A multivalent mRNA monkeypox virus vaccine (BNT166) protects mice and macaques from orthopoxvirus disease.

In response to the 2022 outbreak of mpox driven by unprecedented human-to-human monkeypox virus (MPXV) transmission, we designed BNT166, aiming to create a highly immunogenic, safe, accessible, and scalable next-generation vaccine against MPXV and related orthopoxviruses. To address the multiple viral forms and increase the breadth of immune response, two candidate multivalent mRNA vaccines were evaluated pre-clinically: a quadrivalent vaccine (BNT166a; encoding the MPXV antigens A35, B6, M1, H3) and a trivalent vaccine (BNT166c; without H3). Both candidates induced robust T cell responses and IgG antibodies in mice, including neutralizing antibodies to both MPXV and vaccinia virus. In challenge studies, BNT166a and BNT166c provided complete protection from vaccinia, clade I, and clade IIb MPXV. Furthermore, immunization with BNT166a was 100% effective at preventing death and at suppressing lesions in a lethal clade I MPXV challenge in cynomolgus macaques. These findings support the clinical evaluation of BNT166, now underway (NCT05988203).

Molecular Virology of Orthopoxviruses with Special Reference to Monkeypox Virus.

Poxviruses are large (200-450 nm) and enveloped viruses carrying double-stranded DNA genome with an epidermal cell-specific adaptation. The genus Orthopoxvirus within Poxviridae family constitutes several medically and veterinary important viruses including variola (smallpox), vaccinia, monkeypox virus (MPXV), and cowpox. The monkeypox disease (mpox) has recently emerged as a public health emergency caused by MPXV. An increasing number of human cases of MPXV have been documented in non-endemic nations without any known history of contact with animals brought in from endemic and enzootic regions, nor have they involved travel to an area where the virus was typically prevalent. Here, we review the MPXV replication, virus pathobiology, mechanism of viral infection transmission, virus evasion the host innate immunity and antiviral therapies against Mpox. Moreover, preventive measures including vaccination were discussed and concluded that cross-protection against MPXV may be possible using antibodies that are directed against an Orthopoxvirus. Despite the lack of a specialised antiviral medication, several compounds such as Cidofovir and Ribavirin warrant consideration against mpox.

Type I Interferon: Monkeypox/Mpox Viruses Achilles Heel?

Poxviruses are notorious for having acquired/evolved numerous genes to counteract host innate immunity. Chordopoxviruses have acquired/evolved at least three different inhibitors of host necroptotic death: E3, which blocks ZBP1-dependent necroptotic cell death, and vIRD and vMLKL that inhibit necroptosis downstream of initial cell death signaling. While this suggests the importance of the necroptotic cell death pathway in inhibiting chordopoxvirus replication, several chordopoxviruses have lost one or more of these inhibitory functions. Monkeypox/mpox virus (MPXV) has lost a portion of the N-terminus of its E3 homologue. The N-terminus of the vaccinia virus E3 homologue serves to inhibit activation of the interferon-inducible antiviral protein, ZBP1. This likely makes MPXV unique among the orthopoxviruses in being sensitive to interferon (IFN) treatment in many mammals, including humans, which encode a complete necroptotic cell death pathway. Thus, IFN sensitivity may be the Achille's Heel for viruses like MPXV that cannot fully inhibit IFN-inducible, ZBP1-dependent antiviral pathways.

A plague passing over: Clinical features of the 2022 mpox outbreak in patients of color living with HIV.

INTRODUCTION: Compared with previous geographically localized outbreaks of monkeypox (MPOX), the scale of the 2022 global mpox outbreak has been unprecedented, yet the clinical features of this outbreak remain incompletely characterized. METHODS: We identified patients diagnosed with mpox by polymerase chain reaction (PCR; n = 36) from July to September 2022 at a single, tertiary care institution in the USA. Demographics, clinical presentation, infection course, and histopathologic features were reviewed. RESULTS AND CONCLUSION: Men who have sex with men (89%) and people living with HIV (97%) were disproportionately affected. While fever and chills (56%) were common, some patients (23%) denied any prodromal symptoms. Skin lesions showed a wide range of morphologies, including papules and pustules, and lesions showed localized, not generalized, spread. Erythema was also less appreciable in skin of colour patients (74%). Atypical clinical features and intercurrent skin diseases masked the clinical recognition of several cases, which were ultimately diagnosed by PCR. Biopsies showed viral cytopathic changes consistent with Orthopoxvirus infections. All patients in this case series recovered without complications, although six patients (17%) with severe symptoms were treated with tecovirimat without complication.

The effective factors in human-specific tropism and viral pathogenicity in orthopoxviruses.

The orthopoxvirus (OPV) genus includes several species that infect humans, including variola, monkeypox, vaccinia, and cowpox. Variola and monkeypox are often life-threatening diseases, while vaccinia and cowpox are usually associated with local lesions. The epidemic potential for OPVs may be lower than respiratory-borne viruses or RNA viruses. However, OPVs are notable for their spread and distribution in different environments and among different hosts. The emergence or re-emergence of OPVs in the human population can also occur in wild or domestic animals as intermediate hosts. More effective and safer vaccines for poxvirus can be developed by understanding how immunity is regulated in poxvirus and vaccines for DNA viruses. Downstream events in cells affected by the virus are regulated functionally by a series of characteristics that are affected by host cell interactions and responses of cells against viral infections, including the interferon pathway and apoptosis. Furthermore, infection outcome is greatly influenced by the distinct selection of host-range and immune-modulatory genes that confer the potential for pathogenesis and host-to-host transmission and the distinct host-range properties of each immune-modulatory gene. The present study reviewed the effective factors in human-restricted tropism and virus pathogenicity in OPVs.

Synthesis of esters and amides of 2-aryl-1-hydroxy-4-methyl-1H-imidazole-5-carboxylic acids and study of their antiviral activity against orthopoxviruses.

Smallpox was eradicated >40 years ago but it is not a reason to forget forever about orthopoxviruses pathogenic to humans. Though in 1980 the decision of WHO to cease vaccination against smallpox had seemed logical, it led to the decrease of cross immunity against other infections caused by orthopoxviruses. As a result, in 2022 the multi-country monkeypox outbreak becomes a topic of great concern. In spite of existing FDA-approved drugs for the treatment of such diseases, the search for new small-molecule orthopoxvirus inhibitors continues. In the course of this search a series of novel 2-aryl-1-hydroxyimidazole derivatives containing ester or carboxamide moieties in position 5 of heterocycle has been synthesized and tested for activity against Vaccinia virus in Vero cell culture. Some of the compounds under consideration revealed a selectivity index higher than that of the reference drug Cidofovir. The highest selectivity index SI = 919 was exhibited by ethyl 1-hydroxy-4-methyl-2-[4-(trifluoromethyl)phenyl]-1H-imidazole-5-carboxylate 1f. The most active compound also demonstrated inhibitory activity against the cowpox virus (SI = 20) and the ectromelia virus (SI = 46).

Evaluation of 11 commercially available PCR kits for the detection of monkeypox virus DNA, Berlin, July to September 2022.

Before the international spread of monkeypox in May 2022, PCR kits for the detection of orthopoxviruses, and specifically monkeypox virus, were rarely available. Here we describe the evaluation of 11 recently developed commercially available PCR kits for the detection of monkeypox virus DNA. All tested kits are currently intended for research use only and clinical performance still needs to be assessed in more detail, but all were suitable for diagnostics of monkeypox virus, with variations in specificity rather than sensitivity.

Knowledge of Human Monkeypox and Its Relation to Conspiracy Beliefs among Students in Jordanian Health Schools: Filling the Knowledge Gap on Emerging Zoonotic Viruses.

Background and Objective: The recent multi-country outbreak of human monkeypox (HMPX) in non-endemic regions poses an emerging public health concern. University students in health schools/faculties represent a core knowledgeable group that can be helpful to study from a public health point of view. As future healthcare workers, assessment of their knowledge and attitude towards emerging zoonotic viral infections can be helpful to assess their taught material and courses with potential improvement if gaps in knowledge were identified. Therefore, we aimed to evaluate the level of HMPX knowledge, conspiracy beliefs regarding emerging virus infections, as well as their associated determinants among university students studying Medicine, Nursing, Dentistry, Pharmacy, Medical Laboratory Sciences, and Rehabilitation in Jordanian health schools/faculties. In addition, we sought to evaluate the correlation between HMPX knowledge and the extent of holding conspiracy beliefs regarding emerging viral infection. Materials and Methods: A convenient sample of university students was obtained through an electronic survey distributed in late May 2022 using the chain-referral approach. Assessment of HMPX knowledge and general attitude towards emerging virus infections was based on survey items adopted from previously published literature. Results: The study sample comprised 615 students with a mean age of 20 years and a majority of females (432, 70.2%) and medical students (n = 351, 57.1%). Out of eleven monkeypox knowledge items, three were identified correctly by >70% of the respondents. Only 26.2% of the respondents (n = 161) knew that vaccination to prevent monkeypox is available. Age was significantly associated with better HMPX knowledge for a majority of items. Older age, females, and affiliation to non-medical schools/faculties were associated with harboring higher levels of conspiracy beliefs regarding emerging virus infections. Our data also indicate that lower levels of HMPX knowledge were associated with higher levels of conspiracy beliefs. Conclusion: The current study pointed to generally unsatisfactory levels of knowledge regarding the emerging HMPX among university students in Jordanian health schools/faculties. Conspiracy beliefs regarding emerging virus infections were widely prevalent, and its potential detrimental impact on health behavior should be evaluated in future studies.

Viral Replicative Capacity, Antigen Availability via Hematogenous Spread, and High TFH:TFR Ratios Drive Induction of Potent Neutralizing Antibody Responses.

Live viral vaccines elicit protective, long-lived humoral immunity, but the underlying mechanisms through which this occurs are not fully elucidated. Generation of affinity matured, long-lived protective antibody responses involve close interactions between T follicular helper (TFH) cells, germinal center (GC) B cells, and T follicular regulatory (TFR) cells. We postulated that escalating concentrations of antigens from replicating viruses or live vaccines, spread through the hematogenous route, are essential for the induction and maintenance of long-lived protective antibody responses. Using replicating and poorly replicating or nonreplicating orthopox and influenza A viruses, we show that the magnitude of TFH cell, GC B cell, and neutralizing antibody responses is directly related to virus replicative capacity. Further, we have identified that both lymphoid and circulating TFH:TFR cell ratios during the peak GC response can be used as an early predictor of protective, long-lived antibody response induction. Finally, administration of poorly or nonreplicating viruses to allow hematogenous spread generates significantly stronger TFH:TFR ratios and robust TFH, GC B cell and neutralizing antibody responses.IMPORTANCE Neutralizing antibody response is the best-known correlate of long-term protective immunity for most of the currently licensed clinically effective viral vaccines. However, the host immune and viral factors that are critical for the induction of robust and durable antiviral humoral immune responses are not well understood. Our study provides insight into the dynamics of key cellular mediators of germinal center reaction during live virus infections and the influence of viral replicative capacity on the magnitude of antiviral antibody response and effector function. The significance of our study lies in two key findings. First, the systemic spread of even poorly replicating or nonreplicating viruses to mimic the spread of antigens from replicating viruses due to escalating antigen concentration is fundamental to the induction of durable antibody responses. Second, the TFH:TFR ratio may be used as an early predictor of protective antiviral humoral immune responses long before memory responses are generated.

A many probes-one spot hybridization oligonucleotide microarray.

A variant of the hybridization oligonucleotide microarray, utilizing the principle of many probes-one spot (MPOS-microarrays), is proposed. A case study based on Orthopoxviruses (Variola, Monkeypox, and Ectromelia viruses) demonstrates a considerable increase in the fluorescence signal (up to 100-fold) when several oligonucleotide probes are printed to one spot. Moreover, the specificity of detection also increases (almost 1000-fold), allowing the use of probes that individually lack such high specificity. The optimal probes have a Tm of 32-37 °C and length of 13-15 bases. We suggest that the high specificity and sensitivity of the MPOS-microarray is a result of cooperativity of DNA binding with all probes immobilized in the spot. This variant of DNA detection can be useful for designing biosensors, tools for point-of-care (POC) diagnostics, microbial ecology, analysis of clustered regularly interspaced short palindromic repeats (CRISPR), and others. Graphical abstract ᅟ.

Discovery of a New Class of Inhibitors of Vaccinia Virus Based on (-)-Borneol from Abies sibirica and (+)-Camphor.

A series of the bornyl ester/amide derivatives with N-containing heterocycles were designed and synthesized as vaccinia virus (VV) inhibitors. Bioassay results showed that among the designed compounds, derivatives 6, 13, 14, 34, 36 and 37 showed the best inhibitory activity against VV with the IC50 values of 12.9, 17.9, 3.4, 2.5, 12.5 and 7.5 µm, respectively, and good cytotoxicity. The primary structure-activity relationship (SAR) study suggested that the combination of a saturated N-heterocycle, such as morpholine or 4-methylpiperidine, and a 1,7,7-trimethylbicyclo[2.2.1]heptane scaffold was favorable for antiviral activity.

Antiviral Activity of Lady's Mantle (Alchemilla vulgaris L.) Extracts against Orthopoxviruses.

We studied toxicity and antiviral activity of bioactive substances extracted from the roots (ethylacetate extracts) and aerial parts (ethanol extracts) of lady's mantle (Alchemilla vilgaris L.). Plant extracts are characterized by low toxicity for continuous Vero cell culture, but inhibit the reproduction of orthopoxviruses (vaccinia virus and ectromelia virus) in these cells. Of all studied extracts, ethylacetate extract from lady's mantle roots characterized by the highest content of catechins in comparison with other samples demonstrated the highest activity in vitro towards the studied viruses (neutralization index for vaccinia and ectromelia viruses were 4.0 and 3.5 lg, respectively). The antiviral effect of Alchemilla vulgaris L. extracts was shown to be dose dependent.

Toxicity and Antiviral Activity of the Extracts of Submerged Mycelium of Nematophagous Duddingtonia flagrans Fungus in Vero Cell Culture.

We studied toxicity and antiviral activity of aqueous and ethanol extracts of bioactive substances from the biomass of nematophagous fungus Duddingtonia flagrans prepared by submerged culturing of the mycelium. It is found that both extracts were characterized by low toxicity for cultured Vero cells and inhibited reproduction of DNA-viruses in this cell line. Ethanol extract of the fungus exhibited higher in vitro antiviral activity against Herpes simplex virus type 2, ectromelia virus, and vaccinia virus than water extract, which can be due to higher content of proteins, polysaccharides, flavonols, catechins, or carotenes or more effective their combination. The extracts of cultured mycelium of Duddingtonia flagrans fungus containing a complex of bioactive substances can be used for creation of broad-spectrum antiviral drugs against DNA-viruses.

TRIM5α: A Protean Architect of Viral Recognition and Innate Immunity.

The evolutionary pressures exerted by viral infections have led to the development of various cellular proteins with potent antiviral activities, some of which are known as antiviral restriction factors. TRIpartite Motif-containing protein 5 alpha (TRIM5α) is a well-studied restriction factor of retroviruses that exhibits virus- and host-species-specific functions in protecting against cross-primate transmission of specific lentiviruses. This specificity is achieved at the level of the host gene through positive selection predominantly within its C-terminal B30.2/PRYSPRY domain, which is responsible for the highly specific recognition of retroviral capsids. However, more recent work has challenged this paradigm, demonstrating TRIM5α as a restriction factor for retroelements as well as phylogenetically distinct viral families, acting similarly through the recognition of viral gene products via B30.2/PRYSPRY. This spectrum of antiviral activity raises questions regarding the genetic and structural plasticity of this protein as a mediator of the recognition of a potentially diverse array of viral molecular patterns. This review highlights the dynamic evolutionary footprint of the B30.2/PRYSPRY domain in response to retroviruses while exploring the guided 'specificity' conferred by the totality of TRIM5α's additional domains that may account for its recently identified promiscuity.

[Comparative analysis of the taxonomic classification criteria for a number of groups of pathogenic DNA and RNA viruses based on genomic data].

The basis for criteria of the taxonomic classification of DNA and RNA viruses based on data of the genomic sequencing are viewed in this review. The genomic sequences of viruses, which have genome represented by double-stranded DNA (orthopoxviruses as example), positive-sense single-stranded RNA (alphaviruses and flaviviruses as example), non-segmented negative-sense single-stranded RNA (filoviruses as example), segmented negative-sense single-stranded RNA (arenaviruses and phleboviruses as example) are analyzed. The levels of genetic variability that determine the assignment of compared viruses to taxa of various orders are established for each group of viruses.

Identifying potential monkeypox virus inhibitors: an in silico study targeting the A42R protein.

Monkeypox (now Mpox), a zoonotic disease caused by the monkeypox virus (MPXV) is an emerging threat to global health. In the time span of only six months, from May to October 2022, the number of MPXV cases breached 80,000 and many of the outbreaks occurred in locations that had never previously reported MPXV. Currently there are no FDA-approved MPXV-specific vaccines or treatments, therefore, finding drugs to combat MPXV is of utmost importance. The A42R profilin-like protein of the MPXV is involved in cell development and motility making it a critical drug target. A42R protein is highly conserved across orthopoxviruses, thus A42R inhibitors may work for other family members. This study sought to identify potential A42R inhibitors for MPXV treatment using computational approaches. The energy minimized 3D structure of the A42R profilin-like protein (PDB ID: 4QWO) underwent virtual screening using a library of 36,366 compounds from Traditional Chinese Medicine (TCM), AfroDb, and PubChem databases as well as known inhibitor tecovirimat via AutoDock Vina. A total of seven compounds comprising PubChem CID: 11371962, ZINC000000899909, ZINC000001632866, ZINC000015151344, ZINC000013378519, ZINC000000086470, and ZINC000095486204, predicted to have favorable binding were shortlisted. Molecular docking suggested that all seven proposed compounds have higher binding affinities to A42R (-7.2 to -8.3 kcal/mol) than tecovirimat (-6.7 kcal/mol). This was corroborated by MM/PBSA calculations, with tecovirimat demonstrating the highest binding free energy of -68.694 kJ/mol (lowest binding affinity) compared to the seven shortlisted compounds that ranged from -73.252 to -97.140 kJ/mol. Furthermore, the 7 compounds in complex with A42R demonstrated higher stability than the A42R-tecovirimat complex when subjected to 100 ns molecular dynamics simulations. The protein-ligand interaction maps generated using LigPlot+ suggested that residues Met1, Glu3, Trp4, Ile7, Arg127, Val128, Thr131, and Asn133 are important for binding. These seven compounds were adequately profiled to be potential antivirals via PASS predictions and structural similarity searches. All seven potential lead compounds were scored Pa > Pi for antiviral activity while ZINC000001632866 and ZINC000015151344 were predicted as poxvirus inhibitors with Pa values of 0.315 and 0.215, and Pi values of 0.052 and 0.136, respectively. Further experimental validations of the identified lead compounds are required to corroborate their predicted activity. These seven identified compounds represent solid footing for development of antivirals against MPXV and other orthopoxviruses.

Evaluation of human monkeypox knowledge and beliefs regarding emerging viral infections among healthcare workers.

OBJECTIVES: The purpose of this study was to evaluate possible factors that might be accompanied by high level of human monkey pox (HMPX) knowledge and to explain the relationship between HMPX knowledge and Beliefs regarding emerging viral infections. STUDY DESIGN: A descriptive cross-sectional study was conducted for the implementation of this study. METHODS: Study was conducted at two general hospitals in Mansoura City (Old General Hospital and International Hospital) El Dakahlia Governorate among 620 healthcare workers (HCWs) using a self-managed questionnaire for 1 week (1 to 7 January 2023). The questionnaire has items adapted from the previously published literature to assess HMPX knowledge and Beliefs regarding emerging viral infections. RESULTS: The mean age of the study sample was 27.97 years and most of them were female (86.1%). Physicians and other HCWs (nurses, laboratory technicians, radiographer technicians, and pharmacists) had significantly different levels of knowledge of monkeypox for the majority of the questions. A higher belief was found among two items: viruses are biological weapons manufactured by the superpowers to take global control and the government is misleading the public about the cause of the virus. CONCLUSION: This study discovered lower levels of knowledge of HMPX among HCWs in Egypt. Beliefs about emerging viral infections were widespread, and future research should look into their potential negative impact on health behavior.

Safety and Pharmacokinetics of the Substance of the Anti-Smallpox Drug NIOCH-14 after Oral Administration to Laboratory Animals.

BACKGROUND: Since most of the modern human population has no anti-smallpox immunity, it is extremely important to develop and implement effective drugs for the treatment of smallpox and other orthopoxvirus infections. The objective of this study is to determine the main characteristics of the chemical substance NIOCH-14 and its safety and bioavailability in the body of laboratory animals. METHODS: The safety of NIOCH-14 upon single- or multiple-dose intragastric administration was assessed according to its effect on the main hematological and pathomorphological parameters of laboratory mice and rats. In order to evaluate the pharmacokinetic parameters of NIOCH-14 administered orally, a concentration of ST-246, the active metabolite of NIOCH-14, in mouse blood and organs was determined by tandem mass spectrometry and liquid chromatography. RESULTS: The intragastric administration of NIOCH-14 at a dose of 5 g/kg body weight caused neither death nor signs of intoxication in mice. The intragastric administration of NIOCH-14 to mice and rats at doses of 50 and 150 µg/g body weight either as a single dose or once daily during 30 days did not cause animal death or critical changes in hematological parameters and the microstructure of internal organs. The tissue availability of NIOCH-14 administered orally to the mice at a dose of 50 µg/g body weight, which was calculated according to concentrations of its active metabolite ST-246 for the lungs, liver, kidney, brain, and spleen, was 100, 69.6, 63.3, 26.8 and 20.3%, respectively. The absolute bioavailability of the NIOCH-14 administered orally to mice at a dose of 50 µg/g body weight was 22.8%. CONCLUSION: Along with the previously determined efficacy against orthopoxviruses, including the smallpox virus, the substance NIOCH-14 was shown to be safe and bioavailable in laboratory animal experiments.

Emergence of monkeypox: a worldwide public health crisis.

The human monkeypox virus (MPV), a zoonotic illness that was hitherto solely prevalent in Central and West Africa, has lately been discovered to infect people all over the world and has become a major threat to global health. Humans unintentionally contract this zoonotic orthopoxvirus, which resembles smallpox, when they come into contact with infected animals. Studies show that the illness can also be transferred through frequent proximity, respiratory droplets, and household linens such as towels and bedding. However, MPV infection does not presently have a specified therapy. Smallpox vaccinations provide cross-protection against MPV because of antigenic similarities. Despite scant knowledge of the genesis, epidemiology, and ecology of the illness, the incidence and geographic distribution of monkeypox outbreaks have grown recently. Polymerase chain reaction technique on lesion specimens can be used to detect MPV. Vaccines like ACAM2000, vaccinia immune globulin intravenous (VIG-IV), and JYNNEOS (brand name: Imvamune or Imvanex) as well as FDA-approved antiviral medications such as brincidofovir (brand name: Tembexa), tecovirimat (brand name: TPOXX or ST-246), and cidofovir (brand name: Vistide) are used as therapeutic medications against MPV. In this overview, we provide an outline of the MPV's morphology, evolution, mechanism, transmission, diagnosis, preventative measures, and therapeutic approaches. This study offers the fundamental information required to prevent and manage any further spread of this emerging virus.