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The widespread outbreak of the monkeypox virus (MPXV) recognized in 2022 poses new challenges for public healthcare systems worldwide. With more than 86,000 people infected, there is concern that MPXV may become endemic outside of its original geographical area leading to repeated human spillover infections or continue to be spread person-to-person. Fortunately, classical public health measures (e.g., isolation, contact tracing and quarantine) and vaccination have blunted the spread of the virus, but cases are continuing to be reported in 28 countries in March 2023. We describe here the vaccines and drugs available for the prevention and treatment of MPXV infections. However, although their efficacy against monkeypox (mpox) has been established in animal models, little is known about their efficacy in the current outbreak setting. The continuing opportunity for transmission raises concerns about the potential for evolution of the virus and for expansion beyond the current risk groups. The priorities for action are clear: 1) more data on the efficacy of vaccines and drugs in infected humans must be gathered; 2) global collaborations are necessary to ensure that government authorities work with the private sector in developed and low and middle income countries (LMICs) to provide the availability of treatments and vaccines, especially in historically endemic/enzootic areas; 3) diagnostic and surveillance capacity must be increased to identify areas and populations where the virus is present and may seed resurgence; 4) those at high risk of severe outcomes (e.g., immunocompromised, untreated HIV, pregnant women, and inflammatory skin conditions) must be informed of the risk of infection and be protected from community transmission of MPXV; 5) engagement with the hardest hit communities in a non-stigmatizing way is needed to increase the understanding and acceptance of public health measures; and 6) repositories of monkeypox clinical samples, including blood, fluids, tissues and lesion material must be established for researchers. This MPXV outbreak is a warning that pandemic preparedness plans need additional coordination and resources. We must prepare for continuing transmission, resurgence, and repeated spillovers of MPXV. [ABSTRACT FROM AUTHOR]

The only faculty rank difference that reached statistical significance was at the assistant professor level. The study [[1]] of 1703 academic gastroenterologists shows that there is little difference between the I h i -indices of men and women faculty per rank. Together, these biases compound to unfairly reduce the I h i -index ratings of women; thus, in the study by John [[1]], the women may have outscored the men if the effects of gender bias were removed. [Extracted from the article]

Abstract: We show here that the immunogenicity of the Modified Vaccinia Ankara MVA vaccine strain can be improved by deletion of the A35 gene, without diminishing the ability of the virus to replicate. Deletion of the A35 gene resulted in increased virus-specific immunoglobulin production, class switching to IgG isotypes, and virus-specific IFNγ-secreting splenocytes. The MVA35 deletion virus provided excellent protective efficacy against virulent virus challenge. These results suggest that A35 deletion mutant strains will have superior vaccine performance for poxvirus vaccines as well as platform vaccines for other infectious diseases and cancer. [Copyright &y& Elsevier]

The vaccinia virus A35R gene is highly conserved among poxviruses and encodes a previously uncharacterized hydrophobic acidic protein. Western blotting with anti-A35R peptide antibodies indicated that the protein is expressed early in infection and resolved as a single sharp band of -23 kDa, slightly higher than the 20 kDa predicted from its sequence. The protein band appeared to be the same molecular weight on sodium dodecyl sulfate-polyaerylamide gel electrophoresis, whether expressed in an in vitro transcription/translation system without microsomes or expressed in infected cells, suggesting that it was not glycosylated. A mutant virus with the A35R gene deleted (vA35Δ) formed wild-type-sized plaques on all cell lines tested (human, monkey, mouse, and rabbit); thus, A35R is not required for replication and does not appear to be a host range gene. Although the A35R protein is hydrophobic, it is unlikely to be an integral membrane protein, as it partitioned to the aqueous phase during TX-114 partitioning. The protein could not be detected in virus-infected cell supernatants. A35R localized intracellularly to the virus factories, where the first stages of morphogenesis occur. The vA35Δ mutant formed near-normal levels of the various morphogenie stages of infectious virus particles and supported normal acid-induced fusion of virus-infected cells. Despite normal growth and morphogenesis in vitro, the vA35Δ mutant virus was attenuated in intranasal challenge of mice compared to wild-type and A35R rescue virus. Thus, the intracellular A35R protein plays a role in virulence. The A35R has little homology to any protein outside of poxviruses, suggesting a novel virulence mechanism. [ABSTRACT FROM AUTHOR]

Abstract: Viruses in the family Coronaviridae have elicited new interest, with the outbreaks caused by SARS-HCoV in 2003 and the recent discovery of a new human coronavirus, HCoV-NL63. The genus Torovirus, within the family Coronaviridae, is less well characterized, in part because toroviruses cannot yet be grown in cell culture (except for the Berne virus). In this study, we determined the sequence of the complete genome of Breda-1 (BoTV-1), a bovine torovirus. This is the first complete torovirus genome sequence to be reported. BoTV-1 RNA was amplified using long RT-PCR and the amplicons sequenced. The genome has a length of 28.475kb and consisted mainly of the replicase gene (∼20.2kb) which contains two large overlapping ORFs, ORF1a and ORF1b, encoding polyproteins pp1a and pp1b, respectively. Sequence analysis identified conserved domains within the predicted sequences of pp1a and pp1b. Sequence alignments and protein secondary structure prediction data suggest the presence of a 3C-like serine protease domain with similarity to the arterivirus 3C-like serine protease and a single papain-like cysteine protease domain with similarity to the picornavirus leader protease. The ADRP (APPR-1″) domain – unique to the Coronaviridae – was also located in BoTV pp1a. In addition, several hydrophobic domains were identified that are typical of a nidovirus replicase. Within the pp1b sequence the polymerase and helicase domains were identified, as well as sequences predicted to be involved in ribosomal frameshifting, including the conserved slippery sequence UUUAAAC and two potential pseudoknot structures. [Copyright &y& Elsevier]

Prostaglandins of the E series (PGE) have traditionally been considered as suppressive for immune responses; however, recent data suggest that PGE channels the immune response towards a T helper 2 type response and production of selected immunoglobulin isotypes. Herein, we present data showing that PGE2 and other agents that induce intracellular rises in cAMP significantly increased B lymphocyte IgG1 production (up to sevenfold). PGE2 acted on small resting B cells and on uncommitted B cells expressing high levels of surface IgM to increase the number of cells secreting IgG1. PGE2 even increased IgG1 synthesis by purified B cells in the absence of exogenous IL-4. Finally, PGE2 synergized with IL-4 to induce germline γ1 transcripts through the switch region. This transcription is required for isotype switching. These data support the hypothesis that PGE2 acts on uncommitted resting B cells at the level of germline γ1 transcription to promote class switching to IgG1. PGE2 is an important regulator of the immune response, shifting the balance towards a T helper type 2 response, directing selection of the isotypes produced, and promoting memory cell formation. [Copyright &y& Elsevier]

Phospholipase D (PLD) genes are members of a superfamily that is defined by several highly conserved motifs. PLD in mammals has been proposed to play a role in membrane vesicular trafficking and signal transduction. Using site-directed mutagenesis, 25 point mutants have been made in human PLD1 (hPLD1) and characterized. We find that a motif (HxKxxxxD) and a serine/threonine conserved in all members of the PLD superfamily are critical for PLD biochemical activity, suggesting a possible catalytic mechanism. Functional analysis of catalytically inactive point mutants for yeast PLD demonstrates that the meiotic phenotype ensuing from PLD deficiency in yeast derives from a loss of enzymatic activity. Finally, mutation of an HxKxxxxD motif found in a vaccinia viral protein expressed in the Golgi complex results in loss of efficient vaccinia virus cell-to-cell spreading, implicating the viral protein as a member of the superfamily and suggesting that it encodes a lipid modifying or binding activity. The results suggest that vaccinia virus and hPLD1 may act through analogous mechanisms to effect viral cellular egress and vesicular trafficking, respectively. [ABSTRACT FROM AUTHOR]

Viruses have evolved a myriad of strategies for promoting viral replication, survival and spread. Sequence analysis of the Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV) genome predicts several proteins that are unique to SARS-CoV. The search to understand the high virulence of SARS-CoV compared with related coronaviruses, which cause lesser respiratory illnesses, has recently focused on the unique nsp1 protein of SARS-CoV and suggests evolution of a possible new virulence mechanism in coronaviruses. The SARS-CoV nsp1 protein increases cellular RNA degradation and thus might facilitate SARS-CoV replication or block immune responses. [Copyright &y& Elsevier]

Highlights: [•] Raccoonpoxvirus is much safer than vaccinia virus (WR) in Nude and SCID mice. [•] Raccoonpoxvirus is much safer than vaccinia virus in pregnant mice. [•] Addition of rabies glycoprotein does not increase virulence of raccoonpoxvirus. [•] Raccoonpoxvirus does not persist in mouse organs. [•] Removal of thymidine kinase and hemagglutinin genes attenuates RCNV. [Copyright &y& Elsevier]

Background: With ever increasing numbers of closely related virus genomes being sequenced, it has become desirable to be able to compare two genomes at a level more detailed than gene content because two strains of an organism may share the same set of predicted genes but still differ in their pathogenicity profiles. For example, detailed comparison of multiple isolates of the smallpox virus genome (each approximately 200 kb, with 200 genes) is not feasible without new bioinformatics tools. Results: A software package, Base-By-Base, has been developed that provides visualization tools to enable researchers to 1) rapidly identify and correct alignment errors in large, multiple genome alignments; and 2) generate tabular and graphical output of differences between the genomes at the nucleotide level. Base-By-Base uses detailed annotation information about the aligned genomes and can list each predicted gene with nucleotide differences, display whether variations occur within promoter regions or coding regions and whether these changes result in amino acid substitutions. Base-By-Base can connect to our mySQL database (Virus Orthologous Clusters; VOCs) to retrieve detailed annotation information about the aligned genomes or use information from text files. Conclusion: Base-By-Base enables users to quickly and easily compare large viral genomes; it highlights small differences that may be responsible for important phenotypic differences such as virulence. It is available via the Internet using Java Web Start and runs on Macintosh, PC and Linux operating systems with the Java 1.4 virtual machine. [ABSTRACT FROM AUTHOR]

Pancreatic cancer is the 5th leading cause of cancer deaths, and there are no effective treatments. We developed a poxvirus platform vaccine with improved immunogenicity and inserted the mesothelin gene to create an anti-mesothelin cancer vaccine. Mesothelin expression is mostly restricted to tumors in adult mammals and thus may be a good target for cancer treatment. We show here that the modified vaccinia virus Ankara (MVA) virus expressing mesothelin and the enhanced MVA virus missing the immunosuppressive A35 gene and expressing mesothelin were both safe in mice and were able to induce IFN-gamma secreting T cells in response to mesothelin expressing tumor cells. In addition, the MVA virus has oncolytic properties in vitro as it can replicate in and kill Panc02 pancreatic adenocarcinoma cell line tumor cells, even though it is unable to replicate in most mammalian cells. Deletion of the A35 gene in MVA improved T cell responses as expected. However, we were unable to demonstrate inhibition of Panc02 tumor growth in immunocompetent mice with pre-vaccination of mice, boosts, or even intratumoral injections of the recombinant viruses. Vaccine efficacy may be limited by shedding of mesothelin from tumor cells thus creating a protective screen from the immune system. [ABSTRACT FROM AUTHOR]

Background: Mesothelin has attracted much interest as a tumor specific antigen; it has been reported to promote tumor development and to be a good target for cancer treatment. Most studies to date have used human mesothelin in immunocompromised mice. Since these models do not allow for study of the natural immune response to mesothelin expressing tumors, we have undertaken the characterization of mouse mesothelin so the effects of this protein can be assessed in immunocompetent mouse strains. Methods: We analyzed mouse mesothelin expression, tissue distribution, shedding and biochemistry. In addition we constructed stable mesothelin overexpressing lines of the pancreatic cancer line Panc02 by two methods and tested them for growth and tumorigencity in vitro and in vivo. Results: We show here that mouse mesothelin is similar to human mesothelin in biochemical characteristics, tumor expression and tissue distribution, suggesting the mouse may be a suitable model for study of mesothelin. Stable overexpression of mesothelin in a pancreatic cancer cell line did not increase cell proliferation or anchorageindependent growth in vitro, suggesting that mesothelin is not necessarily a tumor progression factor. Surprisingly overexpression of mesothelin inhibited tumor formation in vivo in immunocompetent mice. Conclusion: The mouse may be a good model for studying mesothelin in the context of an intact immune response. Mesothelin is not necessarily a tumor progression factor, and indeed mesothelin overexpression inhibited tumor growth in immunocompetent mice. [ABSTRACT FROM AUTHOR]

Abstract: The Vaccinia virus gene A35R (Copenhagen designation) is highly conserved in mammalian-tropic poxviruses and is an important virulence factor, but its function was unknown. We show herein that A35 does not affect viral infectivity, apoptosis induction, or replication; however, we found that A35 significantly inhibited MHC class II-restricted antigen presentation, immune priming of T lymphocytes, and subsequent chemokine and cytokine synthesis. A35 localized to endosomes and reduced the amount of a model antigenic peptide displayed in the cleft of class II MHC. In addition, A35 decreased VV specific T cell responses in vivo. Thus, this is the first report identifying a function for the A35 protein in virulence as well as the first report identifying a VV gene that inhibits peptide antigen presentation. [Copyright &y& Elsevier]

Vaccinia virus (VACV) is the current live virus vaccine used to protect humans against smallpox and monkeypox, but its use is contraindicated in several populations because of its virulence. It is therefore important to elucidate the immune evasion mechanisms of VACV. We found that VACV infection of antigen-presenting cells (APCs) significantly decreased major histocompatibility complex (MHC) II antigen presentation and decreased synthesis of 13 chemokines and cytokines, suggesting a potent viral mechanism for immune evasion. In these model systems, responding T cells were not directly affected by virus, indicating that VACV directly affects the APC. VACV significantly decreased nitric oxide production by peritoneal exudate cells and the RAW macrophage cell line in response to lipopolysaccharide (LPS) and interferon (IFN)-γ, decreased class II MHC expression on APCs, and induced apoptosis in macrophages and dendritic cells. However, VACV decreased antigen presentation by 1153 B cells without apparent apoptosis induction, indicating that VACV differentially affects B lymphocytes and other APCs. We show that the key mechanism of VACV inhibition of antigen presentation may be its reduction of antigenic peptide loaded into the cleft of MHC class II molecules. These data indicate that VACV evades the host immune response by impairing critical functions of the APC. [ABSTRACT FROM AUTHOR]

Abstract: Conventional vaccines used for smallpox eradication were often denoted one or another strain of Vaccinia virus (VACV), even though seed virus was sub-cultured multifariously, which rendered the virion population genetically heterogeneous. ACAM2000 cell culture vaccine, recently licensed in the U.S., consists of a biologically vaccine-like VACV homogeneous-sequence clone from the conventional smallpox vaccine Dryvax, which we verified from Dryvax sequence chromatograms is genetically heterogeneous. ACAM2000 VACV and CL3, a mouse-neurovirulent clone from Dryvax, differ by 572 single nucleotide polymorphisms and 53 insertions–deletions of varied size, including a 4.5-kbp deletion in ACAM2000 and a 6.2-kbp deletion in CL3. The sequence diversity between the two clones precludes precisely defining why CL3 is more pathogenic; however, four genes appear significantly dissimilar to account for virulence differences. CL3 encodes intact immunomodulators interferon-α/β and tumor necrosis factor receptors, which are truncated in ACAM2000. CL3 specifies a Cowpox and Variola virus-like ankyrin-repeat protein that might be associated with proteolysis via ubiquitination. And, CL3 shows an elongated thymidylate kinase, similar to the enzyme of the mouse-neurovirulent VACV-WR, a derivative of the New York City Board of Health vaccine, the origin vaccine of Dryvax. Although ACAM2000 encodes most proteins associated with immunization protection, the cloning probably delimited the variant epitopes and other motifs produced by Dryvax due to its VACV genetic heterogeneity. The sequence information for ACAM2000 and CL3 could be significant for resolving the dynamics of their different proteomes and thereby aid development of safer, more effective vaccines. [Copyright &y& Elsevier]

Abstract: Studies indicate that West African and Congo basin isolates of monkeypox virus (MPXV) are genetically distinct. Here, we show Congo basin MPXV-ZAI-V79 is more virulent for cynomolgus monkeys as compared to presumed West African MPXV-COP-58. This finding may explain the lack of case-fatalities in the U.S. 2003 monkeypox outbreak, which was caused by a West African virus. Virulence differences between West African and Congo basin MPXV are further supported by epidemiological analyses that observed a similar prevalence of antibodies in non-vaccinated humans in both regions, while >90% of reported cases occurred in the Congo basin, and no fatal cases were observed outside of this region. To determine the basis for this difference in virulence, we sequenced the genomes of one human West African isolate, and two presumed West African isolates and compared the sequences to Congo basin MPXV-ZAI-96-I-16. The analysis identified D10L, D14L, B10R, B14R, and B19R as possible virulence genes, with D14L (ortholog of vaccinia complement protein) as a leading candidate. [Copyright &y& Elsevier]

Severe acute respiratory syndrome (SARS) emerged from China as an untreatable and rapidly spreading respiratory illness of unknown etiology. Following point source exposure in February 2003, more than a dozen guests infected at a Hog Kong hotel seeded multi-country outbreaks that persisted through the spring of 2003. The World Health Organization responded by invoking traditional public health mea- suers and advanced technologies to control the illness and contain the cause. A novel coronavirus was implicated and its entire genome was sequenced by mid-April 2003. The urgency of responding to this threat focused scientific endeavor and stimulated global collaboration. Through real-time application of accumulating knowledge, the world proved capable of arresting the first pandemic threat of the twenty-first century, despite early respiratory-borne spread and global susceptibility. This review synthesizes lessons learned from this remarkable achievement. These lessons can be applied to re-emergence of SARS or to the next pandemic threat to arise. [ABSTRACT FROM AUTHOR]

Identifies the conserved families of poxvirus orthologous clusters. Alleviation of the need for using multiple Internet sites for analysis; Number of gene families conserved in chordopoxviruses and vertebrate-infecting chordopoxviruses identified; Identification of annotation and sequencing updates in poxvirus genomes.

The vaccinia virus (VACV) A27 protein and its homologs, which are found in a large number of members of the genus Orthopoxvirus (OPXV), are targets of viral neutralization by host antibodies. We have mapped six binding sites (epitopes #1A: aa 32–39, #1B: aa 28–33, #1C: aa 26–31, #1D: 28–34, #4: aa 9–14, and #5: aa 68–71) of A27 specific monoclonal antibodies (mAbs) using peptide arrays. MAbs recognizing epitopes #1A–D and #4 neutralized VACV Elstree in a complement dependent way (50% plaque-reduction: 12.5–200 µg/mL). Fusion of VACV at low pH was blocked through inhibition of epitope #1A. To determine the sequence variability of the six antigenic sites, 391 sequences of A27 protein homologs available were compared. Epitopes #4 and #5 were conserved among most of the OPXVs, while the sequential epitope complex #1A–D was more variable and, therefore, responsible for species-specific epitope characteristics. The accurate and reliable mapping of defined epitopes on immuno-protective proteins such as the A27 of VACV enables phylogenetic studies and insights into OPXV evolution as well as to pave the way to the development of safer vaccines and chemical or biological antivirals. [ABSTRACT FROM AUTHOR]

It was shown previously that the highly conserved vaccinia virus A35 gene is an important virulence factor in respiratory infection of mice. We show here that A35 is also required for full virulence by the intraperitoneal route of infection. A virus mutant in which the A35 gene has been removed replicated normally and elicited improved antibody, gamma interferon-secreting cell, and cytotoxic T-lymphocyte responses compared to wild-type virus, suggesting that A35 increases poxvirus virulence by immunomodulation. The enhanced immune response correlated with an improved control of viral titers in target organs after the development of the specific immune response. Finally, the A35 deletion mutant virus also provided protection from lethal challenge (1,000 50% lethal doses) equal to that of the wild-type virus. Together, these data suggest that A35 deletion viruses will make safer and more efficacious vaccines for poxviruses. In addition, the A35 deletion viruses will serve as improved platform vectors for other infectious diseases and cancer and will be superior vaccine choices for postexposure poxvirus vaccination, as they also provide improved kinetics of the immune response. [ABSTRACT FROM AUTHOR]