DDI - Apresentações orais em encontros internacionais
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- Multi-country and intersectoral assessment of cluster congruence between pipelines for genomics surveillance of foodborne pathogensPublication . Mixão, Verónica; Pinto, Miguel; Brendebach, Holger; Sobral, Daniel; Santos, João Dourado; Radomski, Nicolas; Uldall, Anne Sophie Majgaard; Bomba, Arkadiusz; Pietsch, Michael; Bucciacchio, Andrea; de Ruvo, Andrea; Castelli, Pierluigi; Iwan, Ewelina; Simon, Sandra; Coipan, Claudia E.; Linde, Jörg; Petrovska, Liljana; Kaas, Rolf Sommer; Joensen, Katrine Grimstrup; Nielsen, Sofie Holtsmark; Kiil, Kristoffer; Lagesen, Karin; Di Pasquale, Adriano; Gomes, João Paulo; Deneke, Carlus; Tausch, Simon H.; Borges, VítorDifferent laboratories employ different Whole-Genome Sequencing (WGS) pipelines for Food and Waterborne disease (FWD) surveillance, casting doubt on the comparability of their results and hindering optimal communication at intersectoral and international levels. Through a collaborative effort involving eleven European institutes spanning the food, animal, and human health sectors, we aimed to assess the inter-pipeline clustering congruence across all resolution levels and perform an in-depth comparative analysis of cluster composition at outbreak level for four important foodborne pathogens: Listeria monocytogenes, Salmonella enterica, Escherichia coli, and Campylobacter jejuni. We found a general concordance between allele-based pipelines for all species, except for C. jejuni, where the different resolution power of allele-based schemas led to marked discrepancies. Still, we identified non-negligible differences in outbreak detection and demonstrated how a threshold flexibilization favors the detection of similar outbreak signals by different laboratories. These results, together with the observation that different traditional typing groups (e.g., serotypes) exhibit a remarkably different genetic diversity, represent valuable information for future outbreak case-definitions and WGS-based nomenclature design. This study reinforces the need, while demonstrating the feasibility, of conducting continuous pipeline comparability assessments, and opens good perspectives for a smoother international and intersectoral cooperation towards an efficient One Health FWD surveillance.
- Dengue and Oropouche virus co-infection in a traveller from Cuba to PortugalPublication . Zé-Zé, Líbia; Laranjinha, Joana; Borges, Vítor; Graça, Ana Luísa; Sobral, Daniel; Santos, João Dourado; Carvalho, Ana Cláudia; Faria, Nuno R.; Gomes, João Paulo; Alves, Maria JoãoIn 2024, unprecedented outbreaks of dengue and Oropouche were reported in the Americas. We describe a documented co-infection with dengue and Oropouche viruses in a 35-year-old traveller from Cuba detected in Portugal. RT-PCR and next-generation sequencing confirmed both viruses. Our findings highlight the need for multiplex arboviral diagnostics in travellers from regions with concurrent outbreaks.
- Sustained importance of Streptococcus pneumoniae among pediatric complicated Pneumonia in Portugal (2019-22)Publication . Gomes-Silva, J.; Silva-Costa, Catarina; Pinho, Marcos; Friães, Ana; Ramirez, Mário; Melo-Cristino, José; Portuguese Group for the Study of Streptococcal Infections; Portuguese Study Group of Pediatric Invasive Streptococcal DiseaseBackgrounds: To improve the etiological diagnosis of culture-negative pediatric complicated pneumonia (PCP), we expanded our real-time PCR assay to include other bacterial agents and evaluate potential changes in etiology after 7 years of near universal use of 13-valent conjugate pneumococcal vaccine (PCV13). Methods: We collected 156 culture-negative pleural fluid and empyema samples from children (<18 years), in 62 hospitals in Portugal, from January 2019 to December 2022. Our assay included Streptococcus pneumoniae, Streptococcus pyogenes, Staphylococcus aureus, Mycoplasma pneumoniae, Haemophilus influenzae, Mycobacterium tuberculosis and Streptococcus agalactiae. For S. pneumoniae cases we performed molecular serotyping. Results: Overall, 78 samples were negative for all bacteria tested (50.0%). Among the remaining 78 samples, the majority was positive for S. pneumoniae (n=64, 41.0%). S. pyogenes was found in 7 samples (4.49%), S. aureus in 5 samples (3.20%), H. influenzae in 2 samples (1.28%), and M. pneumoniae in 1 sample (0.64%). We did not detect M. tuberculosis nor S. agalactiae. In 2 samples, we detected the presence of DNA from both S. pneumoniae and another species: S. aureus (n=1) and H. influenzae (n=1). Among the pneumococcal samples, 44 were serotype 3 (56.4%), 5 were serotype 8 (6.41%), 2 were serotype 14 (2.56%) and serotypes 15A, 16F, 19A, 19F and 6C/6D were detected in 1 sample each (1.28% each). The remaining were negative for all serotypes tested (4.49%). Conclusions/Learning Points: After two decades of pneumococcal conjugate vaccine use, S. pneumoniae is still responsible for most culture-negative PCPs, with PCV13 serotype 3 responsible for most cases. Expanding the molecular diagnostic panel to other species allowed the identification of the etiology of only an additional 9.62% of cases, suggesting that bacteria other than S. pneumoniae remain infrequent despite PCV13 use.
- Biofilm Formation by ST17 and ST19 Strains of Streptococcus agalactiaePublication . Silvestre, Inês; Borrego, Maria José; Jordão, LuísaBacterial biofilms are an important virulence factor with a vital role in evasion from the host immune system, colonization and infection. The aim of the present study was to evaluate in vitro the effects of three environmental factors (H+ , glucose and human plasma) in biofilm formation, by carrier and invasive S. agalactiae strains of ST17 and ST19 sequence types, including DNase producers and nonproducers. Bacteria ability to assemble biofilms was classified based on crystal violet assay. Biofilm formation was also monitored by scanning electron microscopy. Depending on the growth medium used, each bacterial isolate could fit in different biofilm production categories. Our data showed that optimal conditions for S. agalactiae biofilm assembly were reached after 48 h incubation at pH 7.6 in the presence of glucose and inactivated human plasma. In the presence of inactivated human plasma, the biofilm biomass of ST19 strains experienced a higher increase than ST17 strains. The composition of the extracellular polymeric matrix of the three strongest biofilm producers (all from ST17) was accessed by enzymatic digestion of mature biofilms and proteins were shown to be the predominant component. The detailed identification of the extracellular protein components should contribute to the development of new therapeutic strategies to fight S. agalactiae infections.
- SARS-CoV-2 seroprevalence studies: the Portuguese experience in last 2 years and next stepsPublication . Rodrigues, Ana Paula; on behalf of the ISN COVID-19 groupPortuguese experience in SARS-CoV-2 seroprevalence studies
- Epidemiology and molecular characterization of invasive disease in children twenty years after the implementation of Haemophilus influenzae serotype b vaccine in Portuguese Immunization ProgrammePublication . Bajanca-Lavado, Maria Paula; Bettencout, Célia; Cunha, Florbela; Gonçalo-Marques, José; Study Group of invasive Haemophilus influenzae disease in of the Pediatric Infection Disease SocietyBackground: Haemophilus influenzae is an important human pathogen responsible for severe childhood invasive disease, despite the implementation of the vaccine against serotype b isolates (Hib), in our National Immunization Programme (NIP), in June 2000. The use of the vaccine lead to a reduction in Hib invasive disease, together with the emergence of non-encapsulated (NTHi), and capsulated non-b-type isolates. This study aims to characterize H. influenzae invasive disease in children, twenty years after the introduction of the Hib vaccine in NIP. Methods Hundred-twenty invasive H. influenzae isolates collected from children in 33 Hospitals, between January 2010 and December 2020, were characterized at the National Reference Laboratory for Haemophilus influenzae. Antibiotic susceptibility was assessed by a microdilution assay. Capsular status was identified by PCR as previously described. MLST was performed as described in the literature. Sequences were analysed and submitted to the MLST website (https://pubmlst.org/hinfluenzae/) for assignment of the sequence type (ST). goeBURST analysis was performed using the PHYLOViZ platform. Results Childhood invasive disease was mainly due to NTHi (55.8%; 67/120), although Hib still in circulation (29.2%; 35/120). Twenty-two cases of vaccine failures were responsible for 62.9% of Hib disease, with 59% of cases occurring in last four years. Non-b capsular types isolates were distributed as follow: 9.2% serotype a (11/120), 1.6% serotype e (2/120) and 4.2% serotype f (5/120). Most isolates were susceptible to all antibiotics studied, with 8.3% (10/120) being ampicillin resistant by β-lactamase producing. MLST revealed, as expected, high genetic variability (77.1%), with 37 different STs among 48 NTHi isolates. In opposition, encapsulated isolates were clonal with Hia assigned to CC23 (ST23-n=6; ST1511-n=1), Hib to CC6 (ST6-n=27, ST190, ST1149 and ST1231 with one isolate each), Hie to CC18 (ST18-n=2) and Hif to CC124 (ST124-n=2, ST1188-n=1). Conclusions Our data suggests that after vaccine implementation, invasive disease among Portuguese children is mainly due to highly genetically diverse, susceptible NTHi isolates. Nevertheless, we are concerned about Hib disease (~30%) despite the higher vaccine coverage observed in our country. Ongoing surveillance should be continued, in order to monitor the burden of the disease, especially Hib, and develop additional public health prevention strategies.
- Molecular identification of fungi from tissue samplesPublication . Sabino, RaquelTissue samples from patients with suspected invasive fungal disease (IFD) should be examined not only by mycological culture but also by microscopy and specific fungal stains should be included. For histopathological diagnosis of fungal infection is required deep knowledge about fungal morphology in tissue and also about the various reactions of the tissue in response to that infection. Highly experienced histopathologists are therefore essential to detect fungal structures and also to recognize tissue reactions associated with IFD, distinguishing them from staining artifacts. This presentation discusses different molecular methods applied to the diagnosis of invasive fungal infections. These methods may help to circunvent some constraints of histology and cultural methods.
- Diagnosis from Tissue: Histology and identification-Quiz the expert session - Q14Publication . Sabino, RaquelHistopathology continues to be a rapid and cost-effective means of providing a presumptive or definitive diagnosis of invasive fungal infections. Tissue samples from patients with suspected invasive fungal disease (IFD) should be examined not only by mycological culture but also by microscopy and specific fungal stains should be included. For histopathological diagnosis of fungal infection is required deep knowledge about fungal morphology in tissue and also about the various reactions of the tissue in response to that infection. Highly experienced histopathologists are therefore essential to detect fungal structures and also to recognize tissue reactions associated with IFD, distinguishing them from staining artifacts. Histopathology of specimens obtained from an affected site, showing a positive result by the presence of distinctive structures associated to specific endemic fungal species, is considered as criteria for proven IFD. On the other hand, observation of yeasts may sometimes point out for a specific IFD but without confidence. Examination of filamentous fungi in histopathological sections can provide important information like the presence of septa, the hyphal diameter or branching angle and melanisation and may thus yield valuable diagnostic clues on the nature of the causative agent. However, a reliable identification of the fungal species based solely on morphological criteria in histopathology is usually impossible. Species identification of fungi detected in histopathological sections should be attempted using immunohistochemical and/or molecular tools. The recently revised EORTC/MSG criteria for diagnosis of IFD from tissue specimens recommends the amplification of fungal DNA by PCR combined by DNA sequencing only when fungal elements are seen by histopathology. During this presentation we will discuss with the audience, in an interactive mode, the advantages and pitfalls of tissue diagnosis, and which tests that can be performed to a more targeted diagnosis.
- Influenza burden estimates in Portugal: seasons 2013/14 to 2018/19Publication . Torres, Ana Rita; Kislaya, Irina; Silva, Susana Pereira; Gomez, Verónica; Machado, Ausenda; Nunes, Baltazar; Guiomar, Raquel; Rodrigues, Ana PaulaReliable influenza burden estimates are essential for a true understanding of the influenza epidemics' impact; informed decision-making and effective risk communication. This study aimed to estimate the influenza burden in Portugal in 2013/14 - 2018/19 seasons.
- Perspectives from a National Reference Laboratory surveillance program on Aspergillus epidemiology and azole resistance” - painel “Fungal infections in Portugal”,Publication . Sabino, RaquelA retrospective laboratory-based study (2017-2018) on Aspergillus surveillance was performed at the national mycology reference laboratory of Portugal. The primary objective were the knowledge of the molecular epidemiology of clinical and environmental Aspergillus isolates. A. fumigatus sensu stricto was the most frequent species in both clinical and environmental sources. A very high frequency of cryptic species was identified. A second objective was the determination of the frequency of azole resistance of A. fumigatus. All isolates included in our collection (2012-2019) identified as belonging to Fumigati section were recovered and screened. The overall frequency of resistance to azoles in A. fumigatus sensu stricto was 3.0% (four clinical and five environmental isolates). The TR34/L98H mutation, linked to environmental transmission route of azole resistance the most frequently detected mutation, found in three environmental and one clinical isolate (1.4%).