The spread of SARS-CoV-2 in Brazil: Brazil has been hard-hit by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic. Candido et al. combined genomic and epidemiological analyses to investigate the impact of nonpharmaceutical interventions (NPIs) in the country. By setting up a network of genomic laboratories using harmonized protocols, the researchers found a 29% positive rate for SARS-CoV-2 among collected samples. More than 100 international introductions of SARS-CoV-2 into Brazil were identified, including three clades introduced from Europe that were already well established before the implementation of NPIs and travel bans. The virus spread from urban centers to the rest of the country, along with a 25% increase in the average distance traveled by air passengers before travel bans, despite an overall drop in short-haul travel. Unfortunately, the evidence confirms that current interventions remain insufficient to keep virus transmission under control in Brazil. Science , this issue p. 1255

Although increasing evidence confirms neuropsychiatric manifestations associated mainly with severe COVID-19 infection, long-term neuropsychiatric dysfunction (recently characterized as part of “long COVID-19” syndrome) has been frequently observed after mild infection. We show the spectrum of cerebral impact of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, ranging from long-term alterations in mildly infected individuals (orbitofrontal cortical atrophy, neurocognitive impairment, excessive fatigue and anxiety symptoms) to severe acute damage confirmed in brain tissue samples extracted from the orbitofrontal region (via endonasal transethmoidal access) from individuals who died of COVID-19. In an independent cohort of 26 individuals who died of COVID-19, we used histopathological signs of brain damage as a guide for possible SARS-CoV-2 brain infection and found that among the 5 individuals who exhibited those signs, all of them had genetic material of the virus in the brain. Brain tissue samples from these five patients also exhibited foci of SARS-CoV-2 infection and replication, particularly in astrocytes. Supporting the hypothesis of astrocyte infection, neural stem cell–derived human astrocytes in vitro are susceptible to SARS-CoV-2 infection through a noncanonical mechanism that involves spike–NRP1 interaction. SARS-CoV-2–infected astrocytes manifested changes in energy metabolism and in key proteins and metabolites used to fuel neurons, as well as in the biogenesis of neurotransmitters. Moreover, human astrocyte infection elicits a secretory phenotype that reduces neuronal viability. Our data support the model in which SARS-CoV-2 reaches the brain, infects astrocytes, and consequently, leads to neuronal death or dysfunction. These deregulated processes could contribute to the structural and functional alterations seen in the brains of COVID-19 patients.

Mutations accrued by SARS-CoV-2 lineage P.1—first detected in Brazil in early January, 2021—include amino acid changes in the receptor-binding domain of the viral spike protein that also are reported in other variants of concern, including B.1.1.7 and B.1.351. We aimed to investigate whether isolates of wild-type P.1 lineage SARS-CoV-2 can escape from neutralising antibodies generated by a polyclonal immune response. We did an immunological study to assess the neutralising effects of antibodies on lineage P.1 and lineage B isolates of SARS-CoV-2, using plasma samples from patients previously infected with or vaccinated against SARS-CoV-2. Two specimens (P.1/28 and P.1/30) containing SARS-CoV-2 lineage P.1 (as confirmed by viral genome sequencing) were obtained from nasopharyngeal and bronchoalveolar lavage samples collected from patients in Manaus, Brazil, and compared against an isolate of SARS-CoV-2 lineage B (SARS.CoV2/SP02.2020) recovered from a patient in Brazil in February, 2020. Isolates were incubated with plasma samples from 21 blood donors who had previously had COVID-19 and from a total of 53 recipients of the chemically inactivated SARS-CoV-2 vaccine CoronaVac: 18 individuals after receipt of a single dose and an additional 20 individuals (38 in total) after receipt of two doses (collected 17–38 days after the most recent dose); and 15 individuals who received two doses during the phase 3 trial of the vaccine (collected 134–230 days after the second dose). Antibody neutralisation of P.1/28, P.1/30, and B isolates by plasma samples were compared in terms of median virus neutralisation titre (VNT50, defined as the reciprocal value of the sample dilution that showed 50% protection against cytopathic effects). In terms of VNT50, plasma from individuals previously infected with SARS-CoV-2 had an 8·6 times lower neutralising capacity against the P.1 isolates (median VNT50 30 [IQR <20–45] for P.1/28 and 30 [<20–40] for P.1/30) than against the lineage B isolate (260 [160–400]), with a binominal model showing significant reductions in lineage P.1 isolates compared with the lineage B isolate (p≤0·0001). Efficient neutralisation of P.1 isolates was not seen with plasma samples collected from individuals vaccinated with a first dose of CoronaVac 20–23 days earlier (VNT50s below the limit of detection [<20] for most plasma samples), a second dose 17–38 days earlier (median VNT50 24 [IQR <20–25] for P.1/28 and 28 [<20–25] for P.1/30), or a second dose 134–260 days earlier (all VNT50s below limit of detection). Median VNT50s against the lineage B isolate were 20 (IQR 20–30) after a first dose of CoronaVac 20–23 days earlier, 75 (<20–263) after a second dose 17–38 days earlier, and 20 (<20–30) after a second dose 134–260 days earlier. In plasma collected 17–38 days after a second dose of CoronaVac, neutralising capacity against both P.1 isolates was significantly decreased (p=0·0051 for P.1/28 and p=0·0336 for P.1/30) compared with that against the lineage B isolate. All data were corroborated by results obtained through plaque reduction neutralisation tests. SARS-CoV-2 lineage P.1 might escape neutralisation by antibodies generated in response to polyclonal stimulation against previously circulating variants of SARS-CoV-2. Continuous genomic surveillance of SARS-CoV-2 combined with antibody neutralisation assays could help to guide national immunisation programmes. São Paulo Research Foundation, Brazilian Ministry of Science, Technology and Innovation and Funding Authority for Studies, Medical Research Council, National Council for Scientific and Technological Development, National Institutes of Health. For the Portuguese translation of the abstract see Supplementary Materials section.

Although increasing evidence confirms neuropsychiatric manifestations associated mainly with severe COVID-19 infection, the long-term neuropsychiatric dysfunction has been frequently observed after mild infection. Here we show the spectrum of the cerebral impact of SARS-CoV-2 infection ranging from long-term alterations in mildly infected individuals (orbitofrontal cortical atrophy, neurocognitive impairment, excessive fatigue and anxiety symptoms) to severe acute damage confirmed in brain tissue samples extracted from the orbitofrontal region (via endonasal trans-ethmoidal approach) from individuals who died of COVID-19. We used surface-based analyses of 3T MRI and identified orbitofrontal cortical atrophy in a group of 81 mildly infected patients (77% referred anosmia or dysgeusia during acute stage) compared to 145 healthy volunteers; this atrophy correlated with symptoms of anxiety and cognitive dysfunction. In an independent cohort of 26 individuals who died of COVID-19, we used histopathological signs of brain damage as a guide for possible SARS-CoV-2 brain infection, and found that among the 5 individuals who exhibited those signs, all of them had genetic material of the virus in the brain. Brain tissue samples from these 5 patients also exhibited foci of SARS-CoV-2 infection and replication, particularly in astrocytes. Supporting the hypothesis of astrocyte infection, neural stem cell-derived human astrocytes in vitro are susceptible to SARS-CoV-2 infection through a non-canonical mechanism that involves spike-NRP1 interaction. SARS-CoV-2-infected astrocytes manifested changes in energy metabolism and in key proteins and metabolites used to fuel neurons, as well as in the biogenesis of neurotransmitters. Moreover, human astrocyte infection elicits a secretory phenotype that reduces neuronal viability. Our data support the model in which SARS-CoV-2 reaches the brain, infects astrocytes and consequently leads to neuronal death or dysfunction. These deregulated processes are also likely to contribute to the structural and functional alterations seen in the brains of COVID-19 patients.

Oropouche orthobunyavirus (OROV) is an emerging arbovirus with a high potential of dissemination in America. Little is known about the role of peripheral blood mononuclear cells (PBMC) response during OROV infection in humans. Thus, to evaluate human leukocytes susceptibility, permissiveness and immune response during OROV infection, we applied RNA hybridization, qRT-PCR and cell-based assays to quantify viral antigens, genome, antigenome and gene expression in different cells. First, we observed OROV replication in human leukocytes lineages as THP-1 monocytes, Jeko-1 B cells and Jurkat T cells. Interestingly, cell viability and viral particle detection are maintained in these cells, even after successive passages. PBMCs from healthy donors were susceptible but the infection was not productive, since neither antigenome nor infectious particle was found in the supernatant of infected PBMCs. In fact, only viral antigens and small quantities of OROV genome were detected at 24 hpi in lymphocytes, monocytes and CD11c⁺ cells. Finally, activation of the Interferon (IFN) response was essential to restrict OROV replication in human PBMCs. Increased expression of type I/III IFNs, ISGs and inflammatory cytokines was detected in the first 24 hpi and viral replication was re-established after blocking IFNAR or treating cells with glucocorticoid. Thus, in short, our results show OROV is able to infect and remain in low titers in human T cells, monocytes, DCs and B cells as a consequence of an effective IFN response after infection, indicating the possibility of leukocytes serving as a trojan horse in specific microenvironments during immunosuppression.

Emerging and re-emerging viruses are a global health concern. Genome sequencing as an approach for monitoring circulating viruses is currently hampered by complex and expensive methods. Untargeted, metagenomic nanopore sequencing can provide genomic information to identify pathogens, prepare for or even prevent outbreaks. SMART (Switching Mechanism at the 5′ end of RNA Template) is a popular method for RNA-Seq but most current methods rely on oligo-dT priming to target polyadenylated mRNA molecules. We have developed two random primed SMART-Seq approaches, ‘SMART-9N’, and a version compatible with barcoded PCR primers available from Oxford Nanopore Technologies, ‘Rapid SMART-9N’, for the detection, characterization, and whole-genome sequencing of RNA viruses. The methods were developed using viral isolates, clinical samples, and compared to a gold-standard amplicon-based method. From a Zika virus isolate the SMART-9N approach recovered 10kb of the 10.8kb RNA genome in a single nanopore read. We also obtained full genome coverage at a high depth coverage using the Rapid SMART-9N, which takes only 10 minutes and costs up to 45% less than other methods. We found the limits of detection of these methods to be 6e00 focus forming units (FFU)/mL with 99.02% and 87.58% genome coverage for SMART-9N and Rapid SMART-9N respectively. Yellow fever virus plasma samples and SARS-CoV-2 nasopharyngeal samples previously confirmed by RT-qPCR with a broad range of Ct-values were selected for validation. Both methods produced greater genome coverage when compared to the multiplex PCR approach and we obtained the longest single read of this study (18.5 kb) with a SARS-CoV-2 clinical sample, 60% of the virus genome using the Rapid SMART-9N method. This work demonstrates that SMART-9N and Rapid SMART-9N are sensitive, low input, and long-read compatible alternatives for RNA virus detection and genome sequencing and Rapid SMART-9N improves the cost, time, and complexity of laboratory work.