Browsing by Author "Sampaio, Daniel"
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- A large outbreak of Legionnaires’ Disease in an industrial town in PortugalPublication . George, Francisco; Shivaji, Tara; Pinto, Catia Sousa; Serra, Luis Antonio Oliveira; Valente, João; Albuquerque, Maria João; Vicêncio, Paula Cristina Olivença; San-Bento, Ana; Diegues, Paulo; Nogueira, Paulo Jorge; Marques, Teresa; Rebelo, Helena; Costa, Filipa; Rodrigues, Raquel; Nunes, Alexandra; Borges, Vitor; Gomes, João Paulo; Sampaio, Daniel; Barreiro, Paula; Duarte, Silvia; Carpinteiro, Dina; Mendonça, Joana; Silva, Catarina; Vieira, Luís; Simões, Maria Joao; Gonçalves, Paulo; Nunes, Baltazar; Dias, Carlos; Machado, Jorge; Almeida, Fernando; Goncalves, Elsa A; Carvalho, Lucilia; Viterbo, Pedro; Jardim, Dilia; Lacasta, Nuno; Boavida, Filomena; Perez, Ana; Santana, Isabel; Matias, Paula; Banza, Nuno; Rabacal, CarlosBackground: We describe the investigation and control of an outbreak of Legionnaires’ disease in Portugal in October, November and December 2014. Methods: Confirmed cases were individuals with pneumonia, laboratory evidence of Legionella pneumophila serogroup 1 and exposure, by residence, occupational or leisure to the affected municipalities. 49 possible sources were reduced to four potential sources, all industries with wet cooling system, following risk assessment. We geo-referenced cases’ residences and the location of cooling towers defining four study areas 10km buffer centered on each cooling tower system. We compared the number of cases with expected numbers, calculated from the outbreak's attack rates applied to 2011 census population. Using Stones’ Test, we tested observed to expected ratios for decline in risk, with distance up to 10km four directions. Isolates of Legionella pneumophila were compared using molecular methods. Results: We identified 403 cases, 377 of which were confirmed, 14 patients died. Patients became ill between 14 October and 2 December. A NE wind and thermal inversion were recorded during the estimated period of exposure. Disease risk was highest in people living south west from all of the industries identified and decreased with distance (p<0.001). 71 clinical isolates demonstrated an identical SBT profile to an isolate from a cooling tower. Whole genome sequencing identified an unusual L. pneumophila subsp. fraseri serogroup 1 as the outbreak causative strain, and confirmed isolates’ relatedness. Conclusions: Industrial wet cooling systems, bacteria with enhanced survival characteristics and a combination of climatic conditions contributed to the second largest outbreak of Legionnaires’ disease recorded internationally.
- Phylogenomic characterization of the causative strain of one of the largest worldwide outbreaks of Legionnaires’ disease occurred in Portugal in 2014Publication . Borges, Vítor; Nunes, Alexandra; Sampaio, Daniel; Vieira, Luís; Gomes, João PauloBackground: Legionnaires’ disease (LD) is a severe pneumonia with a case fatality rate of 8–12%. Transmission usually occurs by inhalation of aerosols or aspiration of water containing Legionella spp, where L. pneumophila serogroup 1 accounts for most of the occasional outbreaks. The world’s second largest outbreak of LD occurred in 2014 in Vila Franca de Xira, Portugal, yielding more than 400 cases and 14 deaths. The L. pneumophila causative strain is from serogroup 1 and displays the novel sequence type (ST) ST1905. Here, we described how whole-genome sequencing (WGS) / Bioinformatics was used for real-time investigation of this LD outbreak as well as for integrating the genetic backbone of the causative strain in the frame of the species phylogeny and diversity.
- Sequenciação de nova geração: o paradigma dos parasitasPublication . Vilares, Anabela; Borges, Vítor; Sampaio, Daniel; Vieira, Luís; Ferreira, Idalina; Martins, Susana; Reis, Tânia; Gomes, João Paulo; Gargaté, Maria JoãoToxoplasma gondii é um protozoário intracelular obrigatório responsável pela Toxoplasmose, infetando diferentes espécies incluindo o Homem. Na Europa e no Norte da América, T. gondii tem uma população clonal constituída por tipo I, II, III. No entanto, estudos recentes têm descrito uma maior diversidade genética, com a identificação de estirpes recombinantes e atípicas mais virulentas. Em Portugal, o conhecimento dos genótipos circulantes de T. gondii é limitado. Neste sentido, estudou-se a diversidade genética de 68 estirpes, incluindo 51 estirpes de T. gondii isoladas em Portugal, a estirpe de referência RH e 16 outras estirpes de referência. Neste estudo realizou-se a genotipagem clássica (Sag2), PCR multiplex de 5 microssatélites e uma combinação de sequenciações por Sanger e NGS de oito loci responsáveis pela virulência. A genotipagem clássica (tipo I, II, III) foi obtida em 100% das estirpes e mostrou que a maioria das estirpes isoladas em Portugal eram do tipo II (70,6%; 36/51) e as restantes 15 do tipo I. A análise de microssatélites por PCR multiplex aumentou o poder discriminatório em apenas mais um tipo identificado. Contudo, quando se adicionaram as metodologias de sequenciação por Sanger e NGS o poder discriminatório aumentou para 36 tipos diferentes. A combinação destas metodologias (Sanger/NGS) permitiram a identificação de uma estrutura mosaico nos isolados de T. gondii em Portugal, não conseguida anteriormente por qualquer uma das outras tecnologias utilizadas.
- Towards a rapid sequencing-based molecular surveillance and mosaicism investigation of Toxoplasma gondiiPublication . Vilares, Anabela; Borges, Vítor; Sampaio, Daniel; Ferreira, Idalina; Martins, Susana; Vieira, Luis; Gargaté, Maria João; Gomes, João PauloAdvances in molecular epidemiology of Toxoplasma gondii are hampered by technical and cost-associated hurdles underlying the acquisition of genomic data from parasites. In order to implement an enhanced genotyping approach for molecular surveillance of T. gondii, we applied a multi-locus amplicon-based sequencing strategy to samples associated with human infection. This approach, targeting genome-dispersed polymorphic loci potentially involved in adaptation and virulence, genetically discriminated almost all 68 studied strains and revealed a scenario of marked genomic mosaicism. Two-thirds (n = 43) of all strains were classified as recombinant, although recombination seemed to be linked to the classical archetypal lineage. While 92% of the Sag2 archetype I strains revealed genetic mosaicism, only 45% of Sag2 archetype II strains were identified as recombinant. Contrarily to the virulence-associated archetype I, most type II strains (regardless of their recombination background) were non-virulent in mouse. Besides Sag2, some of the newly studied loci (namely the type I/I-like alleles of Sag1, B17, PK1, and Sag3 and type III/III-like alleles of TgM-A) constitute promising candidates to rapidly infer T. gondii mouse virulence. Our successful attempt to capture microsatellite length variation launches good perspectives for the straightforward transition from the laborious intensive historical method to more informative next-generation sequencing (NGS)/bioinformatics-based methodologies. Overall, while T. gondii whole-genome sequencing will be hardly feasible in most laboratories, this study shows that a discrete loci panel has the potential to improve the molecular epidemiology of T. gondii towards a better monitoring of circulating genotypes with clinical importance.