RESUMO
Plitidepsin is a host-targeted compound known for inducing a strong anti-SARS-CoV-2 activity, as well as for having the capacity of reducing lung inflammation. Because IL-6 is one of the main cytokines involved in acute respiratory distress syndrome, the effect of plitidepsin in IL-6 secretion in different in vitro and in vivo experimental models was studied. A strong plitidepsin-mediated reduction of IL-6 was found in human monocyte-derived macrophages exposed to nonproductive SARS-CoV-2. In resiquimod (a ligand of TLR7/8)-stimulated THP1 human monocytes, plitidepsin-mediated reductions of IL-6 mRNA and IL-6 levels were also noticed. Additionally, although resiquimod-induced binding to DNA of NF-κB family members was unaffected by plitidepsin, a decrease in the regulated transcription by NF-κB (a key transcription factor involved in the inflammatory cascade) was observed. Furthermore, the phosphorylation of p65 that is required for full transcriptional NF-κB activity was significantly reduced by plitidepsin. Moreover, decreases of IL-6 levels and other proinflammatory cytokines were also seen in either SARS-CoV-2 or H1N1 influenza virus-infected mice, which were treated at low enough plitidepsin doses to not induce antiviral effects. In summary, plitidepsin is a promising therapeutic agent for the treatment of viral infections, not only because of its host-targeted antiviral effect, but also for its immunomodulatory effect, both of which were evidenced in vitro and in vivo by the decrease of proinflammatory cytokines.
Assuntos
Depsipeptídeos , Vírus da Influenza A Subtipo H1N1 , NF-kappa B , Humanos , Animais , Camundongos , NF-kappa B/metabolismo , Interleucina-6/farmacologia , Antivirais/farmacologia , Fatores Imunológicos/farmacologia , Citocinas/metabolismo , SARS-CoV-2/metabolismoRESUMO
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an enveloped positive stranded RNA virus which has caused the recent deadly pandemic called COVID-19. The SARS-CoV-2 virion is coated with a heavily glycosylated Spike glycoprotein which is responsible for attachment and entry into target cells. One, as yet unexploited strategy for preventing SARS-CoV-2 infections, is the targeting of the glycans on Spike. Lectins are carbohydrate-binding proteins produced by plants, algae, and cyanobacteria. Some lectins can neutralize enveloped viruses displaying external glycoproteins, offering an alternative therapeutic approach for the prevention of infection with virulent ß-coronaviruses, such as SARS-CoV-2. Here we show that the cyanobacterial lectin cyanovirin-N (CV-N) can selectively target SARS-CoV-2 Spike oligosaccharides and inhibit SARS-CoV-2 infection in vitro and in vivo. CV-N neutralizes Delta and Omicron variants in vitro better than earlier circulating viral variants. CV-N binds selectively to Spike with a Kd as low as 15 nM and a stoichiometry of 2 CV-N: 1 Spike but does not bind to the receptor binding domain (RBD). Further mapping of CV-N binding sites on Spike shows that select high-mannose oligosaccharides in the S1 domain of Spike are targeted by CV-N. CV-N also reduced viral loads in the nares and lungs in vivo to protect hamsters against a lethal viral challenge. In summary, we present an anti-coronavirus agent that works by an unexploited mechanism and prevents infection by a broad range of SARS-CoV-2 strains.
Assuntos
COVID-19 , SARS-CoV-2 , Animais , Cricetinae , Oligossacarídeos/farmacologia , LectinasRESUMO
People living with human immunodeficiency virus (HIV) are the individuals most affected by the current Monkeypox virus outbreak that was first announced in May 2022. Here we report Pan-pox-specific T-cell responses in a cohort of HIV-1-infected individuals after receiving the nonreplicative, attenuated smallpox vaccine JYNNEOS from Bavarian Nordic. Intradermal (i.d.) and subcutaneous (s.c.) vaccination was safe without major side effects. Dose-sparing i.d. vaccination was superior to s.c. vaccination and promoted T-cell polyfunctionality, and the expression of the gut-homing marker α4ß7 integrin on lymphocytes. HIV-1-infected individuals with CD4 T-cell counts ≤500/mm3 blood required at least a booster vaccination to exhibit efficient virus-specific T-cell responses. The magnitude of the Th1 response after this booster directly correlated with the CD4 T-cell count of the vaccinees. Further studies with a larger number of participants are warranted to confirm and expand our observations.
Assuntos
Infecções por HIV , HIV-1 , Humanos , Linfócitos T CD4-Positivos , VacinaçãoRESUMO
AIM: Aerosols released from the oral cavity help spread the SARS-CoV-2 virus. The use of a mouthwash formulated with an antiviral agent could reduce the viral load in saliva, helping to lower the spread of the virus. The aim of this study was to assess the efficacy of a mouthwash with 0.07% cetylpyridinium chloride (CPC) to reduce the viral load in the saliva of Coronavirus disease 2019 (COVID-19) patients. MATERIALS AND METHODS: In this multi-centre, single-blind, randomized, parallel group clinical trial, 80 COVID-19 patients were enrolled and randomized to two groups, namely test (n = 40) and placebo (n = 40). Saliva samples were collected at baseline and 2 h after rinsing. The samples were analysed by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and an enzyme-linked immunosorbent assay test specific for the nucleocapsid (N) protein of SARS-CoV-2. RESULTS: With RT-qPCR, no significant differences were observed between the placebo group and the test group. However, 2 h after a single rinse, N protein concentration in saliva was significantly higher in the test group, indicating an increase in lysed virus. CONCLUSIONS: The use of 0.07% CPC mouthwash induced a significant increase in N protein detection in the saliva of COVID-19 patients. Lysis of the virus in the mouth could help reduce the transmission of SARS-CoV-2. However, more studies are required to prove this.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , COVID-19/prevenção & controle , Cetilpiridínio/uso terapêutico , Antissépticos Bucais/uso terapêutico , Carga Viral , Método Simples-CegoRESUMO
BACKGROUND: We analyzed humoral and cellular immune responses induced by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) messenger RNA (mRNA) vaccines in people with human immunodeficiency virus (HIV; PWH) who had CD4+ T-cell counts <200/µL (HIV<200 group). METHODS: This prospective cohort study included 58 PWH in the HIV<200 group, 36 with CD4+ T-cell counts >500/µL (HIV>500 group), and 33 HIV-1-negative controls (control group). Antibodies against the SARS-CoV-2 spike protein (anti-S immunoglobulin [Ig] G) and the receptor-binding domain (anti-RBD IgG) were quantified before and 4â weeks after the first and the second doses of BNT162b2 or mRNA-1273 (at week 8). Viral neutralization activity and T-cell responses were also determined. RESULTS: At week 8, anti-S/anti-RBD IgG responses increased in all groups (P < .001). Median (interquartile range) anti-S and anti-RBD IgG levels at week 8 were 153.6 (26.4-654.9) and 171.9 (61.8-425.8) binding antibody units (BAU)/mL, respectively, in the HIV<200 group, compared with 245.6 (145-824) and 555.8 (166.4-1751) BAU/mL in the HIV>500 group and 274.7 (193.7-680.4) and 281.6 (181-831.8) BAU/mL in controls (P < .05). Neutralizing capacity and specific T-cell immune responses were absent or reduced in 33% of those in the HIV<200 group, compared with 3.7% in the HIV>500 group (P < .01). CONCLUSIONS: One-third of PWH with CD4+ T-cell counts <200/µL show low anti-S/anti-RBD IgG levels, reduced in vitro neutralization activity against SARS-CoV-2, and no vaccine-induced T cells after receiving coronavirus disease 2019 mRNA vaccines.
Assuntos
Vacinas contra COVID-19 , COVID-19 , Soropositividade para HIV , Reconstituição Imune , Humanos , Anticorpos Antivirais , Vacina BNT162 , COVID-19/prevenção & controle , Vacinas contra COVID-19/imunologia , Imunoglobulina G , Estudos Prospectivos , SARS-CoV-2 , Vacinação , Imunidade Humoral , Imunidade Celular , Linfócitos TRESUMO
KEY MESSAGE: Rice-produced SD1 retains its physicochemical properties and provides efficient pre-exposure HIV-1 prophylaxis against infection in vitro. Scytovirin (SVN) is an HIV-neutralizing lectin that features two structural domains (SD1 and SD2) that bind to HIV-1 envelope glycoproteins. We expressed SD1 in rice seeds as a potential large-scale production platform and confirmed that rice-derived SD1 binds the HIV-1 envelope glycoprotein gp120 in vitro. We analyzed the thermodynamic properties of SD1 compared to full-size SVN (produced in E. coli) by isothermal titration and differential scanning calorimetry to characterize the specific interactions between SVN/SD1 and gp120 as well as to high-mannose oligosaccharides. SVN bound with moderate affinity (Kd = 1.5 µM) to recombinant gp120, with 2.5-fold weaker affinity to nonamannoside (Kd of 3.9 µM), and with tenfold weaker affinity to tetramannoside (13.8 µM). The melting temperature (Tm) of full-size SVN was 59.1 °C and the enthalpy of unfolding (ΔHunf) was 16.4 kcal/mol, but the Tm fell when SVN bound to nonamannoside (56.5 °C) and twice as much energy was required for unfolding (ΔHunf = 33.5 kcal/mol). Interestingly, binding to tetramannoside destabilized the structure of SD1 (ΔTm ~ 11.5 °C) and doubled the enthalpy of unfolding, suggesting a dimerization event. The similar melting phenomenon shared by SVN and SD1 in the presence of oligomannose confirmed their conserved oligosaccharide-binding mechanisms. SD1 expressed in transgenic rice was able to neutralize HIV-1 in vitro. SD1 expressed in rice, therefore, is suitable as a microbicide component.
Assuntos
HIV-1 , Oryza , Proteínas de Bactérias/metabolismo , Proteínas de Transporte/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Proteína gp120 do Envelope de HIV/genética , Proteína gp120 do Envelope de HIV/metabolismo , Lectinas/química , Lectinas/metabolismo , Proteínas de Membrana/metabolismo , Oryza/genética , Oryza/metabolismo , SindactiliaRESUMO
New therapies to treat or prevent viral infections are essential, as recently observed during the COVID-19 pandemic. Here, we propose a therapeutic strategy based on monoclonal antibodies that block the specific interaction between the host receptor Siglec-1/CD169 and gangliosides embedded in the viral envelope. Antibodies are an excellent option for treating infectious diseases based on their high specificity, strong targeting affinity, and relatively low toxicity. Through a process of humanization, we optimized monoclonal antibodies to eliminate sequence liabilities and performed biophysical characterization. We demonstrated that they maintain their ability to block viral entry into myeloid cells. These molecular improvements during the discovery stage are key if we are to maximize efforts to develop new therapeutic strategies. Humanized monoclonal antibodies targeting CD169 provide new opportunities in the treatment of infections caused by ganglioside-containing enveloped viruses, which pose a constant threat to human health. In contrast with current neutralizing antibodies that bind antigens on the infectious particle, our antibodies can prevent several types of enveloped viruses interacting with host cells because they target the host CD169 protein, thus becoming a potential pan-antiviral therapy.
Assuntos
Anticorpos Monoclonais Humanizados , Antivirais , Lectina 1 Semelhante a Ig de Ligação ao Ácido Siálico , Lectina 1 Semelhante a Ig de Ligação ao Ácido Siálico/imunologia , Humanos , Anticorpos Monoclonais Humanizados/farmacologia , Anticorpos Monoclonais Humanizados/uso terapêutico , Anticorpos Monoclonais Humanizados/imunologia , Antivirais/farmacologia , Antivirais/uso terapêutico , Animais , Tratamento Farmacológico da COVID-19 , Internalização do Vírus/efeitos dos fármacos , SARS-CoV-2/imunologia , SARS-CoV-2/efeitos dos fármacosRESUMO
BACKGROUND: Since the emergence of the global mpox outbreak in May, 2022, more than 90 000 cases have been diagnosed across 110 countries, disproportionately affecting people with HIV. The durability of mpox-specific immunity is unclear and reinfections have been reported. We aimed to compare mpox immune responses up to 6 months after diagnosis in participants with and without HIV and assess their effect on disease severity and viral clearance dynamics. METHODS: This study was embedded within a prospective, observational, multicentre cohort study of viral clearance dynamics among people with mpox in Spain (MoViE). We included women and men aged 18 years or older, who had signs of mpox, and reported having symptom onset within the previous 10 days at the moment of mpox diagnosis from three sex clinics of the Barcelona metropolitan area. Samples from skin ulcers were collected weekly to estimate the time to clear monkeypox virus (MPXV) from skin lesions. Blood samples were taken at diagnosis, 29, 91, and 182 days later for immune analysis. This included quantifying IgG and IgA against three mpox antigens by ELISA, evaluating in-vitro neutralisation, and characterising mpox-specific T-cell responses using interferon γ detecting enzyme-linked immunospot (ELISpot) assay and multiparametric flow cytometry. FINDINGS: Of the 77 originally enrolled participants, we included 33 participants recruited between July 19, and Oct 6, 2022. Participants without HIV (19 [58%] participants) and participants with HIV (14 [42%] participants) had similar clinical severity and time to MPXV clearance in skin lesions. Participants with HIV had a CD4+ T-cell count median of 777 cells per µL (IQR 484-1533), and 11 (78%) of 14 were virally suppressed on antiretroviral therapy. Nine (27%) of 33 participants were age 49 years or older. 15 (45%) of 33 participants were originally from Spain, and all participants were men. Early humoral responses, particularly concentrations and breadth of IgG and IgA, were associated with milder disease and faster viral clearance. Orthopoxvirus-specific T cells count was also positively correlated with MPXV clearance. Antibody titres declined more rapidly in participants with HIV, but T-cell responses against MPXV were sustained up to day 182 after diagnosis, regardless of HIV status. INTERPRETATION: Higher breadth and magnitude of B-cell and T-cell responses are important in facilitating local viral clearance, limiting mpox dissemination, and reducing disease severity in individuals with preserved immune system. Antibodies appear to contribute to early viral control and T-cell responses are sustained over time, which might contribute to milder presentations during reinfection. FUNDING: Fundació Lluita contra les Infeccions, IrsiCaixa, and Consorcio Centro de Investigación Biomédica en Red, Instituto de Salud Carlos III, Ministerio de Ciencia, Innovación e Universidades.
Assuntos
Infecções por HIV , Humanos , Masculino , Espanha/epidemiologia , Infecções por HIV/imunologia , Infecções por HIV/epidemiologia , Estudos Prospectivos , Adulto , Feminino , Pessoa de Meia-Idade , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina A/sangue , Imunoglobulina A/imunologia , Carga Viral , Linfócitos T/imunologiaRESUMO
Safe and effective severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines are crucial to fight against the coronavirus disease 2019 pandemic. Most vaccines are based on a mutated version of the Spike glycoprotein [K986P/V987P (S-2P)] with improved stability, yield and immunogenicity. However, S-2P is still produced at low levels. Here, we describe the V987H mutation that increases by two-fold the production of the recombinant Spike and the exposure of the receptor binding domain (RBD). S-V987H immunogenicity is similar to S-2P in mice and golden Syrian hamsters (GSH), and superior to a monomeric RBD. S-V987H immunization confer full protection against severe disease in K18-hACE2 mice and GSH upon SARS-CoV-2 challenge (D614G or B.1.351 variants). Furthermore, S-V987H immunized K18-hACE2 mice show a faster tissue viral clearance than RBD- or S-2P-vaccinated animals challenged with D614G, B.1.351 or Omicron BQ1.1 variants. Thus, S-V987H protein might be considered for future SARS-CoV-2 vaccines development.
Assuntos
COVID-19 , Melfalan , SARS-CoV-2 , gama-Globulinas , Cricetinae , Animais , Humanos , Camundongos , Mesocricetus , Vacinas contra COVID-19 , COVID-19/prevenção & controle , Glicoproteína da Espícula de Coronavírus/genética , Imunização , Glicoproteínas , Anticorpos Neutralizantes , Anticorpos AntiviraisRESUMO
Sialic-acid-binding immunoglobulin-like lectins are cell surface immune receptors known as Siglecs that play a paramount role as modulators of immunity. In recent years, research has underscored how the underlaying biology of this family of receptors influences the outcome of viral infections. While Siglecs are needed to promote effective antiviral immune responses, they can also pave the way to viral dissemination within tissues. Here, we review how recent preclinical findings focusing on the interplay between Siglecs and viruses may translate into promising broad-spectrum therapeutic interventions or key biomarkers to monitor the course of viral infections.
Assuntos
Lectinas Semelhantes a Imunoglobulina de Ligação ao Ácido Siálico , Viroses , Humanos , Lectinas Semelhantes a Imunoglobulina de Ligação ao Ácido Siálico/genética , Lectinas Semelhantes a Imunoglobulina de Ligação ao Ácido Siálico/metabolismo , Viroses/genéticaRESUMO
The recent monkeypox virus (MPXV) outbreak was of global concern and has mainly affected gay, bisexual and other men who have sex with men (GBMSM). Here we assess prevalence of MPXV in high-risk populations of GBMSM, trans women (TW) and non-binary people without symptoms or with unrecognized monkeypox (Mpox) symptoms, using a self-sampling strategy. Anal and pharyngeal swabs are tested by MPXV real-time PCR and positive samples are tested for cytopathic effect (CPE) in cell culture. 113 individuals participated in the study, 89 (78.76%) were cis men, 17 (15.04%) were TW. The median age was 35.0 years (IQR: 30.0-43.0), 96 (85.02%) individuals were gay or bisexual and 72 (63.72%) were migrants. Seven participants were MPXV positive (6.19% (95% CI: 1.75%-10.64%)). Five tested positive in pharyngeal swabs, one in anal swab and one in both. Six did not present symptoms recognized as MPXV infection. Three samples were positive for CPE, and showed anti-vaccinia pAb staining by FACS and confocal microscopy. This suggests that unrecognized Mpox cases can shed infectious virus. Restricting testing to individuals reporting Mpox symptoms may not be sufficient to contain outbreaks.
Assuntos
Mpox , Minorias Sexuais e de Gênero , Masculino , Humanos , Feminino , Adulto , Espanha/epidemiologia , Homossexualidade Masculina , Mpox/diagnóstico , Mpox/epidemiologia , Monkeypox virus/genéticaRESUMO
The COVID-19 pandemic posed a global health crisis, with new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants weakening vaccine-driven protection. Trained immunity could help tackle COVID-19 disease. Our objective was to analyze whether heat-killed Mycobacterium manresensis (hkMm), an environmental mycobacterium, induces trained immunity and confers protection against SARS-CoV-2 infection. To this end, THP-1 cells and primary monocytes were trained with hkMm. The increased secretion of tumor necrosis factor alpha (TNF-α), interleukin (IL)-6, IL-1ß, and IL-10, metabolic activity, and changes in epigenetic marks suggested hkMm-induced trained immunity in vitro. Healthcare workers at risk of SARS-CoV-2 infection were enrolled into the MANRECOVID19 clinical trial (NCT04452773) and were administered Nyaditum resae (NR, containing hkMm) or placebo. No significant differences in monocyte inflammatory responses or the incidence of SARS-CoV-2 infection were found between the groups, although NR modified the profile of circulating immune cell populations. Our results show that M. manresensis induces trained immunity in vitro but not in vivo when orally administered as NR daily for 14 days.
RESUMO
The SARS-CoV-2 pandemic made evident that there are only a few drugs against coronavirus. Here we aimed to identify a cost-effective antiviral with broad spectrum activity and high safety profile. Starting from a list of 116 drug candidates, we used molecular modelling tools to rank the 44 most promising inhibitors. Next, we tested their efficacy as antivirals against α and ß coronaviruses, such as the HCoV-229E and SARS-CoV-2 variants. Four drugs, OSW-1, U18666A, hydroxypropyl-ß-cyclodextrin (HßCD) and phytol, showed in vitro antiviral activity against HCoV-229E and SARS-CoV-2. The mechanism of action of these compounds was studied by transmission electron microscopy and by fusion assays measuring SARS-CoV-2 pseudoviral entry into target cells. Entry was inhibited by HßCD and U18666A, yet only HßCD inhibited SARS-CoV-2 replication in the pulmonary Calu-3 cells. Compared to the other cyclodextrins, ß-cyclodextrins were the most potent inhibitors, which interfered with viral fusion via cholesterol depletion. ß-cyclodextrins also prevented infection in a human nasal epithelium model ex vivo and had a prophylactic effect in the nasal epithelium of hamsters in vivo. All accumulated data point to ß-cyclodextrins as promising broad-spectrum antivirals against different SARS-CoV-2 variants and distant alphacoronaviruses. Given the wide use of ß-cyclodextrins for drug encapsulation and their high safety profile in humans, our results support their clinical testing as prophylactic antivirals.
Assuntos
COVID-19 , Fármacos Dermatológicos , beta-Ciclodextrinas , Humanos , SARS-CoV-2 , Antivirais/farmacologia , Antivirais/uso terapêutico , beta-Ciclodextrinas/farmacologia , beta-Ciclodextrinas/uso terapêuticoRESUMO
Most COVID-19 vaccines are based on the SARS-CoV-2 Spike glycoprotein (S) or their subunits. However, S shows some structural instability that limits its immunogenicity and production, hampering the development of recombinant S-based vaccines. The introduction of the K986P and V987P (S-2P) mutations increases the production and immunogenicity of the recombinant S trimer, suggesting that these two parameters are related. Nevertheless, S-2P still shows some molecular instability and it is produced with low yield. Here we described a novel set of mutations identified by molecular modeling and located in the S2 region of the S-2P that increase its production up to five-fold. Besides their immunogenicity, the efficacy of two representative S-2P-based mutants, S-29 and S-21, protecting from a heterologous SARS-CoV-2 Beta variant challenge was assayed in K18-hACE2 mice (an animal model of severe SARS-CoV-2 disease) and golden Syrian hamsters (GSH) (a moderate disease model). S-21 induced higher level of WH1 and Delta variants neutralizing antibodies than S-2P in K18-hACE2 mice three days after challenge. Viral load in nasal turbinate and oropharyngeal samples were reduced in S-21 and S-29 vaccinated mice. Despite that, only the S-29 protein protected 100% of K18-hACE2 mice from severe disease. When GSH were analyzed, all immunized animals were protected from disease development irrespectively of the immunogen they received. Therefore, the higher yield of S-29, as well as its improved immunogenicity and efficacy protecting from the highly pathogenic SARS-CoV-2 Beta variant, pinpoint the S-29 mutant as an alternative to the S-2P protein for future SARS-CoV-2 vaccine development.
Assuntos
COVID-19 , SARS-CoV-2 , Cricetinae , Animais , Humanos , Camundongos , SARS-CoV-2/genética , Mesocricetus , COVID-19/prevenção & controle , Vacinas contra COVID-19RESUMO
The SARS-CoV-2 antigen-detecting rapid diagnostic test (Ag-RDTs) is an easy-to-use diagnostic tool to identify the contagious individuals and reduce the new infections. However, to be effective, Ag-RDTs require the detection of distinct variants of concern (VOC) with high analytical sensitivity. Here, we found that the VOC diverge at the nucleocapsid protein used by four commercial Ag-RDTs for the viral detection. Relative to the original D614G variant, there was a 10-fold loss of detection for the Delta and Alpha variants in certain Ag-RDTs, a reduction above the threshold required to isolate the viable virus. However, Beta and Omicron variants did not lose the detection capacity. As the new VOC arise, successful contact tracing requires continuous monitoring of Ag-RDTs performance.
RESUMO
The pandemic caused by the new coronavirus SARS-CoV-2 has made evident the need for broad-spectrum, efficient antiviral treatments to combat emerging and re-emerging viruses. Plitidepsin is an antitumor agent of marine origin that has also shown a potent pre-clinical efficacy against SARS-CoV-2. Plitidepsin targets the host protein eEF1A (eukaryotic translation elongation factor 1 alpha) and affects viral infection at an early, post-entry step. Because electron microscopy is a valuable tool to study virus-cell interactions and the mechanism of action of antiviral drugs, in this work we have used transmission electron microscopy (TEM) to evaluate the effects of plitidepsin in SARS-CoV-2 infection in cultured Vero E6 cells 24 and 48h post-infection. In the absence of plitidepsin, TEM morphological analysis showed double-membrane vesicles (DMVs), organelles that support coronavirus genome replication, single-membrane vesicles with viral particles, large vacuoles with groups of viruses and numerous extracellular virions attached to the plasma membrane. When treated with plitidepsin, no viral structures were found in SARS-CoV-2-infected Vero E6 cells. Immunogold detection of SARS-CoV-2 nucleocapsid (N) protein and double-stranded RNA (dsRNA) provided clear signals in cells infected in the absence of plitidepsin, but complete absence in cells infected and treated with plitidepsin. The present study shows that plitidepsin blocks the biogenesis of viral replication organelles and the morphogenesis of virus progeny. Electron microscopy morphological analysis coupled to immunogold labeling of SARS-CoV-2 products offers a unique approach to understand how antivirals such as plitidepsin work.
Assuntos
Tratamento Farmacológico da COVID-19 , Depsipeptídeos , Animais , Antivirais/uso terapêutico , Chlorocebus aethiops , Depsipeptídeos/farmacologia , Peptídeos Cíclicos , SARS-CoV-2 , Células Vero , Replicação ViralRESUMO
The emerging SARS-CoV-2 variants of concern (VOCs) may display enhanced transmissibility, more severity and/or immune evasion; however, the pathogenesis of these new VOCs in experimental SARS-CoV-2 models or the potential infection of other animal species is not completely understood. Here we infected K18-hACE2 transgenic mice with B.1, B.1.351/Beta, B.1.617.2/Delta and BA.1.1/Omicron isolates and demonstrated heterogeneous infectivity and pathogenesis. B.1.351/Beta variant was the most pathogenic, while BA.1.1/Omicron led to lower viral RNA in the absence of major visible clinical signs. In parallel, we infected wildtype (WT) mice and confirmed that, contrary to B.1 and B.1.617.2/Delta, B.1.351/Beta and BA.1.1/Omicron can infect them. Infection in WT mice coursed without major clinical signs and viral RNA was transient and undetectable in the lungs by day 7 post-infection. In silico modeling supported these findings by predicting B.1.351/Beta receptor binding domain (RBD) mutations result in an increased affinity for both human and murine ACE2 receptors, while BA.1/Omicron RBD mutations only show increased affinity for murine ACE2.
RESUMO
Viruses rely on the cellular machinery to replicate and propagate within newly infected individuals. Thus, viral entry into the host cell sets up the stage for productive infection and disease progression. Different viruses exploit distinct cellular receptors for viral entry; however, numerous viral internalization mechanisms are shared by very diverse viral families. Such is the case of Ebola virus (EBOV), which belongs to the filoviridae family, and the recently emerged coronavirus SARS-CoV-2. These two highly pathogenic viruses can exploit very similar endocytic routes to productively infect target cells. This convergence has sped up the experimental assessment of clinical therapies against SARS-CoV-2 previously found to be effective for EBOV, and facilitated their expedited clinical testing. Here we review how the viral entry processes and subsequent replication and egress strategies of EBOV and SARS-CoV-2 can overlap, and how our previous knowledge on antivirals, antibodies, and vaccines against EBOV has boosted the search for effective countermeasures against the new coronavirus. As preparedness is key to contain forthcoming pandemics, lessons learned over the years by combating life-threatening viruses should help us to quickly deploy effective tools against novel emerging viruses.
RESUMO
HIV-1 cell-to-cell transmission is key for an effective viral replication that evades immunity. This highly infectious mechanism is orchestrated by different cellular targets that utilize a wide variety of processes to efficiently transfer HIV-1 particles. Dendritic cells (DCs) are the most potent antigen presenting cells that initiate antiviral immune responses, but are also the cells with highest capacity to transfer HIV-1. This mechanism, known as trans-infection, relies on the capacity of DCs to capture HIV-1 particles via lectin receptors such as the sialic acid-binding I-type lectin Siglec-1/CD169. The discovery of the molecular interaction of Siglec-1 with sialylated lipids exposed on HIV-1 membranes has enlightened how this receptor can bind to several enveloped viruses. The outcome of these interactions can either mount effective immune responses, boost the productive infection of DCs and favour innate sensing, or fuel viral transmission via trans-infection. Here we review these scenarios focusing on HIV-1 and other enveloped viruses such as Ebola virus or SARS-CoV-2.
RESUMO
BACKGROUND: The banning of mass-gathering indoor events to prevent SARS-CoV-2 spread has had an important effect on local economies. Despite growing evidence on the suitability of antigen-detecting rapid diagnostic tests (Ag-RDT) for mass screening at the event entry, this strategy has not been assessed under controlled conditions. We aimed to assess the effectiveness of a prevention strategy during a live indoor concert. METHODS: We designed a randomised controlled open-label trial to assess the effectiveness of a comprehensive preventive intervention for a mass-gathering indoor event (a live concert) based on systematic same-day screening of attendees with Ag-RDTs, use of facial masks, and adequate air ventilation. The event took place in the Sala Apolo, Barcelona, Spain. Adults aged 18-59 years with a negative result in an Ag-RDT from a nasopharyngeal swab collected immediately before entering the event were randomised 1:1 (block randomisation stratified by age and gender) to either attend the indoor event for 5 hours or go home. Nasopharyngeal specimens used for Ag-RDT screening were analysed by real-time reverse-transcriptase PCR (RT-PCR) and cell culture (Vero E6 cells). 8 days after the event, a nasopharyngeal swab was collected and analysed by Ag-RDT, RT-PCR, and a transcription-mediated amplification test (TMA). The primary outcome was the difference in incidence of RT-PCR-confirmed SARS-CoV-2 infection at 8 days between the control and the intervention groups, assessed in all participants who were randomly assigned, attended the event, and had a valid result for the SARS-CoV-2 test done at follow-up. The trial is registered at ClinicalTrials.gov, NCT04668625. FINDINGS: Participant enrollment took place during the morning of the day of the concert, Dec 12, 2020. Of the 1140 people who responded to the call and were deemed eligible, 1047 were randomly assigned to either enter the music event (experimental group) or continue with normal life (control group). Of the 523 randomly assigned to the experimental group, 465 were included in the analysis of the primary outcome (51 did not enter the event and eight did not take part in the follow-up assessment), and of the 524 randomly assigned to the control group, 495 were included in the final analysis (29 did not take part in the follow-up). At baseline, 15 (3%) of 495 individuals in the control group and 13 (3%) of 465 in the experimental group tested positive on TMA despite a negative Ag-RDT result. The RT-PCR test was positive in one case in each group and cell viral culture was negative in all cases. 8 days after the event, two (<1%) individuals in the control arm had a positive Ag-RDT and PCR result, whereas no Ag-RDT nor RT-PCR positive results were found in the intervention arm. The Bayesian estimate for the incidence between the experimental and control groups was -0·15% (95% CI -0·72 to 0·44). INTERPRETATION: Our study provides preliminary evidence on the safety of indoor mass-gathering events during a COVID-19 outbreak under a comprehensive preventive intervention. The data could help restart cultural activities halted during COVID-19, which might have important sociocultural and economic implications. FUNDING: Primavera Sound Group and the #YoMeCorono Initiative. TRANSLATION: For the Spanish translation of the abstract see Supplementary Materials section.