Dengue fever is among the most significant public health concerns in Brazil. To date, the highest number of Dengue notifications in the Americas has been reported in Brazil, with cases accounting for a total number of 3,418,796 reported cases as of mid-December 2022. Furthermore, the northeastern region of Brazil registered the second-highest incidence of Dengue fever in 2022. Due to the alarming epidemiological scenario, in this study, we used a combination of portable whole-genome sequencing, phylodynamic, and epidemiological analyses to reveal a novel DENV-1 genotype V clade and the persistence of DENV-2 genotype III in the region. We further report the presence of non-synonymous mutations associated with non-structural domains, especially the NS2A (non-structural protein 2A), as well as describe synonymous mutations in envelope and membrane proteins, distributed differently between clades. However, the absence of clinical data at the time of collection and notification, as well as the impossibility of monitoring patients in order to observe worsening or death, restricts our possibility of correlating mutational findings with possible clinical prognoses. Together, these results reinforce the crucial role of genomic surveillance to follow the evolution of circulating DENV strains and understand their spread across the region through inter-regional importation events, likely mediated by human mobility, and also the possible impacts on public health and outbreak management.

Acute febrile illness (AFI) and severe neurological disorders (SNDs) often present diagnostic challenges due to their potential origins from a wide range of infectious agents. Nanopore metagenomics is emerging as a powerful tool for identifying the microorganisms potentially responsible for these undiagnosed clinical cases. In this study, we aim to shed light on the etiological agents underlying AFI and SND cases that conventional diagnostic methods have not been able to fully elucidate. Our approach involved analyzing samples from fourteen hospitalized patients using a comprehensive nanopore metagenomic approach. This process included RNA extraction and enrichment using the SMART-9N protocol, followed by nanopore sequencing. Subsequent steps involved quality control, host DNA/cDNA removal, de novo genome assembly, and taxonomic classification. Our findings in AFI cases revealed a spectrum of disease-associated microbes, including Escherichia coli, Streptococcus sp., Human Immunodeficiency Virus 1 (Subtype B), and Human Pegivirus. Similarly, SND cases revealed the presence of pathogens such as Escherichia coli, Clostridium sp., and Dengue virus type 2 (Genotype-II lineage). This study employed a metagenomic analysis method, demonstrating its efficiency and adaptability in pathogen identification. Our investigation successfully identified pathogens likely associated with AFI and SNDs, underscoring the feasibility of retrieving near-complete genomes from RNA viruses. These findings offer promising prospects for advancing our understanding and control of infectious diseases, by facilitating detailed genomic analysis which is critical for developing targeted interventions and therapeutic strategies.

This study examines a case involving a 7-year-old child who developed dengue symptoms following Qdenga vaccination. Despite initial negative diagnostic results, molecular analysis confirmed an infection with DENV4. Next-generation sequencing detected viral RNA from both DENV2 and DENV4 serotypes, which were identified as vaccine-derived strains using specific primers. Phylogenetic analysis further confirmed that these sequences belonged to the Qdenga vaccine rather than circulating wild-type viruses. This case underscores the critical need for precise diagnostic interpretation in vaccinated individuals to avoid misdiagnosis and to strengthen public health surveillance. A comprehensive understanding of vaccine-induced viremia is essential for refining dengue surveillance, improving diagnostic accuracy, and informing public health strategies in endemic regions.

Dengue virus (DENV) is a major global health threat whose expansion into temperate regions has been facilitated by climate change and vector adaptation. Despite recurrent epidemics in Brazil, genomic surveillance in mosquitoes remains limited, particularly in the South. To address this gap, we implemented a novel urban mosquito-trapping strategy optimized for low-density and peri-domestic environments in Porto Alegre, Rio Grande do Sul. Between April and July 2023, were collected 4,768 Aedes aegypti samples across 16 neighborhoods, generating 2,022 pools. Among these, 41 pools tested positive for DENV, of which 33 pools exhibited RNA integrity suitable for sequencing. Whole-genome sequencing revealed 33 DENV-positive pools, including 29 DENV-1 (Genotype V) and 4 DENV-2 (Genotype II). Two pools contained both serotypes, highlighting the risk of sequential infections in humans. Phylogenetic analyses indicated sustained local transmission alongside multiple introductions, while recurrent mutations, particularly in NS1, NS2A, and NS5, suggested ongoing viral adaptation. These findings represent the first vector-based genomic data for dengue in southern Brazil and demonstrate the utility of mosquito genomics for early outbreak detection, serotype monitoring, and preparedness in emerging transmission zones. Author Summary: Dengue virus (DENV) continues to expand into new regions, driven by climate change and mosquito adaptation. In Brazil, dengue epidemics are recurrent, but genomic surveillance of mosquitoes is still scarce, particularly in the southern states. To fill this gap, we implemented an urban mosquito-trapping strategy designed for low-density and peri-domestic environments in Porto Alegre, Rio Grande do Sul. From April to July 2023, we collected 4,768 Aedes aegypti samples across 16 neighborhoods, generating 2,022 pools. Among these, 41 pools were positive for DENV, of which 33 pools exhibited RNA integrity suitable for sequencing, mainly DENV-1 (Genotype V) and DENV-2 (Genotype II), with two pools showing co-circulation of both serotypes, raising concern for sequential infections in humans. Phylogenetic analysis revealed sustained local transmission combined with multiple virus introductions, and mutations in NS1, NS2A, and NS5 suggested ongoing viral adaptation. Our findings provide the first mosquito-based genomic data for dengue in southern Brazil and highlight how mosquito genomics can strengthen early outbreak detection, serotype monitoring, and epidemic preparedness.

This study examines a case involving a 7-year-old child who developed dengue symptoms following Qdenga vaccination. Despite initial negative diagnostic results, molecular analysis confirmed an infection with DENV4. Next-generation sequencing detected viral RNA from both DENV2 and DENV4 serotypes, which were identified as vaccine-derived strains using specific primers. Phylogenetic analysis further confirmed that these sequences belonged to the Qdenga vaccine rather than circulating wild-type viruses. This case underscores the critical need for precise diagnostic interpretation in vaccinated individuals to avoid misdiagnosis and to strengthen public health surveillance. A comprehensive understanding of vaccine-induced viremia is essential for refining dengue surveillance, improving diagnostic accuracy, and informing public health strategies in endemic regions.