Poxvirus fibromas on African hares.

Small dermal tumors were found on three African hares (Lepus capensis) in the Laikipia District, Kenya. Gross and histopathologic studies revealed similarities to the Shope's fibroma of wild rabbits in North America and fibromas of European hares. Histological examination of the African hare fibromas revealed intracytoplasmic inclusion bodies characteristic of poxviruses and poxvirus virions were demonstrated by electron microscopy of ultrathin sections. Attempts to propagate the virus in rabbit skin, embryonated chicken eggs and cell cultures were unsuccessful.     

Poxvirus tropism

Despite the success of the WHO-led smallpox eradication programme a quarter of a century ago, there remains considerable fear that variola virus, or other related pathogenic poxviruses such as monkeypox, could re-emerge and spread disease in the human population. Even today, we are still mostly ignorant about why most poxvirus infections of vertebrate hosts show strict species specificity, or how zoonotic poxvirus infections occur when poxviruses occasionally leap into novel host species. Poxvirus tropism at the cellular level seems to be regulated by intracellular events downstream of virus binding and entry, rather than at the level of specific host receptors as is the case for many other viruses. This review summarizes our current understanding of poxvirus tropism and host range, and discusses the prospects of exploiting host-restricted poxvirus vectors for vaccines, gene therapy or tissue-targeted oncolytic viral therapies for the treatment of human cancers.     

Time scale of Poxvirus evolution

Unlike in vertebrates and RNA viruses, the molecular clock has not been estimated so far for DNA viruses. The extended conserved central region (102 kb) of the orthopoxvirus genome and the DNA polymerase gene (3 kb) were analyzed in viruses representing several genera of the family Poxviridae. Analysis was based on the known dating of the variola virus (VARV) transfer from Western Africa to South America and previous data on the phylogenetic relatedness of modern West African and South American isolates of VARV. The mutation accumulation rate was for the first time estimated for these DNA viruses at (0.9–1.2) × 10^?6 substitutions per site per year. It was assumed that poxviruses diverged from an ancestor approximately 500,000 years ago to form the recent species and that the ancestor of the genus Orthopoxvirus emerged approximately 300,000 years ago and gave origin to the modern species approximately 14,000 years ago.     

Poxvirus Proteomics and Virus-Host Protein Interactions

Summary: Studies of the functional proteins encoded by the poxvirus genome provide information about the composition of the virus as well as individual virus-virus protein and virus-host protein interactions, which provides insight into viral pathogenesis and drug discovery. Widely used proteomic techniques to identify and characterize specific protein-protein interactions include yeast two-hybrid studies and coimmunoprecipitations. Recently, various mass spectrometry techniques have been employed to identify viral protein components of larger complexes. These methods, combined with structural studies, can provide new information about the putative functions of viral proteins as well as insights into virus-host interaction dynamics. For viral proteins of unknown function, identification of either viral or host binding partners provides clues about their putative function. In this review, we discuss poxvirus proteomics, including the use of proteomic methodologies to identify viral components and virus-host protein interactions. High-throughput global protein expression studies using protein chip technology as well as new methods for validating putative protein-protein interactions are also discussed.     

Inhibition of Poxvirus Replication by Streptovaricin

A component of streptovaricin complex inhibits the replication of poxvirus. To be effective, the inhibitor must be introduced early in the replication cycle; it appears to inhibit early messenger ribonucleic acid synthesis by viral cores, thus interfering with all subsequent events. Neither of the two major components of the complex, streptovaricin A or C, was the active component.     

Induction of Poxvirus Ribonucleic Acid Polymerases

Two distinct ribonucleic acid polymerase activities were induced in HeLa cells by poxvirus infection. These activities differ both in their properties and the time of their appearance after infection. One catalyzes the dAT (copolymer of deoxyadenylate and deoxythymidylate)-primed conversion of adenosine triphosphate and uridine triphosphate into an acid-insoluble product. This enzyme is detectable only if deoxyribonucleic acid synthesis has been blocked. In contrast, the accumulation of progeny genomes is a necessary condition for induction of the second enzyme. The latter activity, which is unmasked by detergent treatment, is found exclusively in maturing virus particles. The possibility that both enzymes are involved in transcribing the viral genome is discussed.     

Poxvirus Orthologous Clusters (POCs)

Poxvirus Orthologous Clusters (POCs) is a JAVA client-server application which accesses an updated database containing all complete poxvirus genomes; it automatically groups orthologous genes into families based on BLASTP scores for assessment by a human database curator. POCs has a user-friendly interface permitting complex SQL queries to retrieve interesting groups of DNA and protein sequences as well as gene families for subsequent interrogation by a variety of integrated tools: BLASTP, BLASTX, TBLASTN, Jalview (multiple alignment), Dotlet (Dotplot), Laj (local alignment), and NAP (nucleotide to amino acid alignment).     

Effect of Rifampicin on Poxvirus Protein Synthesis

Poxvirus strains differ with respect to the effect of rifampicin on viral protein synthesis. Rifampicin severely depressed vaccinia-directed protein synthesis but had little effect on the rate of cowpox-directed protein synthesis, including one late virus-induced enzyme. The spectrum of polypeptides synthesized in cowpox-infected cells was similar in the presence or absence of rifampicin except for one significant difference. After removal of rifampicin, viruslike particles assembled to some extent, even when protein synthesis was inhibited by cycloheximide. However, reversal was more extensive if protein synthesis was allowed.     

The Poxvirus A35 Protein Is an Immunoregulator

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.Poxviruses are large, complex viruses with a broad host range and worldwide distribution (30). Members of the family Poxviridae include variola virus, the causative agent of smallpox, which induced a fatality rate of approximately 30% and killed hundreds of millions of people before its eradication in 1980 (28). Currently, the most dangerous extant human-infecting poxvirus is monkeypox virus, which is commonly found in African rodents. Monkeypox virus causes a smallpox-like illness with a 10% fatality rate. A recent study showed that 1.7% of people in the Likouala region in Africa had monkeypox-specific immunoglobulin M (IgM), indicating a significant ongoing infection rate (25). An outbreak of a low-virulence strain of monkeypox virus occurred in the United States in 2003, causing more than 80 human infections and several hospitalizations (6). This outbreak raises concern that monkeypox virus could establish itself in wild-rodent populations in North America (34), thus creating a local zoonotic reservoir for this emerging pathogen. Of further concern are the facts that monkeypox is spreading more efficiently in humans (18, 24, 31) and that the current poxvirus vaccine is not universally protective against monkeypox infection (27). Both variola and monkeypox viruses are considered bioterrorism and biowarfare concerns and are category A select-agent pathogens. There are also other poxvirus infections that sporadically cause human outbreaks, including Cantagalo virus in South America (10, 41) and buffalopox virus in India (23), and the incidence of tanapox virus appears to be increasing (12, 43). Molluscum contagiosum poxvirus accounts for approximately 300,000 doctor visits each year in the United States alone (29). Thus, the study of virulence mechanisms in this group of viruses is important.The eradication of smallpox was accomplished through the use of the related vaccinia virus (VV) as a live-virus vaccine. Despite its phenomenal success, the public vaccination program was discontinued because of the high incidence of complications due to the virulence of wild-type VV. It is estimated that approximately 25% of the population should not receive this vaccine because of immunodeficiency, eczema, pregnancy, or heart disease (14, 21, 45). Safer vaccines are necessary to protect against emerging or released poxviruses. In addition, poxviruses are being used as platform vaccines for other diseases such as human immunodeficiency virus, malaria, and cancer because they induce a robust immune response and accommodate the insertion of large pieces of foreign DNA. It is therefore of great importance to identify poxvirus virulence genes in order to develop safer and more effective poxvirus vaccines. Replication-defective strains, such as Modified Vaccinia Ankara, have been used in an effort to reduce the risks associated with vaccination (8, 47), but the production of these viruses can be challenging, they require higher doses of vaccine, and their protective efficacy against poxvirus infections in humans is unknown. As poxviruses inhibit the activation of antigen-presenting cells and antigen presentation (26, 38), another way to construct a safer vaccine is to develop replication-competent vaccine strains that exclude immunosuppressive genes (3) while retaining protective antigenic epitopes (17, 32). We show herein that the A35R gene is an excellent candidate for removal from vaccine strains.The VV A35 gene is highly conserved in mammalian-tropic poxviruses, and a sequence identity search has revealed that the protein has little similarity to any other poxvirus protein or any nonpoxvirus protein, suggesting that this gene has an important and novel function (39). We have shown that the A35 gene is not required for viral replication in vitro but is required for full virulence in the mouse model (39).We therefore tested the effects of A35R on immune responses during infection in the mouse model and tested its protective efficacy against virulent challenge.     

Polyglutamine pathogenesis.

An increasing number of neurodegenerative disorders have been found to be caused by expanding CAG triplet repeats that code for polyglutamine. Huntington's disease (HD) is the most common of these disorders and dentatorubral-pallidoluysian atrophy (DRPLA) is very similar to HD, but is caused by mutation in a different gene, making them good models to study. In this review, we will concentrate on the roles of protein aggregation, nuclear localization and proteolytic processing in disease pathogenesis. In cell model studies of HD, we have found that truncated N-terminal portions of huntingtin (the HD gene product) with expanded repeats form more aggregates than longer or full length huntingtin polypeptides. These shorter fragments are also more prone to aggregate in the nucleus and cause more cell toxicity. Further experiments with huntingtin constructs harbouring exogenous nuclear import and nuclear export signals have implicated the nucleus in direct cell toxicity. We have made mouse models of HD and DRPLA using an N-terminal truncation of huntingtin (N171) and full-length atrophin-1 (the DRPLA gene product), respectively. In both models, diffuse neuronal nuclear staining and nuclear inclusion bodies are observed in animals expressing the expanded glutamine repeat protein, further implicating the nucleus as a primary site of neuronal dysfunction. Neuritic pathology is also observed in the HD mice. In the DRPLA mouse model, we have found that truncated fragments of atrophin-1 containing the glutamine repeat accumulate in the nucleus, suggesting that proteolysis may be critical for disease progression. Taken together, these data lead towards a model whereby proteolytic processing, nuclear localization and protein aggregation all contribute to pathogenesis.     

红斑性肢痛症相关痘病毒血清免疫学特征的研究

本文从四个方面综合报道了红斑性肢痛症相关痘病毒(ERPV)血清免疫学特征的研究结果。血清学鉴定表明,ERPV为正痘病毒属内成员,但与正痘病毒属内成员痘苗病毒和鼠痘病毒具有中和抗原表位差异。流行性红斑性肢痛症患者血清中含有抗ERPV的中和抗体和抗ERPVA型包涵体IgG抗体,其检出率明显高于当地未发病者和美国人群的检出率。     

Poxvirus genome evolution by gene gain and loss

The poxviruses (Poxviridae) are a family of viruses with double-stranded DNA genomes and substantial numbers (often >200) of genes per genome. We studied the patterns of gene gain and loss over the evolutionary history of 17 poxvirus complete genomes. A phylogeny based on gene family presence/absence showed good agreement with families based on concatenated amino acid sequences of conserved single-copy genes. Gene duplications in poxviruses were often lineage specific, and the most extensively duplicated viral gene families were found in only a few of the genomes analyzed. A total of 34 gene families were found to include a member in at least one of the poxvirus genomes analyzed and at least one animal genome; in 16 (47%) of these families, there was evidence of recent horizontal gene transfer (HGT) from host to virus. Gene families with evidence of HGT included several involved in host immune defense mechanisms (the MHC class I, interleukin-10, interleukin-24, interleukin-18, the interferon gamma receptor, and tumor necrosis factor receptor II) and others (glutaredoxin and glutathione peroxidase) involved in resistance of cells to oxidative stress. Thus "capture" of host genes by HGT has been a recurrent feature of poxvirus evolution and has played an important role in adapting the virus to survive host antiviral defense mechanisms.     

Some Physiochemical Properties of Yaba Poxvirus Deoxyribonucleic Acid

Yaba tumor poxvirus deoxyribonucleic acid (DNA) has a density of 1.6905 in CsCl and its T(m) value in 0.015 m citrate in saline is 82.3. The guanine plus cytosine content estimated from these properties was taken to be 32.5 +/- 0.5%, a value 2 to 3% less than for DNA from other poxviruses reported to date.     

Immunodiffusion Analysis of Yaba Poxvirus Structural and Associated Antigens

Yaba poxvirus virions were extracted and purified from Rhesus monkey tumors. A saline-soluble virion fraction (Y-xp), obtained by mechanical fractionation of purified virions with an X-press, contained seven components in acrylamide gel electrophoresis; five of these components were reactive in immunodiffusion with whole virion and Y-xp antisera produced in rabbits and monkeys. The saline-insoluble residue remaining after X-press treatment was hydrolyzed with sodium dodecyl sulfate, urea, and 2-mercaptoethanol (SUM). This fraction, Y-sum, contained five components, four of which were demonstrable by immunodiffusion. There was no evidence of antigenic relationships between Y-xp and Y-sum antigens in immunodiffusion. In acrylamide gel electrophoresis, one Y-xp and one Y-sum component had similar mobilities. Y-xp but not Y-sum antisera contained viral-neutralizing antibodies. Virus-free saline extracts of Yaba tumor prepared with Genetron (YS) were essentially devoid of virion structural antigens. They failed to induce precipitating antibodies for virion antigens, were nonreactive in immunodiffusion with virion antisera, and gave low complement-fixation titers with virion antisera. Yaba virion antigens were recovered from the Genetron tumor sediment by SUM and alkaline hydrolysis. Antisera prepared to YS extracts gave a maximum of 17 precipitin lines in immunodiffusion with YS extracts; none was identified as a virion structural antigen. Saline extracts of tumor prepared without Genetron contained immunogenic amounts of 5 virion antigens and 12 to 14 associated antigens. Animals immunized with infected cell culture extracts (virus-free) formed antibodies to six to seven virion antigens. The implications of using extracts of Yaba poxvirus-infected tissues in complement-fixation tests to measure virion antibodies were discussed.