Plasma from patients with dengue-like symptoms was collected in 2013 to 2016 from the Brazilian states of Tocantins and Amapa. 781 samples testing negative for IgM against Dengue, Zika, and Chikungunya viruses and for flaviviruses, alphaviruses and enteroviruses RNA using RT-PCRs were analyzed using viral metagenomics. Viral particles-associated nucleic acids were enriched, randomly amplified, and deep sequenced in 102 mini-pools generating over 2 billion reads. Sequence data was analyzed for the presence of known and novel eukaryotic viral reads. Anelloviruses were detected in 80%, human pegivirus 1 in 19%, and parvovirus B19 in 17% of plasma pools. HIV and enteroviruses were detected in two pools each. Previously uncharacterized viral genomes were also identified, and their presence in single plasma samples confirmed by PCR. Chapparvovirus and ambidensovirus genomes, both in the Parvoviridae family, were partially characterized showing 33% and 34% identity in their NS1 sequences to their closest relative. Molecular surveillance using pre-existing plasma from febrile patients provides a readily scalable approach for the detection of novel, potentially emerging, viruses.
BackgroundIn vitro studies have demonstrated that deletions and point mutations introduced into each 21 bp imperfect repeat of Tax-responsive element (TRE) of the genuine human T-cell leukemia virus type I (HTLV-1) viral promoter abolishes Tax induction. Given these data, we hypothesized that similar mutations may affect the proliferation of HTLV-1infected cells and alter the proviral load (PvL). To test this hypothesis, we conducted a cross-sectional genetic analysis to compare the near-complete LTR nucleotide sequences that cover the TRE1 region in a sample of HTLV-1 asymptomatic carriers with different PvL burden.MethodsA total of 94 asymptomatic HTLV-1 carriers with both sequence from the 5' long terminal repeat (LTR) and a PvL for Tax DNA measured using a sensitive SYBR Green real-time PCR were studied. The 94 subjects were divided into three groups based on PvL measurement: 31 low, 29 intermediate, and 34 high. In addition, each group was compared based on sex, age, and viral genotypes. In another analysis, the median PvLs between individuals infected with mutant and wild-type viruses were compared.ResultsUsing a categorical analysis, a G232A substitution, located in domain A of the TRE-1 motif, was detected in 38.7% (12/31), 27.5% (8/29), and 61.8% (21/34) of subjects with low, intermediate, or high PvLs, respectively. A significant difference in the detection of this mutation was found between subjects with a high or low PvL and between those with a high or intermediate PvL (both p < 0.05), but not between subjects with a low or intermediate PvL (p > 0.05). This result was confirmed by a non-parametric analysis that showed strong evidence for higher PvLs among HTLV-1 positive individuals with the G232A mutation than those without this mutation (p < 0.03). No significant difference was found between the groups in relation to age, sex or viral subtypes (p > 0. 05).ConclusionsThe data described here show that changes in domain A of the HTLV-1 TRE-1 motif resulting in the G232A mutation may increase HTLV-1 replication in a majority of infected subjects.
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