/EEA), HCWs were considered a priority for COVID-19 revaccination during the autumn 2023 campaign [1], and the World Health Organization (WHO) recommended revaccination of HCWs 12 months after their last dose [2]. Because the Omicron sub-lineage XBB.1.5 predominated in spring 2023, the COVID-19 vaccines were adapted to target this emerging strain, and the first XBB.1.5 vaccine was authorised for use in the EU/EEA in August 2023. Omicron BA.2.86/JN.1 emerged in the EU/EEA at the end of 2023, according to data available on the European Respiratory Virus Surveillance Summary (ERVISS) [3]. Evidence for COVID-19 vaccine recommendation in the HCW population remains scarce. Within the Vaccine Effectiveness, Burden and Impact (VEBIS) project, we aimed to measure the COVID-19 vaccine effectiveness (CVE) in HCWs, in the winter season 2023/24.
- Insights into macrolide resistance in Arcobacter butzleri: potential resistance mechanisms and impact on bacterial fitness and virulencePublication . Couto, Francisca; Martins, Inês; Vale, Filipa; Domingues, Fernanda; Oleastro, Mónica; Ferreira, SusanaBackground: Macrolides are recommended for treating the emerging enteropathogen Arcobacter butzleri; nonetheless, this bacterium often exhibits highly variable resistance rates, and the mechanisms behind this resistance phenotype remain largely unexplored. Objectives: To understand the phenotypic and genotypic consequences associated with the acquisition of erythromycin resistance in A. butzleri, as well as the effects on the fitness of this species. Methods: Resistant strains resulting from spontaneous mutations and adaptive laboratory evolution under increasing erythromycin concentrations were examined regarding their cross-resistance and collateral susceptibility profiles. Genetic causes of phenotypic antibiotic resistance were analysed by sequencing and bioinformatics, with functional correlation through ethidium bromide accumulation assays. Growth profiles in the presence and absence of erythromycin, motility and biofilm formation abilities were assessed to detect potential changes in fitness and virulence. Results: Clones from spontaneous mutation rate evolution demonstrated decreased susceptibility to erythromycin and other classes of antibiotics, associated with mutations in the transcriptional repressor areR, causing overexpression of the AreABC efflux pump. In turn, WGS analysis of the evolved strain showed additional mutations in the ribosomal proteins L4 and L22 and in the areR gene. Furthermore, the acquisition of macrolide resistance altered A. butzleri virulence and entailed a high biological cost. Conclusions: The findings of this study have proved that efflux activity contributes synergistically with mutations in the ribosomal proteins L4 and L22 to A. butzleri's high-level macrolide resistance. The results further suggest an impact on the bacterial physiology and virulence, with the increased fitness cost justifying the low worldwide prevalence of high-level resistant circulating strains.
- Screening and in silico characterization of prophages in Helicobacter pylori clinical strainsPublication . Ferreira, Rute; Pinto, Graça; Presa, Eva; Oleastro, Mónica; Silva, Catarina; Vieira, Luís; Sousa, Claúdia; Pires, Diana; Figueiredo, Ceu; Melo, LuísThe increase of antibiotic resistance calls for alternatives to control Helicobacter pylori, a Gram-negative bacterium associated with various gastric diseases. Bacteriophages (phages) can be highly effective in the treatment of pathogenic bacteria. Here, we developed a method to identify prophages in H. pylori genomes aiming at their future use in therapy. A polymerase chain reaction (PCR)-based technique tested five primer pairs on 74 clinical H. pylori strains. After the PCR screening, 14 strains most likely to carry prophages were fully sequenced. After that, a more holistic approach was taken by studying the complete genome of the strains. This study allowed us to identify 12 intact prophage sequences, which were then characterized concerning their morphology, virulence, and antibiotic-resistance genes. To understand the variability of prophages, a phylogenetic analysis using the sequences of all H. pylori phages reported to date was performed. Overall, we increased the efficiency of identifying complete prophages to 54.1 %. Genes with homology to potential virulence factors were identified in some new prophages. Phylogenetic analysis revealed a close relationship among H. pylori-phages, although there are phages with different geographical origins. This study provides a deeper understanding of H. pylori-phages, providing valuable insights into their potential use in therapy.
- Association of BMI, lipid-lowering medication, and age with prevalence of type 2 diabetes in adults with heterozygous familial hypercholesterolaemia: a worldwide cross-sectional studyPublication . Collaboration, European Atherosclerosis Society Familial Hypercholesterolaemia Studies (EAS FHSC)Background: Statins are the cornerstone treatment for patients with heterozygous familial hypercholesterolaemia but research suggests it could increase the risk of type 2 diabetes in the general population. A low prevalence of type 2 diabetes was reported in some familial hypercholesterolaemia cohorts, raising the question of whether these patients are protected against type 2 diabetes. Obesity is a well known risk factor for the development of type 2 diabetes. We aimed to investigate the associations of known key determinants of type 2 diabetes with its prevalence in people with heterozygous familial hypercholesterolaemia. Methods: This worldwide cross-sectional study used individual-level data from the EAS FHSC registry and included adults older than 18 years with a clinical or genetic diagnosis of heterozygous familial hypercholesterolaemia who had data available on age, BMI, and diabetes status. Those with known or suspected homozygous familial hypercholesterolaemia and type 1 diabetes were excluded. The main outcome was prevalence of type 2 diabetes overall and by WHO region, and in relation to obesity (BMI ≥30·0 kg/m2) and lipid-lowering medication as predictors. The study population was divided into 12 risk categories based on age (tertiles), obesity, and receiving statins, and the risk of type 2 diabetes was investigated using logistic regression. Findings: Among 46 683 adults with individual-level data in the FHSC registry, 24 784 with heterozygous familial hypercholesterolaemia were included in the analysis from 44 countries. 19 818 (80%) had a genetically confirmed diagnosis of heterozygous familial hypercholesterolaemia. Type 2 diabetes prevalence in the total population was 5·7% (1415 of 24 784), with 4·1% (817 of 19 818) in the genetically diagnosed cohort. Higher prevalence of type 2 diabetes was observed in the Eastern Mediterranean (58 [29·9%] of 194), South-East Asia and Western Pacific (214 [12·0%] of 1785), and the Americas (166 [8·5%] of 1955) than in Europe (excluding the Netherlands; 527 [8·0%] of 6579). Advancing age, a higher BMI category (obesity and overweight), and use of lipid-lowering medication were associated with a higher risk of type 2 diabetes, independent of sex and LDL cholesterol. Among the 12 risk categories, the probability of developing type 2 diabetes was higher in people in the highest risk category (aged 55-98 years, with obesity, and receiving statins; OR 74·42 [95% CI 47·04-117·73]) than in those in the lowest risk category (aged 18-38 years, without obesity, and not receiving statins). Those who did not have obesity, even if they were in the upper age tertile and receiving statins, had lower risk of type 2 diabetes (OR 24·42 [15·57-38·31]). The corresponding results in the genetically diagnosed cohort were OR 65·04 (40·67-104·02) for those with obesity in the highest risk category and OR 20·07 (12·73-31·65) for those without obesity. Interpretation: Adults with heterozygous familial hypercholesterolaemia in most WHO regions have a higher type 2 diabetes prevalence than in Europe. Obesity markedly increases the risk of diabetes associated with age and use of statins in these patients. Our results suggest that heterozygous familial hypercholesterolaemia does not protect against type 2 diabetes, hence managing obesity is essential to reduce type 2 diabetes in this patient population.
- Impact of gold nanoparticle exposure on genetic materialPublication . Ramos-Pan, Lucía; Touzani, Assia; Fernández-Bertólez, Natalia; Fraga, Sónia; Laffon, Blanca; Valdiglesias, VanessaMetal nanoparticles, with gold nanoparticles (AuNP) at the forefront, have gained immense attention due to their unique properties. At the nanoscale, gold exhibits remarkable physical, chemical, optical, and electronic features, making it ideal for a plethora of applications, including bioimaging, sensing, diagnostics, and drug delivery. Despite their promising utility, concerns have arisen regarding the potential adverse effects of AuNP on human health. Research has indicated that these nanoparticles can accumulate in vital organs and interact with proteins and cellular structures, potentially leading to diverse toxicological manifestations. The precise understanding of these nano-bio interactions is further complicated by the varied physicochemical properties of AuNP that influence their biological effects. This review aims to consolidate the current knowledge on the genotoxic effects of AuNP, shedding light on the underlying mechanisms and factors affecting their toxicity. The search was conducted in PubMed and Web of Science databases. Eventually, 32 studies focusing on the genotoxic effects of AuNP were included in the review. In vitro and in vivo findings revealed that AuNP can induce primary DNA damage, oxidative DNA damage, chromosomal damage, alterations in gene expression, and effects on epigenetic regulation. These effects were found to be influenced by various factors, including nanoparticle size, shape, and surface coating. However, the existing literature also highlights the challenges associated with assessing the genotoxicity of nanomaterials (NM), emphasizing the need for standardized and adapted testing protocols. The interference of nanoparticles with conventional toxicity assays may lead to unreliable results; thus, specific methodologies tailored for NM evaluation must be implemented. In conclusion, while AuNP hold tremendous potential for innovative applications, their safety profile remains a critical concern. Continued research is imperative to elucidate the mechanisms of AuNP induced genotoxicity and develop robust testing protocols, ensuring their safe and effective use in diverse applications.
- Rainfall events and daily mortality across 645 global locations: two stage time series analysisPublication . He, Cheng; Breitner-Busch, Susanne; Huber, Veronika; Chen, Kai; Zhang, Siqi; Gasparrini, Antonio; Bell, Michelle; Kan, Haidong; Royé, Dominic; Armstrong, Ben; Schwartz, Joel; Sera, Francesco; Vicedo-Cabrera, Ana Maria; Honda, Yasushi; Jaakkola, Jouni J.K.; Ryti, Niilo; Kyselý, Jan; Guo, Yuming; Tong, Shilu; de’Donato, Francesca; Michelozzi, Paola; Coelho, Micheline de Sousa Zanotti Staglior; Saldiva, Paulo Hilario Nascimento; Lavigne, Eric; Orru,Hans; Indermitte, Ene; Pascal, Mathilde; Goodman, Patrick; Zeka, Ariana; Kim, Yoonhee; Diaz, Magali Hurtado; Arellano, Eunice Elizabeth Félix; Overcenco, Ala; Klompmaker, Jochem; Rao, Shilpa; Palomares, Alfonso Diz-Lois; Carrasco, Gabriel; Seposo, Xerxes; das Neves Pereira da Silva, Susana; Joana Madureira; Holobaca, Iulian-Horia; Scovronick, Noah; Acquaotta, Fiorella; Kim, Ho; Lee, Whanhee; Hashizume, Masahiro; Tobias, Aurelio; Íñiguez, Carmen; Forsberg, Bertil; Ragettli, Martina S.; Guo, Yue Leon; Pan, Shih-Chun; Osorio, Samuel; Li, Shanshan; Zanobetti, Antonella; Dang, Tran Ngoc; Dung, Do Van; Schneider. AlexandraObjective: To examine the associations between characteristics of daily rainfall (intensity, duration, and frequency) and all cause, cardiovascular, and respiratory mortality. Design: Two stage time series analysis. Setting: 645 locations across 34 countries or regions. Population: Daily mortality data, comprising a total of 109 954 744 all cause, 31 164 161 cardiovascular, and 11 817 278 respiratory deaths from 1980 to 2020. Main outcome measure: Association between daily mortality and rainfall events with return periods (the expected average time between occurrences of an extreme event of a certain magnitude) of one year, two years, and five years, with a 14 day lag period. A continuous relative intensity index was used to generate intensity-response curves to estimate mortality risks at a global scale. Results: During the study period, a total of 50 913 rainfall events with a one year return period, 8362 events with a two year return period, and 3301 events with a five year return period were identified. A day of extreme rainfall with a five year return period was significantly associated with increased daily all cause, cardiovascular, and respiratory mortality, with cumulative relative risks across 0-14 lag days of 1.08 (95% confidence interval 1.05 to 1.11), 1.05 (1.02 to 1.08), and 1.29 (1.19 to 1.39), respectively. Rainfall events with a two year return period were associated with respiratory mortality only, whereas no significant associations were found for events with a one year return period. Non-linear analysis revealed protective effects (relative risk <1) with moderate-heavy rainfall events, shifting to adverse effects (relative risk >1) with extreme intensities. Additionally, mortality risks from extreme rainfall events appeared to be modified by climate type, baseline variability in rainfall, and vegetation coverage, whereas the moderating effects of population density and income level were not significant. Locations with lower variability of baseline rainfall or scarce vegetation coverage showed higher risks. Conclusion: Daily rainfall intensity is associated with varying health effects, with extreme events linked to an increasing relative risk for all cause, cardiovascular, and respiratory mortality. The observed associations varied with local climate and urban infrastructure.
- A Comparative Overview of the Role of Human Ribonucleases in Nonsense-Mediated mRNA DecayPublication . da Costa, Paulo J.; Menezes, Juliane; Guedes, Raquel; Reis, Filipa P.; Teixeira, Alexandre; Saramago, Margarida; Viegas, Sandra C.; Arraiano, Cecília M.; Romão, LuísaEukaryotic cells possess surveillance mechanisms that detect and degrade defective transcripts. Aberrant transcripts include mRNAs with a premature termination codon (PTC), targeted by the nonsense-mediated decay (NMD) pathway, and mRNAs lacking a termination codon, targeted by the nonstop decay (NSD) pathway. The eukaryotic exosome, a ribonucleolytic complex, plays a crucial role in mRNA processing and turnover through its catalytic subunits PM/Scl100 (Rrp6 in yeast), DIS3 (Rrp44 in yeast), and DIS3L1. Additionally, eukaryotic cells have other ribonucleases, such as SMG6 and XRN1, that participate in RNA surveillance. However, the specific pathways through which ribonucleases recognize and degrade mRNAs remain elusive. In this study, we characterized the involvement of human ribonucleases, both nuclear and cytoplasmic, in the mRNA surveillance mechanisms of NMD and NSD. We performed knockdowns of SMG6, PM/Scl100, XRN1, DIS3, and DIS3L1, analyzing the resulting changes in mRNA levels of selected natural NMD targets by RT-qPCR. Additionally, we examined the levels of different human β-globin variants under the same conditions: wild-type, NMD-resistant, NMD-sensitive, and NSD-sensitive. Our results demonstrate that all the studied ribonucleases are involved in the decay of certain endogenous NMD targets. Furthermore, we observed that the ribonucleases SMG6 and DIS3 contribute to the degradation of all β-globin variants, with an exception for βNS in the former case. This is also the case for PM/Scl100, which affects all β-globin variants except the NMD-sensitive variants. In contrast, DIS3L1 and XRN1 show specificity for β-globin WT and NMD-resistant variants. These findings suggest that eukaryotic ribonucleases are target-specific rather than pathway-specific. In addition, our data suggest that ribonucleases play broader roles in mRNA surveillance and degradation mechanisms beyond just NMD and NSD.
- Phenotypic and Genotypic Characterization of Escherichia coli and Salmonella spp. Isolates from Pigs at Slaughterhouse and from Commercial Pork Meat in PortugalPublication . Gonçalves, Carlota; Silveira, Leonor; Rodrigues, João; Furtado, Rosália; Ramos, Sónia; Nunes, Alexandra; Pista, ÂngelaBackground: Foodborne diseases are a serious public health concern, and food-producing animals are a major source of contamination. Methods: The present study analysed Escherichia coli and Salmonella spp. isolated from faecal samples of 100 fattening pigs and from 52 samples of pork meat. Results: The results showed that the majority of the analysed meat samples were considered satisfactory in terms of microbiological quality (92.3% for E. coli and 94.2% for Salmonella spp.). Salmonella spp. was identified in 5.8% of the meat samples, whereas E. coli was detected in 89.5% of all samples (69.2% in meat and 100% in faecal samples). Furthermore, 1.9% of the faecal samples contained Shiga-toxin-producing E. coli and 3.9% contained enterotoxigenic E. coli. All sequenced isolates presented virulence genes for extraintestinal pathogenic E. coli. Moreover, 75.0% of E. coli isolates from meat and 71.8% from faeces samples showed antibiotic resistance, with 40.7% and 51.4%, respectively, being multidrug-resistant (MDR). The most prevalent resistances were to tetracycline, ampicillin, and sulfamethoxazole, and one E. coli isolate showed resistance to extended-spectrum β-lactamase. Conclusions: This study highlights the role of pigs as a potential source of human contamination and the importance of a One Health approach to ensure food safety and to promote public health.
- Toscana Virus in Wild-Caught Sand Flies in Portugal, Findings from the National Vector Surveillance Network, 2023Publication . Amaro, Fátima; Zé-Zé, Líbia; Osório, Hugo Costa; Soares, Patrícia; Silva, Manuel; Freitas, Inês Campos; Alves, Maria JoãoPhlebotomine sand flies play a crucial role in both human and veterinary medicine, acting as vectors for Leishmania parasites and most known phleboviruses. In Portugal, the REVIVE program, a comprehensive national surveillance network under the Ministry of Health, has included sand fly surveys since 2016. REVIVE aims to identify existing sand fly species in the country, determine which pathogens are circulating among them, and provide actionable insights for prevention and control measures when necessary. In this way, annually, from May to October, health technicians collect sand flies across mainland Portugal with CDC light traps. The collected sand flies are sent to the Centre for Vectors and Infectious Diseases Research for species identification and molecular screening of pathogens. On 21 September 2023, Toscana virus (TOSV), a well-known phlebovirus in the Mediterranean region due to its capacity to cause neurological disease, was detected in a pool of 30 sand flies collected in Algarve, the southernmost region of Portugal. A 668 bp partial sequence of the nucleoprotein gene shows similarity with TOSV strains from Spain. To our knowledge, this is the first detection of TOSV in its vector in this country, having previously only been reported in vertebrate hosts. These findings highlight the important role of ongoing surveillance efforts in monitoring and understanding the dynamics of sand fly-borne diseases in Portugal.
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