The roles of CD4+ T cell help, sex, and dose in the induction of protective CD8+ T cells against a lethal poxvirus by mRNA-LNP vaccines.

The role of CD4+ T cells in the induction of protective CD8+ T cells by mRNA lipid nanoparticle (LNP) vaccines is unknown. We used B6 or Tlr9 -/- mice depleted or not of CD4+ T cells and LNP vaccines loaded with mRNAs encoding the ectromelia virus (ECTV) MHC class I H-2 Kb-restricted immunodominant CD8+ T cell epitope TSYKFESV (TSYKFESV mRNA-LNPs) or the ECTV EVM158 protein, which contains TSYKFESV (EVM-158 mRNA-LNPs). Following prime and boost with 10 µg of either vaccine, Kb-TSYKFESV-specific CD8+ T cells fully protected male and female mice from ECTV at 29 (both mRNA-LNPs) or 90 days (EVM158 mRNA-LNPs) post boost (dpb) independently of CD4+ T cells. However, at 29 dpb with 1 µg mRNA-LNPs, males had lower frequencies of Kb-TSYKFESV-specific CD8+ T cells and were much less well protected than females from ECTV, also independently of CD4+ T cells. At 90 dpb with 1 µg EVM158 mRNA-LNPs, the frequencies of Kb-TSYKFESV-specific CD8+ T cells in males and females were similar, and both were similarly partially protected from ECTV, independently of CD4+ T cells. Therefore, at optimal or suboptimal doses of mRNA-LNP vaccines, CD4+ T cell help is unnecessary to induce protective anti-poxvirus CD8+ T cells specific to a dominant epitope. At suboptimal doses, protection of males requires more time to develop.

What the pox? Review of poxviruses affecting humans.

The re-emergence of human mpox with the multi-country outbreak and a recent report of borealpox (previously Alaskapox) resulting in one death has heightened awareness of the significance of the Poxviridae family and their zoonotic potential. This review examines various poxviruses affecting humans, with discussion of less commonly encountered Poxviridae members, including pathogenesis, epidemiology, and diagnostic methods. Poxvirus treatment is beyond the intended scope of this review and will not be discussed.

Animal and bacterial viruses share conserved mechanisms of immune evasion.

Animal and bacterial cells sense and defend against viral infections using evolutionarily conserved antiviral signaling pathways. Here, we show that viruses overcome host signaling using mechanisms of immune evasion that are directly shared across the eukaryotic and prokaryotic kingdoms of life. Structures of animal poxvirus proteins that inhibit host cGAS-STING signaling demonstrate architectural and catalytic active-site homology shared with bacteriophage Acb1 proteins, which inactivate CBASS anti-phage defense. In bacteria, phage Acb1 proteins are viral enzymes that degrade host cyclic nucleotide immune signals. Structural comparisons of poxvirus protein-2'3'-cGAMP and phage Acb1-3'3'-cGAMP complexes reveal a universal mechanism of host nucleotide immune signal degradation and explain kingdom-specific additions that enable viral adaptation. Chimeric bacteriophages confirm that animal poxvirus proteins are sufficient to evade immune signaling in bacteria. Our findings identify a mechanism of immune evasion conserved between animal and bacterial viruses and define shared rules that explain host-virus interactions across multiple kingdoms of life.

Exploring the transcriptomic profile of human monkeypox virus via CAGE and native RNA sequencing approaches.

In this study, we employed short- and long-read sequencing technologies to delineate the transcriptional architecture of the human monkeypox virus and to identify key regulatory elements that govern its gene expression. Specifically, we conducted a transcriptomic analysis to annotate the transcription start sites (TSSs) and transcription end sites (TESs) of the virus by utilizing Cap Analysis of gene expression sequencing on the Illumina platform and direct RNA sequencing on the Oxford Nanopore technology device. Our investigations uncovered significant complexity in the use of alternative TSSs and TESs in viral genes. In this research, we also detected the promoter elements and poly(A) signals associated with the viral genes. Additionally, we identified novel genes in both the left and right variable regions of the viral genome.IMPORTANCEGenerally, gaining insight into how the transcription of a virus is regulated offers insights into the key mechanisms that control its life cycle. The recent outbreak of the human monkeypox virus has underscored the necessity of understanding the basic biology of its causative agent. Our results are pivotal for constructing a comprehensive transcriptomic atlas of the human monkeypox virus, providing valuable resources for future studies.

High Rates of Miscarriage and Stillbirth among Pregnant Women with Clade I Mpox (Monkeypox) Are Confirmed during 2023-2024 DR Congo Outbreak in South Kivu Province.

Mpox (monkeypox) is a neglected tropical disease that has received increased attention since the multi-nation outbreak that began in 2022. The virus is endemic in West and Central Africa, where the Democratic Republic of the Congo (DRC) is the most affected country. Clade I monkeypox virus (MPXV) infection is endemic in the DRC and has an overall case fatality rate of 10.6% among children and adults. A study conducted in Sankuru Province, DRC, from 2007 to 2011 demonstrated that 75% of pregnant women with mpox had miscarriages or stillbirth. Further analysis of a stillborn fetus showed that MPXV could infect both the placenta and fetus, causing congenital infection. No additional cases of Clade I MPXV in pregnant women were reported until a new outbreak occurred in South Kivu Province during 2023 and 2024. Eight pregnant women having Clade I MPXV infection were identified, of whom four had either miscarriages or stillbirth, representing a 50% fetal mortality rate. These reports confirm previous data from the DRC that indicate the capability of Clade I MPXV to affect the fetus, causing congenital infection and fetal loss in a high percentage of cases. In this article, we review both past and new data from the DRC on the effects of Clade I MPXV during pregnancy and discuss the association of mpox with fetal loss.

Swinepox virus: an unusual outbreak in free-range pig farms in Sicily (Italy).

BACKGROUND: Two outbreaks of swinepox were investigated in free-range domestic pig farms located in the northeastern side of Sicily, Italy. The disease is generally self-limiting with a low mortality rate, but morbidity can reach high rates in case of poor sanitary conditions, improper husbandry practices and ectoparasitic infestation. The presented cases are the first ever reported on the island and part of the few cases reported in domestic pigs. CASE PRESENTATION: Carcasses condemned at the slaughterhouse and deceased pigs from Farm A and Farm B respectively, were referred for post-mortem examination and further investigations, with a strong suspect of SwinePox virus (SWPV) infection. Twelve deceased pigs were examined in total, showing poor body condition and pustular lesions scattered all over the cutaneous surfaces. Moreover, pigs from Farm B showed ocular lesions classified from Grade I to IV (from mild conjunctivitis to severe keratoconjunctivitis with corneal oedema, opacity, and ulcers). Final diagnosis was pursued by the microscopic assessment of skin lesions in both farms, which revealed the typical SWPV-lesion appearance, such as severe and disseminated ulcerative dermatitis and suspected inclusion bodies multifocally observed in the epidermis. Moreover, negative staining Electron Microscopy (nsEM) was performed on skin lesions and ocular swabs from Farm B, revealing in two samples the presence of brick-shaped viral particles, 220 nm long and 160 nm wide, with irregularly arranged surface tubules, identified as SWPV. The gene encoding the 482-bp fragment of the virus late transcription factor-3 was detected by PCR and sequencing revealed 99.79% identity and 100% query-cover with a strain previously isolated in Germany. Field clinical assessment was then performed in Farm B, revealing high overcrowding, poor sanitary conditions and improper husbandry practices, which are relevant risk factors for SWPV transmission. CONCLUSIONS: The present is the first case report of SWPV in free-range pigs raised in Sicily, an island of the Southern coast of Italy, and wants to raise awareness on a neglected disease, and cause of animal health and welfare issues.

Comparative analysis of two novel complete genomes of myxoma virus vaccine strains.

Myxoma virus (MYXV) is a double-stranded DNA-containing virus of the family Poxviridae, genus Leporipoxvirus. MYXV is an important model virus for evolutionary and immunological research and a promising oncolytic. In this study, we sequenced and analyzed two complete genomes of MYXV virus vaccine strains B-82 and Rabbivac-B, which are widely used for vaccine production in Russia. Here, we first show that MYXV vaccine strains B-82 and Rabbivac-B share a common origin with the American recombinant MYXV MAV vaccine strain. In addition, our data suggest that the MYXV B-82 and Rabbivac-B strains contain a number of genes at the 5' and 3' ends that are identical to the virulent MYXV Lausanne strain. Several unique genetic signatures were identified in the M013L, M017L, M023, and M121R genes, helping to achieve high genetic resolution between vaccine strains. Overall, these findings highlight the evolutionary flexibility of certain genes in the MYXV genome and provide insights into the molecular epidemiology of the virus and subsequent vaccine development.

Unique ankyrin repeat proteins in the genome of poxviruses-Boon or Wane, a critical review.

Ankyrin repeat is a 33-amino acid motif commonly observed in eukaryotes and, to a lesser extent, in prokaryotes and archaea and rarely in viruses. This motif plays a crucial role in regulating various cellular processes like the cell cycle, transcription, cell signaling, and inflammatory responses through interactions between proteins. Poxviruses exhibit a distinctive feature of containing multiple ankyrin repeat proteins within their genomes. All the genera of poxviruses possess these proteins except molluscipox virus, crocodylidpox virus, and red squirrel poxvirus. An intriguing characteristic has generated notable interest in studying the functions of these proteins within poxvirus biology. Within poxviruses, ankyrin repeat proteins exhibit a distinct configuration, featuring ankyrin repeats in the N-terminal region and a cellular F-box homolog in the C-terminal region, which enables interactions with the cellular Skp, Cullin, F-box containing ubiquitin ligase complex. Through the examination of experimental evidences and discussions from current literature, this review elucidates the organization and role of ankyrin repeat proteins in poxviruses. Various research studies have highlighted the significant importance of these proteins in poxviral pathogenesis and, acting as factors that enhance virulence. Consequently, they represent viable targets for developing genetically altered viruses with decreased virulence, thus displaying potential as candidates for vaccines and antiviral therapeutic development contributing to safer and more effective strategies against poxviral infections.

Host range of a parasitoid wasp is linked to host susceptibility to its mutualistic viral symbiont.

Parasitoid wasps are one of the most species-rich groups of animals on Earth, due to their ability to successfully develop as parasites of nearly all types of insects. Unlike most known parasitoid wasps that specialize towards one or a few host species, Diachasmimorpha longicaudata is a generalist that can survive within multiple genera of tephritid fruit fly hosts, including many globally important pest species. Diachasmimorpha longicaudata has therefore been widely released to suppress pest populations as part of biological control efforts in tropical and subtropical agricultural ecosystems. In this study, we investigated the role of a mutualistic poxvirus in shaping the host range of D. longicaudata across three genera of agricultural pest species: two of which are permissive hosts for D. longicaudata parasitism and one that is a nonpermissive host. We found that permissive hosts Ceratitis capitata and Bactrocera dorsalis were highly susceptible to manual virus injection, displaying rapid virus replication and abundant fly mortality. However, the nonpermissive host Zeugodacus cucurbitae largely overcame virus infection, exhibiting substantially lower mortality and no virus replication. Investigation of transcriptional dynamics during virus infection demonstrated hindered viral gene expression and limited changes in fly gene expression within the nonpermissive host compared with the permissive species, indicating that the host range of the viral symbiont may influence the host range of D. longicaudata wasps. These findings also reveal that viral symbiont activity may be a major contributor to the success of D. longicaudata as a generalist parasitoid species and a globally successful biological control agent.

Poxvirus A51R Proteins Negatively Regulate Microtubule-Dependent Transport by Kinesin-1.

Microtubule (MT)-dependent transport is a critical means of intracellular movement of cellular cargo by kinesin and dynein motors. MT-dependent transport is tightly regulated by cellular MT-associated proteins (MAPs) that directly bind to MTs and either promote or impede motor protein function. Viruses have been widely shown to usurp MT-dependent transport to facilitate their virion movement to sites of replication and/or for exit from the cell. However, it is unclear if viruses also negatively regulate MT-dependent transport. Using single-molecule motility and cellular transport assays, we show that the vaccinia virus (VV)-encoded MAP, A51R, inhibits kinesin-1-dependent transport along MTs in vitro and in cells. This inhibition is selective as the function of kinesin-3 is largely unaffected by VV A51R. Interestingly, we show that A51R promotes the perinuclear accumulation of cellular cargo transported by kinesin-1 such as lysosomes and mitochondria during infection. Moreover, A51R also regulates the release of specialized VV virions that exit the cell using kinesin-1-dependent movement. Using a fluorescently tagged rigor mutant of kinesin-1, we show that these motors accumulate on A51R-stabilized MTs, suggesting these stabilized MTs may form a "kinesin-1 sink" to regulate MT-dependent transport in the cell. Collectively, our findings uncover a new mechanism by which viruses regulate host cytoskeletal processes.

Current Status of Poultry Recombinant Virus Vector Vaccine Development.

Inactivated and live attenuated vaccines are the mainstays of preventing viral poultry diseases. However, the development of recombinant DNA technology in recent years has enabled the generation of recombinant virus vector vaccines, which have the advantages of preventing multiple diseases simultaneously and simplifying the vaccination schedule. More importantly, some can induce a protective immune response in the presence of maternal antibodies and offer long-term immune protection. These advantages compensate for the shortcomings of traditional vaccines. This review describes the construction and characterization of primarily poultry vaccine vectors, including fowl poxvirus (FPV), fowl adenovirus (FAdV), Newcastle disease virus (NDV), Marek's disease virus (MDV), and herpesvirus of turkey (HVT). In addition, the pathogens targeted and the immunoprotective effect of different poultry recombinant virus vector vaccines are also presented. Finally, this review discusses the challenges in developing vector vaccines and proposes strategies for improving immune efficacy.

Poxvirus Immune Evasion.

Poxviruses have evolved a wide array of mechanisms to evade the immune response, and we provide an overview of the different immunomodulatory strategies. Poxviruses prevent the recognition of viral DNA that triggers the immune responses and inhibit signaling pathways within the infected cell. A unique feature of poxviruses is the production of secreted proteins that mimic cytokines and cytokine receptors, acting as decoy receptors to neutralize the activity of cytokines and chemokines. The capacity of these proteins to evade cellular immune responses by inhibiting cytokine activation is complemented by poxviruses' strategies to block natural killer cells and cytotoxic T cells, often through interfering with antigen presentation pathways. Mechanisms that target complement activation are also encoded by poxviruses. Virus-encoded proteins that target immune molecules and pathways play a major role in immune modulation, and their contribution to viral pathogenesis, facilitating virus replication or preventing immunopathology, is discussed.

Differential Diagnosis, Prevention Measures, and Therapeutic Interventions for Enhanced Monkeypox (Mpox) Care.

Monkeypox (Mpox) is a rare viral disease that presents considerable challenges in healthcare settings, necessitating enhanced nursing care for effective management. This review thoroughly explores key aspects related to improving nursing care for Mpox. It commences by examining the background information on Mpox, encompassing its etiology, epidemiology, and modes of transmission. The differential diagnosis of Mpox is investigated, elucidating its clinical presentation, symptoms, and diagnostic methods to differentiate it from similar conditions. Prevention and control measures at both the public health and healthcare levels are scrutinized, including surveillance and reporting, contact tracing, isolation, and vaccination programs. In healthcare settings, infection prevention and control strategies, such as proper utilization of personal protective equipment, hand hygiene, and environmental management, are discussed. Furthermore, therapeutic interventions for Mpox, including symptomatic management, antiviral therapy, and supportive care, are outlined, with a specific emphasis on pain management, fever control, and psychosocial support. Nursing care strategies encompass patient assessment and monitoring, infection prevention strategies, psychosocial support, and patient education. The challenges encountered in enhancing nursing care for Mpox are acknowledged, along with research gaps and areas for further investigation. Finally, innovations in nursing practice for improved care, such as technology integration and simulation-based training, are explored. Enhancing nursing care in Mpox is crucial for positive patient outcomes, reducing transmission risks, and promoting overall well-being. By addressing the unique challenges, conducting further research, and embracing innovative practices, healthcare professionals, particularly nurses, can provide optimal care and contribute to better management of Mpox cases.

Widespread Horizontal Gene Transfer Among Animal Viruses.

The initial objective of this study was to shed light on the evolution of small DNA tumor viruses by analyzing de novo assemblies of publicly available deep sequencing datasets. The survey generated a searchable database of contig snapshots representing more than 100,000 Sequence Read Archive records. Using modern structure-aware search tools, we iteratively broadened the search to include an increasingly wide range of other virus families. The analysis revealed a surprisingly diverse range of chimeras involving different virus groups. In some instances, genes resembling known DNA-replication modules or known virion protein operons were paired with unrecognizable sequences that structural predictions suggest may represent previously unknown replicases and novel virion architectures. Discrete clades of an emerging group called adintoviruses were discovered in datasets representing humans and other primates. As a proof of concept, we show that the contig database is also useful for discovering RNA viruses and candidate archaeal phages. The ancillary searches revealed additional examples of chimerization between different virus groups. The observations support a gene-centric taxonomic framework that should be useful for future virus-hunting efforts.

Poxviruses from the Concept of One Health.

In the last 4 years, the world has experienced two pandemics of bat-borne viruses. Firstly, in 2019 the SARS-CoV-2 pandemic started and has been causing millions of deaths around the world. In 2022, a Monkeypox pandemic rose in various countries of the world. Those pandemics have witnessed movements and initiatives from healthcare and research institutions to establish a worldwide understanding to battle any future pandemics and biological threats. One Health concept is a modern, comprehensive, unifying ways to improve humans, animals, and ecosystems' health. This concept shows how much they are intertwined and related to one another, whether it is an environmental, or a pathological relation. This review aims to describe Poxviridae and its impact on the One Health concept, by studying the underlying causes of how poxviruses can affect the health of animals, humans, and environments. Reviewing the effect of disease transmission between animal to human, human to human, and animal to animal with pox viruses as a third party to achieve a total understanding of infection and viral transmission. Thus, contributing to enhance detection, diagnosis, research, and treatments regarding the application of One Health.

Assembly and Evolution of Poxviruses.

Poxvirus assembly has been an intriguing area of research for several decades. While advancements in experimental techniques continue to yield fresh insights, many questions are still unresolved. Large genome sizes of up to 380 kbp, asymmetrical structure, an exterior lipid bilayer, and a cytoplasmic life cycle are some notable characteristics of these viruses. Inside the particle are two lateral bodies and a protein wall-bound-biconcave core containing the viral nucleocapsid. The assembly progresses through five major stages-endoplasmic reticulum (ER) membrane alteration and rupture, crescent formation, immature virion formation, genome encapsidation, virion maturation and in a subset of viruses, additional envelopment of the virion prior to its dissemination. Several large dsDNA viruses have been shown to follow a comparable sequence of events. In this chapter, we recapitulate our understanding of the poxvirus morphogenesis process while reviewing the most recent advances in the field. We also briefly discuss how virion assembly aids in our knowledge of the evolutionary links between poxviruses and other Nucleocytoplasmic Large DNA Viruses (NCLDVs).

A Multi-pronged Approach to Addressing Global Poxviruses Vaccine Inequity: A Case of Monkeypox.

Monkeypox has been endemic in Congo and Nigeria for at least five decades. Since early May 2022, there have been numerous unprecedented outbreaks throughout the world in places without any previously reported cases. While a majority of the diagnosed cases have been within Europe and the Americas, several cases have occurred in non-endemic African countries. As of December 2022, 82,999 cases had been reported globally, prompting concern among the World Health Organization (WHO) members. While the WHO has not labeled this epidemic a Global Health Emergency, member states have begun to put forward plans to consolidate their emergency vaccine stockpiles and share the limited number of vaccines made by the single FDA-approved manufacturer, Bavarian Nordic. Many countries are concerned about how vaccines will be shared. Some of the larger donor States are positioned to be the biggest beneficiaries of vaccine sharing, while States from areas that have been suffering from the virus since the 1970s have not been allocated any. This pattern of vaccine distribution echoes that seen during the early part of the COVID-19 pandemic. Due to the similarities between Monkeypox and Smallpox, contact precautions and vaccination seem to be effective strategies to combat its rapid spread. We aim to evaluate how an eradication program model similar to that used for Smallpox can be applied to Monkeypox, and whether it can address vaccine inequity. To do this, we use a multi-pronged approach targeting disease surveillance, vaccine awareness, manufacturing, cost, and distribution strategies.

Poxviruses as Agents of Biological Warfare: The Importance of Ensuring Ethical Standards for Research with Viruses.

Historically, biological agents have been used to target various populations. One of the earliest examples could be the catastrophic effect of smallpox in Australia in the eighteenth century (as alleged by some historians). Modern biological techniques can be used to both create or provide protection against various agents of biological warfare. Any microorganism (viruses, bacteria, and fungi) or its toxins can be used as biological agents. Minnesota Department of Health has listed Smallpox (variola major) as a category A bioterrorism agent, even though it has been eradicated in 1980 through an extensive vaccination campaign. Category A agents are considered the highest risk to public health. Laboratory-associated outbreaks of poxviruses could cause unprecedented occupational hazards. Only two WHO-approved BSL-4 facilities in the United States and Russia are allowed to perform research on the variola virus. So, poxviruses present themselves as a classical case of a dual-use dilemma, since research with them can be used for both beneficial and harmful purposes. Although the importance of ethics in scientific research requires no further elaboration, ethical norms assume greater significance during experimentation with poxviruses. In this chapter, we will update the readers on the sensitive nature of conducting research with poxviruses, and how these viruses can be a source of potential biological weapons. Finally, specified ethical guidelines are explored to ensure safe research practices in virology.

Prediction of potential public health risk of the recent multicountry monkeypox outbreak: An update after the end declaration of global public health emergency.

Background and Aims: A double-stranded DNA virus called monkeypox virus (MPV) belonging to the Poxviridae family and Orthopoxvirus genus causes monkeypox (mpox) infection. This virus used to infect only Central, East, and West Africa. However, it has spread to an extent outside Africa recently. The range of MPV outbreaks was so high that on July 23, 2022, the World Health Organization (WHO) declared it a Public Health Emergency of International Concern (PHEIC). About a year later, the WHO notified the end of a global public health emergency for mpox on May 11, 2023. Here, we aimed to assess the current pathogenicity and potential risk of MPV causing public health emergencies. Methods: We searched information from published articles available in PubMed, Scopus, and ScienceDirect. We used monkeypox, mpox, monkeypox outbreak, and monkeypox virus as keywords during the literature search. Results: Many new variants of MPV have emerged throughout the world that created PHEIC for mpox. Considering the low lethality and transmission rate, mpox is no longer a global public health threat. In addition, the availability of therapeutic and preventive measures helped the healthcare authorities fight the mpox infection in an efficient manner. In this review, we have portrayed the history and evolution of mpox from past to present and an idea of its future outcomes. Also, we have discussed the symptoms related to mpox and approved antiviral treatment strategies to fight off the infection in this piece. This review also emphasized the preventive guidelines set by the WHO for patients, caregivers, and healthcare providers to control the outbreak of mpox infection. Conclusion: We believe this article would give an idea about the potential public health threats of the recent multi-country monkeypox outbreak to the healthcare authorities for taking measures accordingly.

Pharmacokinetic and Environmental Risk Assessment of Prime-2-CoV, a Non-Replicating Orf Virus-Based Vaccine against SARS-CoV-2.

Viral vector vaccines represent a substantial advancement in immunization technology, offering numerous benefits over traditional vaccine modalities. The Orf virus (ORFV) strain D1701-VrV is a particularly promising candidate for vaccine development due to its distinctive attributes, such as a good safety profile, the ability to elicit both humoral and cellular immunity, and its favorable genetic and thermal stability. Despite ORFV's theoretical safety advantages, such as its narrow host range and limited systemic spread post-inoculation, a critical gap persists between these theoretical benefits and the empirical evidence regarding its in vivo safety profile. This discrepancy underscores the need for comprehensive preclinical validations to bridge this knowledge gap, especially considering ORFV's use in humans. Our research introduces Prime-2-CoV, an innovative ORFV-based vaccine candidate against COVID-19, designed to elicit a robust immune response by expressing SARS-CoV-2 Nucleocapsid and Spike proteins. Currently under clinical trials, Prime-2-CoV marks the inaugural application of ORFV in human subjects. Addressing the aforementioned safety concerns, our extensive preclinical evaluation, including an environmental risk assessment (ERA) and detailed pharmacokinetic studies in rats and immunocompromised NOG mice, demonstrates Prime-2-CoV's favorable pharmacokinetic profile, negligible environmental impact, and minimal ERA risks. These findings not only affirm the vaccine's safety and efficacy but also pioneer the use of ORFV-based therapeutics, highlighting its potential for wider therapeutic applications.