Browsing by Author "Rebelo-de-Andrade, Helena"
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- Actividade gripal em Portugal no Inverno de 2000/2001 - Análise antigénica e genética das estirpes de vírus influenzaPublication . Pechirra, Pedro; Rebelo-de-Andrade, Helena; Guiomar, Raquel; Ribeiro, Carlos; Coelho, Anabela; Pedro, Sónia; George, FranciscoAs infecções por vírus influenza são uma importante causa de morbilidade em todos os grupos etários e estão associados a uma elevada mortalidade nos idosos e nos indivíduos pertencentes a grupos de risco. No presente estudo analisaram-se os dados da vigilância epidemiológica da gripe durante o Inverno de 2000/2001. Os dados clínicos, epidemiológicos e virológicos referentes aos casos de síndroma gripal foram recolhidos através do Programa Nacional de Vigilância da Gripe que se desenvolve em colaboração com a Direcção Geral da Saúde e integra a informação obtida a partir das redes Médicos-Sentinela e Serviços de Urgência. A análise dos dados recolhidos mostram que, durante a época de Inverno de 2000/2001, a actividade gripal foi baixa, sendo o período epidémico curto e de pequena intensidade e duração. As taxas de incidência do síndrome gripal subiram acima da linha de base durante três semanas e não ultrapassaram os 74 casos por 100 000 habitantes. Os vírus influenza do tipo B foram predominantes, verificando-se a presença simultânea de vírus influenza tipo AH1 e AH3. A caracterização antigénica e genética das estirpes isoladas permitiu confirmar a semelhança entre estas estirpes virais e as estirpes vacinais e detectar a extensão dos drifts antigénicos. Saliente-se que apesar da maioria das estirpes de vírus influenza B serem idênticas antigenicamente à estirpe vacinal, a caracterização genética mostrou uma evolução dirigida para a estirpe B/ Sichuan/379/99 que viria a integrar a vacina antigripal em 2001/2002. Consequentemente, observou-se a co-circulação de duas linhagens diferentes evidenciada pela análise filogenética das estirpes B isoladas no nosso país.
- Adaptive evolution of PB1 from influenza A(H1N1)pdm09 virus towards an enhanced fitnessPublication . Santos, Luís A.; Almeida, Filipe; Gíria, Marta; Trigueiro-Louro, João; Rebelo-de-Andrade, HelenaPB1 influenza virus retain traces of interspecies transmission and adaptation. Previous phylogenetic analyses highlighted mutations L298I, R386K and I517V in PB1 to have putatively ameliorated the A(H1N1)pdm09 adaptation to the human host. This study aimed to evaluate the reversal of these mutations and infer the role of these residues in the virus overall fitness and adaptation. We generate PB1-mutated viruses introducing I298L, K386R and V517I mutations in PB1 and evaluate their phenotypic impact on viral growth and on antigen yield. We observed a decrease in viral growth accompanied by a reduction in hemagglutination titer and neuraminidase activity, in comparison with wt. Our data indicate that the adaptive evolution occurred in the PB1 leads to an improved overall viral fitness; and such biologic advantaged has the potential to be applied to the optimization of influenza vaccine seed prototypes.
- Antibody response to the influenza vaccine in healthcare workersPublication . Sacadura-Leite, Ema; Sousa-Uva, António; Rebelo-de-Andrade, HelenaPeople vaccinated against influenza develop hemagglutination inhibition (HAI) antibodies (Ab) that bind to the virus and neutralize it. Ab titer levels are variable depending on factors insufficiently studied, and tend to decrease over time. In the present study, we analyzed antibodies responses before and after influenza vaccination in nurses working in a hospital, with the aim of: -identifying seroconversion rates to trivalent influenza vaccine one month after immunization; -evaluating if, six months after vaccination, influenza HAI Ab titer fall comparing to one-month post vaccination HAI Ab titer; -studying the association between the lack of HAI Ab response (seroconversion) assessed one month after immunization and: ∘past influenza vaccinations, ∘baseline (before vaccination) HAI Ab titer, ∘baseline (before vaccination) HAI Ab titer ≥ 40 (considered as a protection titer). Hemagglutination inhibition reaction was used to assess specific HAI Ab titers against influenza A(H1N1), A(H3N2) and B virus strains included in the influenza vaccine and we used progressive dilutions of two times, starting on 1:10 until 1:20.480. Seroconversion rates, one month after vaccination, were 66.7% for A(H1N1) strain, 63.2% for A(H3N2) strain and 56.3% for B strain. The most immunogenic strain used in 2007/08 influenza vaccine was A(H1N1). Seroconversion rates after one month were negatively associated with past influenza vaccinations, baseline HAI Ab titers ≥ 40 and baseline HAI Ab titers. Six months after vaccination, 50% of participants showed a drop in HAI Ab titers to an half, for each of the considered strains, but they remain high enough to protect against the disease.
- Antiviral drug profile of seasonal influenza viruses circulating in Portugal from 2004/2005 to 2008/2009 winter seasonPublication . Correia, Vanessa; Rebelo-de-Andrade, Helena; Santos, Luís A.; Lackanby, Angie; Zambon, MariaA research project on antiviral drug resistance of influenza viruses circulating in Portugal has been carried out since 2007. Here, the first results obtained regarding the evaluation of susceptibility to amantadine and oseltamivir are presented. Information about antiviral prescription and exposure was available through the National Influenza Surveillance Programme. Amantadine susceptibility was evaluated by pyrosequencing for known resistance markers on 178 influenza A strains from 2004/2005 to 2006/2007. Susceptibility to oseltamivir was evaluated by 50% inhibitory concentration determination on 340 virus strains from 2004/2005 to 2008/2009, 134 of which were further analyzed by sequencing of the neuraminidase gene. This study revealed that influenza antiviral drugs were rarely prescribed at national level. Resistance to amantadine was observed on only A(H3N2) strain isolated during 2005/2006 and on 38 (74.5%) of the 51 A(H3N2) strains from 2006/2007, all carrying the mutation S31N in theirM2sequence. Oseltamivir resistance was observed in 6 (20.7%) of the 29 A(H1N1) strains from 2007/2008 and in all strains from 2008/2009, which exhibited extremely high IC50 values and carrying the mutation H275Y in their neuraminidase sequence. The national data generated and analyzed in this study may contribute to increase the knowledge on influenza antiviral drug resistance which is a problem of global concern.
- Characterization of influenza A/Fujian/411/2002(H3N2)-like viruses isolated in Portugal between 2003 and 2005Publication . Pechirra, Pedro; Gonçalves, Paulo; Arraiolos, Ana; Coelho, Anabela; Rebelo-de-Andrade, HelenaIn Portugal, influenza surveillance is achieved through the National Influenza Surveillance Programme (NISP), in close collaboration with other European and global surveillance networks. The NISP integrates epidemiological, clinical and virological data based on the information collected by a Network of Sentinel Medical Practitioners and by a network of Emergency Units of Hospitals and Health Care Centres. In this study, genetic and antigenic characterization of influenza A viruses of the A/Fujian/411/2002 lineage, isolated during the 2003/2004 and 2004/2005 influenza winter seasons, in the context of the NISP, are described. Antigenic analysis of A/ Fujian/411/2002-like viruses, first detected and isolated during the 2003/2004 winter season, revealed a close similarity with the reference strains A/Kumamoto/102/2002 and A/Wyoming/ 3/2003. Genetic analysis confirmed this similarity and revealed two different phylogenetic branches. The 2004/2005 influenza A(H3) isolates formed, both antigenic and genetically, a more homogeneous group and were closely related to A/Oslo/807/2004 and A/California/7/2004. During this season, the characterization of the influenza viral strains has shown continuous evolution to variants close related to A/Oslo/807/2004. The majority of amino acid substitutions detected in the haemagglutinin occurred at antigenic sites. This study reflects the contribution of individual countries for the surveillance and knowledge of the molecular epidemiology of the infection, essential for a concerted action towards the global monitoring of the disease. It also reflects the importance of constant monitoring of genetic and antigenic characteristics of circulating influenza strains, which will certainly be a major contribution to the formulation of influenza vaccines.
- Exploring new antiviral targets for influenza and COVID-19: Mapping promising hot spots in viral RNA polymerasesPublication . Figueiredo-Nunes, Inês; Trigueiro-Louro, João; Rebelo-de-Andrade, HelenaInfluenza and COVID-19 are infectious respiratory diseases that represent a major concern to public health with social and economic impact worldwide, for which the available therapeutic options are not satisfactory. The RdRp has a central role in viral replication and thus represents a major target for the development of antiviral approaches. In this study, we focused on Influenza A virus PB1 polymerase protein and the betacoronaviruses nsp12 polymerase protein, considering their functional and structural similarities. We have performed conservation and druggability analysis to map conserved druggable regions, that may have functional or structural importance in these proteins. We disclosed the most promising and new targeting regions for the discovery of new potential polymerase inhibitors. Conserved druggable regions of putative interaction with favipiravir and molnupiravir were also mapped. We have also compared and integrated the current findings with previous research.
- Genomic signatures and antiviral drug susceptibility profile of A(H1N1)pdm09Publication . Giria, Marta; Rebelo-de-Andrade, Helena; Santos, Luís; Martins Correia, Vanessa; Vicente, Sónia; Almeida Santos, MadalenaBACKGROUND: Genetic changes in influenza surface and internal genes can alter viral fitness and virulence. Mutation trend analysis and antiviral drug susceptibility profiling of A(H1N1)pdm09 viruses is essential for risk assessment of emergent strains and disease management. OBJECTIVE: To profile genomic signatures and antiviral drug resistance of A(H1N1)pdm09 viruses and to discuss the potential role of mutated residues in human host adaptation and virulence. STUDY DESIGN: A(H1N1)pdm09 viruses circulating in Portugal during pandemic and post-pandemic periods and 2009/2010 season. Viruses were isolated in MDCK-SIAT1 cell culture and subjected to mutation analysis of surface and internal proteins, and to antiviral drug susceptibility profiling. RESULTS: The A(H1N1)pdm09 strains circulating during the epidemic period in Portugal were resistant to amantadine. The majority of the strains were found to be susceptible to oseltamivir and zanamivir, with five outliers to neuraminidase inhibitors (NAIs) identified. Specific mutation patterns were detected within the functional domains of internal proteins PB2, PB1, PA, NP, NS1, M1 and NS2/NEP, which were common to all isolates and also some cluster-specific. DISCUSSION: Modification of viral genome transcription, replication and apoptosis kinetics, changes in antigenicity and antiviral drug susceptibility are known determinants of virulence. We report several point mutations with putative roles in viral fitness and virulence, and discuss their potential to result in more virulent phenotypes. Monitoring of specific mutations and genetic patterns in influenza viral genes is essential for risk assessing emergent strains, disease epidemiology and public health implications.
- Global update on the susceptibility of human influenza viruses to neuraminidase inhibitors, 2014-2015Publication . Hurt, Aeron C.; Besselaar, Terry G.; Daniels, Rod S.; Ermetal, Burcu; Fry, Alicia; Gubareva, Larisa; Huang, Weijuan; Lackenby, Angie; Lee, Raphael T.C.; Lo, Janice; Maurer-Stroh, Sebastian; Nguyen, Ha T.; Pereyaslov, Dmitriy; Rebelo-de-Andrade, Helena; Siqueira, Marilda M.; Takashita, Emi; Tashiro, Masato; Tilmanis, Danielle; Wang, Dayan; Zhang, Wenqing; Meijer, AdamThe World Health Organization (WHO) Collaborating Centres for Reference and Research on Influenza (WHO CCs) tested 13,312 viruses collected by WHO recognized National Influenza Centres between May 2014 and May 2015 to determine 50% inhibitory concentration (IC50) data for neuraminidase inhibitors (NAIs) oseltamivir, zanamivir, peramivir and laninamivir. Ninety-four per cent of the viruses tested by the WHO CCs were from three WHO regions: Western Pacific, the Americas and Europe. Approximately 0.5% (n = 68) of viruses showed either highly reduced inhibition (HRI) or reduced inhibition (RI) (n = 56) against at least one of the four NAIs. Of the twelve viruses with HRI, six were A(H1N1)pdm09 viruses, three were A(H3N2) viruses and three were B/Yamagata-lineage viruses. The overall frequency of viruses with RI or HRI by the NAIs was lower than that observed in 2013-14 (1.9%), but similar to the 2012-13 period (0.6%). Based on the current analysis, the NAIs remain an appropriate choice for the treatment and prophylaxis of influenza virus infections.
- Heterogeneous Selective Pressure Acting on Influenza B Victoria- and Yamagata-Like HemagglutininsPublication . Nunes, Baltazar; Pechirra, Pedro; Coelho, Anabela; Ribeiro, Carlos; Arraiolos, Ana; Rebelo-de-Andrade, HelenaAs a consequence of immune pressure, influenza virus hemagglutinin presents some of its amino acids under positive selection. Several authors have reported the existence of influenza A hemagglutinin codons under positive selective pressure (PSP). In this framework, the present work objectives were to demonstrate the presence of PSP and evaluate its effects on Victoria- and Yamagata-like influenza B viruses. Methodology adopted consisted in estimating the acceptance rate of nonsynonymous substitutions (ω = dN/dS) that describe the strength of selective pressure and identifying codons that may be positively selected, applying a set of continuous-time Markov chain codon-substitution models. Two groups of HA1 sequences (140 from Yamagata and 60 from Victoria lineage) were used. All the model maximum-likelihood estimates were obtained using codeml software application (PAML 3.15). The hypothesis of no existence of sites under PSP was rejected for both lineages (p<0.001), using likelihood ratio tests. These results demonstrate the presence of positive selection acting on hemagglutinin of both Yamagata- and Victoria-like influenza B viruses. Several different sites were identified to be under PSP on Yamagata and Victoria hemagglutinins. Sites found with a posterior probability >0.95 were codons 197 and 199 in both lineages, codon 75 in the Yamagata lineage, and codon 129 in the Victoria lineage. The detected amino acids are located at or near antigenic sites in influenza A virus H3 hemagglutinin.
- How to dissect viral infections and their interplay with the host-proteome by immunoaffinity and mass spectrometry: A tutorialPublication . Santos, Hugo M.; Carvalho, Luís B.; Lodeiro, Carlos; Martins, Gonçalo; Gomes, Inês L.; Antunes, Wilson D.T.; Correia, Vanessa; Almeida-Santos, Maria M.; Rebelo-de-Andrade, Helena; Matos, António P.A.; Capelo, J.L.The capabilities of bioanalytical mass spectrometry to (i) detect and differentiate viruses at the peptide level whilst maintaining high sample throughput and (ii) to provide diagnosis and prognosis for infected patients are presented as a tutorial in this work to aid analytical chemists and physicians to gain insights into the possibilities offered by current high-resolution mass spectrometry technology and bioinformatics. From (i) sampling to sample treatment; (ii) Matrix-Assisted Laser Desorption Ionization- to Electrospray Ionization -based mass spectrometry; and (iii) from clustering to peptide sequencing; a detailed step-by-step guide is provided and exemplified using SARS-CoV-2 Spike Y839 variant and the variant of concern SARS-CoV-2 Alpha (B.1.1.7 lineage), Influenza B, and Influenza A subtypes AH1N1pdm09 and AH3N2.