Search Results (114)

The monkeypox virus (MPXV) is an emerging zoonotic pathogen responsible for mpox, a disease characterized by some smallpox-like symptoms, typically mild but occasionally fatal. The largest mpox recorded global outbreak began in May 2022, with over 162,000 cases across 140 countries. Herein, we have analyzed the intra-clonal diversity of MPXV obtained from a single skin lesion sample from a male patient (June 2022). Three viral clones were obtained following phenotypic evaluation of MPXV lysis plaque characteristics over a three-course infection in BSC-40 cells. Unlike the vaccinia virus Western Reserve (VACV-WR) strain, MPXV clones did not produce comet-like structures, suggesting reduced extracellular enveloped virus (EEV) morphotype release, which is associated with viral dissemination. Whole-genome sequencing and assembly identified subtle differences among clones. Comparative genomic analyses, including synteny and single nucleotide variation (SNV) calling, revealed intra-clonal differences and divergence from clade I and II references, although the variety of mutations found did not reveal possible variations at the protein level. Altogether, these findings suggest that although similar, it is possible that distinct MPXV variants may circulate together and can be found in a single exanthematous lesion. Full article

The ongoing global monkeypox outbreak since 2022 has highlighted the urgent need for vaccine development. Current vaccination strategies rely on cross-protective immunity provided by orthopoxvirus-based live-attenuated vaccines. However, these vaccines not only exhibit suboptimal efficacy against monkeypox virus (MPXV) but also raise safety concerns, particularly given the significant global overlap between MPXV infections and HIV. Owing to their superior safety profile and accessibility, recombinant subunit vaccines represent a highly promising platform for monkeypox vaccine development. In this study, we developed a subunit vaccine comprising A29L, B6R, and M1R antigens formulated with a proprietary nanoemulsion adjuvant and evaluated its immunogenicity and protective efficacy. In mice immunized with a prime-boost regimen of the three individual antigens combined with the nanoemulsion adjuvant, comparable serum IgG levels against each antigen were elicited. Both A29 and M1 formulations induced serum antibodies with potent neutralizing activity against MPXV and Vaccinia virus Western Reserve strain (VACV-WR). Notably, M1 antiserum exhibited stronger neutralization than A29 antiserum, whereas B6R immune serum showed no significant neutralizing activity. Splenocytes from B6R-immunized mice mounted a robust IFN-γ response, which was markedly lower in those immunized with A29 or M1. All three monovalent vaccines conferred complete survival following an intranasal lethal MPXV challenge, with M1 providing the strongest protection. In a lethal VACV-WR challenge model, only M1 immunization conferred significant protection. Histopathological analysis of lung tissues on day 5 post-infection revealed more pronounced inflammatory features in B6R-immunized mice compared to the nanoemulsion adjuvant control group. Furthermore, the nanoemulsion-adjuvanted bivalent A29L + B6R formulation induced significantly higher IgG and neutralizing antibody titers and demonstrated superior protective efficacy compared to the aluminum hydroxide-adjuvanted formulation. This comparative preclinical evaluation provides important evidence to support the development of a safe and effective subunit vaccine against monkeypox. Full article

Background/Objectives: Monkeypox is endemic to the African continent and has recently garnered global attention due to reported outbreaks in non-endemic nations. No approved drug is available for non-severe cases, and some isolates gained resistance to approved antivirals. In this study, we employed a drug repositioning strategy to evaluate the efficacy of existing FDA-approved antiviral drugs if repurposed for use against emerging Monkeypox, representing a cost-effective method for identifying novel therapeutic interventions. Methods: Methodology including Egyptian virus strain isolation, propagation and titration followed by in vitro studies, molecular docking and molecular dynamics simulations combined with binding free energy were carried out. Twenty-three FDA-approved drugs, including Abacavir, Acyclovir, Amantadine, Chloroquine, Daclatasvir, Dolutegravir, Entecavir, Favipiravir, Hydroxychloroquine, Lamivudine, Molnupiravir, Nevirapine, Oseltamivir, Penciclovir, Remdesivir, Ribavirin, Sofosbuvir, Tenofovir, Valaciclovir, Valganciclovir, Velpatasvir, Zanamivir, and Zidovudine, were screened for potential anti-monkeypox activity in vitro. In silico studies were carried out against three monkeypox proteins, Thymidylate Kinase, A42R Profilin-Like Protein, and VACV D13, to identify their potential targets. Results: In vitro testing showed that two antiviral drugs are positive. The employed computational methods indicate that remdesivir demonstrated superior binding patterns with elevated scores and stable complexes throughout the simulation. Conclusions: Our findings showed that Remdesivir therapeutic compound is potent against the tested strain of MPXV, and exhibited a robust binding affinity for Thymidylate Kinase, A42R Profilin-Like Protein, and VACV D13 enzymes, and thus may potentially be utilized as antiviral for the treatment of monkeypox virus. Full article

Protein kinase R (PKR) is an antiviral protein that is involved in molecular “arms races” with viral antagonists. As a result, some PKR inhibitors, including the vaccinia virus (VACV) protein K3 and its orthologs from other poxviruses only inhibit PKRs of selected species. We previously reported contrasting inhibition patterns of human, sheep, and cow PKRs by VACV K3 and the sheeppox virus (SPPV) K3 ortholog, SPPV 011. Here we show that the differential sensitivities of cow and sheep PKRs to VACV K3 were mediated by only two residues in PKR helix αG. In contrast, SPPV 011 sensitivities were governed by additional residues and regions. Analysis of the PKR sensitivities from 20 mammalian species to VACV K3 and SPPV 011 revealed four different sensitivity patterns: some PKRs were inhibited by only one K3 ortholog, as previously reported, whereas other PKRs were either resistant or sensitive to both inhibitors. Furthermore, we characterized a residue (K45) in VACV K3 that is involved in the species-specific inhibition of PKR. Mutating this residue increased the inhibition of sheep but not human PKR, whereas it decreased the inhibition of mouse PKR, highlighting that a single mutation in a viral protein can result in distinct species-dependent inhibition changes. Full article

The World Health Organization (WHO) declared the mpox (MPX) outbreak a public health emergency of international concern (PHEIC) on 23 July 2022, and 14 August 2024, respectively, underscoring the confirmed and concerning global spread of the disease. A gap exists in our fundamental understanding of the mpox virus (MPXV), despite its genetic relatedness to the variola virus (VARV). This knowledge deficit is evident in the performance of current medical countermeasures; vaccines and antiviral therapies adapted from smallpox programs demonstrate only partial efficacy and are constrained by issues of safety and suboptimal effectiveness against MPXV. In this context, the development of MPX-specific vaccines and antiviral drugs has become a critical priority in the global effort to combat MPX. However, MPXV employs multiple strategies to evade host immune responses, such as producing specific and poxvirus homologous proteins that suppress both innate immunity (including the six principal innate immune signaling pathways and antiviral strategies, notably the interferon [IFN] pathway) and adaptive immunity, thereby complicating vaccine and drug development. Insights from research on vaccinia virus (VACV) and VARV may inform the investigation of MPXV pathogenesis and immune evasion mechanisms. Drawing on relevant scientific literature, this review systematically examines key aspects of MPX infection, pathogenicity, and immune evasion, as well as the coordination between innate and adaptive immune responses. Furthermore, this review elucidates the current application and deployment landscape of the three principal therapeutics and three major vaccines for MPX, aiming to provide a theoretical foundation for future research and development of vaccines and targeted antiviral agents. Full article

Co-infections of Orthopoxviruses (OPVs), such as vaccinia virus (VACV) and monkeypox virus (MPXV), and the human immunodeficiency virus (HIV) can be associated with severe outcomes. Serro’s dairy region, located in Minas Gerais, southeastern Brazil, is an endemic area for VACV, where zoonotic outbreaks affect rural communities. This epidemiological context is especially relevant for at-risk populations, such as people living with HIV (PLHIV). This study aimed to assess the presence of neutralizing antibodies (NAbs) against OPV in PLHIV in this endemic setting. Serum samples were collected from 177 PLHIV in treatment at the specialized service between December 2021 and August 2022. VACV and MPXV NAbs were measured using the plaque reduction neutralization test (PRNT) and VACV-infected cells. The overall occurrence of OPV NAbs was 27.7%. NAbs were higher in individuals born before 1980 (53.3%) than those born after 1980 (1.1%). Among anti-VACV-seropositive individuals, 40.8% also had MPXV NAbs, suggesting cross-immunity. These findings indicate the circulation of VACV in PLHIV and highlight the increased susceptibility to OPV infections among individuals born after the cessation of smallpox vaccination. The results reinforce the importance of continued surveillance of OPV, especially in endemic regions and vulnerable populations. Full article

Despite significant advances in cancer therapy, the prognosis for patients with advanced, disseminated disease remains poor. This underscores the urgent need for novel treatments that not only eliminate tumor cells effectively but also stimulate a strong, durable anti-cancer immune response. Among emerging strategies, oncolytic viruses have shown exceptional promise due to their selective cytotoxicity and their ability to activate T cell-mediated immune responses. In this review, we focus on the vaccinia virus (VACV), a member of the Poxviridae family, which has emerged as a leading candidate in modern oncolytic immunotherapy. We examine the virus’s properties that enable it to evade antiviral defenses and serve as a versatile, potent oncolytic agent. Furthermore, we explore its interactions with various components of the immune system and how these contribute to the induction of a robust T cell-driven response. Finally, we assess current efforts to harness VACV for the treatment of various cancer types and highlight future directions where its application is most likely to succeed. Overall, our goal is to present VACV as a powerful and broadly applicable platform with the potential to transform the landscape of oncology. Full article

Background: The re-emerging mpox virus (MPXV) has spread to numerous countries and raised global concern. There is an urgent need for a safe and effective mRNA vaccine candidate against MPXV infection. Previously, we developed a penta-component mRNA vaccine that contained five distinct antigen-encoded mRNAs encapsulated within lipid nanoparticles (LNPs). Here, we sought to develop a single-chain mRNA vaccine that encodes antigens derived from both intracellular mature virion (IMV) and extracellular enveloped virion (EEV). Methods: A single-chain mRNA vaccine encoding a fusion protein comprising the ectodomains of M1R (eM1R) and A35R (eA35R) (MPXV^eM1-eA35) was developed and characterized, while an admixed formulation of two individual mRNA-LNPs encoding separate antigens was developed as the control (MPXV^eM1+eA35). Meanwhile, based on the same strategy, we designed a single-chain mRNA vaccine encoding dimeric antigens (MPXV^eM1-eA35-Fc). Mice were immunized with two doses of the candidate vaccines, and both humoral and cellular immune responses were evaluated. The protective efficacy of the candidate vaccines was evaluated based on body weight monitoring and tissue viral load measurement after challenge with vaccinia virus (VACV). Results: Immunization with two doses of MPXV^eM1-eA35 elicited robust levels of neutralizing antibodies and antigen-specific cellular immune response. Importantly, MPXV^eM1-eA35 demonstrated protective efficacy in a VACV challenge mouse model and showed superior capacity in preventing weight loss post-challenge compared to MPXV^eM1+eA35. Similarly, MPXV^eM1-eA35-Fc exhibited comparable or superior immunogenicity and protective efficacy compared to the admixed formulations. Conclusions: The single-chain mRNA vaccine elicited a protective immune response in mice, offering significant advantages in terms of manufacturing processes and quality control. Our single-chain mRNA vaccine platform presents a promising strategy for the next generation design of mpox vaccines and contributes to the mitigation of MPXV endemic worldwide. Full article

Background/Objectives: Monkeypox, twice declared a public health emergency of international concern by the WHO, currently lacks approved targeted therapeutics. This study focused on the development of monkeypox virus (MPXV) E8-specific human monoclonal antibodies (mAbs) derived from recipients of the recombinant vaccinia vaccine (rTV), with subsequent evaluation of their cross-neutralizing activity against orthopoxviruses, including the vaccinia virus (VACV) and MPXV. Methods: Three mAbs (C5, C9, and F8) were isolated from rTV vaccinees. Structural mapping characterized their binding domains on the MPXV E8 and VACV D8 proteins. Neutralization potency was assessed against the VACV TianTan strain and MPXV clade IIb. A combo was further evaluated in a VACV-infected mice model for clinical recovery and viral load reduction. Complement-dependent enhancement mechanisms were also investigated in vitro. Results: C9 targets the virion surface region of E8 and both the virion surface region and intravirion region of D8, showing cross-neutralization activity against the MPXV (IC₅₀ = 3.0 μg/mL) and VACV (IC₅₀ = 51.1 ng/mL) in vitro. All three antibodies demonstrated potent neutralization against the VACV in vitro: C5 (IC₅₀ = 3.9 ng/mL), C9 (IC₅₀ = 51.1 ng/mL), and F8 (IC₅₀ = 101.1 ng/mL). Notably, complement enhanced neutralization against the VACV by >50-fold, although no enhancement was observed for the MPXV. In vivo administration accelerated clinical recovery by 24 h and achieved significant viral clearance (0.9-log reduction). Conclusions: E8-targeting mAbs exhibited broad-spectrum neutralization against orthopoxviruses, demonstrating therapeutic potential against both historical (VACV) and emerging (MPXV) pathogens. However, MPXV’s resistance to complement-dependent enhancement highlights the necessity for pathogen-adapted optimization. These findings establish E8 as a critical conserved target for pan-poxvirus VACV and MPXV countermeasure development. Full article

Background: Since the outbreak of mpox in 2022, the disease has spread rapidly worldwide and garnered significant public attention. Vaccination is regarded as an effective measure to prevent the spread of mpox. The success of the COVID-19 mRNA vaccine demonstrates that mRNA-based vaccines represent a rapid and multifunctional platform with considerable potential, and are expected to be a strategy to address mpox spread. Methods: In this study, we screened an mpox multi-antigen-tandem bivalent mRNA vaccine candidate: a lipid nanoparticle-encapsulated mRNA-1017 and mRNA-1995 (mRNA-3012-LNP). We then evaluated the immunogenicity of the mpox virus (MPXV) bivalent mRNA vaccine candidate and its protective efficacy against the vaccinia virus (VACV) in a mouse model. Results: Mice vaccinated with two doses of the mRNA-3012-LNP vaccine exhibited robust binding antibody responses and MPXV-specific Th-1-biased cellular immune responses in vivo. Notably, the boosted immunized mice generated potent neutralizing antibodies against the VACV, effectively protecting them from viral challenge. Additionally, serum transfer protection experiments indicated that serum from mice inoculated with mRNA-3012-LNP was effective in protecting nude mice from VACV challenge. Conclusions: Our results suggest that the mpox bivalent mRNA candidate vaccine mRNA-3012-LNP induces strong immunogenicity and has the potential to serve as a safe and effective vaccine candidate against mpox epidemics. Full article

This study aimed to optimize the electrochemical pretreatment and functionalization of pencil graphite electrodes (PGEs) for the performance evaluation of a transducer applied in initial studies in the development of an immunosensor for vaccinia virus (VACV) detection. The effects of the applied potential, duration, and supporting electrolyte type and concentration on PGE activation were investigated. Functionalization using a polymeric film derived from 2-hydroxybenzamide (2-HXB) was optimized by varying the applied potential, deposition time, and monomer concentration. Optimal activation conditions were found to be +0.90 V in 0.02 M of H₂SO₄ for 300 s, promoting the formation of hydrogenated groups and increasing electrode wettability. For electropolymerization, +1.20 V for 300 s with a 2-HXB concentration of 2.50 mM provided the best results, ensuring proper film formation and adhesion. Scanning electron microscopy revealed a rough, sheet-like surface on the polished PGE, while energy dispersive spectroscopy confirmed poly(2-HXB) adsorption through increased oxygen and nitrogen content on the functionalized electrode. The optimized pretreatment and functionalization conditions significantly influenced the response of the transducer used for VACV detection, demonstrating its crucial role in device development. These findings contribute to the advancement of inexpensive and effective electrochemical transducers and highlight the importance of pretreatment and modification of PGEs in biosensing applications. Full article

Amid the current multi-country mpox outbreak, analyzing monkeypox virus (MPXV) and vaccinia virus (VACV) genomes is vital for understanding evolutionary processes that may impact vaccine efficacy and design. This study aimed to elucidate the phylogenetic relationships and structural features of viral antigens, which are crucial for developing effective vaccines. By aligning 1903 MPXV genomes from the NCBI Virus repository (released between 2022 and 2024), an increase in phylogenetic diversity was observed compared to previous studies. These genomes were grouped into Clade I (25 genomes) and Clade IIB (1898 genomes), with a new Clade I sub-lineage emerging from samples collected in Sud-Kivu province, Democratic Republic of the Congo (DRC). Comparing six key MPXV neutralization determinants (A29, A35, B6, E8, H3, and M1) of a novel 2024 Clade I MPXV isolate to those of the 1996 Zaire isolate revealed remarkable sequence conservation despite spanning 28 years. Homology-based modeling of the Clade I MPXV antigens (A29, A35, E8, H3, and M1) showed high-match identities (84% to 99%) with VACV templates (current mpox vaccine), with several amino acid variants near potential antibody binding sites. Phylogenomic analysis, combined with structural modeling and variant profiling, has yielded valuable insights into the virus and vaccine, guiding vaccine design and functional studies. Full article

Background/Objectives: In preparation for a potential pandemic caused by the H5N1 highly pathogenic avian influenza (HPAI) virus, pre-pandemic vaccines against several viral clades have been developed and stocked worldwide. Although these vaccines are well tolerated, their immunogenicity and cross-reactivity with viruses of different clades can be improved. Methods: To address this aspect, we generated recombinant influenza vaccines against H5-subtype viruses using two different strains of highly attenuated vaccinia virus (VACV) vectors. Results: rLC16m8-mcl2.2 hemagglutinin (HA) and rLC16m8-mcl2.3.4 HA consisted of a recombinant LC16m8 vector encoding the HA protein from clade 2.2 or clade 2.3.4 viruses (respectively); rDIs-mcl2.2 HA consisted of a recombinant DIs vector encoding the HA protein from clade 2.2. A single dose of rLC16m8-mcl2.2 HA showed rapid (1 week after vaccination) and long-term protection (20 months post-vaccination) in mice against the HPAI H5N1 virus. Moreover, cynomolgus macaques immunized with rLC16m8-mcl2.2 HA exhibited long-term protection when challenged with a heterologous clade of the HPAI H5N1 virus. Although the DIs strain is unable to grow in most mammalian cells, rDIs-mcl2.2 HA also showed rapid and long-lasting effects against HPAI H5N1 virus infection. Notably, the protective efficacy of rDIs-mcl2.2 HA was comparable to that of rLC16m8-mcl2.2 HA. Furthermore, these vaccines protected animals previously immunized with VACVs from a lethal challenge with the HPAI H5N1 virus. Conclusions: These results suggest that both rLC16m8-mcl2.2 HA and rDIs-mcl2.2 HA are effective in preventing HPAI H5N1 virus infection, and rDIs-mcl2.2 HA is a promising vaccine candidate against H5 HA-subtype viruses. Full article

The ongoing monkeypox (mpox) disease outbreak has spread to multiple countries in Central Africa and evidence indicates it is driven by a more virulent clade I monkeypox virus (MPXV) strain than the clade II strain associated with the 2022 global mpox outbreak, which led the WHO to declare this mpox outbreak a public health emergency of international concern. The FDA-approved small molecule antiviral tecovirimat (TPOXX) is recommended to treat mpox cases with severe symptoms, but the limited efficacy of TPOXX and the emergence of TPOXX resistant MPXV variants has challenged this medical practice of care and highlighted the urgent need for alternative therapeutic strategies. In this study we have used vaccinia virus (VACV) as a surrogate of MPXV to assess the antiviral efficacy of combination therapy of TPOXX together with mycophenolate mofetil (MMF), an FDA-approved immunosuppressive agent that we have shown to inhibit VACV and MPXV, or the N-myristoyltransferase (NMT) inhibitor IMP-1088. Both MMF and IMP-1088 drugs exhibited strong dose-dependent antiviral activity against VACV and mpox, and potent synergistic effects in conjunction with TPOXX. Our findings support combination therapy of direct-acting (TPOXX) and host-targeted (MMF and IMP-1088) antivirals as a promising approach to treat mpox and prevent the emergence and spread of TPOXX-resistant MPXV variants. Full article

Background: The recent resurgence of mpox in central Africa has been declared a new public health emergency of international concern (PHEIC) requiring coordinated international responses. Vaccination is a priority to expand protection and enhance control strategies, but the vaccine’s need exceeds the currently available doses. Intradermal (ID) administration of one-fifth of the standard modified vaccinia Ankara (MVA-BN) dose was temporarily authorized during the 2022 PHEIC. Studies conducted before 2022 provided evidence about the humoral response against the vaccinia virus (VACV) after vaccination but not against the mpox virus (MPXV). Moreover, no data are available on the T-cell response elicited by MVA-BN administered subcutaneously or intradermally. Methods: We compare the two vaccine administration routes according to reactogenicity (n = 943) and immunogenicity (n = 225) of vaccine recipients attending INMI Spallanzani hospital during the 2022 vaccination campaign in Rome, Italy. Results: We found that the ID route elicited higher titers of MPXV-specific IgG (mean difference of 0.26 log₂, p = 0.05) and nAbs (0.24 log₂, p = 0.08) than the subcutaneous (SC) route one month after the complete vaccination cycle. At the same time, no evidence for a difference in cellular response was found. Conclusions: MVA-BN was globally well tolerated despite higher reactogenicity for the ID than the SC route, especially for the reactions at the local injection site. The ID dose-sparing strategy was proven safe and immunogenic and would make vaccination available to more people. Our data support the current WHO recommendation of using the ID route in low–medium-income countries (LMIC), although response data in people infected with the new 1b clade are urgently needed. Full article