Explore the vast range of titles available.

The incidence of microcephaly in Brazil in 2015 was 20 times higher than in previous years. Congenital microcephaly is associated with genetic factors and several causative agents. Epidemiological data suggest that microcephaly cases in Brazil might be associated with the introduction of Zika virus. We aimed to detect and sequence the Zika virus genome in amniotic fluid samples of two pregnant women in Brazil whose fetuses were diagnosed with microcephaly. In this case study, amniotic fluid samples from two pregnant women from the state of Paraíba in Brazil whose fetuses had been diagnosed with microcephaly were obtained, on the recommendation of the Brazilian health authorities, by ultrasound-guided transabdominal amniocentesis at 28 weeks' gestation. The women had presented at 18 weeks' and 10 weeks' gestation, respectively, with clinical manifestations that could have been symptoms of Zika virus infection, including fever, myalgia, and rash. After the amniotic fluid samples were centrifuged, DNA and RNA were extracted from the purified virus particles before the viral genome was identified by quantitative reverse transcription PCR and viral metagenomic next-generation sequencing. Phylogenetic reconstruction and investigation of recombination events were done by comparing the Brazilian Zika virus genome with sequences from other Zika strains and from flaviviruses that occur in similar regions in Brazil. We detected the Zika virus genome in the amniotic fluid of both pregnant women. The virus was not detected in their urine or serum. Tests for dengue virus, chikungunya virus, Toxoplasma gondii, rubella virus, cytomegalovirus, herpes simplex virus, HIV, Treponema pallidum, and parvovirus B19 were all negative. After sequencing of the complete genome of the Brazilian Zika virus isolated from patient 1, phylogenetic analyses showed that the virus shares 97–100% of its genomic identity with lineages isolated during an outbreak in French Polynesia in 2013, and that in both envelope and NS5 genomic regions, it clustered with sequences from North and South America, southeast Asia, and the Pacific. After assessing the possibility of recombination events between the Zika virus and other flaviviruses, we ruled out the hypothesis that the Brazilian Zika virus genome is a recombinant strain with other mosquito-borne flaviviruses. These findings strengthen the putative association between Zika virus and cases of microcephaly in neonates in Brazil. Moreover, our results suggest that the virus can cross the placental barrier. As a result, Zika virus should be considered as a potential infectious agent for human fetuses. Pathogenesis studies that confirm the tropism of Zika virus for neuronal cells are warranted. Consellho Nacional de Desenvolvimento e Pesquisa (CNPq), Fundação de Amparo a Pesquisa do Estado do Rio de Janeiro (FAPERJ).

Nef is an HIV-1 accessory protein that promotes viral replication and pathogenesis. A key function of Nef is to ensure sustained depletion of CD4 and MHC-I molecules in infected cells by inducing targeting of these proteins to multivesicular bodies (MVBs), and ultimately to lysosomes for degradation. Nef also affects cellular secretory routes promoting its own secretion via exosomes. To better understand the effects of Nef on the exocytic pathway, we investigated whether this viral factor modifies the composition of exosomes released by T lymphocytes. We showed that both CD4 and MHC-I molecules are secreted in exosomes from T cells and that the expression of Nef reduces the amount of these proteins in exosomes. To investigate the functional role for this novel activity of Nef, we performed in vitro HIV-1 infection assays in the presence of distinct populations of exosomes. We demonstrated that exosomes released by CD4+ T cells, but not CD4- T cells, efficiently inhibit HIV-1 infection in vitro. Because CD4 is the main receptor for HIV-1 infection, these results suggest that CD4 molecules displayed on the surface of exosomes can bind to envelope proteins of HIV-1 hindering virus interaction with target cells and infection. Importantly, CD4-depleted exosomes released by CD4+ T cells expressing Nef have a reduced capacity to inhibit HIV-1 infection in vitro. These results provide evidence that Nef promotes HIV-1 infection by reducing the expression of CD4 in exosomes from infected cells, besides the original role of Nef in reducing the CD4 levels at the cell surface.

Brief summary In early symptomatic COVID-19 treatment, high dose oral favipiravir did not accelerate viral clearance. Background Favipiravir, an anti-influenza drug, has in vitro antiviral activity against SARS-CoV-2. Clinical trial evidence to date is inconclusive. Favipiravir has been recommended for the treatment of COVID-19 in some countries. Methods In a multicentre open-label, randomised, controlled, adaptive platform trial, low-risk adult patients with early symptomatic COVID-19 were randomised to one of ten treatment arms including high dose oral favipiravir (3.6g on day 0 followed by 1.6g daily to complete 7 days treatment) or no study drug. The primary outcome was the rate of viral clearance (derived under a linear mixed-effects model from the daily log 10 viral densities in standardised duplicate oropharyngeal swab eluates taken daily over 8 days [18 swabs per patient]), assessed in a modified intention-to-treat population (mITT). The safety population included all patients who received at least one dose of the allocated intervention. This ongoing adaptive platform trial was registered at ClinicalTrials.gov (NCT05041907) on 13/09/2021. Results In the final analysis, the mITT population contained data from 114 patients randomised to favipiravir and 126 patients randomised concurrently to no study drug. Under the linear mixed-effects model fitted to all oropharyngeal viral density estimates in the first 8 days from randomisation (4,318 swabs), there was no difference in the rate of viral clearance between patients given favipiravir and patients receiving no study drug; a -1% (95% credible interval: -14 to 14%) difference. High dose favipiravir was well-tolerated. Interpretation Favipiravir does not accelerate viral clearance in early symptomatic COVID-19. The viral clearance rate estimated from quantitative measurements of oropharyngeal eluate viral densities assesses the antiviral efficacy of drugs in vivo with comparatively few studied patients. Research in context Evidence before this study • The in vivo antiviral effect of favipiravir in patients with early symptomatic COVID-19 was not known. Added value of this study • High-dose favipiravir did not demonstrate antiviral activity in early symptomatic COVID-19. • The rate of viral clearance derived from frequent oropharyngeal swabbing in early COVID-19 can be used to characterise in vivo antiviral efficacy. Implications of all available evidence • In vivo antiviral activity of COVID-19 therapeutics should be used to inform policies and practices.

Coronavirus disease 2019 (COVID-19) pandemic has caused immeasurable impacts on the health and socioeconomic system. The real-time identification and characterization of new Variants of Concern (VOCs) are critical to comprehend its emergence and spread worldwide. In this sense, we carried out a national epidemiological surveillance program in Brazil from April to October 2021. Genotyping by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and sequencing were performed to monitor the dynamics and dissemination of VOCs in samples from 15 federative units. Delta VOC was first detected on June 2021 and took sixteen weeks to replace Gamma. To assess the transmissibility potential of Gamma and Delta VOCs, we studied the dynamics of RT-qPCR cycle threshold (Ct) score in the dominance period of each variant. The data suggest that Delta VOC has a higher transmission rate than Gamma VOC. We also compared relevant symptom patterns in individuals infected with both VOCs. The Delta-infected subjects were less likely to have low oxygen saturation or fatigue, altered results on chest computed tomography, and a propensity for altered X-rays. Altogether, we described the replacement of Gamma by Delta, Delta enhanced transmissibility, and differences in symptom presentation.

The emergence of diverse lineages harboring mutations with functional significance and potentially enhanced transmissibility imposes an increased difficulty on the containment of the SARS-CoV-2 pandemic [...]

Zika virus (ZIKV) is an arthropod-borne virus (arbovirus) member of the Flaviviridae family, which has been associated with the development of the congenital Zika syndrome (CZS). RNA viruses, such as flaviviruses, have been reported to exert a profound impact on host microRNAs (miRNAs). Cellular miRNAs modulated by ZIKV may help identify cellular pathways of relevance to pathogenesis. Here, we screened 754 human cellular miRNAs modulated by ZIKV infection (Brazilian PE strain) in a neuroblastoma cell line. Seven miRNAs (miR-99a*, miR-126*, miR-190b, miR-361-3p, miR-522-3p, miR-299-5p, and miR-1267) were downregulated during ZIKV infection, while miR-145 was upregulated. Furthermore, 11 miRNAs were exclusively expressed in ZIKV-infected (miR-148a, miR-342-5p, miR-598, and miR-708-3p) or mock cells (miR-208, miR-329, miR-432-5p, miR-488, miR-518b, miR-520g, and miR-767-5p). Furthermore, in silico analysis indicated that some central nervous system, cellular migration, and adhesion function-related biological processes were overrepresented in the list of target genes of the miRNAs regulated in ZIKV-infected cells, especially for miR-145 and miR-148a. The induction of miR-145 and miR-148a was confirmed in postmortem brain samples from stillborn with severe CZS. Finally, we determined the expression regulation of microcephaly related genes through RNA interference pathway caused by ZIKV directly on neuron cells.

The E3 ubiquitin ligase Smurf1 catalyzes the ubiquitination and proteasomal degradation of several protein substrates related to inflammatory responses and antiviral signaling. This study investigated the role of Smurf1 in modulating inflammation induced by Betacoronavirus infection. Bone marrow-derived macrophages (BMDMs) from C57BL/6 (wild-type) or Smurf1-deficient (Smurf1−/−) mice were infected with MHV-A59 to evaluate the inflammatory response in vitro. Smurf1 was found to be required to downregulate the macrophage production of pro-inflammatory mediators, including TNF, and CXCL1; to control viral release from infected cells; and to increase cell viability. To assess the impact of Smurf 1 in vivo, we evaluated the infection of mice with MHV-A59 through the intranasal route. Smurf1−/− mice infected with a lethal inoculum of MHV-A59 succumbed earlier to infection. Intranasal inoculation with a 10-fold lower dose of MHV-A59 resulted in hematological parameter alterations in Smurf1−/− mice suggestive of exacerbated systemic inflammation. In the lung parenchyma, Smurf1 expression was essential to promote viral clearance, downregulating IFN-β mRNA and controlling the inflammatory profile of macrophages and neutrophils. Conversely, Smurf1 did not affect IFN-β mRNA regulation in the liver, but it was required to increase TNF and iNOS expression in neutrophils and decrease TNF expression in macrophages. In addition, Smurf1−/− mice exhibited augmented liver injuries, accompanied by high serum levels of alanine aminotransferase (ALT). These findings suggest that Smurf1 plays a critical role in regulating the inflammatory response in macrophages and attenuating systemic inflammation during Betacoronavirus infection.

Hepatic cells are major sites of dengue virus (DENV) replication and liver injury constitutes a characteristic of severe forms of dengue. The role of hepatic cells in dengue pathogenesis is not well established, but since hepatocytes are the major source of plasma proteins, changes in protein secretion by these cells during infection might contribute to disease progression. Previously, we showed that DENV infection alters the secretion pattern of hepatic HepG2 cells, with α-enolase appearing as one of the major proteins secreted in higher levels by infected cells. ELISA analysis demonstrated that DENV infection modulates α-enolase secretion in HepG2 cells in a dose-dependent manner, but has no effect on its gene expression and on the intracellular content of the protein as assessed by PCR and western blot analyses, respectively. Two-dimensional western blots showed that both intracellular and secreted forms of α-enolase appear as five spots, revealing α-enolase isoforms with similar molecular weights but distinct isoeletric points. Remarkably, quantification of each spot content revealed that DENV infection shifts the isoform distribution pattern of secreted α-enolase towards the basic isoforms, whereas the intracellular protein remains unaltered, suggesting that post-translational modifications might be involved in α-enolase secretion by infected cells. These findings provide new insights into the mechanisms underlying α-enolase secretion by hepatic cells and its relationship with the role of liver in dengue pathogenesis. In addition, preliminary results obtained with plasma samples from DENV-infected patients suggest an association between plasma levels of α-enolase and disease severity. Since α-enolase binds plasminogen and modulates its activation, it is plausible to speculate the association of the increase in α-enolase secretion by infected hepatic cells with the haemostatic dysfunction observed in dengue patients including the promotion of fibrinolysis and vascular permeability alterations.

Molnupiravir and ritonavir-boosted nirmatrelvir are the two leading oral COVID-19 antiviral treatments, but their antiviral activities in patients have not been compared directly. The aim of this ongoing platform trial is to compare different antiviral treatments using the rate of viral clearance as the measure of antiviral effect. PLATCOV is an open-label, multicentre, phase 2, randomised, controlled, adaptive pharmacometric platform trial running in Thailand, Brazil, Pakistan, and Laos. The component of the trial reported here was conducted in the Hospital for Tropical Diseases, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand. We recruited low-risk adult patients aged 18–50 years with early symptomatic COVID-19 (<4 days of symptoms). Eligible patients were randomly assigned using block randomisation via a centralised web app to one of seven treatment groups: molnupiravir, ritonavir-boosted nirmatrelvir, casirivimab–imdevimab, tixagevimab–cilgavimab, favipiravir, fluoxetine, or no study drug. The no study drug group comprised a minimum proportion of 20% of patients at all times, with uniform randomisation ratios applied across the active treatment groups. Results for the concurrently randomised molnupiravir, ritonavir-boosted nirmatrelvir, and no study drug groups are reported here. The primary endpoint was the rate of oropharyngeal viral clearance assessed in a modified intention-to-treat population, defined as patients with more than 2 days of follow-up. Safety was assessed in all participants who took at least one dose of the medication. The viral clearance rate was derived under a Bayesian hierarchical linear model fitted to the log10 viral densities in standardised duplicate oropharyngeal swab eluates taken daily over 1 week (18 measurements). Treatment groups with a probability of more than 0·9 that viral clearance was accelerated by more than 20% compared with no drug entered a non-inferiority comparison (with a 10% non-inferiority margin) compared with the platform's current most effective drug. This ongoing trial is registered at ClinicalTrials.gov, NCT05041907. Between June 6, 2022, and Feb 23, 2023, 209 patients in Thailand were enrolled and concurrently randomly assigned to molnupiravir (n=65), ritonavir-boosted nirmatrelvir (n=59), or no study drug (n=85). 129 (62%) of the patients were female and 80 (38%) were male. Relative to the no study drug group, the rates of viral clearance were 37% (95% credible interval 16–65) faster with molnupiravir and 84% (54–119) faster with ritonavir-boosted nirmatrelvir. In the non-inferiority comparison, viral clearance was 25% (10–38) slower with molnupiravir than ritonavir-boosted nirmatrelvir. Molnupiravir was removed from the study platform when it reached the prespecified inferiority margin of 10% compared with ritonavir-boosted nirmatrelvir. Median estimated viral clearance half-lives were 8·5 h (IQR 6·7–10·1) with ritonavir-boosted nirmatrelvir, 11·6 h (8·6–15·4) with molnupiravir, and 15·5 h (11·9–21·2) with no study drug. Viral rebound occurred more frequently following nirmatrelvir (six [10%] of 58) compared with the no study drug (one [1%] of 84; p=0·018) or the molnupiravir (one [2%] of 65; p=0·051) groups. Persistent infections following molnupiravir had more viral mutations (three of nine patients had an increased number of single nucleotide polymorphisms in samples collected at 7 or more days compared with those at baseline) than after nirmatrelvir (zero of three) or no study drug (zero of 19). There were no adverse events of grade 3 or worse, or serious adverse events in any of the reported treatment groups. Both molnupiravir and ritonavir-boosted nirmatrelvir accelerate oropharyngeal SARS-CoV-2 viral clearance in patients with COVID-19, but the antiviral effect of ritonavir-boosted nirmatrelvir was substantially greater. Measurement of oropharyngeal viral clearance rates provides a rapid and well tolerated approach to the assessment and comparison of antiviral drugs in patients with COVID-19. It should be evaluated in other acute viral respiratory infections. Wellcome Trust through the COVID-19 Therapeutics Accelerator.

HIV-1 subtype B is the most prevalent in developed countries and, consequently, it has been extensively studied. On the other hand, subtype C is the most prevalent worldwide and therefore is a reasonable target for future studies. Here we evaluate the acquisition of resistance and the viability of HIV-1 subtype B and C RT clones from different isolates that were subjected to in vitro selection pressure with zidovudine (ZDV) and lamivudine (3TC). MT4 cells were infected with chimeric virus pseudotyped with RT from subtype B and C clones, which were previously subjected to serial passage with increasing concentrations of ZDV and 3TC. The samples collected after each passage were analyzed for the presence of resistance mutations and VL. No differences were found between subtypes B and C in viral load and resistance mutations when these viruses were selected with 3TC. However, the route of mutations and the time to rebound of subtype B and C virus were different when subjected to ZDV treatment. In order to confirm the role of the mutations detected, other clones were generated and subjected to in vitro selection. RT subtype B virus isolates tended to acquire different ZDV resistance mutations (Q151M and D67N or T215Y, D67D/N and F214L) compared to subtype C (D67N, K70R, T215I or T215F). This study suggests that different subtypes have a tendency to react differently to antiretroviral drug selection in vitro. Consequently, the acquisition of resistance in patients undergoing antiretroviral therapy can be dependent on the subtypes composing the viral population.