Intradermally Administered Yellow Fever Vaccine at Reduced Dose Induces a Protective Immune Response: A Randomized Controlled Non-Inferiority Trial

Background:Implementation of yellow fever vaccination is currently hampered by limited supply of vaccine. An alternative route of administration with reduced amounts of vaccine but without loss of vaccine efficacy would boost vaccination programmes.Methods and Findings:A randomized, controlled, non-inferiority trial was conducted in a Dutch university center between August 2005 and February 2007. A total of 155 primary vaccinated and 20 previously vaccinated volunteers participated. Participants were randomly assigned in a 1:1 ratio to receive intradermal (i.d.) vaccination with live attenuated yellow fever 17D vaccine at a reduced dose (1/5th; 0·1 mL) or the conventional subcutaneous (s.c.) vaccination (0·5 mL). Antibody neutralization titers were determined at 2, 4 and 8 weeks and 1 year after vaccination by counting the reduction in virus-induced plaques in the presence of serial serum dilutions. Adverse events were documented in a 3-week dairy. Viraemia was measured 5 days after vaccination. From 2 weeks up to one year after vaccination, the maximum serum-dilution at which 80% of the virus plaques were neutralized, which indicates protection against yellow fever, did not differ between those given a reduced i.d. dose or standard s.c. dose of vaccine. In all cases the WHO standard of seroprotection (i.e. 80% virus neutralization) was reached (in 77/77 and 78/78, respectively). Similar results were found in the previously vaccinated individuals. Viraemia was detected in half of the primary vaccinated participants, which was not predictive of serological response. In revaccinees no viraemia was detected.Conclusions:Intradermal administration of one fifth of the amount of yellow fever vaccine administered subcutaneously results in protective seroimmunity in all volunteers. Albeit this vaccination route should enable vaccination of five-times as many individuals at risk for disease, these results should now be confirmed in field studies in areas with potential yellow fever virus transmission to change vaccination policy.Trial Registration:Nederlands Trial Register ISRCTN46326316

All genera of Flaviviridae host a conserved Xrn1-resistant RNA motif

Computer Systems, Imagery and Medi

The enzymatic activity of the nsp14 exoribonuclease is critical for replication of MERS-CoV and SARS-CoV-2

Coronaviruses (CoVs) stand out for their large RNA genome and complex RNA-synthesizing machinery comprising 16 nonstructural proteins (nsps). The bifunctional nsp14 contains 3'-to-5' exoribonuclease (ExoN) and guanine-N7-methyltransferase (N7-MTase) domains. While the latter presumably supports mRNA capping, ExoN is thought to mediate proofreading during genome replication. In line with such a role, ExoN knockout mutants of mouse hepatitis virus (MHV) and severe acute respiratory syndrome coronavirus (SARS-CoV) were previously reported to have crippled but viable hypermutation phenotypes. Remarkably, using reverse genetics, a large set of corresponding ExoN knockout mutations has now been found to be lethal for another betacoronavirus, Middle East respiratory syndrome coronavirus (MERS-CoV). For 13 mutants, viral progeny could not be recovered, unless-as happened occasionally-reversion had first occurred. Only a single mutant was viable, likely because its E191D substitution is highly conservative. Remarkably, a SARSCoV-2 ExoN knockout mutant was found to be unable to replicate, resembling observations previously made for alphaand gammacoronaviruses, but starkly contrasting with the documented phenotype of ExoN knockout mutants of the closely related SARS-CoV. Subsequently, we established in vitro assays with purified recombinant MERS-CoV nsp14 to monitor its ExoN and N7-MTase activities. All ExoN knockout mutations that proved lethal in reverse genetics were found to severely decrease ExoN activity while not affecting N7-MTase activity. Our study strongly suggests that CoV nsp14 ExoN has an additional function, which apparently is critical for primary viral RNA synthesis and thus differs from the proofreading function that, based on previous MHV and SARS-CoV studies, was proposed to boost longer-term replication fidelity. fidelity.IMPORTANCE The bifunctional nsp14 subunit of the coronavirus replicase contains 3'-to-5' exoribonuclease (ExoN) and guanine-N7-methyltransferase domains. For the betacoronaviruses MHV and SARS-CoV, ExoN was reported to promote the fidelity of genome replication, presumably by mediating a form of proofreading. For these viruses, ExoN knockout mutants are viable while displaying an increased mutation frequency. Strikingly, we have now established that the equivalent ExoN knockout mutants of two other betacoronaviruses, MERS-CoV and SARS-CoV-2, are nonviable, suggesting an additional and critical ExoN function in their replication. This is remarkable in light of the very limited genetic distance between SARS-CoV and SARS-CoV-2, which is highlighted, for example, by 95% amino acid sequence identity in their nsp14 sequences. For (recombinant) MERSCoV nsp14, both its enzymatic activities were evaluated using newly developed in vitro assays that can be used to characterize these key replicative enzymes in more detail and explore their potential as target for antiviral drug development.Molecular basis of virus replication, viral pathogenesis and antiviral strategie

A Yellow Fever 17D virus replicon-based vaccine platform for emerging Coronaviruses

The tremendous global impact of the current SARS-CoV-2 pandemic, as well as other current and recent outbreaks of (re)emerging viruses, emphasize the need for fast-track development of effective vaccines. Yellow fever virus 17D (YF17D) is a live-attenuated virus vaccine with an impressive efficacy record in humans, and therefore, it is a very attractive platform for the development of novel chimeric vaccines against various pathogens. In the present study, we generated a YF17D-based replicon vaccine platform by replacing the prM and E surface proteins of YF17D with antigenic subdomains from the spike (S) proteins of three different betacoronaviruses: MERS-CoV, SARS-CoV and MHV. The prM and E proteins were provided in trans for the packaging of these RNA replicons into single-round infectious particles capable of expressing coronavirus antigens in infected cells. YF17D replicon particles expressing the S1 regions of the MERS-CoV and SARS-CoV spike proteins were immunogenic in mice and elicited (neutralizing) antibody responses against both the YF17D vector and the coronavirus inserts. Thus, YF17D replicon-based vaccines, and their potential DNA- or mRNA-based derivatives, may constitute a promising and particularly safe vaccine platform for current and future emerging coronaviruses.Molecular basis of virus replication, viral pathogenesis and antiviral strategie

Suramin inhibits SARS-CoV-2 infection in cell culture by interfering with early steps of the replication cycle

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic that originated in Wuhan, China, in December 2019 has impacted public health, society, the global economy, and the daily lives of billions of people in an unprecedented manner. There are currently no specific registered antiviral drugs to treat or prevent SARS-CoV-2 infections. Therefore, drug repurposing would be the fastest route to provide at least a temporary solution while better, more specific drugs are being developed. Here, we demonstrate that the antiparasitic drug suramin inhibits SARS-CoV-2 replication, protecting Vero E6 cells with a 50% effective concentration (EC50) of similar to 20 mu M, which is well below the maximum attainable level in human serum. Suramin also decreased the viral load by 2 to 3 logs when Vero E6 cells or cells of a human lung epithelial cell line (Calu-3 2B4 [referred to here as "Calu-3"]) were treated. Time-of-addition and plaque reduction assays performed on Vero E6 cells showed that suramin acts on early steps of the replication cycle, possibly preventing binding or entry of the virus. In a primary human airway epithelial cell culture model, suramin also inhibited the progression of infection. The results of our preclinical study warrant further investigation and suggest that it is worth evaluating whether suramin provides any benefit for COVID-19 patients, which obviously requires safety studies and well-designed, properly controlled randomized clinical trials.Pathogenesis and treatment of chronic pulmonary disease

Quasispecies composition and evolution of a typical Zika virus clinical isolate from Suriname

Molecular Technology and Informatics for Personalised Medicine and Healt

Engineering potent live attenuated coronavirus vaccines by targeted inactivation of the immune evasive viral deubiquitinase

Coronaviruses express a papain-like protease (PLpro) that is required for replicase polyprotein maturation and also serves as a deubiquitinating enzyme (DUB). In this study, using a Middle East respiratory syndrome virus (MERS-CoV) PLpro modified virus in which the DUB is selectively inactivated, we show that the PLpro DUB is an important MERS-CoV interferon antagonist and virulence factor. Although the DUB-negative rMERS-CoVMA replicates robustly in the lungs of human dipeptidyl peptidase 4 knock-in (hDPP4 KI) mice, it does not cause clinical symptoms. Interestingly, a single intranasal vaccination with DUB-negative rMERS-CoVMA induces strong and sustained neutralizing antibody responses and sterilizing immunity after a lethal wt virus challenge. The survival of naive animals also significantly increases when sera from animals vaccinated with the DUB-negative rMERS-CoVMA are passively transferred, prior to receiving a lethal virus dose. These data demonstrate that DUB-negative coronaviruses could be the basis of effective modified live attenuated vaccines.In this work, authors provide a proof-of-concept study showing that deubiquitinating enzyme inactivation in MERS-CoV leads to attenuation in mice, and protection against a lethal challenge.Horizon 2020 (H2020)952373Molecular basis of virus replication, viral pathogenesis and antiviral strategie

Molecular and immunological characterization of a DNA-launched yellow fever virus 17D infectious clone

Molecular basis of virus replication, viral pathogenesis and antiviral strategie