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1

Vaccinia Virus Vaccines: Past, Present and Future  

PubMed Central

Vaccinia virus (VACV) has been used more extensively for human immunization than any other vaccine. For almost two centuries, VACV was employed to provide cross-protection against variola virus, the causative agent of smallpox, until the disease was eradicated in the late 1970s. Since that time, continued research on VACV has produced a number of modified vaccines with improved safety profiles. Attenuation has been achieved through several strategies, including sequential passage in an alternative host, deletion of specific genes or genetic engineering of viral genes encoding immunomodulatory proteins. Some highly attenuated third- and fourth-generation VACV vaccines are now being considered for stockpiling against a possible re-introduction of smallpox through bioterrorism. Researchers have also taken advantage of the ability of the VACV genome to accommodate additional genetic material to produce novel vaccines against a wide variety of infectious agents, including a recombinant VACV encoding the rabies virus glycoprotein that is administered orally to wild animals. This review provides an in-depth examination of these successive generations of VACV vaccines, focusing on how the understanding of poxviral replication and viral gene function permits the deliberate modification of VACV immunogenicity and virulence.

Jacobs, Bertram L.; Langland, Jeffrey O.; Kibler, Karen V.; Denzler, Karen L.; White, Stacy D.; Holechek, Susan A.; Wong, Shukmei; Huynh, Trung; Baskin, Carole R.

2009-01-01

2

Vaccinia Virus: A Tool for Research and Vaccine Development  

NASA Astrophysics Data System (ADS)

Vaccinia virus is no longer needed for smallpox immunization, but now serves as a useful vector for expressing genes within the cytoplasm of eukaryotic cells. As a research tool, recombinant vaccinia viruses are used to synthesize biologically active proteins and analyze structure-function relations, determine the targets of humoral- and cell-mediated immunity, and investigate the immune responses needed for protection against specific infectious diseases. When more data on safety and efficacy are available, recombinant vaccinia and related poxviruses may be candidates for live vaccines and for cancer immunotherapy.

Moss, Bernard

1991-06-01

3

Clinical development of Modified Vaccinia virus Ankara vaccines.  

PubMed

The smallpox vaccine Vaccinia was successfully used to eradicate smallpox, but although very effective, it was a very reactogenic vaccine and responsible for the deaths of one or two people per million vaccinated. Modified Vaccinia virus Ankara (MVA) is a replication-deficient and attenuated derivative, also used in the smallpox eradication campaign and now being developed as a recombinant viral vector to produce vaccines against infectious diseases and cancer. Many clinical trials of these new vaccines have been conducted, and the findings of these trials are reviewed here. The safety of MVA is now well documented, immunogenicity is influenced by the dose and vaccination regimen, and information on the efficacy of MVA-vectored vaccines is now beginning to accumulate. PMID:23523410

Gilbert, Sarah C

2013-09-01

4

Clonal vaccinia virus grown in cell culture as a new smallpox vaccine  

Microsoft Academic Search

Although the smallpox virus was eradicated over 20 years ago, its potential release through bioterrorism has generated renewed interest in vaccination. To develop a modern smallpox vaccine, we have adapted vaccinia virus that was derived from the existing Dryvax vaccine for growth in a human diploid cell line. We characterized six cloned and one uncloned vaccine candidates. One clone, designated

Jian Liu; Konstantin V Pugachev; Gwendolyn A Myers; Brie Coughlin; Paul S Blum; Richard Nichols; Casey Johnson; John Cruz; Jeffrey S Kennedy; Francis A Ennis; Richard Weltzin; Thomas P Monath

2003-01-01

5

Chemotherapy enhances CD8(+) T cell-mediated antitumor immunity induced by vaccination with vaccinia virus.  

PubMed

The use of immunotherapy or chemotherapy alone is generally ineffective against well-established tumors. To overcome this intrinsic resistance against therapy for tumors, we have attempted to combine immunotherapy with chemotherapy. In this study, we tried to induce a rapid antitumor effect via chemoimmunotherapy using a vaccinia viral vaccine as an immunotherapeutic agent with anticancer agents including epigallocatechin-3-gallate (EGCG) and conventional anticancer drugs. Although a combination of vaccinia-mediated vaccination and chemotherapy led to a strong inhibition of tumor growth, monotherapy alone failed to completely cure tumors. In contrast, intravenous injection of cisplatin (CDDP) or cyclophosphamide (CTX) after vaccinia virus vaccination led to complete regression of the established tumors. Interestingly, anticancer drugs appear to augment the antitumor effect of the vaccinia virus-mediated immunotherapy. This effect is mainly associated with the enhanced tumor-specific CD8(+) T cell immune response induced by vaccinia virus, which was demonstrated by antibody depletion. However, anticancer drugs alone failed to induce a significant enhancement of the tumor-specific CD8(+) T cell immune response. Taken together, these results suggest that combining vaccinia virus-based immunotherapy with anticancer drugs is particularly effective against established tumors by increasing the tumor antigen-specific CD8(+) T cell immune response, which is primed by vaccinia virus-mediated vaccination. PMID:17551502

Song, Chung Kil; Han, Hee Dong; Noh, Kyung Hee; Kang, Tae Heung; Park, Yong Sung; Kim, Jin Hee; Park, Eun Suk; Shin, Byung Cheol; Kim, Tae Woo

2007-08-01

6

Human CD4+ T Cell Epitopes from Vaccinia Virus Induced by Vaccination or Infection  

PubMed Central

Despite the importance of vaccinia virus in basic and applied immunology, our knowledge of the human immune response directed against this virus is very limited. CD4+ T cell responses are an important component of immunity induced by current vaccinia-based vaccines, and likely will be required for new subunit vaccine approaches, but to date vaccinia-specific CD4+ T cell responses have been poorly characterized, and CD4+ T cell epitopes have been reported only recently. Classical approaches used to identify T cell epitopes are not practical for large genomes like vaccinia. We developed and validated a highly efficient computational approach that combines prediction of class II MHC-peptide binding activity with prediction of antigen processing and presentation. Using this approach and screening only 36 peptides, we identified 25 epitopes recognized by T cells from vaccinia-immune individuals. Although the predictions were made for HLA-DR1, eight of the peptides were recognized by donors of multiple haplotypes. T cell responses were observed in samples of peripheral blood obtained many years after primary vaccination, and were amplified after booster immunization. Peptides recognized by multiple donors are highly conserved across the poxvirus family, including variola, the causative agent of smallpox, and may be useful in development of a new generation of smallpox vaccines and in the analysis of the immune response elicited to vaccinia virus. Moreover, the epitope identification approach developed here should find application to other large-genome pathogens.

Calvo-Calle, J. Mauricio; Strug, Iwona; Nastke, Maria-Dorothea; Baker, Stephen P; Stern, Lawrence J

2007-01-01

7

Vaccinia viruses: vaccines against smallpox and vectors against infectious diseases and tumors  

PubMed Central

Less than 200 years after its introduction, widespread use of vaccinia virus (VACV) as a smallpox vaccine has eradicated variola virus. Along with the remarkable success of the vaccination program, frequent and sometimes severe adverse reactions to VACV were encountered. After eradication, VACV has been reserved for select populations who might be at significant risk for orthopoxvirus infections. Events over the past decade have renewed concerns over the potential use of variola virus as a biological weapon. Accordingly, interest in VACV and attenuated derivatives has increased, both as vaccines against smallpox and as vectors for other vaccines. This article will focus on new developments in the field of orthopoxvirus immunization and will highlight recent advances in the use of vaccinia viruses as vectors for infectious diseases and malignancies.

Walsh, Stephen R; Dolin, Raphael

2011-01-01

8

Protective efficacy of vaccination by recombinant vaccinia virus against Neospora caninum infection  

Microsoft Academic Search

The recombinant vaccinia viruses expressing the surface protein of Neospora caninum tachyzoite, NcSAG1 or NcSRS2, were constructed. The vaccination with these recombinant viruses could protect effectively the parasite invasion in a mouse model system. The vaccine efficacy of NcSRS2 was higher than that of NcSAG1. The present study indicated that a high level of IgG1 Ab production to parasite is

Yoshifumi Nishikawa; Noboru Inoue; Xuenan Xuan; Hideyuki Nagasawa; Ikuo Igarashi; Kozo Fujisaki; Haruki Otsuka; Takeshi Mikami

2001-01-01

9

Recombinant Vaccinia Virus: Immunization against Multiple Pathogens  

NASA Astrophysics Data System (ADS)

The coding sequences for the hepatitis B virus surface antigen, the herpes simplex virus glycoprotein D, and the influenza virus hemagglutinin were inserted into a single vaccinia virus genome. Rabbits inoculated intravenously or intradermally with this polyvalent vaccinia virus recombinant produced antibodies reactive to all three authentic foreign antigens. In addition, the feasibility of multiple rounds of vaccination with recombinant vaccinia virus was demonstrated.

Perkus, Marion E.; Piccini, Antonia; Lipinskas, Bernard R.; Paoletti, Enzo

1985-09-01

10

Antibody Profiling by Proteome Microarray Reveals the Immunogenicity of the Attenuated Smallpox Vaccine Modified Vaccinia Virus Ankara Is Comparable to That of Dryvax  

Microsoft Academic Search

Modified vaccinia virus Ankara (MVA) is a highly attenuated vaccinia virus that is under consideration as an alternative to the conventional smallpox vaccine Dryvax. MVA was attenuated by extensive passage of vaccinia virus Ankara in chicken embryo fibroblasts. Several immunomodulatory genes and genes that influ- ence host range are deleted or mutated, and replication is aborted in the late stage

D. Huw Davies; Linda S. Wyatt; Frances K. Newman; Patricia L. Earl; Sookhee Chun; Jenny E. Hernandez; Douglas M. Molina; Siddiqua Hirst; Bernard Moss; Sharon E. Frey; Philip L. Felgner

2008-01-01

11

Vaccination with recombinant modified vaccinia virus Ankara protects against measles virus infection in the mouse and cotton rat model  

Microsoft Academic Search

Modified vaccinia virus Ankara (MVA) has been used as an experimental vaccine vector against respiratory infections. We have tested the safety and immunogenicity of a recombinant virus expressing the hemagglutinin of measles virus (MVA-MV-H) using the mouse model of measles virus induced encephalitis and the cotton rat model for respiratory infection. MVA-MV-H proved to induce a TH1 response, neutralizing antibodies

Gerald Weidinger; Marion Ohlmann; Bernd Schlereth; Gerd Sutter; Stefan Niewiesk

2001-01-01

12

Host range, growth property, and virulence of the smallpox vaccine: Vaccinia virus Tian Tan strain  

SciTech Connect

Vaccinia Tian Tan (VTT) was used as a vaccine against smallpox in China for millions of people before 1980, yet the biological characteristics of the virus remain unclear. We have characterized VTT with respect to its host cell range, growth properties in vitro, and virulence in vivo. We found that 11 of the 12 mammalian cell lines studied are permissive to VTT infection whereas one, CHO-K1, is non-permissive. Using electron microscopy and sequence analysis, we found that the restriction of VTT replication in CHO-K1 is at a step before viral maturation probably due to the loss of the V025 gene. Moreover, VTT is significantly less virulent than vaccinia WR but remains neurovirulent in mice and causes significant body weight loss after intranasal inoculation. Our data demonstrate the need for further attenuation of VTT to serve either as a safer smallpox vaccine or as a live vaccine vector for other pathogens.

Fang Qing [Modern Virology Research Center and AIDS Center, National Key Laboratory of Virology, College of Life Sciences, Wuhan University, Hubei 430072 (China); Yang Lin [Modern Virology Research Center and AIDS Center, National Key Laboratory of Virology, College of Life Sciences, Wuhan University, Hubei 430072 (China); Zhu Weijun [Modern Virology Research Center and AIDS Center, National Key Laboratory of Virology, College of Life Sciences, Wuhan University, Hubei 430072 (China); Liu Li [Modern Virology Research Center and AIDS Center, National Key Laboratory of Virology, College of Life Sciences, Wuhan University, Hubei 430072 (China); Wang Haibo [Modern Virology Research Center and AIDS Center, National Key Laboratory of Virology, College of Life Sciences, Wuhan University, Hubei 430072 (China); Yu Wenbo [Modern Virology Research Center and AIDS Center, National Key Laboratory of Virology, College of Life Sciences, Wuhan University, Hubei 430072 (China); Xiao Genfu [Modern Virology Research Center and AIDS Center, National Key Laboratory of Virology, College of Life Sciences, Wuhan University, Hubei 430072 (China); Tien Po [Modern Virology Research Center and AIDS Center, National Key Laboratory of Virology, College of Life Sciences, Wuhan University, Hubei 430072 (China); Zhang Linqi [Modern Virology Research Center and AIDS Center, National Key Laboratory of Virology, College of Life Sciences, Wuhan University, Hubei 430072 (China); Aaron Diamond AIDS Research Center, Rockefeller University, New York, NY 10016 (United States); AIDS Research Center, Chinese Academy of Medical Sciences, Peking Union Medical College, Beijing (China); Chen Zhiwei [Modern Virology Research Center and AIDS Center, National Key Laboratory of Virology, College of Life Sciences, Wuhan University, Hubei 430072 (China) and Aaron Diamond AIDS Research Center, Rockefeller University, New York, NY 10016 (United States)]. E-mail: zchen@adarc.org

2005-05-10

13

Reemergence of Vaccinia Virus during Zoonotic Outbreak, Par? State, Brazil  

PubMed Central

In 2010, vaccinia virus caused an outbreak of bovine vaccinia that affected dairy cattle and rural workers in Pará State, Brazil. Genetic analyses identified the virus as distinct from BeAn58058 vaccinia virus (identified in 1960s) and from smallpox vaccine virus strains. These findings suggest spread of autochthonous group 1 vaccinia virus in this region.

de Assis, Felipe L.; Vinhote, Wagner M.; Barbosa, Jose D.; de Oliveira, Cairo H.S.; de Oliveira, Carlos M.G.; Campos, Karinny F.; Silva, Natalia S.; Trindade, Giliane de Souza

2013-01-01

14

Reemergence of vaccinia virus during Zoonotic outbreak, Pará State, Brazil.  

PubMed

In 2010, vaccinia virus caused an outbreak of bovine vaccinia that affected dairy cattle and rural workers in Pará State, Brazil. Genetic analyses identified the virus as distinct from BeAn58058 vaccinia virus (identified in 1960s) and from smallpox vaccine virus strains. These findings suggest spread of autochthonous group 1 vaccinia virus in this region. PMID:24274374

de Assis, Felipe L; Vinhote, Wagner M; Barbosa, José D; de Oliveira, Cairo H S; de Oliveira, Carlos M G; Campos, Karinny F; Silva, Natália S; Trindade, Giliane de Souza

2013-12-01

15

Recombinant Vaccinia Virus: Immunization against Multiple Pathogens  

Microsoft Academic Search

The coding sequences for the hepatitis B virus surface antigen, the herpes simplex virus glycoprotein D, and the influenza virus hemagglutinin were inserted into a single vaccinia virus genome. Rabbits inoculated intravenously or intradermally with this polyvalent vaccinia virus recombinant produced antibodies reactive to all three authentic foreign antigens. In addition, the feasibility of multiple rounds of vaccination with recombinant

Marion E. Perkus; Antonia Piccini; Bernard R. Lipinskas; Enzo Paoletti

1985-01-01

16

Oral vaccination with vaccinia virus expressing the tick antigen subolesin inhibits tick feeding and transmission of Borrelia burgdorferi.  

PubMed

Immunization with the Ixodes scapularis protein, subolesin, has previously been shown to protect hosts against tick infestation and to decrease acquisition of Anaplsma marginale and Babesia bigemina. Here we report the efficacy of subolesin, a conserved tick protein that can act as a regulator of gene expression, expressed from vaccinia virus for use as an orally delivered reservoir - targeted vaccine for prevention of tick infestation and acquisition/transmission of Borrelia burgdorferi to its tick and mouse hosts. We cloned subolesin into vaccinia virus and showed that it is expressed from mammalian cells infected with the recombinant virus in vitro. We then vaccinated mice by oral gavage. A single dose of the vaccine was sufficient for mice to generate antibody response to subolesin. Vaccination with the subolesin expressing vaccinia virus inhibited tick infestation by 52% compared to control vaccination with vaccinia virus and reduced uptake of B. burgdorferi among the surviving ticks that fed to repletion by 34%. There was a reduction in transmission of B. burgdorferi to uninfected vaccinated mice of 40% compared to controls. These results suggest that subolesin has potential as a component of a reservoir targeted vaccine to decrease B. burgdorferi, Babesia and Anaplasma species infections in their natural hosts. PMID:22864146

Bensaci, Mekki; Bhattacharya, Debaditya; Clark, Roger; Hu, Linden T

2012-09-14

17

Identification of vaccinia virus epitope-specific HLA-A*0201-restricted T cells and comparative analysis of smallpox vaccines  

PubMed Central

Despite worldwide eradication of naturally occurring variola virus, smallpox remains a potential threat to both civilian and military populations. New, safe smallpox vaccines are being developed, and there is an urgent need for methods to evaluate vaccine efficacy after immunization. Here we report the identification of an immunodominant HLA-A*0201-restricted epitope that is recognized by cytotoxic CD8+ T cells and conserved among Orthopoxvirus species including variola virus. This finding has permitted analysis and monitoring of epitope-specific T cell responses after immunization and demonstration of the identified T cell specificity in an A*0201-positive human donor. Vaccination of transgenic mice allowed us to compare the immunogenicity of several vaccinia viruses including highly attenuated, replication-deficient modified vaccinia virus Ankara (MVA). MVA vaccines elicited levels of CD8+ T cell responses that were comparable to those induced by the replication-competent vaccinia virus strains. Finally, we demonstrate that MVA vaccination is fully protective against a lethal respiratory challenge with virulent vaccinia virus strain Western Reserve. Our data provide a basis to rationally estimate immunogenicity of safe, second-generation poxvirus vaccines and suggest that MVA may be a suitable candidate.

Drexler, Ingo; Staib, Caroline; Kastenmuller, Wolfgang; Stevanovic, Stefan; Schmidt, Burkhard; Lemonnier, Francois A.; Rammensee, Hans-Georg; Busch, Dirk H.; Bernhard, Helga; Erfle, Volker; Sutter, Gerd

2003-01-01

18

Failure of the Smallpox Vaccine To Develop a Skin Lesion in Vaccinia Virus-Na?ve Individuals Is Related to Differences in Antibody Profiles before Vaccination, Not After  

PubMed Central

Successful vaccination against smallpox with conventional vaccinia virus is usually determined by the development of a vesicular skin lesion at the site of vaccinia inoculation, called a “take.” Although previous vaccination is known to be associated with attenuation of the take, the immunology that underlies a no-take in vaccinia-naïve individuals is not well understood. We hypothesized that antibody profiling of individuals before and after receiving vaccinia virus would reveal differences between takes and no-takes that may help better explain the phenomenon. Using vaccinia virus proteome microarrays and recombinant protein enzyme-linked immunosorbent assays (ELISAs), we first examined the antibody response in vaccinia-naïve individuals that failed to take after receiving different doses of the replication-competent DryVax and Aventis Pasteur (APSV) smallpox vaccines. Most that received diluted vaccine failed to respond, although four no-takes receiving diluted vaccine and four receiving undiluted vaccine mounted an antibody response. Interestingly, their antibody profiles were not significantly different from those of controls that did show a take. However, we did find elevated antibody titers in no-takes prior to receiving DryVax that were significantly different from those of takes. Although the sample size studied was small, we conclude the failure to take in responders correlates with preexisting immunity of unknown etiology that may attenuate the skin reaction in a way similar to previous smallpox vaccination.

Tan, Xiaolin; Chun, Sookhee; Pablo, Jozelyn; Felgner, Philip; Liang, Xiaowu

2012-01-01

19

Active vaccination with vaccinia virus A33 protects mice against lethal vaccinia and ectromelia viruses but not against cowpoxvirus; elucidation of the specific adaptive immune response.  

PubMed

Vaccinia virus protein A33 (A33VACV) plays an important role in protection against orthopoxviruses, and hence is included in experimental multi-subunit smallpox vaccines. In this study we show that single-dose vaccination with recombinant Sindbis virus expressing A33VACV, is sufficient to protect mice against lethal challenge with vaccinia virus WR (VACV-WR) and ectromelia virus (ECTV) but not against cowpox virus (CPXV), a closely related orthopoxvirus. Moreover, a subunit vaccine based on the cowpox virus A33 ortholog (A33CPXV) failed to protect against cowpox and only partially protected mice against VACV-WR challenge. We mapped regions of sequence variation between A33VACV and A33CPXVand analyzed the role of such variations in protection. We identified a single protective region located between residues 104-120 that harbors a putative H-2Kd T cell epitope as well as a B cell epitope - a target for the neutralizing antibody MAb-1G10 that blocks spreading of extracellular virions. Both epitopes in A33CPXV are mutated and predicted to be non-functional. Whereas vaccination with A33VACV did not induce in-vivo CTL activity to the predicted epitope, inhibition of virus spread in-vitro, and protection from lethal VACV challenge pointed to the B cell epitope highlighting the critical role of residue L118 and of adjacent compensatory residues in protection. This epitope's critical role in protection, as well as its modifications within the orthopoxvirus genus should be taken in context with the failure of A33 to protect against CPXV as demonstrated here. These findings should be considered when developing new subunit vaccines and monoclonal antibody based therapeutics against orthopoxviruses, especially variola virus, the etiologic agent of smallpox. PMID:23842430

Paran, Nir; Lustig, Shlomo; Zvi, Anat; Erez, Noam; Israely, Tomer; Melamed, Sharon; Politi, Boaz; Ben-Nathan, David; Schneider, Paula; Lachmi, Batel; Israeli, Ofir; Stein, Dana; Levin, Reuven; Olshevsky, Udy

2013-01-01

20

Active vaccination with vaccinia virus A33 protects mice against lethal vaccinia and ectromelia viruses but not against cowpoxvirus; elucidation of the specific adaptive immune response  

PubMed Central

Vaccinia virus protein A33 (A33VACV) plays an important role in protection against orthopoxviruses, and hence is included in experimental multi-subunit smallpox vaccines. In this study we show that single-dose vaccination with recombinant Sindbis virus expressing A33VACV, is sufficient to protect mice against lethal challenge with vaccinia virus WR (VACV-WR) and ectromelia virus (ECTV) but not against cowpox virus (CPXV), a closely related orthopoxvirus. Moreover, a subunit vaccine based on the cowpox virus A33 ortholog (A33CPXV) failed to protect against cowpox and only partially protected mice against VACV-WR challenge. We mapped regions of sequence variation between A33VACV and A33CPXVand analyzed the role of such variations in protection. We identified a single protective region located between residues 104–120 that harbors a putative H-2Kd T cell epitope as well as a B cell epitope - a target for the neutralizing antibody MAb-1G10 that blocks spreading of extracellular virions. Both epitopes in A33CPXV are mutated and predicted to be non-functional. Whereas vaccination with A33VACV did not induce in-vivo CTL activity to the predicted epitope, inhibition of virus spread in-vitro, and protection from lethal VACV challenge pointed to the B cell epitope highlighting the critical role of residue L118 and of adjacent compensatory residues in protection. This epitope’s critical role in protection, as well as its modifications within the orthopoxvirus genus should be taken in context with the failure of A33 to protect against CPXV as demonstrated here. These findings should be considered when developing new subunit vaccines and monoclonal antibody based therapeutics against orthopoxviruses, especially variola virus, the etiologic agent of smallpox.

2013-01-01

21

Modified vaccinia virus Ankara as a vaccine against feline coronavirus: immunogenicity and efficacy.  

PubMed

Feline infectious peritonitis virus (FIPV) is a coronavirus that induces a fatal systemic disease mediated by an inappropriate immune response. Most previous vaccination attempts against FIPV were unsuccessful because IgG antibodies against the surface protein enhance the infection. However, two studies have shown that poxvirus vectors (vaccinia WR and canarypox) expressing only the FIPV membrane (M) protein can elicit a partially protective immunity which is supposed to be cell-mediated (Virology 181 (1991) 327; International patent WO 97/20054 (1997)). In our study, we report the construction of another poxvirus, the modified vaccinia virus Ankara (MVA), as an expression vector for the FIPV M protein. In this vector, the M gene has been inserted downstream a strong early/late promoter, whereas the two previously described poxviruses expressed the M protein during their early stage only. The immunogenicity of the recombinant MVA-M was evaluated in the murine model which revealed an effect of the vector on the Th1/Th2 balance. The vaccine was then tested in cats to evaluate its efficacy in an FIPV 79-1146 challenge. Vaccinated kittens developed FIPV-specific antibodies after immunization, however, none of them was protected against FIPV. Our results suggest a crucial role for the type of poxviral promoter that must be used to induce an effective immune response against FIPV. PMID:15123156

Hebben, Matthias; Duquesne, Véronique; Cronier, Joëlle; Rossi, Bernard; Aubert, André

2004-04-01

22

Biosafety aspects of modified vaccinia virus Ankara (MVA)-based vectors used for gene therapy or vaccination.  

PubMed

The modified vaccinia virus Ankara (MVA) strain is a highly attenuated strain of vaccinia virus that has been demonstrated to be safe for humans. MVA is widely considered as the vaccinia virus strain of choice for clinical investigation because of its high safety profile. It also represents an excellent candidate for use as vector system in recombinant vaccine development for gene delivery or vaccination against infectious diseases or tumours, even in immunocompromised individuals. The use of MVA and recombinant MVA vectors must comply with various regulatory requirements, particularly relating to the assessment of potential risks for human health and the environment. The purpose of the present paper is to highlight some biological characteristics of MVA and MVA-based recombinant vectors and to discuss these from a biosafety point of view in the context of the European regulatory framework for genetically modified organisms with emphasis on the assessment of potential risks associated with environmental release. PMID:22342706

Verheust, Céline; Goossens, Martine; Pauwels, Katia; Breyer, Didier

2012-03-30

23

Immune and histopathological responses in animals vaccinated with recombinant vaccinia viruses that express individual genes of human respiratory syncytial virus.  

PubMed

Previous reports have established that vaccinia virus (VV) recombinants expressing G, F, or N protein of respiratory syncytial (RS) virus protect small animals against intranasal challenge with live RS virus. This work demonstrates that a variety of parameters affect the protection induced by recombinant viruses. The route of vaccination, the subtype of challenge virus, and the species used influenced the antibody titers and extent of protection. During these studies, observations were also made on the subclass of antibody generated, and pulmonary histopathological changes induced by challenge after vaccination were noted. The effect of route of inoculation on host response was examined by vaccinating mice intranasally, intraperitoneally, or by scarification with a recombinant VV expressing the RS virus G glycoprotein. Intranasal vaccination induced 25-fold-higher titers of antibody to RS virus in the lung than the intraperitoneal route did, but both routes resulted in complete suppression of virus replication after intranasal challenge 21 days after vaccination. Scarification was a less effective method of vaccination. The antibody induced by recombinant VV in mice was mostly immunoglobulin G2a (IgG2a) with some IgG2b. No antibody to RS virus was detected in the IgA, IgM, IgG1, or IgG3 subclass irrespective of the vaccination route. The G and F glycoproteins were shown to elicit similar subclasses of antibody. However, animals vaccinated with the G and F vectors differed strikingly in their response to challenge by heterologous virus. Mice or cotton rats vaccinated with recombinant VV carrying the G gene of RS virus were protected against challenge only with homologous subtype A virus. Vaccination with a recombinant VV expressing the F glycoprotein induced protection against both homologous and heterologous subtype B virus challenge. The protection induced in mice was greater than that detected in cotton rats, indicating that the host may also affect immunity. Finally, this report describes histological examination of mouse lungs after vaccination and challenge. Vaccinated mice that were subsequently challenged had significantly greater lung lesion scores than unvaccinated challenged mice. The lesions were primarily peribronchiolar and perivascular infiltrations of polymorphonuclear cells and lymphocytes. Further work will establish whether these pulmonary changes are a desirable immune response to virus invasion or a potential immunopathogenic hazard. The results have important implications for planning a strategy of vaccination against RS virus and emphasize potential dangers that may attend the use of recombinant VV as vaccines. PMID:3316707

Stott, E J; Taylor, G; Ball, L A; Anderson, K; Young, K K; King, A M; Wertz, G W

1987-12-01

24

A mouse-based assay for the pre-clinical neurovirulence assessment of vaccinia virus-based smallpox vaccines  

PubMed Central

Post-vaccinal encephalitis, although relatively uncommon, is a known adverse event associated with many live, attenuated smallpox vaccines. Although smallpox vaccination ceased globally in 1980, vaccine manufacture has resumed in response to concerns over the possible use of smallpox virus as an agent of bioterrorism. To better support the production of safer smallpox vaccines, we previously reported the development of a mouse model in which a relatively attenuated vaccine strain (Dryvax®) could be discerned from a more virulent laboratory strain (WR). Here we have further tested the performance of this assay by evaluating the neurovirulence of several vaccinia virus-based smallpox vaccines spanning a known range in neurovirulence for humans. Our data indicate that testing of 10 to 100 pfu of virus in mice following intracranial inoculation reliably assesses the virus’s neurovirulence potential for humans.

Zhang, Cheryl X.; Sauder, Christian; Malik, Tahir; Rubin, Steven A.

2009-01-01

25

Enhanced immunogenicity and protective effect conferred by vaccination with combinations of modified vaccinia virus Ankara and licensed smallpox vaccine Dryvax in a mouse model  

Microsoft Academic Search

Significant adverse events are associated with vaccination with the currently licensed smallpox vaccine. Candidate new-generation smallpox vaccines such as the replication-defective modified vaccinia virus Ankara (MVA) produce very few adverse events in experimental animals and in limited human clinical trials conducted near the end of the smallpox eradication campaign. Efficacy evaluation of such new-generation vaccines will be extraordinarily complex, however,

Clement A. Meseda; Alonzo D. Garcia; Arunima Kumar; Anne E. Mayer; Jody Manischewitz; Lisa R. King; Hana Golding; Michael Merchlinsky; Jerry P. Weir

2005-01-01

26

CD4+ T cells provide intermolecular help to generate robust antibody responses in vaccinia virus-vaccinated humans.  

PubMed

Immunization with vaccinia virus elicits a protective Ab response that is almost completely CD4(+) T cell dependent. A recent study in a rodent model observed a deterministic linkage between Ab and CD4(+) T cell responses to particular vaccinia virus proteins suggesting that CD4(+) T cell help is preferentially provided to B cells with the same protein specificity (Sette et al. 2008. Immunity 28: 847-858). However, a causal linkage between Ab and CD4(+) T cell responses to vaccinia or any other large pathogen in humans has yet to be done. In this study, we measured the Ab and CD4(+) T cell responses against four vaccinia viral proteins (A27L, A33R, B5R, and L1R) known to be strongly targeted by humoral and cellular responses induced by vaccinia virus vaccination in 90 recently vaccinated and 7 long-term vaccinia-immunized human donors. Our data indicate that there is no direct linkage between Ab and CD4(+) T cell responses against each individual protein in both short-term and long-term immunized donors. Together with the observation that the presence of immune responses to these four proteins is linked together within donors, our data suggest that in vaccinia-immunized humans, individual viral proteins are not the primary recognition unit of CD4(+) T cell help for B cells. Therefore, we have for the first time, to our knowledge, shown evidence that CD4(+) T cells provide intermolecular (also known as noncognate or heterotypic) help to generate robust Ab responses against four vaccinia viral proteins in humans. PMID:23667112

Yin, Liusong; Calvo-Calle, J Mauricio; Cruz, John; Newman, Frances K; Frey, Sharon E; Ennis, Francis A; Stern, Lawrence J

2013-06-15

27

Oral vaccination with modified vaccinia virus Ankara attached covalently to TMPEG-modified cationic liposomes overcomes pre-existing poxvirus immunity from recombinant vaccinia immunization  

PubMed Central

Development of a safe and effective vaccine for induction of mucosal immunity to the human immunodeficiency virus (HIV) envelope glycoprotein (Env, gp160) represents the best hope for containing the spread of an HIV epidemic worldwide. The highly attenuated modified vaccinia virus Ankara (MVA) is a laboratory virus well suited as a safe vaccine vector. However, the presence of pre-existing immunity to Vaccinia virus in the adult population represents a hindrance that limits the application of the MVA vector for inducing immunity to HIV antigens. Here, cationic liposomes were covalently attached to the surface of recombinant MVA expressing the HIV-1 strain IIIB Env glycoprotein and ?-galactosidase (MVAIIIB/?-gal) using tresylmonomethoxypolyethylene glycol (TMPEG) grafted into a lipid membrane without compromising viral infectivity in vitro and in vivo. The orally administered MVAIIIB/?-gal–TMPEG/liposome complexes were capable of delivering the transgenes to mucosal tissues in mice with pre-existing poxvirus immunity based on ?-galactosidase gene expression in intestinal tissues measured 18 h after infection. Importantly, the MVAIIIB/?-gal–TMPEG/liposome complexes enhanced Env-specific cellular and humoral immune responses in the mucosal and systemic tissues after repeated oral immunization of BALB/c mice. This approach may prove useful for induction of protective immunity against infectious diseases and cancer in populations with pre-existing immunity to vaccinia from smallpox vaccination.

Naito, Toshio; Kaneko, Yutaro; Kozbor, Danuta

2008-01-01

28

Infectious vaccinia virus recombinants that express hepatitis B virus surface antigen  

NASA Astrophysics Data System (ADS)

Potential live vaccines against hepatitis B virus have been produced. The coding sequence for hepatitis B virus surface antigen (HBsAg) has been inserted into the vaccinia virus genome under control of vaccinia virus early promoters. Cells infected with these vaccinia virus recombinants synthesize and excrete HBsAg and vaccinated rabbits rapidly produce antibodies to HBsAg.

Smith, Geoffrey L.; Mackett, Michael; Moss, Bernard

1983-04-01

29

Modified Vaccinia Virus Ankara (MVA) as Production Platform for Vaccines against Influenza and Other Viral Respiratory Diseases  

PubMed Central

Respiratory viruses infections caused by influenza viruses, human parainfluenza virus (hPIV), respiratory syncytial virus (RSV) and coronaviruses are an eminent threat for public health. Currently, there are no licensed vaccines available for hPIV, RSV and coronaviruses, and the available seasonal influenza vaccines have considerable limitations. With regard to pandemic preparedness, it is important that procedures are in place to respond rapidly and produce tailor made vaccines against these respiratory viruses on short notice. Moreover, especially for influenza there is great need for the development of a universal vaccine that induces broad protective immunity against influenza viruses of various subtypes. Modified Vaccinia Virus Ankara (MVA) is a replication-deficient viral vector that holds great promise as a vaccine platform. MVA can encode one or more foreign antigens and thus functions as a multivalent vaccine. The vector can be used at biosafety level 1, has intrinsic adjuvant capacities and induces humoral and cellular immune responses. However, there are some practical and regulatory issues that need to be addressed in order to develop MVA-based vaccines on short notice at the verge of a pandemic. In this review, we discuss promising novel influenza virus vaccine targets and the use of MVA for vaccine development against various respiratory viruses.

Altenburg, Arwen F.; Kreijtz, Joost H. C. M.; de Vries, Rory D.; Song, Fei; Fux, Robert; Rimmelzwaan, Guus F.; Sutter, Gerd; Volz, Asisa

2014-01-01

30

Modified Vaccinia Virus Ankara (MVA) as Production Platform for Vaccines against Influenza and Other Viral Respiratory Diseases.  

PubMed

Respiratory viruses infections caused by influenza viruses, human parainfluenza virus (hPIV), respiratory syncytial virus (RSV) and coronaviruses are an eminent threat for public health. Currently, there are no licensed vaccines available for hPIV, RSV and coronaviruses, and the available seasonal influenza vaccines have considerable limitations. With regard to pandemic preparedness, it is important that procedures are in place to respond rapidly and produce tailor made vaccines against these respiratory viruses on short notice. Moreover, especially for influenza there is great need for the development of a universal vaccine that induces broad protective immunity against influenza viruses of various subtypes. Modified Vaccinia Virus Ankara (MVA) is a replication-deficient viral vector that holds great promise as a vaccine platform. MVA can encode one or more foreign antigens and thus functions as a multivalent vaccine. The vector can be used at biosafety level 1, has intrinsic adjuvant capacities and induces humoral and cellular immune responses. However, there are some practical and regulatory issues that need to be addressed in order to develop MVA-based vaccines on short notice at the verge of a pandemic. In this review, we discuss promising novel influenza virus vaccine targets and the use of MVA for vaccine development against various respiratory viruses. PMID:25036462

Altenburg, Arwen F; Kreijtz, Joost H C M; de Vries, Rory D; Song, Fei; Fux, Robert; Rimmelzwaan, Guus F; Sutter, Gerd; Volz, Asisa

2014-01-01

31

Quantification of antibody responses against multiple antigens of the two infectious forms of Vaccinia virus provides a benchmark for smallpox vaccination  

Microsoft Academic Search

Smallpox was eradicated without an adequate understanding of how vaccination induced protection. In response to possible bioterrorism with smallpox, the UK government vaccinated ?300 health care workers with vaccinia virus (VACV) strain Lister. Antibody responses were analyzed using ELISA for multiple surface antigens of the extracellular enveloped virus (EEV) and the intracellular mature virus (IMV), plaque reduction neutralization and a

Mike M Pütz; Claire M Midgley; Mansun Law; Geoffrey L Smith

2006-01-01

32

Preclinical evaluation of a modified vaccinia virus Ankara (MVA)-based vaccine against influenza A\\/H5N1 viruses  

Microsoft Academic Search

Highly pathogenic avian influenza viruses of the H5N1 subtype are responsible for an increasing number of infections in humans since 2003. More than 60% of the infections is lethal and new infections are reported frequently. In the light of the pandemic threat caused by these events the rapid availability of safe and effective vaccines is desirable. Modified vaccinia virus Ankara

J. H. C. M. Kreijtz; Y. Suezer; G. de Mutsert; J. M. A. van den Brand; G. van Amerongen; B. S. Schnierle; T. Kuiken; R. A. M. Fouchier; J. Löwer; A. D. M. E. Osterhaus; G. Sutter; G. F. Rimmelzwaan

2009-01-01

33

Safety mechanism assisted by the repressor of tetracycline (SMART) vaccinia virus vectors for vaccines and therapeutics  

PubMed Central

Replication-competent viruses, such as Vaccinia virus (VACV), are powerful tools for the development of oncolytic viral therapies and elicit superior immune responses when used as vaccine and immunotherapeutic vectors. However, severe complications from uncontrolled viral replication can occur, particularly in immunocompromised individuals or in those with other predisposing conditions. VACVs constitutively expressing interferon-? (IFN-?) replicate in cell culture indistinguishably from control viruses; however, they replicate in vivo to low or undetectable levels, and are rapidly cleared even in immunodeficient animals. In an effort to develop safe and highly effective replication-competent VACV vectors, we established a system to inducibly express IFN-?. Our SMART (safety mechanism assisted by the repressor of tetracycline) vectors are designed to express the tetracycline repressor under a constitutive VACV promoter and IFN-? under engineered tetracycline-inducible promoters. Immunodeficient SCID mice inoculated with VACVs not expressing IFN-? demonstrated severe weight loss, whereas those given VACVs expressing IFN-? under constitutive VACV promoters showed no signs of infection. Most importantly, mice inoculated with a VACV expressing the IFN-? gene under an inducible promoter remained healthy in the presence of doxycycline, but exhibited severe weight loss in the absence of doxycycline. In this study, we developed a safety mechanism for VACV based on the conditional expression of IFN-? under a tightly controlled tetracycline-inducible VACV promoter for use in vaccines and oncolytic cancer therapies.

Grigg, Patricia; Titong, Allison; Jones, Leslie A.; Yilma, Tilahun D.; Verardi, Paulo H.

2013-01-01

34

Effect of the Deletion of Genes Encoding Proteins of the Extracellular Virion Form of Vaccinia Virus on Vaccine Immunogenicity and Protective Effectiveness in the Mouse Model  

PubMed Central

Antibodies to both infectious forms of vaccinia virus, the mature virion (MV) and the enveloped virion (EV), as well as cell-mediated immune response appear to be important for protection against smallpox. EV virus particles, although more labile and less numerous than MV, are important for dissemination and spread of virus in infected hosts and thus important in virus pathogenesis. The importance of the EV A33 and B5 proteins for vaccine induced immunity and protection in a murine intranasal challenge model was evaluated by deletion of both the A33R and B5R genes in a vaccine-derived strain of vaccinia virus. Deletion of either A33R or B5R resulted in viruses with a small plaque phenotype and reduced virus yields, as reported previously, whereas deletion of both EV protein-encoding genes resulted in a virus that formed small infection foci that were detectable and quantifiable only by immunostaining and an even more dramatic decrease in total virus yield in cell culture. Deletion of B5R, either as a single gene knockout or in the double EV gene knockout virus, resulted in a loss of EV neutralizing activity, but all EV gene knockout viruses still induced a robust neutralizing activity against the vaccinia MV form of the virus. The effect of elimination of A33 and/or B5 on the protection afforded by vaccination was evaluated by intranasal challenge with a lethal dose of either vaccinia virus WR or IHD-J, a strain of vaccinia virus that produces relatively higher amounts of EV virus. The results from multiple experiments, using a range of vaccination doses and virus challenge doses, and using mortality, morbidity, and virus dissemination as endpoints, indicate that the absence of A33 and B5 have little effect on the ability of a vaccinia vaccine virus to provide protection against a lethal intranasal challenge in a mouse model.

Meseda, Clement A.; Campbell, Joseph; Kumar, Arunima; Garcia, Alonzo D.; Merchlinsky, Michael; Weir, Jerry P.

2013-01-01

35

Effect of the deletion of genes encoding proteins of the extracellular virion form of vaccinia virus on vaccine immunogenicity and protective effectiveness in the mouse model.  

PubMed

Antibodies to both infectious forms of vaccinia virus, the mature virion (MV) and the enveloped virion (EV), as well as cell-mediated immune response appear to be important for protection against smallpox. EV virus particles, although more labile and less numerous than MV, are important for dissemination and spread of virus in infected hosts and thus important in virus pathogenesis. The importance of the EV A33 and B5 proteins for vaccine induced immunity and protection in a murine intranasal challenge model was evaluated by deletion of both the A33R and B5R genes in a vaccine-derived strain of vaccinia virus. Deletion of either A33R or B5R resulted in viruses with a small plaque phenotype and reduced virus yields, as reported previously, whereas deletion of both EV protein-encoding genes resulted in a virus that formed small infection foci that were detectable and quantifiable only by immunostaining and an even more dramatic decrease in total virus yield in cell culture. Deletion of B5R, either as a single gene knockout or in the double EV gene knockout virus, resulted in a loss of EV neutralizing activity, but all EV gene knockout viruses still induced a robust neutralizing activity against the vaccinia MV form of the virus. The effect of elimination of A33 and/or B5 on the protection afforded by vaccination was evaluated by intranasal challenge with a lethal dose of either vaccinia virus WR or IHD-J, a strain of vaccinia virus that produces relatively higher amounts of EV virus. The results from multiple experiments, using a range of vaccination doses and virus challenge doses, and using mortality, morbidity, and virus dissemination as endpoints, indicate that the absence of A33 and B5 have little effect on the ability of a vaccinia vaccine virus to provide protection against a lethal intranasal challenge in a mouse model. PMID:23785523

Meseda, Clement A; Campbell, Joseph; Kumar, Arunima; Garcia, Alonzo D; Merchlinsky, Michael; Weir, Jerry P

2013-01-01

36

Genomic sequence and virulence of clonal isolates of vaccinia virus Tiantan, the Chinese smallpox vaccine strain.  

PubMed

Despite the worldwide eradication of smallpox in 1979, the potential bioterrorism threat from variola virus and the ongoing use of vaccinia virus (VACV) as a vector for vaccine development argue for continued research on VACV. In China, the VACV Tiantan strain (TT) was used in the smallpox eradication campaign. Its progeny strain is currently being used to develop a human immunodeficiency virus (HIV) vaccine. Here we sequenced the full genomes of five TT clones isolated by plaque purification from the TT (752-1) viral stock. Phylogenetic analysis with other commonly used VACV strains showed that TT (752-1) and its clones clustered and exhibited higher sequence diversity than that found in Dryvax clones. The ?190 kbp genomes of TT appeared to encode 273 open reading frames (ORFs). ORFs located in the middle of the genome were more conserved than those located at the two termini, where many virulence and immunomodulation associated genes reside. Several patterns of nucleotide changes including point mutations, insertions and deletions were identified. The polymorphisms in seven virulence-associated proteins and six immunomodulation-related proteins were analyzed. We also investigated the neuro- and skin- virulence of TT clones in mice and rabbits, respectively. The TT clones exhibited significantly less virulence than the New York City Board of Health (NYCBH) strain, as evidenced by less extensive weight loss and morbidity in mice as well as produced smaller skin lesions and lower incidence of putrescence in rabbits. The complete genome sequences, ORF annotations, and phenotypic diversity yielded from this study aid our understanding of the Chinese historic TT strain and are useful for HIV vaccine projects employing TT as a vector. PMID:23593246

Zhang, Qicheng; Tian, Meijuan; Feng, Yi; Zhao, Kai; Xu, Jing; Liu, Ying; Shao, Yiming

2013-01-01

37

Genomic Sequence and Virulence of Clonal Isolates of Vaccinia Virus Tiantan, the Chinese Smallpox Vaccine Strain  

PubMed Central

Despite the worldwide eradication of smallpox in 1979, the potential bioterrorism threat from variola virus and the ongoing use of vaccinia virus (VACV) as a vector for vaccine development argue for continued research on VACV. In China, the VACV Tiantan strain (TT) was used in the smallpox eradication campaign. Its progeny strain is currently being used to develop a human immunodeficiency virus (HIV) vaccine. Here we sequenced the full genomes of five TT clones isolated by plaque purification from the TT (752-1) viral stock. Phylogenetic analysis with other commonly used VACV strains showed that TT (752-1) and its clones clustered and exhibited higher sequence diversity than that found in Dryvax clones. The ?190 kbp genomes of TT appeared to encode 273 open reading frames (ORFs). ORFs located in the middle of the genome were more conserved than those located at the two termini, where many virulence and immunomodulation associated genes reside. Several patterns of nucleotide changes including point mutations, insertions and deletions were identified. The polymorphisms in seven virulence-associated proteins and six immunomodulation-related proteins were analyzed. We also investigated the neuro- and skin- virulence of TT clones in mice and rabbits, respectively. The TT clones exhibited significantly less virulence than the New York City Board of Health (NYCBH) strain, as evidenced by less extensive weight loss and morbidity in mice as well as produced smaller skin lesions and lower incidence of putrescence in rabbits. The complete genome sequences, ORF annotations, and phenotypic diversity yielded from this study aid our understanding of the Chinese historic TT strain and are useful for HIV vaccine projects employing TT as a vector.

Zhang, Qicheng; Tian, Meijuan; Feng, Yi; Zhao, Kai; Xu, Jing; Liu, Ying; Shao, Yiming

2013-01-01

38

Modified Vaccinia Virus Ankara Immunization Protects against Lethal Challenge with Recombinant Vaccinia Virus Expressing Murine Interleukin4  

Microsoft Academic Search

Recent events have raised concern over the use of pathogens, including variola virus, as biological weapons. Vaccination with Dryvax is associated with serious side effects and is contraindicated for many people, and the development of a safer effective smallpox vaccine is necessary. We evaluated an attenuated vaccinia virus, modified vaccinia virus Ankara (MVA), by use of a murine model to

Lewis H. McCurdy; John A. Rutigliano; Teresa R. Johnson; Man Chen; Barney S. Graham

2004-01-01

39

Discovery of Naturally Processed and HLA-Presented Class I Peptides from Vaccinia Virus Infection using Mass Spectrometry for Vaccine Development  

PubMed Central

An important approach for developing a safer smallpox vaccine is to identify naturally processed immunogenic vaccinia-derived peptides rather than live whole vaccinia virus. We used two-dimensional liquid chromatography coupled to mass spectrometry to identify 116 vaccinia peptides, encoded by 61 open reading frames, from a B-cell line (homozygous for HLA class I A*0201, B*1501, and C*03) after infection with vaccinia virus (Dryvax). Importantly, 68 of these peptides are conserved in variola, providing insight into the peptides that induce protection against smallpox. Twenty-one of these 68 conserved peptides were 11 amino acids long or longer, outside of the range of most predictive algorithms. Thus, direct identification of naturally processed and presented HLA peptides gives important information not provided by current computational methods for identifying potential vaccinia epitopes.

Johnson, Kenneth L.; Ovsyannikova, Inna G.; Mason, Christopher J.; Bergen, H. Robert; Poland, Gregory A.

2009-01-01

40

Induction of Antibody Responses to African Horse Sickness Virus (AHSV) in Ponies after Vaccination with Recombinant Modified Vaccinia Ankara (MVA)  

PubMed Central

Background African horse sickness virus (AHSV) causes a non-contagious, infectious disease in equids, with mortality rates that can exceed 90% in susceptible horse populations. AHSV vaccines play a crucial role in the control of the disease; however, there are concerns over the use of polyvalent live attenuated vaccines particularly in areas where AHSV is not endemic. Therefore, it is important to consider alternative approaches for AHSV vaccine development. We have carried out a pilot study to investigate the ability of recombinant modified vaccinia Ankara (MVA) vaccines expressing VP2, VP7 or NS3 genes of AHSV to stimulate immune responses against AHSV antigens in the horse. Methodology/Principal Findings VP2, VP7 and NS3 genes from AHSV-4/Madrid87 were cloned into the vaccinia transfer vector pSC11 and recombinant MVA viruses generated. Antigen expression or transcription of the AHSV genes from cells infected with the recombinant viruses was confirmed. Pairs of ponies were vaccinated with MVAVP2, MVAVP7 or MVANS3 and both MVA vector and AHSV antigen-specific antibody responses were analysed. Vaccination with MVAVP2 induced a strong AHSV neutralising antibody response (VN titre up to a value of 2). MVAVP7 also induced AHSV antigen–specific responses, detected by western blotting. NS3 specific antibody responses were not detected. Conclusions This pilot study demonstrates the immunogenicity of recombinant MVA vectored AHSV vaccines, in particular MVAVP2, and indicates that further work to investigate whether these vaccines would confer protection from lethal AHSV challenge in the horse is justifiable.

Maan, Sushila; Rao, Shujing; Mertens, Peter; Blacklaws, Barbara; Davis-Poynter, Nick; Wood, James; Castillo-Olivares, Javier

2009-01-01

41

Identification and preliminary characterization of vaccinia virus (Dryvax) antigens recognized by vaccinia immune globulin  

Microsoft Academic Search

Using vaccinia immune globulin (VIG), a high-titer antibody preparation from immunized subjects, we demonstrate that the humoral immune response in humans is directed against numerous antigens in the Dryvax vaccine strain. Western blot and immunoprecipitation analyses revealed highly antigenic proteins associated with both the extracellular enveloped virus and intracellular mature virus forms. The modified vaccinia virus Ankara (MVA), a new

Agnes Jones-Trower; Alonzo Garcia; Clement A. Meseda; Yong He; Carol Weiss; Arunima Kumar; Jerry P. Weir; Michael Merchlinsky

2005-01-01

42

Priming-Boosting Vaccination with Recombinant Mycobacterium bovis Bacillus Calmette-Guerin and a Nonreplicating Vaccinia Virus Recombinant Leads to Long-Lasting and Effective Immunity  

Microsoft Academic Search

Virus-specific T-cell responses can limit immunodeficiency virus type 1 (HIV-1) transmission and prevent disease progression and so could serve as the basis for an affordable, safe, and effective vaccine in humans. To assess their potential for a vaccine, we used Mycobacterium bovis bacillus Calmette-Guerin (BCG)-Tokyo and a replication-deficient vaccinia virus strain (DIs) as vectors to express full-length gag from simian

Yasushi Ami; Yasuyuki Izumi; Kazuhiro Matsuo; Kenji Someya; Masaru Kanekiyo; Shigeo Horibata; Naoto Yoshino; Koji Sakai; Katsuaki Shinohara; Sohkichi Matsumoto; Takeshi Yamada; Shudo Yamazaki; Naoki Yamamoto; Mitsuo Honda

2005-01-01

43

Evaluation of modified vaccinia virus Ankara based recombinant SARS vaccine in ferrets  

Microsoft Academic Search

Severe acute respiratory syndrome (SARS) caused by a newly identified coronavirus (SARS-CoV) remains a threat to cause epidemics as evidenced by recent sporadic cases in China. In this communication, we evaluated the efficacy and safety of two SARS vaccine candidates based on the recombinant modified vaccinia Ankara (MVA) expressing SARS-CoV spike or nucleocapsid proteins in ferrets. No clinical signs were

Markus Czub; Hana Weingartl; Stefanie Czub; Runtao He; Jingxin Cao

2005-01-01

44

DNA and Modified Vaccinia Virus Ankara Vaccines Encoding Multiple Cytotoxic and Helper T-Lymphocyte Epitopes of Human Immunodeficiency Virus Type 1 (HIV-1) Are Safe but Weakly Immunogenic in HIV-1-Uninfected, Vaccinia Virus-Naive Adults  

PubMed Central

We evaluated a DNA plasmid-vectored vaccine and a recombinant modified vaccinia virus Ankara vaccine (MVA-mBN32), each encoding cytotoxic and helper T-lymphocyte epitopes of human immunodeficiency virus type 1 (HIV-1) in a randomized, double-blinded, placebo-controlled trial in 36 HIV-1-uninfected adults using a heterologous prime-boost schedule. HIV-1-specific cellular immune responses, measured as interleukin-2 and/or gamma interferon production, were induced in 1 (4%) of 28 subjects after the first MVA-mBN32 immunization and in 3 (12%) of 25 subjects after the second MVA-mBN32 immunization. Among these responders, polyfunctional T-cell responses, including the production of tumor necrosis factor alpha and perforin, were detected. Vaccinia virus-specific antibodies were induced to the MVA vector in 27 (93%) of 29 and 26 (93%) of 28 subjects after the first and second immunizations with MVA-mBN32. These peptide-based vaccines were safe but were ineffective at inducing HIV-1-specific immune responses and induced much weaker responses than MVA vaccines expressing the entire open reading frames of HIV-1 proteins.

Newman, Mark J.; deCamp, Allan; Hay, Christine Mhorag; De Rosa, Stephen C.; Noonan, Elizabeth; Livingston, Brian D.; Fuchs, Jonathan D.; Kalams, Spyros A.; Cassis-Ghavami, Farah L.

2012-01-01

45

Passatempo Virus, a Vaccinia Virus Strain, Brazil  

PubMed Central

Passatempo virus was isolated during a zoonotic outbreak. Biologic features and molecular characterization of hemagglutinin, thymidine kinase, and vaccinia growth factor genes suggested a vaccinia virus infection, which strengthens the idea of the reemergence and circulation of vaccinia virus in Brazil. Molecular polymorphisms indicated that Passatempo virus is a different isolate.

Leite, Juliana A.; Drumond, Betania P.; Trindade, Giliane S.; Lobato, Zelia I.P.; da Fonseca, Flavio G.; dos Santos, Joao R.; Madureira, Marieta C.; Guedes, Maria I.M.C.; Ferreira, Jaqueline M.S.; Bonjardim, Claudio A.; Ferreira, Paulo C.P.

2005-01-01

46

Protection of mice from fatal measles encephalitis by vaccination with vaccinia virus recombinants encoding either the hemagglutinin or the fusion protein.  

PubMed Central

Vaccinia virus recombinants encoding the hemagglutinin or fusion protein of measles virus have been constructed. Infection of cell cultures with the recombinants led to the synthesis of authentic measles proteins as judged by their electrophoretic mobility, recognition by antibodies, glycosylation, proteolytic cleavage, and presentation on the cell surface. Mice vaccinated with a single dose of the recombinant encoding the hemagglutinin protein developed antibodies capable of both inhibiting hemagglutination activity and neutralizing measles virus, whereas animals vaccinated with the recombinant encoding the fusion protein developed measles neutralizing antibodies. Mice vaccinated with either of the recombinants resisted a normally lethal intracerebral inoculation of a cell-associated measles virus subacute sclerosing panencephalitis strain. Images

Drillien, R; Spehner, D; Kirn, A; Giraudon, P; Buckland, R; Wild, F; Lecocq, J P

1988-01-01

47

Safety and Immunogenicity of DNA Prime and Modified Vaccinia Ankara Virus-HIV Subtype C Vaccine Boost in Healthy Adults  

PubMed Central

A randomized, double-blind, placebo-controlled phase I trial was conducted in 32 HIV-uninfected healthy volunteers to assess the safety and immunogenicity of 3 doses of DNA vaccine (Advax) plus 1 dose of recombinant modified vaccinia virus Ankara (MVA) (TBC-M4) or 3 doses of TBC-M4 alone (groups A and B, respectively). Both vaccine regimens were found to be safe and well tolerated. Gamma interferon (IFN-?) enzyme-linked immunosorbent spot (ELISPOT) assay responses were detected in 1/10 (10%) individuals in group A after three Advax primes and in 9/9 individuals (100%) after the MVA boost. In group B, IFN-? ELISPOT responses were detected in 6/12 (50%) and 7/11 (64%) individuals after the second and third MVA vaccinations, respectively. Responses to all vaccine components, but predominantly to Env, were seen. The breadth and magnitude of the T cell response and viral inhibition were greater in group A than in group B, indicating that the quality of the T-cell response was enhanced by the DNA prime. Intracellular cytokine staining indicated that the T-cell responses were polyfunctional but were skewed toward Env with a CD4+ phenotype. At 2 weeks after the last vaccination, HIV-specific antibody responses were detected in all (100%) group B and 1/11 (9.1%) group A vaccinees. Vaccinia virus-specific responses were detected in all (100%) group B and 2/11 (18.2%) group A vaccinees. In conclusion, HIV-specific T-cell responses were seen in the majority of volunteers in groups A and B but with a trend toward greater quality of the T-cell response in group A. Antibody responses were better in group B than in group A.

Hayes, Peter; Gilmour, Jill; von Lieven, Andrea; Gill, Dilbinder; Clark, Lorna; Kopycinski, Jakub; Cheeseman, Hannah; Chung, Amy; Alter, Galit; Dally, Len; Zachariah, Devika; Lombardo, Angela; Ackland, James; Sayeed, Eddy; Jackson, Akil; Boffito, Marta; Gazzard, Brian; Fast, Patricia E.; Laufer, Dagna

2013-01-01

48

Development of a replication-deficient recombinant vaccinia virus vaccine effective against parainfluenza virus 3 infection in an animal model  

Microsoft Academic Search

The highly attenuated, replication-deficient, modified vaccinia virus Ankara (MVA) was used to express the fusion (F) and\\/or hemagglutinin-neuraminidase (HN) glycoproteins of parainfluenza virus 3 (PIV3). Initial recombinant viruses in which the HN gene was regulated by a very strong synthetic earlyllate promoter replicated poorly in permissive chick embryo cells evidently due to toxic levels of the gene product. This result

Linda S. Wyatt; Scott T. Shors; Brian R. Murphy; Bernard Moss

1996-01-01

49

Direct comparison of antigen production and induction of apoptosis by canarypox virus- and modified vaccinia virus ankara-human immunodeficiency virus vaccine vectors.  

PubMed

Recombinant poxvirus vectors are undergoing intensive evaluation as vaccine candidates for a variety of infectious pathogens. Avipoxviruses, such as canarypox virus, are replication deficient in mammalian cells by virtue of a poorly understood species-specific restriction. Highly attenuated vaccinia virus strains such as modified vaccinia virus Ankara (MVA) are similarly unable to complete replication in most mammalian cells but have an abortive-late phenotype, in that the block to replication occurs post-virus-specific DNA replication. In this study, an identical expression cassette for human immunodeficiency virus gag, pro, and env coding sequences was placed in canarypox virus and MVA vector backbones in order to directly compare vector-borne expression and to analyze differences in vector-host cell interactions. Antigen production by recombinant MVA was shown to be greater than that from recombinant canarypox virus in the mammalian cell lines and in the primary human cells tested. This observation was primarily due to a longer duration of antigen production in recombinant MVA-infected cells. Apoptosis induction was found to be more profound with the empty canarypox virus vector than with MVA. Remarkably, however, the inclusion of a gag/pro/env expression cassette altered the kinetics of apoptosis induction in recombinant MVA-infected cells to levels equal to those found in canarypox virus-infected cells. Antigen production by MVA was noted to be greater in human dendritic cells and resulted in enhanced T-cell stimulation in an in vitro antigen presentation assay. These results reveal differences in poxvirus vector-host cell interactions that should be relevant to their use as immunization vehicles. PMID:17409140

Zhang, Xiugen; Cassis-Ghavami, Farah; Eller, Mike; Currier, Jeff; Slike, Bonnie M; Chen, Xuemin; Tartaglia, James; Marovich, Mary; Spearman, Paul

2007-07-01

50

Modified Vaccinia Virus Ankara Recombinants Are as Potent as Vaccinia Recombinants in Diversified Prime and Boost Vaccine Regimens to Elicit Therapeutic Antitumor Responses  

Microsoft Academic Search

Cancer vaccine regimens use various strategies to enhance immune responses to specific tumor-associated antigens (TAAs), including the increasing use of recombinant poxviruses (vaccinia (rV) and fowlpox (rF)) for delivery of the TAA to the immune system. However, the use of replication competent vectors with the potential of adverse reactions have made attenuation a priority for next-generation vaccine strategies. Mod- ified

James W. Hodge; Diane J. Poole; Wilhelmina M. Aarts; Alicia Gomez Yafal; Linda Gritz; Jeffrey Schlom

2003-01-01

51

Vaccination of horses with a recombinant modified vaccinia Ankara virus (MVA) expressing African horse sickness (AHS) virus major capsid protein VP2 provides complete clinical protection against challenge  

PubMed Central

African horse sickness virus (AHSV) is an arthropod-borne pathogen that infects all species of equidae and causes high mortality in horses. Previously, a recombinant modified vaccinia Ankara (MVA) virus expressing the protein VP2 of AHSV serotype 4 was shown to induce virus neutralising antibodies in horses and protected interferon alpha receptor gene knock-out mice (IFNAR ?/?) against virulent AHSV challenge. This study builds on the previous work, examining the protective efficacy of MVA-VP2 vaccination in the natural host of AHSV infection. A study group of 4 horses was vaccinated twice with a recombinant MVA virus expressing the major capsid protein (VP2) of AHSV serotype 9. Vaccinated animals and a control group of unvaccinated horses were then challenged with a virulent strain of AHSV-9. The vaccinated animals were completely protected against clinical disease and also against viraemia as measured by standard end-point dilution assays. In contrast, all control horses presented viraemia after challenge and succumbed to the infection. These results demonstrate the potential of recombinant MVA viruses expressing the outer capsid VP2 of AHSV as a protective vaccine against AHSV infection in the field.

Alberca, Berta; Bachanek-Bankowska, Katarzyna; Cabana, Marta; Calvo-Pinilla, Eva; Viaplana, Elisenda; Frost, Lorraine; Gubbins, Simon; Urniza, Alicia; Mertens, Peter; Castillo-Olivares, Javier

2014-01-01

52

Vaccination of horses with a recombinant modified vaccinia Ankara virus (MVA) expressing African horse sickness (AHS) virus major capsid protein VP2 provides complete clinical protection against challenge.  

PubMed

African horse sickness virus (AHSV) is an arthropod-borne pathogen that infects all species of equidae and causes high mortality in horses. Previously, a recombinant modified vaccinia Ankara (MVA) virus expressing the protein VP2 of AHSV serotype 4 was shown to induce virus neutralising antibodies in horses and protected interferon alpha receptor gene knock-out mice (IFNAR -/-) against virulent AHSV challenge. This study builds on the previous work, examining the protective efficacy of MVA-VP2 vaccination in the natural host of AHSV infection. A study group of 4 horses was vaccinated twice with a recombinant MVA virus expressing the major capsid protein (VP2) of AHSV serotype 9. Vaccinated animals and a control group of unvaccinated horses were then challenged with a virulent strain of AHSV-9. The vaccinated animals were completely protected against clinical disease and also against viraemia as measured by standard end-point dilution assays. In contrast, all control horses presented viraemia after challenge and succumbed to the infection. These results demonstrate the potential of recombinant MVA viruses expressing the outer capsid VP2 of AHSV as a protective vaccine against AHSV infection in the field. PMID:24837765

Alberca, Berta; Bachanek-Bankowska, Katarzyna; Cabana, Marta; Calvo-Pinilla, Eva; Viaplana, Elisenda; Frost, Lorraine; Gubbins, Simon; Urniza, Alicia; Mertens, Peter; Castillo-Olivares, Javier

2014-06-17

53

Oral vaccination of the fox against rabies using a live recombinant vaccinia virus  

Microsoft Academic Search

Rabies, a viral disease affecting all warm-blooded animals, is prevalent in most parts of the world1, where it propagates amongst wild animals, particularly the fox and dog. The public health and economic consequences of infection in man and livestock are well known. Attempts to control the disease by vaccinating wild carnivores with inactivated or attenuated rabies virus remain controversial, and

J. Blancou; M. P. Kieny; R. Lathe; J. P. Lecocq; P. P. Pastore; J. P. Soulebot; P. Desmettre

1986-01-01

54

DNA/MVA HIV-1/AIDS Vaccine Elicits Long-Lived Vaccinia Virus-Specific Immunity and Confers Protection Against a Lethal Monkeypox Challenge  

PubMed Central

Modified vaccinia Ankara (MVA) is being tested in humans as an alternative to the current smallpox vaccine Dryvax. Here, we compare the magnitude and longevity of protective immune responses elicited by a DNA/MVA HIV-1 vaccine with those elicited by Dryvax using a monkeypox virus/macaque model. The DNA/MVA vaccine elicited similar levels of vaccinia virus (VV)-specific antibody and 5–10-fold lower levels of VV-specific cellular responses than Dryvax. This MVA-elicited cellular and humoral immunity was long lived. A subset of the DNA/MVA - and Dryvax- vaccinated macaques were subjected to a lethal monkeypox virus challenge at 3 years after vaccination. All of the vaccinated monkeys survived, whereas the unvaccinated controls succumbed to monkeypox. The viral control correlated with early post challenge levels of monkeypox-specific neutralizing antibody but not with VV-specific cellular immune response. Thus, our results demonstrate the elicitation of long lasting protective immunity for a lethal monkeypox challenge by a DNA/MVA HIV-1 vaccine.

Nigam, Pragati; Earl, Patricia L.; Americo, Jeffrey L.; Sharma, Sunita; Wyatt, Linda S.; Edghill-Spano, Yvette; Chennareddi, Lakshmi S.; Silvera, Peter; Moss, Bernard; Robinson, Harriet L.; Amara, Rama Rao

2007-01-01

55

Protective Immunity to Vaccinia Virus Induced by Vaccination with Multiple Recombinant Outer Membrane Proteins of Intracellular and Extracellular Virions  

Microsoft Academic Search

Infectious intracellular and extracellular forms of vaccinia virus have different outer membrane proteins, presenting multiple targets to the immune system. We investigated the immunogenicity of soluble forms of L1, an outer membrane protein of the intracellular mature virus, and of A33 and B5, outer membrane proteins of the extracellular enveloped virus. The recombinant proteins, in 10-g amounts mixed with a

Christiana Fogg; Shlomo Lustig; J. Charles Whitbeck; Roselyn J. Eisenberg; Gary H. Cohen; Bernard Moss

2004-01-01

56

The nonreplicating smallpox candidate vaccines defective vaccinia Lister (dVV-L) and modified vaccinia Ankara (MVA) elicit robust long-term protection  

Microsoft Academic Search

Current smallpox vaccines are live vaccinia viruses that replicate in the vaccinee inducing immunity against the deadly disease smallpox. Replication resulting in virus spread within the host, however, is the major cause of severe postvaccinal adverse events. Therefore, attenuated strains such as modified vaccinia Ankara (MVA) or LC16m8 are candidates as next generation vaccines. These strains are usually grown in

S. Coulibaly; P. Brühl; J. Mayrhofer; K. Schmid; M. Gerencer; F. G. Falkner

2005-01-01

57

Broad and potent cellular and humoral immune responses after a second late HIV-modified vaccinia virus ankara vaccination in HIV-DNA-primed and HIV-modified vaccinia virus Ankara-boosted Swedish vaccinees.  

PubMed

We have previously shown that an HIV vaccine regimen including three HIV-DNA immunizations and a single HIV-modified vaccinia virus Ankara (MVA) boost was safe and highly immunogenic in Swedish volunteers. A median 38 months after the first HIV-MVA vaccination, 24 volunteers received 10(8) plaque-forming units of HIV-MVA. The vaccine was well tolerated. Two weeks after this HIV-MVA vaccination, 18 (82%) of 22 evaluable vaccinees were interferon (IFN)-? enzyme-linked immunospot (ELISpot) reactive: 18 to Gag and 10 (45%) to Env. A median minimal epitope count of 4 to Gag or Env was found in a subset of 10 vaccinees. Intracellular cytokine staining revealed CD4(+) and/or CD8(+) T cell responses in 23 (95%) of 24 vaccinees, 19 to Gag and 19 to Env. The frequency of HIV-specific CD4(+) and CD8(+) T cell responses was equally high (75%). A high proportion of CD4(+) and CD8(+) T cell responses to Gag was polyfunctional with production of three or more cytokines (40% and 60%, respectively). Of the Env-specific CD4(+) T cells 40% were polyfunctional. Strong lymphoproliferative responses to Aldrithiol-2 (AT-2)-treated subtype A, B, C, and A_E virus were demonstrable in 21 (95%) of 22 vaccinees. All vaccinees developed binding antibodies to Env and Gag. Neutralizing antibodies were detected in a peripheral blood mononuclear cell (PBMC)-based assay against subtype B and CRF01_AE viruses. The neutralizing antibody response rates were influenced by the vaccine dose and/or mode of delivery used at the previous HIV-MVA vaccination. Thus, a second late HIV-MVA boost induced strong and broad cellular immune responses and improved antibody responses. The data support further exploration of this vaccine concept. PMID:24090081

Nilsson, Charlotta; Godoy-Ramirez, Karina; Hejdeman, Bo; Bråve, Andreas; Gudmundsdotter, Lindvi; Hallengärd, David; Currier, Jeffrey R; Wieczorek, Lindsay; Hasselrot, Klara; Earl, Patricia L; Polonis, Victoria R; Marovich, Mary A; Robb, Merlin L; Sandström, Eric; Wahren, Britta; Biberfeld, Gunnel

2014-03-01

58

Expression of rabies virus glycoprotein from a recombinant vaccinia virus  

Microsoft Academic Search

Rabies is one of the oldest diseases known to man, but its successful control has remained elusive. Although effective vaccines of tissue culture origin against rabies do exist1, such preparations are expensive. Live vaccinia virus (VV) recombinants expressing influenza or hepatitis B antigens have recently been used to immunize against these diseases2-4. We have now used this approach to produce

M. P. Kieny; R. Lathe; R. Drillien; D. Spehner; S. Skory; D. Schmitt; T. Wiktor; H. Koprowski; J. P. Lecocq

1984-01-01

59

Risk of Vaccinia Transfer to the Hands of Vaccinated Persons after Smallpox Immunization  

Microsoft Academic Search

Transmission of vaccinia virus after smallpox vaccination is a concern. We conducted a prospective examination of the protection afforded by vaccination-site bandages in recently vaccinated individuals. After smallpox vaccination, inoculation sites were covered with 2 occlusive dressings. Site assessment and bandage changes occurred every 3-5 days until the site was healed. At each visit, specimens from the vaccination site, outer

Thomas R. Talbot; Ellis Ziel; Jennifer K. Doersam; Bonnie LaFleur; Sharon Tollefson; Kathryn M. Edwards

2004-01-01

60

Towards a universal vaccine for avian influenza: Protective efficacy of modified Vaccinia virus Ankara and Adenovirus vaccines expressing conserved influenza antigens in chickens challenged with low pathogenic avian influenza virus  

PubMed Central

Current vaccines targeting surface proteins can drive antigenic variation resulting either in the emergence of more highly pathogenic viruses or of antigenically distinct viruses that escape control by vaccination and thereby persist in the host population. Influenza vaccines typically target the highly mutable surface proteins and do not provide protection against heterologous challenge. Vaccines which induce immune responses against conserved influenza epitopes may confer protection against heterologous challenge. We report here the results of vaccination with recombinant modified Vaccinia virus Ankara (MVA) and Adenovirus (Ad) expressing a fusion construct of nucleoprotein and matrix protein (NP + M1). Prime and boost vaccination regimes were trialled in different ages of chicken and were found to be safe and immunogenic. Interferon-? (IFN-?) ELISpot was used to assess the cellular immune response post secondary vaccination. In ovo Ad prime followed by a 4 week post hatch MVA boost was identified as the most immunogenic regime in one outbred and two inbred lines of chicken. Following vaccination, one inbred line (C15I) was challenged with low pathogenic avian influenza (LPAI) H7N7 (A/Turkey/England/1977). Birds receiving a primary vaccination with Ad-NP + M1 and a secondary vaccination with MVA-NP + M1 exhibited reduced cloacal shedding as measured by plaque assay at 7 days post infection compared with birds vaccinated with recombinant viruses containing irrelevant antigen. This preliminary indication of efficacy demonstrates proof of concept in birds; induction of T cell responses in chickens by viral vectors containing internal influenza antigens may be a productive strategy for the development of vaccines to induce heterologous protection against influenza in poultry.

Boyd, Amy C.; Ruiz-Hernandez, Raul; Peroval, Marylene Y.; Carson, Connor; Balkissoon, Devanand; Staines, Karen; Turner, Alison V.; Hill, Adrian V.S.; Gilbert, Sarah C.; Butter, Colin

2013-01-01

61

Towards a universal vaccine for avian influenza: protective efficacy of modified Vaccinia virus Ankara and Adenovirus vaccines expressing conserved influenza antigens in chickens challenged with low pathogenic avian influenza virus.  

PubMed

Current vaccines targeting surface proteins can drive antigenic variation resulting either in the emergence of more highly pathogenic viruses or of antigenically distinct viruses that escape control by vaccination and thereby persist in the host population. Influenza vaccines typically target the highly mutable surface proteins and do not provide protection against heterologous challenge. Vaccines which induce immune responses against conserved influenza epitopes may confer protection against heterologous challenge. We report here the results of vaccination with recombinant modified Vaccinia virus Ankara (MVA) and Adenovirus (Ad) expressing a fusion construct of nucleoprotein and matrix protein (NP+M1). Prime and boost vaccination regimes were trialled in different ages of chicken and were found to be safe and immunogenic. Interferon-? (IFN-?) ELISpot was used to assess the cellular immune response post secondary vaccination. In ovo Ad prime followed by a 4 week post hatch MVA boost was identified as the most immunogenic regime in one outbred and two inbred lines of chicken. Following vaccination, one inbred line (C15I) was challenged with low pathogenic avian influenza (LPAI) H7N7 (A/Turkey/England/1977). Birds receiving a primary vaccination with Ad-NP+M1 and a secondary vaccination with MVA-NP+M1 exhibited reduced cloacal shedding as measured by plaque assay at 7 days post infection compared with birds vaccinated with recombinant viruses containing irrelevant antigen. This preliminary indication of efficacy demonstrates proof of concept in birds; induction of T cell responses in chickens by viral vectors containing internal influenza antigens may be a productive strategy for the development of vaccines to induce heterologous protection against influenza in poultry. PMID:23200938

Boyd, Amy C; Ruiz-Hernandez, Raul; Peroval, Marylene Y; Carson, Connor; Balkissoon, Devanand; Staines, Karen; Turner, Alison V; Hill, Adrian V S; Gilbert, Sarah C; Butter, Colin

2013-01-11

62

L1R, A27L, A33R and B5R vaccinia virus genes expressed by fowlpox recombinants as putative novel orthopoxvirus vaccines  

PubMed Central

Background The traditional smallpox vaccine, administered by scarification, was discontinued in the general population from 1980, because of the absence of new smallpox cases. However, the development of an effective prophylactic vaccine against smallpox is still necessary, to protect from the threat of deliberate release of the variola virus for bioterrorism and from new zoonotic infections, and to improve the safety of the traditional vaccine. Preventive vaccination still remains the most effective control and new vectors have been developed to generate recombinant vaccines against smallpox that induce the same immunogenicity as the traditional one. As protective antibodies are mainly directed against the surface proteins of the two infectious forms of vaccinia, the intracellular mature virions and the extracellular virions, combined proteins from these viral forms can be used to better elicit a complete and protective immunity. Methods Four novel viral recombinants were constructed based on the fowlpox genetic background, which independently express the vaccinia virus L1 and A27 proteins present on the mature virions, and the A33 and B5 proteins present on the extracellular virions. The correct expression of the transgenes was determined by RT-PCR, Western blotting, and immunofluorescence. Results and conclusions Using immunoprecipitation and Western blotting, the ability of the proteins expressed by the four novel FPL1R, FPA27L, FPA33R and FPB5R recombinants to be recognized by VV-specific hyperimmune mouse sera was demonstrated. By neutralisation assays, recombinant virus particles released by infected chick embryo fibroblasts were shown not be recognised by hyperimmune sera. This thus demonstrates that the L1R, A27L, A33R and B5R gene products are not inserted into the new viral progeny. Fowlpox virus replicates only in avian species, but it is permissive for entry and transgene expression in mammalian cells, while being immunologically non–cross-reactive with vaccinia virus. These recombinants might therefore represent safer and more promising immunogens that can circumvent neutralisation by vector-generated immunity in smallpox-vaccine-experienced humans.

2013-01-01

63

Prevention of Infection by a Granulocyte-Macrophage Colony-Stimulating Factor Co-Expressing DNA/Modified Vaccinia Ankara Simian Immunodeficiency Virus Vaccine  

PubMed Central

A simian immunodeficiency virus (SIV) vaccine coexpressing granulocyte-macrophage colony stimulating factor (GM-CSF) prevented infection in 71% of macaques that received 12 rectal challenges. The SIVsmE660 challenge had the tropism of incident human immunodeficiency virus (HIV) infections and a similar genetic distance from the SIV239 vaccine as intraclade HIV isolates. The heterologous prime-boost vaccine regimen used recombinant DNA for priming and recombinant modified vaccinia Ankara for boosting. Co-expression of GM-CSF in the DNA prime enhanced the avidity of elicited immunoglobulin G for SIV envelope glycoproteins, the titers of neutralizing antibody for easy-to-neutralize SIV isolates, and antibody-dependent cellular cytotoxicity. Impressively, the co-expressed GM-CSF increased vaccine-induced prevention of infection from 25% in the non–GM-CSF co-expressing vaccine group to 71% in the GM-CSF co-expressing vaccine group. The prevention of infection showed a strong correlation with the avidity of the elicited Env-specific antibody for the Env of the SIVsmE660 challenge virus (r = 0.9; P < .0001).

Lai, Lilin; Kwa, SueFen; Kozlowski, Pamela A.; Montefiori, David C.; Ferrari, Guido; Johnson, Welkin E.; Hirsch, Vanessa; Villinger, Francois; Chennareddi, Lakshmi; Earl, Patricia L.; Moss, Bernard; Amara, Rama Rao

2011-01-01

64

Characterization and use of mammalian-expressed vaccinia virus extracellular membrane proteins for quantification of the humoral immune response to smallpox vaccines.  

PubMed

The licensed smallpox vaccine Dryvax is used as the standard in comparative immunogenicity and protection studies of new smallpox vaccine candidates. Although the correlates of protection against smallpox are unknown, recent studies have shown that a humoral response against the intracellular mature virion and extracellular enveloped virion (EV) forms of vaccinia virus is crucial for protection. Using a recombinant Semliki Forest virus (rSFV) vector system, we expressed a set of full-length EV proteins for the development of EV antigen-specific enzyme-linked immunosorbent assays (ELISAs) and the production of monospecific antisera. The EV-specific ELISAs were used to evaluate the EV humoral response elicited by Dryvax and the nonreplicating modified vaccinia virus Ankara (MVA) in mouse vaccination experiments comparing doses and routes of vaccination. Quantitatively similar titers of antibodies against EV antigens A33R, A56R, and B5R were measured in mice vaccinated with Dryvax and MVA when MVA was administered at a dose of 10(8) plaque-forming units. Further, a substantial increase in the EV-specific antibody response was induced in mice inoculated with MVA by using a prime-boost schedule. Finally, we investigated the abilities of the EV-expressing rSFV vectors to elicit the production of polyclonal monospecific antisera against the corresponding EV proteins in mice. The monospecific serum antibody levels against A33R, A56R, and B5R were measurably higher than the antibody levels induced by Dryvax. The resulting polyclonal antisera were used in Western blot analysis and immunofluorescence assays, indicating that rSFV particles are useful vectors for generating monospecific antisera. PMID:17596428

García, Alonzo D; Meseda, Clement A; Mayer, Anne E; Kumar, Arunima; Merchlinsky, Michael; Weir, Jerry P

2007-08-01

65

Structure of Intracellular Mature Vaccinia Virus Visualized by In Situ Atomic Force Microscopy  

Microsoft Academic Search

Vaccinia virus, the basis of the smallpox vaccine, is one of the largest viruses to replicate in humans. We have used in situ atomic force microscopy (AFM) to directly visualize fully hydrated, intact intracellular mature vaccinia virus (IMV) virions and chemical and enzymatic treatment products thereof. The latter included virion cores, core-enveloping coats, and core substructures. The isolated coats appeared

A. J. Malkin; A. McPherson; P. D. Gershon

2003-01-01

66

Adsorption of recombinant poxvirus L1-protein to aluminum hydroxide/CpG vaccine adjuvants enhances immune responses and protection of mice from vaccinia virus challenge.  

PubMed

The stockpiling of live vaccinia virus vaccines has enhanced biopreparedness against the intentional or accidental release of smallpox. Ongoing research on future generation smallpox vaccines is providing key insights into protective immune responses as well as important information about subunit-vaccine design strategies. For protein-based recombinant subunit vaccines, the formulation and stability of candidate antigens with different adjuvants are important factors to consider for vaccine design. In this work, a non-tagged secreted L1-protein, a target antigen on mature virus, was expressed using recombinant baculovirus technology and purified. To identify optimal formulation conditions for L1, a series of biophysical studies was performed over a range of pH and temperature conditions. The overall physical stability profile was summarized in an empirical phase diagram. Another critical question to address for development of an adjuvanted vaccine was if immunogenicity and protection could be affected by the interactions and binding of L1 to aluminum salts (Alhydrogel) with and without a second adjuvant, CpG. We thus designed a series of vaccine formulations with different binding interactions between the L1 and the two adjuvants, and then performed a series of vaccination-challenge experiments in mice including measurement of antibody responses and post-challenge weight loss and survival. We found that better humoral responses and protection were conferred with vaccine formulations when the L1-protein was adsorbed to Alhydrogel. These data demonstrate that designing vaccine formulation conditions to maximize antigen-adjuvant interactions is a key factor in smallpox subunit-vaccine immunogenicity and protection. PMID:23153450

Xiao, Yuhong; Zeng, Yuhong; Alexander, Edward; Mehta, Shyam; Joshi, Sangeeta B; Buchman, George W; Volkin, David B; Middaugh, C Russell; Isaacs, Stuart N

2013-01-01

67

Adsorption of recombinant poxvirus L1-protein to aluminum hydroxide/CpG vaccine adjuvants enhances immune responses and protection of mice from vaccinia virus challenge  

PubMed Central

The stockpiling of live vaccinia virus vaccines has enhanced biopreparedness against the intentional or accidental release of smallpox. Ongoing research on future generation smallpox vaccines is providing key insights into protective immune responses as well as important information about subunit vaccine design strategies. For protein-based recombinant subunit vaccines, the formulation and stability of candidate antigens with different adjuvants are important factors to consider for vaccine design. In this work, a non-tagged secreted L1-protein, a target antigen on mature virus, was expressed using recombinant baculovirus technology and purified. To identify optimal formulation conditions for L1, a series of biophysical studies was performed over a range of pH and temperature conditions. The overall physical stability profile was summarized in an empirical phase diagram. Another critical question to address for development of an adjuvanted-vaccine was if immunogenicity and protection could be affected by the interactions and binding of L1 to aluminum salts (Alhydrogel) with and without a second adjuvant, CpG. We thus designed a series of vaccine formulations with different binding interactions between the L1 and the two adjuvants, and then performed a series of vaccination-challenge experiments in mice including measurement of antibody responses and post-challenge weight-loss and survival. We found that better humoral responses and protection were conferred with vaccine formulations when the L1-protein was adsorbed to Alhydrogel. These data demonstrate that designing vaccine formulation conditions to maximize antigen-adjuvant interactions is a key factor in smallpox subunit vaccine immunogenicity and protection.

Xiao, Yuhong; Zeng, Yuhong; Alexander, Edward; Mehta, Shyam; Joshi, Sangeeta B.; Buchman, George W.; Volkin, David B.; Middaugh, C. Russell; Isaacs, Stuart N.

2012-01-01

68

Phase 1 Safety and Immunogenicity Testing of DNA and Recombinant Modified Vaccinia Ankara Vaccines Expressing HIV-1 Virus-like Particles  

PubMed Central

Background.?Recombinant DNA and modified vaccinia virus Ankara (rMVA) vaccines represent a promising approach to an HIV/AIDS vaccine. This Phase 1 clinical trial compared the safety and immunogenicity of a rMVA vaccine administered with and without DNA vaccine priming Methods.?GeoVax pGA2/JS7 DNA (D) and MVA/HIV62 (M) vaccines encode noninfectious virus-like particles. Intramuscular needle injections were used to deliver placebo, 2 doses of DNA followed by 2 doses of rMVA (DDMM), one dose of DNA followed by 2 doses of rMVA (DMM), or 3 doses of rMVA (MMM) to HIV-seronegative participants. Results.?Local and systemic symptoms were mild or moderate. Immune response rates for CD4 + and CD8 + T cells were highest in the DDMM group and lowest in the MMM group (77% vs 43% CD4 + and 42% vs 17% CD8 +). In contrast, response rates for Env binding and neutralizing Ab were highest in the MMM group. The DMM group had intermediate response rates. A 1/10th-dose DDMM regimen induced similar T cell but reduced Ab response rates compared with the full-dose DDMM. Conclusions.?MVA62 was well tolerated and elicited different patterns of T cell and Ab responses when administered alone or in combination with the JS7 DNA vaccine.

Elizaga, Marnie L.; Sato, Alicia; Qin, Li; Cardinali, Massimo; Hay, Christine M.; Hural, John; DeRosa, Stephen C.; DeFawe, Olivier D.; Tomaras, Georgia D.; Montefiori, David C.; Xu, Yongxian; Lai, Lilin; Kalams, Spyros A.; Baden, Lindsey R.; Frey, Sharon E.; Blattner, William A.; Wyatt, Linda S.; Moss, Bernard; Robinson, Harriet L.

2011-01-01

69

Patterns of viral replication correlate with outcome in simian immunodeficiency virus (SIV)-infected macaques: effect of prior immunization with a trivalent SIV vaccine in modified vaccinia virus Ankara.  

PubMed Central

The dynamics of plasma viremia were explored in a group of 12 simian immunodeficiency virus (SIV)-infected rhesus macaques (Macaca mulatta) that had received prior immunization with either nonrecombinant or trivalent (gag-pol, env) SIV-recombinant vaccinia viruses. Three distinct patterns of viral replication observed during and following primary viremia accounted for significant differences in survival times. High-level primary plasma viremia with subsequently increasing viremia was associated with rapid progression to AIDS (n = 2). A high-level primary plasma virus load with a transient decline and subsequent progressive increase in viremia in the post-acute phase of infection was associated with progression to AIDS within a year (n = 6). Low levels of primary plasma viremia followed by sustained restriction of virus replication were associated with maintenance of normal lymphocyte subsets and intact lymphoid architecture (n = 4), reminiscent of the profile observed in human immunodeficiency virus type 1-infected long-term nonprogressors. Three of four macaques that showed this pattern had been immunized with an SIV recombinant derived from the attenuated vaccinia virus, modified vaccinia virus Ankara. These data link the dynamics and extent of virus replication to disease course and suggest that sustained suppression of virus promotes long-term, asymptomatic survival of SIV-infected macaques. These findings also suggest that vaccine modulation of host immunity may have profound beneficial effects on the subsequent disease course, even if sterilizing immunity is not achieved.

Hirsch, V M; Fuerst, T R; Sutter, G; Carroll, M W; Yang, L C; Goldstein, S; Piatak, M; Elkins, W R; Alvord, W G; Montefiori, D C; Moss, B; Lifson, J D

1996-01-01

70

Peptide vaccination of mice immune to LCMV or vaccinia virus causes serious CD8+ T cell-mediated, TNF-dependent immunopathology  

PubMed Central

CD8+ T cells play a key role in clearing primary virus infections and in protecting against subsequent challenge. The potent antiviral effects of these cells make them important components of vaccine-induced immunity and, because of this, peptide vaccines often contain epitopes designed to induce strong CD8+ T cell responses. However, the same effector functions that protect the host also can be harmful if they are not tightly regulated, and virus-specific CD8+ T cells are a frequent cause of immunopathology. Here, we report that the administration of peptide to virus-immune recipient mice can lead to the synchronous activation of preexisting virus-specific CD8+ T cells with serious, and even lethal, consequences. Mice infected with LCMV or vaccinia virus developed rapid and profound hypothermia following injection of cognate synthetic peptides, and LCMV-infected mice frequently died within hours. Detailed analyses of the LCMV infected mice revealed enterocyte apoptosis and implicated TNF produced by peptide-specific CD8+ T cells as the major mediator of disease. The caspase inhibitor zVADfmk had no demonstrable effect on the development of hypothermia, but diminished enterocyte apoptosis and greatly reduced the number of deaths. These findings, if similarly observed in patients, counsel caution when administering powerful immunogens such as peptide vaccines to individuals who may have a large preexisting pool of epitope-specific CD8+ T cells.

Liu, Fei; Feuer, Ralph; Hassett, Daniel E.; Whitton, J. Lindsay

2006-01-01

71

Studies of a prophylactic HIV1 vaccine candidate based on modified vaccinia virus Ankara (MVA) with and without DNA priming: Effects of dosage and route on safety and immunogenicity  

Microsoft Academic Search

BackgroundTwo parallel studies evaluated safety and immunogenicity of a prophylactic HIV-1 vaccine in 192 HIV-seronegative, low-risk volunteers. Modified vaccinia virus Ankara (MVA) and plasmid DNA (pTHr) expressed HIV-1 clade A gag p24 and p17 fused to a string of 25 overlapping CD8+ T cell epitopes (HIVA).

Barry S. Peters; Walter Jaoko; Eftyhia Vardas; George Panayotakopoulos; Patricia Fast; Claudia Schmidt; Jill Gilmour; Mampedi Bogoshi; Gloria Omosa-Manyonyi; Len Dally; Linda Klavinskis; Bashir Farah; Tony Tarragona; Pierre-Alexandre Bart; Andrew Robinson; Colleen Pieterse; Wendy Stevens; Richard Thomas; Burc Barin; Andrew J. McMichael; James A. McIntyre; Giuseppe Pantaleo; Tomáš Hanke; Job Bwayo

2007-01-01

72

Vaccination of mice with a modified Vaccinia Ankara (MVA) virus expressing the African horse sickness virus (AHSV) capsid protein VP2 induces virus neutralising antibodies that confer protection against AHSV upon passive immunisation.  

PubMed

In previous studies we showed that a recombinant Modified Vaccinia Ankara (MVA) virus expressing the protein VP2 of AHSV serotype 4 (MVA-VP2) induced virus neutralising antibodies in horses and protected interferon alpha receptor gene knock-out mice (IFNAR-/-) against challenge. We continued these studies and determined, in the IFNAR-/- mouse model, whether the antibody responses induced by MVA-VP2 vaccination play a key role in protection against AHSV. Thus, groups of mice were vaccinated with wild type MVA (MVA-wt) or MVA-VP2 and the antisera from these mice were used in a passive immunisation experiment. Donor antisera from (a) MVA-wt; (b) MVA-VP2 vaccinated; or (c) MVA-VP2 vaccinated and AHSV infected mice, were transferred to AHSV non-immune recipient mice. The recipients were challenged with virulent AHSV together with MVA-VP2 vaccinated and MVA-wt vaccinated control animals and the levels of protection against AHSV-4 were compared between all these groups. The results showed that following AHSV challenge, mice that were passively immunised with MVA-VP2 vaccinated antisera were highly protected against AHSV disease and had lower levels of viraemia than recipients of MVA-wt antisera. Our study indicates that MVA-VP2 vaccination induces a highly protective humoral immune response against AHSV. PMID:24333835

Calvo-Pinilla, Eva; de la Poza, Francisco; Gubbins, Simon; Mertens, Peter Paul Clement; Ortego, Javier; Castillo-Olivares, Javier

2014-02-13

73

Vaccination with recombinant vaccinia viruses expressing ICP27 induces protective immunity against herpes simplex virus through CD4+ Th1+ T cells.  

PubMed Central

This study was designed to evaluate the efficacy and mechanisms of protection mediated by recombinant vaccinia viruses encoding immediate-early (IE) proteins of herpes simplex virus type 2 (HSV-2). Three mouse strains were immunized against the IE proteins ICP27, ICP0, and ICP4, and mice were challenged intracutaneously in the zosteriform model with HSV-2 strain MS. Protection was observed only following immunization with the ICP27 construct and then only in the BALB/c mouse strain. Protection in BALB/c mice was ablated by CD4+ T-cell suppression but remained intact in animals depleted of CD8+ T cells. Moreover, protection could be afforded to SCID nude recipients with CD4+ but not CD8+ T cells from ICP27-immunized mice. Only BALB/c mice developed a delayed-type hypersensitivity reaction to HSV-2, and in vitro measurements of humoral and cell-mediated immunity revealed response patterns to ICP27 and HSV that differed between protected BALB/c and unprotected mouse strains. Accordingly, BALB/c responses showed antigen-induced cytokine profiles dominated by type 1 cytokines, whereas C57BL/6 and C3H/HeN mice generated cytokine responses mainly of the type 2 variety. Our results may indicate that protection against zosterification is mainly mediated by CD4+ T cells that express a type 1 cytokine profile and that protective vaccines against HSV which effectively induce such T-cell responses should be chosen.

Manickan, E; Francotte, M; Kuklin, N; Dewerchin, M; Molitor, C; Gheysen, D; Slaoui, M; Rouse, B T

1995-01-01

74

Chimpanzee/human mAbs to vaccinia virus B5 protein neutralize vaccinia and smallpox viruses and protect mice against vaccinia virus.  

PubMed

Chimpanzee Fabs against the B5 envelope glycoprotein of vaccinia virus were isolated and converted into complete mAbs with human gamma 1 heavy chain constant regions. The two mAbs (8AH8AL and 8AH7AL) displayed high binding affinities to B5 (Kd of 0.2 and 0.7 nM). The mAb 8AH8AL inhibited the spread of vaccinia virus as well as variola virus (the causative agent of smallpox) in vitro, protected mice from subsequent intranasal challenge with virulent vaccinia virus, protected mice when administered 2 days after challenge, and provided significantly greater protection than that afforded by a previously isolated rat anti-B5 mAb (19C2) or by vaccinia immune globulin. The mAb bound to a conformational epitope between amino acids 20 and 130 of B5. These chimpanzee/human anti-B5 mAbs may be useful in the prevention and treatment of vaccinia virus-induced complications of vaccination against smallpox and may also be effective in the immunoprophylaxis and immunotherapy of smallpox. PMID:16436502

Chen, Zhaochun; Earl, Patricia; Americo, Jeffrey; Damon, Inger; Smith, Scott K; Zhou, Yi-Hua; Yu, Fujuan; Sebrell, Andrew; Emerson, Suzanne; Cohen, Gary; Eisenberg, Roselyn J; Svitel, Juraj; Schuck, Peter; Satterfield, William; Moss, Bernard; Purcell, Robert

2006-02-01

75

Highly attenuated modified vaccinia virus Ankara (MVA) as an effective recombinant vector: a Murine tumor model  

Microsoft Academic Search

Modified vaccinia virus Ankara (MVA), a highly attenuated strain of vaccinia virus (VV) that is unable to replicate in most mammalian cells, was evaluated as an expression vector for a model tumor associated antigen (TAA) and as a potential anti-cancer vaccine. We employed an experimental murine model in which an adenocarcinoma tumor line, CT26. CL25, was stably transfected with a

Miles W. Carroll; Willem W. Overwijk; Ronald S. Chamberlain; Steven A. Rosenberg; Bernard Moss; Nicholas P. Restifo

1997-01-01

76

Protective efficacy of Modified Vaccinia virus Ankara in preclinical studies.  

PubMed

Modified Vaccinia virus Ankara (MVA) is a tissue culture-derived, highly attenuated strain of vaccinia virus (VACV) exhibiting characteristic defective replication in cells from mammalian hosts. In the 1960s MVA was originally generated as a candidate virus for safer vaccination against smallpox. Now, MVA is widely used in experimental vaccine development targeting important infectious diseases and cancer. Versatile technologies for genetic engineering, large-scale production, and quality control facilitate R&D of recombinant and non-recombinant MVA vaccines matching today's requirements for new biomedical products. Such vaccines are attractive candidates for delivering antigens from pathogens against which no, or no effective vaccine is available, including emerging infections caused by highly pathogenic influenza viruses, chikungunya virus, West Nile virus or zoonotic orthopoxviruses. Other directions are seeking valuable vaccines against highly complex diseases such as AIDS, malaria, and tuberculosis. Here, we highlight examples of MVA candidate vaccines against infectious diseases, and review the efforts made to assess both the efficacy of vaccination and immune correlates of protection in preclinical studies. PMID:23523402

Volz, Asisa; Sutter, Gerd

2013-09-01

77

Attenuated and Replication-Competent Vaccinia Virus Strains M65 and M101 with Distinct Biology and Immunogenicity as Potential Vaccine Candidates against Pathogens  

PubMed Central

Replication-competent poxvirus vectors with an attenuation phenotype and with a high immunogenic capacity of the foreign expressed antigen are being pursued as novel vaccine vectors against different pathogens. In this investigation, we have examined the replication and immunogenic characteristics of two vaccinia virus (VACV) mutants, M65 and M101. These mutants were generated after 65 and 101 serial passages of persistently infected Friend erythroleukemia (FEL) cells. In cultured cells of different origins, the mutants are replication competent and have growth kinetics similar to or slightly reduced in comparison with those of the parental Western Reserve (WR) virus strain. In normal and immune-suppressed infected mice, the mutants showed different levels of attenuation and pathogenicity in comparison with WR and modified vaccinia Ankara (MVA) strains. Wide genome analysis after deep sequencing revealed selected genomic deletions and mutations in a number of viral open reading frames (ORFs). Mice immunized in a DNA prime/mutant boost regimen with viral vectors expressing the LACK (Leishmania homologue for receptors of activated C kinase) antigen of Leishmania infantum showed protection or a delay in the onset of cutaneous leishmaniasis. Protection was similar to that triggered by MVA-LACK. In immunized mice, both polyfunctional CD4+ and CD8+ T cells with an effector memory phenotype were activated by the two mutants, but the DNA-LACK/M65-LACK protocol preferentially induced CD4+ whereas DNA-LACK/M101-LACK preferentially induced CD8+ T cell responses. Altogether, our findings showed the adaptive changes of the WR genome during long-term virus-host cell interaction and how the replication competency of M65 and M101 mutants confers distinct biological properties and immunogenicity in mice compared to those of the MVA strain. These mutants could have applicability for understanding VACV biology and as potential vaccine vectors against pathogens and tumors.

Sanchez-Sampedro, Lucas; Gomez, Carmen Elena; Mejias-Perez, Ernesto; Perez-Jimenez, Eva; Oliveros, Juan Carlos

2013-01-01

78

Persistence of Vaccinia at the Site of Smallpox Vaccination.  

National Technical Information Service (NTIS)

Persistence of vaccinia at vaccination sites may help determine the risk associated with secondary transmission. Culture, PCR, and antigen detection were performed on serial vaccination site swab specimens. On day 21 after vaccination, 37% of volunteers w...

C. Hawkes G. V. Ludwig J. F. Cummings M. Klote M. E. Polhemus

2008-01-01

79

Highly Effective Control of an AIDS Virus Challenge in Macaques by Using Vesicular Stomatitis Virus and Modified Vaccinia Virus Ankara Vaccine Vectors in a Single-Boost Protocol  

Microsoft Academic Search

Previous studies have shown that vaccination and boosting of rhesus macaques with attenuated vesicular stomatitis virus (VSV) vectors encoding Env and Gag proteins of simian immunodeficiency virus-human immunodeficiency virus (SHIV) hybrid viruses protect rhesus macaques from AIDS after challenge with the highly pathogenic SHIV 89.6P (23). In the present study, we compared the effectiveness of a single prime-boost protocol consisting

Elizabeth Ramsburg; Nina F. Rose; Preston A. Marx; Megan Mefford; Douglas F. Nixon; Walter J. Moretto; David Montefiori; Patricia Earl; Bernard Moss; John K. Rose

2004-01-01

80

Identification of vaccinia virus epitope-specific HLA-A*0201-restricted T cells and comparative analysis of smallpox vaccines  

Microsoft Academic Search

Despite worldwide eradication of naturally occurring variola virus, smallpox remains a potential threat to both civilian and military populations. New, safe smallpox vaccines are being developed, and there is an urgent need for methods to evaluate vaccine efficacy after immunization. Here we report the identification of an immunodominant HLA-A*0201-restricted epitope that is recognized by cytotoxic CD8+ T cells and conserved

Ingo Drexler; Caroline Staib; Wolfgang Kastenmüller; Stefan Stevanovi; Burkhard Schmidt; François A. Lemonnier; Hans-Georg Rammensee; Dirk H. Busch; Helga Bernhard; Volker Erfle; Gerd Sutter

2003-01-01

81

Vaccinia Virus Infection in Monkeys, Brazilian Amazon  

PubMed Central

To detect orthopoxvirus in the Brazilian Amazon, we conducted a serosurvey of 344 wild animals. Neutralizing antibodies against orthopoxvirus were detected by plaque-reduction neutralizing tests in 84 serum samples. Amplicons from 6 monkey samples were sequenced. These amplicons identified vaccinia virus genetically similar to strains from bovine vaccinia outbreaks in Brazil.

Abrahao, Jonatas S.; Silva-Fernandes, Andre T.; Lima, Larissa S.; Campos, Rafael K.; Guedes, Maria I.M.C.; Cota, Marcela M.G.; Assis, Felipe L.; Borges, Iara A.; Souza-Junior, Milton F.; Lobato, Zelia I.P.; Bonjardim, Claudio A.; Ferreira, Paulo C.P.; Trindade, Giliane S.

2010-01-01

82

A Modified Vaccinia Ankara Virus (MVA) Vaccine Expressing African Horse Sickness Virus (AHSV) VP2 Protects Against AHSV Challenge in an IFNAR -/- Mouse Model  

PubMed Central

African horse sickness (AHS) is a lethal viral disease of equids, which is transmitted by Culicoides midges that become infected after biting a viraemic host. The use of live attenuated vaccines has been vital for the control of this disease in endemic regions. However, there are safety concerns over their use in non-endemic countries. Research efforts over the last two decades have therefore focused on developing alternative vaccines based on recombinant baculovirus or live viral vectors expressing structural components of the AHS virion. However, ethical and financial considerations, relating to the use of infected horses in high biosecurity installations, have made progress very slow. We have therefore assessed the potential of an experimental mouse-model for AHSV infection for vaccine and immunology research. We initially characterised AHSV infection in this model, then tested the protective efficacy of a recombinant vaccine based on modified vaccinia Ankara expressing AHS-4 VP2 (MVA-VP2).

Castillo-Olivares, Javier; Calvo-Pinilla, Eva; Casanova, Isabel; Bachanek-Bankowska, Katarzyna; Chiam, Rachael; Maan, Sushila; Nieto, Jose Maria; Ortego, Javier; Mertens, Peter Paul Clement

2011-01-01

83

Rapid spreading and immune evasion by vaccinia virus.  

PubMed

Vaccinia virus (VACV) is the live vaccine that was used to eradicate smallpox, a feat achieved in 1977 and certified by the World Health Organization in 1980. Since 1980, research with VACV has continued in part because of the development of techniques to genetically manipulate VACV and create live VACV strains expressing foreign genes. These recombinant VACVs can be used as live vaccines against other infectious diseases and cancers, and as a powerful tool to study virus pathogenesis, immunology, cell biology, and virus-host interactions. This short article describes two examples of how enduring interest in VACV has revealed new features of VACV biology and the immune system. PMID:24595611

Smith, Geoffrey L

2014-01-01

84

A Consecutive Priming-Boosting Vaccination of Mice with Simian Immunodeficiency Virus (SIV) gag/pol DNA and Recombinant Vaccinia Virus Strain DIs Elicits Effective Anti-SIV Immunity  

PubMed Central

To evaluate immunity induced by a novel DNA prime-boost regimen, we constructed a DNA plasmid encoding the gag and pol genes from simian immunodeficiency virus (SIV) (SIVgag/pol DNA), in addition to a replication-deficient vaccinia virus strain DIs recombinant expressing SIV gag and pol genes (rDIsSIVgag/pol). In mice, priming with SIVgag/pol DNA, followed by rDIsSIVgag/pol induced an SIV-specific lymphoproliferative response that was mediated by a CD4+-T-lymphocyte subset. Immunization with either vaccine alone was insufficient to induce high levels of proliferation or Th1 responses in the animals. The prime-boost regimen also induced SIV Gag-specific cellular responses based on gamma interferon secretion, as well as cytotoxic-T-lymphocyte responses. Thus, the regimen of DNA priming and recombinant DIs boosting induced Th1-type cell-mediated immunity, which was associated with resistance to viral challenge with wild-type vaccinia virus expressing SIVgag/pol, suggesting that this new regimen may hold promise as a safe and effective vaccine against human immunodeficiency virus type 1.

Someya, Kenji; Xin, Ke-Qin; Matsuo, Kazuhiro; Okuda, Kenji; Yamamoto, Naoki; Honda, Mitsuo

2004-01-01

85

A Phase 1 study to evaluate the safety and immunogenicity of a recombinant HIV type 1 subtype C-modified vaccinia Ankara virus vaccine candidate in Indian volunteers.  

PubMed

A recombinant modified vaccinia Ankara virus vaccine candidate (TBC-M4) expressing HIV-1 subtype C env, gag, tat-rev, and nef-RT genes was tested in a randomized, double-blind, dose escalation Phase I trial in 32 HIV-uninfected healthy volunteers who received three intramuscular injections of TBC-M4 at 0, 1, and 6 months of 5 x 10(7) plaque-forming units (pfu) (low dosage, LD) (n = 12) or 2.5 x 10(8) pfu (high dosage, HD) (n = 12) or placebo (n = 8). Local and systemic reactogenicity was experienced by approximately 67% and 83% of vaccine recipients, respectively. The reactogenicity events were mostly mild in severity. Severe but transient systemic reactogenicity was seen in one volunteer of the HD group. No vaccine-related serious adverse events or events suggesting perimyocarditis were seen. A higher frequency of local reactogenicity events was observed in the HD group. Cumulative HIV-specific IFN-gamma ELISPOT responses were detected in frozen PBMCs from 9/11 (82%), 12/12 (100%), and 1/8 (13%) volunteers after the third injection of the LD, HD, and placebo groups, respectively. Most of the responses were to gag and env proteins (maximum of 430 SFU/10(6) PBMCs) persisting across multiple time points. HIV-specific ELISA antibody responses were detected in 10/11, 12/12, and 0/8 volunteers post-third vaccination, in the LD, HD, and placebo groups, respectively. No neutralizing activity against HIV-1 subtype C isolates was detected. TBC-M4 appears to be generally safe and well-tolerated. The immune response detected was dose dependent, modest in magnitude, and directed mostly to env and gag proteins, suggesting further evaluation of this vaccine in a prime-boost regimen. PMID:19943789

Ramanathan, Vadakkuppatu Devasenapathi; Kumar, Makesh; Mahalingam, Jayashri; Sathyamoorthy, Pattabiraman; Narayanan, Paranji Ramaiyengar; Solomon, Suniti; Panicali, Dennis; Chakrabarty, Sekhar; Cox, Josephine; Sayeed, Eddy; Ackland, James; Verlinde, Carl; Vooijs, Dani; Loughran, Kelley; Barin, Burc; Lombardo, Angela; Gilmour, Jill; Stevens, Gwynneth; Smith, Michelle Seth; Tarragona-Fiol, Tony; Hayes, Peter; Kochhar, Sonali; Excler, Jean-Louis; Fast, Patricia

2009-11-01

86

Protection of Rhesus Monkeys from Fatal Lassa Fever by Vaccination with a Recombinant Vaccinia Virus Containing the Lassa Virus Glycoprotein Gene  

Microsoft Academic Search

Lassa fever is an acute febrile disease of West Africa, where there are as many as 300,000 infections a year and an estimated 3000 deaths. As control of the rodent host is impracticable at present, the best immediate prospect is vaccination. We tested as potential vaccines in rhesus monkeys a closely related virus, Mopeia virus (two monkeys), and a recombinant

S. P. Fisher-Hoch; J. B. McCormick; D. Auperin; B. G. Brown; M. Castor; G. Perez; S. Ruo; A. Conaty; L. Brammer; S. Bauer

1989-01-01

87

In vitro host range, multiplication and virion forms of recombinant viruses obtained from co-infection in vitro with a vaccinia-vectored influenza vaccine and a naturally occurring cowpox virus isolate  

PubMed Central

Background Poxvirus-vectored vaccines against infectious diseases and cancer are currently under development. We hypothesized that the extensive use of poxvirus-vectored vaccine in future might result in co-infection and recombination between the vaccine virus and naturally occurring poxviruses, resulting in hybrid viruses with unpredictable characteristics. Previously, we confirmed that co-infecting in vitro a Modified vaccinia virus Ankara (MVA) strain engineered to express influenza virus haemagglutinin (HA) and nucleoprotein (NP) genes with a naturally occurring cowpox virus (CPXV-NOH1) resulted in recombinant progeny viruses (H Hansen, MI Okeke, Ø Nilssen, T Traavik, Vaccine 23: 499–506, 2004). In this study we analyzed the biological properties of parental and progeny hybrid viruses. Results Five CPXV/MVA progeny viruses were isolated based on plaque phenotype and the expression of influenza virus HA protein. Progeny hybrid viruses displayed in vitro cell line tropism of CPXV-NOH1, but not that of MVA. The HA transgene or its expression was lost on serial passage of transgenic viruses and the speed at which HA expression was lost varied with cell lines. The HA transgene in the progeny viruses or its expression was stable in African Green Monkey derived Vero cells but became unstable in rat derived IEC-6 cells. Hybrid viruses lacking the HA transgene have higher levels of virus multiplication in mammalian cell lines and produced more enveloped virions than the transgene positive progenitor virus strain. Analysis of the subcellular localization of the transgenic HA protein showed that neither virus strain nor cell line have effect on the subcellular targets of the HA protein. The influenza virus HA protein was targeted to enveloped virions, plasma membrane, Golgi apparatus and cytoplasmic vesicles. Conclusion Our results suggest that homologous recombination between poxvirus-vectored vaccine and naturally circulating poxviruses, genetic instability of the transgene, accumulation of non-transgene expressing vectors or hybrid virus progenies, as well as cell line/type specific selection against the transgene are potential complications that may result if poxvirus vectored vaccines are extensively used in animals and man.

Okeke, Malachy Ifeanyi; Nilssen, ?ivind; Moens, Ugo; Tryland, Morten; Traavik, Terje

2009-01-01

88

A protein-based smallpox vaccine protects mice from vaccinia and ectromelia virus challenges when given as a prime and single boost  

Microsoft Academic Search

The heightened concern about the intentional release of variola virus has led to the need to develop safer smallpox vaccines. While subunit vaccine strategies are safer than live virus vaccines, subunit vaccines have been hampered by the need for multiple boosts to confer optimal protection. Here we developed a protein-based subunit vaccine strategy that provides rapid protection in mouse models

Yuhong Xiao; Lydia Aldaz-Carroll; Alexandra M. Ortiz; J. Charles Whitbeck; Edward Alexander; Huan Lou; Heather L. Davis; Thomas J. Braciale; Roselyn J. Eisenberg; Gary H. Cohen; Stuart N. Isaacs

2007-01-01

89

Reduction of Simian-Human Immunodeficiency Virus 89.6P Viremia in Rhesus Monkeys by Recombinant Modified Vaccinia Virus Ankara Vaccination  

Microsoft Academic Search

Since cytotoxic T lymphocytes (CTLs) are critical for controlling human immunodeficiency virus type 1 (HIV-1) replication in infected individuals, candidate HIV-1 vaccines should elicit virus-specific CTL re- sponses. In this report, we study the immune responses elicited in rhesus monkeys by a recombinant poxvirus vaccine and the degree of protection afforded against a pathogenic simian-human immunodeficiency virus SHIV-89.6P challenge. Immunization

DAN H. BAROUCH; SAMPA SANTRA; MARCELO J. KURODA; JORN E. SCHMITZ; RONALD PLISHKA; ALICIA BUCKLER-WHITE; ALICIA E. GAITAN; REBEKAH ZIN; JAE-HWAN NAM; LINDA S. WYATT; MICHELLE A. LIFTON; CHRISTINE E. NICKERSON; BERNARD MOSS; DAVID C. MONTEFIORI; VANESSA M. HIRSCH; NORMAN L. LETVIN

2001-01-01

90

Photoinactivation of Vaccinia Virus with Rose Bengal  

Microsoft Academic Search

SUMMARY Photoinactivation of vaccinia virus was complete but slower with rose bengal than with methylene blue as the sensitizing dye. Photoinactivation was inhibited and changes in dye absorption spectra occurred when protein but not when nucleic acid was added to rose bengal + virus mixtures, with methylene blue+virus mixtures nucleic aid but not protein was inhibitory. Photo- inactivation with methylene

G. S. Turner; C. Kaplan

1968-01-01

91

Origin of the Vaccinia Virus Hemagglutinin 1  

PubMed Central

The relationship between the vaccinia virus hemagglutinin and hemadsorption was examined. Hemagglutinin synthesis was temporally related to the appearance of the hemadsorption reaction. Only chicken erythrocytes, which reacted with hemagglutinin, hemadsorbed to infected cells, and both of these reactions were inhibited by Ca2+. The distribution of the vaccinia hemagglutinin and 5?-adenosine monophosphatase, a plasma membrane marker enzyme, in sucrose gradients was similar. Plasma membrane ghosts derived from infected cells hemadsorbed erythrocytes and yielded hemagglutinin upon sonic disruption. These data suggest that the majority of vaccinia hemagglutinin is derived from the plasma-membrane of the infected cell.

Blackman, Kenneth E.; Bubel, H. Curt

1972-01-01

92

Modified Vaccinia Virus Ankara Immunization Protects against Lethal Challenge with Recombinant Vaccinia Virus Expressing Murine Interleukin-4  

PubMed Central

Recent events have raised concern over the use of pathogens, including variola virus, as biological weapons. Vaccination with Dryvax is associated with serious side effects and is contraindicated for many people, and the development of a safer effective smallpox vaccine is necessary. We evaluated an attenuated vaccinia virus, modified vaccinia virus Ankara (MVA), by use of a murine model to determine its efficacy against an intradermal (i.d.) or intranasal (i.n.) challenge with vaccinia virus (vSC8) or a recombinant vaccinia virus expressing murine interleukin-4 that exhibits enhanced virulence (vSC8-mIL4). After an i.d. challenge, 15 of 16 mice who were inoculated with phosphate-buffered saline developed lesions, one dose of intramuscularly administered MVA was partially protective (3 of 16 mice developed lesions), and the administration of two or three doses of MVA was completely protective (0 of 16 mice developed lesions). In unimmunized mice, an i.n. challenge with vSC8 caused a significant but self-limited illness, while vSC8-mIL4 resulted in lethal infections. Immunization with one or two doses of MVA prevented illness and reduced virus titers in mice who were challenged with either vSC8 or vSC8-mIL4. MVA induced a dose-related neutralizing antibody and vaccinia virus-specific CD8+-T-cell response. Mice immunized with MVA were fully protected from a low-dose vSC8-mIL4 challenge despite a depletion of CD4+ cells, CD8+ cells, or both T-cell subsets or an antibody deficiency. CD4+- or CD8+-T-cell depletion reduced the protection against a high-dose vSC8-mIL4 challenge, and the depletion of both T-cell subsets was associated with severe illness and higher vaccinia virus titers. Thus, MVA induces broad humoral and cellular immune responses that can independently protect against a molecularly modified lethal poxvirus challenge in mice. These data support the continued development of MVA as an alternative candidate vaccine for smallpox.

McCurdy, Lewis H.; Rutigliano, John A.; Johnson, Teresa R.; Chen, Man; Graham, Barney S.

2004-01-01

93

Deletion of Specific Immune-Modulatory Genes from Modified Vaccinia Virus Ankara-Based HIV Vaccines Engenders Improved Immunogenicity in Rhesus Macaques  

PubMed Central

Modified vaccinia virus Ankara (MVA) is a safe, attenuated orthopoxvirus that is being developed as a vaccine vector but has demonstrated limited immunogenicity in several early-phase clinical trials. Our objective was to rationally improve the immunogenicity of MVA-based HIV/AIDS vaccines via the targeted deletion of specific poxvirus immune-modulatory genes. Vaccines expressing codon-optimized HIV subtype C consensus Env and Gag antigens were generated from MVA vector backbones that (i) harbor simultaneous deletions of four viral immune-modulatory genes, encoding an interleukin-18 (IL-18) binding protein, an IL-1? receptor, a dominant negative Toll/IL-1 signaling adapter, and CC-chemokine binding protein (MVA?4-HIV); (ii) harbor a deletion of an additional (fifth) viral gene, encoding uracil-DNA glycosylase (MVA?5-HIV); or (iii) represent the parental MVA backbone as a control (MVA-HIV). We performed head-to-head comparisons of the cellular and humoral immune responses that were elicited by these vectors during homologous prime-boost immunization regimens utilizing either high-dose (2 × 108 PFU) or low-dose (1 × 107 PFU) intramuscular immunization of rhesus macaques. At all time points, a majority of the HIV-specific T cell responses, elicited by all vectors, were directed against Env, rather than Gag, determinants, as previously observed with other vector systems. Both modified vectors elicited up to 6-fold-higher frequencies of HIV-specific CD8 and CD4 T cell responses and up to 25-fold-higher titers of Env (gp120)-specific binding (nonneutralizing) antibody responses that were relatively transient in nature. While the correlates of protection against HIV infection remain incompletely defined, our results indicate that the rational deletion of specific genes from MVA vectors can positively alter their cellular and humoral immunogenicity profiles in nonhuman primates.

O'Mara, Leigh A.; Gangadhara, Sailaja; McQuoid, Monica; Zhang, Xiugen; Zheng, Rui; Gill, Kiran; Verma, Meena; Yu, Tianwei; Johnson, Brent; Li, Bing; Derdeyn, Cynthia A.; Ibegbu, Chris; Altman, John D.; Hunter, Eric; Feinberg, Mark B.

2012-01-01

94

Deletion of specific immune-modulatory genes from modified vaccinia virus Ankara-based HIV vaccines engenders improved immunogenicity in rhesus macaques.  

PubMed

Modified vaccinia virus Ankara (MVA) is a safe, attenuated orthopoxvirus that is being developed as a vaccine vector but has demonstrated limited immunogenicity in several early-phase clinical trials. Our objective was to rationally improve the immunogenicity of MVA-based HIV/AIDS vaccines via the targeted deletion of specific poxvirus immune-modulatory genes. Vaccines expressing codon-optimized HIV subtype C consensus Env and Gag antigens were generated from MVA vector backbones that (i) harbor simultaneous deletions of four viral immune-modulatory genes, encoding an interleukin-18 (IL-18) binding protein, an IL-1? receptor, a dominant negative Toll/IL-1 signaling adapter, and CC-chemokine binding protein (MVA?4-HIV); (ii) harbor a deletion of an additional (fifth) viral gene, encoding uracil-DNA glycosylase (MVA?5-HIV); or (iii) represent the parental MVA backbone as a control (MVA-HIV). We performed head-to-head comparisons of the cellular and humoral immune responses that were elicited by these vectors during homologous prime-boost immunization regimens utilizing either high-dose (2 × 10(8) PFU) or low-dose (1 × 10(7) PFU) intramuscular immunization of rhesus macaques. At all time points, a majority of the HIV-specific T cell responses, elicited by all vectors, were directed against Env, rather than Gag, determinants, as previously observed with other vector systems. Both modified vectors elicited up to 6-fold-higher frequencies of HIV-specific CD8 and CD4 T cell responses and up to 25-fold-higher titers of Env (gp120)-specific binding (nonneutralizing) antibody responses that were relatively transient in nature. While the correlates of protection against HIV infection remain incompletely defined, our results indicate that the rational deletion of specific genes from MVA vectors can positively alter their cellular and humoral immunogenicity profiles in nonhuman primates. PMID:22973033

Garber, David A; O'Mara, Leigh A; Gangadhara, Sailaja; McQuoid, Monica; Zhang, Xiugen; Zheng, Rui; Gill, Kiran; Verma, Meena; Yu, Tianwei; Johnson, Brent; Li, Bing; Derdeyn, Cynthia A; Ibegbu, Chris; Altman, John D; Hunter, Eric; Feinberg, Mark B

2012-12-01

95

Long-term protective immunity to rinderpest in cattle following a single vaccination with a recombinant vaccinia virus expressing the virus haemagglutinin protein  

Microsoft Academic Search

A recombinant vaccine, produced by using a highly attenuated smallpox vaccine (LC16mO) as a vector and which expresses the rinderpest virus (RPV) haemagglutinin protein, has been developed. The properties of this vaccine, including its heat stability, efficacy in short-term trials, safety and genetic stability, have been confirmed in an earlier report. In the present study, the duration of the protective

Kazue Ohishi; Kenjiro Inui; Thomas Barrett; Kazuya Yamanouchi

96

Environmental persistence of vaccinia virus on materials.  

PubMed

Smallpox is caused by the variola virus, and ranks as one of the most serious diseases that could originate from a biological weapon. However, limited data exist on the persistence of variola and related viruses on materials (that may act as fomites), under controlled environmental conditions. To fill these data gaps, we determined the persistence of the vaccinia virus (an established surrogate for the variola virus) as a function of temperature, relative humidity and material. Experiments were conducted with vaccinia virus in a freeze-dried form, using four materials under four sets of environmental conditions. After elapsed times ranging from 1 to 56 days, the virus was extracted from small coupons and quantified via plaque-forming units (PFU). The vaccinia virus was most persistent at low temperature and low relative humidity, with greater than 10(4) PFU recovered from glass, galvanized steel and painted cinder block at 56 days (equivalent to only a c. 2 log reduction). Thus, vaccinia virus may persist from weeks to months, depending on the material and environmental conditions. This study may aid those responsible for infection control to make informed decisions regarding the need for environmental decontamination following the release of an agent such as variola. PMID:23815079

Wood, J P; Choi, Y W; Wendling, M Q; Rogers, J V; Chappie, D J

2013-11-01

97

Characterization of chimpanzee/human monoclonal antibodies to vaccinia virus A33 glycoprotein and its variola virus homolog in vitro and in a vaccinia virus mouse protection model.  

PubMed

Three distinct chimpanzee Fabs against the A33 envelope glycoprotein of vaccinia virus were isolated and converted into complete monoclonal antibodies (MAbs) with human gamma 1 heavy-chain constant regions. The three MAbs (6C, 12C, and 12F) displayed high binding affinities to A33 (K(d) of 0.14 nM to 20 nM) and may recognize the same epitope, which was determined to be conformational and located within amino acid residues 99 to 185 at the C terminus of A33. One or more of the MAbs were shown to reduce the spread of vaccinia virus as well as variola virus (the causative agent of smallpox) in vitro and to more effectively protect mice when administered before or 2 days after intranasal challenge with virulent vaccinia virus than a previously isolated mouse anti-A33 MAb (1G10) or vaccinia virus immunoglobulin. The protective efficacy afforded by anti-A33 MAb was comparable to that of a previously isolated chimpanzee/human anti-B5 MAb. The combination of anti-A33 MAb and anti-B5 MAb did not synergize the protective efficacy. These chimpanzee/human anti-A33 MAbs may be useful in the prevention and treatment of vaccinia virus-induced complications of vaccination against smallpox and may also be effective in the immunoprophylaxis and immunotherapy of smallpox and other orthopoxvirus diseases. PMID:17581986

Chen, Zhaochun; Earl, Patricia; Americo, Jeffrey; Damon, Inger; Smith, Scott K; Yu, Fujuan; Sebrell, Andrew; Emerson, Suzanne; Cohen, Gary; Eisenberg, Roselyn J; Gorshkova, Inna; Schuck, Peter; Satterfield, William; Moss, Bernard; Purcell, Robert

2007-09-01

98

Protection of mice and swine from pseudorabies virus conferred by vaccinia virus-based recombinants.  

PubMed Central

Glycoproteins gp50, gII, and gIII of pseudorabies virus (PRV) were expressed either individually or in combination by vaccinia virus recombinants. In vitro analysis by immunoprecipitation and immunofluorescence demonstrated the expression of a gII protein of approximately 120 kDa that was proteolytically processed to the gIIb (67- to 74-kDa) and gIIc (58-kDa) mature protein species similar to those observed in PRV-infected cells. Additionally, the proper expression of the 90-kDa gIII and 50-kDa gp50 was observed. All three of these PRV-derived glycoproteins were detectable on the surface of vaccinia virus-PRV recombinant-infected cells. In vivo, mice were protected against a virulent PRV challenge after immunization with the PRV glycoprotein-expressing vaccinia virus recombinants. The coexpression of gII and gIII by a single vaccinia virus recombinant resulted in a significantly reduced vaccination dose required to protect mice against PRV challenge. Inoculation of piglets with the various vaccinia virus-PRV glycoprotein recombinants also resulted in protection against virulent PRV challenge as measured by weight gain. The simultaneous expression of gII and gp50 in swine resulted in a significantly enhanced level of protection as evaluated by weight evolution following challenge with live PRV. Images

Riviere, M; Tartaglia, J; Perkus, M E; Norton, E K; Bongermino, C M; Lacoste, F; Duret, C; Desmettre, P; Paoletti, E

1992-01-01

99

Single Radial Immunodiffusion Test for Detecting Antibodies Against Surface Antigens of Intracellular and Extracellular Vaccinia Virus  

Microsoft Academic Search

SUMMARY Antibodies to surface antigens of intracellular naked vaccinia virus (INV) and in limited studies extracellular enveloped virus (EEV) were determined by single radial immunodiffusion tests (SRDT) with immobilized virions in agarose gels. Antibodies to INV were demonstrable in rabbit hyperimmune sera (one to four visible zones), smallpox convalescent sera and sera from re-vaccinated individuals. A difference in specificity of

V. J. Prakash; E. Norrby; L. Payne

1977-01-01

100

Novel Recombinant Mycobacterium bovis BCG, Ovine Atadenovirus, and Modified Vaccinia Virus Ankara Vaccines Combine To Induce Robust Human Immunodeficiency Virus-Specific CD4 and CD8 T-Cell Responses in Rhesus Macaques?  

PubMed Central

Mycobacterium bovis bacillus Calmette-Guérin (BCG), which elicits a degree of protective immunity against tuberculosis, is the most widely used vaccine in the world. Due to its persistence and immunogenicity, BCG has been proposed as a vector for vaccines against other infections, including HIV-1. BCG has a very good safety record, although it can cause disseminated disease in immunocompromised individuals. Here, we constructed a recombinant BCG vector expressing HIV-1 clade A-derived immunogen HIVA using the recently described safer and more immunogenic BCG strain AERAS-401 as the parental mycobacterium. Using routine ex vivo T-cell assays, BCG.HIVA401 as a stand-alone vaccine induced undetectable and weak CD8 T-cell responses in BALB/c mice and rhesus macaques, respectively. However, when BCG.HIVA401 was used as a priming component in heterologous vaccination regimens together with recombinant modified vaccinia virus Ankara-vectored MVA.HIVA and ovine atadenovirus-vectored OAdV.HIVA vaccines, robust HIV-1-specific T-cell responses were elicited. These high-frequency T-cell responses were broadly directed and capable of proliferation in response to recall antigen. Furthermore, multiple antigen-specific T-cell clonotypes were efficiently recruited into the memory pool. These desirable features are thought to be associated with good control of HIV-1 infection. In addition, strong and persistent T-cell responses specific for the BCG-derived purified protein derivative (PPD) antigen were induced. This work is the first demonstration of immunogenicity for two novel vaccine vectors and the corresponding candidate HIV-1 vaccines BCG.HIVA401 and OAdV.HIVA in nonhuman primates. These results strongly support their further exploration.

Rosario, Maximillian; Hopkins, Richard; Fulkerson, John; Borthwick, Nicola; Quigley, Maire F.; Joseph, Joan; Douek, Daniel C.; Greenaway, Hui Yee; Venturi, Vanessa; Gostick, Emma; Price, David A.; Both, Gerald W.; Sadoff, Jerald C.; Hanke, Tomas

2010-01-01

101

Replication of Modified Vaccinia Virus Ankara in Primary Chicken Embryo Fibroblasts Requires Expression of the Interferon Resistance Gene E3L  

Microsoft Academic Search

Highly attenuated modified vaccinia virus Ankara (MVA) serves as a candidate vaccine to immunize against infectious diseases and cancer. MVA was randomly obtained by serial growth in cultures of chicken embryo fibroblasts (CEF), resulting in the loss of substantial genomic information including many genes regulating virus-host interactions. The vaccinia virus interferon (IFN) resistance gene E3L is among the few conserved

Simone Hornemann; Olof Harlin; Caroline Staib; Sigrid Kisling; Volker Erfle; Bernd Kaspers; Georg Hacker; Gerd Sutter

2003-01-01

102

Interleukin12 (IL12) Enhancement of the Cellular Immune Response against Human Immunodeficiency Virus Type 1 Env Antigen in a DNA Prime\\/Vaccinia Virus Boost Vaccine Regimen Is Time and Dose Dependent: Suppressive Effects of IL12 Boost Are Mediated by Nitric Oxide  

Microsoft Academic Search

We previously demonstrated that codelivery of interleukin-12 (IL-12) with the human immunodeficiency virus type 1 (HIV-1) Env antigen from a recombinant vaccinia virus (rVV) can enhance the specific anti-Env cell-mediated immune (CMI) response. In the present study, we have investigated the effects of IL-12 in mice when it is expressed in a DNA prime\\/VV boost vaccine regimen. The delivery of

M. MAGDALENA GHERARDI; JUAN C. RAMIREZ; MARIANO ESTEBAN

2000-01-01

103

Immunization with Modified Vaccinia Virus Ankara-Based Recombinant Vaccine against Severe Acute Respiratory Syndrome Is Associated with Enhanced Hepatitis in Ferrets  

Microsoft Academic Search

Severe acute respiratory syndrome (SARS) caused by a newly identified coronavirus (SARS-CoV) is a serious emerging human infectious disease. In this report, we immunized ferrets (Mustela putorius furo) with recom- binant modified vaccinia virus Ankara (rMVA) expressing the SARS-CoV spike (S) protein. Immunized ferrets developed a more rapid and vigorous neutralizing antibody response than control animals after challenge with SARS-CoV;

Hana Weingartl; Markus Czub; Stefanie Czub; James Neufeld; Peter Marszal; Jason Gren; Greg Smith; Shane Jones; Roxanne Proulx; Yvonne Deschambault; Elsie Grudeski; Anton Andonov; Runtao He; Yan Li; John Copps; Allen Grolla; Daryl Dick; Jody Berry; Shelley Ganske; Lisa Manning; Jingxin Cao

2004-01-01

104

Different contribution of co-stimulatory molecules B7.1 and B7.2 to the immune response to recombinant modified vaccinia virus ankara vaccine expressing prM/E proteins of Japanese encephalitis virus and two hepatitis B virus vaccines.  

PubMed

This study clarifies the role of co-stimulatory molecules B7.1 and B7.2 in the immune response to 3 types of vaccines: a/ recombinant modified Vaccinia virus Ankara (MVA) (vJH9) expressing prM/E proteins of Japanese encephalitis virus (JEV), b/ recombinant yeast-expressed Hepatitis B virus (YHBV), c/ human plasma-derived Hepatitis B virus (PHBV). We constructed plasmids expressing B7.1 and B7.2 molecules and found that the expression level of B7.2 protein in transfected CHO-k1 cells was higher than that of B7.1 protein. Mice were co-injected with vaccines vJH9, YHBV and PHBV and plasmids expressing B7.1 or B7.2, respectively, and specific antibody titers for each vaccine were monitored at days 7, 14 and 28 post injection (p.i.). In mice injected with vJH9 vaccine and both B7 plasmids, plasmid B7.2 induced a higher anti-JEV immune response than plasmid B7.1. This implies that the stimulation of the B7.2 immune pathway may be a feasible method of boosting protective immunity against a recombinant viral vaccine. Both B7 molecules were able to induce a specific anti-HBV immune response using YHBV vaccine. On the other hand, B7 molecules had little effect to the specific antibody induction in PHBV vaccination. These results suggested that the contribution of B7.1 and B7.2 molecules in an immune response depended on the character and status of the presenting antigen. PMID:17900219

NAM, J H; BANG, H S; CHO, H W; CHUNG, Y H

2007-01-01

105

Immunological Memory after Exposure to Variola Virus, Monkeypox Virus, and Vaccinia Virus  

PubMed Central

We compared cellular and humoral immunity to vaccinia virus (VV) in individuals exposed to 3 different orthopoxviruses: 154 individuals previously vaccinated with VV, 7 individuals with a history of monkeypox virus infection, and 8 individuals with a history of variola virus infection. Among individuals vaccinated >20 years prior, 9 (14%) of 66 individuals demonstrated VV-specific interferon (IFN)-? enzyme-linked immunospot (ELISPOT) assay responses; 21 (50%) of 42 had lymphoproliferative (LP) responses, and 29 (97%) of 30 had VV-specific neutralizing antibodies. One year after monkeypox virus infection, 6 of 7 individuals had IFN-? ELISPOT responses, all had VV-specific LP responses, and 3 of 7 had VV-specific neutralizing antibodies. Of 8 individuals with a history of variola virus infection, 1 had a VV-specific IFN-? ELISPOT response, 4 had LP responses against whole VV, 7 had LP responses against heat-denatured vaccinia antigen, and 7 had VV-specific neutralizing antibodies. Survivors of variola virus infection demonstrated VV-specific CD4 memory cell responses and neutralizing antibodies >40 years after infection.

Sivapalasingam, Sumathi; Kennedy, Jeffrey S.; Borkowsky, William; Valentine, Fred; Zhan, Ming-Xia; Pazoles, Pamela; Paolino, Anna; Ennis, Francis A.; Steigbigel, Neal H.

2007-01-01

106

Safety and Tolerability of Conserved Region Vaccines Vectored by Plasmid DNA, Simian Adenovirus and Modified Vaccinia Virus Ankara Administered to Human Immunodeficiency Virus Type 1-Uninfected Adults in a Randomized, Single-Blind Phase I Trial  

PubMed Central

Trial Design HIV-1 vaccine development has advanced slowly due to viral antigenic diversity, poor immunogenicity and recently, safety concerns associated with human adenovirus serotype-5 vectors. To tackle HIV-1 variation, we designed a unique T-cell immunogen HIVconsv from functionally conserved regions of the HIV-1 proteome, which were presented to the immune system using a heterologous prime-boost combination of plasmid DNA, a non-replicating simian (chimpanzee) adenovirus ChAdV-63 and a non-replicating poxvirus, modified vaccinia virus Ankara. A block-randomized, single-blind, placebo-controlled phase I trial HIV-CORE 002 administered for the first time candidate HIV-1- vaccines or placebo to 32 healthy HIV-1/2-uninfected adults in Oxford, UK and elicited high frequencies of HIV-1-specific T cells capable of inhibiting HIV-1 replication in vitro. Here, detail safety and tolerability of these vaccines are reported. Methods Local and systemic reactogenicity data were collected using structured interviews and study-specific diary cards. Data on all other adverse events were collected using open questions. Serum neutralizing antibody titres to ChAdV-63 were determined before and after vaccination. Results Two volunteers withdrew for vaccine-unrelated reasons. No vaccine-related serious adverse events or reactions occurred during 190 person-months of follow-up. Local and systemic events after vaccination occurred in 27/32 individuals and most were mild (severity grade 1) and predominantly transient (<48 hours). Myalgia and flu-like symptoms were more strongly associated with MVA than ChAdV63 or DNA vectors and more common in vaccine recipients than in placebo. There were no intercurrent HIV-1 infections during follow-up. 2/24 volunteers had low ChAdV-63-neutralizing titres at baseline and 7 increased their titres to over 200 with a median (range) of 633 (231-1533) post-vaccination, which is of no safety concern. Conclusions These data demonstrate safety and good tolerability of the pSG2.HIVconsv DNA, ChAdV63.HIVconsv and MVA.HIVconsv vaccines and together with their high immunogenicity support their further development towards efficacy studies. Trial Registration ClinicalTrials.gov NCT01151319

Hayton, Emma-Jo; Rose, Annie; Ibrahimsa, Umar; Del Sorbo, Mariarosaria; Capone, Stefania; Crook, Alison; Black, Antony P.; Dorrell, Lucy; Hanke, Tomas

2014-01-01

107

Susceptibility of Vaccinia Virus to Chemical Disinfectants  

PubMed Central

Vaccinia virus (VACV) is the cause of bovine vaccinia (BV), an emerging zoonotic disease that affects dairy cows and milkers. Some chemical disinfectants have been used on farms affected by BV to disinfect cow teats and milkers' hands. To date, there is no information about the efficacy of disinfectants against VACV. Therefore, this study aimed to assess the virucidal activity of some active disinfectants commonly used in the field. Sodium hypochlorite, quaternary ammonium combined with chlorhexidine, and quaternary ammonium combined with glutaraldehyde were effective in inactivating the virus at all concentrations tested. Iodine and quaternary ammonium as the only active component were partially effective. The presence of bovine feces as organic matter and light decreased the effectiveness of sodium hypochlorite. These results show that an appropriated disinfection and asepsis of teats and hands may be helpful in the control and prevention of BV and other infections with VACV.

de Oliveira, Tercia Moreira Ludolfo; Rehfeld, Izabelle Silva; Coelho Guedes, Maria Isabel Maldonado; Ferreira, Jaqueline Maria Siqueira; Kroon, Erna Geessien; Lobato, Zelia Ines Portela

2011-01-01

108

Can vaccinia virus be replaced by MVA virus for testing virucidal activity of chemical disinfectants?  

PubMed Central

Background Vaccinia virus strain Lister Elstree (VACV) is a test virus in the DVV/RKI guidelines as representative of the stable enveloped viruses. Since the potential risk of laboratory-acquired infections with VACV persists and since the adverse effects of vaccination with VACV are described, the replacement of VACV by the modified vaccinia Ankara strain (MVA) was studied by testing the activity of different chemical biocides in three German laboratories. Methods The inactivating properties of different chemical biocides (peracetic acid, aldehydes and alcohols) were tested in a quantitative suspension test according to the DVV/RKI guideline. All tests were performed with a protein load of 10% fetal calf serum with both viruses in parallel using different concentrations and contact times. Residual virus was determined by endpoint dilution method. Results The chemical biocides exhibited similar virucidal activity against VACV and MVA. In three cases intra-laboratory differences were determined between VACV and MVA - 40% (v/v) ethanol and 30% (v/v) isopropanol are more active against MVA, whereas MVA seems more stable than VACV when testing with 0.05% glutardialdehyde. Test accuracy across the three participating laboratories was high. Remarkably inter-laboratory differences in the reduction factor were only observed in two cases. Conclusions Our data provide valuable information for the replacement of VACV by MVA for testing chemical biocides and disinfectants. Because MVA does not replicate in humans this would eliminate the potential risk of inadvertent inoculation with vaccinia virus and disease in non-vaccinated laboratory workers.

2010-01-01

109

Applications of Pox Virus Vectors to Vaccination: An Update  

Microsoft Academic Search

Recombinant pox viruses have been generated for vaccination against heterologous pathogens. Amongst these, the following are notable examples. (i) The engineering of the Copenhagen strain of vaccinia virus to express the rabies virus glycoprotein. When applied in baits, this recombinant has been shown to vaccinate the red fox in Europe and raccoons in the United States, stemming the spread of

Enzo Paoletti

1996-01-01

110

Development of a novel, guinea pig-specific IFN-? ELISPOT assay and characterization of guinea pig cytomegalovirus GP83-specific cellular immune responses following immunization with a modified vaccinia virus Ankara (MVA)-vectored GP83 vaccine.  

PubMed

The guinea pig (Cavia porcellus) provides a useful animal model for studying the pathogenesis of many infectious diseases, and for preclinical evaluation of vaccines. However, guinea pig models are limited by the lack of immunological reagents required for characterization and quantification of antigen-specific T cell responses. To address this deficiency, an enzyme-linked immunospot (ELISPOT) assay for guinea pig interferon (IFN)-? was developed to measure antigen/epitope-specific T cell responses to guinea pig cytomegalovirus (GPCMV) vaccines. Using splenocytes harvested from animals vaccinated with a modified vaccinia virus Ankara (MVA) vector encoding the GPCMV GP83 (homolog of human CMV pp65 [gpUL83]) protein, we were able to enumerate and map antigen-specific responses, both in vaccinated as well as GPCMV-infected animals, using a panel of GP83-specific peptides. Several potential immunodominant GP83-specific peptides were identified, including one epitope, LGIVHFFDN, that was noted in all guinea pigs that had a detectable CD8+ response to GP83. Development of a guinea pig IFN-? ELISPOT should be useful in characterization of additional T cell-specific responses to GPCMV, as well as other pathogens. This information in turn can help focus future experimental evaluation of immunization strategies, both for GPCMV as well as for other vaccine-preventable illnesses studied in the guinea pig model. PMID:24856783

Gillis, Peter A; Hernandez-Alvarado, Nelmary; Gnanandarajah, Josephine S; Wussow, Felix; Diamond, Don J; Schleiss, Mark R

2014-06-30

111

Monkeypox: Experimental Infection in Chimpanzee (Pan Satyrus) and Immunization with Vaccinia Virus.  

National Technical Information Service (NTIS)

The efficacy of vaccinia viral vaccine in protecting subhuman primates against monkeypox was demonstrated. Chimpanzees given vaccinia viral (smallpox) vaccine were protected against challenge inoculation of virulent poxvirus isolated from an orangutan (Po...

S. McConnell R. L. Hickman W. L. Wooding D. L. Huxsoll

1967-01-01

112

Vaccinia Virus Recombinant Expressing Herpes Simplex Virus Type 1 Glycoprotein D Prevents Latent Herpes in Mice  

NASA Astrophysics Data System (ADS)

In humans, herpes simplex virus causes a primary infection and then often a latent ganglionic infection that persists for life. Because these latent infections can recur periodically, vaccines are needed that can protect against both primary and latent herpes simplex infections. Infectious vaccinia virus recombinants that contain the herpes simplex virus type 1 (HSV-1) glycoprotein D gene under control of defined early or late vaccinia virus promoters were constructed. Tissue culture cells infected with these recombinant viruses synthesized a glycosylated protein that had the same mass (60,000 daltons) as the glycoprotein D produced by HSV-1. Immunization of mice with one of these recombinant viruses by intradermal, subcutaneous, or intraperitoneal routes resulted in the production of antibodies that neutralized HSV-1 and protected the mice against subsequent lethal challenge with HSV-1 or HSV-2. Immunization with the recombinant virus also protected the majority of the mice against the development of a latent HSV-1 infection of the trigeminal ganglia. This is the first demonstration that a genetically engineered vaccine can prevent the development of latency.

Cremer, Kenneth J.; Mackett, Michael; Wohlenberg, Charles; Notkins, Abner Louis; Moss, Bernard

1985-05-01

113

Methodology for the Efficient Generation of Fluorescently Tagged Vaccinia Virus Proteins  

PubMed Central

Tagging of viral proteins with fluorescent proteins has proven an indispensable approach to furthering our understanding of virus-host interactions. Vaccinia virus (VACV), the live vaccine used in the eradication of smallpox, is particularly amenable to fluorescent live-cell microscopy owing to its large virion size and the ease with which it can be engineered at the genome level. We report here an optimized protocol for generating recombinant viruses. The minimal requirements for targeted homologous recombination during vaccinia replication were determined, which allows the simplification of construct generation. This enabled the alliance of transient dominant selection (TDS) with a fluorescent reporter and metabolic selection to provide a rapid and modular approach to fluorescently label viral proteins. By streamlining the generation of fluorescent recombinant viruses, we are able to facilitate downstream applications such as advanced imaging analysis of many aspects of the virus-host interplay that occurs during virus replication.

Lynn, Helena; Yamamoto, Yui; Horsington, Jacquelyn; Newsome, Timothy P.

2014-01-01

114

Methodology for the efficient generation of fluorescently tagged vaccinia virus proteins.  

PubMed

Tagging of viral proteins with fluorescent proteins has proven an indispensable approach to furthering our understanding of virus-host interactions. Vaccinia virus (VACV), the live vaccine used in the eradication of smallpox, is particularly amenable to fluorescent live-cell microscopy owing to its large virion size and the ease with which it can be engineered at the genome level. We report here an optimized protocol for generating recombinant viruses. The minimal requirements for targeted homologous recombination during vaccinia replication were determined, which allows the simplification of construct generation. This enabled the alliance of transient dominant selection (TDS) with a fluorescent reporter and metabolic selection to provide a rapid and modular approach to fluorescently label viral proteins. By streamlining the generation of fluorescent recombinant viruses, we are able to facilitate downstream applications such as advanced imaging analysis of many aspects of the virus-host interplay that occurs during virus replication. PMID:24473272

Marzook, N Bishara; Procter, Dean J; Lynn, Helena; Yamamoto, Yui; Horsington, Jacquelyn; Newsome, Timothy P

2014-01-01

115

A Differential Role for Macropinocytosis in Mediating Entry of the Two Forms of Vaccinia Virus into Dendritic Cells  

Microsoft Academic Search

Vaccinia virus (VACV) is being developed as a recombinant viral vaccine vector for several key pathogens. Dendritic cells (DCs) are specialised antigen presenting cells that are crucial for the initiation of primary immune responses; however, the mechanisms of uptake of VACV by these cells are unclear. Therefore we examined the binding and entry of both the intracellular mature virus (MV)

Kerrie J. Sandgren; John Wilkinson; Monica Miranda-Saksena; Gerald M. McInerney; Karen Byth-Wilson; Phillip J. Robinson; Anthony L. Cunningham

2010-01-01

116

Polypeptide composition of extracellular enveloped vaccinia virus.  

PubMed Central

Extracellular enveloped vaccinia (EEV) virus grown in SIRC and in HeLa cells was purified by consecutive equilibrium centrifugations in sucrose and cesium chloride gradients. A higher degree of purity was obtained with virus material prepared in SIRC cells. The polypeptides of purified EEV and INV (intracellular naked vaccinia) virus were compared in polyacrylamide slab gel electrophoresis. Three proteins (200,000 molecular weight [200K], 95K, and 13K) detected in HeLa-derived INV were absent in EEV. In addition, two INV proteins (65K and 30K) occurred in reduced concentrations in EEV, white another INV protein (27K) was increased in EEV. INV from SIRC cells showed similar alterations of these proteins (with the exception of the 30K and 13K proteins). Detergent treatment, ether extraction, and Pronase treatment showed that these six proteins are located at the surface of INV and are not cecessary for infectivity. Eight proteins (210K, 110K, 89K, 42K, 37K, 21.5K, 21K, and 20K) were detected in EEV that were absent from inv. Brij-58 treatment was employed to remove the envelope from EEV, resulting in the formation of naked particles and an envelope fraction which were separated on cesium chloride gradients. The envelope fractions contained all eight proteins. Seven of the eight proteins were glycoproteins, with the 37K protein being the only unglycosylated protein. It is concluded that a processing of surface INV particle proteins occurs during evelopment. The resultant EEV particle is comprised of an INV particle with a modified surface composition enclosed in an envelope containing virus-specific proteins unique to EEV. Images

Payne, L

1978-01-01

117

USE OF RECOMBINANT VACCINIA-RABIES GLYCOPROTEIN VIRUS FOR ORAL VACCINATION OF WILDLIFE AGAINST RABIES: INNOCUITY TO SEVERAL NONTARGET BAIT CONSUMING SPECIES  

Microsoft Academic Search

The pathiogenicity of a vaccimiia recombinant virus expressing the rabies glycoprotein (\\\\'VTGgRAB) was tested in several wild amiimal species which could compete with the natural rabies host, the red fox (Vulpes vulpes) ins comisuming vaccine baits in Europe. The following species were included in this study: wild boar (Sus scrofa), Eurasian badger (Meles meles), wood mouse (Apodenius sylvaticus), yellow-necked mouse

Bernard Brochier; Jean Blancou; Isabelle Thomas; Bernard Languet; Marc Artois; Marie-Paule Kieny; Jean-Pierre Lecocq; Philippe Desmettre; Paul-Pierre Pastoret

118

Quantitation of CD8 ? T Cell Responses to Newly Identified HLA-A * 0201-restricted T Cell Epitopes Conserved Among Vaccinia and Variola (Smallpox) Viruses  

Microsoft Academic Search

Immunization with vaccinia virus resulted in long-lasting protection against smallpox and was the approach used to eliminate natural smallpox infections worldwide. Due to the concern about the potential use of smallpox virus as a bioweapon, smallpox vaccination is currently be- ing reintroduced. Severe complications from vaccination were associated with congenital or acquired T cell deficiencies, but not with congenital agammaglobulinemia,

Masanori Terajima; John Cruz; Gregory Raines; Elizabeth D. Kilpatrick; Jeffrey S. Kennedy; Alan L. Rothman; Francis A. Ennis

119

A Novel High-Throughput Vaccinia Virus Neutralization Assay and Preexisting Immunity in Populations from Different Geographic Regions in China  

PubMed Central

Background Pre-existing immunity to Vaccinia Tian Tan virus (VTT) resulting from a large vaccination campaign against smallpox prior to the early 1980s in China, has been a major issue for application of VTT-vector based vaccines. It is essential to establish a sensitive and high-throughput neutralization assay to understand the epidemiology of Vaccinia-specific immunity in current populations in China. Methodology/Principal Findings A new anti-Vaccinia virus (VACV) neutralization assay that used the attenuated replication-competent VTT carrying the firefly luciferase gene of Photinus pyralis (rTV-Fluc) was established and standardized for critical parameters that included the choice of cell line, viral infection dose, and the infection time. The current study evaluated the maintenance of virus-specific immunity after smallpox vaccination by conducting a non-randomized, cross-sectional analysis of antiviral antibody-mediated immune responses in volunteers examined 30–55 years after vaccination. The rTV-Fluc neutralization assay was able to detect neutralizing antibodies (NAbs) against Vaccinia virus without the ability to differentiate strains of Vaccinia virus. We showed that the neutralizing titers measured by our assay were similar to those obtained by the traditional plaque reduction neutralization test (PRNT). Using this assay, we found a low prevalence of NAb to VTT (7.6%) in individuals born before 1980 from Beijing and Anhui provinces in China, and when present, anti-VTT NAb titers were low. No NAbs were detected in all 222 samples from individuals born after 1980. There was no significant difference observed for titer or prevalence by gender, age range and geographic origin. Conclusion A simplified, sensitive, standardized, reproducible, and high-throughput assay was developed for the quantitation of NAbs against different Vaccinia strains. The current study provides useful insights for the future development of VTT-based vaccination in Beijing and Anhui provinces of China.

Liu, Qiang; Huang, Weijin; Nie, Jianhui; Zhu, Rong; Gao, Dongying; Song, Aijing; Meng, Shufang; Xu, Xuemei; Wang, Youchun

2012-01-01

120

Pathogeneses of respiratory infections with virulent and attenuated vaccinia viruses  

Microsoft Academic Search

BACKGROUND: Respiratory infection with the neurovirulent vaccinia virus (VV) strain Western Reserve (WR) results in an acute infection of the lung followed by dissemination of the virus to other organs and causes lethality in mice. The mechanisms of lethality are not well-understood. In this study, we analyzed virus replication and host immune responses after intranasal infection with lethal and non-lethal

Daisuke Hayasaka; Francis A Ennis; Masanori Terajima

2007-01-01

121

Recombinant modified vaccinia virus Ankara expressing the hemagglutinin gene confers protection against homologous and heterologous H5N1 influenza virus infections in macaques  

Microsoft Academic Search

Background. Highly pathogenic avian influenza viruses of the H5N1 subtype have been responsible for an increasing number of infections in humans since 2003. More than 60% of infected individuals die, and new infections are reported frequently. In light of the pandemic threat caused by these events, the rapid availability of safe and effective vaccines is desirable. Modified vaccinia virus Ankara

Y. Suezer; G. de Mutsert; Brand van den J. M. A; G. van Amerongen; T. Kuiken; J. Löwer; G. Sutter

2009-01-01

122

Use of Vaccinia Virus in the Treatment of Metastatic Malignant Melanoma  

PubMed Central

Of 19 patients with proved metastases from malignant melanoma treated by inoculations of smallpox vaccine, intradermal deposits disappeared completely in six out of ten cases. Five of these remained well 2 to 22 months after initial treatment. The response was limited strictly to the site of inoculation. The mechanism of action of vaccinia virus in malignant melanoma is not clear. ImagesFig. 1Fig. 2Fig. 3Fig. 4Fig. 5Fig. 6

Hunter-Craig, I.; Newton, K. A.; Westbury, G.; Lacey, B. W.

1970-01-01

123

Use of a recombinant vaccinia virus expressing interferon gamma for post-exposure protection against vaccinia and ectromelia viruses.  

PubMed

Post-exposure vaccination with vaccinia virus (VACV) has been suggested to be effective in minimizing death if administered within four days of smallpox exposure. While there is anecdotal evidence for efficacy of post-exposure vaccination this has not been definitively studied in humans. In this study, we analyzed post-exposure prophylaxis using several attenuated recombinant VACV in a mouse model. A recombinant VACV expressing murine interferon gamma (IFN-?) was most effective for post-exposure protection of mice infected with VACV and ectromelia virus (ECTV). Untreated animals infected with VACV exhibited severe weight loss and morbidity leading to 100% mortality by 8 to 10 days post-infection. Animals treated one day post-infection had milder symptoms, decreased weight loss and morbidity, and 100% survival. Treatment on days 2 or 3 post-infection resulted in 40% and 20% survival, respectively. Similar results were seen in ECTV-infected mice. Despite the differences in survival rates in the VACV model, the viral load was similar in both treated and untreated mice while treated mice displayed a high level of IFN-? in the serum. These results suggest that protection provided by IFN-? expressed by VACV may be mediated by its immunoregulatory activities rather than its antiviral effects. These results highlight the importance of IFN-? as a modulator of the immune response for post-exposure prophylaxis and could be used potentially as another post-exposure prophylaxis tool to prevent morbidity following infection with smallpox and other orthopoxviruses. PMID:24147092

Holechek, Susan A; Denzler, Karen L; Heck, Michael C; Schriewer, Jill; Buller, R Mark; Legrand, Fatema A; Verardi, Paulo H; Jones, Leslie A; Yilma, Tilahun; Jacobs, Bertram L

2013-01-01

124

Real-time PCR assay to identify variants of Vaccinia virus: Implications for the diagnosis of bovine vaccinia in Brazil  

Microsoft Academic Search

Naturally occurring infections of Vaccinia virus (VACV) have been recognized in Brazil during the past 10 years. Human Brazilian Vaccinia virus (BVV) infections typically occur as a zoonosis transferred from affected dairy cows to their handlers. Outbreaks have caused notable economic losses to the rural community in the region. The origins of BVV are unclear but previous analyses have shown

Giliane de Souza Trindade; Yu Li; Victoria A. Olson; Ginny Emerson; Russell L. Regnery; Flavio Guimaraes da Fonseca; Erna Geessien Kroon; Inger Damon

2008-01-01

125

IMMUNE RESPONSE IN HUMANS AFTER VACCINATION WITH VACCINIA VIRUS: GENERATION OF A VIRUS-SPECIFIC CYTOTOXIC ACTIVITY BY HUMAN PERIPHERAL LYMPHOCYTES  

Microsoft Academic Search

The outcome of a virus infection mainly relates to the respective characteris- tics of the viruses, the cells they replicate in, and the immune system of the host (1-4). The immune response of man is usually analyzed in terms of humoral or cellular reactions. Much data have accumulated on humoral responses to viral infections; in fact, a rise in antibody

LUC H. PERRIN; ROLF M. ZINKERNAGEL; MICHAEL B. A. OLDSTONE

126

Middle East Respiratory Syndrome Coronavirus Spike Protein Delivered by Modified Vaccinia Virus Ankara Efficiently Induces Virus-Neutralizing Antibodies  

PubMed Central

Middle East respiratory syndrome coronavirus (MERS-CoV) has recently emerged as a causative agent of severe respiratory disease in humans. Here, we constructed recombinant modified vaccinia virus Ankara (MVA) expressing full-length MERS-CoV spike (S) protein (MVA-MERS-S). The genetic stability and growth characteristics of MVA-MERS-S make it a suitable candidate vaccine for clinical testing. Vaccinated mice produced high levels of serum antibodies neutralizing MERS-CoV. Thus, MVA-MERS-S may serve for further development of an emergency vaccine against MERS-CoV.

Song, Fei; Fux, Robert; Provacia, Lisette B.; Volz, Asisa; Eickmann, Markus; Becker, Stephan; Osterhaus, Albert D. M. E.; Haagmans, Bart L.

2013-01-01

127

Local delivery of recombinant vaccinia virus encoding for neu counteracts growth of mammary tumors more efficiently than systemic delivery in neu transgenic mice  

PubMed Central

Recombinant vaccinia virus has been widely employed as a cancer vaccine in several clinical trials. In this study we explored, employing BALB/c mice transgenic for the rat neu oncogene, the ability of the recombinant vaccinia virus neu (rV-neuT) vaccine to inhibit growth of neu+ mammary carcinomas and whether the efficacy of vaccination was dependent on: a) carcinogenesis stage at which the vaccination was initiated; b) number of vaccinations and c) route of delivery (systemic vs local). BALB-neuT mice were vaccinated one, two and three times by subcutaneous (s.c) and intramammary gland (im.g) injection with rV-neuT or V-wt (wild type vaccinia virus) starting at the stage in which mouse mammary gland displays atypical hyperplasia, carcinoma in situ or invasive carcinoma. We demonstrated that vaccination using rV-neuT was more effective when started at an earlier stage of mammary carcinogenesis and after three vaccinations. im.g vaccination was more effective than s.c vaccination in inhibiting mammary carcinogenesis, eliciting anti-Neu antibodies, increasing anti-Neu IgG2a/G3 isotypes and inducing antibodies able to trigger mammary tumor cell apoptosis and antibody dependent cellular cytotoxicity. The better protective ability of rV-neuT im.g vaccination was associated with its capacity to induce a superior degree of in vivo mammary cancer cell apoptosis. Our research suggests that intratumoral vaccination using recombinant vaccinia virus could be employed to increase the activity of a genetic cancer vaccine. This study may have important implications for the design of cancer vaccine protocols for the treatment of breast cancer and of accessible tumors using recombinant vaccinia virus.

Masuelli, Laura; Marzocchella, Laura; Focaccetti, Chiara; Lista, Florigio; Nardi, Alessandra; Scardino, Antonio; Mattei, Maurizio; Turriziani, Mario; Modesti, Mauro; Forni, Guido; Schlom, Jeffrey; Modesti, Andrea; Bei, Roberto

2012-01-01

128

[Mumps vaccine virus transmission].  

PubMed

In this work we report the mumps vaccine virus shedding based on the laboratory confirmed cases of the mumps virus (MuV) infection. The likely epidemiological sources of the transmitted mumps virus were children who were recently vaccinated with the mumps vaccine containing Leningrad-Zagreb or Leningrad-3 MuV. The etiology of the described cases of the horizontal transmission of both mumps vaccine viruses was confirmed by PCR with the sequential restriction analysis. PMID:24772647

Otrashevskaia, E V; Kulak, M V; Otrashevskaia, A V; Karpov, I A; Fisenko, E G; Ignat'ev, G M

2013-01-01

129

Genetically stable and fully effective smallpox vaccine strain constructed from highly attenuated vaccinia LC16m8  

Microsoft Academic Search

A highly attenuated LC16m8 (m8) smallpox vaccine has been licensed in Japan because of its extremely low neurovirulence profile, which is comparable to that of replication incompetent strains of vaccinia virus. From 1973 to 1975, m8 was administrated to >100,000 infants where it induced levels of immunity similar to that of the originating Lister strain, without any serious side effects.

Minoru Kidokoro; Masato Tashiro; Hisatoshi Shida

2005-01-01

130

Secondary and tertiary transmission of vaccinia virus after sexual contact with a smallpox vaccinee--San Diego, California, 2012.  

PubMed

On June 24, 2012, CDC notified Public Health Services, County of San Diego Health and Human Services Agency, of a suspected case of vaccinia virus infection transmitted by sexual contact. The case had been reported to CDC by an infectious disease specialist who had requested vaccinia immune globulin intravenous (VIGIV) (Cangene Corporation, Berwyn, Pennsylvania) for a patient with lesions suspicious for vaccinia. The patient reported two recent sexual contacts: one with a partner who recently had been vaccinated against smallpox and a later encounter with an unvaccinated partner. Infections resulting from secondary transmission of vaccinia virus from the smallpox vaccinee to the patient and subsequent tertiary transmission of the virus from the patient to the unvaccinated partner were confirmed by the County of San Diego Public Health Laboratory. The smallpox vaccine had been administered under the U.S. Department of Defense smallpox vaccination program. The vaccinee did not experience vaccine-associated complications; however, the secondary and tertiary patients were hospitalized and treated with VIGIV. No further transmission was known to have occurred. This report describes the epidemiology and clinical course of the secondary and tertiary cases and efforts to prevent further transmission to contacts. PMID:23446513

2013-03-01

131

A chitosan hydrogel-based cancer drug delivery system exhibits synergistic antitumor effects by combining with a vaccinia viral vaccine.  

PubMed

Cancer treatment combining chemotherapy and immunotherapy has been vigorously exploited to further improve cancer therapeutic efficacy. This study investigated a new chemoimmunotherapy approach utilizing hydrogel as a local anti-cancer drug delivery system. Chitosan hydrogel containing doxorubicin (CH-DOX) and vaccinia virus vaccine expressing Sig/E7/LAMP-1 (Vac-Sig/E7/LAMP-1) were used as chemoimmunotherapeutic agents. It was found that intratumoral injection of CH-DOX effectively inhibited tumor growth itself and, in addition, exhibited a synergistic antitumor effect in combination with a vaccinia virus-based vaccine. This combination did not decrease but rather increased the number of tumor-specific CD8(+) T cells primed by vaccinia virus-mediated vaccination; the resulting antitumor effects were further improved up to 60 days as compared with monotherapy after tumor challenge, and the survival of tumor-bearing mice was dramatically prolonged. This study is a pioneer report that demonstrates the use of a biodegradable hydrogel system as an anti-cancer drug delivery system for successful chemoimmunotherapy. It is hoped that, this study can provide a foundation for a rational approach to improve antitumor efficacy of chemoimmunotherapy. PMID:17897800

Han, Hee Dong; Song, Chung Kil; Park, Yong Sung; Noh, Kyung Hee; Kim, Jin Hee; Hwang, Taewon; Kim, Tae Woo; Shin, Byung Cheol

2008-02-28

132

Immunogenicity of a recombinant vaccinia virus expressing envelope a glycoprotein of bovine leukaemia virus.  

PubMed

We constructed a recombinant vaccinia virus (RVV) expressing envelope (env) glycoprotein (gp51) of bovine leukaemia virus (BLV): the expression of gp51 was detected by Western blot of the lysates of rabbit kidney cells infected with the RVV. The rabbits inoculated intradermally with the RVV alone failed to induce detectable anti-gp51 antibodies even 10 weeks after immunization. However, when these animals were boosted with inactivated BLV virion in saline, significant levels of anti-gp51 antibodies were induced as shown both in Western blot and immunodiffusion analyses. In these animals, antibodies against gag product (p24) were not detected. On the other hand, the rabbits inoculated with wild-type vaccinia virus and boosted similarly three times with the BLV virion in saline did not induce detectable anti-gp51 antibodies at all. The present experiment revealed that the RVV possessed the capability to endow immunological memory without inducing apparent anti-gp51 antibody responses, meaning that the RVV activated helper T cells far more strongly than B cells. The applicability of the RVV to vaccine development is discussed. PMID:2848378

Ohishi, K; Maruyama, T; Shida, H; Nishimaki, J; Miki, K; Sagata, N; Ikawa, Y; Sugimoto, M

1988-10-01

133

Intratumoral Injection of Therapeutic HPV Vaccinia Vaccine Following Cisplatin Enhances HPV-specific Antitumor Effects  

PubMed Central

Despite theconventional treatments of radiation therapy and chemotherapy, the five-year survival rates for patients withadvanced stage cervical cancers remain low. Cancer immunotherapy has emerged as an alternative, innovative therapythat may improve survival. Here we utilize a preclinical HPV-16 E6/E7-expressing tumor model, TC-1,and employ the chemotherapeutic agent cisplatin to generate an accumulation of CD11c+ dendritic cells in tumor loci making it an ideal location for the administration of therapeutic vaccines. Following cisplatin treatment, we tested different routes of administration of a therapeutic HPV vaccinia vaccine encoding HPV-16 E7 antigen (CRT/E7-VV).We found that TC-1tumor-bearing C57BL/6 mice treated with cisplatin and intratumoral injection of CRT/E7-VV significantly increased E7-specific CD8+ T cells in the blood and generated potent local and systemic antitumor immune responsescompared to mice receiving cisplatin and CRT/E7-VV intraperitoneally or mice treated with cisplatin alone. We further extended our study using a clinical grade recombinant vaccinia vaccine encoding HPV-16/18 E6/E7 antigens (TA-HPV).We found that intratumoral injection with TA-HPV following cisplatin treatment also led to increased E7-specific CD8+ T cells in the blood as well as significantly decreased tumor size compared to intratumoral injection with wild type vaccinia virus. Our study has strong implications for future clinical translation using intratumoralinjection of TA-HPV in conjunction with the current treatment strategies for patients with advanced cervical cancer.

Lee, Sung Yong; Kang, Tae Heung; Knoff, Jayne; Huang, Zhuomin; Soong, Ruey-Shyang; Alvarez, Ronald D.; Hung, Chien-Fu; Wu, Tzyy-Choou

2013-01-01

134

Antiviral treatment is more effective than smallpox vaccination upon lethal monkeypox virus infection  

Microsoft Academic Search

There is concern that variola virus, the aetiological agent of smallpox, may be used as a biological weapon. For this reason several countries are now stockpiling (vaccinia virus-based) smallpox vaccine. Although the preventive use of smallpox vaccination has been well documented, little is known about its efficacy when used after exposure to the virus. Here we compare the effectiveness of

Koert J. Stittelaar; Johan Neyts; Lieve Naesens; Geert van Amerongen; Rob F. van Lavieren; Antonin Holý; Erik de Clercq; Edwin Fries; Chantal Maas; Paul G. H. Mulder; Ben A. M. van der Zeijst; Albert D. M. E. Osterhaus

2006-01-01

135

Expression and characterization of bovine lactoperoxidase by recombinant vaccinia virus  

PubMed Central

Lactoperoxidase (LPO) is a 78 kDa heme-containing oxidation–reduction enzyme present in milk, found in physiological fluids of mammals. LPO has an antimicrobial activity, and presumably contribute to the protective functions of milk against infectious diseases. In this study, recombinant vaccinia virus expressing bovine LPO (vv/bLPO) was constructed. In rabbit kidney (RK13) cells infected with vv/bLPO, recombinant bLPO was detected in both cell extracts and culture supernatants. Tunicamycin treatment decreased the molecular weight of recombinant bLPO, indicating that recombinant bLPO contains a N-linked glycosylation site. The replication of recombinant vaccinia viruses expressing bovine lactoferrin (vv/bLF) at a multiplicity of infection (moi) of 5 plaque-forming units (PFU)/cell was inhibited by antiviral activity of recombinant bLF, suggesting that vv/bLF has an antiviral effect against vaccinia virus. On the other hand, the replication of vv/bLPO at a moi of 5 PFU/cell was not inhibited by antiviral activity of recombinant bLPO, indicating that this recombinant virus could be used as a suitable viral vector. These results indicate that a combination of bLPO and vaccinia virus vector may be useful for medical and veterinary applications in vivo.

Xuan, Xuenan; Kojima, Asato; Igarashi, Ikuo; Fujisaki, Kozo; Shimazaki, Kei-ichi

2009-01-01

136

Clinical responses to smallpox vaccine in vaccinia-naive and previously vaccinated populations: undiluted and diluted Lancy-Vaxina vaccine in a single-blind, randomized, prospective trial.  

PubMed

We conducted a single-blind, randomized trial of 2 dilutions (1:1 or 1:10) of Lancy-Vaxina vaccine (Berna Biotech) in vaccinia-naive persons (n=36) and persons previously vaccinated >25 years ago (n=76). All vaccinees responded successfully to the vaccination. There were no significant differences in the size of the skin lesions, the number of adverse events, the amount of viral shedding, or the level of antibody responses between the undiluted (n=56) and diluted (n = 56) vaccine groups. Compared with vaccinia-naive persons, previously vaccinated persons exhibited significantly smaller and more rapidly evolving skin lesions and fewer adverse events. Previously vaccinated persons had significantly higher neutralizing antibody levels before the administration of the study vaccine than vaccinia-naive persons, and viral shedding from lesions in previously vaccinated persons was lower and diminished more rapidly than from lesions in vaccinia-naive persons. PMID:16107961

Kim, Sung-Han; Yeo, Sang-Gu; Jang, Hee-Chang; Park, Wan-Beom; Lee, Chang-Seop; Lee, Ki-Deok; Kim, Hong-Bin; Kim, Nam-Joong; Kim, Young-Taek; Jee, Youngmee; Cho, Haewol; Oh, Myoung-don; Choe, Kang-Won

2005-09-15

137

Clinical signs, diagnosis, and case reports of Vaccinia virus infections  

Microsoft Academic Search

Vaccinia virus is responsible for a zoonosis that usually affects cattle and human beings in Brazil. The initial clinical signs of the infection are focal red skin areas, fever, and general symptoms similar to those of a cold. Then, pustules and ulcerated lesions surrounded by edema and erythema follow, as well as local lymphadenopathy that can last for weeks. Cure

Daniela Carla Medeiros-Silva; Eduardo Augusto dos Santos Moreira-Silva; Juliana de Assis Silva Gomes; Flávio Guimarães da Fonseca; Rodrigo Correa-Oliveira

2010-01-01

138

Characterization of an attenuated TE3L-deficient vaccinia virus Tian Tan strain.  

PubMed

An attenuated vaccinia virus (VACV), TE3L(-)VTT, was evaluated for virulence and safety to determine its potential use as a vaccine or as a recombinant virus vector to express foreign genes. The virulence of TE3L(-)VTT was compared with that of the wild-type VTT both in vivo and in vitro. The humoral and cellular immune responses were detected in a mouse model to test the vaccine efficacy of the TE3L mutant. The results suggested that deletion of the TE3L gene decreased the virulence and neurovirulence significantly in mice and rabbit models, yet retained the immunogenicity. Thus, the deletion of TE3L improved the safety of the VTT vector; this approach may yield a valuable resource for studies of recombinant VACV-vectored vaccines. PMID:23084929

Wang, Yuhang; Kan, Shifu; Du, Shouwen; Qi, Yanxin; Wang, Jinhui; Liu, Liming; Ji, Huifan; He, Dongyun; Wu, Na; Li, Chang; Chi, Baorong; Li, Xiao; Jin, Ningyi

2012-12-01

139

Hydroxyurea-resistant vaccinia virus: overproduction of ribonucleotide reductase  

SciTech Connect

Repeated passage of vaccinia virus in increasing concentrations of hydroxyurea followed by plaque purification resulted in the isolation of variants capable of growth in 5 mM hydroxyurea, a drug concentration which inhibited the reproduction of wild-type vaccinia virus 1000-fold. Analyses of viral protein synthesis by using (/sup 35/S)methionine pulse-labeling at intervals throughout the infection cycle revealed that all isolates overproduced a 34,000-molecular-weight (MW) early polypeptide. Measurement of ribonucleoside-diphosphate reductase activity after infection indicated that 4- to 10-fold more activity was induced by hydroxyurea-resistant viruses than by the wild-type virus. A two-step partial purification resulted in a substantial enrichment for the 34,000-MW protein from extracts of wild-type and hydroxyurea-resistant-virus-infected, but not mock-infected, cells. In the presence of the drug, the isolates incorporated (/sup 3/H)thymidine into DNA earlier and a rate substantially greater than that of the wild type, although the onset of DNA synthesis was delayed in both cases. The drug resistance trait was markedly unstable in all isolates. In the absence of selective pressure, plaque-purified isolated readily segregated progeny that displayed a wide range of resistance phenotypes. The results of this study indicate that vaccinia virus encodes a subunit of ribonucleotide reductase which is 34,000-MW early protein whose overproduction confers hydroxyurea resistance on reproducing viruses.

Slabaugh, M.B.; Mathews, C.K.

1986-11-01

140

Capturing the Natural Diversity of the Human Antibody Response against Vaccinia Virus ?  

PubMed Central

The eradication of smallpox (variola) and the subsequent cessation of routine vaccination have left modern society vulnerable to bioterrorism employing this devastating contagious disease. The existing, licensed vaccines based on live vaccinia virus (VACV) are contraindicated for a substantial number of people, and prophylactic vaccination of large populations is not reasonable when there is little risk of exposure. Consequently, there is an emerging need to develop efficient and safe therapeutics to be used shortly before or after exposure, either alone or in combination with vaccination. We have characterized the human antibody response to smallpox vaccine (VACV Lister) in immunized volunteers and isolated a large number of VACV-specific antibodies that recognize a variety of different VACV antigens. Using this broad antibody panel, we have generated a fully human, recombinant analogue to plasma-derived vaccinia immunoglobulin (VIG), which mirrors the diversity and specificity of the human antibody immune response and offers the advantage of unlimited supply and reproducible specificity and activity. The recombinant VIG was found to display a high specific binding activity toward VACV antigens, potent in vitro VACV neutralizing activity, and a highly protective efficacy against VACV challenge in the mouse tail lesion model when given either prophylactically or therapeutically. Altogether, the results suggest that this compound has the potential to be used as an effective postexposure prophylaxis or treatment of disease caused by orthopoxviruses.

Lantto, Johan; Haahr Hansen, Margit; Rasmussen, S?ren Kofoed; Steinaa, Lucilla; Poulsen, Tine R.; Duggan, Jackie; Dennis, Mike; Naylor, Irene; Easterbrook, Linda; Bregenholt, S?ren; Haurum, John; Jensen, Allan

2011-01-01

141

Chasing Jenner's Vaccine: Revisiting Cowpox Virus Classification  

PubMed Central

Cowpox virus (CPXV) is described as the source of the first vaccine used to prevent the onset and spread of an infectious disease. It is one of the earliest described members of the genus Orthopoxvirus, which includes the viruses that cause smallpox and monkeypox in humans. Both the historic and current literature describe “cowpox” as a disease with a single etiologic agent. Genotypic data presented herein indicate that CPXV is not a single species, but a composite of several (up to 5) species that can infect cows, humans, and other animals. The practice of naming agents after the host in which the resultant disease manifests obfuscates the true taxonomic relationships of “cowpox” isolates. These data support the elevation of as many as four new species within the traditional “cowpox” group and suggest that both wild and modern vaccine strains of Vaccinia virus are most closely related to CPXV of continental Europe rather than the United Kingdom, the homeland of the vaccine.

Carroll, Darin S.; Emerson, Ginny L.; Li, Yu; Sammons, Scott; Olson, Victoria; Frace, Michael; Nakazawa, Yoshinori; Czerny, Claus Peter; Tryland, Morten; Kolodziejek, Jolanta; Nowotny, Norbert; Olsen-Rasmussen, Melissa; Khristova, Marina; Govil, Dhwani; Karem, Kevin; Damon, Inger K.; Meyer, Hermann

2011-01-01

142

Vaccination against Canine Distemper Virus Infection in Infant Ferrets with and without Maternal Antibody Protection, Using Recombinant Attenuated Poxvirus Vaccines  

Microsoft Academic Search

Canine distemper virus (CDV) infection of ferrets is clinically and immunologically similar to measles, making this a useful model for the human disease. The model was used to determine if parenteral or mucosal immunization of infant ferrets at 3 and 6 weeks of age with attenuated vaccinia virus (NYVAC) or canarypox virus (ALVAC) vaccine strains expressing the CDV hemagglutinin (H)

JANET WELTER; JILL TAYLOR; JAMES TARTAGLIA; ENZO PAOLETTI; CHARLES B. STEPHENSEN

2000-01-01

143

Attenuated vaccinia virus-circumsporozoite protein recombinants confer protection against rodent malaria.  

PubMed Central

NYVAC-based vaccinia virus recombinants expressing the circumsporozoite protein (CSP) were evaluated in the Plasmodium berghei rodent malaria model system. Immunization of mice with a NYVAC-based CSP recombinant elicited a high level of protection (60 to 100%). Protection did not correlate with CS repeat-specific antibody responses and was abrogated by in vivo CD8+ T-cell depletion. Protection was not enhanced by modification of the subcellular localization of CSP. These results suggest the potential of poxvirus-based vectors for the development of vaccine candidates for human malaria.

Lanar, D E; Tine, J A; de Taisne, C; Seguin, M C; Cox, W I; Winslow, J P; Ware, L A; Kauffman, E B; Gordon, D; Ballou, W R; Paoletti, E; Sadoff, J C

1996-01-01

144

IL-15 adjuvanted multivalent vaccinia-based universal influenza vaccine requires CD4+ T cells for heterosubtypic protection.  

PubMed

Current influenza vaccines are ineffective against novel viruses and the source or the strain of the next outbreak of influenza is unpredictable; therefore, establishing universal immunity by vaccination to limit the impact of influenza remains a high priority. To meet this challenge, a novel vaccine has been developed using the immunogenic live vaccinia virus as a vaccine vector, expressing multiple H5N1 viral proteins (HA, NA, M1, M2, and NP) together with IL-15 as a molecular adjuvant. Previously, this vaccine demonstrated robust sterile cross-clade protection in mice against H5 influenza viruses, and herein its use has been extended to mediate heterosubtypic immunity toward viruses from both group 1 and 2 HA lineages. The vaccine protected mice against lethal challenge by increasing survival and significantly reducing lung viral loads against the most recent human H7N9, seasonal H3N2, pandemic-2009 H1N1, and highly pathogenic H7N7 influenza A viruses. Influenza-specific antibodies elicited by the vaccine failed to neutralize heterologous viruses and were unable to confer protection by passive transfer. Importantly, heterologous influenza-specific CD4(+) and CD8(+) T-cell responses that were elicited by the vaccine were effectively recalled and amplified following viral challenge in the lungs and periphery. Selective depletion of T-cell subsets in the immunized mice revealed an important role for CD4(+) T cells in heterosubtypic protection, despite low sequence conservation among known MHC-II restricted epitopes across different influenza viruses. This study illustrates the potential utility of our multivalent Wyeth/IL-15/5Flu as a universal influenza vaccine with a correlate of protective immunity that is independent of neutralizing antibodies. PMID:24706798

Valkenburg, Sophie A; Li, Olive T W; Mak, Polly W Y; Mok, Chris K P; Nicholls, John M; Guan, Yi; Waldmann, Thomas A; Peiris, J S Malik; Perera, Liyanage P; Poon, Leo L M

2014-04-15

145

Recombinant Modified Vaccinia Virus Ankara Expressing the Surface gp120 of Simian Immunodeficiency Virus (SIV) Primes for a Rapid Neutralizing Antibody Response to SIV Infection in Macaques  

Microsoft Academic Search

Neutralizing antibodies were assessed before and after intravenous challenge with pathogenic SIVsmE660 in rhesus macaques that had been immunized with recombinant modified vaccinia virus Ankara expressing one or more simian immunodeficiency virus gene products (MVA-SIV). Animals received either MVA-gag-pol, MVA-env, MVA-gag-pol-env, or nonrecombinant MVA. Although no animals were completely protected from infection with SIV, animals immunized with recombinant MVA-SIV vaccines

ILNOUR OURMANOV; MIROSLAWA BILSKA; VANESSA M. HIRSCH; DAVID C. MONTEFIORI

2000-01-01

146

Strong HIV-Specific CD4+ and CD8+ T-Lymphocyte Proliferative Responses in Healthy Individuals Immunized with an HIV-1 DNA Vaccine and Boosted with Recombinant Modified Vaccinia Virus Ankara Expressing HIV-1 Genes?  

PubMed Central

We investigated HIV-1 vaccine-induced lymphoproliferative responses in healthy volunteers immunized intradermally or intramuscularly (with or without adjuvant granulocyte-macrophage colony-stimulating factor [GM-CSF] protein) with DNA expressing HIV-1 gag, env, rev, and rt at months 0, 1, and 3 using a Biojector and boosted at 9 months with modified vaccinia virus Ankara (MVA) expressing heterologous HIV-1 gag, env, and pol (HIV-MVA). Lymphoproliferative responses to aldrithiol-2 (AT-2)-inactivated-HIV-1 antigen were tested by a [3H]thymidine uptake assay and a flow-cytometric assay of specific cell-mediated immune response in activated whole blood (FASCIA-WB) 2 weeks after the HIV-MVA boost (n = 38). A FASCIA using peripheral blood mononuclear cells (FASCIA-PBMC) was also employed (n = 14). Thirty-five of 38 (92%) vaccinees were reactive by the [3H]thymidine uptake assay. Thirty-two of 38 (84%) vaccinees were reactive by the CD4+ T-cell FASCIA-WB, and 7 of 38 (18%) also exhibited CD8+ T-cell responses. There was strong correlation between the proliferative responses measured by the [3H]thymidine uptake assay and CD4+ T-cell FASCIA-WB (r = 0.68; P < 0.01). Fourteen vaccinees were analyzed using all three assays. Ten of 14 (71%) and 11/14 (79%) demonstrated CD4+ T-cell responses in FASCIA-WB and FASCIA-PBMC, respectively. CD8+ T-cell reactivity was observed in 3/14 (21%) and 7/14 (50%) using the FASCIA-WB and FASCIA-PBMC, respectively. All 14 were reactive by the [3H]thymidine uptake assay. The overall HIV-specific T-cell proliferative response in the vaccinees employing any of the assays was 100% (38/38). A standardized FASCIA-PBMC, which allows simultaneous phenotyping, may be an option to the [3H]thymidine uptake assay for assessment of vaccine-induced T-cell proliferation, especially in isotope-restricted settings.

Aboud, Said; Nilsson, Charlotta; Karlen, Katarina; Marovich, Mary; Wahren, Britta; Sandstrom, Eric; Gaines, Hans; Biberfeld, Gunnel; Godoy-Ramirez, Karina

2010-01-01

147

The Cowpox Virus-Encoded Homolog of the Vaccinia Virus Complement Control Protein Is an Inflammation Modulatory Protein  

Microsoft Academic Search

Vaccinia virus complement control protein (VCP) is encoded by vaccinia virus, with its homolog encoded by other pathogenic poxviruses including variola virus. Since rodents are the primary reservoir hosts of cowpox virus (CPV) and since CPV encodes a highly conserved functional homolog of VCP, termed here the inflammation modulatory protein (IMP), the effects of injection of CPV into the footpads

Cathie G. Miller; Sergei N. Shchelkunov; Girish J. Kotwal

1997-01-01

148

Evaluation of modified vaccinia virus Ankara expressing VP2 protein of infectious bursal disease virus as an immunogen in chickens  

PubMed Central

A recombinant modified vaccinia Ankara (MVA) virus expressing mature viral protein 2 (VP2) of the infectious bursal disease virus (IBDV) was constructed to develop MVA-based vaccines for poultry. We demonstrated that this recombinant virus was able to induce a specific immune response by observing the production of anti-IBDV-seroneutralizing antibodies in specific pathogen-free chickens. Besides, as the epitopes of VP2 responsible to induce IBDV-neutralizing antibodies are discontinuous, our results suggest that VP2 protein expressed from MVA-VP2 maintained the correct conformational structure. To our knowledge, this is the first report on the usefulness of MVA-based vectors for developing recombinant vaccines for poultry.

Zajac, Maria Paula Del Medico; Taboga, Oscar Alberto; Calamante, Gabriela

2012-01-01

149

Vaccinia Virus Inhibits T Cell Receptor-Dependent Responses by Human ?? T Cells  

PubMed Central

Vaccinia virus (VV) is an effective vaccine and vector but has evolved multiple mechanisms for evading host immunity. We characterized the interactions of VV (TianTan and New York City Board of Health strains) with human ?? T cells because of the role they play in immune control of this virus. Exposure to VV failed to trigger proliferative responses in ?? T cells from unprimed individuals, but it was an unexpected finding that VV blocked responses to model antigens by the V?2V?2 T cell subset. Infectious or ultraviolet light–inactivated VV inhibited proliferative V?2V?2 T cell responses to phosphoantigens and tumor cells, prevented cytolysis of Daudi B cells, and reduced cytokine production. Inhibiting V?2V?2 T cells may be a mechanism for evading host immunity and increasing VV virulence. Increased VV replication or expression in the absence of ?? T cell responses might contribute to its potency as a vaccine against poxvirus and recombinant antigens.

Li, Haishan; Deetz, Carl O.; Zapata, Juan Carlos; Cairo, Cristiana; Hebbeler, Andrew M.; Propp, Nadia; Salvato, Maria S.; Shao, Yiming; Pauza, C. David

2008-01-01

150

In vitro activity of cidofovir against the emerging Cantagalo virus and the smallpox vaccine strain IOC  

Microsoft Academic Search

The antiviral effect of cidofovir was evaluated against two strains of vaccinia virus: the field strain Cantagalo virus (CTGV) and the smallpox vaccine IOC. The drug severely inhibited virus replication, revealing an EC50 (drug concentration required to inhibit 50% of virus replication) of 7.68?M and 9.66?M, respectively, for CTGV and vaccine strain IOC. Similarly, other field isolates of Cantagalo-like viruses

Desyreé Murta Jesus; Nissin Moussatché; Clarissa R. Damaso

2009-01-01

151

Detection of vaccinia virus DNA by quartz crystal microbalance.  

PubMed

In this study, we describe a detection system for the indirect detection of vaccinia virus by DNA analysis. The system uses quartz crystal microbalance (QCM) as the detection technique and polymerase chain reaction (PCR) for amplification. Different immobilization strategies for the capture probe on the quartz chip are studied. For the QCM detection of hybridisation, the influence of the structure and length of target DNA is analyzed. For the detection of DNA from an amplification product, an efficient denaturation procedure is developed. On the basis of these investigations, vaccinia virus DNA is detected with only a low number of amplification rounds and a short analysis time. Specificity can be clearly shown. To enhance the signal strength and to have a further proof of specificity, a gold nanoparticle-tagged enhancer sequence can be used. PMID:21820997

Kleo, K; Kapp, A; Ascher, L; Lisdat, F

2011-11-15

152

Optimisation of Prime-Boost Immunization in Mice Using Novel Protein-Based and Recombinant Vaccinia (Tiantan)-Based HBV Vaccine  

PubMed Central

Background A therapeutic vaccine for chronic hepatitis B virus (HBV) infection that enhances virus-specific cellular immune responses is urgently needed. The “prime–boost” regimen is a widely used vaccine strategy against many persistence infections. However, few reports have addressed this strategy applying for HBV therapeutic vaccine development. Methodology/Principal Findings To develop an effective HBV therapeutic vaccine, we constructed a recombinant vaccinia virus (Tiantan) containing the S+PreS1 fusion antigen (RVJSS1) combined with the HBV particle-like subunit vaccine HBVSS1 to explore the most effective prime–boost regimen against HBV. The immune responses to different prime–boost regimens were assessed in C57BL/C mice by ELISA, ELISpot assay and Intracellular cytokine staining analysis. Among the combinations tested, an HBV protein particle vaccine priming and recombinant vaccinia virus boosting strategy accelerated specific seroconversion and produced high antibody (anti-PreS1, anti-S antibody) titres as well as the strongest multi-antigen (PreS1, and S)-specific cellular immune response. HBSS1 protein prime/RVJSS1 boost immunization was also generated more significant level of both CD4+ and CD8+ T cell responses for Th1 cytokines (TNF-? and IFN-?). Conclusions The HBSS1 protein-vaccine prime plus RVJSS1 vector boost elicits specific antibody as well as CD4 and CD8 cells secreting Th1-like cytokines, and these immune responses may be important parameters for the future HBV therapeutic vaccines.

Deng, Yao; Wen, Bo; Wang, Wen; Xiong, Shaoqing; Ruan, Li; Tan, Wenjie

2012-01-01

153

Protein Composition of the Vaccinia Virus Mature Virion  

SciTech Connect

The protein content of vaccinia virus mature virions, purified by rate zonal and isopycnic centrifugation and solubilized by SDS or a solution of urea and thiourea, was determined by the accurate mass and time tag technology which uses both tandem mass spectrometry and Fourier transform-ion cyclotron resonance mass spectrometry to detect tryptic peptides separated by high-resolution liquid chromatography. Eighty vaccinia virus-encoded proteins representing 37% of the 218 genes annotated in the complete genome sequence were detected in at least three analyses. Ten proteins accounted for approximately 80% of the mass, while the least abundant proteins made up 1% or less of the mass. Thirteen identified proteins were not previously reported as components of virions. On the other hand, 8 previously described virion proteins were not detected here, presumably due to technical reasons including small size and hydrophobicity. In addition to vaccinia virus-encoded proteins, 24 host proteins omitting isoforms were detected. The most abundant of these were cytoskeletal proteins, heat shock proteins, and proteins involved in translation.

Resch, Wolfgang; Hixson, Kim K.; Moore, Ronald J.; Lipton, Mary S.; Moss, Bernard

2007-02-05

154

Priming and boosting immunity to respiratory syncytial virus by recombinant replication-defective vaccinia virus MVA  

Microsoft Academic Search

Intranasal and intramuscular immunizations of mice with the highly attenuated MVA strain of vaccinia virus expressing the respiratory syncytial virus (RSV) F or G glycoprotein induced higher RSV antibody titers than those achieved by infection with RSV and greatly restricted the replication of RS challenge virus in both the upper and lower respiratory tracts. In addition, a recombinant MVA expressing

Linda S. Wyatt; Stephen S. Whitehead; Katherine A. Venanzi; Brian R. Murphy; Bernard Moss

1999-01-01

155

Efficacy and safety of a modified vaccinia Ankara (MVA) vectored plague vaccine in mice  

PubMed Central

The efficacy and safety of plague vaccines based on the modified vaccinia Ankara (MVA) viral vector was evaluated. MVA recombinants were constructed expressing Yersinia pestis antigens under the translational control of the encephalomyocarditis virus (EMCV) internal ribosomal entry site (IRES) and/or fused to the tissue plasminogen activator (tPA) secretory signal. A MVA/Y. pestis recombinant that expressed a truncated version of the low-calcium response V antigen (MVA/IRES/tPA/V307), conferred significant protection (87.5%–100%) against intranasal or intraperitoneal challenge with CO92 (encapsulated) or Java 9 (non-encapsulated) strains of Y pestis, respectively. In contrast, a MVA/Y. pestis recombinant that expressed the full-length V antigen provided only 37.5% protection against challenge with CO92 or Java 9 strains, respectively. Interestingly, a MVA/Y. pestis recombinant that expressed the capsular protein (F1) did not elicit significant antibody titers but still conferred 50% and 25% protection against CO92 or Java 9 challenge, respectively. The MVA/Y. pestis recombinant viruses did not demonstrate any mortality or morbidity in SCID mice. Based on their safety and efficacy in mice, these MVA/Y. pestis recombinants are candidates for further development as biodefense and public health vaccines.

Brewoo, Joseph N.; Powell, Tim D.; Stinchcomb, Dan T.; Osorio, Jorge E.

2010-01-01

156

Effects of different promoters on the virulence and immunogenicity of a HIV-1 Env-expressing recombinant vaccinia vaccine.  

PubMed

Previously, we developed a vaccination regimen that involves priming with recombinant vaccinia virus LC16m8? (rm8?) strain followed by boosting with a Sendai virus-containing vector. This protocol induced both humoral and cellular immune responses against the HIV-1 envelope protein. The current study aims to optimize this regimen by comparing the immunogenicity and safety of two rm8? strains that express HIV-1 Env under the control of a moderate promoter, p7.5, or a strong promoter, pSFJ1-10. m8?-p7.5-JRCSFenv synthesized less gp160 but showed significantly higher growth potential than m8?-pSFJ-JRCSFenv. The two different rm8? strains induced antigen-specific immunity; however, m8?-pSFJ-JRCSFenv elicited a stronger anti-Env antibody response whereas m8?-p7.5-JRCSFenv induced a stronger Env-specific cytotoxic T lymphocyte response. Both strains were less virulent than the parental m8? strain, suggesting that they would be safe for use in humans. These findings indicate the vaccine can be optimized to induce favorable immune responses (either cellular or humoral), and forms the basis for the rational design of an AIDS vaccine using recombinant vaccinia as the delivery vector. PMID:24370703

Isshiki, Mao; Zhang, Xianfeng; Sato, Hirotaka; Ohashi, Takashi; Inoue, Makoto; Shida, Hisatoshi

2014-02-01

157

Expression of Brucella Antigens in Vaccinia Virus to Prevent Brucellosis in Humans: Protection Studies in Mice.  

National Technical Information Service (NTIS)

The current work has the overall objectives of genetically engineering mono and poly-valent, IL-12 producing vaccinia/Brucella recombinant vaccines using synthetic E/L promoters and to test such vaccinia recombinants for their ability to induce a protecti...

G. G. Schurig

1999-01-01

158

Immunogenicity and safety of the vaccinia virus LC16m8? vector expressing SIV Gag under a strong or moderate promoter in a recombinant BCG prime-recombinant vaccinia virus boost protocol.  

PubMed

We compared the effect of the very strong pSFJ1-10 and moderately strong p7.5 promoters on the immunogenicity and pathogenicity of the replication-competent vaccinia virus (VV) LC16m8? (m8?) vector harboring the SIV gag gene in a vaccination regimen consisting of a recombinant BCG-SIVGag (rBCG-SIVGag) prime followed by a recombinant vaccinia boost. m8?/pSFJ/SIVGag synthesized more Gag protein than m8?/p7.5/SIVGag but replicated less efficiently in vitro. In addition, m8?/pSFJ/SIVGag was less pathogenic and elicited Gag-specific IFN-?(+), CD107a(+), CD8(+) cells more efficiently than m8?/p7.5/SIVGag. Vaccination by this regimen elicited long-lasting Gag-specific CD8(+) T cells, the majority of which showed a CCR7(-) phenotype at over 8 weeks post-boost. Tetramer staining analyses revealed maintenance of Gag specific tetramer(+), CD62L(-), CD8(+) T cells for long time in vaccinated mice. However, Gag expression increased the neurotoxicity of the vaccinia vector, indicating the necessity of safety testing for each recombinant VV. We propose that this recombinant BCG prime-m8?/pSFJ/HIVGag boost regimen would be a promising vaccination procedure for preventing HIV infection. PMID:23731631

Sato, Hirotaka; Jing, Chen; Isshiki, Mao; Matsuo, Kazuhiro; Kidokoro, Minoru; Takamura, Shiki; Zhang, Xianfeng; Ohashi, Takashi; Shida, Hisatoshi

2013-08-01

159

Safety, immunogenicity and efficacy of modified vaccinia Ankara (MVA) against Dryvax challenge in vaccinia-naïve and vaccinia-immune individuals.  

PubMed

Modified vaccinia Ankara (MVA) was evaluated as an alternative to Dryvax in vaccinia-naïve and vaccinia-immune adult volunteers. Subjects received intramuscular MVA or placebo followed by Dryvax challenge at 3 months. Two or more doses of MVA prior to Dryvax reduced severity of lesion formation, decreased magnitude and duration of viral shedding, and augmented post-Dryvax vaccinia-specific CD8(+) T cell responses and extracellular enveloped virus protein-specific antibody responses. MVA vaccination is safe and immunogenic and improves the safety and immunogenicity of subsequent Dryvax vaccination supporting the potential for using MVA as a vaccine in the general population to improve immunity to orthopoxviruses. PMID:17126963

Parrino, Janie; McCurdy, Lewis H; Larkin, Brenda D; Gordon, Ingelise J; Rucker, Steven E; Enama, Mary E; Koup, Richard A; Roederer, Mario; Bailer, Robert T; Moodie, Zoe; Gu, Lin; Yan, Lihan; Graham, Barney S

2007-02-01

160

Environmental Risk Assessment of Clinical Trials Involving Modified Vaccinia Virus Ankara (MVA)-Based Vectors  

PubMed Central

The modified vaccinia virus Ankara (MVA) strain, which has been developed as a vaccine against smallpox, is since the nineties widely tested in clinical trials as recombinant vector for vaccination or gene therapy applications. Although MVA is renowned for its safety, several biosafety aspects need to be considered when performing the risk assessment of a recombinant MVA (rMVA). This paper presents the biosafety issues and the main lessons learned from the evaluation of the clinical trials with rMVA performed in Belgium. Factors such as the specific characteristics of the rMVA, the inserted foreign sequences/transgene, its ability for reconversion, recombination and dissemination in the population and the environment are the main points of attention. Measures to prevent or manage identified risks are also discussed.

Goossens, Martine; Pauwels, Katia; Willemarck, Nicolas; Breyer, Didier

2013-01-01

161

Environmental risk assessment of clinical trials involving modified vaccinia virus Ankara (MVA)-based vectors.  

PubMed

The modified vaccinia virus Ankara (MVA) strain, which has been developed as a vaccine against smallpox, is since the nineties widely tested in clinical trials as recombinant vector for vaccination or gene therapy applications. Although MVA is renowned for its safety, several biosafety aspects need to be considered when performing the risk assessment of a recombinant MVA (rMVA). This paper presents the biosafety issues and the main lessons learned from the evaluation of the clinical trials with rMVA performed in Belgium. Factors such as the specific characteristics of the rMVA, the inserted foreign sequences/transgene, its ability for reconversion, recombination and dissemination in the population and the environment are the main points of attention. Measures to prevent or manage identified risks are also discussed. PMID:24397528

Goossens, Martine; Pauwels, Katia; Willemarck, Nicolas; Breyer, Didier

2013-12-01

162

Characterization of UVC light sensitivity of vaccinia virus.  

PubMed

Interest in airborne smallpox transmission has been renewed because of concerns regarding the potential use of smallpox virus as a biothreat agent. Air disinfection via upper-room 254-nm germicidal UV (UVC) light in public buildings may reduce the impact of primary agent releases, prevent secondary airborne transmission, and be effective prior to the time when public health authorities are aware of a smallpox outbreak. We characterized the susceptibility of vaccinia virus aerosols, as a surrogate for smallpox, to UVC light by using a benchtop, one-pass aerosol chamber. We evaluated virus susceptibility to UVC doses ranging from 0.1 to 3.2 J/m(2), three relative humidity (RH) levels (20%, 60%, and 80%), and suspensions of virus in either water or synthetic respiratory fluid. Dose-response plots show that vaccinia virus susceptibility increased with decreasing RH. These plots also show a significant nonlinear component and a poor fit when using a first-order decay model but show a reasonable fit when we assume that virus susceptibility follows a log-normal distribution. The overall effects of RH (P < 0.0001) and the suspending medium (P = 0.014) were statistically significant. When controlling for the suspending medium, the RH remained a significant factor (P < 0.0001) and the effect of the suspending medium was significant overall (P < 0.0001) after controlling for RH. Virus susceptibility did not appear to be a function of virus particle size. This work provides an essential scientific basis for the design of effective upper-room UVC installations for the prevention of airborne infection transmission of smallpox virus by characterizing the susceptibility of an important orthopoxvirus to UVC exposure. PMID:17644645

McDevitt, James J; Lai, Ka Man; Rudnick, Stephen N; Houseman, E Andres; First, Melvin W; Milton, Donald K

2007-09-01

163

Attenuation and immunogenicity of host-range extended modified vaccinia virus Ankara recombinants.  

PubMed

Modified vaccinia virus Ankara (MVA) is being widely investigated as a safe smallpox vaccine and as an expression vector to produce vaccines against other infectious diseases and cancer. MVA was isolated following more than 500 passages in chick embryo fibroblasts and suffered several major deletions and numerous small mutations resulting in replication defects in human and most other mammalian cells as well as severe attenuation of pathogenicity. Due to the host range restriction, primary chick embryo fibroblasts are routinely used for production of MVA-based vaccines. While a replication defect undoubtedly contributes to safety of MVA, it is worth considering whether host range and attenuation are partially separable properties. Marker rescue transfection experiments resulted in the creation of recombinant MVAs with extended mammalian cell host range. Here, we characterize two host-range extended rMVAs and show that they (i) have acquired the ability to stably replicate in Vero cells, which are frequently used as a cell substrate for vaccine manufacture, (ii) are severely attenuated in immunocompetent and immunodeficient mouse strains following intranasal infection, (iii) are more pathogenic than MVA but less pathogenic than the ACAM2000 vaccine strain at high intracranial doses, (iv) do not form lesions upon tail scratch in mice in contrast to ACAM2000 and (v) induce protective humoral and cell-mediated immune responses similar to MVA. The extended host range of rMVAs may be useful for vaccine production. PMID:23928462

Melamed, Sharon; Wyatt, Linda S; Kastenmayer, Robin J; Moss, Bernard

2013-09-23

164

Inhibitors of C5 Complement Enhance Vaccinia Virus Oncolysis  

PubMed Central

Genetically engineered tumor-selective vaccinia virus (VV) has been demonstrated to be a highly effective oncolytic agent, but immune clearance may limit its therapeutic potential. As previously demonstrated, immunosuppression can lead to significant enhancement of viral recovery and therapeutic effect, but the magnitude of complement-mediated viral inactivation has not been fully elucidated and warrants further investigation. Using fluorescent microscopy and quantitative plaque assays, we have determined complement's key role in viral clearance and its multi-faceted means to pathogen destruction. Complement can lead to direct viral destruction and inhibition of viral uptake into cells, even in the absence of anti-vaccinia antibodies. Our data demonstrate C5 to be integral to the clearance pathway, and its inhibition by Staphylococcal superantigen-like protein (SSL7) leads to a 90-fold and 150-fold enhancement of VV infectivity in both the presence and absence of anti-VV antibodies, respectively. This study suggests that complement inhibition may reduce vaccinia viral neutralization and may be critical to future in vivo work.

Magge, Deepa; Guo, Z. Sheng; O'Malley, Mark E.; Francis, Lily; Ravindranathan, Roshni; Bartlett, David L.

2014-01-01

165

Serological study of vaccinia virus reservoirs in areas with and without official reports of outbreaks in cattle and humans in São Paulo, Brazil.  

PubMed

Vaccinia virus (VACV), the etiological agent of an exanthematic disease, has been associated with several bovine outbreaks in Brazil since the end of the global vaccination campaign against smallpox. It was previously believed that the vaccine virus used for the WHO global campaign had adapted to an unknown wild reservoir and was sporadically re-emerging in outbreaks in cattle and milkers. At present, it is known that Brazilian VACV is phylogenetically different from the vaccinia virus vaccinal strain, but its origin remains unknown. This study assessed the seroprevalence of orthopoxviruses in domestic and wild animals and farmers from 47 farms in three cities in the southwest region of the state of São Paulo with or without official reports of outbreaks in cattle or humans. Our data indicate a low seroprevalence of antibodies in wild animals and raise interesting questions about the real potential of wild rodents and marsupials as VACV reservoirs, suggesting other routes through which VACV can be spread. PMID:23760628

Peres, Marina Gea; Bacchiega, Thais Silva; Appolinário, Camila Michele; Vicente, Acácia Ferreira; Allendorf, Susan Dora; Antunes, João Marcelo Azevedo Paula; Moreira, Sabrina Almeida; Legatti, Emerson; Fonseca, Clóvis Rinaldo; Pituco, Edviges Maristela; Okuda, Liria Hiromi; Pantoja, José Carlos de Figueiredo; Ferreira, Fernando; Megid, Jane

2013-12-01

166

Smallpox DNA Vaccine Delivered by Novel Skin Electroporation Device Protects Mice Against Intranasal Poxvirus Challenge.  

National Technical Information Service (NTIS)

Previously, we demonstrated that an experimental smallpox DNA vaccine comprised of four vaccinia virus genes (4pox) administered by gene gun elicited protective immunity in mice challenged with vaccinia virus, and in nonhuman primates challenged with monk...

A. D. King A. M. Ferro J. W. Golden J. W. Hooper

2006-01-01

167

42 CFR 102.21 - Smallpox (Vaccinia) Vaccine Injury Table.  

Code of Federal Regulations, 2012 CFR

...illness, disability, injury, or condition) Time interval for first symptom or manifestation...vaccinia in contacts (C). Please note that these time intervals do not refer to time periods for the date of diagnosis of the...

2012-10-01

168

Characterization of Hepatitis E Virus Recombinant ORF2 Proteins Expressed by Vaccinia Viruses  

PubMed Central

Hepatitis E virus (HEV), an enterically transmitted pathogen, is one of the major causes of acute hepatitis in humans worldwide, being responsible for outbreaks and epidemics in regions with suboptimal sanitary conditions, in many of which it is endemic. In industrialized countries, hepatitis E is rarely reported, but recent studies have revealed quite high human seroprevalence rates and the possibility of porcine zoonotic transmission. There is currently no specific therapy or licensed vaccine against HEV infection, and little is known about its intracellular growth cycle, as until very recently no efficient cell culture system has been available. In the present study, vaccinia viruses have been used to express recombinant HEV ORF2 proteins, allowing the study of their glycosylation patterns and subcellular localization. Furthermore, the expressed proteins have been shown to be good antigens for diagnostic purposes and to elicit high and long-lasting specific anti-HEV titers of antibodies in mice that are passively transferred to the offspring by both transplacental and lactation routes.

Jimenez de Oya, Nereida; Escribano-Romero, Estela; Blazquez, Ana-Belen; Lorenzo, Maria; Martin-Acebes, Miguel A.; Blasco, Rafael

2012-01-01

169

Characterization of hepatitis E virus recombinant ORF2 proteins expressed by vaccinia viruses.  

PubMed

Hepatitis E virus (HEV), an enterically transmitted pathogen, is one of the major causes of acute hepatitis in humans worldwide, being responsible for outbreaks and epidemics in regions with suboptimal sanitary conditions, in many of which it is endemic. In industrialized countries, hepatitis E is rarely reported, but recent studies have revealed quite high human seroprevalence rates and the possibility of porcine zoonotic transmission. There is currently no specific therapy or licensed vaccine against HEV infection, and little is known about its intracellular growth cycle, as until very recently no efficient cell culture system has been available. In the present study, vaccinia viruses have been used to express recombinant HEV ORF2 proteins, allowing the study of their glycosylation patterns and subcellular localization. Furthermore, the expressed proteins have been shown to be good antigens for diagnostic purposes and to elicit high and long-lasting specific anti-HEV titers of antibodies in mice that are passively transferred to the offspring by both transplacental and lactation routes. PMID:22593167

Jiménez de Oya, Nereida; Escribano-Romero, Estela; Blázquez, Ana-Belén; Lorenzo, María; Martín-Acebes, Miguel A; Blasco, Rafael; Saiz, Juan-Carlos

2012-08-01

170

Generation of Recombinant Vaccinia Viruses via Green Fluorescent Protein Selection  

PubMed Central

We developed a rapid method to generate recombinant vaccinia viruses (rVVs) based upon a bicistronic cassette encoding the gene for green fluorescent protein (GFP) and a foreign gene of interest separated by an internal ribosome entry site (IRES). As proof-of-concept, we inserted a mutant env gene of human immunodeficiency virus (HIV) into the cassette, which was cloned into the vaccinia virus (VV) insertion vector pSC59 under the control of the early-late VV synthetic promoter and flanked by disrupted tk gene sequences. To generate rVVs, 293T cells were inoculated with wild-type (wt) VV, followed by transfection of the modified pSC59 vector containing the bicistronic cassette, which allows expression of GFP and the protein of interest. Next, GFP-positive cells were isolated by flow cytometry or by picking under a fluorescent microscope. Thymidine kinase–deficient (Tk?) 143B cells were then exposed to lysates of GFP-positive 293T cells and cultured in the presence of bromodeoxyuridine. This selection allows only Tk? rVV to remain viable. We demonstrated the success of this GFP selection strategy by expressing high levels of mutant HIV Env. Our approach shortens the time needed to generate rVVs and represents a practical approach to generate recombinant proteins.

Popov, Sergei; Mirshahidi, Saied; Essono, Sosthene; Song, Ruijiang; Wang, Xiaowei

2009-01-01

171

Deoxyadenosine reverses hydroxyurea inhibition of vaccinia virus growth.  

PubMed Central

Hydroxyurea, an inhibitor of ribonucleotide reductase, blocks replication of vaccinia virus. However, when medium containing hydroxyurea and dialyzed serum was supplemented with deoxyadenosine, the block to viral reproduction was circumvented, provided that an inhibitor of adenosine deaminase was also present. Deoxyguanosine, deoxycytidine, and deoxythymidine were ineffective alone and did not augment the deoxyadenosine effect. In fact, increasing concentrations of deoxyguanosine and deoxythymidine, but not deoxycytidine, eliminated the deoxyadenosine rescue, an effect that was reversed by the addition of low concentrations of deoxycytidine. These results suggested that the inhibition of viral replication by hydroxyurea was primarily due to a deficiency of dATP. Deoxyribonucleoside triphosphate pools in vaccinia virus-infected cells were measured at the height of viral DNA synthesis after a synchronous infection. With 0.5 mM hydroxyurea, the dATP pool was greater than 90% depleted, the dCTP and dGTP pools were 40 to 50% reduced, and the dTTP pool was increased. Assay of ribonucleotide reductase activity in intact virus-infected cells suggested that hydroxyurea may differentially affect reduction of the various substrates of the enzyme.

Slabaugh, M B; Howell, M L; Wang, Y; Mathews, C K

1991-01-01

172

Protection against lethal Japanese encephalitis virus infection of mice by immunization with the highly attenuated MVA strain of vaccinia virus expressing JEV prM and E genes.  

PubMed

Genes encoding the glycosylated precursor of the membrane (prM) and envelope (E) proteins of a Korean strain of Japanese encephalitis virus (JEV) were inserted into the genome of the host-range restricted, highly attenuated, and safety-tested MVA strain of vaccinia virus. MVA recombinants containing the JEV genes, under strong synthetic or modified H5 vaccinia virus promoters, were isolated. Synthesis of JEV prM and E proteins was detected by immunofluorescence microscopy, flow cytometry, and polyacrylamide gel electrophoresis. Mice inoculated and boosted by various routes with either of the MVA recombinants produced JEV neutralizing antibodies, that had titres comparable with those induced by an inactivated JEV vaccine, as well as haemagglutination-inhibiting antibodies. Mice immunized with 2 x 10(6) infectious units of MVA/JEV recombinants by intramuscular or intraperitoneal routes were completely protected against a 10(5) LD50 JEV challenge at 9 weeks of age. PMID:9987162

Nam, J H; Wyatt, L S; Chae, S L; Cho, H W; Park, Y K; Moss, B

1999-01-21

173

Identification of Non-Nucleoside DNA Synthesis Inhibitors of Vaccinia Virus by High-Throughput Screening  

PubMed Central

Variola virus, the causative agent of smallpox, is a potential bio-weapon. The development of new antiviral compounds for smallpox prophylaxis and treatment is critical, especially since the virus can acquire resistance to the drugs that are currently available. We have identified novel small chemical inhibitors that target DNA synthesis of vaccinia, the prototypical poxvirus. Robotic high-throughput screening of 49,663 compounds and follow-up studies identified very potent inhibitors of vaccinia DNA synthesis, with IC50 values as low as 0.5 ?M. Cell-based assays showed that 16 inhibitors effectively blocked vaccinia infection with minimal cytotoxicity. Three inhibitors had selectivity indexes that approximate that of cidofovir. These new non-nucleoside inhibitors are expected to interfere with components of the vaccinia DNA synthesis apparatus that are distinct from cidofovir. Based on the high sequence similarity between the proteins of vaccinia and variola viruses, these new inhibitors are anticipated to be equally effective against smallpox.

Ciustea, Mihai; Silverman, Janice Elaine Y.; Shudofsky, Abigail M. Druck; Ricciardi, Robert P.

2009-01-01

174

Lister strain of vaccinia virus armed with endostatin-angiostatin fusion gene as a novel therapeutic agent for human pancreatic cancer  

PubMed Central

Summary Survival following pancreatic cancer remains poor despite incremental advances in surgical and adjuvant therapy, and new strategies for treatment are needed. Oncolytic virotherapy is an attractive approach for cancer treatment. In this study, we have evaluated the effectiveness of the Lister vaccine strain of vaccinia virus armed with the endostatin-angiostatin fusion gene (VVhEA) as a novel therapeutic approach for pancreatic cancer. The Lister vaccine strain of vaccinia virus was effective against all human pancreatic carcinoma cells tested in vitro, especially those insensitive to oncolytic adenovirus. The virus displayed inherently high selectivity for cancer cells, sparing normal cells both in vitro and in vivo, with effective infection of tumors after both intravenous (IV) and intratumoral (IT) administration. The expression of endostatin-angiostatin fusion protein was confirmed in a pancreatic cancer model both in vitro and in vivo, with evidence of inhibition of angiogenesis. This novel vaccinia virus demonstrated significant antitumor potency in vivo against the Suit-2 model by IT administration. The present study suggests that the novel Lister strain of vaccinia virus armed with the endostatin-angiostatin fusion gene is a potential therapeutic agent for pancreatic cancer.

Tysome, James R; Briat, Arnaud; Alusi, Ghassan; Cao, Fengyu; Gao, Dongling; Yu, Jinxia; Wang, Pengju; Yang, Shaolong; Dong, Ziming; Wang, Shengdian; Deng, Liufu; Francis, Jennelle; Timiryasova, Tatyana; Fodor, Istvan; Lemoine, Nick R; Wang, Yaohe

2009-01-01

175

Molecular Chaperone Hsp90 Is Important for Vaccinia Virus Growth in Cells  

Microsoft Academic Search

Molecular chaperones assist protein folding, and some chaperones are induced by heat, nutrient depletion, or pathogen invasion. This study investigates the role played by Hsp90 in the life cycle of vaccinia virus. The titer of vaccinia intracellular mature virions (IMV) was reduced by 2 orders of magnitude in RK13 cells treated with geldanamycin (GA), which blocks the ATPase activity of

Jan-Jong Hung; Che-Sheng Chung; Wen Chang

2002-01-01

176

Evidence for Protection against Chronic Hepatitis C Virus Infection in Chimpanzees by Immunization with Replicating Recombinant Vaccinia Virus?  

PubMed Central

Given the failures of nonreplicating vaccines against chronic hepatitis C virus (HCV) infection, we hypothesized that a replicating viral vector may provide protective immunity. Four chimpanzees were immunized transdermally twice with recombinant vaccinia viruses (rVV) expressing HCV genes. After challenge with 24 50% chimpanzee infective doses of homologous HCV, the two control animals that had received only the parental VV developed chronic HCV infection. All four immunized animals resolved HCV infection. The difference in the rate of chronicity between the immunized and the control animals was close to statistical significance (P = 0.067). Immunized animals developed vigorous gamma interferon enzyme-linked immunospot responses and moderate proliferative responses. To investigate cross-genotype protection, the immunized recovered chimpanzees were challenged with a pool of six major HCV genotypes. During the acute phase after the multigenotype challenge, all animals had high-titer viremia in which genotype 4 dominated (87%), followed by genotype 5 (13%). However, after fluctuating low-level viremia, the viremia finally turned negative or persisted at very low levels. This study suggests the potential efficacy of replicating recombinant vaccinia virus-based immunization against chronic HCV infection.

Youn, Jin-Won; Hu, Yu-Wen; Tricoche, Nancy; Pfahler, Wolfram; Shata, Mohamed Tarek; Dreux, Marlene; Cosset, Francois-Loic; Folgori, Antonella; Lee, Dong-Hun; Brotman, Betsy; Prince, Alfred M.

2008-01-01

177

Vaccinia Virus Activation of CCR5 Invokes Tyrosine Phosphorylation Signaling Events That Support Virus Replication  

PubMed Central

Vaccinia virus, a poxvirus, produces structurally distinct forms of virions for which the immediate events following cell entry are ill-defined. We provide evidence that intracellular mature virus (IMV) enters both permissive and nonpermissive T-cell lines and that introduction of CCR5 into nonpermissive mouse fibroblasts or human primary T cells renders the cells permissive for vaccinia replication. Notably, T cells expressing CCR5 in which tyrosine 339 in the intracellular region is replaced by phenylalanine no longer support virus replication or virus-inducible activation of specific host cell signaling effectors IRS-2, Grb2, and Erk1/2. We show that following IMV entry into the cell, the intact but not the tyrosine-deficient CCR5 is rapidly internalized and colocalizes with virus. This colocalization precedes virus-inducible signaling and replication.

Rahbar, Ramtin; Murooka, Thomas T.; Hinek, Anna A.; Galligan, Carole L.; Sassano, Antonella; Yu, Celeste; Srivastava, Kishore; Platanias, Leonidas C.; Fish, Eleanor N.

2006-01-01

178

Vaccinia virus activation of CCR5 invokes tyrosine phosphorylation signaling events that support virus replication.  

PubMed

Vaccinia virus, a poxvirus, produces structurally distinct forms of virions for which the immediate events following cell entry are ill-defined. We provide evidence that intracellular mature virus (IMV) enters both permissive and nonpermissive T-cell lines and that introduction of CCR5 into nonpermissive mouse fibroblasts or human primary T cells renders the cells permissive for vaccinia replication. Notably, T cells expressing CCR5 in which tyrosine 339 in the intracellular region is replaced by phenylalanine no longer support virus replication or virus-inducible activation of specific host cell signaling effectors IRS-2, Grb2, and Erk1/2. We show that following IMV entry into the cell, the intact but not the tyrosine-deficient CCR5 is rapidly internalized and colocalizes with virus. This colocalization precedes virus-inducible signaling and replication. PMID:16809330

Rahbar, Ramtin; Murooka, Thomas T; Hinek, Anna A; Galligan, Carole L; Sassano, Antonella; Yu, Celeste; Srivastava, Kishore; Platanias, Leonidas C; Fish, Eleanor N

2006-07-01

179

Expression of Herpes Simplex Virus 1 Glycoprotein B by a Recombinant Vaccinia Virus and Protection of Mice against Lethal Herpes Simplex Virus 1 Infection  

NASA Astrophysics Data System (ADS)

The herpes simplex virus 1 (HSV-1) strain F gene encoding glycoprotein gB was isolated and modified at the 5' end by in vitro oligonucleotide-directed mutagenesis. The modified gB gene was inserted into the vaccinia virus genome and expressed under the control of a vaccinia virus promoter. The mature gB glycoprotein produced by the vaccinia virus recombinant was glycosylated, was expressed at the cell surface, and was indistinguishable from authentic HSV-1 gB in terms of electrophoretic mobility. Mice immunized intradermally with the recombinant vaccinia virus produced gB-specific neutralizing antibodies and were resistant to a lethal HSV-1 challenge.

Cantin, Edouard M.; Eberle, Richard; Baldick, Joseph L.; Moss, Bernard; Willey, Dru E.; Notkins, Abner L.; Openshaw, Harry

1987-08-01

180

Profile of natural killer cells after a previous natural Vaccinia virus infection in an in vitro viral re-exposure.  

PubMed

The present study compares the profile of NK cells in an in vitro re-exposure by Vaccinia virus (VACV), in groups that have had a previous vaccination or natural infection. Our data suggests that stimulation with VACV triggers a cytotoxic response by NK cells marked by an increase of NCRs: NKp30, NKp44, and NKp46 in infected (vaccinated and unvaccinated) subjects and in non-infected vaccinated patients, when compared with non-infected unvaccinated individuals. However, the degranulation and secretion processes are inhibited in infected (vaccinated and unvaccinated) subjects and in the non-infected vaccinated patients, when compared with non-infected unvaccinated individuals. We demonstrated that stimulation with VACV downregulates the percentage of expression of Perforin, Granzyme A, and CD107a, but upregulate CD94 in infected (vaccinated and unvaccinated) subjects and in non-infected vaccinated patients, when compared with non-infected unvaccinated individuals. Furthermore, the percentage of IFN-?(+) NK cells was significantly lower in non-infected unvaccinated subjects, when compared with infected (vaccinated and unvaccinated) and non-infected vaccinated individuals. Our results also show that the percentage of TNF-?(+) NK cells was significantly higher in infected (vaccinated and unvaccinated) subjects and in non-infected vaccinated patients, when compared with non-infected unvaccinated individuals, after in vitro stimulation with UV-inactivated VACV. Our data suggest that the expression of NCRs NKp30, NKp44, NKp46 and cytokines by NK cells are important in the innate response against VACV. PMID:24530576

Moreira-Silva, Eduardo Augusto Dos Santos; Medeiros-Silva, Daniela Carla; Gomes, Juliana de Assis Silva; Fonseca, Flávio Guimarães da; Correa-Oliveira, Rodrigo

2014-05-01

181

Vaccinia virus strain differences in cell attachment and entry  

SciTech Connect

Vaccinia virus (VACV) strain WR can enter cells by a low pH endosomal pathway or direct fusion with the plasma membrane at neutral pH. Here, we compared attachment and entry of five VACV strains in six cell lines and discovered two major patterns. Only WR exhibited pH 5-enhanced rate of entry following neutral pH adsorption to cells, which correlated with sensitivity to bafilomycin A1, an inhibitor of endosomal acidification. Entry of IHD-J, Copenhagen and Elstree strains were neither accelerated by pH 5 treatment nor prevented by bafilomycin A1. Entry of the Wyeth strain, although not augmented by pH 5, was inhibited by bafilomycin A1. WR and Wyeth were both relatively resistant to the negative effects of heparin on entry, whereas the other strains were extremely sensitive due to inhibition of cell binding. The relative sensitivities of individual vaccinia virus strains to heparin correlated inversely with their abilities to bind to and enter glycosaminoglycan-deficient sog9 cells but not other cell lines tested. These results suggested that that IHD-J, Copenhagen and Elstree have a more limited ability than WR and Wyeth to use the low pH endosomal pathway and are more dependent on binding to glycosaminoglycans for cell attachment.

Bengali, Zain; Townsley, Alan C. [Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 33 North Drive, Bethesda, MD 20892-3210 (United States); Moss, Bernard, E-mail: bmoss@nih.go [Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 33 North Drive, Bethesda, MD 20892-3210 (United States)

2009-06-20

182

Vaccinia virus strain differences in cell attachment and entry.  

PubMed

Vaccinia virus (VACV) strain WR can enter cells by a low pH endosomal pathway or direct fusion with the plasma membrane at neutral pH. Here, we compared attachment and entry of five VACV strains in six cell lines and discovered two major patterns. Only WR exhibited pH 5-enhanced rate of entry following neutral pH adsorption to cells, which correlated with sensitivity to bafilomycin A1, an inhibitor of endosomal acidification. Entry of IHD-J, Copenhagen and Elstree strains were neither accelerated by pH 5 treatment nor prevented by bafilomycin A1. Entry of the Wyeth strain, although not augmented by pH 5, was inhibited by bafilomycin A1. WR and Wyeth were both relatively resistant to the negative effects of heparin on entry, whereas the other strains were extremely sensitive due to inhibition of cell binding. The relative sensitivities of individual vaccinia virus strains to heparin correlated inversely with their abilities to bind to and enter glycosaminoglycan-deficient sog9 cells but not other cell lines tested. These results suggested that that IHD-J, Copenhagen and Elstree have a more limited ability than WR and Wyeth to use the low pH endosomal pathway and are more dependent on binding to glycosaminoglycans for cell attachment. PMID:19428041

Bengali, Zain; Townsley, Alan C; Moss, Bernard

2009-06-20

183

Immunotherapeutic potential of oncolytic vaccinia virus  

Microsoft Academic Search

There has recently been resurgence in interest for the use of replication-selective (oncolytic) viruses for the treatment\\u000a of cancers. This has been fueled by positive clinical data and the promise provided by next-generation vectors that are better\\u000a targeted and display enhanced therapeutic potential. One factor that has led to more effective oncolytic vectors has been\\u000a a greater appreciation of their

Steve H. Thorne

2011-01-01

184

Immunotherapeutic Potential of Oncolytic Vaccinia Virus  

PubMed Central

The concept of oncolytic viral therapy was based on the hypothesis that engineering tumor-selectivity into the replication potential of viruses would permit direct destruction of tumor cells as a result of viral-mediated lysis, resulting in amplification of the therapy exclusively within the tumor environment. The immune response raised by the virus was not only considered to be necessary for the safety of the approach, but also something of a hindrance to optimal therapeutic activity and repeat dosing. However, the pre-clinical and subsequent clinical success of several oncolytic viruses expressing selected cytokines has demonstrated the potential for harnessing the immune response as an additional and beneficial mechanism of therapeutic activity within the platform. Over the last few years, a variety of novel approaches have been incorporated to try to enhance this immunotherapeutic activity. Several innovative and subtle approaches have moved far beyond the expression of a single cytokine transgene, with the hope of optimizing anti-tumor immunity while having minimal detrimental impact on viral oncolytic activity.

Thorne, Steve H.

2014-01-01

185

Group 1 Vaccinia virus zoonotic outbreak in Maranhao State, Brazil.  

PubMed

In Brazil, several exanthematic autochthone Vaccinia virus (VACV) outbreaks affecting dairy cattle and rural workers have been reported since 1999. Although outbreaks had been first described in the Brazilian Southeast, VACV outbreaks were notified in all Brazilian regions in < 10 years. However, in this context, VACV outbreaks had not been described in some Brazilian States, likely because of a lack of notification, or yet unknown epidemiological reasons. Here, we describe the first VACV outbreak in Maranhão State, northeastern Brazil. The virus isolated from this outbreak showed several biological and molecular features that resemble other Group 1 Brazilian VACV, including a deletion signature in the A56R gene. This study raises new questions about diversity and epidemiology of Brazilian VACV. PMID:24166043

Oliveira, Danilo Bretas; Assis, Felipe Lopes; Ferreira, Paulo Cesar Peregrino; Bonjardim, Cláudio Antônio; de Souza Trindade, Giliane; Kroon, Erna Geessien; Abrahão, Jônatas Santos

2013-12-01

186

Immune Modulation in Primary Vaccinia virus Zoonotic Human Infections  

PubMed Central

In 2010, the WHO celebrated the 30th anniversary of the smallpox eradication. Ironically, infections caused by viruses related to smallpox are being increasingly reported worldwide, including Monkeypox, Cowpox, and Vaccinia virus (VACV). Little is known about the human immunological responses elicited during acute infections caused by orthopoxviruses. We have followed VACV zoonotic outbreaks taking place in Brazil and analyzed cellular immune responses in patients acutely infected by VACV. Results indicated that these patients show a biased immune modulation when compared to noninfected controls. Amounts of B cells are low and less activated in infected patients. Although present, T CD4+ cells are also less activated when compared to noninfected individuals, and so are monocytes/macrophages. Similar results were obtained when Balb/C mice were experimentally infected with a VACV sample isolated during the zoonotic outbreaks. Taking together, the data suggest that zoonotic VACVs modulate specific immune cell compartments during an acute infection in humans.

Gomes, Juliana Assis Silva; de Araujo, Fernanda Fortes; Trindade, Giliane de Souza; Quinan, Barbara Resende; Drumond, Betania Paiva; Ferreira, Jaqueline Maria Siqueira; Mota, Bruno Eduardo Fernandes; Nogueira, Mauricio Lacerda; Kroon, Erna Geessien; Abrahao, Jonatas Santos; Correa-Oliveira, Rodrigo; da Fonseca, Flavio Guimaraes

2012-01-01

187

Proteomic Basis of the Antibody Response to Monkeypox Virus Infection Examined in Cynomolgus Macaques and a Comparison to Human Smallpox Vaccination.  

National Technical Information Service (NTIS)

Monkeypox is a zoonotic viral disease that occurs primarily in Central and West Africa. A recent outbreak in the United States heightened public health concerns for susceptible human populations. Vaccinating with vaccinia virus to prevent smallpox is also...

A. Tikhonov, B. Schweitzer, C. Pugh, G. Chen, S. Keasey

2010-01-01

188

The DC receptor DNGR-1 mediates cross-priming of CTLs during vaccinia virus infection in mice.  

PubMed

In order to prime T cells, DCs integrate signals emanating directly from pathogens and from their noxious action on the host. DNGR-1 (CLEC9A) is a DC-restricted receptor that detects dead cells. Therefore, we investigated the possibility that DNGR-1 affects immunity to cytopathic viruses. DNGR-1 was essential for cross-presentation of dying vaccinia virus-infected (VACV-infected) cells to CD8(+) T cells in vitro. Following injection of VACV or VACV-infected cells into mice, DNGR-1 detected the ligand in dying infected cells and mediated cross-priming of anti-VACV CD8(+) T cells. Loss of DNGR-1 impaired the CD8+ cytotoxic response to VACV, especially against those virus strains that are most dependent on cross-presentation. The decrease in total anti-VACV CTL activity was associated with a profound increase in viral load and delayed resolution of the primary lesion. In addition, lack of DNGR-1 markedly diminished protection from infection induced by vaccination with the modified vaccinia Ankara (MVA) strain. DNGR-1 thus contributes to anti-VACV immunity, following both primary infection and vaccination. The non-redundant ability of DNGR-1 to regulate cross-presentation of viral antigens suggests that this form of regulation of antiviral immunity could be exploited for vaccination. PMID:22505455

Iborra, Salvador; Izquierdo, Helena M; Martínez-López, María; Blanco-Menéndez, Noelia; Reis e Sousa, Caetano; Sancho, David

2012-05-01

189

SHORT REPORT: ISOLATION OF TWO VACCINIA VIRUS STRAINS FROM A SINGLE BOVINE VACCINIA OUTBREAK IN RURAL AREA FROM BRAZIL: IMPLICATIONS ON THE EMERGENCE OF ZOONOTIC ORTHOPOXVIRUSES  

Microsoft Academic Search

Abstract. Outbreaks,of bovine,vaccinia disease caused,by circulation of Vaccinia virus (VACV) strains have been a common occurrence in Brazil in the recent years, being an important emergent zoonosis. During a single outbreak that took place in 2001, two genetically different VACV strains were isolated and named Guarani P1 virus (GP1V) and Guarani P2 virus (GP2V). Molecular diagnosis was done,through,restriction fragment,length polymorphism,(RFLP)

Giliane S. Trindade; Zélia I. P. Lobato; Betânia P. Drumond; Juliana A. Leite; Ricardo C. Trigueiro; Maria I. M. C. Guedes; Flávio G. Da Fonseca; João R. Dos Santos; Cláudio A. Bonjardim; Paulo C. P. Ferreira; Erna G. Kroon

190

[Synthesis and antiviral evaluation against Vaccinia virus of new N¹-oxide analogues of 5'-noraristeromycin].  

PubMed

New N¹-benzyl esters of N¹-oxide analogues of 5'-noraristeromycin were synthesized and tested as potential inhibitors of S-adenosyl-L-homocysteine hydrolase in Vaccinia virus infected cell systems. PMID:21317946

Matiugina, E S; Seley-Radtke, K L; Andronova, V L; Galegov, G A; Kochetkov, S N; Khandazhinskaia, A L

2010-01-01

191

Blocking of Production and Activity of Interferon by an Inhibitor Induced by Vaccinia Virus.  

National Technical Information Service (NTIS)

When monolayer cultures of chicken fibroblasts are infected with a used strain of vaccinia virus, the production of an inhibitor blocking the formation and effect of interferon is stimulated. The blocking effect of the inhibitor becomes apparent when it i...

T. A. Bektemirov A. E. Gummenik

1967-01-01

192

Construction of Poxviruses as Cloning Vectors: Insertion of the Thymidine Kinase Gene from Herpes Simplex Virus into the DNA of Infectious Vaccinia Virus  

NASA Astrophysics Data System (ADS)

We have constructed recombinant vaccinia viruses containing the thymidine kinase gene from herpes simplex virus. The gene was inserted into the genome of a variant of vaccinia virus that had undergone spontaneous deletion as well as into the 120-megadalton genome of the large prototypic vaccinia variant. This was accomplished via in vivo recombination by contransfection of eukaryotic tissue culture cells with cloned BamHI-digested thymidine kinase gene from herpes simplex virus containing flanking vaccinia virus DNA sequences and infectious rescuing vaccinia virus. Pure populations of the recombinant viruses were obtained by replica filter techniques or by growth of the recombinant virus in biochemically selective medium. The herpes simplex virus thymidine kinase gene, as an insert in vaccinia virus, is transcribed in vivo and in vitro, and the fidelity of in vivo transcription into a functional gene product was detected by the phosphorylation of 5-[125I]iodo-2'-deoxycytidine.

Panicali, Dennis; Paoletti, Enzo

1982-08-01

193

Expanding the Repertoire of Modified Vaccinia Ankara-Based Vaccine Vectors via Genetic Complementation Strategies  

PubMed Central

Background Modified Vaccinia virus Ankara (MVA) is a safe, highly attenuated orthopoxvirus that is being developed as a recombinant vaccine vector for immunization against a number of infectious diseases and cancers. However, the expression by MVA vectors of large numbers of poxvirus antigens, which display immunodominance over vectored antigens-of-interest for the priming of T cell responses, and the induction of vector-neutralizing antibodies, which curtail the efficacy of subsequent booster immunizations, remain as significant impediments to the overall utility of such vaccines. Thus, genetic approaches that enable the derivation of MVA vectors that are antigenically less complex may allow for rational improvement of MVA-based vaccines. Principal Findings We have developed a genetic complementation system that enables the deletion of essential viral genes from the MVA genome, thereby allowing us to generate MVA vaccine vectors that are antigenically less complex. Using this system, we deleted the essential uracil-DNA-glycosylase (udg) gene from MVA and propagated this otherwise replication-defective variant on a complementing cell line that constitutively expresses the poxvirus udg gene and that was derived from a newly identified continuous cell line that is permissive for growth of wild type MVA. The resulting virus, MVA?udg, does not replicate its DNA genome or express late viral gene products during infection of non-complementing cells in culture. As proof-of-concept for immunological ‘focusing’, we demonstrate that immunization of mice with MVA?udg elicits CD8+ T cell responses that are directed against a restricted repertoire of vector antigens, as compared to immunization with parental MVA. Immunization of rhesus macaques with MVA?udg-gag, a udg? recombinant virus that expresses an HIV subtype-B consensus gag transgene, elicited significantly higher frequencies of Gag-specific CD8 and CD4 T cells following both primary (2–4-fold) and booster (2-fold) immunizations as compared to the udg+ control virus MVA-gag, as determined by intracellular cytokine assay. In contrast, levels of HIV Gag-specific antibodies were elicited similarly in macaques following immunization with MVA?udg-gag and MVA-gag. Furthermore, both udg? and udg+ MVA vectors induced comparatively similar titers of MVA-specific neutralizing antibody responses following immunization of mice (over a 4-log range: 104–108 PFU) and rhesus macaques. These results suggest that the generation of MVA-specific neutralizing antibody responses are largely driven by input MVA antigens, rather than those that are synthesized de novo during infection, and that the processes governing the generation of antiviral antibody responses are more readily saturated by viral antigen than are those that elicit CD8+ T cell responses. Significance Our identification of a spontaneously-immortalized (but not transformed) chicken embryo fibroblast cell line (DF-1) that is fully permissive for MVA growth and that can be engineered to stably express MVA genes provides the basis for a genetic system for MVA. DF-1 cells (and derivatives thereof) constitute viable alternatives, for the manufacture of MVA-based vaccines, to primary CEFs – the conventional cell substrate for MVA vaccines that is not amenable to genetic complementation strategies due to these cells' finite lifespan in culture. The establishment of a genetic system for MVA, as illustrated here to allow udg deletion, enables the generation of novel replication-defective MVA mutants and expands the repertoire of genetic viral variants that can now be explored as improved vaccine vectors.

Garber, David A.; O'Mara, Leigh A.; Zhao, Jun; Gangadhara, Sailaja; An, InChul; Feinberg, Mark B.

2009-01-01

194

Vaccinia Virus Infection Attenuates Innate Immune Responses and Antigen Presentation by Epidermal Dendritic Cells  

Microsoft Academic Search

Langerhans cells (LCs) are antigen-presenting cells in the skin that play sentinel roles in host immune defense by secreting proinflammatory molecules and activating T cells. Here we studied the interaction of vaccinia virus with XS52 cells, a murine epidermis-derived dendritic cell line that serves as a surrogate model for LCs. We found that vaccinia virus productively infects XS52 cells, yet

Liang Deng; Peihong Dai; Wanhong Ding; Richard D. Granstein; Stewart Shuman

2006-01-01

195

Comparison on Virulence and Immunogenicity of Two Recombinant Vaccinia Vaccines, Tian Tan and Guang9 Strains, Expressing the HIV-1 Envelope Gene  

PubMed Central

Background The vaccinia virus Guang9 strain (VG9), derived from the vaccinia virus Tian Tan strain (VTT) has been found to be less virulent than VTT. Methodology/Principal Findings To investigate whether VG9 could be a potential replicating virus vector, the TK genes in VG9 and VTT were replaced with the HIV-1 envelope gene via homologous recombination, resulting in the recombinant viruses, VG9-E and VTT-E. The biology, virulence, humoral and cellular immunological responses of VG9-E and VTT-E were evaluated. Our results indicated no obvious difference in range of host cells and diffusion between two recombinant viruses. Neurovirulence for VG9-E in weanling and suckling mice, and skin virulence in rabbits, were lower than that of VTT-E. The humoral immune responses, including binding antibody and neutralizing antibody responses, induced by VG9-E were not significantly different from those for VTT-E whilst IFN-? response which represented cellular immune response induced by VG9-E was significantly higher than that did by VTT-E. Conclusions/Significance Our results indicated that VG9-E was less virulent, yet induced higher cellular immune response than VTT-E. Therefore, it could be an ideal replicating vaccinia vector for HIV vaccine research and development.

Zhu, Rong; Huang, Weijin; Wang, Wenbo; Liu, Qiang; Nie, Jianhui; Meng, Shufang; Yu, Yongxin; Wang, Youchun

2012-01-01

196

Intravenous Inoculation of Replication-Deficient Recombinant Vaccinia Virus DIs Expressing Simian Immunodeficiency Virus Gag Controls Highly Pathogenic Simian-Human Immunodeficiency Virus in Monkeys  

PubMed Central

To be effective, a vaccine against human immunodeficiency virus type 1 (HIV-1) must induce virus-specific T-cell responses and it must be safe for use in humans. To address these issues, we developed a recombinant vaccinia virus DIs vaccine (rDIsSIVGag), which is nonreplicative in mammalian cells and expresses the full-length gag gene of simian immunodeficiency virus (SIV). Intravenous inoculation of 106 PFU of rDIsSIVGag in cynomologus macaques induced significant levels of gamma interferon (IFN-?) spot-forming cells (SFC) specific for SIV Gag. Antigen-specific lymphocyte proliferative responses were also induced and were temporally associated with the peak of IFN-? SFC activity in each macaque. In contrast, macaques immunized with a vector control (rDIsLacZ) showed no significant induction of antigen-specific immune responses. After challenge with a highly pathogenic simian-human immunodeficiency virus (SHIV), CD4+ T lymphocytes were maintained in the peripheral blood and lymphoid tissues of the immunized macaques. The viral set point in plasma was also reduced in these animals, which may be related to the enhancement of virus-specific intracellular IFN-?+ CD8+ cell numbers and increased antibody titers after SHIV challenge. These results demonstrate that recombinant DIs has potential for use as an HIV/AIDS vaccine.

Izumi, Yasuyuki; Ami, Yasushi; Matsuo, Kazuhiro; Someya, Kenji; Sata, Tetsutaro; Yamamoto, Naoki; Honda, Mitsuo

2003-01-01

197

Coated microneedle arrays for transcutaneous delivery of live virus vaccines.  

PubMed

Vaccines are sensitive biologics that require continuous refrigerated storage to maintain their viability. The vast majority of vaccines are also administered using needles and syringes. The need for cold chain storage and the significant logistics surrounding needle-and-syringe vaccination is constraining the success of immunization programs. Recombinant live viral vectors are a promising platform for the development of vaccines against a number of infectious diseases, however these viruses must retain infectivity to be effective. Microneedles offer an effective and painless method for delivery of vaccines directly into skin that in the future could provide solutions to current vaccination issues. Here we investigated methods of coating live recombinant adenovirus and modified vaccinia virus Ankara (MVA) vectors onto solid microneedle arrays. An effective spray-coating method, using conventional pharmaceutical processes, was developed, in tandem with suitable sugar-based formulations, which produces arrays with a unique coating of viable virus in a dry form around the shaft of each microneedle on the array. Administration of live virus-coated microneedle arrays successfully resulted in virus delivery, transcutaneous infection and induced an antibody or CD8(+) T cell response in mice that was comparable to that obtained by needle-and-syringe intradermal immunization. To our knowledge, this is the first report of successful vaccination with recombinant live viral vectored vaccines coated on microneedle delivery devices. PMID:22245683

Vrdoljak, Anto; McGrath, Marie G; Carey, John B; Draper, Simon J; Hill, Adrian V S; O'Mahony, Conor; Crean, Abina M; Moore, Anne C

2012-04-10

198

Novel vaccines against influenza viruses  

PubMed Central

Killed and live attenuated influenza virus vaccines are effective in preventing and curbing the spread of influenza epidemics when the strains present in the vaccines are closely matched with the predicted epidemic strains. These vaccines are primarily targeted to induce immunity to the variable major target antigen, hemagglutinin (HA) of influenza virus. However, current vaccines are not effective in preventing the emergence of new pandemic or highly virulent viruses. New approaches are being investigated to develop universal influenza virus vaccines as well as to apply more effective vaccine delivery methods. Conserved vaccine targets including the influenza M2 ion channel protein and HA stalk domains are being developed using recombinant technologies to improve the level of cross protection. In addition, recent studies provide evidence that vaccine supplements can provide avenues to further improve current vaccination.

Kang, Sang-Moo; Song, Jae-Min; Compans, Richard W.

2011-01-01

199

Robust Intrapulmonary CD8 T Cell Responses and Protection with an Attenuated N1L Deleted Vaccinia Virus  

PubMed Central

Background Vaccinia viruses have been used as a model for viral disease and as a protective live vaccine. Methodology and Principal Findings We investigated the immunogenicity of an attenuated strain of vaccinia virus engineered to inactivate the N1L gene (vGK5). Using the intranasal route, this recombinant virus was 2 logs less virulent compared to the wildtype VACV-WR. Infection by the intranasal, intraperitoneal, and tail scarification routes resulted in the robust induction of cytolytic virus-specific CD8 T cells in the spleens and the lungs. VACV-specific antibodies were also detected in the sera of mice infected 3–5 months prior with the attenuated vGK5 virus. Finally, mice immunized with vGK5 were significantly protected when challenged with a lethal dose of VACV-WR. Conclusions These results indicate that the attenuated vGK5 virus protects against subsequent infection and suggest that the N1L protein limits the strength of the early antiviral CD8 T cell response following respiratory infection.

Mathew, Anuja; O'Bryan, Joel; Marshall, William; Kotwal, Girish J.; Terajima, Masanori; Green, Sharone; Rothman, Alan L.; Ennis, Francis A.

2008-01-01

200

Anti-metastatic effect of oncolysates from murine melanoma cells transfected with recombinant vaccinia virus encoding human IL2  

Microsoft Academic Search

Oncolysates, debris of tumor cells, have been proven to be effective in active immunotherapy of cancer. In this experiment,\\u000a the oncolysates from murine melanoma cells B16-F10 transfected by recombinant vaccinia viruses encoding human IL-2(IL-2VBO)\\u000a were used as vaccine. After treatment of tumor bearing mice with pulmonary metastases by intravenous injection of IL-2VBO\\u000a or rVV-IL-2, higher level IL-2 activity was detected

Tao Wan; Xuetao Cao; Dianwen Ju; Yizhi Yu; Qun Tao; Hong Lei

1997-01-01

201

Induction of multifunctional human immunodeficiency virus type 1 (HIV-1)-specific T cells capable of proliferation in healthy subjects by using a prime-boost regimen of DNA- and modified vaccinia virus Ankara-vectored vaccines expressing HIV-1 Gag coupled to CD8+ T-cell epitopes.  

PubMed

A double-blind randomized phase I trial was conducted in human immunodeficiency virus type 1 (HIV-1)-negative subjects receiving vaccines vectored by plasmid DNA and modified vaccinia virus Ankara (MVA) expressing HIV-1 p24/p17 gag linked to a string of CD8(+) T-cell epitopes. The trial had two groups. One group received either two doses of MVA.HIVA (2x MVA.HIVA) (n=8) or two doses of placebo (2x placebo) (n=4). The second group received 2x pTHr.HIVA followed by one dose of MVA.HIVA (n=8) or 3x placebo (n=4). In the pTHr.HIVA-MVA.HIVA group, HIV-1-specific T-cell responses peaked 1 week after MVA.HIVA vaccination in both ex vivo gamma interferon (IFN-gamma) ELISPOT (group mean, 210 spot-forming cells/10(6) cells) and proliferation (group mean stimulation index, 37), with assays detecting positive responses in four out of eight and five out of eight subjects, respectively. No HIV-1-specific T-cell responses were detected in either assay in the 2x MVA.HIVA group or subjects receiving placebo. Using a highly sensitive and reproducible cultured IFN-gamma ELISPOT assay, positive responses mainly mediated by CD4(+) T cells were detected in eight out of eight vaccinees in the pTHr.HIVA-MVA.HIVA group and four out of eight vaccinees in the 2x MVA.HIVA group. Importantly, no false-positive responses were detected in the eight subjects receiving placebo. Of the 12 responders, 11 developed responses to previously identified immunodominant CD4(+) T-cell epitopes, with 6 volunteers having responses to more than one epitope. Five out of 12 responders also developed CD8(+) T-cell responses to the epitope string. Induced T cells produced a variety of anti-viral cytokines, including tumor necrosis factor alpha and macrophage inflammatory protein 1 beta. These data demonstrate that prime-boost vaccination with recombinant DNA and MVA vectors can induce multifunctional HIV-1-specific T cells in the majority of vaccinees. PMID:16641265

Goonetilleke, Nilu; Moore, Stephen; Dally, Len; Winstone, Nicola; Cebere, Inese; Mahmoud, Abdul; Pinheiro, Susana; Gillespie, Geraldine; Brown, Denise; Loach, Vanessa; Roberts, Joanna; Guimaraes-Walker, Ana; Hayes, Peter; Loughran, Kelley; Smith, Carole; De Bont, Jan; Verlinde, Carl; Vooijs, Danii; Schmidt, Claudia; Boaz, Mark; Gilmour, Jill; Fast, Pat; Dorrell, Lucy; Hanke, Tomas; McMichael, Andrew J

2006-05-01

202

Induction of Multifunctional Human Immunodeficiency Virus Type 1 (HIV-1)-Specific T Cells Capable of Proliferation in Healthy Subjects by Using a Prime-Boost Regimen of DNA- and Modified Vaccinia Virus Ankara-Vectored Vaccines Expressing HIV-1 Gag Coupled to CD8+ T-Cell Epitopes†  

PubMed Central

A double-blind randomized phase I trial was conducted in human immunodeficiency virus type 1 (HIV-1)-negative subjects receiving vaccines vectored by plasmid DNA and modified vaccinia virus Ankara (MVA) expressing HIV-1 p24/p17 gag linked to a string of CD8+ T-cell epitopes. The trial had two groups. One group received either two doses of MVA.HIVA (2× MVA.HIVA) (n = 8) or two doses of placebo (2× placebo) (n = 4). The second group received 2× pTHr.HIVA followed by one dose of MVA.HIVA (n = 8) or 3× placebo (n = 4). In the pTHr.HIVA-MVA.HIVA group, HIV-1-specific T-cell responses peaked 1 week after MVA.HIVA vaccination in both ex vivo gamma interferon (IFN-?) ELISPOT (group mean, 210 spot-forming cells/106 cells) and proliferation (group mean stimulation index, 37), with assays detecting positive responses in four out of eight and five out of eight subjects, respectively. No HIV-1-specific T-cell responses were detected in either assay in the 2× MVA.HIVA group or subjects receiving placebo. Using a highly sensitive and reproducible cultured IFN-? ELISPOT assay, positive responses mainly mediated by CD4+ T cells were detected in eight out of eight vaccinees in the pTHr.HIVA-MVA.HIVA group and four out of eight vaccinees in the 2× MVA.HIVA group. Importantly, no false-positive responses were detected in the eight subjects receiving placebo. Of the 12 responders, 11 developed responses to previously identified immunodominant CD4+ T-cell epitopes, with 6 volunteers having responses to more than one epitope. Five out of 12 responders also developed CD8+ T-cell responses to the epitope string. Induced T cells produced a variety of anti-viral cytokines, including tumor necrosis factor alpha and macrophage inflammatory protein 1?. These data demonstrate that prime-boost vaccination with recombinant DNA and MVA vectors can induce multifunctional HIV-1-specific T cells in the majority of vaccinees.

Goonetilleke, Nilu; Moore, Stephen; Dally, Len; Winstone, Nicola; Cebere, Inese; Mahmoud, Abdul; Pinheiro, Susana; Gillespie, Geraldine; Brown, Denise; Loach, Vanessa; Roberts, Joanna; Guimaraes-Walker, Ana; Hayes, Peter; Loughran, Kelley; Smith, Carole; De Bont, Jan; Verlinde, Carl; Vooijs, Danii; Schmidt, Claudia; Boaz, Mark; Gilmour, Jill; Fast, Pat; Dorrell, Lucy; Hanke, Tomas; McMichael, Andrew J.

2006-01-01

203

Identification of Novel Antipoxviral Agents: Mitoxantrone Inhibits Vaccinia Virus Replication by Blocking Virion Assembly?  

PubMed Central

The bioterror threat of a smallpox outbreak in an unvaccinated population has mobilized efforts to develop new antipoxviral agents. By screening a library of known drugs, we identified 13 compounds that inhibited vaccinia virus replication at noncytotoxic doses. The anticancer drug mitoxantrone is unique among the inhibitors identified in that it has no apparent impact on viral gene expression. Rather, it blocks processing of viral structural proteins and assembly of mature progeny virions. The isolation of mitoxantrone-resistant vaccinia strains underscores that a viral protein is the likely target of the drug. Whole-genome sequencing of mitoxantrone-resistant viruses pinpointed missense mutations in the N-terminal domain of vaccinia DNA ligase. Despite its favorable activity in cell culture, mitoxantrone administered intraperitoneally at the maximum tolerated dose failed to protect mice against a lethal intranasal infection with vaccinia virus.

Deng, Liang; Dai, Peihong; Ciro, Anthony; Smee, Donald F.; Djaballah, Hakim; Shuman, Stewart

2007-01-01

204

Differences in Virus-Induced Cell Morphology and in Virus Maturation between MVA and Other Strains (WR, Ankara, and NYCBH) of Vaccinia Virus in Infected Human Cells  

PubMed Central

Live recombinants based on attenuated modified vaccinia virus Ankara (MVA) are potential vaccine candidates against a broad spectrum of diseases and tumors. To better understand the efficacy of MVA as a human vaccine, we analyzed by confocal and electron microscopy approaches MVA-induced morphological changes and morphogenetic stages during infection of human HeLa cells in comparison to other strains of vaccinia virus (VV): the wild-type Western Reserve (WR), Ankara, and the New York City Board of Health (NYCBH) strains. Confocal microscopy studies revealed that MVA infection alters the cytoskeleton producing elongated cells (bipolar), which do not form the characteristic actin tails. Few virions are detected in the projections connecting neighboring cells. In contrast, cells infected with the WR, Ankara, and NYCBH strains exhibit a stellated (multipolar) or rounded morphology with actin tails. A detailed transmission electron microscopy analysis of HeLa cells infected with MVA showed important differences in fine ultrastructure and amounts of the viral intermediates compared to cells infected with the other VV strains. In HeLa cells infected with MVA, the most abundant viral forms are intracellular immature virus, with few intermediates reaching the intracellular mature virus (IMV) form, at various stages of maturation, which exhibit a more rounded shape than IMVs from cells infected with the other VV strains. The “IMVs” from MVA-infected cells have an abnormal internal structure (“atypical” viruses) with potential alterations in the core-envelope interactions and are unable to significantly acquire the additional double envelope to render intracellular envelope virus. The presence of potential cell-associated envelope virus is very scarce. Our findings revealed that MVA in human cells promotes characteristic morphological changes to the cells and is able to reach the IMV stage, but these virions were not structurally normal and the subsequent steps in the morphogenetic pathway are blocked.

Gallego-Gomez, Juan Carlos; Risco, Cristina; Rodriguez, Dolores; Cabezas, Pilar; Guerra, Susana; Carrascosa, Jose L.; Esteban, Mariano

2003-01-01

205

A comparison of the antigens present on the surface of virus released artificially from chick cells infected with vaccinia virus, and cowpox virus and its white pock mutant  

PubMed Central

Antisera prepared against vaccinia and cowpox viruses were absorbed with purified suspensions of vaccinia virus, red cowpox and white cowpox viruses. They were then tested for their ability to neutralize the viruses, and to precipitate the virus soluble antigens. The results showed that some virus specific antigens were not virus surface components and that some components were present on the surface of all three viruses. However, certain components were detected on the surface of vaccinia virus but not on the surface of cowpox virus, and vice versa. Some evidence for the existence of a vaccinia-specific surface component was also obtained. Comparisons between results of cross-neutralization tests and immunodiffusion tests on the absorbed sera indicated that antibody to a number of antigens, including the classical LS, and the cowpox-specific d antigen play no part in the process of poxvirus neutralization. ImagesFig. AFig. BFig. CFig. DFig. EFig. FFig. G

Baxby, Derrick

1972-01-01

206

Vaccinia virus exhibits cell-type-dependent entry characteristics  

PubMed Central

Differing and sometimes conflicting data have been reported regarding several aspects of vaccinia virus (VV) entry. To address this, we used a ?-galactosidase reporter virus to monitor virus entry into multiple cell types under varying conditions. Entry into HeLa, B78H1 and L cells was strongly inhibited by heparin whereas entry into Vero and BSC-1 cells was unaffected. Bafilomycin also exhibited variable and cell-type-specific effects on VV entry. Entry into B78H1 and BSC-1 cells was strongly inhibited by bafilomycin whereas entry into Vero and HeLa cells was only partially inhibited suggesting the co-existence of both pH-dependent and pH-independent VV entry pathways in these cell types. Finally, entry into HeLa, B78H1, L and BSC-1 cells exhibited a lag of 6–9 min whereas this delay was undetectable in Vero cells. Our results suggest that VV exploits multiple cell attachment and entry pathways allowing it to infect a broad range of cells.

Whitbeck, J. Charles; Foo, Chwan-Hong; de Leon, Manuel Ponce; Eisenberg, Roselyn J.; Cohen, Gary H.

2014-01-01

207

Elucidating and Minimizing the Loss by Recombinant Vaccinia Virus of Human Immunodeficiency Virus Gene Expression Resulting from Spontaneous Mutations and Positive Selection ?  

PubMed Central

While characterizing modified vaccinia virus recombinants (rMVAs) containing human immunodeficiency virus env and gag-pol genes, we detected nonexpressing mutants by immunostaining individual plaques. In many cases, the numbers of mutants increased during successive passages, indicating strong selection pressure. This phenomenon provided an opportunity to investigate the formation of spontaneous mutations in vaccinia virus, which encodes its own cytoplasmic replication system, and a challenge to reduce the occurrence of mutations for vaccine production. Analysis of virus from individual plaques indicated that loss of expression was due to frameshift mutations, mostly by addition or deletion of a single nucleotide in runs of four to six Gs or Cs, and large deletions that included MVA DNA flanking the recombinant gene. Interruption of the runs of Gs and Cs by silent codon alterations and moving the recombinant gene to a site between essential, highly conserved MVA genes eliminated or reduced frameshifts and viable deletion mutants, respectively. The rapidity at which nonexpressing mutants accumulated depended on the individual env and gag-pol genes and their suppressive effects on virus replication. Both the extracellular and transmembrane domains contributed to the selection of nonexpressing Env mutants. Stability of an unstable Env was improved by swapping external or transmembrane domains with a more stable Env. Most dramatically, removal of the transmembrane and cytoplasmic domains stabilized even the most highly unstable Env. Understanding the causes of instability and taking preemptive actions will facilitate the development of rMVA and other poxviruses as human and veterinary recombinant vaccines.

Wyatt, Linda S.; Earl, Patricia L.; Xiao, Wei; Americo, Jeffrey L.; Cotter, Catherine A.; Vogt, Jennifer; Moss, Bernard

2009-01-01

208

Novel avian influenza virus vaccines  

Microsoft Academic Search

Summary Current vaccines against avian influenza (AI) virus infections are primarily based on classical inactivated whole-virus preparations. Although administration of these vaccines can protect poultry from clinical disease, sterile immunity is not achieved under field conditions, allowing for undetected virus spread and evolution under immune cover. Therefore, there is an urgent need for a robust and reliable system of differentiation

W. Fuchs; A. Römer-Oberdörfer; J. Veits; T. C. Mettenleiter

2009-01-01

209

Characterization of Chimpanzee\\/Human Monoclonal Antibodies to Vaccinia Virus A33 Glycoprotein and Its Variola Virus Homolog In Vitro and in a Vaccinia Virus Mouse Protection Model  

Microsoft Academic Search

Three distinct chimpanzee Fabs against the A33 envelope glycoprotein of vaccinia virus were isolated and converted into complete monoclonal antibodies (MAbs) with human 1 heavy-chain constant regions. The three MAbs (6C, 12C, and 12F) displayed high binding affinities to A33 (Kd of 0.14 nM to 20 nM) and may recognize the same epitope, which was determined to be conformational and

Zhaochun Chen; Patricia Earl; Jeffrey Americo; Inger Damon; Scott K. Smith; Fujuan Yu; Andrew Sebrell; Suzanne Emerson; Gary Cohen; Roselyn J. Eisenberg; Inna Gorshkova; Peter Schuck; William Satterfield; Bernard Moss; Robert Purcell

2007-01-01

210

Vaccine strategies against Babesia bovis based on prime-boost immunizations in mice with modified vaccinia Ankara vector and recombinant proteins.  

PubMed

In this study, a recombinant modified vaccinia virus Ankara vector expressing a chimeric multi-antigen was obtained and evaluated as a candidate vaccine in homologous and heterologous prime-boost immunizations with a recombinant protein cocktail. The chimeric multi-antigen comprises immunodominant B and T cell regions of three Babesia bovis proteins. Humoral and cellular immune responses were evaluated in mice to compare the immunogenicity induced by different immunization schemes. The best vaccination scheme was achieved with a prime of protein cocktail and a boost with the recombinant virus. This scheme induced high level of specific IgG antibodies and secreted IFN and a high degree of activation of IFN?(+) CD4(+) and CD8(+) specific T cells. This is the first report in which a novel vaccine candidate was constructed based on a rationally designed multi-antigen and evaluated in a prime-boost regime, optimizing the immune response necessary for protection against bovine babesiosis. PMID:24968152

Jaramillo Ortiz, José Manuel; Del Médico Zajac, María Paula; Zanetti, Flavia Adriana; Molinari, María Paula; Gravisaco, María José; Calamante, Gabriela; Wilkowsky, Silvina Elizabeth

2014-08-01

211

Safety, Immunogenicity and Efficacy of Modified Vaccinia Ankara (MVA) Against Dryvax® Challenge in Vaccinia-Na?ve and Vaccinia-Immune Individuals  

PubMed Central

Modified vaccinia Ankara (MVA) was evaluated as an alternative to Dryvax® in vaccinia-naïve and immune adult volunteers. Subjects received intramuscular MVA or placebo followed by Dryvax® challenge at 3 months. Two or more doses of MVA prior to Dryvax® reduced severity of lesion formation, decreased magnitude and duration of viral shedding, and augmented post-Dryvax® vaccinia-specific CD8+ T cell responses and extracellular enveloped virus protein-specific antibody responses. MVA vaccination is safe and immunogenic and improves the safety and immunogenicity of subsequent Dryvax® vaccination supporting the potential for using MVA as a vaccine in the general population to improve immunity to orthopoxviruses.

Parrino, Janie; McCurdy, Lewis H.; Larkin, Brenda D.; Gordon, Ingelise J.; Rucker, Steven E.; Enama, Mary E.; Koup, Richard A.; Roederer, Mario; Bailer, Robert T.; Moodie, Zoe; Gu, Lin; Yan, Lihan; Graham, Barney S.

2007-01-01

212

Cowpox virus but not Vaccinia virus induces secretion of CXCL1, IL-8 and IL-6 and chemotaxis of monocytes in vitro.  

PubMed

Orthopoxviruses are large DNA viruses which can cause disease in numerous host species. Today, after eradication of Variola virus and the end of vaccination against smallpox, zoonotic Orthopoxvirus infections are emerging as potential threat to human health. The most common causes of zoonotic Orthopoxvirus infections are Cowpox virus in Europe, Monkeypox virus in Africa and Vaccinia virus in South America. Although all three viruses are genetically and antigenically closely related, the human diseases caused by each virus differ considerably. This observation may reflect different capabilities of these viruses to modulate the hosts' immune response. Therefore, we aimed at characterizing the specific cytokine response induced by Orthopoxvirus infection in vitro. We analysed the gene expression of nine human pro-inflammatory cytokines and chemokines in response to infection of HeLa cells and could identify an upregulation of cytokine gene expression following Cowpox virus and Monkeypox virus infection but not following Vaccinia virus infection. This was verified by a strong induction of especially IL-6, IL-8 and CXCL1 secretion into the cell culture supernatant following Cowpox virus infection. We could further show that supernatants derived from Cowpox virus-infected cells exhibit an increased chemotactic activity towards monocytic and macrophage-like cells. On the one hand, increased cytokine secretion by Cowpox virus-infected cells and subsequent monocyte/macrophage recruitment may contribute to host defence and facilitate clearance of the infection. On the other hand, given the assumed important role of circulating macrophages in viral spread, this may also point towards a mechanism facilitating delivery of the virus to further tissues in vivo. PMID:23207068

Bourquain, Daniel; Nitsche, Andreas

2013-01-01

213

Intrafamilial Transmission of Vaccinia virus during a Bovine Vaccinia Outbreak in Brazil: A New Insight in Viral Transmission Chain.  

PubMed

Bovine vaccinia (BV) is an emerging zoonosis caused by the Vaccinia virus (VACV), genus Orthopoxvirus (OPV), Poxviridae family. In general, human cases are related to direct contact with sick cattle but there is a lack of information about human-to-human transmission of VACV during BV outbreaks. In this study, we epidemiologically and molecularly show a case of VACV transmission between humans in São Francisco de Itabapoana County, Rio de Janeiro state. Our group collected samples from the patients, a 49-year-old patient and his son. Our results showed that patients had developed anti-OPV IgG or IgM antibodies and presented neutralizing antibodies against OPV. The VACV isolates displayed high identity (99.9%) and were grouped in the same phylogenetic tree branch. Our data indicate that human-to-human VACV transmission occurred during a BV outbreak, raising new questions about the risk factors of the VACV transmission chain. PMID:24615135

Pereira Oliveira, Graziele; Tavares Silva Fernandes, André; Lopes de Assis, Felipe; Augusto Alves, Pedro; Moreira Franco Luiz, Ana Paula; Barcelos Figueiredo, Leandra; Costa de Almeida, Cláudia Maria; Pires Ferreira Travassos, Carlos Eurico; de Souza Trindade, Giliane; Santos Abrahão, Jônatas; Geessien Kroon, Erna

2014-06-01

214

DIFFERENTIAL IMMUNOGENICITY OF VACCINIA AND HIV-1 COMPONENTS OF A HUMAN RECOMBINANT VACCINE IN MUCOSAL AND BLOOD COMPARTMENTS  

PubMed Central

Mucosal immune responses induced by HIV-1 vaccines are likely critical for prevention. We report a Phase 1 safety and immunogenicity trial in 8 participants using the vaccinia-based TBC-3B vaccine given subcutaneously to determine the relationship between HIV-1 specific systemic and gastrointestinal mucosal responses. Across all subjects, detectable levels of blood vaccinia- and HIV-1-specific antibodies were elicited but none were seen mucosally. While the vaccinia component was immunogenic for CD8+ T lymphocyte (CTL) responses in both blood and mucosa, it was greater in blood. The HIV-1 component of the vaccine was poorly immunogenic in both blood and mucosa. Although only 8 volunteers were studied intensively, the discordance between mucosal and blood responses may highlight mechanisms contributing to recent vaccine failures.

Anton, Peter A.; Ibarrondo, F Javier; Boscardin, W. John; Zhou, Ying; Schwartz, Elissa J.; Ng, Hwee L.; Hausner, Mary Ann; Shih, Roger; Elliott, Julie; Hultin, Patricia M.; Hultin, Lance E.; Price, Charles; Fuerst, Marie; Adler, Amy; Wong, Johnson T.; Yang, Otto O.; Jamieson, Beth D.

2008-01-01

215

Differential immunogenicity of vaccinia and HIV-1 components of a human recombinant vaccine in mucosal and blood compartments.  

PubMed

Mucosal immune responses induced by HIV-1 vaccines are likely critical for prevention. We report a Phase 1 safety and immunogenicity trial in eight participants using the vaccinia-based TBC-3B vaccine given subcutaneously to determine the relationship between HIV-1 specific systemic and gastrointestinal mucosal responses. Across all subjects, detectable levels of blood vaccinia- and HIV-1-specific antibodies were elicited but none were seen mucosally. While the vaccinia component was immunogenic for CD8(+) T lymphocyte (CTL) responses in both blood and mucosa, it was greater in blood. The HIV-1 component of the vaccine was poorly immunogenic in both blood and mucosa. Although only eight volunteers were studied intensively, the discordance between mucosal and blood responses may highlight mechanisms contributing to recent vaccine failures. PMID:18621451

Anton, Peter A; Ibarrondo, F Javier; Boscardin, W John; Zhou, Ying; Schwartz, Elissa J; Ng, Hwee L; Hausner, Mary Ann; Shih, Roger; Elliott, Julie; Hultin, Patricia M; Hultin, Lance E; Price, Charles; Fuerst, Marie; Adler, Amy; Wong, Johnson T; Yang, Otto O; Jamieson, Beth D

2008-08-18

216

Nigericin is a potent inhibitor of the early stage of vaccinia virus replication.  

PubMed

Poxviruses remain a significant public health concern due to their potential use as bioterrorist agents and the spread of animal borne poxviruses, such as monkeypox virus, to humans. Thus, the identification of small molecule inhibitors of poxvirus replication is warranted. Vaccinia virus is the prototypic member of the Orthopoxvirus genus, which also includes variola and monkeypox virus. In this study, we demonstrate that the carboxylic ionophore nigericin is a potent inhibitor of vaccinia virus replication in several human cell lines. In HeLa cells, we found that the 50% inhibitory concentration of nigericin against vaccinia virus was 7.9 nM, with a selectivity index of 1038. We present data demonstrating that nigericin targets vaccinia virus replication at a post-entry stage. While nigericin moderately inhibits both early vaccinia gene transcription and translation, viral DNA replication and intermediate and late gene expression are severely compromised in the presence of nigericin. Our results demonstrate that nigericin has the potential to be further developed into an effective antiviral to treat poxvirus infections. PMID:20951746

Myskiw, Chad; Piper, Jessica; Huzarewich, Rhiannon; Booth, Tim F; Cao, Jingxin; He, Runtao

2010-12-01

217

Expression of dengue virus structural proteins and nonstructural protein NS1 by a recombinant vaccinia virus.  

PubMed

A recombinant vaccinia virus containing cloned DNA sequences coding for the three structural proteins and nonstructural proteins NS1 and NS2a of dengue type 4 virus was constructed. Infection of CV-1 cells with this recombinant virus produced dengue virus structural proteins as well as the nonstructural protein NS1. These proteins were precipitated by specific antisera and exhibited the same molecular size and glycosylation patterns as authentic dengue virus proteins. Infection of cotton rats with the recombinant virus induced NS1 antibodies in 1 of 11 animals. However, an immune response to the PreM and E glycoproteins was not detected. A reduced level of gene expression was probably the reason for the limited serologic response to these dengue virus antigens. PMID:3316711

Zhao, B T; Prince, G; Horswood, R; Eckels, K; Summers, P; Chanock, R; Lai, C J

1987-12-01

218

Reverse Genetics of SARS-Related Coronavirus Using Vaccinia Virus-Based Recombination  

PubMed Central

Severe acute respiratory syndrome (SARS) is a zoonotic disease caused by SARS-related coronavirus (SARS-CoV) that emerged in 2002 to become a global health concern. Although the original outbreak was controlled by classical public health measures, there is a real risk that another SARS-CoV could re-emerge from its natural reservoir, either in its original form or as a more virulent or pathogenic strain; in which case, the virus would be difficult to control in the absence of any effective antiviral drugs or vaccines. Using the well-studied SARS-CoV isolate HKU-39849, we developed a vaccinia virus-based SARS-CoV reverse genetic system that is both robust and biosafe. The SARS-CoV genome was cloned in separate vaccinia virus vectors, (vSARS-CoV-5prime and vSARS-CoV-3prime) as two cDNAs that were subsequently ligated to create a genome-length SARS-CoV cDNA template for in vitro transcription of SARS-CoV infectious RNA transcripts. Transfection of the RNA transcripts into permissive cells led to the recovery of infectious virus (recSARS-CoV). Characterization of the plaques produced by recSARS-CoV showed that they were similar in size to the parental SARS-CoV isolate HKU-39849 but smaller than the SARS-CoV isolate Frankfurt-1. Comparative analysis of replication kinetics showed that the kinetics of recSARS-CoV replication are similar to those of SARS-CoV Frankfurt-1, although the titers of virus released into the culture supernatant are approximately 10-fold less. The reverse genetic system was finally used to generate a recSARS-CoV reporter virus expressing Renilla luciferase in order to facilitate the analysis of SARS-CoV gene expression in human dendritic cells (hDCs). In parallel, a Renilla luciferase gene was also inserted into the genome of human coronavirus 229E (HCoV-229E). Using this approach, we demonstrate that, in contrast to HCoV-229E, SARS-CoV is not able to mediate efficient heterologous gene expression in hDCs.

Zevenhoven, Jessika C.; Weber, Friedemann; Zust, Roland; Kuri, Thomas; Dijkman, Ronald; Chang, Guohui; Siddell, Stuart G.; Snijder, Eric J.; Thiel, Volker; Davidson, Andrew D.

2012-01-01

219

The N-terminus of vaccinia virus host range protein C7L is essential for function  

PubMed Central

Vaccinia virus (VACV), a member of the Poxviridae family of large double-stranded DNA viruses, is being used as a smallpox vaccine as well as an expression vector for immunization against other infectious diseases and cancer. The host range of wild type VACV is very broad among mammalian cells. C7L is a host range gene identified in VACV and is well conserved in mammalian poxviruses except for parapoxviruses and molluscum contagiosum virus. The molecular mechanisms by which the C7L gene exerts host range function are not well understood. The C7L protein does not have any known conserved domains or show sequence similarity to cellular proteins or viral proteins other than the C7L homologues in mammalian poxviruses. We generated recombinant vaccinia viruses carrying deletion mutants of the C7L gene using NYVAC as a parental strain and found that the N-terminus is essential for host range function of C7L, which is consistent with a previous report that showed homology among C7L homologues are greater near the N-terminus than the C-terminus.

Terajima, Masanori; Urban, Stina L.; Leporati, Anita M.

2012-01-01

220

Truncation of gene F5L partially masks rescue of vaccinia virus strain MVA growth on mammalian cells by restricting plaque size.  

PubMed

Modified vaccinia virus Ankara (MVA) is a candidate vaccine vector that is severely attenuated due to mutations acquired during several hundred rounds of serial passage in vitro. A previous study used marker rescue to produce a set of MVA recombinants with improved replication on mammalian cells. Here, we extended the characterization of these rescued MVA strains and identified vaccinia virus (VACV) gene F5L as a determinant of plaque morphology but not replication in vitro. F5 joins a growing group of VACV proteins that influence plaque formation more strongly than virus replication and which are disrupted in MVA. These defective genes in MVA confound the interpretation of marker rescue experiments designed to map mutations responsible for the attenuation of this important VACV strain. PMID:24145605

Dobson, Bianca M; Tscharke, David C

2014-02-01

221

Immunogenicity of the Plasmodium falciparum glutamate-rich protein expressed by vaccinia virus.  

PubMed Central

The glurp gene of Plasmodium falciparum F32 has been inserted into a vaccinia virus, and the recombinant virus was designated VVG4. Expression of glurp in VVG4-infected Vero cells was analyzed by immunoprecipitation and revealed a primary GLURP product of approximately 220,000 Da; GLURP was detected both intracellularly and in culture supernatants. To study the immunogenicity of vaccinia virus-expressed GLURP, mice were immunized with VVG4 and serum samples were analyzed for antibody reactivity with three polypeptides, covering almost the entire GLURP molecule; these three polypeptides were produced in recombinant form in Escherichia coli. The immune response was primarily directed against a carboxy-terminal repeat region. The mouse anti-GLURP serum recognized authentic GLURP by immunoprecipitation analysis from P. falciparum grown in vitro. These results demonstrate that vaccinia virus-expressed glurp product can induce a humoral immune response against GLURP derived from blood-stage parasites. Images

Theisen, M; Cox, G; H?gh, B; Jepsen, S; Vuust, J

1994-01-01

222

Systemically administered DNA and fowlpox recombinants expressing four vaccinia virus genes although immunogenic do not protect mice against the highly pathogenic IHD-J vaccinia strain.  

PubMed

The first-generation smallpox vaccine was based on live vaccinia virus (VV) and it successfully eradicated the disease worldwide. Therefore, it was not administered any more after 1980, as smallpox no longer existed as a natural infection. However, emerging threats by terrorist organisations has prompted new programmes for second-generation vaccine development based on attenuated VV strains, which have been shown to cause rare but serious adverse events in immunocompromised patients. Considering the closely related animal poxviruses that might also be used as bioweapons, and the increasing number of unvaccinated young people and AIDS-affected immunocompromised subjects, a safer and more effective smallpox vaccine is still required. New avipoxvirus-based vectors should improve the safety of conventional vaccines, and protect from newly emerging zoonotic orthopoxvirus diseases and from the threat of deliberate release of variola or monkeypox virus in a bioterrorist attack. In this study, DNA and fowlpox recombinants expressing the L1R, A27L, A33R and B5R genes were constructed and evaluated in a pre-clinical trial in mouse, following six prime/boost immunisation regimens, to compare their immunogenicity and protective efficacy against a challenge with the lethal VV IHD-J strain. Although higher numbers of VV-specific IFN?-producing T lymphocytes were observed in the protected mice, the cytotoxic T-lymphocyte response and the presence of neutralising antibodies did not always correlate with protection. In spite of previous successful results in mice, rabbits and monkeys, where SIV/HIV transgenes were expressed by the fowlpox vector, the immune response elicited by these recombinants was low, and most of the mice were not protected. PMID:24050999

Bissa, Massimiliano; Pacchioni, Sole Maria; Zanotto, Carlo; De Giuli Morghen, Carlo; Illiano, Elena; Granucci, Francesca; Zanoni, Ivan; Broggi, Achille; Radaelli, Antonia

2013-12-26

223

High, broad, polyfunctional, and durable T cell immune responses induced in mice by a novel hepatitis C virus (HCV) vaccine candidate (MVA-HCV) based on modified vaccinia virus Ankara expressing the nearly full-length HCV genome.  

PubMed

A major goal in the control of hepatitis C infection is the development of a vaccine. Here, we have developed a novel HCV vaccine candidate based on the highly attenuated poxvirus vector MVA (referred to as MVA-HCV) expressing the nearly full-length (7.9-kbp) HCV sequence, with the aim to target almost all of the T and B cell determinants described for HCV. In infected cells, MVA-HCV produces a polyprotein that is subsequently processed into the structural and nonstructural HCV proteins, triggering the cytoplasmic accumulation of dense membrane aggregates. In both C57BL/6 and transgenic HLA-A2-vaccinated mice, MVA-HCV induced high, broad, polyfunctional, and long-lasting HCV-specific T cell immune responses. The vaccine-induced T cell response was mainly mediated by CD8 T cells; however, although lower in magnitude, the CD4(+) T cells were highly polyfunctional. In homologous protocol (MVA-HCV/MVA-HCV) the main CD8(+) T cell target was p7+NS2, whereas in heterologous combination (DNA-HCV/MVA-HCV) the main target was NS3. Antigenic responses were also detected against other HCV proteins (Core, E1-E2, and NS4), but the magnitude of the responses was dependent on the protocol used. The majority of the HCV-induced CD8(+) T cells were triple or quadruple cytokine producers. The MVA-HCV vaccine induced memory CD8(+) T cell responses with an effector memory phenotype. Overall, our data showed that MVA-HCV induced broad, highly polyfunctional, and durable T cell responses of a magnitude and quality that might be associated with protective immunity and open the path for future considerations of MVA-HCV as a prophylactic and/or therapeutic vaccine candidate against HCV. PMID:23596307

Gómez, Carmen E; Perdiguero, Beatriz; Cepeda, María Victoria; Mingorance, Lidia; García-Arriaza, Juan; Vandermeeren, Andrea; Sorzano, Carlos Óscar S; Esteban, Mariano

2013-07-01

224

Glomulin: a permissivity factor for vaccinia virus infection.  

PubMed

In earlier studies we provided evidence that vaccinia virus (VACV) phosphorylation-activation of host cell signaling effectors is critical for subsequent viral replication. In this report, using mass spectrometry-based proteomics, we have identified 387 host cell proteins that co-immunoprecipitate with VACV in infected, permissive PM1.CCR5 human T cells. Among these, glomulin was distinguishable based on its known interaction with a tyrosine kinase receptor, c-Met, its ability to become tyrosine-phosphorylated, and its association with signaling effectors. siRNA knockdown of glomulin expression in PM1.CCR5 T cells reduces VACV infection. Glomulin interacts with the inactive, nonphosphorylated form of c-MET. We demonstrate that treatment of PM1.CCR5 T cells with a c-Met phosphorylation inhibitor leads to a significant reduction in VACV infectivity. Additionally, inhibition of phosphorylation of c-Met abrogates VACV-inducible phosphorylation of Erk 1/2 and IRS-2, signaling effectors identified as critical for VACV infection. These data identify glomulin as a permissivity factor for VACV infection and as a potential therapeutic target for inhibition of VACV infection. PMID:22280104

Rahbar, Ramtin; Rogers, Erin; Murooka, Thomas; Kislinger, Thomas; Fish, Eleanor N

2012-03-01

225

Stunned Silence: Gene Expression Programs in Human Cells Infected with Monkeypox or Vaccinia Virus  

PubMed Central

Poxviruses use an arsenal of molecular weapons to evade detection and disarm host immune responses. We used DNA microarrays to investigate the gene expression responses to infection by monkeypox virus (MPV), an emerging human pathogen, and Vaccinia virus (VAC), a widely used model and vaccine organism, in primary human macrophages, primary human fibroblasts and HeLa cells. Even as the overwhelmingly infected cells approached their demise, with extensive cytopathic changes, their gene expression programs appeared almost oblivious to poxvirus infection. Although killed (gamma-irradiated) MPV potently induced a transcriptional program characteristic of the interferon response, no such response was observed during infection with either live MPV or VAC. Moreover, while the gene expression response of infected cells to stimulation with ionomycin plus phorbol 12-myristate 13-acetate (PMA), or poly (I-C) was largely unimpaired by infection with MPV, a cluster of pro-inflammatory genes were a notable exception. Poly(I-C) induction of genes involved in alerting the innate immune system to the infectious threat, including TNF-alpha, IL-1 alpha and beta, CCL5 and IL-6, were suppressed by infection with live MPV. Thus, MPV selectively inhibits expression of genes with critical roles in cell-signaling pathways that activate innate immune responses, as part of its strategy for stealthy infection.

Rubins, Kathleen H.; Hensley, Lisa E.; Relman, David A.; Brown, Patrick O.

2011-01-01

226

Biochemical and Biophysical Properties of a Putative Hub Protein Expressed by Vaccinia Virus*  

PubMed Central

H5 is a constitutively expressed, phosphorylated vaccinia virus protein that has been implicated in viral DNA replication, post-replicative gene expression, and virus assembly. For the purpose of understanding the role of H5 in vaccinia biology, we have characterized its biochemical and biophysical properties. Previously, we have demonstrated that H5 is associated with an endoribonucleolytic activity. In this study, we have shown that this cleavage results in a 3?-OH end suitable for polyadenylation of the nascent transcript, corroborating a role for H5 in vaccinia transcription termination. Furthermore, we have shown that H5 is intrinsically disordered, with an elongated rod-shaped structure that preferentially binds double-stranded nucleic acids in a sequence nonspecific manner. The dynamic phosphorylation status of H5 influences this structure and has implications for the role of H5 in multiple processes during virus replication.

Kay, Nicole E.; Bainbridge, Travis W.; Condit, Richard C.; Bubb, Michael R.; Judd, Reuben E.; Venkatakrishnan, Balasubramanian; McKenna, Robert; D'Costa, Susan M.

2013-01-01

227

Studies using a recombinant vaccinia virus expressing the circumsporozoite protein of Plasmodium berghei.  

PubMed

A recombinant vaccinia virus was constructed which expressed the circumsporozoite protein of Plasmodium berghei. Four different strains of mice belonging to different haplotypes were immunized with the recombinant virus. The antibody response to the circumsporozoite protein as well as to vaccinia virus varied among the strains, independently of each other. The anti-circumsporozoite protein titers were comparable to that obtained on immunization with irradiated sporozoites. Spleen cells from H2d mice immunized with P. berghei sporozoites showed a significant proliferative response when cultured in vitro with a low multiplicity of the recombinant vaccinia virus. A weak cytotoxic T lymphocyte response specifically targeting the circumsporozoite protein could be identified in spleens of BALB/c (H2d) mice immunized with vaccinia virus when BALB 3T3 cells transformed with a plasmid expressing the circumsporozoite protein under control of the simian virus 40 promoter were used as target cells in the cytotoxic T lymphocyte assay. However, none of the recombinant virus-immunized animals could be protected from a challenge of sporozoites even at the lowest dose of parasite used. PMID:1779992

Satchidanandam, V; Zavala, F; Moss, B

1991-09-01

228

Expression of the F glycoprotein of respiratory syncytial virus by a recombinant vaccinia virus: comparison of the individual contributions of the F and G glycoproteins to host immunity.  

PubMed

A cDNA clone representing the mRNA coding sequence of the fusion glycoprotein (F) gene of human respiratory syncytial virus (RSV) was constructed and inserted into the thymidine kinase gene of vaccinia virus (WR strain) under the control of a vaccinia virus promoter. The resulting recombinant vaccinia virus, vaccinia F, expressed the F1 and F2 cleavage products (48 and 20 kDa, respectively) of the F glycoprotein in cell culture. F1 and F2 were indistinguishable from their authentic RSV counterparts with respect to glycosylation, disulfide linkage, electrophoretic mobility, cell-surface expression, and antigenic specificity. Cotton rats infected intradermally with vaccinia F developed a high titer of serum F-specific antibodies, which neutralized infectivity of RSV. This neutralizing antibody response exceeded that induced by infection of the respiratory tract with RSV and was 6-fold higher than that induced by vaccinia G, a recombinant vaccinia virus that expressed the RSV G glycoprotein gene. Immunization with vaccinia F stimulated almost complete resistance to replication of RSV in the lower respiratory tract as well as significant resistance in the upper respiratory tract. The degree of resistance conferred by vaccinia F exceeded that induced by vaccinia G. PMID:3532115

Olmsted, R A; Elango, N; Prince, G A; Murphy, B R; Johnson, P R; Moss, B; Chanock, R M; Collins, P L

1986-10-01

229

The immunogenicity of recombinant LC16mO vaccinia virus harboring HBsAg gene in mice.  

PubMed

It was found that hepatitis B surface antigen (HBsAg) expressed by recombinant vaccinia virus (RVV), rProHBmO143, harboring HBsAg gene was immunologically similar to plasma-derived HBsAg and immunogenicity of the rProHBmO143 was possible to evaluate by the skin scarification (SS) method using BALB/c mice. When we compared the immunogenicity of 10(8) TCID50 of the rProHBmO143 by the SS method with that of 0.125 ml of the plasma-derived hepatitis B vaccine (HB vaccine) given by intraperitoneal inoculation, the anti-HBs antibody eliciting ability of its RVV was almost the same as that of the HB vaccine with maintenance of high antibody titers, and the antibody responses rose further by re-inoculation in association with HB vaccine, especially by using its RVV as a priming. Also, no virus was recovered from the liver, spleen or brain of the mice inoculated with rProHBmO143 by the SS method. Furthermore, in mice inoculated with rProHBmO143 and then inoculated with RVV harboring Japanese encephalitis virus (JEV) gene 24-weeks later, no effect was recognized on duration of anti-HBs antibody persistence while anti-JEV antibody is being produced. These results suggest that the rProHBmO143 is likely to become a practical live vaccine; a different immunization schedule to protect against hepatitis B virus and two or more kinds of RVV vaccines may be usable for the same animal or humans at intervals of some years. PMID:1888498

Watanabe, K; Morita, M; Sato, T; Yausi, K; Kojima, A

1991-04-01

230

Antibodies against the extracellular enveloped virus B5R protein are mainly responsible for the EEV neutralizing capacity of vaccinia immune globulin.  

PubMed

In the event of smallpox bioterrorism, widespread vaccination may be required. Vaccinia immune globulin (VIG) has been used to treat complications from the smallpox vaccine. While the potency of VIG was defined by its ability to neutralize intracellular mature virus, a second form of vaccinia called the extracellular enveloped virus (EEV) is critical for virus spread in the host. The B5R-protein is one of many EEV-specific proteins. Immunoprecipitation and ELISA revealed that VIG recognizes the B5R-protein. An EEV plaque-reduction assay using a recombinant vaccinia that lacks the majority of the extracellular domain of B5R showed that the ability of VIG to neutralize EEV is principally directed at B5R. In addition, absorbing out the anti-B5R antibody present in VIG through the addition of recombinant B5R protein abrogated VIG's ability to significantly neutralize wild-type EEV. This work demonstrates the prominent role of B5R as a target of EEV-neutralizing activity of human antibodies. PMID:15246280

Bell, Edward; Shamim, Mohammad; Whitbeck, J Charles; Sfyroera, Georgia; Lambris, John D; Isaacs, Stuart N

2004-08-01

231

Effect of glycosylation inhibitors on the release of enveloped vaccinia virus.  

PubMed Central

Addition of 1 to 10 mM 2-deoxy-D-glucose (2-dg) or glucosamine (gln) to the growth medium of vaccinia virus-infected cells inhibited the release of extracellular enveloped vaccinia virus (EEV) without affecting the production of intracellular naked vaccinia virus (INV) particles. In contrast, INV infectivity (particles per PFU) was decreased sevenfold by 50 mM 2-dg. Treatment with 2-dg reduced but did not eliminate glycosylation of the INV 37,000-molecular-weight glycoprotein. The kinetics of sensitivity to inhibitor addition experiments and inhibitor reversal experiments indicated that EEV release was dependent on glycosylation before 8 h postinfection. This was supported by polyacrylamide gel electrophoretic analysis of the synthesis kinetics for cell membrane-associated vaccinia glycoproteins in 2-dg-inhibited infected cells. The dependence of vaccinia protein glycosylation before 8 h postinfection for efficient EEV release was observed in spite of the fact that the period of greatest glycoprotein synthesis was 8 to 12 h postinfection. The presence of 2-dg resulted in an incompletely glycosylated 89,000-molecular-weight glycoprotein, as indicated by a reduction in the apparent glycoprotein molecular weight. The morphological event affected by the inhibitors was the acquisition by INV of a double-membrane structure from the Golgi apparatus. This morphological intermediate is necessary for release of EEV. Images

Payne, L G; Kristensson, K

1982-01-01

232

TRAF2 Facilitates Vaccinia Virus Replication by Promoting Rapid Virus Entry  

PubMed Central

ABSTRACT Tumor necrosis factor receptor (TNFR)-associated factor 2 (TRAF2) is a pivotal intracellular mediator of signaling pathways downstream of TNFR1 and -2 with known pro- and antiviral effects. We investigated its role in the replication of the prototype poxvirus vaccinia virus (VACV). Loss of TRAF2 expression, either through small interfering RNA treatment of HeLa cells or through genetic knockout in murine embryonic fibroblasts (MEFs), led to significant reductions in VACV growth following low-multiplicity infection. In single-cycle infections, there was delayed production of both early and late VACV proteins as well as accelerated virus-induced alterations to cell morphology, indicating that TRAF2 influences early stages of virus replication. Consistent with an early role, uncoating assays showed normal virus attachment but delayed virus entry in the absence of TRAF2. Although alterations to c-Jun N-terminal kinase (JNK) signaling were apparent in VACV-infected TRAF2?/? MEFs, treatment of wild-type cells with a JNK inhibitor did not affect virus entry. Instead, treatment with an inhibitor of endosomal acidification greatly reduced virus entry into TRAF2?/? MEFs, suggesting that VACV is reliant on the endosomal route of entry in the absence of TRAF2. Thus, TRAF2 is a proviral factor for VACV that plays a role in promoting efficient viral entry, most likely via the plasma membrane. IMPORTANCE Tumor necrosis factor receptor-associated factors (TRAFs) are key facilitators of intracellular signaling with roles in innate and adaptive immunity and stress responses. We have discovered that TRAF2 is a proviral factor in vaccinia virus replication in both HeLa cells and mouse embryonic fibroblasts and that its influence is exercised through promotion of efficient virus entry.

Haga, Ismar R.; Pechenick Jowers, Tali; Griffiths, Samantha J.; Haas, Juergen

2014-01-01

233

Aurintricarboxylic acid inhibits the early stage of vaccinia virus replication by targeting both cellular and viral factors.  

PubMed

Aurintricarboxylic acid (ATA) has been shown to inhibit the replication of viruses from several different families, including human immunodeficiency virus, vesicular stomatitis virus, and the coronavirus causing severe acute respiratory syndrome. This study characterizes the inhibitory effect of ATA on vaccinia virus replication in HeLa, Huh7, and AD293 cells. Vaccinia virus replication is significantly abrogated upon ATA treatment, which is associated with the inhibition of early viral gene transcription. This inhibitory effect may be attributed to two findings. First, ATA blocks the phosphorylation of extracellular signal-regulated kinase 1/2, an event shown to be essential for vaccinia virus replication. Second, ATA inhibits the phosphatase activity of the viral enzyme H1L, which is required to initiate viral transcription. Thus, ATA inhibits vaccinia virus replication by targeting both cellular and viral factors essential for the early stage of replication. PMID:17192307

Myskiw, Chad; Deschambault, Yvon; Jefferies, Kristel; He, Runtao; Cao, Jingxin

2007-03-01

234

Aurintricarboxylic Acid Inhibits the Early Stage of Vaccinia Virus Replication by Targeting both Cellular and Viral Factors?  

PubMed Central

Aurintricarboxylic acid (ATA) has been shown to inhibit the replication of viruses from several different families, including human immunodeficiency virus, vesicular stomatitis virus, and the coronavirus causing severe acute respiratory syndrome. This study characterizes the inhibitory effect of ATA on vaccinia virus replication in HeLa, Huh7, and AD293 cells. Vaccinia virus replication is significantly abrogated upon ATA treatment, which is associated with the inhibition of early viral gene transcription. This inhibitory effect may be attributed to two findings. First, ATA blocks the phosphorylation of extracellular signal-regulated kinase 1/2, an event shown to be essential for vaccinia virus replication. Second, ATA inhibits the phosphatase activity of the viral enzyme H1L, which is required to initiate viral transcription. Thus, ATA inhibits vaccinia virus replication by targeting both cellular and viral factors essential for the early stage of replication.

Myskiw, Chad; Deschambault, Yvon; Jefferies, Kristel; He, Runtao; Cao, Jingxin

2007-01-01

235

Genetic strain modification of a live rabies virus vaccine widely used in Europe for wildlife oral vaccination.  

PubMed

In Europe, the main reservoir and vector of rabies has been the red fox (Vulpes vulpes). Oral immunization of foxes with live vaccines, using attenuated rabies strains (SAD B19, SAD Bern), apathogenic mutants of an attenuated strain (SAG2) and the vaccinia-rabies glycoprotein recombinant virus vaccine (V-RG), has been shown to be the most effective method for the control and elimination of rabies. Among all vaccines currently used for wildlife oral vaccination, one vaccine (marketed as SAD Bern strain) has been widely used in Europe since 1992 with the distribution of 17million of baits in 2011. Because of the potential environmental safety risk of a live virus which could revert to virulence, the full genome sequencing of this vaccine was undertaken and the sequence was characterized and compared with those of referenced rabies viruses. The vaccine showed higher similarity to the strains belonging to the SAD B19 vaccine virus strains than to the SAD Bern vaccines. This study is the first one reporting on virus strain identity changes in this attenuated vaccine. PMID:23899697

Cliquet, Florence; Robardet, Emmanuelle; Picard Meyer, Evelyne

2013-10-01

236

Canine Distemper Virus (CDV) Infection of Ferrets as a Model for TestingMorbillivirusVaccine Strategies: NYVAC- and ALVAC-Based CDV Recombinants Protect against Symptomatic Infection  

Microsoft Academic Search

Canine distemper virus (CDV) infection of ferrets causes an acute systemic disease involving multiple organ systems, including the respiratory tract, lymphoid system, and central nervous system (CNS). We have tested candidate CDV vaccines incorporating the fusion (F) and hemagglutinin (HA) proteins in the highly attenu- ated NYVAC strain of vaccinia virus and in the ALVAC strain of canarypox virus, which

CHARLES B. STEPHENSEN; JANET WELTER; SUBHASHCHANDRA R. THAKER; JILL TAYLOR

237

Nitric Oxide Production Is Increased during Murine Vaccinia Virus Infection, but May Not Be Essential for Virus Clearance  

Microsoft Academic Search

Recent reports have highlighted a potential antiviral activity for nitric oxide (NO). The purpose of this study was to investigate the production of NO in mice during vaccinia virus (VV) or herpes simplex virus type 1 infection, and to assess the role of NO in clearance of VV. Reactive nitrogen intermediates (RNI; NO and its stable oxidation products, nitrite and

MICHAEL S. ROLPH; IAN A. RAMSHAW; KIRK A. ROCKETT; JANET RUBY; WILLIAM B. COWDEN

1996-01-01

238

Evaluating vaccinia virus cytokine co-expression in TLR GKO mice  

Microsoft Academic Search

Using Toll-like receptor (TLR) and MyD88 gene knock-out (GKO) mice the effect of TLRs and MyD88 on virus replication, interferon (IFN)-? production, natural killer (NK) cell and CD8T cell responses were assessed following ectromelia virus (ECTV) and recombinant vaccinia virus (rVV) infection. The capacity for rVVs encoding cytokines to restore immune function in MyD88?\\/? mice was clearly demonstrated. Results showed

Duncan B Sutherland; Charani Ranasinghe; Matthias Regner; Simon Phipps; Klaus I Matthaei; Stephanie L Day; Ian A Ramshaw

2011-01-01

239

Myopericarditis following Smallpox Vaccination.  

National Technical Information Service (NTIS)

Myopericarditis has been a rare or unrecognized event after smallpox vaccinations with the New York City Board of Health strain of vaccinia virus (Dryvax; Wyeth Laboratories, Marietta, Pennsylvania). In this article, the authors report an attributable inc...

J. E. Atwood M. K. Arness R. E. Eckart S. S. Love T. S. Wells

2004-01-01

240

Intracellular processing of the porcine coronavirus transmissible gastroenteritis virus spike protein expressed by recombinant vaccinia virus.  

PubMed

The Spike (S) protein from a virulent British field isolate of porcine transmissible gastroenteritis virus (TGEV) FS772/70 was constructed from cDNA and inserted into the vaccinia virus (VV) thymidine kinase gene locus under the control of the VV early/late gene P7.5k promoter. Recombinant S protein was synthesized as an endo-beta-N-acetylglucosaminidase H (Endo H)-sensitive glycoprotein with high mannose simple oligosaccharides (gp 190) that underwent post-translational modification to an Endo H-resistant glycoprotein with complex oligosaccharides (gp210). Immunofluorescence analysis demonstrated that the majority of recombinant S protein was retained at the Golgi but some S protein was expressed on the plasma membrane. Monoclonal antibodies (mAbs) raised against native S protein reacted with this recombinant S protein; also, mice infected with the recombinant vaccinia virus (rVV) expressing the S protein induced TGEV neutralizing antibodies. A truncated S protein (S delta) was also expressed in rVV-infected cells by introducing a deletion into the S protein cDNA that removed 292 amino acids from the C-terminus. The S delta protein (gp 170) was shown to be antigenically similar to TGEV S protein by immunofluorescence and immunoprecipitation tests but was retained in the endoplasmic reticulum and not expressed on the cell surface. PMID:1850927

Pulford, D J; Britton, P

1991-06-01

241

Recombination-Mediated Genetic Engineering of a Bacterial Artificial Chromosome Clone of Modified Vaccinia virus Ankara (MVA)  

PubMed Central

The production, manipulation and rescue of a bacterial artificial chromosome clone of Vaccinia virus (VAC-BAC) in order to expedite construction of expression vectors and mutagenesis of the genome has been described (Domi & Moss, 2002, PNAS 99 12415–20). The genomic BAC clone was ‘rescued’ back to infectious virus using a Fowlpox virus helper to supply transcriptional machinery. We apply here a similar approach to the attenuated strain Modified Vaccinia virus Ankara (MVA), now widely used as a safe non-replicating recombinant vaccine vector in mammals, including humans. Four apparently full-length, rescuable clones were obtained, which had indistinguishable immunogenicity in mice. One clone was shotgun sequenced and found to be identical to the parent. We employed GalK recombination-mediated genetic engineering (recombineering) of MVA-BAC to delete five selected viral genes. Deletion of C12L, A44L, A46R or B7R did not significantly affect CD8+ T cell immunogenicity in BALB/c mice, but deletion of B15R enhanced specific CD8+ T cell responses to one of two endogenous viral epitopes (from the E2 and F2 proteins), in accordance with published work (Staib et al., 2005, J. Gen. Virol. 86, 1997–2006). In addition, we found a higher frequency of triple-positive IFN-?, TNF-? and IL-2 secreting E3-specific CD8+ T-cells 8 weeks after vaccination with MVA lacking B15R. Furthermore, a recombinant vaccine capable of inducing CD8+ T cells against an epitope from Plasmodium berghei was created using GalK counterselection to insert an antigen expression cassette lacking a tandem marker gene into the traditional thymidine kinase locus of MVA-BAC. MVA continues to feature prominently in clinical trials of recombinant vaccines against diseases such as HIV-AIDS, malaria and tuberculosis. Here we demonstrate in proof-of-concept experiments that MVA-BAC recombineering is a viable route to more rapid and efficient generation of new candidate mutant and recombinant vaccines based on a clinically deployable viral vector.

Cottingham, Matthew G.; Andersen, Rikke F.; Spencer, Alexandra J.; Saurya, Saroj; Furze, Julie; Hill, Adrian V. S.; Gilbert, Sarah C.

2008-01-01

242

Safety and Immunogenicity of LC16m8, an Attenuated Smallpox Vaccine in Vaccinia-Naive Adults  

PubMed Central

Introduction.?LC16m8 is an attenuated cell culture–adapted Lister vaccinia smallpox vaccine missing the B5R protein and licensed for use in Japan. Methods.?We conducted a phase I/II clinical trial that compared the safety and immunogenicity of LC16m8 with Dryvax in vaccinia-naive participants. Adverse events were assessed, as were electrocardiography and laboratory testing for cardiotoxicity and viral culturing of the vaccination sites. Neutralization titers to vaccinia, monkeypox, and variola major were assessed and cell-mediated immune responses were measured by interferon (IFN)–? enzyme-linked immunosorbent spot and lymphoproliferation assays. Results.?Local and systemic reactions after vaccination with LC16m8 were similar to those reported after Dryvax. No clinically significant abnormalities consistent with cardiac toxicity were seen for either vaccine. Both vaccines achieved antivaccinia, antivariola, and antimonkeypox neutralizing antibody titers >1:40, although the mean plaque reduction neutralization titer of LC16m8 at day 30 after vaccination was significantly lower than Dryvax for anti-NYCBH vaccinia (P < .01), antimonkeypox (P < .001), and antivariola (P < .001). LC16m8 produced robust cellular immune responses that trended higher than Dryvax for lymphoproliferation (P = .06), but lower for IFN-? ELISPOT (P = .02). Conclusions.?LC16m8 generates neutralizing antibody titers to multiple poxviruses, including vaccinia, monkeypox, and variola major, and broad T-cell responses, indicating that LC16m8 may have efficacy in protecting individuals from smallpox. Clinical Trials Registration.?NCT00103584.

Kennedy, Jeffrey S.; Gurwith, Marc; Dekker, Cornelia L.; Frey, Sharon E.; Edwards, Kathryn M.; Kenner, Julie; Lock, Michael; Empig, Cyril; Morikawa, Shigeru; Saijo, Masayuki; Yokote, Hiroyuki; Karem, Kevin; Damon, Inger; Perlroth, Mark

2011-01-01

243

Intranodal Immunization With a Vaccinia Virus Encoding Multiple Antigenic Epitopes and Costimulatory Molecules in Metastatic Melanoma  

PubMed Central

Recombinant vaccinia virus (rVV) encoding tumor-associated antigens (TAAs) and adhesion or costimulatory molecules may represent important immunogenic reagents for cancer immunotherapy. Recently, intranodal (IN) antigen administration was suggested to be more immunogenic than intradermal (ID) vaccination. However, IN rVV administration has not been attempted so far. We used a rVV encoding gp100280–288, Melan-A/MART-127–35 and tyrosinase1–9 HLA-A0201 restricted epitopes and CD80 and CD86 costimulatory molecules in stage III and IV melanoma patients in a phase 1/2 trial. Of 15 patients initiating treatment, including two cycles of IN immunization, each comprising one rVV administration and three recall injections of the corresponding peptides, accompanied by subcutaneous granulocyte macrophage–colony stimulating factor supplementation, five withdrew due to progressing disease. Of 10 remaining patients seven showed evidence of induction of cytotoxic T lymphocytes (CTLs) directed against at least one epitope under investigation, as detectable by limiting dilution analysis (LDA) of specific precursors and multimer staining. Adverse reactions were mild (National Cancer Institute (NCI) grade 1–2) and mainly represented by fever, skin rashes, and pruritus. These data indicate that IN administration of rVV encoding melanoma-associated epitopes and costimulatory molecules is safe and immunogenic.

Adamina, Michel; Rosenthal, Rachel; Weber, Walter P; Frey, Daniel M; Viehl, Carsten T; Bolli, Martin; Huegli, Rolf W; Jacob, Augustinus L; Heberer, Michael; Oertli, Daniel; Marti, Walter; Spagnoli, Giulio C; Zajac, Paul

2009-01-01

244

Vaccinia Virus Entry, Exit, and Interaction with Differentiated Human Airway Epithelia?  

PubMed Central

Variola virus, the causative agent of smallpox, enters and exits the host via the respiratory route. To better understand the pathogenesis of poxvirus infection and its interaction with respiratory epithelia, we used vaccinia virus and examined its interaction with primary cultures of well-differentiated human airway epithelia. We found that vaccinia virus preferentially infected the epithelia through the basolateral membrane and released viral progeny across the apical membrane. Despite infection and virus production, epithelia retained tight junctions, transepithelial electrical conductance, and a steep transepithelial concentration gradient of virus, indicating integrity of the epithelial barrier. In fact, during the first four days of infection, epithelial height and cell number increased. These morphological changes and maintenance of epithelial integrity required vaccinia virus growth factor, which was released basolaterally, where it activated epidermal growth factor 1 receptors. These data suggest a complex interaction between the virus and differentiated airway epithelia; the virus preferentially enters the cells basolaterally, exits apically, and maintains epithelial integrity by stimulating growth factor receptors.

Vermeer, Paola D.; McHugh, Julia; Rokhlina, Tatiana; Vermeer, Daniel W.; Zabner, Joseph; Welsh, Michael J.

2007-01-01

245

One-step selection of Vaccinia virus-binding DNA aptamers by MonoLEX  

PubMed Central

Background As a new class of therapeutic and diagnostic reagents, more than fifteen years ago RNA and DNA aptamers were identified as binding molecules to numerous small compounds, proteins and rarely even to complete pathogen particles. Most aptamers were isolated from complex libraries of synthetic nucleic acids by a process termed SELEX based on several selection and amplification steps. Here we report the application of a new one-step selection method (MonoLEX) to acquire high-affinity DNA aptamers binding Vaccinia virus used as a model organism for complex target structures. Results The selection against complete Vaccinia virus particles resulted in a 64-base DNA aptamer specifically binding to orthopoxviruses as validated by dot blot analysis, Surface Plasmon Resonance, Fluorescence Correlation Spectroscopy and real-time PCR, following an aptamer blotting assay. The same oligonucleotide showed the ability to inhibit in vitro infection of Vaccinia virus and other orthopoxviruses in a concentration-dependent manner. Conclusion The MonoLEX method is a straightforward procedure as demonstrated here for the identification of a high-affinity DNA aptamer binding Vaccinia virus. MonoLEX comprises a single affinity chromatography step, followed by subsequent physical segmentation of the affinity resin and a single final PCR amplification step of bound aptamers. Therefore, this procedure improves the selection of high affinity aptamers by reducing the competition between aptamers of different affinities during the PCR step, indicating an advantage for the single-round MonoLEX method.

Nitsche, Andreas; Kurth, Andreas; Dunkhorst, Anna; Panke, Oliver; Sielaff, Hendrik; Junge, Wolfgang; Muth, Doreen; Scheller, Frieder; Stocklein, Walter; Dahmen, Claudia; Pauli, Georg; Kage, Andreas

2007-01-01

246

Secondary and Tertiary Transmission of Vaccinia Virus from US Military Service Member  

PubMed Central

During February and March 2010, the New York State Department of Health investigated secondary and tertiary vaccinia contact transmission from a military vaccinee to 4 close contacts. Identification of these cases underscores the need for strict adherence to postvaccination infection control guidance to avoid transmission of the live virus.

Hidalgo, Christina M.; Sullivan-Frohm, Ann; Schulte, Cynthia; Davis, Stephen; Kelly-Cirino, Cassandra; Egan, Christina; Wilkins, Kimberly; Emerson, Ginny L.; Noyes, Kimberly; Blog, Debra

2011-01-01

247

Partial deletion of the human host range gene in the attenuated vaccinia virus MVA  

Microsoft Academic Search

Summary The genome of a strongly attenuated vaccinia virus strain, MVA, was investigated by Southern blot and sequence analyses. Three major deletions, relative to the WR strain, were localized in MVA DNA. The deletions occurred near both ends of the viral genome and one of them affected a 55 K as well as the 32 K human host range gene.

W. Altenburger; C. P. Süter; Judit Altenburger

1989-01-01

248

A Loss of Function Analysis of Host Factors Influencing Vaccinia virus Replication by RNA Interference  

PubMed Central

Vaccinia virus (VACV) is a large, cytoplasmic, double-stranded DNA virus that requires complex interactions with host proteins in order to replicate. To explore these interactions a functional high throughput small interfering RNA (siRNA) screen targeting 6719 druggable cellular genes was undertaken to identify host factors (HF) influencing the replication and spread of an eGFP-tagged VACV. The experimental design incorporated a low multiplicity of infection, thereby enhancing detection of cellular proteins involved in cell-to-cell spread of VACV. The screen revealed 153 pro- and 149 anti-viral HFs that strongly influenced VACV replication. These HFs were investigated further by comparisons with transcriptional profiling data sets and HFs identified in RNAi screens of other viruses. In addition, functional and pathway analysis of the entire screen was carried out to highlight cellular mechanisms involved in VACV replication. This revealed, as anticipated, that many pro-viral HFs are involved in translation of mRNA and, unexpectedly, suggested that a range of proteins involved in cellular transcriptional processes and several DNA repair pathways possess anti-viral activity. Multiple components of the AMPK complex were found to act as pro-viral HFs, while several septins, a group of highly conserved GTP binding proteins with a role in sequestering intracellular bacteria, were identified as strong anti-viral VACV HFs. This screen has identified novel and previously unexplored roles for cellular factors in poxvirus replication. This advancement in our understanding of the VACV life cycle provides a reliable knowledge base for the improvement of poxvirus-based vaccine vectors and development of anti-viral theraputics.

Gonzalez, Orland; Haga, Ismar R.; Pechenick Jowers, Tali; Reynolds, Danielle K.; Wildenhain, Jan; Tekotte, Hille; Auer, Manfred; Tyers, Mike; Ghazal, Peter; Zimmer, Ralf; Haas, Jurgen

2014-01-01

249

A Differential Role for Macropinocytosis in Mediating Entry of the Two Forms of Vaccinia Virus into Dendritic Cells  

PubMed Central

Vaccinia virus (VACV) is being developed as a recombinant viral vaccine vector for several key pathogens. Dendritic cells (DCs) are specialised antigen presenting cells that are crucial for the initiation of primary immune responses; however, the mechanisms of uptake of VACV by these cells are unclear. Therefore we examined the binding and entry of both the intracellular mature virus (MV) and extracellular enveloped virus (EV) forms of VACV into vesicular compartments of monocyte-derived DCs. Using a panel of inhibitors, flow cytometry and confocal microscopy we have shown that neither MV nor EV binds to the highly expressed C-type lectin receptors on DCs that are responsible for capturing many other viruses. We also found that both forms of VACV enter DCs via a clathrin-, caveolin-, flotillin- and dynamin-independent pathway that is dependent on actin, intracellular calcium and host-cell cholesterol. Both MV and EV entry were inhibited by the macropinocytosis inhibitors rottlerin and dimethyl amiloride and depended on phosphotidylinositol-3-kinase (PI(3)K), and both colocalised with dextran but not transferrin. VACV was not delivered to the classical endolysosomal pathway, failing to colocalise with EEA1 or Lamp2. Finally, expression of early viral genes was not affected by bafilomycin A, indicating that the virus does not depend on low pH to deliver cores to the cytoplasm. From these collective results we conclude that VACV enters DCs via macropinocytosis. However, MV was consistently less sensitive to inhibition and is likely to utilise at least one other entry pathway. Definition and future manipulation of these pathways may assist in enhancing the activity of recombinant vaccinia vectors through effects on antigen presentation.

Sandgren, Kerrie J.; Wilkinson, John; Miranda-Saksena, Monica; McInerney, Gerald M.; Byth-Wilson, Karen; Robinson, Phillip J.; Cunningham, Anthony L.

2010-01-01

250

Nosocomial Vaccinia Infection  

PubMed Central

Although hospital-associated spread of vaccinia has been reported in the past, there have been no recent reports. This paper describes hospital-associated spread of vaccinia virus infection, supplies data on the environmental survival of vaccinia virus and offers recommendations for the management of patients with vaccinia that may minimize the hazard of infection in other high-risk patients. ImagesFigure 1.Figure 2.Figure 3.

Johnson, Rudolph H.; Krupp, Jan R.; Hoffman, Andrew R.; Koplan, Jeffrey P.; Nakano, James H.; Merigan, Thomas C.

1976-01-01

251

Prospective Surveillance for Cardiac Adverse Events in Healthy Adults Receiving Modified Vaccinia Ankara Vaccines: A Systematic Review  

PubMed Central

Background Vaccinia-associated myo/pericarditis was observed during the US smallpox vaccination (DryVax) campaign initiated in 2002. A highly-attenuated vaccinia strain, modified vaccinia Ankara (MVA) has been evaluated in clinical trials as a safer alternative to DryVax and as a vector for recombinant vaccines. Due to the lack of prospectively collected cardiac safety data, the US Food and Drug Administration required cardiac screening and surveillance in all clinical trials of MVA since 2004. Here, we report cardiac safety surveillance from 6 phase I trials of MVA vaccines. Methods Four clinical research organizations contributed cardiac safety data using common surveillance methods in trials administering MVA or recombinant MVA vaccines to healthy participants. ‘Routine cardiac investigations’ (ECGs and cardiac enzymes obtained 2 weeks after injections of MVA or MVA-HIV recombinants, or placebo-controls), and ‘Symptom-driven cardiac investigations’ are reported. The outcome measure is the number of participants who met the CDC-case definition for vaccinia-related myo/pericarditis or who experienced cardiac adverse events from an MVA vaccine. Results Four hundred twenty-five study participants had post-vaccination safety data analyzed, 382 received at least one MVA-containing vaccine and 43 received placebo; 717 routine ECGs and 930 cardiac troponin assays were performed. Forty-five MVA recipients (12%) had additional cardiac testing performed; 22 for cardiac symptoms, 19 for ECG/laboratory changes, and 4 for cardiac symptoms with an ECG/laboratory change. No participant had evidence of symptomatic or asymptomatic myo/pericarditis meeting the CDC-case definition and judged to be related to an MVA vaccine. Conclusions Prospective surveillance of MVA recipients for myo/pericarditis did not detect cardiac adverse reactions in 382 study participants. Trial Registration ClinicalTrials.gov NCT00082446?NCT003766090?NCT00252148?NCT00083603?NCT00301184?NCT00428337

Elizaga, Marnie L.; Vasan, Sandhya; Marovich, Mary A.; Sato, Alicia H.; Lawrence, Dale N.; Chaitman, Bernard R.; Frey, Sharon E.; Keefer, Michael C.

2013-01-01

252

Advances in virus research  

SciTech Connect

This book contains eight chapters. Some of the titles are: Initiation of viral DNA replication; Vaccinia: virus, vector, vaccine; The pre-S region of hepadnavirus envelope proteins; and Archaebacterial viruses.

Maramorosch, K. (Rutgers--the State Univ., New Brunswick, NJ (USA)); Murphy, F.A. (Centers for Disease Control, Atlanta, GA (USA)); Shatkin, A.J. (Rutgers-UMDNJ, Piscataway, NJ (US))

1988-01-01

253

Analysis of Vaccinia Virus-Host Protein-Protein Interactions: Validations of Yeast Two-Hybrid Screenings  

PubMed Central

Vaccinia virus, a large double-stranded DNA virus, is the prototype of the Orthopoxvirus genus, which includes several pathogenic poxviruses of humans, such as monkeypox virus and variola virus. Here, we report a comprehensive yeast two-hybrid (Y2H) screening for the protein?protein interactions between vaccinia and human proteins. A total of 109 novel vaccinia?human protein interactions were detected among 33 viral proteins. To validate subsets of those interactions, we constructed an ORFeome library of vaccinia virus strain WR using the Gateway plasmid cloning system. By co-expressing selected vaccinia and host proteins in a variety of expression systems, we found that at least 17 of the Y2H hits identified between vaccinia and human proteins can be verified by independent methods using GST pull-down assays, representing a 63% validation rate for the Y2H hits examined (17/27). Because the cloned ORFs are conveniently transferable from the entry vectors to various destination expression vectors, the vaccinia ORFeome library will be a useful resource for future high-throughput functional proteomic experiments.

2009-01-01

254

Detection of camel pox and vaccinia viruses by polymerase chain reaction  

Microsoft Academic Search

PCR following two methods of DNA extraction was used to confirm the growth of camel pox virus (CPV) and vaccinia virus in\\u000a cell culture and chorioallantoic membrane. Results were compared with the commonly used neutralization test. The first method\\u000a of DNA extraction was accomplished by using viral DNA in tissue culture supernatant and Chorioallantoic membrane, which was\\u000a released by initial

Hanan M. Sheikh Ali; A. I. Khalafalla; A. H. Nimir

2009-01-01

255

An intact signal peptide on dengue virus E protein enhances immunogenicity for CD8(+) T cells and antibody when expressed from modified vaccinia Ankara.  

PubMed

Dengue is a global public health concern and this is aggravated by a lack of vaccines or antiviral therapies. Despite the well-known role of CD8(+) T cells in the immunopathogenesis of Dengue virus (DENV), only recent studies have highlighted the importance of this arm of the immune response in protection against the disease. Thus, the majority of DENV vaccine candidates are designed to achieve protective titers of neutralizing antibodies, with less regard for cellular responses. Here, we used a mouse model to investigate CD8(+) T cell and humoral responses to a set of potential DENV vaccines based on recombinant modified vaccinia virus Ankara (rMVA). To enable this study, we identified two CD8(+) T cell epitopes in the DENV-3 E protein in C57BL/6 mice. Using these we found that all the rMVA vaccines elicited DENV-specific CD8(+) T cells that were cytotoxic in vivo and polyfunctional in vitro. Moreover, vaccines expressing the E protein with an intact signal peptide sequence elicited more DENV-specific CD8(+) T cells than those expressing E proteins in the cytoplasm. Significantly, it was these same ER-targeted E protein vaccines that elicited antibody responses. Our results support the further development of rMVA vaccines expressing DENV E proteins and add to the tools available for dengue vaccine development. PMID:24726244

Quinan, Bárbara R; Flesch, Inge E A; Pinho, Tânia M G; Coelho, Fabiana M; Tscharke, David C; da Fonseca, Flávio G

2014-05-23

256

The vaccinia virus-encoded Bcl-2 homologues do not act as direct Bax inhibitors.  

PubMed

Many viruses, including members of several poxvirus genera, encode inhibitors that block apoptosis by simultaneously binding the proapoptotic Bcl-2 proteins Bak and Bax. The Orthopoxvirus vaccinia virus encodes the Bcl-2-like F1 protein, which sequesters Bak but not Bax. However, N1, a potent virulence factor, is reported to be antiapoptotic and to interact with Bax. Here we investigated whether vaccinia virus inhibits Bak/Bax-dependent apoptosis via the cooperative action of F1 and N1. We found that Western Reserve (WR) and ?N1L viruses inhibited drug- and infection-induced apoptosis equally. Meanwhile, infections with ?F1L or ?N1L/F1L virus resulted in similar levels of Bax activation and apoptosis. Outside the context of infection, N1 did not block drug- or Bax-induced cell death or interact with Bax. In addition to F1 and N1, vaccinia virus encodes further structural homologs of Bcl-2 proteins that are conserved in orthopoxviruses, including A46, A52, B14, C1, C6, C16/B22, K7, and N2. However, we found that these do not associate with Bax or inhibit drug-induced cell death. Based on our findings that N1 is not an antiapoptotic protein, we propose that the F1 orthologs represent the only orthopoxvirus Bcl-2 homolog to directly inhibit the Bak/Bax checkpoint. PMID:22013032

Postigo, Antonio; Way, Michael

2012-01-01

257

The Vaccinia Virus-Encoded Bcl-2 Homologues Do Not Act as Direct Bax Inhibitors  

PubMed Central

Many viruses, including members of several poxvirus genera, encode inhibitors that block apoptosis by simultaneously binding the proapoptotic Bcl-2 proteins Bak and Bax. The Orthopoxvirus vaccinia virus encodes the Bcl-2-like F1 protein, which sequesters Bak but not Bax. However, N1, a potent virulence factor, is reported to be antiapoptotic and to interact with Bax. Here we investigated whether vaccinia virus inhibits Bak/Bax-dependent apoptosis via the cooperative action of F1 and N1. We found that Western Reserve (WR) and ?N1L viruses inhibited drug- and infection-induced apoptosis equally. Meanwhile, infections with ?F1L or ?N1L/F1L virus resulted in similar levels of Bax activation and apoptosis. Outside the context of infection, N1 did not block drug- or Bax-induced cell death or interact with Bax. In addition to F1 and N1, vaccinia virus encodes further structural homologs of Bcl-2 proteins that are conserved in orthopoxviruses, including A46, A52, B14, C1, C6, C16/B22, K7, and N2. However, we found that these do not associate with Bax or inhibit drug-induced cell death. Based on our findings that N1 is not an antiapoptotic protein, we propose that the F1 orthologs represent the only orthopoxvirus Bcl-2 homolog to directly inhibit the Bak/Bax checkpoint.

Postigo, Antonio

2012-01-01

258

Tyrosine phosphorylation of phospholipase C-gamma 1 by vaccinia virus growth factor.  

PubMed

Vaccinia virus growth factor (VGF), a highly glycosylated polypeptide encoded in the genome of vaccinia virus, shares amino acid sequence homology and functional properties with cellular growth factors, EGF and TGF-alpha. Although the mitogenic activity of the purified or synthetic VGF suggested that the factor may be beneficial for viral replication by stimulation of host cell growth, neither the role in virus life cycle nor the step next to the EGF receptor activation had been firmly established. We found tyrosine-phosphorylations of PLC-gamma 1 and concomitant increase of the phosphoinositides level in the human epidermoidal A431 cells either treated with purified VGF or infected with vaccinia virus. In contrast to the wild-type virus, a VGF- mutant virus did not induce tyrosine phosphorylation of PLC-gamma 1. Phosphopeptide analysis indicated that the phosphorylation of the PLC-gamma 1 by VGF includes tyrosine residues identical to those phosphorylated by EGF. These results suggest that tyrosine phosphorylation of PLC-gamma 1, mediated by VGF, leads to activation of PLC-gamma 1 and a concomitant increase in phosphatidylinositol turnover. PMID:8525617

Kim, H S; Lee, Y H; Min, D S; Chang, J S; Ryu, S H; Ahn, B Y; Suh, P G

1995-12-01

259

Vaccinia Virus Infection Disarms the Mitochondrion-Mediated Pathway of the Apoptotic Cascade by Modulating the Permeability Transition Pore  

PubMed Central

Many viruses have evolved strategies that target crucial components within the apoptotic cascade. One of the best studied is the caspase 8 inhibitor, crmA/Spi-2, encoded by members of the poxvirus family. Since many proapoptotic stimuli induce apoptosis through a mitochondrion-dependent, caspase 8-independent pathway, we hypothesized that vaccinia virus would encode a mechanism to directly modulate the mitochondrial apoptotic pathway. In support of this, we observed that Jurkat cells, which undergo Fas-mediated apoptosis exclusively through the mitochondrial route, were resistant to Fas-induced death following infection with a crmA/Spi-2-deficient strain of vaccinia virus. In addition, vaccinia virus-infected cells subjected to the proapoptotic stimulus staurosporine exhibited decreased levels of both cytochrome c released from the mitochondria and caspase 3 activation. In all cases we found that the loss of the mitochondrial membrane potential, which occurs as a result of opening the multimeric permeability transition pore complex, was prevented in vaccinia virus-infected cells. Moreover, vaccinia virus infection specifically inhibited opening of the permeability transition pore following treatment with the permeability transition pore ligand atractyloside and t-butylhydroperoxide. These studies indicate that vaccinia virus infection directly impacts the mitochondrial apoptotic cascade by influencing the permeability transition pore.

Wasilenko, Shawn T.; Meyers, Adrienne F. A.; Vander Helm, Kathleen; Barry, Michele

2001-01-01

260

Detailed kinetics of immune responses to a new cell culture-derived smallpox vaccine in vaccinia-naïve adults.  

PubMed

The aim of the present study was to investigate the kinetics of humoral and cell-mediated immune responses to a new cell culture-derived smallpox vaccine (CJ-50300, CJ Corporation, South Korea) in 18 vaccinia-naïve volunteers. All subjects achieved positive humoral immune responses (plaque reduction neutralizing antibody assay) 28 days after vaccination, and cell-mediated immune responses (ELISPOT assay) 14 days after vaccination. Humoral immune responses increased up to 28 days after vaccination and were maintained up to 56 days after vaccination. In contrast, cell-mediated immune responses increased up to 14 days after vaccination and steadily decreased to 56 days after vaccination [Clinical Trial No. NCT 00336635]. PMID:17597266

Kim, Sung-Han; Choi, Su-Jin; Park, Wan Beom; Kim, Hong-Bin; Kim, Nam-Joong; Oh, Myoung-Don; Choe, Kang-Won

2007-08-14

261

Enteric immunization of mice against influenza with recombinant vaccinia.  

PubMed Central

Intrajejunal administration to mice of a recombinant vaccinia virus containing the influenza virus hemagglutinin gene induced IgA antibody in nasal, gut, and vaginal secretions. It also induced IgG antibody in serum and cell-mediated immunity. The immunization provided significant protection against an influenza virus challenge. This work suggests that enteric-coated recombinant vaccinia could be an orally administered, inexpensive, multivalent, temperature-stable, safe, and effective vaccine for children that could be particularly useful in developing nations, where multiple injections are not easily administered. Oral administration of vaccines should also reduce children's fear of shots at the doctor's office.

Meitin, C A; Bender, B S; Small, P A

1994-01-01

262

Enteric Immunization of Mice Against Influenza with Recombinant Vaccinia  

NASA Astrophysics Data System (ADS)

Intrajejunal administration to mice of a recombinant vaccinia virus containing the influenza virus hemagglutinin gene induced IgA antibody in nasal, gut, and vaginal secretions. It also induced IgG antibody in serum and cell-mediated immunity. The immunization provided significant protection against an influenza virus challenge. This work suggests that enteric-coated recombinant vaccinia could be an orally administered, inexpensive, multivalent, temperature-stable, safe, and effective vaccine for children that could be particularly useful in developing nations, where multiple injections are not easily administered. Oral administration of vaccines should also reduce children's fear of shots at the doctor's office.

Meitin, Catherine A.; Bender, Bradley S.; Small, Parker A., Jr.

1994-11-01

263

Permissivity of the NCI-60 cancer cell lines to oncolytic Vaccinia Virus GLV-1h68  

PubMed Central

Background Oncolytic viral therapy represents an alternative therapeutic strategy for the treatment of cancer. We previously described GLV-1h68, a modified Vaccinia Virus with exclusive tropism for tumor cells, and we observed a cell line-specific relationship between the ability of GLV-1h68 to replicate in vitro and its ability to colonize and eliminate tumor in vivo. Methods In the current study we surveyed the in vitro permissivity to GLV-1h68 replication of the NCI-60 panel of cell lines. Selected cell lines were also tested for permissivity to another Vaccinia Virus and a vesicular stomatitis virus (VSV) strain. In order to identify correlates of permissity to viral infection, we measured transcriptional profiles of the cell lines prior infection. Results We observed highly heterogeneous permissivity to VACV infection amongst the cell lines. The heterogeneity of permissivity was independent of tissue with the exception of B cell derivation. Cell lines were also tested for permissivity to another Vaccinia Virus and a vesicular stomatitis virus (VSV) strain and a significant correlation was found suggesting a common permissive phenotype. While no clear transcriptional pattern could be identified as predictor of permissivity to infection, some associations were observed suggesting multifactorial basis permissivity to viral infection. Conclusions Our findings have implications for the design of oncolytic therapies for cancer and offer insights into the nature of permissivity of tumor cells to viral infection.

2011-01-01

264

Expression of bovine leukaemia virus envelope gene by recombinant vaccinia viruses.  

PubMed

Recombinant vaccinia viruses (VV) containing the envelope gene of bovine leukaemia virus (BLV) were constructed. Three virus constructs were designed: VV-BLV1 which contained the open reading frame for envelope glycoprotein gp51 alone, under control of VVP7.5 promoter; VV-BLV2 and VV-BLV3 contained the entire gene (gp51 + gp30) coding sequence downstream of VP7.5 and the fowlpox virus early/late promoter (PFE/L) respectively. All three VV recombinants expressed envelope glycoproteins as determined by the agar gel diffusion assay. By immunofluorescence techniques it was shown that while VV-BLV2 and VV-BLV3 expressed envelope glycoprotein on the surface of virus-infected cells, VV-BLV1 failed to do so. Rabbits inoculated with VV-BLV1 failed to show an anti envelope glycoprotein antibody response, however, significant levels of antibodies against envelope glycoprotein were detected in sera from rabbits inoculated with VV-BLV2 and VV-BLV3. PMID:1963249

Kumar, S; Andrew, M E; Boyle, D B; Brandon, R B; Lavin, M F; Daniel, R C

1990-10-01

265

Oncolytic vaccinia virus demonstrates antiangiogenic effects mediated by targeting of VEGF.  

PubMed

Oncolytic vaccinia virus has been shown to induce a profound, rapid and tumor-specific vascular collapse in both preclinical models and clinical studies; however, a complete examination of the kinetics and levels of collapse and revascularization has not been described previously. Contrast-enhanced ultrasound was used to follow tumor perfusion levels in mouse tumor models at times after vaccinia therapy. It was observed that revascularization after viral therapy was dramatically delayed and did not occur until after viral clearance. This indicated that oncolytic vaccinia may possess a previously undescribed antiangiogenic potential that might synergize with the reported anti-vascular effects. Despite a rapid loss of perfusion and widespread hypoxia within the tumor, it was observed that VEGF levels in the tumor were suppressed throughout the period of active viral infection. Although tumor vasculature could eventually reform after the viral therapy was cleared in mouse models, anti-tumor effects could be significantly enhanced through additional combination with anti-VEGF therapies. This was initially examined using a gene therapy approach (Ad-Flk1-Fc) to target VEGF directly, demonstrating that the timing of application of the antiangiogenic therapy was critical. However, it is also known that oncolytic vaccinia sensitizes tumors to tyrosine kinase inhibitors (TKI) in the clinic through an unknown mechanism. It is possible this phenomenon may be mediated through the antiangiogenic effects of the TKIs. This was modeled in mouse tumors using sunitinib in combination with oncolytic vaccinia. It was observed that prevention of angiogenesis mediated by oncolytic vaccinia can be utilized to enhance the TKI therapy. PMID:24474587

Hou, Weizhou; Chen, Hannah; Rojas, Juan; Sampath, Padma; H Thorne, Stephen

2014-09-01

266

Dominant negative selection of vaccinia virus using a thymidine kinase/thymidylate kinase fusion gene and the prodrug azidothymidine  

SciTech Connect

The Escherichia coli thymidine kinase/thymidylate kinase (tk/tmk) fusion gene encodes an enzyme that efficiently converts the prodrug 3'-azido-2',3'-dideoxythymidine (AZT) into its toxic triphosphate derivative, a substance which stops DNA chain elongation. Integration of this marker gene into vaccinia virus that normally is not inhibited by AZT allowed the establishment of a powerful selection procedure for recombinant viruses. In contrast to the conventional vaccinia thymidine kinase (tk) selection that is performed in tk-negative cell lines, AZT selection can be performed in normal (tk-positive) cell lines. The technique is especially useful for the generation of replication-deficient vaccinia viruses and may also be used for gene knock-out studies of essential vaccinia genes.

Holzer, Georg W. [Baxter BioScience/Vaccines, Biomedical Research Center, Uferstrasse 15, A-2304 Orth/Donau (Austria); Mayrhofer, Josef [Baxter BioScience/Vaccines, Biomedical Research Center, Uferstrasse 15, A-2304 Orth/Donau (Austria); Gritschenberger, Werner [Baxter BioScience/Vaccines, Biomedical Research Center, Uferstrasse 15, A-2304 Orth/Donau (Austria); Falkner, Falko G. [Baxter BioScience/Vaccines, Biomedical Research Center, Uferstrasse 15, A-2304 Orth/Donau (Austria)]. E-mail: falknef@baxter.com

2005-07-05

267

Identification of a Bohle iridovirus thymidine kinase gene and demonstration of activity using vaccinia virus.  

PubMed

In recent years interest in the family Iridoviridae has been renewed by the identification of a number of viruses, particularly from the genus Ranavirus, associated with disease in a range of poikilotherms. Ranaviruses have been isolated from amphibian, piscine and reptilian species. Here we describe an open reading frame (ORF) identified in the genome of Bohle iridovirus (BIV) which contains a nucleotide binding motif conserved within the thymidine kinase (TK) genes of iridoviruses from other genera (lymphocystis disease virus, LCDV, type species of the genus Lymphocystivirus; Chilo iridescent virus, CIV, type species of the genus Iridovirus). The ability of this putative gene to express a functional TK was confirmed by rescue of a TK negative mutant vaccinia virus in the presence of selective media, when expression was controlled by a vaccinia virus promoter. The sequence of the BIV TK was compared with the homologous sequences from epizootic haematopoietic necrosis virus (EHNV), a virus associated with disease in fish, from Wamena iridovirus (WIV) associated with systemic disease in green pythons, and from frog virus 3 (FV3) the ranavirus type species. Comparisons between these sequences and those available from other ranaviruses, other iridoviruses, other DNA viruses and cellular TKs are presented. PMID:15883656

Coupar, B E H; Goldie, S G; Hyatt, A D; Pallister, J A

2005-09-01

268

9 CFR 113.211 - Feline Rhinotracheitis Vaccine, Killed Virus.  

Code of Federal Regulations, 2010 CFR

... 2010-01-01 2010-01-01 false Feline Rhinotracheitis Vaccine, Killed Virus...REQUIREMENTS Killed Virus Vaccines § 113.211 Feline Rhinotracheitis Vaccine, Killed Virus. Feline Rhinotracheitis Vaccine, Killed...

2010-01-01

269

9 CFR 113.203 - Feline Panleukopenia Vaccine, Killed Virus.  

Code of Federal Regulations, 2010 CFR

... 2009-01-01 2009-01-01 false Feline Panleukopenia Vaccine, Killed Virus. 113...REQUIREMENTS Killed Virus Vaccines § 113.203 Feline Panleukopenia Vaccine, Killed Virus. Feline Panleukopenia Vaccine, Killed...

2009-01-01

270

9 CFR 113.203 - Feline Panleukopenia Vaccine, Killed Virus.  

Code of Federal Regulations, 2010 CFR

... 2010-01-01 2010-01-01 false Feline Panleukopenia Vaccine, Killed Virus. 113...REQUIREMENTS Killed Virus Vaccines § 113.203 Feline Panleukopenia Vaccine, Killed Virus. Feline Panleukopenia Vaccine, Killed...

2010-01-01

271

9 CFR 113.211 - Feline Rhinotracheitis Vaccine, Killed Virus.  

Code of Federal Regulations, 2010 CFR

... 2009-01-01 2009-01-01 false Feline Rhinotracheitis Vaccine, Killed Virus...REQUIREMENTS Killed Virus Vaccines § 113.211 Feline Rhinotracheitis Vaccine, Killed Virus. Feline Rhinotracheitis Vaccine, Killed...

2009-01-01

272

Human cytomegalovirus UL97 kinase confers ganciclovir susceptibility to recombinant vaccinia virus.  

PubMed Central

We analyzed whether the phosphotransferase encoded by the UL97 open reading frame of human cytomegalovirus (HCMV) alone is sufficient to confer ganciclovir (GCV) susceptibility to a foreign virus. Two vaccinia virus recombinants (T1 and A5) containing the UL97 open reading frames from a GCV-sensitive HCMV and from a GCV-resistant strain were constructed. T1 exhibited a GCV-sensitive phenotype in plaque reduction assays, whereas A5 did not. Moreover, T1-infected cell cultures showed a strongly increased incorporation of [14C]GCV triphosphate into macromolecular DNA, compared with recombinant A5 or vaccinia virus controls, which could be inhibited by the addition of guanosine. This shows that UL97 kinase is the only additional gene product required to make vaccinia virus susceptible to GCV, and guanosine seems to be one natural substrate for the enzyme. The system described here should be very helpful for fast and detailed functional analyses of UL97 mutations found in GCV-resistant HCMV isolates. Images

Metzger, C; Michel, D; Schneider, K; Luske, A; Schlicht, H J; Mertens, T

1994-01-01

273

Sedimentation of an RNA polymerase complex from vaccinia virus that specifically initiates and terminates transcription.  

PubMed Central

A high-molecular-weight protein complex that is capable of accurate transcription initiation and termination of vaccinia virus early genes without additional factors was demonstrated. The complex was solubilized by disruption of purified virions, freed of DNA by passage through a DEAE-cellulose column, and isolated by glycerol gradient sedimentation. All detectable RNA polymerase activity was associated with the transcription complex, whereas the majority of enzymes released from virus cores including mRNA (nucleoside-2'-O)methyltransferase, poly(A) polymerase, topoisomerase, nucleoside triphosphate phosphohydrolase II, protein kinase, and single-strand DNase sedimented more slowly. Activities corresponding to two enzymes, mRNA guanylyltransferase (capping enzyme) and nucleoside triphosphate phosphohydrolase I (DNA-dependent ATPase), partially sedimented with the complex. Silver-stained polyacrylamide gels, immunoblots, and autoradiographs confirmed the presence of subunits of vaccinia virus RNA polymerase, mRNA guanylyltransferase, and nucleoside triphosphate phosphohydrolase I, as well as additional unidentified polypeptides, in fractions with transcriptase activity. A possible role for the DNA-dependent ATPase was suggested by studies with ATP analogs with gamma-S or nonhydrolyzable beta-gamma-phosphodiester bonds. These analogs were used by vaccinia virus RNA polymerase to nonspecifically transcribe single-stranded DNA templates but did not support accurate transcription of early genes by the complex. Transcription also was sensitive to high concentrations of novobiocin; however, this effect could be attributed to inhibition of RNA polymerase or ATPase activities rather than topoisomerase. Images

Broyles, S S; Moss, B

1987-01-01

274

Vaccinia reporter viruses for quantifying viral function at all stages of gene expression.  

PubMed

Poxviruses are a family of double stranded DNA viruses that include active human pathogens such as monkeypox, molluscum contagiousum, and Contagalo virus. The family also includes the smallpox virus, Variola. Due to the complexity of poxvirus replication, many questions still remain regarding their gene expression strategy. In this article we describe the conceptualization and usage of recombinant vaccinia viruses that enable real-time measurement of single and multiple stages of viral gene expression in a high-throughput format. This is enabled through the use of spectrally distinct fluorescent proteins as reporters for each of three stages of viral replication. These viruses provide a high signal-to-noise ratio while retaining stage specific expression patterns, enabling plate-based assays and microscopic observations of virus propagation and replication. These tools have uses for antiviral discovery, studies of the virus-host interaction, and evolutionary biology. PMID:24894622

Rozelle, Daniel K; Filone, Claire Marie; Dower, Ken; Connor, John H

2014-01-01

275

Antagonizing activity of vaccinia virus E3L against human interferons in Huh7 cells  

Microsoft Academic Search

The E3L protein of vaccinia virus (VV) is well known for its capacity to evade cellular innate antiviral immunity related to interferon (IFN), for example PKR and RNaseL mediated antiviral activities. However, due to the limited range of cells that support VV E3L deletion mutant replication, the full capacity of E3L inhibiting the innate immune response induced by IFNs remains

Janilyn Arsenio; Yvon Deschambault; Jingxin Cao

2008-01-01

276

Inhibition of I?B Kinase by Vaccinia Virus Virulence Factor B14  

Microsoft Academic Search

The I?B kinase (IKK) complex is a key regulator of signal transduction pathways leading to the induction of NF-?B-dependent gene expression and production of pro-inflammatory cytokines. It therefore represents a major target for the development of anti-inflammatory therapeutic drugs and may be targeted by pathogens seeking to diminish the host response to infection. Previously, the vaccinia virus (VACV) strain Western

Ron A.-J. Chen; Grigory Ryzhakov; Samantha Cooray; Felix Randow; Geoffrey L. Smith

2008-01-01

277

Preferential virosomal location of underphosphorylated H5R protein synthesized in vaccinia virus-infected cells  

Microsoft Academic Search

The phosphorylation state of vaccinia virus (VV) protein H5R synthesized in infected cells was in- vestigated by two-dimensional gel electrophoresis. Most of the H5R protein was underphosphorylated (pI 5- 9t o 6 -8) and, on centrifugation of cell lysates, was associated with virosomes sedimenting with nuclei. However, about a quarter of the H5R protein synthesized was highly phosphorylated (pI 5-5),

Georges Beaud; Ruth Beaud

278

Complementation Analysis of the Dales Collection of Vaccinia Virus Temperature-Sensitive Mutants  

Microsoft Academic Search

A collection of randomly generated temperature-sensitive (ts) vaccinia virus (strain IHD-W) mutants were reported by S. Dales et al., (1978, Virology, 84, 403–428) in 1978 and characterized by electron microscopy. We have performed further genetic analysis on the Dales collection of mutants to make the mutants more useful to the scientific community. We obtained the entire Dales collection, 97 mutants,

Cari A. Lackner; Susan M. D'Costa; Charles Buck; Richard C. Condit

2003-01-01

279

Induction of HIV Immunity in the Genital Tract After Intranasal Delivery of a MVA Vector: Enhanced Immunogenicity After DNA Prime-Modified Vaccinia Virus Ankara Boost Immunization Schedule1  

Microsoft Academic Search

Vaccines intended to prevent mucosal transmission of HIV should be able to induce multiple immune effectors in the host including Abs and cell-mediated immune responses at mucosal sites. The aim of this study was to characterize and to enhance the immu- nogenicity of a recombinant modified vaccinia virus Ankara (MVA) expressing HIV-1 Env IIIB Ag (MVAenv) inoculated in BALB\\/c mice

M. Magdalena Gherardi; Eva Perez-Jimenez; JoseLuis Najera; Mariano Esteban

280

Strikingly poor CD8(+) T-cell immunogenicity of vaccinia virus strain MVA in BALB/c mice.  

PubMed

Vaccinia virus (VACV) strain MVA is a highly attenuated vector for vaccines that is being explored in clinical trials. We compared the CD8(+) T-cell immunogenicity of MVA with that of a virulent laboratory strain of VACV (strain WR) in BALB/c mice by examining epitope-specific responses as well as estimating the total number of activated CD8(+) T cells, irrespective of specificity. We found that MVA elicited total CD8(+) T-cell responses that were reduced by at least 20-fold compared with strain WR in BALB/c mice. In C57Bl/6 mice, we also found a substantial difference in immunogenicity between these VACV strains, but it was more modest at around fivefold. Of note, the size of responses to the virulent WR virus was similar in both strains of mice suggesting that BALB/c mice can mount robust CD8(+) T-cell responses to VACV. Although the data for total responses clearly showed that MVA overall is poorly immunogenic in BALB/c mice, we found one epitope for which strong responses were made irrespective of virus strain. Therefore, in the context of a vaccine, some recombinant epitopes may have similar immunogenicity when expressed from MVA and other strains of VACV, but we would expect these to be exceptions. These data show clearly the substantial difference in immunogenicity between MVA and virulent VACV strains and suggest that the impact of host genetics on responses to attenuated vaccine vectors like MVA requires more consideration. PMID:24566805

Russell, Tiffany A; Tscharke, David C

2014-05-01

281

Characterization of a New Vaccinia virus Isolate Reveals the C23L Gene as a Putative Genetic Marker for Autochthonous Group 1 Brazilian Vaccinia virus  

PubMed Central

Since 1999, several Vaccinia virus (VACV) isolates, the etiological agents of bovine vaccinia (BV), have been frequently isolated and characterized with various biological and molecular methods. The results from these approaches have grouped these VACV isolates into two different clusters. This dichotomy has elicited debates surrounding the origin of the Brazilian VACV and its epidemiological significance. To ascertain vital information to settle these debates, we and other research groups have made efforts to identify molecular markers to discriminate VACV from other viruses of the genus Orthopoxvirus (OPV) and other VACV-BR groups. In this way, some genes have been identified as useful markers to discriminate between the VACV-BR groups. However, new markers are needed to infer ancestry and to correlate each sample or group with its unique epidemiological and biological features. The aims of this work were to characterize a new VACV isolate (VACV DMTV-2005) molecularly and biologically using conserved and non-conserved gene analyses for phylogenetic inference and to search for new genes that would elucidate the VACV-BR dichotomy. The VACV DMTV-2005 isolate reported in this study is biologically and phylogenetically clustered with other strains of Group 1 VACV-BR, the most prevalent VACV group that was isolated during the bovine vaccinia outbreaks in Brazil. Sequence analysis of C23L, the gene that encodes for the CC-chemokine-binding protein, revealed a ten-nucleotide deletion, which is a new Group 1 Brazilian VACV genetic marker. This deletion in the C23L open reading frame produces a premature stop-codon that is shared by all Group 1 VACV-BR strains and may also reflect the VACV-BR dichotomy; the deletion can also be considered to be a putative genetic marker for non-virulent Brazilian VACV isolates and may be used for the detection and molecular characterization of new isolates.

Oliveira, Danilo B.; Franco-Luiz, Ana P. M.; Campos, Rafael K.; Guedes, Maria I. M.; Fonseca, Flavio G.; Trindade, Giliane S.; Drumond, Betania P.; Kroon, Erna G.; Abrahao, Jonatas S.

2012-01-01

282

Vaccinia virus gene A18R encodes an essential DNA helicase.  

PubMed Central

The vaccinia virus A18R protein is a DNA-dependent ATPase that contains the canonical sequence motifs associated with the DEXH group of DNA and RNA helicases. Investigation of A18R protein function during infection indicated it functions in the early and late phases of vaccinia virus transcription. The A18R protein shares sequence similarity with the mammalian DNA helicase ERCC3. The ERCC3 protein has a dual function: it is a component of the transcription factor TFIIH and is an essential participant in the cellular nucleotide excision repair pathway. Here we present evidence that the A18R protein is a DNA helicase that unwinds duplex DNA in a 3'-to-5' direction. The A18R helicase was inactive on RNA-DNA and RNA-RNA hybrids. The A18R unwinding activity was most efficient on DNA substrates with lengths of 20 nucleotides or less, and its unwinding activity was not stimulated by the addition of Escherichia coli single-strand-binding protein (SSB), the bacteriophage T4 gene 32 SSB, or the vaccinia virus I3L protein, a putative SSB. We have used an electrophoretic gel mobility shift assay to show that the A18R protein forms a stable complex with single-stranded DNA, and to a lesser extent RNA, in a reaction that does not require ATP.

Simpson, D A; Condit, R C

1995-01-01

283

Live Virus Smallpox Vaccine  

MedlinePLUS

... for Clinicians Brucella Lab Info Surveillance & Investigation Cholera Ebola virus E. coli Food safety threats Glanders Lassa fever Marburg virus Melioidosis Plague Case Definitions and Report Forms Diagnosis & Evaluation Infection Control Lab Testing Surveillance & Investigation Publications & ...

284

Vaccinia virus protein C4 inhibits NF-?B activation and promotes virus virulence  

PubMed Central

Vaccinia virus (VACV) strain Western Reserve protein C4 has been characterized and its function and contribution to virus virulence assessed. Bioinformatic analysis showed that C4 is conserved in six orthopoxvirus species and shares 43?% amino acid identity with VACV protein C16, a known virulence factor. A recombinant VACV expressing a C-terminally tagged version of C4 showed that, like C16, this 37 kDa protein is expressed early during infection and localizes to both the cytoplasm and the nucleus. Functional assays using a firefly luciferase reporter plasmid under the control of a nuclear factor kappa B (NF-?B)-dependent promoter demonstrated that C4 inhibits NF-?B activation at, or downstream of, the inhibitor of kappa kinase (IKK) complex. Consistent with this, C4 inhibited interleukin-1?-induced translocation of p65 into the nucleus. A VACV lacking the C4L gene (v?C4) showed no significant differences from wild-type virus in growth kinetics or spread in cell culture, but had reduced virulence in a murine intranasal model of infection. v?C4-infected mice exhibited fewer symptoms, lost less weight and recovered 7 days earlier than animals infected with control viruses expressing C4. Furthermore, bronchoalveolar lavage fluid from v?C4-infected mice had increased cell numbers at day 5 post-infection, which correlated with reduced lung virus titres from this time onward. C4 represents the ninth VACV protein to inhibit NF-?B activation and remarkably, in every case examined, loss of each protein individually caused an alteration in virus virulence, despite the presence of other NF-?B inhibitors.

Ember, Stuart W. J.; Ren, Hongwei; Ferguson, Brian J.

2012-01-01

285

Studies of Respiratory Syncytial Virus Vaccines.  

National Technical Information Service (NTIS)

Studies of Respiratory Syncytial virus vaccines included both biological and physical aspects with the principle objective being a study of the banding of RS virus in the density gradient ultracentrifuge. Various studies on virus propagation both in the B...

R. N. Hull C. B. Reimer L. F. Ellis

1966-01-01

286

Recombinant vaccines against bluetongue virus.  

PubMed

Bluetongue (BT) is a hemorrhagic disease of ruminants caused by bluetongue virus (BTV), the prototype member of the genus Orbivirus within the family Reoviridae and is transmitted via biting midges of the genus Culicoides. BTV can be found on all continents except Antarctica, and up to 26 immunologically distinct BTV serotypes have been identified. Live attenuated and inactivated BTV vaccines have been used over the years with different degrees of success. The multiple outbreaks of BTV in Mediterranean Europe in the last two decades and the incursion of BTV-8 in Northern Europe in 2008 has re-stimulated the interest to develop improved vaccination strategies against BTV. In particular, safer, cross-reactive, more efficacious vaccines with differential diagnostic capability have been pursued by multiple BTV research groups and vaccine manufacturers. A wide variety of recombinant BTV vaccine prototypes have been investigated, ranging from baculovirus-expressed sub-unit vaccines to the use of live viral vectors. This article gives a brief overview of all these modern approaches to develop vaccines against BTV including some recent unpublished data. PMID:24287057

Calvo-Pinilla, Eva; Castillo-Olivares, Javier; Jabbar, Tamara; Ortego, Javier; de la Poza, Francisco; Marín-López, Alejandro

2014-03-01

287

Vaccinia virus, herpes simplex virus, and carcinogens induce DNA amplification in a human cell line and support replication of a helpervirus dependent parvovirus  

SciTech Connect

The SV40-transformed human kidney cell line, NB-E, amplifies integrated as well as episomal SV40 DNA upon treatment with chemical (DMBA) or physical (uv irradiation) carcinogens (initiators) as well as after infection with herpes simplex virus (HSV) type 1 or with vaccinia virus. In addition it is shown that vaccinia virus induces SV40 DNA amplification also in the SV40-transformed Chinese hamster embryo cell line, CO631. These findings demonstrate that human cells similar to Chinese hamster cells amplify integrated DNA sequences after treatment with carcinogens or infection with specific viruses. Furthermore, a poxvirus--vaccinia virus--similar to herpes group viruses induces DNA amplification. As reported for other systems, the vaccinia virus-induced DNA amplification in NB-E cells is inhibited by coinfection with adeno-associated virus (AAV) type 5. This is in line with previous studies on inhibition of carcinogen- or HSV-induced DNA amplification in CO631 cells. The experiments also demonstrate that vaccinia virus, in addition to herpes and adenoviruses acts as a helper virus for replication and structural antigen synthesis of AAV-5 in NB-E cells.

Schlehofer, J.R.; Ehrbar, M.; zur Hausen, H.

1986-07-15

288

Subunit Recombinant Vaccine Protects against Monkeypox 1  

Microsoft Academic Search

The smallpox vaccine Dryvax, a live vaccinia virus (VACV), protects against smallpox and monkeypox, but is contraindicated in immunocompromised individuals. Because Abs to VACV mediate protection, a live virus vaccine could be substituted by a safe subunit protein-based vaccine able to induce a protective Ab response. We immunized rhesus macaques with plasmid DNA encoding the monkeypox orthologs of the VACV

Jean-Michel Heraud; Yvette Edghill-Smith; Victor Ayala; Irene Kalisz; Janie Parrino; Vaniambadi S. Kalyanaraman; Jody Manischewitz; Lisa R. King; Anna Hryniewicz; Christopher J. Trindade; Meredith Hassett; Wen-Po Tsai; David Venzon; Aysegul Nalca; Monica Vaccari; Peter Silvera; Mike Bray; Barney S. Graham; Hana Golding; Jay W. Hooper; Genoveffa Franchini

289

The vaccinia virus C12L protein inhibits mouse IL18 and promotes virus virulence in the murine intranasal model  

Microsoft Academic Search

A bioassay that measured the interleukin (IL)-12-induced production of interferon (IFN)-c from mouse splenocytes was used to identify a soluble factor in the supernatants of vaccinia virus (VV)- infected cells that inhibited the production of IFN-c. This soluble factor was expressed by 14 out of 16 VV strains including the Western Reserve (WR) strain, but strains Copenhagen and Tashkent and

Julian A. Symons; Elizabeth Adams; David C. Tscharke; Patrick C. Reading; Herman Waldmann; Geoffrey L. Smith

290

Intratumoral delivery of recombinant vaccinia virus encoding for ErbB2/Neu inhibits the growth of salivary gland carcinoma cells  

PubMed Central

Background The antitumor activity induced by intratumoral vaccination with poxvirus expressing a tumor antigen was shown to be superior to that induced by subcutaneous vaccination. Salivary gland carcinomas overexpress ErbB2. Trastuzumab, a monoclonal antibody to ErbB2, was proposed for salivary gland tumors treatment. We explored the effectiveness of intratumoral vaccination with the recombinant vaccinia virus ErbB2/Neu (rV-neuT) vaccine in hampering the growth of transplanted Neu-overexpressing BALB-neuT salivary gland cancer cells (SALTO) in BALB-neuT mice. Methods BALB-neuT male mice were subcutaneously injected with SALTO tumor cells and intratumorally vaccinated twice with different doses of either rV-neuT or V-wt (wild-type). Tumors were measured weekly. The presence of anti-ErbB2/Neu antibodies was assayed by ELISA, immunoprecipitation or indirect immunofluorescence. Biological activity of immune sera was investigated by analyzing antibody-dependent cellular cytotoxicity (ADCC), SALTO cells proliferation and apoptosis, ErbB2/Neu receptor down regulation and ERK1/2 phosphorylation. Anti-Neu T cell immunity was investigated by determining the release of IL-2 and IFN-gamma in T cells supernatant. Survival curves were determined using the Kaplan-Meier method and compared using the log-rank test. Differences in tumor volumes, number of apoptotic cells, titer of the serum, percentage of ADCC were evaluated through a two-tailed Student’s t-test. Results rV-neuT intratumoral vaccination was able to inhibit the growth of SALTO cancer cells in a dose-dependent manner. The anti-Neu serum titer paralleled in vivo antitumor activity of rV-neuT vaccinated mice. rV-neuT immune serum was able to mediate ADCC, inhibition of SALTO cells proliferation, down regulation of the ErbB2/Neu receptor, inhibition of ERK1/2 phosphorylation and induction of apoptosis, thus suggesting potential mechanisms of in vivo tumor growth interference. In addition, spleen T cells of rV-neuT vaccinated mice released IFN-gamma and IL-2 upon in vitro stimulation with several Neu-specific peptides located in the extracellular domain of Neu sequence. Conclusions rV-neuT intratumoral vaccination could be employed to induce an efficient antitumor response and reject transplanted salivary gland tumors. Our findings may have important implications for the design of cancer vaccine protocols for the treatment of salivary gland tumors and other accessible tumors using intratumoral injection of recombinant vaccinia virus.

2014-01-01

291

Non-replicating recombinant vaccinia virus expressing CD80 to enhance T-cell stimulation.  

PubMed

The following method describes the generation of a recombinant vaccinia virus expressing a costimulatory molecule (human CD80 or B7.1).The procedure first requires the cloning, by classical methods not described here, of the gene of interest, e.g. CD80, into a vaccinia shuttle plasmid under the control of a virus-specific promoter enabling a transcription during the early phase of infection. Flanking the insert, the plasmid contains viral sequences and a selection maker needed for the insertion into the viral genome. The successive plaque isolation of recombinant virus on cell monolayer described here is based on the transient "gpt" selection system which enables other insertions in different loci of the same virus. Finally, after verification amplification and titration of the recombinant vector, replication will be impaired by a psoralen-UV treatment in order to produce a non-replicating virus. Expression and function of inserts, following infection of cells, are verified by specific phenotypic and functional assays. PMID:19048219

Zajac, Paul

2009-01-01

292

Production and characterization of mammalian virus-like particles from modified vaccinia virus Ankara vectors expressing influenza H5N1 hemagglutinin and neuraminidase.  

PubMed

Several studies have described the production of influenza virus-like particles (VLP) using a variety of platform systems. These VLPs are non-replicating particles that spontaneously self-assemble from expressed influenza virus proteins and have been proposed as vaccine candidates for both seasonal and pandemic influenza. Although still in the early stages of development and evaluation as influenza vaccines, influenza VLPs have a variety of other valuable uses such as examining and understanding correlates of protection against influenza and investigating virus-cell interactions. The most common production system for influenza VLPs is the baculovirus-insect cell expression which has several attractive features including the ease in which new gene combinations can be constructed, the immunogenicity elicited and protection afforded by the produced VLPs, and the scalability offered by the system. However, there are differences between the influenza VLPs produced by baculovirus expression systems in insect cells and the influenza viruses produced for use as current vaccines or the virus produced during a productive clinical infection. We describe here the development of a modified vaccinia virus Ankara (MVA) system to generate mammalian influenza VLPs containing influenza H5N1 proteins. The MVA vector system is flexible for manipulating and generating various VLP constructs, expresses high level of influenza hemagglutinin (HA), neuraminidase (NA), and matrix (M) proteins, and can be scaled up to produce VLPs in quantities sufficient for in vivo studies. We show that mammalian VLPs are generated from recombinant MVA vectors expressing H5N1 HA alone, but that increased VLP production can be achieved if NA is co-expressed. These mammalian H5N1 influenza VLPs have properties in common with live virus, as shown by electron microscopy analysis, their ability to hemagglutinate red blood cells, express neuraminidase activity, and to bind influenza specific antibodies. Importantly, these VLPs are able to elicit a protective immune response in a mouse challenge model, suggesting their utility in dissecting the correlates of immunity in such models. Mammalian derived VLPs may also provide a useful tool for studying virus-cell interactions and may have potential for development as pandemic vaccines. PMID:22465746

Schmeisser, Falko; Adamo, Joan E; Blumberg, Benjamin; Friedman, Rachel; Muller, Jacqueline; Soto, Jackeline; Weir, Jerry P

2012-05-14

293

Genome Sequence of WAU86/88-1, a New Variant of Vaccinia Virus Lister Strain from Poland  

PubMed Central

The poxviruses Warsaw Agricultural University 86 (WAU86) and 88-1 (WAU88-1) were isolated in 1986 to 1988 from separate outbreaks in laboratory mice in Poland and described as ectromelia virus isolates. The genome sequences of these poxviruses reveal that they are almost identical and represent a novel variant of the vaccinia virus Lister strain.

Mavian, Carla; Lopez-Bueno, Alberto

2014-01-01

294

CD4 and CD8 T cells participate in the immune memory response against Vaccinia virus after a previous natural infection.  

PubMed

The present study evaluates the immune response of memory CD4(+) and CD8(+) T cells from patients following a natural Vaccinia virus (VACV) infection. A total of 42 individuals were involved in the study being: 22 previously infected individuals (vaccinated or not against smallpox) and 20 non-infected individuals (vaccinated or not). A short-term in vitro stimulation with UV-inactivated VACV of whole blood cells was performed. Our study showed that previously infected individuals have a lower percentage of CD4(+) T cells expressing lymph-node homing receptors (CD4(+)CD62L(+)CCR7(+)) and higher percentage of memory CD4(+) T cells subsets (CD4(+)CD45RO(High)) when compared with non-infected subjects, after in vitro viral stimulation. We also showed that infected individuals presented higher percentages of CD4(+) and CD8(+) memory T lymphocytes expressing IFN-? when compared to non-infected individuals. We verified that the percentage of CD4(+) and CD8(+) T memory cells expressing TNF-? was higher in infected and non-infected vaccinated subjects when compared with non-infected unvaccinated individual. We also observed that previously infected individuals have higher percentages of CD8(+) T cells expressing lymph-node homing receptors (CCR7(+) and CD62L(+)) and that the memory T cells expressing IFN-? and TNF-? were at higher percentages in the whole blood cells from infected and non-infected vaccinated individuals, when compared to unvaccinated non-infected subjects. Thus, our findings suggest that CD4(+) and CD8(+) T cells are involved in the immune memory response against Vaccinia virus natural infection. PMID:24600565

Medeiros-Silva, Daniela Carla; Dos Santos Moreira-Silva, Eduardo Augusto; Assis Silva Gomes, Juliana de; da Fonseca, Flávio Guimarães; Correa-Oliveira, Rodrigo

2013-01-01

295

Virus-encoded ectopic CD74 enhances poxvirus vaccine efficacy.  

PubMed

Vaccinia virus (VV) has been used globally as a vaccine to eradicate smallpox. Widespread use of this viral vaccine has been tempered in recent years because of its immuno-evasive properties, with restrictions prohibiting VV inoculation of individuals with immune deficiencies or atopic skin diseases. VV infection is known to perturb several pathways for immune recognition including MHC class II (MHCII) and CD1d-restricted antigen presentation. MHCII and CD1d molecules associate with a conserved intracellular chaperone, CD74, also known as invariant chain. Upon VV infection, cellular CD74 levels are significantly reduced in antigen-presenting cells, consistent with the observed destabilization of MHCII molecules. In the current study, the ability of sustained CD74 expression to overcome VV-induced suppression of antigen presentation was investigated. Viral inhibition of MHCII antigen presentation could be partially ameliorated by ectopic expression of CD74 or by infection of cells with a recombinant VV encoding murine CD74 (mCD74-VV). In contrast, virus-induced disruptions in CD1d-mediated antigen presentation persisted even with sustained CD74 expression. Mice immunized with the recombinant mCD74-VV displayed greater protection during VV challenge and more robust anti-VV antibody responses. Together, these observations suggest that recombinant VV vaccines encoding CD74 may be useful tools to improve CD4? T-cell responses to viral and tumour antigens. PMID:24205828

Walline, Crystal C; Deffit, Sarah N; Wang, Nan; Guindon, Lynette M; Crotzer, Victoria L; Liu, Jianyun; Hollister, Kristin; Eisenlohr, Laurence C; Brutkiewicz, Randy R; Kaplan, Mark H; Blum, Janice S

2014-04-01

296

RNAi screening reveals proteasome- and Cullin3-dependent stages in vaccinia virus infection.  

PubMed

A two-step, automated, high-throughput RNAi silencing screen was used to identify host cell factors required during vaccinia virus infection. Validation and analysis of clustered hits revealed previously unknown processes during virus entry, including a mechanism for genome uncoating. Viral core proteins were found to be already ubiquitinated during virus assembly. After entering the cytosol of an uninfected cell, the viral DNA was released from the core through the activity of the cell's proteasomes. Next, a Cullin3-based ubiquitin ligase mediated a further round of ubiquitination and proteasome action. This was needed in order to initiate viral DNA replication. The results accentuate the value of large-scale RNAi screens in providing directions for detailed cell biological investigation of complex pathways. The list of cell functions required during poxvirus infection will, moreover, provide a resource for future virus-host cell interaction studies and for the discovery of antivirals. PMID:23084750

Mercer, Jason; Snijder, Berend; Sacher, Raphael; Burkard, Christine; Bleck, Christopher Karl Ernst; Stahlberg, Henning; Pelkmans, Lucas; Helenius, Ari

2012-10-25

297

The heterogeneity of human antibody responses to vaccinia virus revealed through use of focused protein arrays.  

PubMed

The renewed interest in strategies to combat infectious agents with epidemic potential has led to a re-examination of vaccination protocols against smallpox. To help define which antigens elicit a human antibody response, we have targeted proteins known or predicted to be presented on the surface of the intracellular mature virion (IMV) or the extracellular enveloped virion (EEV). The predicted ectodomains were expressed in a mammalian in vitro coupled transcription/translation reaction using tRNA(lys) precharged with lysine-epsilon-biotin followed by solid phase immobilization on 384-well neutravidin-coated plates. The generated array is highly specific and sensitive in a micro-ELISA format. By comparison of binding of vaccinia-immune sera to the reticulocyte lysate-produced proteins and to secreted post-translationally modified proteins, we demonstrate that for several proteins including the EEV proteins B5 and A33, proper recognition is dependent upon appropriate folding, with little dependence upon glycosylation per se. We further demonstrate that the humoral immune response to vaccinia among different individuals is not uniform in specificity or strength, as different IMV and EEV targets predominate within the group of immunogenic proteins. This heterogeneity likely results from the diversity of HLA Class II alleles and CD4 T helper cell epitopes stimulating B cell antibody production. Our findings have important implications both for design of new recombinant subunit vaccines as well as for methods of assaying the human antibody response utilizing recombinant proteins produced in vitro. PMID:19146908

Duke-Cohan, Jonathan S; Wollenick, Kristin; Witten, Elizabeth A; Seaman, Michael S; Baden, Lindsey R; Dolin, Raphael; Reinherz, Ellis L

2009-02-18

298

Vaccinia virus protein C16 acts intracellularly to modulate the host response and promote virulence  

PubMed Central

The vaccinia virus (VACV) strain Western Reserve C16 protein has been characterized and its effects on virus replication and virulence have been determined. The C16L gene is present in the inverted terminal repeat and so is one of the few VACV genes that are diploid. The C16 protein is highly conserved between different VACV strains, and also in the orthopoxviruses variola virus, ectromelia virus, horsepox virus and cowpox virus. C16 is a 37.5?kDa protein, which is expressed early during infection and localizes to the cell nucleus and cytoplasm of infected and transfected cells. The loss of the C16L gene had no effect on virus growth kinetics but did reduce plaque size slightly. Furthermore, the virulence of a virus lacking C16L (v?C16) was reduced in a murine intranasal model compared with control viruses and there were reduced virus titres from 4?days post-infection. In the absence of C16, the recruitment of inflammatory cells in the lung and bronchoalveolar lavage was increased early after infection (day 3) and more CD4+ and CD8+ T cells expressed the CD69 activation marker. Conversely, late after infection with v?C16 (day 10) there were fewer T cells remaining, indicating more rapid clearance of infection. Collectively, these data indicate that C16 diminishes the immune response and is an intracellular immunomodulator.

Fahy, Aodhnait S.; Clark, Richard H.; Glyde, Emily F.; Smith, Geoffrey L.

2008-01-01

299

Kunjin Virus Replicon Vaccine Vectors Induce Protective CD8+ T-Cell Immunity  

PubMed Central

The ability of self-replicating RNA (replicon) vaccine vectors derived from the Australian flavivirus Kunjin (KUN) to induce protective ?? CD8+ T-cell responses was examined. KUN replicons encoding a model immunogen were delivered by three different vaccine modalities: (i) as naked RNA transcribed in vitro, (ii) as plasmid DNA constructed to allow in vivo transcription of replicon RNA by cellular RNA polymerase II (DNA based), and (iii) as replicon RNA encapsidated into virus-like particles. A single immunization with any of these KUN replicon vaccines induced CD8+ T-cell responses at levels comparable to those induced by recombinant vaccinia virus encoding the same immunogen. Immunization with only 0.1 ?g of DNA-based KUN replicons elicited CD8+ T-cell responses similar to those seen after immunization with 100 ?g of a conventional DNA vaccine. Naked RNA immunization with KUN replicons also protected mice against challenges with recombinant vaccinia virus and B16 tumor cells. These results demonstrate the value of KUN replicon vectors for inducing protective antiviral and anticancer CD8+ T-cell responses.

Anraku, Itaru; Harvey, Tracey J.; Linedale, Richard; Gardner, Joy; Harrich, David; Suhrbier, Andreas; Khromykh, Alexander A.

2002-01-01

300

Effects of granulocyte-macrophage colony stimulating factor gene encoded vaccinia virus vector on murine pulmonary metastatic melanoma  

Microsoft Academic Search

A recombinant vaccinia virus expressing marine granulocyte-macrophage colony-stimulating factor (VVGM-CSF) was tested for\\u000a its antitumor activity. Murine pulmonary metastasis was established by injecting 2×105 B16F10 melanoma cells into the tail vein of C57BL\\/6 mice. Three days after B16F10 inoculation, WGM-CSF or VVTK, a thymidine\\u000a kinase gene deficient control vaccinia virus, were injected intraperitoneally twice weekly for 2 weeks. Two weeks

Dianwen Ju; Xuetao Cao; Tao Wan; Shihua Ma; Baomei Wang; Yizhi Yu; Tianxing Ye

1997-01-01

301

One More Piece in the VACV Ecological Puzzle: Could Peridomestic Rodents Be the Link between Wildlife and Bovine Vaccinia Outbreaks in Brazil?  

Microsoft Academic Search

BackgroundDespite the fact that smallpox eradication was declared by the World Health Organization (WHO) in 1980, other poxviruses have emerged and re-emerged, with significant public health and economic impacts. Vaccinia virus (VACV), a poxvirus used during the WHO smallpox vaccination campaign, has been involved in zoonotic infections in Brazilian rural areas (Bovine Vaccinia outbreaks – BV), affecting dairy cattle and

Jônatas S. Abrahão; Maria Isabel M. Guedes; Giliane S. Trindade; Flávio G. Fonseca; Rafael K. Campos; Bruno F. Mota; Zélia I. P. Lobato; André T. Silva-Fernandes; Gisele O. L. Rodrigues; Larissa S. Lima; Paulo C. P. Ferreira; Cláudio A. Bonjardim; Erna G. Kroon; Joel Mark Montgomery

2009-01-01

302

Identification of the DNA sequences encoding the large subunit of the mRNA-capping enzyme of vaccinia virus.  

PubMed Central

The DNA sequences encoding the large subunit of the mRNA-capping enzyme of vaccinia virus were located on the viral genome. The formation of an enzyme-guanylate covalent intermediate labeled with [alpha-32P]GTP allowed the identification of the large subunit of the capping enzyme and was used to monitor the appearance of the enzyme during the infectious cycle. This assay confirmed that after vaccinia infection, a novel 84,000-molecular-weight polypeptide corresponding to the large subunit was rapidly synthesized before viral DNA replication. Hybrid-selected cell-free translation of early viral mRNA established that vaccinia virus encoded a polypeptide identical in molecular weight with the 32P-labeled 84,000-molecular-weight polypeptide found in vaccinia virions. Like the authentic capping enzyme, this virus-encoded cell-free translation product bound specifically to DNA-cellulose. A comparison of the partial proteolytic digestion fragments generated by V8 protease, chymotrypsin, and trypsin demonstrated that the 32P-labeled large subunit and the [35S]methionine-labeled cell-free translation product were identical. The mRNA encoding the large subunit of the capping enzyme was located 3.1 kilobase pairs to the left of the HindIII D restriction fragment of the vaccinia genome. Furthermore, the mRNA was determined to be 3.0 kilobases in size, and its 5' and 3' termini were precisely located by S1 nuclease analysis. Images

Morgan, J R; Cohen, L K; Roberts, B E

1984-01-01

303

Towards a Vaccine Against Ebola Virus.  

National Technical Information Service (NTIS)

Ebola virus infection causes hemorrhagic fever with high mortality rates in humans and nonhuman primates. Currently, there are no vaccines or therapies approved for human use. Outbreaks of Ebola virus have been infrequent, largely confined to remote locat...

T. W. Geisbert P. B. Jahrling

2003-01-01

304

A source of glycosylated human T-cell lymphotropic virus type 1 envelope protein: expression of gp46 by the vaccinia virus/T7 polymerase system.  

PubMed

Heterologous expression of the human T-cell lymphotropic virus type 1 (HTLV-1) envelope surface glycoprotein (gp46) in a vaccinia virus/T7 polymerase system resulted in the production of authentic recombinant gp46. Five differentially glycosylated forms of the surface envelope protein were produced by this mammalian system, as demonstrated by tunicamycin inhibition of N-glycosylation and N-glycan removal with endoglycosidase H and glycopeptidase F. These studies revealed that all four potential N-glycosylation sites in gp46 were used for oligosaccharide modification and that the oligosaccharides were mannose-rich and/or hybrid in composition. Conformational integrity of the recombinant HTLV-1 envelope protein was determined by the ability to bind to various HTLV-1-infected human sera and a panel of conformational-dependent human monoclonal antibodies under nondenaturing conditions. Furthermore, this recombinant gp46 was recognized by a series of HTLV-2-infected human sera and sera from a Pan paniscus chimpanzee infected with the distantly related simian T-cell lymphotropic virus STLVpan-p. Maintenance of highly conserved conformational epitopes in the recombinant HTLV-1 envelope protein structure suggests that it may serve as a useful diagnostic reagent and an effective vaccine candidate. PMID:8892853

Arp, J; LeVatte, M; Rowe, J; Perkins, S; King, E; Leystra-Lantz, C; Foung, S K; Dekaban, G A

1996-11-01

305

Membrane-bound complement regulatory activity is decreased on vaccinia virus-infected cells.  

PubMed Central

Decay accelerating factor (DAF), membrane cofactor protein (MCP), complement receptor 1 and mouse Crry are cell surface-bound complement regulatory proteins capable of inhibiting C3 convertase activity on cell membranes, and therefore provide a substantial protection from attack by homologous complement activated either by the classical or by the alternative pathway. Decrease in complement regulatory activity might lead to spontaneous complement deposition and subsequent cell injury. MoAb 5I2 can inhibit the complement regulatory activity of molecules on rat cells, resulting in deposition of homologous complement. The antigen recognized by 5I2 MoAb in rats is homologous to mouse Crry. Fifteen to 20 h after infection with vaccinia virus, in vitro cultured KDH-8 rat hepatoma cells show a strong decrease in expression of Crry-like antigen, and proved to be sensitive to complement deposition when 1:5 diluted normal rat serum was added to the culture medium as a source of complement. Addition of complement to the cultured KDH-8 cells infected with a very low dose of vaccinia virus (1 plaque-forming unit (PFU)/1000 cells) substantially reduced spreading of virus infection in the cell culture, while inactivation of complement by heat or zymosan treatment abrogated the protective effect.

Baranyi, L; Okada, N; Baranji, K; Takizawa, H; Okada, H

1994-01-01

306

Correlates Between Host and Viral Transcriptional Program Associated with Different Oncolytic Vaccinia Virus Isolates  

PubMed Central

Abstract Vaccinia virus (VACV) has emerged as an attractive tool in oncolytic virotherapy. VACV replication efficiency plays a crucial role in the therapeutic outcome. However, little is known about the influence of host factors on viral replication efficiency and permissiveness of a host cell line to infection and oncolysis. In this study, replication of the attenuated VACV GLV-1h68 strain and three wild-type VACV isolates was determined in two autologous human melanoma cell lines (888-MEL and 1936-MEL). Host gene expression and viral gene expression in infected cells were evaluated via respective expression array platforms. Microarray analyses followed by sequential statistical approaches characterized human genes that change specifically due to virus infection. Viral gene transcription correlated with viral replication in a time-dependent manner. A set of human genes revealed strong correlations with the respective viral gene expression. Finally we identified a set of human genes with possible predictive value for viral replication in an independent dataset. The results demonstrate a probable correlation between viral replication, early gene expression, and the respective host response, and thus a possible involvement of human host factors in viral early replication. The characterization of human target genes that influence viral replication could help answer the question of host cell permissiveness to oncolytic virotherapy and provide important information for the development of novel recombinant vaccinia viruses with improved features to enhance replication rate and hence trigger therapeutic outcome.

Reinboth, Jennifer; Ascierto, Maria L.; Chen, Nanhai G.; Zhang, Qian; Yu, Yong A.; Aguilar, Richard J.; Carretero, Rafael; Worschech, Andrea; Zhao, Yingdong; Wang, Ena; Marincola, Francesco M.

2012-01-01

307

The Vaccinia Virus A18R DNA Helicase Is a Postreplicative Negative Transcription Elongation Factor  

PubMed Central

Loss of vaccinia virus A18R gene function results in an aberrant transcription profile termed promiscuous transcription, defined as transcription within regions of the genome which are normally transcriptionally silent late during infection. Promiscuous transcription results in an increase in the intracellular concentration of double-stranded RNA, which in turn results in activation of the cellular 2-5A pathway and subsequent RNase L-catalyzed degradation of viral and cellular RNAs. One of three hypotheses could account for promiscuous transcription: (i) reactivation of early promoters late during infection, (ii) random transcription initiation, (iii) readthrough transcription from upstream promoters. Transcriptional analysis of several viral genes, presented here, argues strongly against the first two hypotheses. We have tested the readthrough hypothesis by conducting a detailed transcriptional analysis of a region of the vaccinia virus genome which contains three early genes (M1L, M2L, and K1L) positioned directly downstream of the intermediate gene, K2L. The results show that mutation of the A18R gene results in increased readthrough transcription of the M1L gene originating from the K2L intermediate promoter. A18R mutant infection of RNase L knockout mouse fibroblast (KO3) cells does not result in 2-5A pathway activation, yet the virus mutant is defective in late viral gene expression and remains temperature sensitive. These results demonstrate that the A18R gene product is a negative transcription elongation factor for postreplicative viral genes.

Xiang, Ying; Simpson, David A.; Spiegel, Jason; Zhou, Aimin; Silverman, Robert H.; Condit, Richard C.

1998-01-01

308

Deletion of C7L and K1L genes leads to significantly decreased virulence of recombinant vaccinia virus TianTan.  

PubMed

The vaccinia virus TianTan (VTT) has been modified as an HIV vaccine vector in China and has shown excellent performance in immunogenicity and safety. However, its adverse effects in immunosuppressed individuals warrant the search for a safer vector in the following clinic trails. In this study, we deleted the C7L and K1L genes of VTT and constructed six recombinant vaccinia strains VTT?C7L, VTT?K1L, VTT?C7LK1L, VTKgpe?C7L, VTKgpe?K1L and VTT?C7LK1L-gag. The pathogenicity and immunogenicity of these recombinants were evaluated in mouse and rabbit models. Comparing to parental VTT, VTT?C7L and VTT?K1L showed significantly decreased replication capability in CEF, Vero, BHK-21 and HeLa cell lines. In particular, replication of VTT?C7LK1L decreased more than 10-fold in all four cell lines. The virulence of all these mutants were decreased in BALB/c mouse and rabbit models; VTT?C7LK1L once again showed the greatest attenuation, having resulted in no evident damage in mice and erythema of only 0.4 cm diameter in rabbits, compared to 1.48 cm for VTT. VTKgpe?C7L, VTKgpe?K1L and VTT?C7LK1L-gag elicited as strong cellular and humoral responses against HIV genes as did VTKgpe, while humoral immune response against the vaccinia itself was reduced by 4-8-fold. These data show that deletion of C7L and K1L genes leads to significantly decreased virulence without compromising animal host immunogenicity, and may thus be key to creating a more safe and effective HIV vaccine vector. PMID:23840887

Liu, Zheng; Wang, Shuhui; Zhang, Qicheng; Tian, Meijuan; Hou, Jue; Wang, Rongmin; Liu, Chang; Ji, Xu; Liu, Ying; Shao, Yiming

2013-01-01

309

Deletion of C7L and K1L Genes Leads to Significantly Decreased Virulence of Recombinant Vaccinia Virus TianTan  

PubMed Central

The vaccinia virus TianTan (VTT) has been modified as an HIV vaccine vector in China and has shown excellent performance in immunogenicity and safety. However, its adverse effects in immunosuppressed individuals warrant the search for a safer vector in the following clinic trails. In this study, we deleted the C7L and K1L genes of VTT and constructed six recombinant vaccinia strains VTT?C7L, VTT?K1L, VTT?C7LK1L, VTKgpe?C7L, VTKgpe?K1L and VTT?C7LK1L-gag. The pathogenicity and immunogenicity of these recombinants were evaluated in mouse and rabbit models. Comparing to parental VTT, VTT?C7L and VTT?K1L showed significantly decreased replication capability in CEF, Vero, BHK-21 and HeLa cell lines. In particular, replication of VTT?C7LK1L decreased more than 10-fold in all four cell lines. The virulence of all these mutants were decreased in BALB/c mouse and rabbit models; VTT?C7LK1L once again showed the greatest attenuation, having resulted in no evident damage in mice and erythema of only 0.4 cm diameter in rabbits, compared to 1.48 cm for VTT. VTKgpe?C7L, VTKgpe?K1L and VTT?C7LK1L-gag elicited as strong cellular and humoral responses against HIV genes as did VTKgpe, while humoral immune response against the vaccinia itself was reduced by 4-8-fold. These data show that deletion of C7L and K1L genes leads to significantly decreased virulence without compromising animal host immunogenicity, and may thus be key to creating a more safe and effective HIV vaccine vector.

Liu, Zheng; Wang, Shuhui; Zhang, Qicheng; Tian, Meijuan; Hou, Jue; Wang, Rongmin; Liu, Chang; Ji, Xu; Liu, Ying; Shao, Yiming

2013-01-01

310

Plaque development by Vaccinia-viruses on the chicken chorio-allantois under the influence of Cyclophosphamide  

Microsoft Academic Search

The development of Vaccinia-virus plaques on the chorioallantois of embryonated eggs was studied under the influence of active cyclophosphamide metabolites. The viruses were inoculated on the 9th or the 13th incubation day. Cyclophosphamide was injected in doses of 62.5–500 µg in 0.2 ml NaCl into the yolk sac either 7 hrs after the virus inoculation or 3, 6, or 9

K. Norpoth; M. Huth; U. Witting; G. Maass

1975-01-01

311

Transmission of vaccinia virus, possibly through sexual contact, to a woman at high risk for adverse complications.  

PubMed

Severe adverse events, including eczema vaccinatum (EV), can result after smallpox vaccination. Persons at risk for EV include those with underlying dermatologic conditions, such as atopic dermatitis. We investigated a case of vaccinia infection, possibly acquired during sexual contact with a recently vaccinated military service member, in a female Maryland resident with atopic dermatitis. The U.S. Department of Defense's Vaccine Healthcare Centers Network (VHCN) and the Centers for Disease Control and Prevention (CDC) worked in conjunction with the patient's physician and the Maryland Department of Health and Mental Hygiene (DHMH) to confirm the diagnosis, ensure treatment, and prevent further transmission. Specimens collected from the patient were tested at the DHMH laboratories and were positive by real-time polymerase chain reaction for nonvariola orthopoxvirus. Testing at the CDC verified the presence of vaccinia-specific DNA signatures. Continuing spread of the patient's lesions led to the administration of vaccinia immune globulin and strict infection control measures to prevent tertiary transmission to vulnerable family members, also with atopic dermatitis. VHCN contacted the service member to reinforce vaccination site care and hygiene. This case underscores the importance of prevaccination education for those receiving the smallpox vaccine to protect contacts at risk for developing severe adverse reactions. PMID:24306023

Said, Maria A; Haile, Charles; Palabindala, Venkataraman; Barker, Naomi; Myers, Robert; Thompson, Ruth; Wilson, Lucy; Allan-Martinez, Frances; Montgomery, Jay; Monroe, Benjamin; Tack, Danielle; Reynolds, Mary; Damon, Inger; Blythe, David

2013-12-01

312

Inhibition of Vaccinia virus entry by a broad spectrum antiviral peptide  

SciTech Connect

Concerns about the possible use of Variola virus, the causative agent of smallpox, as a weapon for bioterrorism have led to renewed efforts to identify new antivirals against orthopoxviruses. We identified a peptide, EB, which inhibited infection by Vaccinia virus with an EC{sub 50} of 15 muM. A control peptide, EBX, identical in composition to EB but differing in sequence, was inactive (EC{sub 50} > 200 muM), indicating sequence specificity. The inhibition was reversed upon removal of the peptide, and EB treatment had no effect on the physical integrity of virus particles as determined by electron microscopy. Viral adsorption was unaffected by the presence of EB, and the addition of EB post-entry had no effect on viral titers or on early gene expression. The addition of EB post-adsorption resulted in the inhibition of beta-galactosidase expression from an early viral promoter with an EC{sub 50} of 45 muM. A significant reduction in virus entry was detected in the presence of the peptide when the number of viral cores released into the cytoplasm was quantified. Electron microscopy indicated that 88% of the virions remained on the surface of cells in the presence of EB, compared to 37% in the control (p < 0.001). EB also blocked fusion-from-within, suggesting that virus infection is inhibited at the fusion step. Analysis of EB derivatives suggested that peptide length may be important for the activity of EB. The EB peptide is, to our knowledge, the first known small molecule inhibitor of Vaccinia virus entry.

Altmann, S.E.; Jones, J.C. [Microbiology Doctoral Training Program, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706 (United States); Schultz-Cherry, S. [Microbiology Doctoral Training Program, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706 (United States); Department of Medical Microbiology and Immunology, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706 (United States); Brandt, C.R., E-mail: crbrandt@wisc.ed [Microbiology Doctoral Training Program, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706 (United States); Department of Ophthalmology and Visual Sciences, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706 (United States); Department of Medical Microbiology and Immunology, University of Wisconsin School of Medicine and Public Health, Madison, WI 53706 (United States)

2009-06-05

313

Multiserotype protection elicited by a combinatorial prime-boost vaccination strategy against bluetongue virus.  

PubMed

Bluetongue virus (BTV) belongs to the genus Orbivirus within the family Reoviridae. The development of vector-based vaccines expressing conserved protective antigens results in increased immune activation and could reduce the number of multiserotype vaccinations required, therefore providing a cost-effective product. Recent recombinant DNA technology has allowed the development of novel strategies to develop marker and safe vaccines against BTV. We have now engineered naked DNAs and recombinant modified vaccinia virus Ankara (rMVA) expressing VP2, VP7 and NS1 proteins from BTV-4. IFNAR((-/-)) mice inoculated with DNA/rMVA-VP2,-VP7-NS1 in an heterologous prime boost vaccination strategy generated significant levels of antibodies specific of VP2, VP7, and NS1, including those with neutralizing activity against BTV-4. In addition, vaccination stimulated specific CD8(+) T cell responses against these three BTV proteins. Importantly, the vaccine combination expressing NS1, VP2 and VP7 proteins of BTV-4, elicited sterile protection against a lethal dose of homologous BTV-4 infection. Remarkably, the vaccine induced cross-protection against lethal doses of heterologous BTV-8 and BTV-1 suggesting that the DNA/rMVA-VP2,-VP7,-NS1 marker vaccine is a promising multiserotype vaccine against BTV. PMID:22514660

Calvo-Pinilla, Eva; Navasa, Nicolás; Anguita, Juan; Ortego, Javier

2012-01-01

314

Multiserotype Protection Elicited by a Combinatorial Prime-Boost Vaccination Strategy against Bluetongue Virus  

PubMed Central

Bluetongue virus (BTV) belongs to the genus Orbivirus within the family Reoviridae. The development of vector-based vaccines expressing conserved protective antigens results in increased immune activation and could reduce the number of multiserotype vaccinations required, therefore providing a cost-effective product. Recent recombinant DNA technology has allowed the development of novel strategies to develop marker and safe vaccines against BTV. We have now engineered naked DNAs and recombinant modified vaccinia virus Ankara (rMVA) expressing VP2, VP7 and NS1 proteins from BTV-4. IFNAR(?/?) mice inoculated with DNA/rMVA-VP2,-VP7-NS1 in an heterologous prime boost vaccination strategy generated significant levels of antibodies specific of VP2, VP7, and NS1, including those with neutralizing activity against BTV-4. In addition, vaccination stimulated specific CD8+ T cell responses against these three BTV proteins. Importantly, the vaccine combination expressing NS1, VP2 and VP7 proteins of BTV-4, elicited sterile protection against a lethal dose of homologous BTV-4 infection. Remarkably, the vaccine induced cross-protection against lethal doses of heterologous BTV-8 and BTV-1 suggesting that the DNA/rMVA-VP2,-VP7,-NS1 marker vaccine is a promising multiserotype vaccine against BTV.

Calvo-Pinilla, Eva; Navasa, Nicolas; Anguita, Juan; Ortego, Javier

2012-01-01

315

Immunoglobulin-like transcript 2 (ILT2) is a biomarker of therapeutic response to oncolytic immunotherapy with vaccinia viruses  

PubMed Central

Background Oncolytic viruses represent a novel form of cancer immunotherapy. Vaccinia viruses encoding human T cell co-stimulatory molecules have demonstrated clinical activity in phase I clinical trials in patients with advanced melanoma. However, predictive biomarkers of therapeutic response have not yet been identified. Methods A customized microarray was performed to identify changes in peripheral blood mononuclear cell (PBMC) gene expression upon exposure to recombinant oncolytic vaccinia viruses. Up-regulated and down-regulated genes were identified and selected for further analysis using PBMC samples from normal donors and oncolytic virus-treated patients before and after viral injection. Quantitative PCR and flow cytometry of defined T cell subsets was performed to evaluate expression patterns and clinical correlations. Results The microarray identified 301 genes that were up-regulated and 960 genes that were down-regulated in T cells after exposure to oncolytic vaccinia virus. The B7.1 gene was highly up-regulated and the immunoglobulin-like transcript 2 (ILT2) gene was highly down-regulated by vaccinia-B7.1, which was consistent with the known inverse regulation of these two genes. We observed an inverse association between ILT2 expression in the tumor microenvironment and clinical response and further identified ILT2 as a marker of regulatory CD4+ and suppressor CD8+ T cell responses and whose down-regulation was predictive of therapeutic responses in patients treated with oncolytic virus immunotherapy. Conclusions ILT2 is a new putative biomarker of T cell and clinical response in patients treated with oncolytic vaccinia virus immunotherapy. Further confirmation of ILT2 as a biomarker requires prospective validation in a larger series of clinical trials.

2014-01-01

316

Novel vaccine strategies against emerging viruses  

PubMed Central

One of the main public health concerns of emerging viruses is their potential introduction into and sustained circulation among populations of immunologically naïve, susceptible hosts. The induction of protective immunity through vaccination can be a powerful tool to prevent this concern by conferring protection to the population at risk. Conventional approaches to develop vaccines against emerging pathogens have significant limitations: lack of experimental tools for several emerging viruses of concern, poor immunogenicity, safety issues, or lack of cross-protection against antigenic variants. The unpredictability of the emergence of future virus threats demands the capability to rapidly develop safe, effective vaccines. We describe some recent advances in new vaccine strategies that are being explored as alternatives to classical attenuated and inactivated vaccines, and provide examples of potential novel vaccines for emerging viruses. These approaches might be applied to the control of many other emerging pathogens.

Garcia-Sastre, Adolfo; Mena, Ignacio

2013-01-01

317

Vaccination with a fusion protein that introduces HIV-1 gag antigen into a multitrimer CD40L construct results in enhanced CD8+ T cell responses and protection from viral challenge by vaccinia-gag.  

PubMed

CD40 ligand (CD40L, CD154) is a membrane protein that is important for the activation of dendritic cells (DCs) and DC-induced CD8(+) T cell responses. To be active, CD40L must cluster CD40 receptors on responding cells. To produce a soluble form of CD40L that clusters CD40 receptors necessitates the use of a multitrimer construct. With this in mind, a tripartite fusion protein was made from surfactant protein D (SPD), HIV-1 Gag as a test antigen, and CD40L, where SPD serves as a scaffold for the multitrimer protein complex. This SPD-Gag-CD40L protein activated CD40-bearing cells and bone marrow-derived DCs in vitro. Compared to a plasmid for Gag antigen alone (pGag), DNA vaccination of mice with pSPD-Gag-CD40L induced an increased number of Gag-specific CD8(+) T cells with increased avidity for major histocompatibility complex class I-restricted Gag peptide and improved vaccine-induced protection from challenge by vaccinia-Gag virus. The importance of the multitrimeric nature of the complex was shown using a plasmid lacking the N terminus of SPD that produced a single trimer fusion protein. This plasmid, pTrimer-Gag-CD40L, was only weakly active on CD40-bearing cells and did not elicit strong CD8(+) T cell responses or improve protection from vaccinia-Gag challenge. An adenovirus 5 (Ad5) vaccine incorporating SPD-Gag-CD40L was much stronger than Ad5 expressing Gag alone (Ad5-Gag) and induced complete protection (i.e., sterilizing immunity) from vaccinia-Gag challenge. Overall, these results show the potential of a new vaccine design in which antigen is introduced into a construct that expresses a multitrimer soluble form of CD40L, leading to strongly protective CD8(+) T cell responses. PMID:24227853

Gupta, Sachin; Termini, James M; Raffa, Francesca N; Williams, Cindi-Ann; Kornbluth, Richard S; Stone, Geoffrey W

2014-02-01

318

Mutations in active-site residues of the uracil-DNA glycosylase encoded by vaccinia virus are incompatible with virus viability.  

PubMed Central

The D4R gene of vaccinia virus encodes a functional uracil-DNA glycosylase that is essential for viral viability (D. T. Stuart, C. Upton, M. A. Higman, E. G. Niles, and G. McFadden, J. Virol. 67:2503-2513, 1993), and a D4R mutant, ts4149, confers a conditional lethal defect in viral DNA replication (A. K. Millns, M. S. Carpenter, and A. M. DeLange, Virology 198:504-513, 1994). The mutant ts4149 protein was expressed in vitro and assayed for uracil-DNA glycosylase activity. Less than 6% of wild-type activity was observed at permissive temperatures, but the ts4149 protein was completely inactive at the nonpermissive temperature. Mutagenesis of the ts4149 gene back to wild type (Arg-179-->Gly) restored full activity. The ts4149 protein was considerably reduced in lysates of cells infected at the permissive temperature, and its activity was undetectable, even in the presence of the uracil glycosylase inhibitor protein, which inhibits the host uracil-DNA glycosylases but not that of vaccinia virus. Thus the ts4149 protein is thermolabile, correlating uracil removal with vaccinia virus DNA replication. Three active-site amino acids of the vaccinia virus uracil-DNA glycosylase were mutated (Asp-68-->Asn, Asn-120-->Val, and His-181-->Leu), producing proteins that were completely defective in uracil excision but still retained the ability to bind DNA. Each mutated D4R gene was transfected into vaccinia virus ts4149-infected cells in order to assess the recombination events that allowed virus survival at 40 degrees C. Genetic analysis and sequencing studies revealed that the only viruses to survive were those in which recombination eliminated the mutant locus. We conclude that the uracil cleavage activity of the D4R protein is essential for its function in vaccinia virus DNA replication, suggesting that the removal of uracil residues plays an obligatory role.

Ellison, K S; Peng, W; McFadden, G

1996-01-01

319

Efficacy of a Plasmodium vivax Malaria Vaccine Using ChAd63 and Modified Vaccinia Ankara Expressing Thrombospondin-Related Anonymous Protein as Assessed with Transgenic Plasmodium berghei Parasites  

PubMed Central

Plasmodium vivax is the world's most widely distributed malaria parasite and a potential cause of morbidity and mortality for approximately 2.85 billion people living mainly in Southeast Asia and Latin America. Despite this dramatic burden, very few vaccines have been assessed in humans. The clinically relevant vectors modified vaccinia virus Ankara (MVA) and the chimpanzee adenovirus ChAd63 are promising delivery systems for malaria vaccines due to their safety profiles and proven ability to induce protective immune responses against Plasmodium falciparum thrombospondin-related anonymous protein (TRAP) in clinical trials. Here, we describe the development of new recombinant ChAd63 and MVA vectors expressing P. vivax TRAP (PvTRAP) and show their ability to induce high antibody titers and T cell responses in mice. In addition, we report a novel way of assessing the efficacy of new candidate vaccines against P. vivax using a fully infectious transgenic Plasmodium berghei parasite expressing P. vivax TRAP to allow studies of vaccine efficacy and protective mechanisms in rodents. Using this model, we found that both CD8+ T cells and antibodies mediated protection against malaria using virus-vectored vaccines. Our data indicate that ChAd63 and MVA expressing PvTRAP are good preerythrocytic-stage vaccine candidates with potential for future clinical application.

Bauza, Karolis; Malinauskas, Tomas; Pfander, Claudia; Anar, Burcu; Jones, E. Yvonne; Billker, Oliver; Hill, Adrian V. S.

2014-01-01

320

Efficacy of a Plasmodium vivax malaria vaccine using ChAd63 and modified vaccinia Ankara expressing thrombospondin-related anonymous protein as assessed with transgenic Plasmodium berghei parasites.  

PubMed

Plasmodium vivax is the world's most widely distributed malaria parasite and a potential cause of morbidity and mortality for approximately 2.85 billion people living mainly in Southeast Asia and Latin America. Despite this dramatic burden, very few vaccines have been assessed in humans. The clinically relevant vectors modified vaccinia virus Ankara (MVA) and the chimpanzee adenovirus ChAd63 are promising delivery systems for malaria vaccines due to their safety profiles and proven ability to induce protective immune responses against Plasmodium falciparum thrombospondin-related anonymous protein (TRAP) in clinical trials. Here, we describe the development of new recombinant ChAd63 and MVA vectors expressing P. vivax TRAP (PvTRAP) and show their ability to induce high antibody titers and T cell responses in mice. In addition, we report a novel way of assessing the efficacy of new candidate vaccines against P. vivax using a fully infectious transgenic Plasmodium berghei parasite expressing P. vivax TRAP to allow studies of vaccine efficacy and protective mechanisms in rodents. Using this model, we found that both CD8+ T cells and antibodies mediated protection against malaria using virus-vectored vaccines. Our data indicate that ChAd63 and MVA expressing PvTRAP are good preerythrocytic-stage vaccine candidates with potential for future clinical application. PMID:24379295

Bauza, Karolis; Malinauskas, Tomas; Pfander, Claudia; Anar, Burcu; Jones, E Yvonne; Billker, Oliver; Hill, Adrian V S; Reyes-Sandoval, Arturo

2014-03-01

321

Effect of aldosterone on the amplification of oncolytic vaccinia virus in human cancer lines  

PubMed Central

Background/Aims JX-594 is an oncolytic virus derived from the Wyeth vaccinia strain that causes replication-dependent cytolysis and antitumor immunity. Starting with a cross-examination of clinical-trial samples from advanced hepatocellular carcinoma patients having high levels of aldosterone and virus amplification in JX-594 treatment, we investigated the association between virus amplification and aldosterone in human cancer cell lines. Methods Cell proliferation was determined by a cell-counting-kit-based colorimetric assay, and vaccinia virus quantitation was performed by quantitative polymerase chain reaction (qPCR) and a viral plaque assay. Also, the intracellular pH was measured using a pH-sensitive dye. Results Simultaneous treatment with JX-594 and aldosterone significantly increased viral replication in A2780, PC-3, and HepG2 cell lines, but not in U2OS cell lines. Furthermore, the aldosterone treatment time altered the JX-594 replication according to the cell line. The JX-594 replication peaked after 48 and 24 hours of treatment in PC-3 and HepG2 cells, respectively. qPCR showed that JX-594 entry across the plasma membrane was increased, however, the changes are not significant by the treatment. This was inhibited by treatment with spironolactone (an aldosterone-receptor inhibitor). JX-594 entry was significantly decreased by treatment with EIPA [5-(N-ethyl-N-isopropyl)amiloride; a Na+/H+-exchange inhibitor], but aldosterone significantly restored JX-594 entry even in the presence of EIPA. Intracellular alkalization was observed after aldosterone treatment but was acidified by EIPA treatment. Conclusions Aldosterone stimulates JX-594 amplification via increased virus entry by affecting the H+ gradient.

Lee, Hyun Ju; Rho, Jasung; Gui, Shao Ran; Kim, Mi Kyung; Lee, Yu Kyoung; Lee, Yeon Sook; Kim, Jeong Eun; Cho, Euna; Hwang, Tae-Ho

2011-01-01

322

Molecular Characterization of the Host Defense Activity of the Barrier to Autointegration Factor against Vaccinia Virus ?  

PubMed Central

The barrier to autointegration factor (BAF) is an essential cellular protein with functions in mitotic nuclear reassembly, retroviral preintegration complex stability, and transcriptional regulation. Molecular properties of BAF include the ability to bind double-stranded DNA in a sequence-independent manner, homodimerize, and bind proteins containing a LEM domain. These capabilities allow BAF to compact DNA and assemble higher-order nucleoprotein complexes, the nature of which is poorly understood. Recently, it was revealed that BAF also acts as a potent host defense against poxviral DNA replication in the cytoplasm. Here, we extend these observations by examining the molecular mechanism through which BAF acts as a host defense against vaccinia virus replication and cytoplasmic DNA in general. Interestingly, BAF rapidly relocalizes to transfected DNA from a variety of sources, demonstrating that BAF's activity as a host defense factor is not limited to poxviral infection. BAF's relocalization to cytoplasmic foreign DNA is highly dependent upon its DNA binding and dimerization properties but does not appear to require its LEM domain binding activity. However, the LEM domain protein emerin is recruited to cytoplasmic DNA in a BAF-dependent manner during both transfection and vaccinia virus infection. Finally, we demonstrate that the DNA binding and dimerization capabilities of BAF are essential for its function as an antipoxviral effector, while the presence of emerin is not required. Together, these data provide further mechanistic insight into which of BAF's molecular properties are employed by cells to impair the replication of poxviruses or respond to foreign DNA in general.

Ibrahim, Nouhou; Wicklund, April; Wiebe, Matthew S.

2011-01-01

323

Phenotypic characterization of mutants in vaccinia virus gene G2R, a putative transcription elongation factor.  

PubMed Central

The phenotypic defects of two mutants of vaccinia virus, the lesions of which map to gene G2R, were characterized in vivo, and the results suggest a role for the G2R protein in viral transcription elongation. Both a temperature-sensitive mutant, Cts56, and an isatin-beta-thiosemicarbazone-dependent deletion mutant, G2A, in gene G2R have a characteristic and unique defect in late viral gene expression. The G2R mutants synthesize early viral RNA, early viral proteins, and viral DNA normally under nonpermissive conditions. In G2R mutants, late viral protein synthesis begins at the normal time, low-molecular-weight viral proteins are synthesized in normal quantities, but synthesis of high-molecular-weight viral proteins is reduced in amount. Intermediate and late promoter utilization is normal in G2R mutants, but intermediate and late RNAs are reduced in size. The reduction in length of the intermediate and late mRNAs represents a truncation of mRNA 3' ends. Thus, intermediate and late RNAs are too short to encode large proteins but long enough to encode small proteins, therefore accounting for the protein synthesis phenotype. These results suggest that the G2R protein acts to regulate the elongation potential of the viral RNA polymerase late during a vaccinia virus infection.

Black, E P; Condit, R C

1996-01-01

324

Detection of Vaccinia virus in blood and faeces of experimentally infected cows.  

PubMed

Bovine vaccinia (BV), a zoonosis caused by Vaccinia virus (VACV), affects dairy cattle and milkers, causing economic, veterinary and human health impacts. Despite such impacts, there are no experimental studies about the pathogenesis of BV in cows to assess whether there is a systemic spread of the virus and whether there are different ways of VACV shedding. Trying to answer some of these questions, a study was proposed using experimental inoculation of VACV in cows. All experimentally infected cows developed lesions compatible with VACV infection in cattle. Two of the six animals presented VACV DNA in blood and faecal samples, starting at the 2nd and the 3rd day post-infection (d.p.i.), respectively, and lasting until the 36th d.p.i., in an intermittent way. This study provides new evidence that VACV can be detected in blood and faeces of infected cows, suggesting that BV could be a systemic disease, and also bringing new information about the epidemiology and pathogenesis of BV. PMID:22909142

Guedes, M I M C; Rehfeld, I S; de Oliveira, T M L; Assis, F L; Matos, A C D; Abrahão, J S; Kroon, E G; Lobato, Z I P

2013-12-01

325

Anisotropic cell-to-cell spread of vaccinia virus on microgrooved substrate.  

PubMed

Cell-to-cell spread of virus is a comprehensive process with involvement of cellular actin cytoskeleton and substrate topography can affect the arrangement of cytoskeleton via contact guidance, yet interaction among virus, cytoskeleton and substrate topography is still unknown. To investigate the virus-cell-substrate interaction, we designed a microgrooved poly(dimethyl siloxane) (PDMS) substrate for the study of vaccinia virus (VACV) cell-to-cell spread and the remodeling of cellular actin cytoskeleton in viral infection process. Interestingly, VACV-induced plaques on microgrooved substrate were elliptical instead of circular plaques on smooth substrate, suggesting an anisotropic cell-to-cell spread of VACV. The spread rate was faster in the direction parallel to microgroove and slower in the direction perpendicular to microgroove than that on smooth substrate. Host cells cultured on microgrooved surface showed significant alignment and elongation in the axis parallel to microgrooves. Cell elongation is one reason for anisotropic spread but could not totally explain the phenomenon. Actin fibers in infected cells maintained alignment and VACV-induced actin tails tipped with virions were oriented along the direction parallel to microgroove. These results suggested that substrate topography can affect infected cells and these effects will guide the spread of virus via orientation of actin cytoskeleton. This work opens a window for understanding virus response to substrate topography, and has potential implications on revealing virus-cell-substrate interactions in vivo. PMID:24685266

Xu, Na; Wang, Ji; Zhang, Zhen-Feng; Pang, Dai-Wen; Wang, Han-Zhong; Zhang, Zhi-Ling

2014-06-01

326

A36-dependent Actin Filament Nucleation Promotes Release of Vaccinia Virus  

PubMed Central

Cell-to-cell transmission of vaccinia virus can be mediated by enveloped virions that remain attached to the outer surface of the cell or those released into the medium. During egress, the outer membrane of the double-enveloped virus fuses with the plasma membrane leaving extracellular virus attached to the cell surface via viral envelope proteins. Here we report that F-actin nucleation by the viral protein A36 promotes the disengagement of virus attachment and release of enveloped virus. Cells infected with the A36YdF virus, which has mutations at two critical tyrosine residues abrogating localised actin nucleation, displayed a 10-fold reduction in virus release. We examined A36YdF infected cells by transmission electron microscopy and observed that during release, virus appeared trapped in small invaginations at the plasma membrane. To further characterise the mechanism by which actin nucleation drives the dissociation of enveloped virus from the cell surface, we examined recombinant viruses by super-resolution microscopy. Fluorescently-tagged A36 was visualised at sub-viral resolution to image cell-virus attachment in mutant and parental backgrounds. We confirmed that A36YdF extracellular virus remained closely associated to the plasma membrane in small membrane pits. Virus-induced actin nucleation reduced the extent of association, thereby promoting the untethering of virus from the cell surface. Virus release can be enhanced via a point mutation in the luminal region of B5 (P189S), another virus envelope protein. We found that the B5P189S mutation led to reduced contact between extracellular virus and the host membrane during release, even in the absence of virus-induced actin nucleation. Our results posit that during release virus is tightly tethered to the host cell through interactions mediated by viral envelope proteins. Untethering of virus into the surrounding extracellular space requires these interactions be relieved, either through the force of actin nucleation or by mutations in luminal proteins that weaken these interactions.

Horsington, Jacquelyn; Lynn, Helena; Turnbull, Lynne; Cheng, Delfine; Braet, Filip; Diefenbach, Russell J.; Whitchurch, Cynthia B.; Karupiah, Guna; Newsome, Timothy P.

2013-01-01

327

Expression of the E3L Gene of Vaccinia Virus in Transgenic Mice Decreases Host Resistance to Vaccinia Virus and Leishmania major Infections?  

PubMed Central

The E3L gene of vaccinia virus (VACV) encodes the E3 protein that in cultured cells inhibits the activation of interferon (IFN)-induced proteins, double-stranded RNA-dependent protein kinase (PKR), 2?-5?-oligoadenylate synthetase/RNase L (2-5A system) and adenosine deaminase (ADAR-1), thus helping the virus to evade host responses. Here, we have characterized the in vivo E3 functions in a murine inducible cell culture system (E3L-TetOFF) and in transgenic mice (TgE3L). Inducible E3 expression in cultured cells conferred on cells resistance to the antiviral action of IFN against different viruses, while expression of the E3L gene in TgE3L mice triggered enhanced sensitivity of the animals to pathogens. Virus infection monitored in TgE3L mice by different inoculation routes (intraperitoneal and tail scarification) showed that transgenic mice became more susceptible to VACV infection than control mice. TgE3L mice were also more susceptible to Leishmania major infection, leading to an increase in parasitemia compared to control mice. The enhanced sensitivity of TgE3L mice to VACV and L. major infections occurred together with alterations in the host immune system, as revealed by decreased T-cell responses to viral antigens in the spleen and lymph nodes and by differences in the levels of specific innate cell populations. These results demonstrate that expression of the E3L gene in transgenic mice partly reverses the resistance of the host to viral and parasitic infections and that these effects are associated with immune alterations.

Domingo-Gil, Elena; Perez-Jimenez, Eva; Ventoso, Ivan; Najera, Jose L.; Esteban, Mariano

2008-01-01

328

Vaccinia virus F1L protein promotes virulence by inhibiting inflammasome activation  

PubMed Central

Host innate immune responses to DNA viruses involve members of the nucleotide-binding domain, leucine-rich repeat and pyrin domain containing protein (NLRP) family, which form “inflammasomes” that activate caspase-1, resulting in proteolytic activation of cytokines interleukin (IL)-1? and IL-18. We hypothesized that DNA viruses would target inflammasomes to overcome host defense. A Vaccinia virus (VACV) B-cell CLL/lymphoma 2 (Bcl-2) homolog, F1L, was demonstrated to bind and inhibit the NLR family member NLRP1 in vitro. Moreover, infection of macrophages in culture with virus lacking F1L (?F1L) caused increased caspase-1 activation and IL-1? secretion compared with wild-type virus. Virulence of ?F1L virus was attenuated in vivo, causing altered febrile responses, increased proteolytic processing of caspase-1, and more rapid inflammation in lungs of infected mice without affecting cell death or virus replication. Furthermore, we found that a hexapeptide from F1L is necessary and sufficient for inhibiting the NLRP1 inflammasome in vitro, thus identifying a peptidyl motif required for binding and inhibiting NLRP1. The functional importance of this NLRP1-binding motif was further confirmed by studies of recombinant ?F1L viruses reconstituted either with the wild-type F1L or a F1L mutant that fails to bind NLRP1. Cellular infection with wild-type F1L reconstituted virus-suppressed IL-1? production, whereas mutant F1L did not. In contrast, both wild-type and mutant versions of F1L equally suppressed apoptosis. In vivo, the NLR nonbinding F1L mutant virus exhibited an attenuated phenotype similar to ?F1L virus, thus confirming the importance of F1L interactions with NLRP1 for viral pathogenicity in mice. Altogether, these findings reveal a unique viral mechanism for evading host innate immune responses.

Gerlic, Motti; Faustin, Benjamin; Postigo, Antonio; Yu, Eric Chi-Wang; Proell, Martina; Gombosuren, Naran; Krajewska, Maryla; Flynn, Rachel; Croft, Michael; Way, Michael; Satterthwait, Arnold; Liddington, Robert C.; Salek-Ardakani, Shahram; Matsuzawa, Shu-ichi; Reed, John C.

2013-01-01

329

Expression of the highly conserved vaccinia virus E6 protein is required for virion morphogenesis  

SciTech Connect

The vaccinia virus E6R gene (VACVWR062) is conserved in all members of the poxvirus family and encodes a protein associated with the mature virion. We confirmed this association and provided evidence for an internal location. An inducible mutant that conditionally expresses E6 was constructed. In the absence of inducer, plaque formation and virus production were severely inhibited in several cell lines, whereas some replication occurred in others. This difference could be due to variation in the stringency of repression, since we could not isolate a stable deletion mutant even in the more 'permissive' cells. Under non-permissive conditions, viral late proteins were synthesized but processing of core proteins was inefficient, indicative of an assembly block. Transmission electron microscopy of sections of cells infected with the mutant in the absence of inducer revealed morphogenetic defects with crescents and empty immature virions adjacent to dense inclusions of viroplasm. Mature virions were infrequent and cores appeared to have lucent centers.

Resch, Wolfgang; Weisberg, Andrea S. [Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892-3210 (United States); Moss, Bernard, E-mail: bmoss@nih.go [Laboratory of Viral Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892-3210 (United States)

2009-04-10

330

RNA Virus Reverse Genetics and Vaccine Design  

PubMed Central

RNA viruses are capable of rapid spread and severe or potentially lethal disease in both animals and humans. The development of reverse genetics systems for manipulation and study of RNA virus genomes has provided platforms for designing and optimizing viral mutants for vaccine development. Here, we review the impact of RNA virus reverse genetics systems on past and current efforts to design effective and safe viral therapeutics and vaccines.

Stobart, Christopher C.; Moore, Martin L.

2014-01-01

331

RNA virus reverse genetics and vaccine design.  

PubMed

RNA viruses are capable of rapid spread and severe or potentially lethal disease in both animals and humans. The development of reverse genetics systems for manipulation and study of RNA virus genomes has provided platforms for designing and optimizing viral mutants for vaccine development. Here, we review the impact of RNA virus reverse genetics systems on past and current efforts to design effective and safe viral therapeutics and vaccines. PMID:24967693

Stobart, Christopher C; Moore, Martin L

2014-01-01

332

Urticaria, exanthems, and other benign dermatologic reactions to smallpox vaccination in adults.  

PubMed

A phase 1 smallpox vaccine trial involving 350 adult volunteers was conducted. Of these subjects, 250 were naive to vaccinia virus vaccine (i.e., "vaccinia naive"). Volunteers received a new cell-cultured smallpox vaccine or a live vaccinia virus vaccine. Nine self-limiting rashes (3.6%) were observed in the vaccinia-naive group. None of the vaccinia-experienced patients had a rash. Rashes appeared 6-19 days after vaccination and had 5 different clinical presentations. Five volunteers had urticarial rashes that resolved within 4-15 days, 1 had an exanthem that lasted 20 days, and 1 each presented with folliculitis, contact dermatitis, and erythematous papules found only on the hands and fingers. Volunteers reported pruritus, tingling, and occasional headaches. Relief was obtained with antihistamine and acetaminophen therapy. No volunteer experienced fever or significant discomfort. PMID:15034827

Greenberg, Richard N; Schosser, Robert H; Plummer, Elizabeth A; Roberts, Sara E; Caldwell, Malissia A; Hargis, Dana L; Rudy, David W; Evans, Martin E; Hopkins, Robert J

2004-04-01

333

Evaluating vaccinia virus cytokine co-expression in TLR GKO mice.  

PubMed

Using Toll-like receptor (TLR) and MyD88 gene knock-out (GKO) mice the effect of TLRs and MyD88 on virus replication, interferon (IFN)-? production, natural killer (NK) cell and CD8T cell responses were assessed following ectromelia virus (ECTV) and recombinant vaccinia virus (rVV) infection. The capacity for rVVs encoding cytokines to restore immune function in MyD88(-/-) mice was clearly demonstrated. Results showed that TLR2(-/-), TLR4(-/-)and TLR7(-/-) mice survived ECTV infection whereas MyD88(-/-) and TLR9(-/-)mice, in contrast, were highly susceptible. Next, following infection with rVV, MyD88(-/-) mice elicited reduced serum IFN-?, NK cell and CD8T cell responses compared with wild-type mice, whereas TLR9(-/-) mice showed elevated CD8T cell responses. When MyD88(-/-)mice were infected with rVV co-expressing IFN-? these mice were able to restore IFN-? levels and CD8T cell responses but not NK cell activation. Interestingly, even though rVV co-expressing interleukin (IL)-2 enhanced NK cell activation in MyD88(-/-) mice, this was not associated with an antiviral effect, as observed in normal mice. Surprisingly, co-infection with rVV IL-2/rVV IL-12, but not rVV IL-2/rVV IFN-?, restored the attenuated phenotype of rVV IL-2 in MyD88(-/-) mice indicating that the IL-2/IL-12 combination promotes antiviral responses. Our results clearly show that the CD8T cell defect observed in MyD88(-/-) mice to vaccinia virus infection can be restored by rVV-encoding IFN-? demonstrating the critical role of this cytokine in T cell mediated immunity and illustrates that the model can provide an effective platform for the elucidation of cytokine immunobiology. PMID:21173782

Sutherland, Duncan B; Ranasinghe, Charani; Regner, Matthias; Phipps, Simon; Matthaei, Klaus I; Day, Stephanie L; Ramshaw, Ian A

2011-08-01

334

Tumor vascularization is critical for oncolytic vaccinia virus treatment of peritoneal carcinomatosis.  

PubMed

Peritoneal carcinomatosis (PC) represents a significant clinical challenge for which there are few treatment options. Oncolytic viruses are ideal candidates for PC treatment because of their high tumor specificity, excellent safety profile and suitability for peritoneal delivery. Here, we described the use of vvDD-SR-RFP, a recombinant vaccinia virus, in xenograft and syngeneic models of colorectal PC. Colorectal cancer cell lines were highly susceptible to vvDD-SR-RFP replication and cytotoxicity. Intraperitoneal delivery of vvDD-SR-RFP on Day 12 to mice with colorectal carcinomatosis significantly improved survival whereas survival was not improved following virus treatment on Day 8, when tumors were smaller. Immunohistochemistry revealed early tumors had a poorly distributed network of blood vessels and lower proliferation index compared to later tumors. Virus infection was also restricted to tumor rims following Day 8 treatment, whereas it was disseminated in tumors treated on Day 12. Additionally, direct infection of tumor endothelium was observed and virus infection correlated with a loss of endothelial staining and induction of cell death. Our results demonstrate that tumor vasculature has a critical role in virus delivery and tumor response. This will have significant implications in the clinical setting, both in understanding timing of therapies and in designing combination treatment strategies. PMID:23893655

Ottolino-Perry, Kathryn; Tang, Nan; Head, Renee; Ng, Calvin; Arulanandam, Rozanne; Angarita, Fernando A; Acuna, Sergio A; Chen, Yonghong; Bell, John; Dacosta, Ralph S; McCart, J Andrea

2014-02-01

335

Comparative analysis of poxvirus orthologues of the vaccinia virus E3 protein: modulation of protein kinase R activity, cytokine responses, and virus pathogenicity.  

PubMed

Poxviruses are important human and animal pathogens that have evolved elaborate strategies for antagonizing host innate and adaptive immunity. The E3 protein of vaccinia virus, the prototypic member of the orthopoxviruses, functions as an inhibitor of innate immune signaling and is essential for vaccinia virus replication in vivo and in many human cell culture systems. However, the function of orthologues of E3 expressed by poxviruses of other genera with different host specificity remains largely unknown. In the present study, we characterized the E3 orthologues from sheeppox virus, yaba monkey tumor virus, swinepox virus, and myxoma virus for their ability to modulate protein kinase R (PKR) function, cytokine responses and virus pathogenicity. We found that the E3 orthologues of myxoma virus and swinepox virus could suppress PKR activation and interferon (IFN)-induced antiviral activities and restore the host range function of E3 in HeLa cells. In contrast, the E3 orthologues from sheeppox virus and yaba monkey tumor virus were unable to inhibit PKR activation. While the sheeppox orthologue was unable to restore the host range function of E3, the yaba monkey tumor virus orthologue partially restored E3-deficient vaccinia virus replication in HeLa cells, correlated with its ability to suppress IFN-induced antiviral activities. Moreover, poxvirus E3 orthologues show varying ability to inhibit the induction of antiviral and proinflammatory cytokines. Despite these in vitro results, none of the E3 orthologues tested was capable of restoring pathogenicity to E3-deficient vaccinia virus in vivo. PMID:21917954

Myskiw, Chad; Arsenio, Janilyn; Hammett, Craig; van Bruggen, Rebekah; Deschambault, Yvon; Beausoleil, Nicole; Babiuk, Shawn; Cao, Jingxin

2011-12-01

336

Comparative Analysis of Poxvirus Orthologues of the Vaccinia Virus E3 Protein: Modulation of Protein Kinase R Activity, Cytokine Responses, and Virus Pathogenicity?  

PubMed Central

Poxviruses are important human and animal pathogens that have evolved elaborate strategies for antagonizing host innate and adaptive immunity. The E3 protein of vaccinia virus, the prototypic member of the orthopoxviruses, functions as an inhibitor of innate immune signaling and is essential for vaccinia virus replication in vivo and in many human cell culture systems. However, the function of orthologues of E3 expressed by poxviruses of other genera with different host specificity remains largely unknown. In the present study, we characterized the E3 orthologues from sheeppox virus, yaba monkey tumor virus, swinepox virus, and myxoma virus for their ability to modulate protein kinase R (PKR) function, cytokine responses and virus pathogenicity. We found that the E3 orthologues of myxoma virus and swinepox virus could suppress PKR activation and interferon (IFN)-induced antiviral activities and restore the host range function of E3 in HeLa cells. In contrast, the E3 orthologues from sheeppox virus and yaba monkey tumor virus were unable to inhibit PKR activation. While the sheeppox orthologue was unable to restore the host range function of E3, the yaba monkey tumor virus orthologue partially restored E3-deficient vaccinia virus replication in HeLa cells, correlated with its ability to suppress IFN-induced antiviral activities. Moreover, poxvirus E3 orthologues show varying ability to inhibit the induction of antiviral and proinflammatory cytokines. Despite these in vitro results, none of the E3 orthologues tested was capable of restoring pathogenicity to E3-deficient vaccinia virus in vivo.

Myskiw, Chad; Arsenio, Janilyn; Hammett, Craig; van Bruggen, Rebekah; Deschambault, Yvon; Beausoleil, Nicole; Babiuk, Shawn; Cao, Jingxin

2011-01-01

337

Identification by mass spectroscopy of three major early proteins associated with virosomes in vaccinia virus-infected cells  

Microsoft Academic Search

Virosomes are cytoplasmic sites of replication of vaccinia virus DNA and were prepared from virus-infected HeLa cells. The early virosomal proteins were 35S-labelled and SDS polyacrylamide gel electrophoresis revealed the presence of three major early 35S-labelled proteins of 34, 24 and 45 kDa. The masses of molecules present in the 34 and 24 kDa proteins were measured by the convenient

Adriana Murcia-Nicolas; Gérard Bolbach; Jean-Claude Blais; Georges Beaud

1999-01-01

338

Vaccinia Virus 15-Kilodalton (A14L) Protein Is Essential for Assembly and Attachment of Viral Crescents to Virosomes  

Microsoft Academic Search

Early stages in vaccinia virus (VV) assembly involve the recruitment of cellular membranes from the endoplasmic reticulum-Golgi intermediate compartment (ERGIC) to virus factories (or virosomes). The key viral factors involved in this process are not yet known. We have previously identified and characterized two viral proteins, of 21 kDa (A17L gene) and 15 kDa (A14L gene), that associate with tubulovesicular

JUAN RAMON RODRIGUEZ; CRISTINA RISCO; MARIANO ESTEBAN; DOLORES RODRIGUEZ; Centro Nacional

1998-01-01

339

Identification ofrpo3O, a Vaccinia VirusRNA Polymerase Gene withStructural Similarity toa Eucaryotic Transcription Elongation Factor  

Microsoft Academic Search

Eucaryotic transcription factors that stimulate RNA polymerase II byincreasing theefficiency ofelongation ofspecifically orrandomly initiated RNA chains havebeenisolated andcharacterized. We haveidentified a 30-kilodalton (kDa)vaccinia virus-encoded protein withapparent homology toSIT, a 34-kDamammalian transcriptional elongation factor. Inaddition toaminoacidsequencesimilarities, bothproteins contain C-terminal putative zincfinger domains. Identification ofthegene,rpo3O, encoding thevaccinia virus protein was achieved byusing antibody tothepurified viral RNA polymerase forimmunoprecipitation oftheinvitro translation products ofinvivo-synthesized

BYUNG-YOON AHN; PAUL D. GERSHON; BERNARD MOSS

1990-01-01

340

JNK-deficiency enhanced oncolytic vaccinia virus replication and blocked activation of double-stranded RNA-dependent protein kinase  

Microsoft Academic Search

Vaccinia virus has recently been used as an expression vector for gene delivery and an oncolytic agent for cancer therapy. Although it has been established that interferon-induced double-stranded RNA (dsRNA)-activated protein kinase (PKR) and RNase L interfere with viral replication, little else is known about the other host factors that might affect viral replication and virus-mediated host cell killing. In

W Hu; W Hofstetter; W Guo; H Li; A Pataer; H H Peng; Z S Guo; D L Bartlett; A Lin; S G Swisher; B Fang

2008-01-01

341

Control of vaccinia virus skin lesions by long-term-maintained IFN-?+TNF-?+ effector/memory CD4+ lymphocytes in humans  

PubMed Central

Vaccinia virus (VV) vaccination is used to immunize against smallpox and historically was considered to have been successful if a skin lesion formed at the vaccination site. While antibody responses have been widely proposed as a correlate of efficacy and protection in humans, the role of cellular and humoral immunity in VV-associated skin lesion formation was unknown. We therefore investigated whether long-term residual humoral and cellular immune memory to VV, persisting 30 years after vaccination, could control VV-induced skin lesion in revaccinated individuals. Here, we have shown that residual VV-specific IFN-?+TNF-?+ or IFN-?+IL-2+ CD4+ lymphocytes but not CD8+ effector/memory lymphocytes expressing a skin-homing marker are inversely associated with the size of the skin lesion formed in response to revaccination. Indeed, high numbers of residual effector T cells were associated with lower VV skin lesion size after revaccination. In contrast, long-term residual VV-specific neutralizing antibody (NAbs) titers did not affect skin lesion formation. However, the size of the skin lesion strongly correlated with high levels of NAbs boosted after revaccination. These findings demonstrate a potential role for VV-specific CD4+ responses at the site of VV-associated skin lesion, thereby providing new insight into immune responses at these sites and potentially contributing to the development of new approaches to measure the efficacy of VV vaccination.

Puissant-Lubrano, Benedicte; Bossi, Philippe; Gay, Frederick; Crance, Jean-Marc; Bonduelle, Olivia; Garin, Daniel; Bricaire, Francois; Autran, Brigitte; Combadiere, Behazine

2010-01-01

342

Comparison of host cell gene expression in cowpox, monkeypox or vaccinia virus-infected cells reveals virus-specific regulation of immune response genes  

PubMed Central

Background Animal-borne orthopoxviruses, like monkeypox, vaccinia and the closely related cowpox virus, are all capable of causing zoonotic infections in humans, representing a potential threat to human health. The disease caused by each virus differs in terms of symptoms and severity, but little is yet know about the reasons for these varying phenotypes. They may be explained by the unique repertoire of immune and host cell modulating factors encoded by each virus. In this study, we analysed the specific modulation of the host cell’s gene expression profile by cowpox, monkeypox and vaccinia virus infection. We aimed to identify mechanisms that are either common to orthopoxvirus infection or specific to certain orthopoxvirus species, allowing a more detailed description of differences in virus-host cell interactions between individual orthopoxviruses. To this end, we analysed changes in host cell gene expression of HeLa cells in response to infection with cowpox, monkeypox and vaccinia virus, using whole-genome gene expression microarrays, and compared these to each other and to non-infected cells. Results Despite a dominating non-responsiveness of cellular transcription towards orthopoxvirus infection, we could identify several clusters of infection-modulated genes. These clusters are either commonly regulated by orthopoxvirus infection or are uniquely regulated by infection with a specific orthopoxvirus, with major differences being observed in immune response genes. Most noticeable was an induction of genes involved in leukocyte migration and activation in cowpox and monkeypox virus-infected cells, which was not observed following vaccinia virus infection. Conclusion Despite their close genetic relationship, the expression profiles induced by infection with different orthopoxviruses vary significantly. It may be speculated that these differences at the cellular level contribute to the individual characteristics of cowpox, monkeypox and vaccinia virus infections in certain host species.

2013-01-01

343

Therapeutic Potential of Vaccinia Hyper Immune Sera in Mouse Models of Lethal Orthopoxviruses Infection  

Microsoft Academic Search

\\u000a Vaccinia Immune Globulin (VIG) is currently used to treat severe complications of smallpox vaccines. In this study we compare\\u000a the therapeutic potential of vaccinia virus rabbit hyper immune sera (RHIS) with that of human VIG. The clearance rate of\\u000a RHIS from mouse circulation is only slightly slower than that of VIG (t\\u000a 1\\/2=10 and 7.5 days respectively). Like VIG, passively

Sharon Melamed; Nir Paran; Tomer Israely; Noam Erez; Shaul Reuveny; Arie Ordentlich; Shlomo Lustig

344

Vaccinia Virus H3L Envelope Protein Is a Major Target of Neutralizing Antibodies in Humans and Elicits Protection against Lethal Challenge in Mice  

Microsoft Academic Search

The smallpox vaccine is the prototypic vaccine, yet the viral targets critical for vaccine-mediated protection remain unclear in humans. We have produced protein microarrays of a near-complete vaccinia proteome and used them to determine the major antigen specificities of the human humoral immune response to the smallpox vaccine (Dryvax). H3L, an intracellular mature virion envelope protein, was consistently recognized by

D. Huw Davies; Megan M. McCausland; Conrad Valdez; Devan Huynh; Jenny E. Hernandez; Yunxiang Mu; Siddiqua Hirst; Luis Villarreal; Philip L. Felgner; Shane Crotty

2005-01-01

345

The eukaryotic translation initiation factor 4E is not modified during the course of vaccinia virus replication.  

PubMed

The ability of vaccinia virus to inhibit processes of cap-dependent translational initiation by inactivating the eukaryotic translation initiation factor 4E (eIF-4E) has been examined. Analyses of the quantities of eIF-4E present in either uninfected mouse L929 cells or vaccinia virus-infected cells showed that during the first 12 hr of virus replication, when there is a marked decrease in host gene expression in infected cells, there is no change in the total amount of eIF-4E present. Analyses of eIF-4E that was metabolically labeled with [32P] and then purified by affinity chromatography using m7GTP-Sepharose 4B, indicated that neither the incorporation of radiolabel into eIF-4E nor the amounts of eIF-4E capable of binding to cap structures changed significantly during virus replication. Immunodetection of phosphorylated and unphosphorylated eIF-4E in cell lysates fractionated by two-dimensional gel electrophoresis showed that the steady-state levels of phosphorylated and unphosphorylated forms of eIF-4E were similar in uninfected and virus-infected cells. These results suggest that vaccinia virus does not gain preferential translation of viral mRNAs over other mRNAs in the cell by reducing either eIF-4E phosphorylation or its ability to bind to the cap structure. PMID:1585661

Gierman, T M; Frederickson, R M; Sonenberg, N; Pickup, D J

1992-06-01

346

Smallpox DNA vaccine delivered by novel skin electroporation device protects mice against intranasal poxvirus challenge  

Microsoft Academic Search

Previously, we demonstrated that an experimental smallpox DNA vaccine comprised of four vaccinia virus genes (4pox) administered by gene gun elicited protective immunity in mice challenged with vaccinia virus, and in nonhuman primates challenged with monkeypox virus (Hooper JW, et al. Smallpox DNA vaccine protects nonhuman primates against lethal monkeypox. J Virol 2004;78:4433–43). Here, we report that this 4pox DNA

Jay W. Hooper; Joseph W. Golden; Anthony M. Ferro; Alan D. King

2007-01-01

347

Clinical, hematological and biochemical parameters of dairy cows experimentally infected with Vaccinia virus.  

PubMed

Vaccinia virus (VACV) is the etiological agent of bovine vaccinia (BV), an important zoonosis that affects dairy cattle. There are many aspects of the disease that remain unknown, and aiming to answer some of these questions, the clinical, hematological, and biochemical parameters of VACV experimentally infected cows were evaluated. In the first part of the study, lactating cows were infected with VACV-GP2 strain. In the second part, animals previously infected with VACV-GP2 were divided into two treatment groups: Group 1, immunosuppressed cows; and Group 2, re-infected cows. In this study, BV could be experimentally reproduced, with similar lesions as observed in natural infections. Moreover, a short incubation period and local lymphadenopathy were also observed. VACV could be detected by PCR and isolated from scabs taken from teat lesions of all inoculated and re-inoculated animals. Lymphocytosis and neutrophilia were observed in all animals from the first part of the experiment, and lymphopenia and relative neutrophilia were observed in the immunosuppressed animals. Detection of viral DNA in oral mucosa lesions suggests that viral reactivation might occur in immunosuppressed animals. Moreover, clinical disease with teat lesions may occur in previously VACV-infected cows under the experimental conditions of the present study. PMID:23747141

Rehfeld, Izabelle S; Guedes, Maria Isabel M C; Matos, Ana Carolina D; de Oliveira, Tércia M L; Rivetti, Anselmo V; Moura, Ana Carolina J; Paes, Paulo Ricardo O; do Lago, Luiz Alberto; Kroon, Erna G; Lobato, Zélia Inês P

2013-10-01

348

Vaccinia Virus Virulence Factor N1L is a Novel Promising Target for Antiviral Therapeutic Intervention  

PubMed Central

The 14 kDa homodimeric N1L protein is a potent vaccinia and variola (smallpox) virulence factor. It is not essential for viral replication, but it causes a strong attenuation of viral production in culture when deleted. The N1L protein is predicted to contain the BH3-like binding domain characteristic of Bcl-2 family proteins, and it is able to bind the BH3 peptides. Its overexpression has been reported to prevent infected cells from committing apoptosis. Therefore, interfering with the N1L apoptotic blockade may be a legitimate therapeutic strategy affecting the viral growth. By using in silico ligand docking and an array of in vitro assays, we have identified sub-micromolar (600 nM) N1L antagonists, belonging to the family of polyphenols. Their affinity is comparable to that of the BH3 peptides (70 nM ÷ 1000 nM). We have also identified the natural polyphenol resveratrol as a moderate N1L inhibitor. Finally, we show that our ligands efficiently inhibit growth of vaccinia virus.

Cheltsov, Anton V.; Aoyagi, Mika; Aleshin, Alexander; Chi-Wang, Yu Eric; Gilliland, Taylor; Zhai, Dayong; Bobkov, Andrey A.; Reed, John C.; Liddington, Robert C.; Abagyan, Ruben

2010-01-01

349

Multiple Phosphatidylinositol 3-Kinases Regulate Vaccinia Virus Morphogenesis  

PubMed Central

Poxvirus morphogenesis is a complex process that involves the successive wrapping of the virus in host cell membranes. We screened by plaque assay a focused library of kinase inhibitors for those that caused a reduction in viral growth and identified several compounds that selectively inhibit phosphatidylinositol 3-kinase (PI3K). Previous studies demonstrated that PI3Ks mediate poxviral entry. Using growth curves and electron microscopy in conjunction with inhibitors, we show that that PI3Ks additionally regulate morphogenesis at two distinct steps: immature to mature virion (IMV) transition, and IMV envelopment to form intracellular enveloped virions (IEV). Cells derived from animals lacking the p85 regulatory subunit of Type I PI3Ks (p85??/???/?) presented phenotypes similar to those observed with PI3K inhibitors. In addition, VV appear to redundantly use PI3Ks, as PI3K inhibitors further reduce plaque size and number in p85??/???/? cells. Together, these data provide evidence for a novel regulatory mechanism for virion morphogenesis involving phosphatidylinositol dynamics and may represent a new therapeutic target to contain poxviruses.

McNulty, Shannon; Bornmann, William; Schriewer, Jill; Werner, Chas; Smith, Scott K.; Olson, Victoria A.; Damon, Inger K.; Buller, R. Mark; Heuser, John; Kalman, Daniel

2010-01-01

350

Immunological characterization of a modified vaccinia virus Ankara vector expressing the human papillomavirus 16 E1 protein.  

PubMed

Women showing normal cytology but diagnosed with a persistent high-risk human papillomavirus (HR-HPV) infection have a higher risk of developing high-grade cervical intraepithelial neoplasia and cervical cancer than noninfected women. As no therapeutic management other than surveillance is offered to these women, there is a major challenge to develop novel targeted therapies dedicated to the treatment of these patients. As such, E1 and E2 antigens, expressed early in the HPV life cycle, represent very interesting candidates. Both proteins are necessary for maintaining coordinated viral replication and gene synthesis during the differentiation process of the epithelium and are essential for the virus to complete its normal and propagative replication cycle. In the present study, we evaluated a new active targeted immunotherapeutic, a modified vaccinia virus Ankara (MVA) vector containing the E1 sequence of HPV16, aimed at inducing cellular immune responses with the potential to help and clear persistent HPV16-related infection. We carried out an extensive comparative time course analysis of the cellular immune responses induced by different schedules of immunization in C57BL/6 mice. We showed that multiple injections of MVA-E1 allowed sustained HPV16 E1-specific cellular immune responses in vaccinated mice and had no impact on the exhaustion phenotype of the generated HPV16 E1-specific CD8? T cells, but they led to the differentiation of multifunctional effector T cells with high cytotoxic capacity. This study provides proof of concept that an MVA expressing HPV16 E1 can induce robust and long-lasting E1-specific responses and warrants further development of this candidate. PMID:24307238

Remy-Ziller, Christelle; Germain, Claire; Spindler, Anita; Hoffmann, Chantal; Silvestre, Nathalie; Rooke, Ronald; Bonnefoy, Jean-Yves; Préville, Xavier

2014-02-01

351

Vaccinia virus-mediated melanin production allows MR and optoacoustic deep tissue imaging and laser-induced thermotherapy of cancer  

PubMed Central

We reported earlier the delivery of antiangiogenic single chain antibodies by using oncolytic vaccinia virus strains to enhance their therapeutic efficacy. Here, we provide evidence that gene-evoked production of melanin can be used as a therapeutic and diagnostic mediator, as exemplified by insertion of only one or two genes into the genome of an oncolytic vaccinia virus strain. We found that produced melanin is an excellent reporter for optical imaging without addition of substrate. Melanin production also facilitated deep tissue optoacoustic imaging as well as MRI. In addition, melanin was shown to be a suitable target for laser-induced thermotherapy and enhanced oncolytic viral therapy. In conclusion, melanin as a mediator for thermotherapy and reporter for different imaging modalities may soon become a versatile alternative to replace fluorescent proteins also in other biological systems. After ongoing extensive preclinical studies, melanin overproducing oncolytic virus strains might be used in clinical trials in patients with cancer.

Stritzker, Jochen; Kirscher, Lorenz; Scadeng, Miriam; Deliolanis, Nikolaos C.; Morscher, Stefan; Symvoulidis, Panagiotis; Schaefer, Karin; Zhang, Qian; Buckel, Lisa; Hess, Michael; Donat, Ulrike; Bradley, William G.; Ntziachristos, Vasilis; Szalay, Aladar A.

2013-01-01

352

Vaccinia virus A19 protein participates in the transformation of spherical immature particles to barrel-shaped infectious virions.  

PubMed

The A19L open reading frame of vaccinia virus encodes a 9-kDa protein that is conserved in all sequenced chordopoxviruses, yet until now it has not been specifically characterized in any species. We appended an epitope tag after the start codon of the A19L open reading frame without compromising infectivity. The protein was synthesized after viral DNA replication and was phosphorylated independently of the vaccinia virus F10 kinase. The A19 protein was present in purified virions and was largely resistant to nonionic detergent extraction, suggesting a location within the core. A conditional lethal mutant virus was constructed by placing the A19 open reading frame under the control of the Escherichia coli lac repressor system. A19 synthesis and infectious virus formation were dependent on inducer. In the absence of inducer, virion morphogenesis was interrupted, and spherical dense particles that had greatly reduced amounts of the D13 scaffold accumulated in place of barrel-shaped mature virions. The infectivity of purified A19-deficient particles was more than 2 log units less than that of A19-containing virions. Nevertheless, the A19-deficient particles contained DNA, and except for the absence of A19 and decreased core protein processing, they appeared to have a similar protein composition as A19-containing virions. Thus, the A19 protein participates in the maturation of immature vaccinia virus virions to infectious particles. PMID:23885081

Satheshkumar, P S; Weisberg, Andrea S; Moss, Bernard

2013-10-01

353

DNA Vaccines Against Influenza Viruses  

Microsoft Academic Search

\\u000a As an attractive alternative to conventional vaccines, DNA vaccines play a critical role in inducing protection against several\\u000a infectious diseases. In this review, we discuss the advantages that DNA vaccines offer in comparison to conventional protein-based\\u000a vaccines. We discuss strategies to improve the potency and efficacy of DNA vaccines. Specifically, we focus on the potential\\u000a use of DNA-based vaccines to

Jin Hyang Kim; Joshy Jacob

354

Structure of the uracil complex of Vaccinia virus uracil DNA glycosylase  

PubMed Central

Poxvirus uracil DNA glycosylases are the most diverse members of the family I uracil DNA glycosylases (UNGs). The crystal structure of the uracil complex of Vaccinia virus uracil DNA glycosylase (D4) was determined at 2.03?Å resolution. One uracil molecule was located in the active-site pocket in each of the 12 noncrystallographic symmetry-related D4 subunits. Although the UNGs of the poxviruses (including D4) feature significant differences in the characteristic motifs designated for uracil recognition and in the base-excision mechanism, the architecture of the active-site pocket in D4 is very similar to that in UNGs of other organisms. Overall, the interactions of the bound uracil with the active-site residues are also similar to the interactions previously observed in the structures of human and Escherichia coli UNG.

Schormann, N.; Banerjee, S.; Ricciardi, R.; Chattopadhyay, D.

2013-01-01

355

Regulation of vaccinia virus E3 protein by small ubiquitin-like modifier proteins.  

PubMed

The vaccinia virus (VACV) E3 protein is essential for virulence and has antiapoptotic activity and the ability to impair the host innate immune response. Here we demonstrate that E3 interacts with SUMO1 through a small ubiquitin-like modifier (SUMO)-interacting motif (SIM). SIM integrity is required for maintaining the stability of the viral protein and for the covalent conjugation of E3 to SUMO1 or SUMO2, a modification that has a negative effect on the E3 transcriptional transactivation of the p53-upregulated modulator of apoptosis (PUMA) and APAF-1 genes. We also demonstrate that E3 is ubiquitinated, a modification that does not destabilize the wild-type protein but triggers the degradation of an E3-?SIM mutant. This report constitutes the first demonstration of the important roles that both SUMO and ubiquitin play in the regulation of the VACV protein E3. PMID:21957283

González-Santamaría, José; Campagna, Michela; García, María Angel; Marcos-Villar, Laura; González, Dolores; Gallego, Pedro; Lopitz-Otsoa, Fernando; Guerra, Susana; Rodríguez, Manuel S; Esteban, Mariano; Rivas, Carmen

2011-12-01

356

Regulation of Vaccinia Virus E3 Protein by Small Ubiquitin-Like Modifier Proteins ? †  

PubMed Central

The vaccinia virus (VACV) E3 protein is essential for virulence and has antiapoptotic activity and the ability to impair the host innate immune response. Here we demonstrate that E3 interacts with SUMO1 through a small ubiquitin-like modifier (SUMO)-interacting motif (SIM). SIM integrity is required for maintaining the stability of the viral protein and for the covalent conjugation of E3 to SUMO1 or SUMO2, a modification that has a negative effect on the E3 transcriptional transactivation of the p53-upregulated modulator of apoptosis (PUMA) and APAF-1 genes. We also demonstrate that E3 is ubiquitinated, a modification that does not destabilize the wild-type protein but triggers the degradation of an E3-?SIM mutant. This report constitutes the first demonstration of the important roles that both SUMO and ubiquitin play in the regulation of the VACV protein E3.

Gonzalez-Santamaria, Jose; Campagna, Michela; Garcia, Maria Angel; Marcos-Villar, Laura; Gonzalez, Dolores; Gallego, Pedro; Lopitz-Otsoa, Fernando; Guerra, Susana; Rodriguez, Manuel S.; Esteban, Mariano; Rivas, Carmen

2011-01-01

357

Introduction of the Six Major Genomic Deletions of Modified Vaccinia Virus Ankara (MVA) into the Parental Vaccinia Virus Is Not Sufficient To Reproduce an MVA-Like Phenotype in Cell Culture and in Mice?  

PubMed Central

Modified vaccinia virus Ankara (MVA) has a highly restricted host range in cell culture and is apathogenic in vivo. MVA was derived from the parental chorioallantois vaccinia virus Ankara (CVA) by more than 570 passages in chicken embryo fibroblast (CEF) cells. During CEF cell passaging, six major deletions comprising 24,668 nucleotides occurred in the CVA genome. We have cloned both the MVA and the parental CVA genome as bacterial artificial chromosomes (BACs) and have sequentially introduced the six major MVA deletions into the cloned CVA genome. Reconstituted mutant CVA viruses containing up to six major MVA deletions showed no detectable replication restriction in 12 of 14 mammalian cell lines tested; the exceptions were rabbit cell lines RK13 and SIRC. In mice, CVA mutants with up to three deletions showed slightly enhanced virulence, suggesting that gene deletion in replicating vaccinia virus (VACV) can result in gain of fitness in vivo. CVA mutants containing five or all six deletions were still pathogenic, with a moderate degree of attenuation. Deletion V was mainly responsible for the attenuated phenotype of these mutants. In conclusion, loss or truncation of all 31 open reading frames in the six major deletions is not sufficient to reproduce the specific MVA phenotype of strong attenuation and highly restricted host range. Mutations in viral genes outside or in association with the six major deletions appear to contribute significantly to this phenotype. Host range restriction and avirulence of MVA are most likely a cooperative effect of gene deletions and mutations involving the major deletions.

Meisinger-Henschel, Christine; Spath, Michaela; Lukassen, Susanne; Wolferstatter, Michael; Kachelriess, Heike; Baur, Karen; Dirmeier, Ulrike; Wagner, Markus; Chaplin, Paul; Suter, Mark; Hausmann, Jurgen

2010-01-01

358

T-cell engager-armed oncolytic vaccinia virus significantly enhances antitumor therapy.  

PubMed

Oncolytic vaccinia virus (VV) therapy has shown promise in preclinical models and in clinical studies. However, complete responses have rarely been observed. This lack of efficacy is most likely due to suboptimal virus spread through the tumor resulting in limited tumor cell destruction. We reasoned that redirecting T cells to the tumor has the potential to improve the antitumor activity of oncolytic VVs. We, therefore, constructed a VV encoding a secretory bispecific T-cell engager consisting of two single- chain variable fragments specific for CD3 and the tumor cell surface antigen EphA2 (EphA2-T-cell engager-armed VV (EphA2-TEA-VV)). In vitro, EphA2-TEA-VV's ability to replicate and induce oncolysis was similar to that of unmodified virus. However, only tumor cells infected with EphA2-TEA-VV induced T-cell activation as judged by the secretion of interferon-? and interleukin-2. In coculture assays, EphA2-TEA-VV not only killed infected tumor cells, but in the presence of T cells, it also induced bystander killing of noninfected tumor cells. In vivo, EphA2-TEA-VV plus T cells had potent antitumor activity in comparison with control VV plus T cells in a lung cancer xenograft model. Thus, arming oncolytic VVs with T-cell engagers may represent a promising approach to improve oncolytic virus therapy. PMID:24135899

Yu, Feng; Wang, Xingbing; Guo, Z Sheng; Bartlett, David L; Gottschalk, Stephen M; Song, Xiao-Tong

2014-01-01

359

Silver nanoparticles inhibit vaccinia virus infection by preventing viral entry through a macropinocytosis-dependent mechanism.  

PubMed

Silver nanoparticles have been shown to inhibit viruses. However, very little is known about the mechanism of antiviral activity. This study tested the hypothesis that 25-nm silver nanoparticles inhibited Vaccinia virus replication by preventing viral entry. Plaque reduction, confocal microscopy, and beta-galactosidase reporter gene assays were used to examine viral attachment and entry in the presence and absence of silver nanoparticles. To explore the mechanism of inhibition, viral entry experiments were conducted with silver nanoparticles and small interfering RNAs designed to silence the gene coding for p21-activated kinase 1, a key mediator of macropinocytosis. The silver nanoparticles caused a 4- to 5-log reduction in viral titer at concentrations that were not toxic to cells. Virus was capable of adsorbing to cells but could not enter cells in the presence of silver nanoparticles. Virus particles that had adsorbed to cells in the presence of silver nanoparticles were found to be infectious upon removal from the cells, indicating lack of direct virucidal effect. The half maximal inhibitory concentration for viral entry in the presence of silver nanoparticles was 27.4+/-3.3 microg/ml. When macropinocytosis was blocked, this inhibition was significantly reduced. Thus, macropinocytosis was required for the full antiviral effect. For the first time, this study points to the novel result that a cellular process involved in viral entry is responsible for the antiviral effects of silver nanoparticles. PMID:23980510

Trefry, John C; Wooley, Dawn P

2013-09-01

360

Vaccines in Development against West Nile Virus  

PubMed Central

West Nile encephalitis emerged in 1999 in the United States, then rapidly spread through the North American continent causing severe disease in human and horses. Since then, outbreaks appeared in Europe, and in 2012, the United States experienced a new severe outbreak reporting a total of 5,387 cases of West Nile virus (WNV) disease in humans, including 243 deaths. So far, no human vaccine is available to control new WNV outbreaks and to avoid worldwide spreading. In this review, we discuss the state-of-the-art of West Nile vaccine development and the potential of a novel safe and effective approach based on recombinant live attenuated measles virus (MV) vaccine. MV vaccine is a live attenuated negative-stranded RNA virus proven as one of the safest, most stable and effective human vaccines. We previously described a vector derived from the Schwarz MV vaccine strain that stably expresses antigens from emerging arboviruses, such as dengue, West Nile or chikungunya viruses, and is strongly immunogenic in animal models, even in the presence of MV pre-existing immunity. A single administration of a recombinant MV vaccine expressing the secreted form of WNV envelope glycoprotein elicited protective immunity in mice and non-human primates as early as two weeks after immunization, indicating its potential as a human vaccine.

Brandler, Samantha; Tangy, Frederic

2013-01-01

361

Molecular characterization of the host defense activity of the barrier to autointegration factor against vaccinia virus.  

PubMed

The barrier to autointegration factor (BAF) is an essential cellular protein with functions in mitotic nuclear reassembly, retroviral preintegration complex stability, and transcriptional regulation. Molecular properties of BAF include the ability to bind double-stranded DNA in a sequence-independent manner, homodimerize, and bind proteins containing a LEM domain. These capabilities allow BAF to compact DNA and assemble higher-order nucleoprotein complexes, the nature of which is poorly understood. Recently, it was revealed that BAF also acts as a potent host defense against poxviral DNA replication in the cytoplasm. Here, we extend these observations by examining the molecular mechanism through which BAF acts as a host defense against vaccinia virus replication and cytoplasmic DNA in general. Interestingly, BAF rapidly relocalizes to transfected DNA from a variety of sources, demonstrating that BAF's activity as a host defense factor is not limited to poxviral infection. BAF's relocalization to cytoplasmic foreign DNA is highly dependent upon its DNA binding and dimerization properties but does not appear to require its LEM domain binding activity. However, the LEM domain protein emerin is recruited to cytoplasmic DNA in a BAF-dependent manner during both transfection and vaccinia virus infection. Finally, we demonstrate that the DNA binding and dimerization capabilities of BAF are essential for its function as an antipoxviral effector, while the presence of emerin is not required. Together, these data provide further mechanistic insight into which of BAF's molecular properties are employed by cells to impair the replication of poxviruses or respond to foreign DNA in general. PMID:21880762

Ibrahim, Nouhou; Wicklund, April; Wiebe, Matthew S

2011-11-01

362

Inactivated infectious haematopoietic necrosis virus (IHNV) vaccines.  

PubMed

The inactivation dynamics of infectious haematopoietic necrosis virus (IHNV) by b-propiolactone (BPL), binary ethylenimine (BEI), formaldehyde or heat and the antigenic and immunogenic properties of the inactivated vaccines were evaluated. Chemical treatment of IHNV with 2.7 mm BPL, 1.5 mm BEI or 50 mm formaldehyde abolished virus infectivity within 48 h whereas heat treatment at 50 or 100 degrees C rendered the virus innocuous within 30 min. The inactivated IHNV vaccines were recognized by rainbow trout, Oncorhynchus mykiss, IHNV-specific antibodies and were differentially recognized by antigenic site I or antigenic site II IHNV glycoprotein-specific neutralizing monoclonal antibodies. The BPL inactivated whole virus vaccine was highly efficacious in vaccinated rainbow trout challenged by waterborne exposure to IHNV 7, 28, 42 or 56 days (15 degrees C) after immunization. The formaldehyde inactivated whole virus vaccine was efficacious 7 or 11 days after vaccination of rainbow trout but performed inconsistently when tested at later time points. The other vaccines tested were not efficacious. PMID:18752542

Anderson, E; Clouthier, S; Shewmaker, W; Weighall, A; LaPatra, S

2008-10-01

363

Enhancement of MHC class I-restricted peptide-specific T cell induction by a DNA prime\\/MVA boost vaccination regime  

Microsoft Academic Search

Human immunodeficiency virus (HIV) vaccine candidates were previously constructed as a string of cytotoxic T lymphocyte (CTL) epitopes delivered and expressed using DNA and modified virus Ankara (MVA; an attenuated vaccinia virus) vectors. These vaccines were shown to induce interferon (IFN)-?-producing and cytolytic CD8+ T cells after a single vaccine administration. In the course of this work, immunization protocols were

T. Hanke; T. J. Blanchard; J. Schneider; C. M. Hannan; M. Becker; S. C. Gilbert; A. V. S. Hill; G. L. Smith; A. McMichael

1998-01-01

364

Comparing Adjuvanted H28 and Modified Vaccinia Virus Ankara Expressing H28 in a Mouse and a Non-Human Primate Tuberculosis Model  

PubMed Central

Here we report for the first time on the immunogenicity and protective efficacy of a vaccine strategy involving the adjuvanted fusion protein “H28” (consisting of Ag85B-TB10.4-Rv2660c) and Modified Vaccinia Virus Ankara expressing H28. We show that a heterologous prime-boost regimen involving priming with H28 in a Th1 adjuvant followed by boosting with H28 expressed by MVA (H28/MVA28) induced the highest percentage of IFN-? expressing T cells, the highest production of IFN-? per single cell and the highest induction of CD8 T cells compared to either of the vaccines given alone. In contrast, in mice vaccinated with adjuvanted recombinant H28 alone (H28/H28) we observed the highest production of IL-2 per single cell and the highest frequency of antigen specific TNF-?/IL-2 expressing CD4 T cells pre and post infection. Interestingly, TNF-?/IL-2 expressing central memory-like CD4 T cells showed a significant positive correlation with protection at week 6 post infection, whereas the opposite was observed for post infection CD4 T cells producing only IFN-?. Moreover, as a BCG booster vaccine in a clinically relevant non-human primate TB model, the H28/H28 vaccine strategy induced a slightly more prominent reduction of clinical disease and pathology for up to one year post infection compared to H28/MVA28. Taken together, our data showed that the adjuvanted subunit and MVA strategies led to different T cell subset combinations pre and post infection and that TNF-?/IL-2 double producing but not IFN-? single producing CD4 T cell subsets correlated with protection in the mouse TB model. Moreover, our data demonstrated that the H28 vaccine antigen was able to induce strong protection in both a mouse and a non-human primate TB model.

Billeskov, Rolf; Christensen, Jan P.; Aagaard, Claus; Andersen, Peter; Dietrich, Jes

2013-01-01

365

Homology between DNA polymerases of poxviruses, herpesviruses, and adenoviruses: nucleotide sequence of the vaccinia virus DNA polymerase gene.  

PubMed Central

A 5400-base-pair segment of the vaccinia virus genome was sequenced and an open reading frame of 938 codons was found precisely where the DNA polymerase had been mapped by transfer of a phosphonoacetate-resistance marker. A single nucleotide substitution changing glycine at position 347 to aspartic acid accounts for the drug resistance of the mutant vaccinia virus. The 5' end of the DNA polymerase mRNA was located 80 base pairs before the methionine codon initiating the open reading frame. Correspondence between the predicted Mr 108,577 polypeptide and the 110,000 purified enzyme indicates that little or no proteolytic processing occurs. Extensive homology, extending over 435 amino acids, was found upon comparing the DNA polymerase of vaccinia virus and DNA polymerase of Epstein-Barr virus. A highly conserved sequence of 14 amino acids in the carboxyl-terminal regions of the above DNA polymerases is also present at a similar location in adenovirus DNA polymerase. This structure, which is predicted to form a turn flanked by beta-pleated sheets, may form part of an essential binding or catalytic site that accounts for its presence in DNA polymerases of poxviruses, herpesviruses, and adenoviruses. Images

Earl, P L; Jones, E V; Moss, B

1986-01-01

366

Repression of Vaccinia Virus Holliday Junction Resolvase Inhibits Processing of Viral DNA into Unit-Length Genomes  

PubMed Central

The vaccinia virus A22R gene encodes a protein that is homologous to the bacterial enzyme RuvC and specifically cleaves and resolves four-way DNA Holliday junctions into linear duplex products. To investigate the role of the vaccinia virus Holliday junction resolvase during an infection, we constructed two recombinant viruses: vA22-HA, which has a short C-terminal epitope tag appended to the A22R open reading frame, and vA22i, in which the original A22R gene is deleted and replaced by an inducible copy. Polyacrylamide gel electrophoresis and Western blot analysis of extracts and purified virions from cells infected with vA22-HA revealed that the resolvase was expressed after the onset of DNA replication and incorporated into virion cores. vA22i exhibited a conditional replication defect. In the absence of an inducer, (i) viral protein synthesis was unaffected, (ii) late-stage viral DNA replication was reduced, (iii) most of the newly synthesized viral DNA remained in a branched or concatemeric form that caused it to be trapped at the application site during pulsed-field gel electrophoresis, (iv) cleavage of concatemer junctions was inhibited, and (v) virion morphogenesis was arrested at an immature stage. These data indicated multiple roles for the vaccinia virus Holliday junction resolvase in the replication and processing of viral DNA into unit-length genomes.

Garcia, Alonzo D.; Moss, Bernard

2001-01-01

367

Genetically engineered poxviruses for recombinant gene expression, vaccination, and safety.  

PubMed Central

Vaccinia virus, no longer required for immunization against smallpox, now serves as a unique vector for expressing genes within the cytoplasm of mammalian cells. As a research tool, recombinant vaccinia viruses are used to synthesize and analyze the structure-function relationships of proteins, determine the targets of humoral and cell-mediated immunity, and investigate the types of immune response needed for protection against specific infectious diseases and cancer. The vaccine potential of recombinant vaccinia virus has been realized in the form of an effective oral wild-life rabies vaccine, although no product for humans has been licensed. A genetically altered vaccinia virus that is unable to replicate in mammalian cells and produces diminished cytopathic effects retains the capacity for high-level gene expression and immunogenicity while promising exceptional safety for laboratory workers and potential vaccine recipients. Images Fig. 1 Fig. 2

Moss, B

1996-01-01

368

Genetically Engineered Poxviruses for Recombinant Gene Expression, Vaccination, and Safety  

NASA Astrophysics Data System (ADS)

Vaccinia virus, no longer required for immunization against smallpox, now serves as a unique vector for expressing genes within the cytoplasm of mammalian cells. As a research tool, recombinant vaccinia viruses are used to synthesize and analyze the structure--function relationships of proteins, determine the targets of humoral and cell-mediated immunity, and investigate the types of immune response needed for protection against specific infectious diseases and cancer. The vaccine potential of recombinant vaccinia virus has been realized in the form of an effective oral wild-life rabies vaccine, although no product for humans has been licensed. A genetically altered vaccinia virus that is unable to replicate in mammalian cells and produces diminished cytopathic effects retains the capacity for high-level gene expression and immunogenicity while promising exceptional safety for laboratory workers and potential vaccine recipients.

Moss, Bernard

1996-10-01

369

Vaccine and adjuvant design for emerging viruses  

PubMed Central

Vaccination is currently the most effective strategy to medically control viral diseases. However, developing vaccines is a long and expensive process and traditional methods, such as attenuating wild-type viruses by serial passage, may not be suitable for all viruses and may lead to vaccine safety considerations, particularly in the case of the vaccination of particular patient groups, such as the immunocompromised and the elderly. In particular, developing vaccines against emerging viral pathogens adds a further level of complexity, as they may only be administered to small groups of people or only in response to a specific event or threat, limiting our ability to study and evaluate responses. In this commentary, we discuss how novel techniques may be used to engineer a new generation of vaccine candidates as we move toward a more targeted vaccine design strategy, driven by our understanding of the mechanisms of viral pathogenesis, attenuation and the signaling events which are required to develop a lasting, protective immunity. We will also briefly discuss the potential future role of vaccine adjuvants, which could be used to bridge the gap between vaccine safety and lasting immunity from a single vaccination.

McAuley, Alexander J

2011-01-01

370

Increased ATP generation in the host cell is required for efficient vaccinia virus production.  

PubMed

To search for cellular genes up-regulated by vaccinia virus (VV) infection, differential display-reverse transcription-polymerase chain reaction (ddRT-PCR) assays were used to examine the expression of mRNAs from mock-infected and VV-infected HeLa cells. Two mitochondrial genes for proteins that are part of the electron transport chain that generates ATP, ND4 and CO II, were up-regulated after VV infection. Up-regulation of ND4 level by VV infection was confirmed by Western blotting analysis. Up-regulation of ND4 was reduced by the MAPK inhibitor, apigenin, which has been demonstrated elsewhere to inhibit VV replication. The induction of ND4 expression occurred after viral DNA replication since ara C, an inhibitor of poxviral DNA replication, could block this induction. ATP production was increased in the host cells after VV infection. Moreover, 4.5 microM oligomycin, an inhibitor of ATP production, reduced the ATP level 13 hr after virus infection to that of mock-infected cells and inhibited viral protein expression and virus production, suggesting that increased ATP production is required for efficient VV production. Our results further suggest that induction of ND4 expression is through a Bcl-2 independent pathway. PMID:19725950

Chang, Chia-Wei; Li, Hui-Chun; Hsu, Che-Fang; Chang, Chiao-Yen; Lo, Shih-Yen

2009-01-01

371

Increased ATP generation in the host cell is required for efficient vaccinia virus production  

PubMed Central

To search for cellular genes up-regulated by vaccinia virus (VV) infection, differential display-reverse transcription-polymerase chain reaction (ddRT-PCR) assays were used to examine the expression of mRNAs from mock-infected and VV-infected HeLa cells. Two mitochondrial genes for proteins that are part of the electron transport chain that generates ATP, ND4 and CO II, were up-regulated after VV infection. Up-regulation of ND4 level by VV infection was confirmed by Western blotting analysis. Up-regulation of ND4 was reduced by the MAPK inhibitor, apigenin, which has been demonstrated elsewhere to inhibit VV replication. The induction of ND4 expression occurred after viral DNA replication since ara C, an inhibitor of poxviral DNA replication, could block this induction. ATP production was increased in the host cells after VV infection. Moreover, 4.5 ?M oligomycin, an inhibitor of ATP production, reduced the ATP level 13 hr after virus infection to that of mock-infected cells and inhibited viral protein expression and virus production, suggesting that increased ATP production is required for efficient VV production. Our results further suggest that induction of ND4 expression is through a Bcl-2 independent pathway.

Chang, Chia-Wei; Li, Hui-Chun; Hsu, Che-Fang; Chang, Chiao-Yen; Lo, Shih-Yen

2009-01-01

372

Role for CCR5 in dissemination of vaccinia virus in vivo.  

PubMed

In an earlier report, we provided evidence that expression of CCR5 by primary human T cells renders them permissive for vaccinia virus (VACV) replication. This may represent a mechanism for dissemination throughout the lymphatic system. To test this hypothesis, wild-type CCR5(+/+) and CCR5 null mice were challenged with VACV by intranasal inoculation. In time course studies using different infective doses of VACV, we identified viral replication in the lungs of both CCR5(+/+) and CCR5(-/-) mice, yet there were diminished viral loads in the spleens and brains of CCR5(-/-) mice compared with CCR5(+/+) mice. Moreover, in association with VACV infection, we provide evidence for CD4+ and CD8+ T-cell as well as CD11c+ and F4/80+ cell infiltration into the lungs of CCR5(+/+) but not CCR5(-/-) mice, and we show that the CCR5-expressing T cells harbor virus. We demonstrate that this CCR5 dependence is VACV specific, since CCR5(-/-) mice are as susceptible to intranasal influenza virus (A/WSN/33) infection as CCR5(+/+) mice. In a final series of experiments, we provide evidence that adoptive transfer of CCR5(+/+) bone marrow leukocytes into CCR5(-/-) mice restores VACV permissiveness, with evidence of lung and spleen infection. Taken together, our data suggest a novel role for CCR5 in VACV dissemination in vivo. PMID:19073715

Rahbar, Ramtin; Murooka, Thomas T; Fish, Eleanor N

2009-03-01

373

Role for CCR5 in Dissemination of Vaccinia Virus In Vivo?  

PubMed Central

In an earlier report, we provided evidence that expression of CCR5 by primary human T cells renders them permissive for vaccinia virus (VACV) replication. This may represent a mechanism for dissemination throughout the lymphatic system. To test this hypothesis, wild-type CCR5+/+ and CCR5 null mice were challenged with VACV by intranasal inoculation. In time course studies using different infective doses of VACV, we identified viral replication in the lungs of both CCR5+/+ and CCR5?/? mice, yet there were diminished viral loads in the spleens and brains of CCR5?/? mice compared with CCR5+/+ mice. Moreover, in association with VACV infection, we provide evidence for CD4+ and CD8+ T-cell as well as CD11c+ and F4/80+ cell infiltration into the lungs of CCR5+/+ but not CCR5?/? mice, and we show that the CCR5-expressing T cells harbor virus. We demonstrate that this CCR5 dependence is VACV specific, since CCR5?/? mice are as susceptible to intranasal influenza virus (A/WSN/33) infection as CCR5+/+ mice. In a final series of experiments, we provide evidence that adoptive transfer of CCR5+/+ bone marrow leukocytes into CCR5?/? mice restores VACV permissiveness, with evidence of lung and spleen infection. Taken together, our data suggest a novel role for CCR5 in VACV dissemination in vivo.

Rahbar, Ramtin; Murooka, Thomas T.; Fish, Eleanor N.

2009-01-01

374

Modified Vaccinia Virus Ankara Triggers Type I IFN Production in Murine Conventional Dendritic Cells via a cGAS/STING-Mediated Cytosolic DNA-Sensing Pathway  

PubMed Central

Modified vaccinia virus Ankara (MVA) is an attenuated poxvirus that has been engineered as a vaccine against infectious agents and cancers. Our goal is to understand how MVA modulates innate immunity in dendritic cells (DCs), which can provide insights to vaccine design. In this study, using murine bone marrow-derived dendritic cells, we assessed type I interferon (IFN) gene induction and protein secretion in response to MVA infection. We report that MVA infection elicits the production of type I IFN in murine conventional dendritic cells (cDCs), but not in plasmacytoid dendritic cells (pDCs). Transcription factors IRF3 (IFN regulatory factor 3) and IRF7, and the positive feedback loop mediated by IFNAR1 (IFN alpha/beta receptor 1), are required for the induction. MVA induction of type I IFN is fully dependent on STING (stimulator of IFN genes) and the newly discovered cytosolic DNA sensor cGAS (cyclic guanosine monophosphate-adenosine monophosphate synthase). MVA infection of cDCs triggers phosphorylation of TBK1 (Tank-binding kinase 1) and IRF3, which is abolished in the absence of cGAS and STING. Furthermore, intravenous delivery of MVA induces type I IFN in wild-type mice, but not in mice lacking STING or IRF3. Treatment of cDCs with inhibitors of endosomal and lysosomal acidification or the lysosomal enzyme Cathepsin B attenuated MVA-induced type I IFN production, indicating that lysosomal enzymatic processing of virions is important for MVA sensing. Taken together, our results demonstrate a critical role of the cGAS/STING-mediated cytosolic DNA-sensing pathway for type I IFN induction in cDCs by MVA. We present evidence that vaccinia virulence factors E3 and N1 inhibit the activation of IRF3 and the induction of IFNB gene in MVA-infected cDCs.

Cao, Hua; Avogadri, Francesca; Dai, Lianpan; Drexler, Ingo; Joyce, Johanna A.; Li, Xiao-Dong; Chen, Zhijian; Merghoub, Taha; Shuman, Stewart; Deng, Liang

2014-01-01

375

Modified vaccinia virus Ankara triggers type I IFN production in murine conventional dendritic cells via a cGAS/STING-mediated cytosolic DNA-sensing pathway.  

PubMed

Modified vaccinia virus Ankara (MVA) is an attenuated poxvirus that has been engineered as a vaccine against infectious agents and cancers. Our goal is to understand how MVA modulates innate immunity in dendritic cells (DCs), which can provide insights to vaccine design. In this study, using murine bone marrow-derived dendritic cells, we assessed type I interferon (IFN) gene induction and protein secretion in response to MVA infection. We report that MVA infection elicits the production of type I IFN in murine conventional dendritic cells (cDCs), but not in plasmacytoid dendritic cells (pDCs). Transcription factors IRF3 (IFN regulatory factor 3) and IRF7, and the positive feedback loop mediated by IFNAR1 (IFN alpha/beta receptor 1), are required for the induction. MVA induction of type I IFN is fully dependent on STING (stimulator of IFN genes) and the newly discovered cytosolic DNA sensor cGAS (cyclic guanosine monophosphate-adenosine monophosphate synthase). MVA infection of cDCs triggers phosphorylation of TBK1 (Tank-binding kinase 1) and IRF3, which is abolished in the absence of cGAS and STING. Furthermore, intravenous delivery of MVA induces type I IFN in wild-type mice, but not in mice lacking STING or IRF3. Treatment of cDCs with inhibitors of endosomal and lysosomal acidification or the lysosomal enzyme Cathepsin B attenuated MVA-induced type I IFN production, indicating that lysosomal enzymatic processing of virions is important for MVA sensing. Taken together, our results demonstrate a critical role of the cGAS/STING-mediated cytosolic DNA-sensing pathway for type I IFN induction in cDCs by MVA. We present evidence that vaccinia virulence factors E3 and N1 inhibit the activation of IRF3 and the induction of IFNB gene in MVA-infected cDCs. PMID:24743339

Dai, Peihong; Wang, Weiyi; Cao, Hua; Avogadri, Francesca; Dai, Lianpan; Drexler, Ingo; Joyce, Johanna A; Li, Xiao-Dong; Chen, Zhijian; Merghoub, Taha; Shuman, Stewart; Deng, Liang

2014-04-01

376

Emerging Respiratory Viruses: Challenges and Vaccine Strategies  

PubMed Central

The current threat of avian influenza to the human population, the potential for the reemergence of severe acute respiratory syndrome (SARS)-associated coronavirus, and the identification of multiple novel respiratory viruses underline the necessity for the development of therapeutic and preventive strategies to combat viral infection. Vaccine development is a key component in the prevention of widespread viral infection and in the reduction of morbidity