Viral metagenomics is increasingly being used for the identification of emerging and re-emerging viral pathogens in clinical samples with unknown etiology. The objective of this study was to shield light on the metavirome composition in clinical samples obtained from patients with clinical history compatible with an arboviral infection, but that presented inconclusive results when tested using RT-qPCR. The inconclusive amplification results might be an indication of the presence of an emerging arboviral agent that is inefficiently amplified by conventional PCR techniques. A total of eight serum samples with inconclusive amplification results for the routinely tested arboviruses—dengue (DENV), Zika (ZIKV), and Chikungunya (CHIKV) obtained during DENV and CHIKV outbreaks registered in the state of Alagoas, Northeast Brazil between July and August 2021—were submitted to metagenomic next-generation sequencing assay using NextSeq 2000 and bioinformatic pipeline for viral discovery. The performed bioinformatic analysis revealed the presence of two arboviruses: DENV type 2 (DENV-2) and CHIKV with a high genome coverage. Further, the metavirome of those samples revealed the presence of multiple commensal viruses apparently without clinical significance. The phylogenetic analysis demonstrated that the DENV-2 genome belonged to the Asian/American genotype and clustered with other Brazilian strains. The identified CHIKV genome was taxonomically assigned as ECSA genotype, which is circulating in Brazil. Together, our results reinforce the utility of metagenomics as a valuable tool for viral identification in samples with inconclusive arboviral amplification. Viral metagenomics is one of the most potent methods for the identification of emerging arboviruses.

Fulminant hepatitis is a severe clinical disease characterized by a marked decline in liver function and encephalopathy. In a previous survey, using metagenomics in a group of 27 patients with this clinical condition, we observed an expressive quantity of reads of the Human pegivirus-1 (HPgV-1). Therefore, the objective of this study was to evaluate the frequency, molecular features, and HPgV-1 circulating genotypes in patients with fulminant hepatitis. After testing the collected plasma samples, we discovered twelve samples (44.4%) that were positive for HPgV-1 RNA (using both real-time and nested PCR). The positive samples presented a mean cycle threshold (Ct) of 28.5 (±7.3). Genotyping assignments revealed that all HPgV-1 positive samples belonged to the HPgV-1 genotype 2 (both subgenotypes 2A and 2B were identified). Although HPgV-1 is considered a commensal virus, little is known regarding its prevalence and genotypes in cases of fulminant hepatitis. More research is needed to understand whether HPgV-1 can be implicated in clinical disorders and infectious diseases.

Viral infections are one of the most important causes of morbidity and mortality among patients undergoing allogeneic hematopoietic stem cell transplantation (HSCT). Immunosuppression may lead to the reactivation of latent viruses or the acquisition of new infections, resulting in severe clinical outcomes. The early detection of viral reactivations is crucial for effective patient management and post-transplant care. In this study, we employed next-generation metagenomics to assess changes in viral abundance and detect clinically significant viruses in allogeneic HSCT patients. A total of 20 patients from the Transplant Unit of the University Hospital of the Faculty of Medicine of Ribeirão Preto, University of São Paulo were included, with plasma samples collected at three time points: D + 0 (pre-transplantation), D + 30 (30 days post-transplantation), and D + 100 (~100 days post-transplantation). A higher presence of clinically relevant viruses, such as the cytomegalovirus (CMV), the Epstein-Barr virus (EBV) and adenoviruses, were predominantly detected at D + 30. The diversity of commensal viruses, primarily anelloviruses, increased gradually, with the highest abundance and variability detected at D + 100. Viruses with clinical importance for HSCT, including CMV, adenovirus and EBV, were confirmed and characterized at the molecular level, showing generally high cycle threshold values. Our findings demonstrate a rise in anellovirus abundance following allogeneic HSCT, with the highest levels observed at D + 100. Notably, D + 30 was identified as a critical time point for the reactivation of clinically significant viruses. This study underscores the potential of metagenomics in the identification of clinically relevant viruses and highlights the importance of monitoring latent viruses in immunocompromised populations, including allogeneic HSCT patients.