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Sample records for acam2000 smallpox vaccinia

  1. ACAM2000™: The new smallpox vaccine for United States Strategic National Stockpile

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    Aysegul Nalca

    2010-05-01

    Full Text Available Aysegul Nalca, Elizabeth E ZumbrunCenter for Aerobiological Sciences, US Army Medical Research Institute of Infectious Diseases (USAMRIID, Fort Detrick, MD, USAAbstract: Smallpox was eradicated more than 30 years ago, but heightened concerns over bioterrorism have brought smallpox and smallpox vaccination back to the forefront. The previously licensed smallpox vaccine in the United States, Dryvax® (Wyeth Laboratories, Inc., was highly effective, but the supply was insufficient to vaccinate the entire current US population. Additionally, Dryvax® had a questionable safety profile since it consisted of a pool of vaccinia virus strains with varying degrees of virulence, and was grown on the skin of calves, an outdated technique that poses an unnecessary risk of contamination. The US government has therefore recently supported development of an improved live vaccinia virus smallpox vaccine. This initiative has resulted in the development of ACAM2000™ (Acambis, Inc.™, a single plaque-purified vaccinia virus derivative of Dryvax®, aseptically propagated in cell culture. Preclinical and clinical trials reported in 2008 demonstrated that ACAM2000™ has comparable immunogenicity to that of Dryvax®, and causes a similar frequency of adverse events. Furthermore, like Dryvax®, ACAM2000™ vaccination has been shown by careful cardiac screening to result in an unexpectedly high rate of myocarditis and pericarditis. ACAM2000™ received US Food and Drug Administration (FDA approval in August 2007, and replaced Dryvax® for all smallpox vaccinations in February 2008. Currently, over 200 million doses of ACAM2000™ have been produced for the US Strategic National Stockpile. This review of ACAM2000™ addresses the production, characterization, clinical trials, and adverse events associated with this new smallpox vaccine.Keywords: smallpox, vaccinia, variola, vaccine, efficacy, safety

  2. Human antibody responses to the polyclonal Dryvax vaccine for smallpox prevention can be distinguished from responses to the monoclonal replacement vaccine ACAM2000.

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    Pugh, Christine; Keasey, Sarah; Korman, Lawrence; Pittman, Phillip R; Ulrich, Robert G

    2014-06-01

    Dryvax (Wyeth Laboratories, Inc., Marietta, PA) is representative of the vaccinia virus preparations that were previously used for preventing smallpox. While Dryvax was highly effective, the national supply stocks were depleted, and there were manufacturing concerns regarding sterility and the clonal heterogeneity of the vaccine. ACAM2000 (Acambis, Inc./Sanofi-Pasteur Biologics Co., Cambridge, MA), a single-plaque-purified vaccinia virus derivative of Dryvax, recently replaced the polyclonal smallpox vaccine for use in the United States. A substantial amount of sequence heterogeneity exists within the polyclonal proteome of Dryvax, including proteins that are missing from ACAM2000. Reasoning that a detailed comparison of antibody responses to the polyclonal and monoclonal vaccines may be useful for identifying unique properties of each antibody response, we utilized a protein microarray comprised of approximately 94% of the vaccinia poxvirus proteome (245 proteins) to measure protein-specific antibody responses of 71 individuals receiving a single vaccination with ACAM2000 or Dryvax. We observed robust antibody responses to 21 poxvirus proteins in vaccinated individuals, including 11 proteins that distinguished Dryvax responses from ACAM2000. Analysis of protein sequences from Dryvax clones revealed amino acid level differences in these 11 antigenic proteins and suggested that sequence variation and clonal heterogeneity may contribute to the observed differences between Dryvax and ACAM2000 antibody responses.

  3. Treatment with the smallpox antiviral tecovirimat (ST-246) alone or in combination with ACAM2000 vaccination is effective as a postsymptomatic therapy for monkeypox virus infection.

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    Berhanu, Aklile; Prigge, Jonathan T; Silvera, Peter M; Honeychurch, Kady M; Hruby, Dennis E; Grosenbach, Douglas W

    2015-07-01

    The therapeutic efficacies of smallpox vaccine ACAM2000 and antiviral tecovirimat given alone or in combination starting on day 3 postinfection were compared in a cynomolgus macaque model of lethal monkeypox virus infection. Postexposure administration of ACAM2000 alone did not provide any protection against severe monkeypox disease or mortality. In contrast, postexposure treatment with tecovirimat alone or in combination with ACAM2000 provided full protection. Additionally, tecovirimat treatment delayed until day 4, 5, or 6 postinfection was 83% (days 4 and 5) or 50% (day 6) effective.

  4. Use of Vaccinia Virus Smallpox Vaccine in Laboratory and Health Care Personnel at Risk for Occupational Exposure to Orthopoxviruses - Recommendations of the Advisory Committee on Immunization Practices (ACIP), 2015.

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    Petersen, Brett W; Harms, Tiara J; Reynolds, Mary G; Harrison, Lee H

    2016-03-18

    On June 25, 2015, the Advisory Committee on Immunization Practices (ACIP) recommended routine vaccination with live smallpox (vaccinia) vaccine (ACAM2000) for laboratory personnel who directly handle 1) cultures or 2) animals contaminated or infected with replication-competent vaccinia virus, recombinant vaccinia viruses derived from replication-competent vaccinia strains (i.e., those that are capable of causing clinical infection and producing infectious virus in humans), or other orthopoxviruses that infect humans (e.g., monkeypox, cowpox, and variola) (recommendation category: A, evidence type 2 [Box]). Health care personnel (e.g., physicians and nurses) who currently treat or anticipate treating patients with vaccinia virus infections and whose contact with replication-competent vaccinia viruses is limited to contaminated materials (e.g., dressings) and persons administering ACAM2000 smallpox vaccine who adhere to appropriate infection prevention measures can be offered vaccination with ACAM2000 (recommendation category: B, evidence type 2 [Box]). These revised recommendations update the previous ACIP recommendations for nonemergency use of vaccinia virus smallpox vaccine for laboratory and health care personnel at risk for occupational exposure to orthopoxviruses (1). Since 2001, when the previous ACIP recommendations were developed, ACAM2000 has replaced Dryvax as the only smallpox vaccine licensed by the U.S. Food and Drug Administration (FDA) and available for use in the United States (2). These recommendations contain information on ACAM2000 and its use in laboratory and health care personnel at risk for occupational exposure to orthopoxviruses.

  5. Smallpox vaccine, ACAM2000: Sites and duration of viral shedding and effect of povidone iodine on scarification site shedding and immune response.

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    Pittman, Phillip R; Garman, Patrick M; Kim, Sung-Han; Schmader, Trevor J; Nieding, William J; Pike, Jason G; Knight, Ryan; Johnston, Sara C; Huggins, John W; Kortepeter, Mark G; Korman, Lawrence; Ranadive, Manmohan; Quinn, Xiaofei; Meyers, Mitchell S

    2015-06-12

    The U.S. Department of Defense vaccinates personnel deployed to high-risk areas with the vaccinia virus (VACV)-based smallpox vaccine. Autoinoculations and secondary and tertiary transmissions due to VACV shedding from the vaccination site continue to occur despite education of vaccinees on the risks of such infections. The objectives of this study were to investigate, in naïve smallpox vaccinees, (a) whether the vaccination site can remain contagious after the scab separates and (b) whether the application of povidone iodine ointment (PIO) to the vaccination site inactivates VACV without affecting the immune response. These objectives were tested in 60 individuals scheduled to receive smallpox vaccine. Thirty individuals (control) did not receive PIO; 30 subjects (treatment) received PIO starting on post-vaccination day 7. Counter to current dogma, this study showed that VACV continues to shed from the vaccination site after the scab separates. Overall viral shedding levels in the PIO group were significantly lower than those in the control group (p=0.0045), and PIO significantly reduced the duration of viral shedding (median duration 14.5 days and 21 days in the PIO and control groups, respectively; p=0.0444). At least 10% of control subjects continued to shed VACV at day 28, and 3.4% continued to shed the virus at day 42. PIO reduced the proportion of subjects shedding virus from the vaccination site from day 8 until days 21-23 compared with control subjects. Groups did not differ significantly in the proportion of subjects mounting an immune response, as measured by neutralizing antibodies, IgM, IgG, and interferon-gamma enzyme-linked immunospot assay. When applied to the vaccination site starting on day 7, PIO reduced viral shedding without altering the immune response. The use of PIO in addition to a semipermeable dressing may reduce the rates of autoinoculation and contact transmission originating from the vaccination site in smallpox-vaccinated individuals.

  6. New smallpox vaccines for an ancient scourge.

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    Frey, Sharon E

    2014-01-01

    The potential use of variola virus, a Class A agent of bioterrorism, remains a concern. In an effort to prepare for a possible smallpox outbreak due to an intentional release of variola, the U.S. government and industry have been evaluating vaccines stored in the National Strategic Stockpile including cell culture grown ACAM2000 and modified vaccinia Ankara, IMVAMUNE, in clinical studies. This paper discusses smallpox vaccines studies conducted at the Saint Louis University Center for Vaccine Development.

  7. Cutaneous responses to vaccinia in individuals with previous smallpox vaccination.

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    Simpson, Eric L; Hercher, Michelle; Hammarlund, Erika K; Lewis, Matthew W; Slifka, Mark K; Hanifin, Jon M

    2007-09-01

    The durability of immune responses to smallpox vaccine is a subject of considerable debate. We compared cutaneous vaccinia responses in patients vaccinated in the distant past with vaccine-naïve individuals using serial close-up photographs. The previously vaccinated group had a significantly reduced time course and milder cutaneous reactions. Vaccinated individuals appear to maintain clinically detectable immunity against vaccinia for at least 20 years after smallpox vaccination.

  8. Genomic sequence and analysis of a vaccinia virus isolate from a patient with a smallpox vaccine-related complication

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    Damon Inger

    2006-10-01

    Full Text Available Abstract Background Vaccinia virus (VACV-DUKE was isolated from a lesion on a 54 year old female who presented to a doctor at the Duke University Medical Center. She was diagnosed with progressive vaccinia and treated with vaccinia immune globulin. The availability of the VACV-DUKE genome sequence permits a first time genomic comparison of a VACV isolate associated with a smallpox vaccine complication with the sequence of culture-derived clonal isolates of the Dryvax vaccine. Results This study showed that VACV-DUKE is most similar to VACV-ACAM2000 and CLONE3, two VACV clones isolated from the Dryvax® vaccine stock confirming VACV-DUKE as an isolate from Dryvax®. However, VACV-DUKE is unique because it is, to date, the only Dryvax® clone isolated from a patient experiencing a vaccine-associated complication. The 199,960 bp VACV-DUKE genome encodes 225 open reading frames, including 178 intact genes and 47 gene fragments. Between VACV-DUKE and the other Dryvax® isolates, the major genomic differences are in fragmentation of the ankyrin-like, and kelch-like genes, presence of a full-length Interferon-α/β receptor gene, and the absence of a duplication of 12 ORFs in the inverted terminal repeat. Excluding this region, the DNA sequence of VACV-DUKE differs from the other two Dryvax® isolates by less than 0.4%. DNA sequencing also indicated that there was little heterogeneity in the sample, supporting the hypothesis that virus from an individual lesion is clonal in origin despite the fact that the vaccine is a mixed population. Conclusion Virus in lesions that result from progressive vaccinia following vaccination with Dryvax are likely clonal in origin. The genomic sequence of VACV-DUKE is overall very similar to that of Dryvax® cell culture-derived clonal isolates. Furthermore, with the sequences of multiple clones from Dryvax® we can begin to appreciate the diversity of the viral population in the smallpox vaccine.

  9. Analysis of variola and vaccinia virus neutralization assays for smallpox vaccines.

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    Hughes, Christine M; Newman, Frances K; Davidson, Whitni B; Olson, Victoria A; Smith, Scott K; Holman, Robert C; Yan, Lihan; Frey, Sharon E; Belshe, Robert B; Karem, Kevin L; Damon, Inger K

    2012-07-01

    Possible smallpox reemergence drives research for third-generation vaccines that effectively neutralize variola virus. A comparison of neutralization assays using different substrates, variola and vaccinia (Dryvax and modified vaccinia Ankara [MVA]), showed significantly different 90% neutralization titers; Dryvax underestimated while MVA overestimated variola neutralization. Third-generation vaccines may rely upon neutralization as a correlate of protection.

  10. Vaccinia virus induces rapid necrosis in keratinocytes by a STAT3-dependent mechanism.

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    Yong He

    Full Text Available Humans with a dominant negative mutation in STAT3 are susceptible to severe skin infections, suggesting an essential role for STAT3 signaling in defense against cutaneous pathogens.To focus on innate antiviral defenses in keratinocytes, we used a standard model of cutaneous infection of severe combined immunodeficient mice with the current smallpox vaccine, ACAM-2000. In parallel, early events post-infection with the smallpox vaccine ACAM-2000 were investigated in cultured keratinocytes of human and mouse origin.Mice treated topically with a STAT3 inhibitor (Stattic developed larger vaccinia lesions with higher virus titers and died more rapidly than untreated controls. Cultured human and murine keratinocytes infected with ACAM-2000 underwent rapid necrosis, but when treated with Stattic or with inhibitors of RIP1 kinase or caspase-1, they survived longer, produced higher titers of virus, and showed reduced activation of type I interferon responses and inflammatory cytokines release. Treatment with inhibitors of RIP1 kinase and STAT3, but not caspase-1, also reduced the inflammatory response of keratinocytes to TLR ligands. Vaccinia growth properties in Vero cells, which are known to be defective in some antiviral responses, were unaffected by inhibition of RIP1K, caspase-1, or STAT3.Our findings indicate that keratinocytes suppress the replication and spread of vaccinia virus by undergoing rapid programmed cell death, in a process requiring STAT3. These data offer a new framework for understanding susceptibility to skin infection in patients with STAT3 mutations. Interventions which promote prompt necroptosis/pyroptosis of infected keratinocytes may reduce risks associated with vaccination with live vaccinia virus.

  11. Clinical guidance for smallpox vaccine use in a postevent vaccination program.

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    Petersen, Brett W; Damon, Inger K; Pertowski, Carol A; Meaney-Delman, Dana; Guarnizo, Julie T; Beigi, Richard H; Edwards, Kathryn M; Fisher, Margaret C; Frey, Sharon E; Lynfield, Ruth; Willoughby, Rodney E

    2015-02-20

    This report outlines recommendations for the clinical use of the three smallpox vaccines stored in the U.S. Strategic National Stockpile for persons who are exposed to smallpox virus or at high risk for smallpox infection during a postevent vaccination program following an intentional or accidental release of the virus. No absolute contraindications exist for smallpox vaccination in a postevent setting. However, several relative contraindications exist among persons with certain medical conditions. CDC recommendations for smallpox vaccine use were developed in consideration of the risk for smallpox infection, risk for an adverse event following vaccination, and benefit from vaccination. Smallpox vaccines are made from live vaccinia viruses that protect against smallpox disease. They do not contain variola virus, the causative agent of smallpox. The three smallpox vaccines stockpiled are ACAM2000, Aventis Pasteur Smallpox Vaccine (APSV), and Imvamune. Surveillance and containment activities including vaccination with replication-competent smallpox vaccine (i.e., vaccine viruses capable of replicating in mammalian cells such as ACAM2000 and APSV) will be the primary response strategy for achieving epidemic control. Persons exposed to smallpox virus are at high risk for developing and transmitting smallpox and should be vaccinated with a replication-competent smallpox vaccine unless severely immunodeficient. Because of a high likelihood of a poor immune response and an increased risk for adverse events, smallpox vaccination should be avoided in persons with severe immunodeficiency who are not expected to benefit from vaccine, including bone marrow transplant recipients within 4 months of transplantation, persons infected with HIV with CD4 cell counts smallpox virus exposure in persons with severe immunodeficiency. Persons without a known smallpox virus exposure might still be at high risk for developing smallpox infection depending on the magnitude of the outbreak and

  12. HLA alleles associated with the adaptive immune response to smallpox vaccine: a replication study.

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    Ovsyannikova, Inna G; Pankratz, V Shane; Salk, Hannah M; Kennedy, Richard B; Poland, Gregory A

    2014-09-01

    We previously reported HLA allelic associations with vaccinia virus (VACV)-induced adaptive immune responses in a cohort of healthy individuals (n = 1,071 subjects) after a single dose of the licensed smallpox (Dryvax) vaccine. This study demonstrated that specific HLA alleles were significantly associated with VACV-induced neutralizing antibody (NA) titers (HLA-B*13:02, *38:02, *44:03, *48:01, and HLA-DQB1*03:02, *06:04) and cytokine (HLA-DRB1*01:03, *03:01, *10:01, *13:01, *15:01) immune responses. We undertook an independent study of 1,053 healthy individuals and examined associations between HLA alleles and measures of adaptive immunity after a single dose of Dryvax-derived ACAM2000 vaccine to evaluate previously discovered HLA allelic associations from the Dryvax study and determine if these associations are replicated with ACAM2000. Females had significantly higher NA titers than male subjects in both study cohorts [median ID50 discovery cohort 159 (93, 256) vs. 125 (75, 186), p smallpox vaccine-induced adaptive immune responses are significantly influenced by HLA gene polymorphisms. These data provide information for functional studies and design of novel candidate smallpox vaccines.

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

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    Fang, Qing; Yang, Lin; Zhu, Weijun; Liu, Li; Wang, Haibo; Yu, Wenbo; Xiao, Genfu; Tien, Po; Zhang, Linqi; Chen, Zhiwei

    2005-05-10

    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.

  14. Cardiac safety of Modified Vaccinia Ankara for vaccination against smallpox in a young, healthy study population.

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    Eva-Maria Zitzmann-Roth

    Full Text Available Conventional smallpox vaccines based on replicating vaccinia virus (VV strains (e.g. Lister Elstree, NYCBOH are associated with a high incidence of myo-/pericarditis, a severe inflammatory cardiac complication. A new smallpox vaccine candidate based on a non-replicating Modified Vaccinia Ankara (MVA poxvirus has been assessed for cardiac safety in a large placebo-controlled clinical trial.Cardiac safety of one and two doses of MVA compared to placebo was assessed in 745 healthy subjects. Vaccinia-naïve subjects received either one dose of MVA and one dose of placebo, two doses of MVA, or two doses of placebo by subcutaneous injection four weeks apart; vaccinia-experienced subjects received a single dose of MVA. Solicited and unsolicited adverse events (AE and cardiac safety parameters (recorded as Adverse Events of Special Interest, AESI were monitored after each injection.A total of 5 possibly related AESI (3 cases of palpitations, 2 of tachycardia were reported during the study. No case of myo- or pericarditis occurred. One possibly related serious AE (SAE was reported during the 6-month follow-up period (sarcoidosis. The most frequently observed AEs were injection site reactions.Vaccination with MVA was safe and well tolerated and did not increase the risk for development of myo-/pericarditis.ClinicalTrials.gov NCT00316524.

  15. 42 CFR 102.21 - Smallpox (Vaccinia) Vaccine Injury Table.

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    2010-10-01

    ... inflammatory cells in the dermis of the skin at the vaccination or inoculation site. The diagnosis of PV may be... the mother that results from the placental transmission of the vaccinia virus during any time in the... membrane lesion containing an accumulation of white blood cells. (8) Recipient means a person to whom the...

  16. A vaccinia virus renaissance: new vaccine and immunotherapeutic uses after smallpox eradication.

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    Verardi, Paulo H; Titong, Allison; Hagen, Caitlin J

    2012-07-01

    In 1796, Edward Jenner introduced the concept of vaccination with cowpox virus, an Orthopoxvirus within the family Poxviridae that elicits cross protective immunity against related orthopoxviruses, including smallpox virus (variola virus). Over time, vaccinia virus (VACV) replaced cowpox virus as the smallpox vaccine, and vaccination efforts eventually led to the successful global eradication of smallpox in 1979. VACV has many characteristics that make it an excellent vaccine and that were crucial for the successful eradication of smallpox, including (1) its exceptional thermal stability (a very important but uncommon characteristic in live vaccines), (2) its ability to elicit strong humoral and cell-mediated immune responses, (3) the fact that it is easy to propagate, and (4) that it is not oncogenic, given that VACV replication occurs exclusively within the host cell cytoplasm and there is no evidence that the viral genome integrates into the host genome. Since the eradication of smallpox, VACV has experienced a renaissance of interest as a viral vector for the development of recombinant vaccines, immunotherapies, and oncolytic therapies, as well as the development of next-generation smallpox vaccines. This revival is mainly due to the successful use and extensive characterization of VACV as a vaccine during the smallpox eradication campaign, along with the ability to genetically manipulate its large dsDNA genome while retaining infectivity and immunogenicity, its wide mammalian host range, and its natural tropism for tumor cells that allows its use as an oncolytic vector. This review provides an overview of new uses of VACV that are currently being explored for the development of vaccines, immunotherapeutics, and oncolytic virotherapies.

  17. Bioluminescent imaging of vaccinia virus infection in immunocompetent and immunodeficient rats as a model for human smallpox

    OpenAIRE

    Qiang Liu; Changfa Fan; Shuya Zhou; Yanan Guo; Qin Zuo; Jian Ma; Susu Liu; Xi Wu; Zexu Peng; Tao Fan; Chaoshe Guo; Yuelei Shen; Weijin Huang; Baowen Li; Zhengming He

    2015-01-01

    Due to the increasing concern of using smallpox virus as biological weapons for terrorist attack, there is renewed interest in studying the pathogenesis of human smallpox and development of new therapies. Animal models are highly demanded for efficacy and safety examination of new vaccines and therapeutic drugs. Here, we demonstrated that both wild type and immunodeficient rats infected with an engineered vaccinia virus carrying Firefly luciferase reporter gene (rTV-Fluc) could recapitulate i...

  18. ACAM2000(TM): The New Smallpox Vaccine for United States Strategic National Stockpile

    Science.gov (United States)

    2010-01-01

    lupus erythematosus . Am J Med. 1990;89(4):411–419. 53. Woodruff JF. Viral myocarditis. A review. Am J Pathol. 1980;101(2): 425–484. 54. Berhanu A, King...attempting to limit vaccination to those without immunodeficiencies, eczema, or pregnancy . Among 730,580 DOD vaccinees, three cases of post... lupus , or infections such as HIV or parvovirus B19.49 Interestingly, patients with lupus or parvovirus B19 infections are predisposed to myocarditis

  19. Impaired innate, humoral, and cellular immunity despite a take in smallpox vaccine recipients.

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    Kennedy, Richard B; Poland, Gregory A; Ovsyannikova, Inna G; Oberg, Ann L; Asmann, Yan W; Grill, Diane E; Vierkant, Robert A; Jacobson, Robert M

    2016-06-14

    Smallpox vaccine is highly effective, inducing protective immunity to smallpox and diseases caused by related orthopoxviruses. Smallpox vaccine efficacy was historically defined by the appearance of a lesion or "take" at the vaccine site, which leaves behind a characteristic scar. Both the take and scar are readily recognizable and were used during the eradication effort to indicate successful vaccination and to categorize individuals as "protected." However, the development of a typical vaccine take may not equate to the successful development of a robust, protective immune response. In this report, we examined two large (>1000) cohorts of recipients of either Dryvax(®) or ACAM2000 using a testing and replication study design and identified subgroups of individuals who had documented vaccine takes, but who failed to develop robust neutralizing antibody titers. Examination of these individuals revealed that they had suboptimal cellular immune responses as well. Further testing indicated these low responders had a diminished innate antiviral gene expression pattern (IFNA1, CXCL10, CXCL11, OASL) upon in vitro stimulation with vaccinia virus, perhaps indicative of a dysregulated innate response. Our results suggest that poor activation of innate antiviral pathways may result in suboptimal immune responses to the smallpox vaccine. These genes and pathways may serve as suitable targets for adjuvants in new attenuated smallpox vaccines and/or effective antiviral therapy targets against poxvirus infections.

  20. Humoral Immunity to Primary Smallpox Vaccination: Impact of Childhood versus Adult Immunization on Vaccinia Vector Vaccine Development in Military Populations.

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    Slike, Bonnie M; Creegan, Matthew; Marovich, Mary; Ngauy, Viseth

    2017-01-01

    Modified Vaccinia virus has been shown to be a safe and immunogenic vector platform for delivery of HIV vaccines. Use of this vector is of particular importance to the military, with the implementation of a large scale smallpox vaccination campaign in 2002 in active duty and key civilian personnel in response to potential bioterrorist activities. Humoral immunity to smallpox vaccination was previously shown to be long lasting (up to 75 years) and protective. However, using vaccinia-vectored vaccine delivery for other diseases on a background of anti-vector antibodies (i.e. pre-existing immunity) may limit their use as a vaccine platform, especially in the military. In this pilot study, we examined the durability of vaccinia antibody responses in adult primary vaccinees in a healthy military population using a standard ELISA assay and a novel dendritic cell neutralization assay. We found binding and neutralizing antibody (NAb) responses to vaccinia waned after 5-10 years in a group of 475 active duty military, born after 1972, who were vaccinated as adults with Dryvax®. These responses decreased from a geometric mean titer (GMT) of 250 to baseline (vaccination. This contrasted with a comparator group of adults, ages 35-49, who were vaccinated with Dryvax® as children. In the childhood vaccinees, titers persisted for >30 years with a GMT of 210 (range 112-3234). This data suggests limited durability of antibody responses in adult vaccinees compared to those vaccinated in childhood and further that adult vaccinia recipients may benefit similarly from receipt of a vaccinia based vaccine as those who are vaccinia naïve. Our findings may have implications for the smallpox vaccination schedule and support the ongoing development of this promising viral vector in a military vaccination program.

  1. 76 FR 49776 - The Development and Evaluation of Next-Generation Smallpox Vaccines; Public Workshop

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    2011-08-11

    ... cell culture technology under modern manufacturing practices and standards. Both ACAM2000 and Dryvax... response to smallpox vaccines and development of animal models for demonstration of effectiveness of next...-generation smallpox vaccines were prepared on the skin of calves or other animals or in chicken eggs...

  2. Genome Sequence of Vaccinia virus Strain Lister-Butantan, a Lister Vaccine Variant Used during a Smallpox Eradication Campaign in Brazil

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    Assis, Felipe; Trindade, Giliane; Drumond, Betânia; Frace, Mike; Sammons, Scott; Emerson, Ginny; Li, Yu; Carroll, Darin; Batra, Dhwani; Kroon, Erna

    2016-01-01

    Here, we report the 187.8-kb genome sequence of Vaccinia virus Lister-Butantan, which was used in Brazil during the WHO smallpox eradication campaign. Its genome showed an average similarity of 98.18% with the original Lister isolate, highlighting the low divergence among related Vaccinia virus vaccine strains, even after several passages in animals and cell culture. PMID:27340056

  3. Bioluminescent imaging of vaccinia virus infection in immunocompetent and immunodeficient rats as a model for human smallpox

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    Liu, Qiang; Fan, Changfa; Zhou, Shuya; Guo, Yanan; Zuo, Qin; Ma, Jian; Liu, Susu; Wu, Xi; Peng, Zexu; Fan, Tao; Guo, Chaoshe; Shen, Yuelei; Huang, Weijin; Li, Baowen; He, Zhengming; Wang, Youchun

    2015-01-01

    Due to the increasing concern of using smallpox virus as biological weapons for terrorist attack, there is renewed interest in studying the pathogenesis of human smallpox and development of new therapies. Animal models are highly demanded for efficacy and safety examination of new vaccines and therapeutic drugs. Here, we demonstrated that both wild type and immunodeficient rats infected with an engineered vaccinia virus carrying Firefly luciferase reporter gene (rTV-Fluc) could recapitulate infectious and clinical features of human smallpox. Vaccinia viral infection in wild type Sprague-Dawley (SD) rats displayed a diffusible pattern in various organs, including liver, head and limbs. The intensity of bioluminescence generated from rTV-Fluc correlated well with viral loads in tissues. Moreover, neutralizing antibodies had a protective effect against virus reinfection. The recombination activating gene 2 (Rag2) knockout rats generated by transcription activator-like effector nucleases (TALENs) technology were further used to examine the infectivity of the rTV-Fluc in immunodeficient populations. Here we demonstrated that Rag2-/- rats were more susceptible to rTV-Fluc than SD rats with a slower virus clearance rate. Therefore, the rTV-Fluc/SD rats and rTV-Fluc/Rag2-/- rats are suitable visualization models, which recapitulate wild type or immunodeficient populations respectively, for testing human smallpox vaccine and antiviral drugs. PMID:26235050

  4. Safety and immunogenicity of LC16m8, an attenuated smallpox vaccine in vaccinia-naive adults.

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    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; Greenberg, Richard N

    2011-11-01

    LC16m8 is an attenuated cell culture-adapted Lister vaccinia smallpox vaccine missing the B5R protein and licensed for use in Japan. 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)-γ enzyme-linked immunosorbent spot and lymphoproliferation assays. 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 smallpox. Clinical Trials Registration. NCT00103584.

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

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

    2013-01-01

    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.

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

    Directory of Open Access Journals (Sweden)

    Qicheng Zhang

    Full Text Available 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.

  7. Adverse Events Post Smallpox-Vaccination: Insights from Tail Scarification Infection in Mice with Vaccinia virus

    Science.gov (United States)

    Mota, Bruno E. F.; Gallardo-Romero, Nadia; Trindade, Giliane; Keckler, M. Shannon; Karem, Kevin; Carroll, Darin; Campos, Marco A.; Vieira, Leda Q.; da Fonseca, Flávio G.; Ferreira, Paulo C. P.; Bonjardim, Cláudio A.; Damon, Inger K.; Kroon, Erna G.

    2011-01-01

    Adverse events upon smallpox vaccination with fully-replicative strains of Vaccinia virus (VACV) comprise an array of clinical manifestations that occur primarily in immunocompromised patients leading to significant host morbidity/mortality. The expansion of immune-suppressed populations and the possible release of Variola virus as a bioterrorist act have given rise to concerns over vaccination complications should more widespread vaccination be reinitiated. Our goal was to evaluate the components of the host immune system that are sufficient to prevent morbidity/mortality in a murine model of tail scarification, which mimics immunological and clinical features of smallpox vaccination in humans. Infection of C57BL/6 wild-type mice led to a strictly localized infection, with complete viral clearance by day 28 p.i. On the other hand, infection of T and B-cell deficient mice (Rag1−/−) produced a severe disease, with uncontrolled viral replication at the inoculation site and dissemination to internal organs. Infection of B-cell deficient animals (µMT) produced no mortality. However, viral clearance in µMT animals was delayed compared to WT animals, with detectable viral titers in tail and internal organs late in infection. Treatment of Rag1−/− with rabbit hyperimmune anti-vaccinia serum had a subtle effect on the morbidity/mortality of this strain, but it was effective in reduce viral titers in ovaries. Finally, NUDE athymic mice showed a similar outcome of infection as Rag1−/−, and passive transfer of WT T cells to Rag1−/− animals proved fully effective in preventing morbidity/mortality. These results strongly suggest that both T and B cells are important in the immune response to primary VACV infection in mice, and that T-cells are required to control the infection at the inoculation site and providing help for B-cells to produce antibodies, which help to prevent viral dissemination. These insights might prove helpful to better identify individuals

  8. Adverse events post smallpox-vaccination: insights from tail scarification infection in mice with Vaccinia virus.

    Directory of Open Access Journals (Sweden)

    Bruno E F Mota

    Full Text Available Adverse events upon smallpox vaccination with fully-replicative strains of Vaccinia virus (VACV comprise an array of clinical manifestations that occur primarily in immunocompromised patients leading to significant host morbidity/mortality. The expansion of immune-suppressed populations and the possible release of Variola virus as a bioterrorist act have given rise to concerns over vaccination complications should more widespread vaccination be reinitiated. Our goal was to evaluate the components of the host immune system that are sufficient to prevent morbidity/mortality in a murine model of tail scarification, which mimics immunological and clinical features of smallpox vaccination in humans. Infection of C57BL/6 wild-type mice led to a strictly localized infection, with complete viral clearance by day 28 p.i. On the other hand, infection of T and B-cell deficient mice (Rag1(-/- produced a severe disease, with uncontrolled viral replication at the inoculation site and dissemination to internal organs. Infection of B-cell deficient animals (µMT produced no mortality. However, viral clearance in µMT animals was delayed compared to WT animals, with detectable viral titers in tail and internal organs late in infection. Treatment of Rag1(-/- with rabbit hyperimmune anti-vaccinia serum had a subtle effect on the morbidity/mortality of this strain, but it was effective in reduce viral titers in ovaries. Finally, NUDE athymic mice showed a similar outcome of infection as Rag1(-/-, and passive transfer of WT T cells to Rag1(-/- animals proved fully effective in preventing morbidity/mortality. These results strongly suggest that both T and B cells are important in the immune response to primary VACV infection in mice, and that T-cells are required to control the infection at the inoculation site and providing help for B-cells to produce antibodies, which help to prevent viral dissemination. These insights might prove helpful to better identify

  9. Co-administration of the broad-spectrum antiviral, brincidofovir (CMX001), with smallpox vaccine does not compromise vaccine protection in mice challenged with ectromelia virus.

    Science.gov (United States)

    Parker, Scott; Crump, Ryan; Foster, Scott; Hartzler, Hollyce; Hembrador, Ed; Lanier, E Randall; Painter, George; Schriewer, Jill; Trost, Lawrence C; Buller, R Mark

    2014-11-01

    Natural orthopoxvirus outbreaks such as vaccinia, cowpox, cattlepox and buffalopox continue to cause morbidity in the human population. Monkeypox virus remains a significant agent of morbidity and mortality in Africa. Furthermore, monkeypox virus's broad host-range and expanding environs make it of particular concern as an emerging human pathogen. Monkeypox virus and variola virus (the etiological agent of smallpox) are both potential agents of bioterrorism. The first line response to orthopoxvirus disease is through vaccination with first-generation and second-generation vaccines, such as Dryvax and ACAM2000. Although these vaccines provide excellent protection, their widespread use is impeded by the high level of adverse events associated with vaccination using live, attenuated virus. It is possible that vaccines could be used in combination with antiviral drugs to reduce the incidence and severity of vaccine-associated adverse events, or as a preventive in individuals with uncertain exposure status or contraindication to vaccination. We have used the intranasal mousepox (ectromelia) model to evaluate the efficacy of vaccination with Dryvax or ACAM2000 in conjunction with treatment using the broad spectrum antiviral, brincidofovir (BCV, CMX001). We found that co-treatment with BCV reduced the severity of vaccination-associated lesion development. Although the immune response to vaccination was quantifiably attenuated, vaccination combined with BCV treatment did not alter the development of full protective immunity, even when administered two days following ectromelia challenge. Studies with a non-replicating vaccine, ACAM3000 (MVA), confirmed that BCV's mechanism of attenuating the immune response following vaccination with live virus was, as expected, by limiting viral replication and not through inhibition of the immune system. These studies suggest that, in the setting of post-exposure prophylaxis, co-administration of BCV with vaccination should be considered

  10. Extent of Systemic Spread Determines CD8+ T Cell Immunodominance for Laboratory Strains, Smallpox Vaccines, and Zoonotic Isolates of Vaccinia Virus.

    Science.gov (United States)

    Flesch, Inge E A; Hollett, Natasha A; Wong, Yik Chun; Quinan, Bárbara Resende; Howard, Debbie; da Fonseca, Flávio G; Tscharke, David C

    2015-09-01

    CD8(+) T cells that recognize virus-derived peptides presented on MHC class I are vital antiviral effectors. Such peptides presented by any given virus vary greatly in immunogenicity, allowing them to be ranked in an immunodominance hierarchy. However, the full range of parameters that determine immunodominance and the underlying mechanisms remain unknown. In this study, we show across a range of vaccinia virus strains, including the current clonal smallpox vaccine, that the ability of a strain to spread systemically correlated with reduced immunodominance. Reduction in immunodominance was observed both in the lymphoid system and at the primary site of infection. Mechanistically, reduced immunodominance was associated with more robust priming and especially priming in the spleen. Finally, we show this is not just a property of vaccine and laboratory strains of virus, because an association between virulence and immunodominance was also observed in isolates from an outbreak of zoonotic vaccinia virus that occurred in Brazil.

  11. Mucosal Immunization Induces a Higher Level of Lasting Neutralizing Antibody Response in Mice by a Replication-Competent Smallpox Vaccine: Vaccinia Tiantan Strain

    Directory of Open Access Journals (Sweden)

    Bin Lu

    2011-01-01

    Full Text Available The possible bioterrorism threat using the variola virus, the causative agent of smallpox, has promoted us to further investigate the immunogenicity profiles of existing vaccines. Here, we study for the first time the immunogenicity profile of a replication-competent smallpox vaccine (vaccinia Tiantan, VTT strain for inducing neutralizing antibodies (Nabs through mucosal vaccination, which is noninvasive and has a critical implication for massive vaccination programs. Four different routes of vaccination were tested in parallel including intramuscular (i.m., intranasal (i.n., oral (i.o., and subcutaneous (s.c. inoculations in mice. We found that one time vaccination with an optimal dose of VTT was able to induce anti-VTT Nabs via each of the four routes. Higher levels of antiviral Nabs, however, were induced via the i.n. and i.o. inoculations when compared with the i.m. and s.c. routes. Moreover, the i.n. and i.o. vaccinations also induced higher sustained levels of Nabs overtime, which conferred better protections against homologous or alternating mucosal routes of viral challenges six months post vaccination. The VTT-induced immunity via all four routes, however, was partially effective against the intramuscular viral challenge. Our data have implications for understanding the potential application of mucosal smallpox vaccination and for developing VTT-based vaccines to overcome preexisting antivaccinia immunity.

  12. Mucosal immunization induces a higher level of lasting neutralizing antibody response in mice by a replication-competent smallpox vaccine: vaccinia Tiantan strain.

    Science.gov (United States)

    Lu, Bin; Yu, Wenbo; Huang, Xiaoxing; Wang, Haibo; Liu, Li; Chen, Zhiwei

    2011-01-01

    The possible bioterrorism threat using the variola virus, the causative agent of smallpox, has promoted us to further investigate the immunogenicity profiles of existing vaccines. Here, we study for the first time the immunogenicity profile of a replication-competent smallpox vaccine (vaccinia Tiantan, VTT strain) for inducing neutralizing antibodies (Nabs) through mucosal vaccination, which is noninvasive and has a critical implication for massive vaccination programs. Four different routes of vaccination were tested in parallel including intramuscular (i.m.), intranasal (i.n.), oral (i.o.), and subcutaneous (s.c.) inoculations in mice. We found that one time vaccination with an optimal dose of VTT was able to induce anti-VTT Nabs via each of the four routes. Higher levels of antiviral Nabs, however, were induced via the i.n. and i.o. inoculations when compared with the i.m. and s.c. routes. Moreover, the i.n. and i.o. vaccinations also induced higher sustained levels of Nabs overtime, which conferred better protections against homologous or alternating mucosal routes of viral challenges six months post vaccination. The VTT-induced immunity via all four routes, however, was partially effective against the intramuscular viral challenge. Our data have implications for understanding the potential application of mucosal smallpox vaccination and for developing VTT-based vaccines to overcome preexisting antivaccinia immunity.

  13. The strategic use of novel smallpox vaccines in the post-eradication world.

    Science.gov (United States)

    Golden, Joseph W; Hooper, Jay W

    2011-07-01

    We still face a threat of orthopoxviruses in the form of biological weapons and emerging zoonoses. Therefore, there is a need to maintain a comprehensive defense strategy to counter the low-probability, high-impact threat of smallpox, as well as the ongoing threat of naturally occurring orthopoxvirus disease. The currently licensed live-virus smallpox vaccine ACAM2000 is effective, but associated with serious and even life-threatening adverse events. The health threat posed by this vaccine, and other previously licensed vaccines, has prevented many first responders, and even many in the military, from receiving a vaccine against smallpox. At the same time, global immunity produced during the smallpox eradication campaign is waning. Here, we review novel subunit/component vaccines and how they might play roles in unconventional strategies to defend against emerging orthopoxvirus diseases throughout the world and against smallpox used as a weapon of mass destruction.

  14. 42 CFR 102.54 - Documentation the representative of the estate of a deceased smallpox vaccine recipient or...

    Science.gov (United States)

    2010-10-01

    ... deceased smallpox vaccine recipient or vaccinia contact must submit to be deemed eligible by the Secretary... VACCINES SMALLPOX COMPENSATION PROGRAM Required Documentation To Be Deemed Eligible § 102.54 Documentation the representative of the estate of a deceased smallpox vaccine recipient or vaccinia contact...

  15. A Randomized, Double-Blind, Placebo-Controlled Phase II Trial Investigating the Safety and Immunogenicity of Modified Vaccinia Ankara Smallpox Vaccine (MVA-BN® in 56-80-Year-Old Subjects.

    Directory of Open Access Journals (Sweden)

    Richard N Greenberg

    Full Text Available Modified Vaccinia Ankara MVA-BN® is a live, highly attenuated, viral vaccine under advanced development as a non-replicating smallpox vaccine. In this Phase II trial, the safety and immunogenicity of Modified Vaccinia Ankara MVA-BN® (MVA was assessed in a 56-80 years old population.MVA with a virus titer of 1 x 108 TCID50/dose was administered via subcutaneous injection to 56-80 year old vaccinia-experienced subjects (N = 120. Subjects received either two injections of MVA (MM group or one injection of Placebo and one injection of MVA (PM group four weeks apart. Safety was evaluated by assessment of adverse events (AE, focused physical exams, electrocardiogram recordings and safety laboratories. Solicited AEs consisted of a set of pre-defined expected local reactions (erythema, swelling, pain, pruritus, and induration and systemic symptoms (body temperature, headache, myalgia, nausea and fatigue and were recorded on a memory aid for an 8-day period following each injection. The immunogenicity of the vaccine was evaluated in terms of humoral immune responses measured with a vaccinia-specific enzyme-linked immunosorbent assay (ELISA and a plaque reduction neutralization test (PRNT before and at different time points after vaccination.Vaccinations were well tolerated by all subjects. No serious adverse event related to MVA and no case of myopericarditis was reported. The overall incidence of unsolicited AEs was similar in both groups. For both groups immunogenicity responses two weeks after the final vaccination (i.e. Visit 4 were as follows: Seroconversion (SC rates (doubling of titers from baseline in vaccine specific antibody titers measured by ELISA were 83.3% in Group MM and 82.8% in Group PM (difference 0.6% with 95% exact CI [-13.8%, 15.0%], and 90.0% for Group MM and 77.6% for Group PM measured by PRNT (difference 12.4% with 95% CI of [-1.1%, 27.0%]. Geometric mean titers (GMT measured by ELISA two weeks after the final vaccination for

  16. A Randomized, Double-Blind, Placebo-Controlled Phase II Trial Investigating the Safety and Immunogenicity of Modified Vaccinia Ankara Smallpox Vaccine (MVA-BN®) in 56-80-Year-Old Subjects

    Science.gov (United States)

    Greenberg, Richard N.; Hay, Christine M.; Stapleton, Jack T.; Marbury, Thomas C.; Wagner, Eva; Kreitmeir, Eva; von Krempelhuber, Alfred; Young, Philip; Nichols, Richard; Meyer, Thomas P.; Weigl, Josef; Virgin, Garth; Arndtz-Wiedemann, Nathaly; Chaplin, Paul

    2016-01-01

    Background Modified Vaccinia Ankara MVA-BN® is a live, highly attenuated, viral vaccine under advanced development as a non-replicating smallpox vaccine. In this Phase II trial, the safety and immunogenicity of Modified Vaccinia Ankara MVA-BN® (MVA) was assessed in a 56–80 years old population. Methods MVA with a virus titer of 1 x 108 TCID50/dose was administered via subcutaneous injection to 56–80 year old vaccinia-experienced subjects (N = 120). Subjects received either two injections of MVA (MM group) or one injection of Placebo and one injection of MVA (PM group) four weeks apart. Safety was evaluated by assessment of adverse events (AE), focused physical exams, electrocardiogram recordings and safety laboratories. Solicited AEs consisted of a set of pre-defined expected local reactions (erythema, swelling, pain, pruritus, and induration) and systemic symptoms (body temperature, headache, myalgia, nausea and fatigue) and were recorded on a memory aid for an 8-day period following each injection. The immunogenicity of the vaccine was evaluated in terms of humoral immune responses measured with a vaccinia-specific enzyme-linked immunosorbent assay (ELISA) and a plaque reduction neutralization test (PRNT) before and at different time points after vaccination. Results Vaccinations were well tolerated by all subjects. No serious adverse event related to MVA and no case of myopericarditis was reported. The overall incidence of unsolicited AEs was similar in both groups. For both groups immunogenicity responses two weeks after the final vaccination (i.e. Visit 4) were as follows: Seroconversion (SC) rates (doubling of titers from baseline) in vaccine specific antibody titers measured by ELISA were 83.3% in Group MM and 82.8% in Group PM (difference 0.6% with 95% exact CI [-13.8%, 15.0%]), and 90.0% for Group MM and 77.6% for Group PM measured by PRNT (difference 12.4% with 95% CI of [-1.1%, 27.0%]). Geometric mean titers (GMT) measured by ELISA two weeks after

  17. Vaccinia virus infections in martial arts gym, Maryland, USA, 2008.

    Science.gov (United States)

    Hughes, Christine M; Blythe, David; Li, Yu; Reddy, Ramani; Jordan, Carol; Edwards, Cindy; Adams, Celia; Conners, Holly; Rasa, Catherine; Wilby, Sue; Russell, Jamaal; Russo, Kelly S; Somsel, Patricia; Wiedbrauk, Danny L; Dougherty, Cindy; Allen, Christopher; Frace, Mike; Emerson, Ginny; Olson, Victoria A; Smith, Scott K; Braden, Zachary; Abel, Jason; Davidson, Whitni; Reynolds, Mary; Damon, Inger K

    2011-04-01

    Vaccinia virus is an orthopoxvirus used in the live vaccine against smallpox. Vaccinia virus infections can be transmissible and can cause severe complications in those with weakened immune systems. We report on a cluster of 4 cases of vaccinia virus infection in Maryland, USA, likely acquired at a martial arts gym.

  18. Third-generation smallpox vaccines: challenges in the absence of clinical smallpox.

    Science.gov (United States)

    Meseda, Clement A; Weir, Jerry P

    2010-09-01

    Smallpox, a disease caused by variola virus, is estimated to have killed hundreds of millions to billions of people before it was certified as eradicated in 1980. However, there has been renewed interest in smallpox vaccine development due in part to zoonotic poxvirus infections and the possibility of a re-emergence of smallpox, as well as the fact that first-generation smallpox vaccines are associated with relatively rare, but severe, adverse reactions in some vaccinees. An understanding of the immune mechanisms of vaccine protection and the use of suitable animal models for vaccine efficacy assessment are paramount to the development of safer and effective smallpox vaccines. This article focuses on studies aimed at understanding the immune responses elicited by vaccinia virus and the various animal models that can be used to evaluate smallpox vaccine efficacy. Harnessing this information is necessary to assess the effectiveness and potential usefulness of new-generation smallpox vaccines.

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

    NARCIS (Netherlands)

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

    2006-01-01

    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 ab

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

    NARCIS (Netherlands)

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

    2006-01-01

    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 ab

  1. Smallpox: clinical highlights and considerations for vaccination.

    Directory of Open Access Journals (Sweden)

    Mahoney M

    2003-01-01

    Full Text Available Smallpox virus has gained considerable attention as a potential bioterrorism agent. Recommendations for smallpox (vaccinia vaccination presume a low risk for use of smallpox as a terrorist biological agent and vaccination is currently recommended for selected groups of individuals such as health care workers, public health authorities, and emergency/rescue workers, among others. Information about adverse reactions to the smallpox vaccine is based upon studies completed during the 1950s and 1960s. The prevalence of various diseases has changed over the last four decades and new disease entities have been described during this period. The smallpox vaccination may be contra-indicated in many of these conditions. This has made pre-screening of potential vaccines necessary. It is believed that at present, the risks of vaccine-associated complications far outweigh the potential benefits of vaccination in the general population.

  2. Smallpox vaccines: targets of protective immunity.

    Science.gov (United States)

    Moss, Bernard

    2011-01-01

    The eradication of smallpox, one of the great triumphs of medicine, was accomplished through the prophylactic administration of live vaccinia virus, a comparatively benign relative of variola virus, the causative agent of smallpox. Nevertheless, recent fears that variola virus may be used as a biological weapon together with the present susceptibility of unimmunized populations have spurred the development of new-generation vaccines that are safer than the original and can be produced by modern methods. Predicting the efficacy of such vaccines in the absence of human smallpox, however, depends on understanding the correlates of protection. This review outlines the biology of poxviruses with particular relevance to vaccine development, describes protein targets of humoral and cellular immunity, compares animal models of orthopoxvirus disease with human smallpox, and considers the status of second- and third-generation smallpox vaccines.

  3. The immunology of smallpox vaccines.

    Science.gov (United States)

    Kennedy, Richard B; Ovsyannikova, Inna G; Jacobson, Robert M; Poland, Gregory A

    2009-06-01

    In spite of the eradication of smallpox over 30 years ago; orthopox viruses such as smallpox and monkeypox remain serious public health threats both through the possibility of bioterrorism and the intentional release of smallpox and through natural outbreaks of emerging infectious diseases such as monkeypox. The eradication effort was largely made possible by the availability of an effective vaccine based on the immunologically cross-protective vaccinia virus. Although the concept of vaccination dates back to the late 1800s with Edward Jenner, it is only in the past decade that modern immunologic tools have been applied toward deciphering poxvirus immunity. Smallpox vaccines containing vaccinia virus elicit strong humoral and cellular immune responses that confer cross-protective immunity against variola virus for decades after immunization. Recent studies have focused on: establishing the longevity of poxvirus-specific immunity, defining key immune epitopes targeted by T and B cells, developing subunit-based vaccines, and developing genotypic and phenotypic immune response profiles that predict either vaccine response or adverse events following immunization.

  4. Complete Genome Sequence of Vaccinia Virus Strain L-IVP

    Science.gov (United States)

    Shvalov, Alexander N.; Sivolobova, Galina F.; Kuligina, Elena V.

    2016-01-01

    Most of the live vaccine doses of vaccinia virus donated to the Intensified Smallpox Eradication Programme after 1971 were prepared using the L-IVP strain. A mixture of three clones of the L-IVP strain was sequenced using MySEQ. Consensus sequence similarity with the vaccinia virus Lister strain is 99.5%. PMID:27174282

  5. Vaccine Basics (Smallpox)

    Science.gov (United States)

    ... Search Form Controls Cancel Submit Search the CDC Smallpox Note: Javascript is disabled or is not supported ... message, please visit this page: About CDC.gov . Smallpox About Smallpox History of Smallpox Spread and Eradication ...

  6. Signs and Symptoms (Smallpox)

    Science.gov (United States)

    ... Search Form Controls Cancel Submit Search the CDC Smallpox Note: Javascript is disabled or is not supported ... message, please visit this page: About CDC.gov . Smallpox About Smallpox History of Smallpox Spread and Eradication ...

  7. Recent advances in the study of live attenuated cell-cultured smallpox vaccine LC16m8.

    Science.gov (United States)

    Eto, Akiko; Saito, Tomoya; Yokote, Hiroyuki; Kurane, Ichiro; Kanatani, Yasuhiro

    2015-11-01

    LC16m8 is a live, attenuated, cell-cultured smallpox vaccine that was developed and licensed in Japan in the 1970s, but was not used in the campaign to eradicate smallpox. In the early 2000s, the potential threat of bioterrorism led to reconsideration of the need for a smallpox vaccine. Subsequently, LC16m8 production was restarted in Japan in 2002, requiring re-evaluation of its safety and efficacy. Approximately 50,000 children in the 1970s and about 3500 healthy adults in the 2000s were vaccinated with LC16m8 in Japan, and 153 adults have been vaccinated with LC16m8 or Dryvax in phase I/II clinical trials in the USA. These studies confirmed the safety and efficacy of LC16m8, while several studies in animal models have shown that LC16m8 protects the host against viral challenge. The World Health Organization Strategic Advisory Group of Experts on Immunization recommended LC16m8, together with ACAM2000, as a stockpile vaccine in 2013. In addition, LC16m8 is expected to be a viable alternative to first-generation smallpox vaccines to prevent human monkeypox.

  8. What is Smallpox?

    Science.gov (United States)

    ... this page: About CDC.gov . Smallpox About Smallpox History of Smallpox Spread and Eradication of Smallpox Transmission Signs and Symptoms Prevention and Treatment Smallpox Vaccine Basics Vaccine Safety Side Effects of Vaccination Who Should Get a Smallpox Vaccination? Bioterrorism The ...

  9. Side-by-side comparison of gene-based smallpox vaccine with MVA in nonhuman primates.

    Directory of Open Access Journals (Sweden)

    Joseph W Golden

    Full Text Available Orthopoxviruses remain a threat as biological weapons and zoonoses. The licensed live-virus vaccine is associated with serious health risks, making its general usage unacceptable. Attenuated vaccines are being developed as alternatives, the most advanced of which is modified-vaccinia virus Ankara (MVA. We previously developed a gene-based vaccine, termed 4pox, which targets four orthopoxvirus antigens, A33, B5, A27 and L1. This vaccine protects mice and non-human primates from lethal orthopoxvirus disease. Here, we investigated the capacity of the molecular adjuvants GM-CSF and Escherichia coli heat-labile enterotoxin (LT to enhance the efficacy of the 4pox gene-based vaccine. Both adjuvants significantly increased protective antibody responses in mice. We directly compared the 4pox plus LT vaccine against MVA in a monkeypox virus (MPXV nonhuman primate (NHP challenge model. NHPs were vaccinated twice with MVA by intramuscular injection or the 4pox/LT vaccine delivered using a disposable gene gun device. As a positive control, one NHP was vaccinated with ACAM2000. NHPs vaccinated with each vaccine developed anti-orthopoxvirus antibody responses, including those against the 4pox antigens. After MPXV intravenous challenge, all control NHPs developed severe disease, while the ACAM2000 vaccinated animal was well protected. All NHPs vaccinated with MVA were protected from lethality, but three of five developed severe disease and all animals shed virus. All five NHPs vaccinated with 4pox/LT survived and only one developed severe disease. None of the 4pox/LT-vaccinated animals shed virus. Our findings show, for the first time, that a subunit orthopoxvirus vaccine delivered by the same schedule can provide a degree of protection at least as high as that of MVA.

  10. Side-by-Side Comparison of Gene-Based Smallpox Vaccine with MVA in Nonhuman Primates

    Science.gov (United States)

    Golden, Joseph W.; Josleyn, Matthew; Mucker, Eric M.; Hung, Chien-Fu; Loudon, Peter T.; Wu, T. C.; Hooper, Jay W.

    2012-01-01

    Orthopoxviruses remain a threat as biological weapons and zoonoses. The licensed live-virus vaccine is associated with serious health risks, making its general usage unacceptable. Attenuated vaccines are being developed as alternatives, the most advanced of which is modified-vaccinia virus Ankara (MVA). We previously developed a gene-based vaccine, termed 4pox, which targets four orthopoxvirus antigens, A33, B5, A27 and L1. This vaccine protects mice and non-human primates from lethal orthopoxvirus disease. Here, we investigated the capacity of the molecular adjuvants GM-CSF and Escherichia coli heat-labile enterotoxin (LT) to enhance the efficacy of the 4pox gene-based vaccine. Both adjuvants significantly increased protective antibody responses in mice. We directly compared the 4pox plus LT vaccine against MVA in a monkeypox virus (MPXV) nonhuman primate (NHP) challenge model. NHPs were vaccinated twice with MVA by intramuscular injection or the 4pox/LT vaccine delivered using a disposable gene gun device. As a positive control, one NHP was vaccinated with ACAM2000. NHPs vaccinated with each vaccine developed anti-orthopoxvirus antibody responses, including those against the 4pox antigens. After MPXV intravenous challenge, all control NHPs developed severe disease, while the ACAM2000 vaccinated animal was well protected. All NHPs vaccinated with MVA were protected from lethality, but three of five developed severe disease and all animals shed virus. All five NHPs vaccinated with 4pox/LT survived and only one developed severe disease. None of the 4pox/LT-vaccinated animals shed virus. Our findings show, for the first time, that a subunit orthopoxvirus vaccine delivered by the same schedule can provide a degree of protection at least as high as that of MVA. PMID:22860117

  11. Smallpox vaccination and bioterrorism with pox viruses.

    Science.gov (United States)

    Mayr, Anton

    2003-10-01

    Bioterrorist attacks occupy a special place amongst the innumerable potential types of terrorist attack, with the intentional release of pox viruses being especially feared in this connection. Apart from the variola virus, the agent responsible for smallpox in humans, the monkeypox virus and numerous other animal pox viruses pose potential risks for humans and animals. This risk scenario also includes recombinations between the various pox viruses, changes in hosts and genetically engineered manipulations of pox viruses. For over 200 years, the method of choice for combatting smallpox was via vaccination with a reproductive, original vaccinia virus. Worldwide eradication of smallpox at the end of the 1970s and the discontinuation of routine smallpox vaccination in 1980 can be credited to such vaccination. Unfortunately, these vaccinations were associated with a large number of postvaccinal impairments, sometimes resulting in death (e.g. postvaccinal encephalitis). The only way to restrict such postvaccinal complications was to carry out initial vaccination within the first 2 postnatal years. Initial vaccination at a later age led to such a sharp increase in the number of vaccines with complications that vaccination had to be discouraged. The dilemma of the smallpox vaccine stocks stems from the fact that a large portion of these stocks are produced with the same vaccinia strains as before. This is irresponsible, especially as the percentage of immune-suppressed persons in the population, for whom vaccination-related complications pose an especial threat, is increasing. One solution to the dilemma of the smallpox vaccine stocks is the MVA strain. It is harmless, protects humans and animals equally well against smallpox and can be applied parenterally.

  12. Smallpox vaccine: the good, the bad, and the ugly.

    Science.gov (United States)

    Belongia, Edward A; Naleway, Allison L

    2003-04-01

    Smallpox inarguably shaped the course of human history by killing countless millions in both the Old World and the New World. Dr. Edward Jenner's discovery of vaccination in the late 18th century, and the global eradication of smallpox in the 1970s, rank among the greatest achievements in human history. Amidst recent growing concerns about bioterrorism, smallpox vaccination has resurfaced from the history books to become a topic of major importance. Inoculation with vaccinia virus is highly effective for the prevention of smallpox infection, but it is associated with several known side effects that range from mild and self-limited to severe and life-threatening. As the United States moves forward with plans to vaccinate selected health care workers and the military, and perhaps offer the vaccination to all citizens in the future, it is important to fully understand and appreciate the history, risks, and benefits of smallpox vaccination.

  13. Modified Vaccinia Ankara Virus Vaccination Provides Long-Term Protection against Nasal Rabbitpox Virus Challenge

    Science.gov (United States)

    Jones, Dorothy I.; McGee, Charles E.; Sample, Christopher J.; Sempowski, Gregory D.; Pickup, David J.

    2016-01-01

    Modified vaccinia Ankara virus (MVA) is a smallpox vaccine candidate. This study was performed to determine if MVA vaccination provides long-term protection against rabbitpox virus (RPXV) challenge, an animal model of smallpox. Two doses of MVA provided 100% protection against a lethal intranasal RPXV challenge administered 9 months after vaccination. PMID:27146001

  14. Modified vaccinia virus Ankara protects macaques against respiratory challenge with monkeypox virus.

    NARCIS (Netherlands)

    K.J. Stittelaar (Koert); G. van Amerongen (Geert); I. Kondova (Ivanela); R.F. van Lavieren (Rob); F.H. Pistoor (Frank); H.G.M. Niesters (Bert); G.J.J. van Doornum (Gerard); B.A.M. van der Zeijst (Ben); L. Mateo (Luis); P.J. Chaplin (Paul); A.D.M.E. Osterhaus (Albert); T. Kuiken (Thijs)

    2005-01-01

    textabstractThe use of classical smallpox vaccines based on vaccinia virus (VV) is associated with severe complications in both naive and immune individuals. Modified vaccinia virus Ankara (MVA), a highly attenuated replication-deficient strain of VV, has been proven to be safe in humans and immunoc

  15. Modified vaccinia virus Ankara protects macaques against respiratory challenge with monkeypox virus

    NARCIS (Netherlands)

    Stittelaar, Koert J; van Amerongen, Geert; Kondova, Ivanela; Kuiken, Thijs; van Lavieren, Rob F; Pistoor, Frank H M; Niesters, Hubert G M; van Doornum, Gerard; van der Zeijst, Ben A M; Mateo, Luis; Chaplin, Paul J; Osterhaus, Albert D M E

    2005-01-01

    The use of classical smallpox vaccines based on vaccinia virus (VV) is associated with severe complications in both naive and immune individuals. Modified vaccinia virus Ankara (MVA), a highly attenuated replication-deficient strain of VV, has been proven to be safe in humans and immunocompromised a

  16. Modified vaccinia virus Ankara protects macaques against respiratory challenge with monkeypox virus

    NARCIS (Netherlands)

    Stittelaar, Koert J; van Amerongen, Geert; Kondova, Ivanela; Kuiken, Thijs; van Lavieren, Rob F; Pistoor, Frank H M; Niesters, Hubert G M; van Doornum, Gerard; van der Zeijst, Ben A M; Mateo, Luis; Chaplin, Paul J; Osterhaus, Albert D M E

    2005-01-01

    The use of classical smallpox vaccines based on vaccinia virus (VV) is associated with severe complications in both naive and immune individuals. Modified vaccinia virus Ankara (MVA), a highly attenuated replication-deficient strain of VV, has been proven to be safe in humans and immunocompromised a

  17. Brazilian Vaccinia Viruses and Their Origins

    Centers for Disease Control (CDC) Podcasts

    2007-07-30

    Smallpox was eradicated more than 25 years ago, but live viruses used in vaccines may have survived to cause animal and human illness today. Dr. Inger Damon, Acting Branch Chief of the Poxvirus and Rabies Branch at CDC, discusses efforts to determine origins and spread of vaccinia viruses in Brazil.  Created: 7/30/2007 by Emerging Infectious Diseases.   Date Released: 7/30/2007.

  18. Vaccinia Virus: A Tool for Research and Vaccine Development

    Science.gov (United States)

    Moss, Bernard

    1991-06-01

    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.

  19. Should remaining stockpiles of smallpox virus (variola) be destroyed?

    Science.gov (United States)

    Weinstein, Raymond S

    2011-04-01

    In 2011, the World Health Organization will recommend the fate of existing smallpox stockpiles, but circumstances have changed since the complete destruction of these cultures was first proposed. Recent studies suggest that variola and its experimental surrogate, vaccinia, have a remarkable ability to modify the human immune response through complex mechanisms that scientists are only just beginning to unravel. Further study that might require intact virus is essential. Moreover, modern science now has the capability to recreate smallpox or a smallpox-like organism in the laboratory in addition to the risk of nature re-creating it as it did once before. These factors strongly suggest that relegating smallpox to the autoclave of extinction would be ill advised.

  20. Immunomodulator-based enhancement of anti smallpox immune responses.

    Science.gov (United States)

    Martínez, Osmarie; Miranda, Eric; Ramírez, Maite; Santos, Saritza; Rivera, Carlos; Vázquez, Luis; Sánchez, Tomás; Tremblay, Raymond L; Ríos-Olivares, Eddy; Otero, Miguel

    2015-01-01

    The current live vaccinia virus vaccine used in the prevention of smallpox is contraindicated for millions of immune-compromised individuals. Although vaccination with the current smallpox vaccine produces protective immunity, it might result in mild to serious health complications for some vaccinees. Thus, there is a critical need for the production of a safe virus-free vaccine against smallpox that is available to everyone. For that reason, we investigated the impact of imiquimod and resiquimod (Toll-like receptors agonists), and the codon-usage optimization of the vaccinia virus A27L gene in the enhancement of the immune response, with intent of producing a safe, virus-free DNA vaccine coding for the A27 vaccinia virus protein. We analyzed the cellular-immune response by measuring the IFN-γ production of splenocytes by ELISPOT, the humoral-immune responses measuring total IgG and IgG2a/IgG1 ratios by ELISA, and the TH1 and TH2 cytokine profiles by ELISA, in mice immunized with our vaccine formulation. The proposed vaccine formulation enhanced the A27L vaccine-mediated production of IFN-γ on mouse spleens, and increased the humoral immunity with a TH1-biased response. Also, our vaccine induced a TH1 cytokine milieu, which is important against viral infections. These results support the efforts to find a new mechanism to enhance an immune response against smallpox, through the implementation of a safe, virus-free DNA vaccination platform.

  1. Smallpox vaccine revisited.

    Science.gov (United States)

    Capriotti, Teri

    2002-12-01

    Smallpox is a serious contagious disease which is back in the public eye. Yet, most health care providers are unprepared for its return. Nurses will be key health care professionals in a smallpox outbreak or vaccination program.

  2. Evaluation of smallpox vaccines using variola neutralization.

    Science.gov (United States)

    Damon, Inger K; Davidson, Whitni B; Hughes, Christine M; Olson, Victoria A; Smith, Scott K; Holman, Robert C; Frey, Sharon E; Newman, Frances; Belshe, Robert B; Yan, Lihan; Karem, Kevin

    2009-08-01

    The search for a 'third'-generation smallpox vaccine has resulted in the development and characterization of several vaccine candidates. A significant barrier to acceptance is the absence of challenge models showing induction of correlates of protective immunity against variola virus. In this light, virus neutralization provides one of few experimental methods to show specific 'in vitro' activity of vaccines against variola virus. Here, we provide characterization of the ability of a modified vaccinia virus Ankara vaccine to induce variola virus-neutralizing antibodies, and we provide comparison with the neutralization elicited by standard Dryvax vaccination.

  3. Vaccinations against smallpox and tuberculosis are associated with better long-term survival

    DEFF Research Database (Denmark)

    Rieckmann, Andreas; Villumsen, Marie; Sørup, Signe;

    2016-01-01

    BACKGROUND: When vaccinations with vaccinia against smallpox and Bacillus Calmette-Guérin (BCG) against tuberculosis were phased out in some high-income countries around 1980, the impact on overall mortality was not examined. Recent studies from low-income countries have suggested that these vacc......BACKGROUND: When vaccinations with vaccinia against smallpox and Bacillus Calmette-Guérin (BCG) against tuberculosis were phased out in some high-income countries around 1980, the impact on overall mortality was not examined. Recent studies from low-income countries have suggested...

  4. Antiviral immunity following smallpox virus infection: a case-control study.

    Science.gov (United States)

    Hammarlund, Erika; Lewis, Matthew W; Hanifin, Jon M; Mori, Motomi; Koudelka, Caroline W; Slifka, Mark K

    2010-12-01

    Outbreaks of smallpox (i.e., caused by variola virus) resulted in up to 30% mortality, but those who survived smallpox infection were regarded as immune for life. Early studies described the levels of neutralizing antibodies induced after infection, but smallpox was eradicated before contemporary methods for quantifying T-cell memory were developed. To better understand the levels and duration of immunity after smallpox infection, we performed a case-control study comparing antiviral CD4(+) and CD8(+) T-cell responses and neutralizing antibody levels of 24 smallpox survivors with the antiviral immunity observed in 60 smallpox-vaccinated (i.e., vaccinia virus-immune) control subjects. We found that the duration of immunity following smallpox infection was remarkably similar to that observed after smallpox vaccination, with antiviral T-cell responses that declined slowly over time and antiviral antibody responses that remained stable for decades after recovery from infection. These results indicate that severe, potentially life-threatening disease is not required for the development of sustainable long-term immunity. This study shows that the levels of immunity induced following smallpox vaccination are comparable in magnitude to that achieved through natural variola virus infection, and this may explain the notable success of vaccination in eradicating smallpox, one of the world's most lethal diseases.

  5. Antiviral Immunity following Smallpox Virus Infection: a Case-Control Study▿

    Science.gov (United States)

    Hammarlund, Erika; Lewis, Matthew W.; Hanifin, Jon M.; Mori, Motomi; Koudelka, Caroline W.; Slifka, Mark K.

    2010-01-01

    Outbreaks of smallpox (i.e., caused by variola virus) resulted in up to 30% mortality, but those who survived smallpox infection were regarded as immune for life. Early studies described the levels of neutralizing antibodies induced after infection, but smallpox was eradicated before contemporary methods for quantifying T-cell memory were developed. To better understand the levels and duration of immunity after smallpox infection, we performed a case-control study comparing antiviral CD4+ and CD8+ T-cell responses and neutralizing antibody levels of 24 smallpox survivors with the antiviral immunity observed in 60 smallpox-vaccinated (i.e., vaccinia virus-immune) control subjects. We found that the duration of immunity following smallpox infection was remarkably similar to that observed after smallpox vaccination, with antiviral T-cell responses that declined slowly over time and antiviral antibody responses that remained stable for decades after recovery from infection. These results indicate that severe, potentially life-threatening disease is not required for the development of sustainable long-term immunity. This study shows that the levels of immunity induced following smallpox vaccination are comparable in magnitude to that achieved through natural variola virus infection, and this may explain the notable success of vaccination in eradicating smallpox, one of the world's most lethal diseases. PMID:20926574

  6. Frequency of adverse events after vaccination with different vaccinia strains.

    Directory of Open Access Journals (Sweden)

    Mirjam Kretzschmar

    2006-08-01

    Full Text Available BACKGROUND: Large quantities of smallpox vaccine have been stockpiled to protect entire nations against a possible reintroduction of smallpox. Planning for an appropriate use of these stockpiled vaccines in response to a smallpox outbreak requires a rational assessment of the risks of vaccination-related adverse events, compared to the risk of contracting an infection. Although considerable effort has been made to understand the dynamics of smallpox transmission in modern societies, little attention has been paid to estimating the frequency of adverse events due to smallpox vaccination. Studies exploring the consequences of smallpox vaccination strategies have commonly used a frequency of approximately one death per million vaccinations, which is based on a study of vaccination with the New York City Board of Health (NYCBH strain of vaccinia virus. However, a multitude of historical studies of smallpox vaccination with other vaccinia strains suggest that there are strain-related differences in the frequency of adverse events after vaccination. Because many countries have stockpiled vaccine based on the Lister strain of vaccinia virus, a quantitative evaluation of the adverse effects of such vaccines is essential for emergency response planning. We conducted a systematic review and statistical analysis of historical data concerning vaccination against smallpox with different strains of vaccinia virus. METHODS AND FINDINGS: We analyzed historical vaccination data extracted from the literature. We extracted data on the frequency of postvaccinal encephalitis and death with respect to vaccinia strain and age of vaccinees. Using a hierarchical Bayesian approach for meta-analysis, we estimated the expected frequencies of postvaccinal encephalitis and death with respect to age at vaccination for smallpox vaccines based on the NYCBH and Lister vaccinia strains. We found large heterogeneity between findings from different studies and a time-period effect

  7. Mucosal Vaccination Overcomes the Barrier to Recombinant Vaccinia Immunization Caused by Preexisting Poxvirus Immunity

    Science.gov (United States)

    Belyakov, Igor M.; Moss, Bernard; Strober, Warren; Berzofsky, Jay A.

    1999-04-01

    Overcoming preexisting immunity to vaccinia virus in the adult population is a key requirement for development of otherwise potent recombinant vaccinia vaccines. Based on our observation that s.c. immunization with vaccinia induces cellular and antibody immunity to vaccinia only in systemic lymphoid tissue and not in mucosal sites, we hypothesized that the mucosal immune system remains naive to vaccinia and therefore amenable to immunization with recombinant vaccinia vectors despite earlier vaccinia exposure. We show that mucosal immunization of vaccinia-immune BALB/c mice with recombinant vaccinia expressing HIV gp160 induced specific serum antibody and strong HIV-specific cytotoxic T lymphocyte responses. These responses occurred not only in mucosal but also in systemic lymphoid tissue, whereas systemic immunization was ineffective under these circumstances. In this context, intrarectal immunization was more effective than intranasal immunization. Boosting with a second dose of recombinant vaccinia was also more effective via the mucosal route. The systemic HIV-specific cytotoxic T lymphocyte response was enhanced by coadministration of IL-12 at the mucosal site. These results also demonstrate the independent compartmentalization of the mucosal versus systemic immune systems and the asymmetric trafficking of lymphocytes between them. This approach to circumvent previous vaccinia immunity may be useful for induction of protective immunity against infectious diseases and cancer in the sizable populations with preexisting immunity to vaccinia from smallpox vaccination.

  8. In vitro characterization of a nineteenth-century therapy for smallpox.

    Science.gov (United States)

    Arndt, William; Mitnik, Chandra; Denzler, Karen L; White, Stacy; Waters, Robert; Jacobs, Bertram L; Rochon, Yvan; Olson, Victoria A; Damon, Inger K; Langland, Jeffrey O

    2012-01-01

    In the nineteenth century, smallpox ravaged through the United States and Canada. At this time, a botanical preparation, derived from the carnivorous plant Sarracenia purpurea, was proclaimed as being a successful therapy for smallpox infections. The work described characterizes the antipoxvirus activity associated with this botanical extract against vaccinia virus, monkeypox virus and variola virus, the causative agent of smallpox. Our work demonstrates the in vitro characterization of Sarracenia purpurea as the first effective inhibitor of poxvirus replication at the level of early viral transcription. With the renewed threat of poxvirus-related infections, our results indicate Sarracenia purpurea may act as another defensive measure against Orthopoxvirus infections.

  9. In vitro characterization of a nineteenth-century therapy for smallpox.

    Directory of Open Access Journals (Sweden)

    William Arndt

    Full Text Available In the nineteenth century, smallpox ravaged through the United States and Canada. At this time, a botanical preparation, derived from the carnivorous plant Sarracenia purpurea, was proclaimed as being a successful therapy for smallpox infections. The work described characterizes the antipoxvirus activity associated with this botanical extract against vaccinia virus, monkeypox virus and variola virus, the causative agent of smallpox. Our work demonstrates the in vitro characterization of Sarracenia purpurea as the first effective inhibitor of poxvirus replication at the level of early viral transcription. With the renewed threat of poxvirus-related infections, our results indicate Sarracenia purpurea may act as another defensive measure against Orthopoxvirus infections.

  10. Smallpox virus plaque phenotypes: genetic, geographical and case fatality relationships.

    Science.gov (United States)

    Olson, Victoria A; Karem, Kevin L; Smith, Scott K; Hughes, Christine M; Damon, Inger K

    2009-04-01

    Smallpox (infection with Orthopoxvirus variola) remains a feared illness more than 25 years after its eradication. Historically, case-fatality rates (CFRs) varied between outbreaks (<1 to approximately 40 %), the reasons for which are incompletely understood. The extracellular enveloped virus (EEV) form of orthopoxvirus progeny is hypothesized to disseminate infection. Investigations with the closely related Orthopoxvirus vaccinia have associated increased comet formation (EEV production) with increased mouse mortality (pathogenicity). Other vaccinia virus genetic manipulations which affect EEV production inconsistently support this association. However, antisera against vaccinia virus envelope protect mice from lethal challenge, further supporting a critical role for EEV in pathogenicity. Here, we show that the increased comet formation phenotypes of a diverse collection of variola viruses associate with strain phylogeny and geographical origin, but not with increased outbreak-related CFRs; within clades, there may be an association of plaque size with CFR. The mechanisms for variola virus pathogenicity probably involves multiple host and pathogen factors.

  11. Mozart and smallpox.

    Science.gov (United States)

    Zegers, Richard H C

    2007-01-01

    In 1767 at 11 years of age, composer Wolfgang Amadeus Mozart contracted smallpox, allegedly causing him temporary blindness. Although now eradicated, smallpox in those days had a high mortality rate, and the history of classical music would have been very different if Mozart had become permanently blind, or died, as a result of the disease.

  12. Smallpox: can we still learn from the journey to eradication?

    Science.gov (United States)

    Smith, Kendall A

    2013-05-01

    One of the most celebrated achievements of immunology and modern medicine is the eradication of the dreaded plague smallpox. From the introduction of smallpox vaccination by Edward Jenner, to its popularization by Louis Pasteur, to the eradication effort led by Donald Henderson, this story has many lessons for us today, including the characteristics of the disease and vaccine that permitted its eradication, and the obviousness of the vaccine as a vector for other intractable Infectious diseases. The disease itself, interpreted in the light of modern molecular immunology, is an obvious immunopathological disease, which occurs after a latent interval of 1-2 weeks, and manifests as a systemic cell-mediated delayed type hypersensitivity (DTH) syndrome. The vaccine that slayed this dragon was given the name vaccinia, and was thought to have evolved from cowpox virus, but is now known to be most closely related to a poxvirus isolated from a horse. Of interest is the fact that of the various isolates of orthopox viruses, only variola, vaccinia and monkeypox viruses can infect humans. In contrast to the systemic disease of variola, vaccinia only replicates locally at the site of inoculation, and causes a localized DTH response that usually peaks after 7-10 days. This difference in the pathogenicity of variola vs. vaccinia is thought to be due to the capacity of variola to circumvent innate immunity, which allows it to disseminate widely before the adaptive immune response occurs. Thus, the fact that vaccinia virus is attenuated compared to variola, but is still replication competent, makes for its remarkable efficacy as a vaccine, as the localized infection activates all of the cells and molecules of both innate and adaptive immunity. Accordingly vaccinia itself, and not modified replication incompetent vaccina, is the hope for use as a vector in the eradication of additional pathogenic microbes from the globe.

  13. Potential virulence determinants in terminal regions of variola smallpox virus genome.

    Science.gov (United States)

    Massung, R F; Esposito, J J; Liu, L I; Qi, J; Utterback, T R; Knight, J C; Aubin, L; Yuran, T E; Parsons, J M; Loparev, V N

    Smallpox eradication culminated the most successful antimicrobial campaign in medical history. To characterize further the linear double-stranded DNA genome of the aetiological agent of smallpox, we have determined the entire nucleotide sequence of the highly virulent variola major virus, strain Bangladesh-1975 (VAR-BSH; 186,102 base pairs, 33.7% G + C; Genbank accession number, L22579). Here we highlight features of the molecule and focus on a few of the 187 putative proteins that probably contribute to pathogenicity and virus host-range properties. One hundred and fifty proteins were markedly similar to those of vaccinia virus (smallpox vaccine), for which a complete sequence has been reported for strain Copenhagen (VAC-CPN; 191,636 base pairs, 33.3% G + C). The remaining 37 proteins reflected variola-specific sequences or open reading frame divergences for variant proteins, which are often truncated or elongated compared with their vaccinia counterparts.

  14. Immunomodulator-based enhancement of anti smallpox immune responses.

    Directory of Open Access Journals (Sweden)

    Osmarie Martínez

    Full Text Available The current live vaccinia virus vaccine used in the prevention of smallpox is contraindicated for millions of immune-compromised individuals. Although vaccination with the current smallpox vaccine produces protective immunity, it might result in mild to serious health complications for some vaccinees. Thus, there is a critical need for the production of a safe virus-free vaccine against smallpox that is available to everyone. For that reason, we investigated the impact of imiquimod and resiquimod (Toll-like receptors agonists, and the codon-usage optimization of the vaccinia virus A27L gene in the enhancement of the immune response, with intent of producing a safe, virus-free DNA vaccine coding for the A27 vaccinia virus protein.We analyzed the cellular-immune response by measuring the IFN-γ production of splenocytes by ELISPOT, the humoral-immune responses measuring total IgG and IgG2a/IgG1 ratios by ELISA, and the TH1 and TH2 cytokine profiles by ELISA, in mice immunized with our vaccine formulation.The proposed vaccine formulation enhanced the A27L vaccine-mediated production of IFN-γ on mouse spleens, and increased the humoral immunity with a TH1-biased response. Also, our vaccine induced a TH1 cytokine milieu, which is important against viral infections.These results support the efforts to find a new mechanism to enhance an immune response against smallpox, through the implementation of a safe, virus-free DNA vaccination platform.

  15. Immunogenicity and protection efficacy of subunit-based smallpox vaccines using variola major antigens.

    Science.gov (United States)

    Sakhatskyy, Pavlo; Wang, Shixia; Zhang, Chuanyou; Chou, Te-Hui; Kishko, Michael; Lu, Shan

    2008-02-05

    The viral strain responsible for smallpox infection is variola major (VARV). As a result of the successful eradication of smallpox with the vaccinia virus (VACV), the general population is no longer required to receive a smallpox vaccine, and will have no protection against smallpox. This lack of immunity is a concern due to the potential for use of smallpox as a biological weapon. Considerable progress has been made in the development of subunit-based smallpox vaccines resulting from the identification of VACV protective antigens. It also offers the possibility of using antigens from VARV to formulate the next generation subunit-based smallpox vaccines. Here, we show that codon-optimized DNA vaccines expressing three VARV antigens (A30, B7 and F8) and their recombinant protein counterparts elicited high-titer, cross-reactive, VACV neutralizing antibody responses in mice. Vaccinated mice were protected from intraperitoneal and intranasal challenges with VACV. These results suggest the feasibility of a subunit smallpox vaccine based on VARV antigen sequences to induce immunity against poxvirus infection.

  16. Planning for smallpox outbreaks

    Science.gov (United States)

    Ferguson, Neil M.; Keeling, Matt J.; John Edmunds, W.; Gani, Raymond; Grenfell, Bryan T.; Anderson, Roy M.; Leach, Steve

    2003-10-01

    Mathematical models of viral transmission and control are important tools for assessing the threat posed by deliberate release of the smallpox virus and the best means of containing an outbreak. Models must balance biological realism against limitations of knowledge, and uncertainties need to be accurately communicated to policy-makers. Smallpox poses the particular challenge that key biological, social and spatial factors affecting disease spread in contemporary populations must be elucidated largely from historical studies undertaken before disease eradication in 1979. We review the use of models in smallpox planning within the broader epidemiological context set by recent outbreaks of both novel and re-emerging pathogens.

  17. Susceptibility of different leukocyte cell types to Vaccinia virus infection

    Directory of Open Access Journals (Sweden)

    Sánchez-Puig Juana M

    2004-11-01

    Full Text Available Abstract Background Vaccinia virus, the prototype member of the family Poxviridae, was used extensively in the past as the Smallpox vaccine, and is currently considered as a candidate vector for new recombinant vaccines. Vaccinia virus has a wide host range, and is known to infect cultures of a variety of cell lines of mammalian origin. However, little is known about the virus tropism in human leukocyte populations. We report here that various cell types within leukocyte populations have widely different susceptibility to infection with vaccinia virus. Results We have investigated the ability of vaccinia virus to infect human PBLs by using virus recombinants expressing green fluorescent protein (GFP, and monoclonal antibodies specific for PBL subpopulations. Flow cytometry allowed the identification of infected cells within the PBL mixture 1–5 hours after infection. Antibody labeling revealed that different cell populations had very different infection rates. Monocytes showed the highest percentage of infected cells, followed by B lymphocytes and NK cells. In contrast to those cell types, the rate of infection of T lymphocytes was low. Comparison of vaccinia virus strains WR and MVA showed that both strains infected efficiently the monocyte population, although producing different expression levels. Our results suggest that MVA was less efficient than WR in infecting NK cells and B lymphocytes. Overall, both WR and MVA consistently showed a strong preference for the infection of non-T cells. Conclusions When infecting fresh human PBL preparations, vaccinia virus showed a strong bias towards the infection of monocytes, followed by B lymphocytes and NK cells. In contrast, very poor infection of T lymphocytes was detected. These finding may have important implications both in our understanding of poxvirus pathogenesis and in the development of improved smallpox vaccines.

  18. The rediscovery of smallpox.

    Science.gov (United States)

    Thèves, C; Biagini, P; Crubézy, E

    2014-03-01

    Smallpox is an infectious disease that is unique to humans, caused by a poxvirus. It is one of the most lethal of diseases; the virus variant Variola major has a mortality rate of 30%. People surviving this disease have life-long consequences, but also assured immunity. Historically, smallpox was recognized early in human populations. This led to prevention attempts--variolation, quarantine, and the isolation of infected subjects--until Jenner's discovery of the first steps of vaccination in the 18th century. After vaccination campaigns throughout the 19th and 20th centuries, the WHO declared the eradication of smallpox in 1980. With the development of microscopy techniques, the structural characterization of the virus began in the early 20th century. In 1990, the genomes of different smallpox viruses were determined; viruses could be classified in order to investigate their origin, diffusion, and evolution. To study the evolution and possible re-emergence of this viral pathogen, however, researchers can only use viral genomes collected during the 20th century. Cases of smallpox in ancient periods are sometimes well documented, so palaeomicrobiology and, more precisely, the study of ancient smallpox viral strains could be an exceptional opportunity. The analysis of poxvirus fragmented genomes could give new insights into the genetic evolution of the poxvirus. Recently, small fragments of the poxvirus genome were detected. With the genetic information obtained, a new phylogeny of smallpox virus was described. The interest in conducting studies on ancient strains is discussed, in order to explore the natural history of this disease. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

  19. Neurologic Complications of Smallpox Vaccination

    Directory of Open Access Journals (Sweden)

    J Gordon Millichap

    2003-06-01

    Full Text Available Smallpox and smallpox vaccination is reviewed from the Departments of Neurology, Yale University School of Medicine, New Haven, CT, and University of New Mexico School of Medicine, Albuquerque.

  20. Mankind's Magnificent Milestone: Smallpox Eradication.

    Science.gov (United States)

    Small, Parker A., Jr.; Small, Natalie S.

    1996-01-01

    Illustrates the complex interactions between disease, societal attitudes, and technology by looking at the history of smallpox. Describes one of mankind's most magnificent accomplishments--the eradication of smallpox from the earth. (JRH)

  1. Small(pox) success?

    Science.gov (United States)

    Birn, Anne-Emanuelle

    2011-02-01

    The 30th anniversary of the World Health Organization's (WHO) official certification of smallpox eradication was marked by a slew of events hailing the campaign's dramatic tale of technological and organizational triumph against an ancient scourge. Yet commemorations also serve as moments of critical reflection. This article questions the acclaim showered upon smallpox eradication as the single greatest public health success in history. It examines how and why smallpox eradication and WHO's concurrent social justice-oriented primary health care approach (following from the Declaration of Alma-Ata) became competing paradigms. It synthesizes critiques of eradication's shortcomings and debunks some of the myths surrounding the global eradication campaign as a public health priority and necessity, and as a Cold War victory of cooperation. The article concludes with thoughts on integrating technical and social-political aspects of health within the context of welfare states as the means to achieving widespread and enduring global public health success.

  2. Small(pox success?

    Directory of Open Access Journals (Sweden)

    Anne-Emanuelle Birn

    Full Text Available The 30th anniversary of the World Health Organization's (WHO official certification of smallpox eradication was marked by a slew of events hailing the campaign's dramatic tale of technological and organizational triumph against an ancient scourge. Yet commemorations also serve as moments of critical reflection. This article questions the acclaim showered upon smallpox eradication as the single greatest public health success in history. It examines how and why smallpox eradication and WHO's concurrent social justice-oriented primary health care approach (following from the Declaration of Alma-Ata became competing paradigms. It synthesizes critiques of eradication's shortcomings and debunks some of the myths surrounding the global eradication campaign as a public health priority and necessity, and as a Cold War victory of cooperation. The article concludes with thoughts on integrating technical and social-political aspects of health within the context of welfare states as the means to achieving widespread and enduring global public health success.

  3. Human CD4+ T cell epitopes from vaccinia virus induced by vaccination or infection.

    Directory of Open Access Journals (Sweden)

    J Mauricio Calvo-Calle

    2007-10-01

    Full Text Available 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.

  4. Molecular Smallpox Vaccine Delivered by Alphavirus Replicons Elicits Protective Immunity in Mice and Non-human Primates

    Science.gov (United States)

    Hooper, Jay W.; Ferro, Anthony M.; Golden, Joseph W.; Silvera, Peter; Dudek, Jeanne; Alterson, Kim; Custer, Max; Rivers, Bryan; Morris, John; Owens, Gary; Smith, Jonathan F.; Kamrud, Kurt I.

    2009-01-01

    Naturally occurring smallpox was eradicated as a result of successful vaccination campaigns during the 1960s and 70s. Because of its highly contagious nature and high mortality rate, smallpox has significant potential as a biological weapon. Unfortunately, the current vaccine for orthopoxviruses is contraindicated for large portions of the population. Thus, there is a need for new, safe, and effective orthopoxvirus vaccines. Alphavirus replicon vectors, derived from strains of Venezuelan equine encephalitis virus, are being used to develop alternatives to the current smallpox vaccine. Here, we demonstrated that virus-like replicon particles (VRP) expressing the vaccinia virus A33R, B5R, A27L, and L1R genes elicited protective immunity in mice comparable to vaccination with live-vaccinia virus. Furthermore, cynomolgus macaques vaccinated with a combination of the four poxvirus VRPs (4pox-VRP) developed antibody responses to each antigen. These antibody responses were able to neutralize and inhibit the spread of both vaccinia virus and monkeypox virus. Macaques vaccinated with 4pox-VRP, flu HA VRP (negative control), or live-vaccinia virus (positive control) were challenged intravenously with 5 × 106 PFU of monkeypox virus 1 month after the second VRP vaccination. Four of the six negative control animals succumbed to monkeypox and the remaining two animals demonstrated either severe or grave disease. Importantly, all 10 macaques vaccinated with the 4pox-VRP vaccine survived without developing severe disease. These findings revealed that a single-boost VRP smallpox vaccine shows promise as a safe alternative to the currently licensed live-vaccinia virus smallpox vaccine. PMID:19833247

  5. Molecular smallpox vaccine delivered by alphavirus replicons elicits protective immunity in mice and non-human primates.

    Science.gov (United States)

    Hooper, Jay W; Ferro, Anthony M; Golden, Joseph W; Silvera, Peter; Dudek, Jeanne; Alterson, Kim; Custer, Max; Rivers, Bryan; Morris, John; Owens, Gary; Smith, Jonathan F; Kamrud, Kurt I

    2009-12-11

    Naturally occurring smallpox was eradicated as a result of successful vaccination campaigns during the 1960s and 1970s. Because of its highly contagious nature and high mortality rate, smallpox has significant potential as a biological weapon. Unfortunately, the current vaccine for orthopoxviruses is contraindicated for large portions of the population. Thus, there is a need for new, safe, and effective orthopoxvirus vaccines. Alphavirus replicon vectors, derived from strains of Venezuelan equine encephalitis virus, are being used to develop alternatives to the current smallpox vaccine. Here, we demonstrated that virus-like replicon particles (VRPs) expressing the vaccinia virus A33R, B5R, A27L, and L1R genes elicited protective immunity in mice comparable to vaccination with live-vaccinia virus. Furthermore, cynomolgus macaques vaccinated with a combination of the four poxvirus VRPs (4pox-VRP) developed antibody responses to each antigen. These antibody responses were able to neutralize and inhibit the spread of both vaccinia virus and monkeypox virus. Macaques vaccinated with 4pox-VRP, flu HA VRP (negative control), or live-vaccinia virus (positive control) were challenged intravenously with 5 x 10(6)pfu of monkeypox virus 1 month after the second VRP vaccination. Four of the six negative control animals succumbed to monkeypox and the remaining two animals demonstrated either severe or grave disease. Importantly, all 10 macaques vaccinated with the 4pox-VRP vaccine survived without developing severe disease. These findings revealed that a single-boost VRP smallpox vaccine shows promise as a safe alternative to the currently licensed live-vaccinia virus smallpox vaccine.

  6. GM-CSF production allows the identification of immunoprevalent antigens recognized by human CD4+ T cells following smallpox vaccination.

    Directory of Open Access Journals (Sweden)

    Valeria Judkowski

    Full Text Available The threat of bioterrorism with smallpox and the broad use of vaccinia vectors for other vaccines have led to the resurgence in the study of vaccinia immunological memory. The importance of the role of CD4+ T cells in the control of vaccinia infection is well known. However, more CD8+ than CD4+ T cell epitopes recognized by human subjects immunized with vaccinia virus have been reported. This could be, in part, due to the fact that most of the studies that have identified human CD4+ specific protein-derived fragments or peptides have used IFN-γ production to evaluate vaccinia specific T cell responses. Based on these findings, we reasoned that analyzing a large panel of cytokines would permit us to generate a more complete analysis of the CD4 T cell responses. The results presented provide clear evidence that TNF-α is an excellent readout of vaccinia specificity and that other cytokines such as GM-CSF can be used to evaluate the reactivity of CD4+ T cells in response to vaccinia antigens. Furthermore, using these cytokines as readout of vaccinia specificity, we present the identification of novel peptides from immunoprevalent vaccinia proteins recognized by CD4+ T cells derived from smallpox vaccinated human subjects. In conclusion, we describe a "T cell-driven" methodology that can be implemented to determine the specificity of the T cell response upon vaccination or infection. Together, the single pathogen in vitro stimulation, the selection of CD4+ T cells specific to the pathogen by limiting dilution, the evaluation of pathogen specificity by detecting multiple cytokines, and the screening of the clones with synthetic combinatorial libraries, constitutes a novel and valuable approach for the elucidation of human CD4+ T cell specificity in response to large pathogens.

  7. Immunization with vaccinia virus induces polyfunctional and phenotypically distinctive CD8+ T cell responses

    Science.gov (United States)

    Precopio, Melissa L.; Betts, Michael R.; Parrino, Janie; Price, David A.; Gostick, Emma; Ambrozak, David R.; Asher, Tedi E.; Douek, Daniel C.; Harari, Alexandre; Pantaleo, Giuseppe; Bailer, Robert; Graham, Barney S.; Roederer, Mario; Koup, Richard A.

    2007-01-01

    Vaccinia virus immunization provides lifelong protection against smallpox, but the mechanisms of this exquisite protection are unknown. We used polychromatic flow cytometry to characterize the functional and phenotypic profile of CD8+ T cells induced by vaccinia virus immunization in a comparative vaccine trial of modified vaccinia virus Ankara (MVA) versus Dryvax immunization in which protection was assessed against subsequent Dryvax challenge. Vaccinia virus–specific CD8+ T cells induced by both MVA and Dryvax were highly polyfunctional; they degranulated and produced interferon γ, interleukin 2, macrophage inflammatory protein 1β, and tumor necrosis factor α after antigenic stimulation. Responding CD8+ T cells exhibited an unusual phenotype (CD45RO−CD27intermediate). The unique phenotype and high degree of polyfunctionality induced by vaccinia virus also extended to inserted HIV gene products of recombinant NYVAC. This quality of the CD8+ T cell response may be at least partially responsible for the profound efficacy of these vaccines in protection against smallpox and serves as a benchmark against which other vaccines can be evaluated. PMID:17535971

  8. Vaccinations against smallpox and tuberculosis are associated with better long-term survival

    DEFF Research Database (Denmark)

    Rieckmann, Andreas; Villumsen, Marie; Sørup, Signe

    2016-01-01

    BACKGROUND: When vaccinations with vaccinia against smallpox and Bacillus Calmette-Guérin (BCG) against tuberculosis were phased out in some high-income countries around 1980, the impact on overall mortality was not examined. Recent studies from low-income countries have suggested that these vacc......BACKGROUND: When vaccinations with vaccinia against smallpox and Bacillus Calmette-Guérin (BCG) against tuberculosis were phased out in some high-income countries around 1980, the impact on overall mortality was not examined. Recent studies from low-income countries have suggested...... that these vaccines are associated with mortality reductions, not explained by specific disease protection. We examined whether vaccinia and BCG administered in childhood were associated with long-term mortality reductions in a high-income population. METHODS: In this case-cohort study, we followed 47 622...... schoolchildren from Copenhagen, Denmark, born 1965 to 1976, from their first health examination to 2010. This cohort experienced the phase-out of vaccinia and BCG vaccination programmes. RESULTS: A sub-cohort of 5 316 individuals (699 excluded) was followed for 164 450 person-years (0.2% were lost to follow...

  9. Pre-event Smallpox Vaccination for Healthcare Workers Revisited – the Need for a Carefully Screened Multidisciplinary Cadre

    Energy Technology Data Exchange (ETDEWEB)

    Malone, JD D.

    2007-03-01

    Abstract As healthcare institutions are a focus of smallpox transmission early in an epidemic, several mathematical models support pre-event smallpox vaccination of healthcare workers (HCWs). The deciding factor for HCW voluntary vaccination is the risk of disease exposure versus the risk of vaccine adverse events. In a United States military population, with careful screening to exclude atopic dermatitis/eczema and immunosuppression, over 1 million vaccinia vaccinations were delivered with 1 fatality attributed to vaccination. Among 37,901 U.S. civilian volunteer healthcare workers vaccinated, 100 serious adverse events were reported including 10 ischemic cardiac episodes and six myocardial infarctions – 2 were fatal. This older population had a higher rate of adverse events due to age related coronary artery disease. T-cell mediated inflammatory processes, induced by live vaccinia vaccination, may have a role in the observed acute coronary artery events. With exclusion of individuals at risk for coronary artery disease, atopic dermatitis/eczema, and immunosuppression, HCWs can be smallpox vaccinated with minimal risk. A smallpox pre-vaccinated multidisciplinary cadre (physician, nurse, infection control practitioner, technician) will supply leadership to deal with fear and uncertainty while limiting spread and initial mortality of smallpox. Stochastic – from the Greek meaning “skillful in aiming” – is currently interpreted as arising from chance and involving probability. This issue’s article “Containing a large bioterrorist smallpox attack: a computer simulation approach” by Longini et al. is a discrete time, stochastic computer simulation model that offers additional planning guidance for a smallpox (variola virus) outbreak (1). Although interpretation of the model’s information may differ, Longini’s article concludes “Given that surveillance and containment measures are in place, preemptive vaccination of hospital workers would further

  10. From Crescent to Mature Virion: Vaccinia Virus Assembly and Maturation

    Directory of Open Access Journals (Sweden)

    Liang Liu

    2014-10-01

    Full Text Available Vaccinia virus (VACV has achieved unprecedented success as a live viral vaccine for smallpox which mitigated eradication of the disease. Vaccinia virus has a complex virion morphology and recent advances have been made to answer some of the key outstanding questions, in particular, the origin and biogenesis of the virion membrane, the transformation from immature virion (IV to mature virus (MV, and the role of several novel genes, which were previously uncharacterized, but have now been shown to be essential for VACV virion formation. This new knowledge will undoubtedly contribute to the rational design of safe, immunogenic vaccine candidates, or effective antivirals in the future. This review endeavors to provide an update on our current knowledge of the VACV maturation processes with a specific focus on the initiation of VACV replication through to the formation of mature virions.

  11. Oncolytic vaccinia therapy of squamous cell carcinoma

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    Yu Yong A

    2009-07-01

    Full Text Available Abstract Background Novel therapies are necessary to improve outcomes for patients with squamous cell carcinomas (SCC of the head and neck. Historically, vaccinia virus was administered widely to humans as a vaccine and led to the eradication of smallpox. We examined the therapeutic effects of an attenuated, replication-competent vaccinia virus (GLV-1h68 as an oncolytic agent against a panel of six human head and neck SCC cell lines. Results All six cell lines supported viral transgene expression (β-galactosidase, green fluorescent protein, and luciferase as early as 6 hours after viral exposure. Efficient transgene expression and viral replication (>150-fold titer increase over 72 hrs were observed in four of the cell lines. At a multiplicity of infection (MOI of 1, GLV-1h68 was highly cytotoxic to the four cell lines, resulting in ≥ 90% cytotoxicity over 6 days, and the remaining two cell lines exhibited >45% cytotoxicity. Even at a very low MOI of 0.01, three cell lines still demonstrated >60% cell death over 6 days. A single injection of GLV-1h68 (5 × 106 pfu intratumorally into MSKQLL2 xenografts in mice exhibited localized intratumoral luciferase activity peaking at days 2–4, with gradual resolution over 10 days and no evidence of spread to normal organs. Treated animals exhibited near-complete tumor regression over a 24-day period without any observed toxicity, while control animals demonstrated rapid tumor progression. Conclusion These results demonstrate significant oncolytic efficacy by an attenuated vaccinia virus for infecting and lysing head and neck SCC both in vitro and in vivo, and support its continued investigation in future clinical trials.

  12. Defending against smallpox: a focus on vaccines.

    Science.gov (United States)

    Voigt, Emily A; Kennedy, Richard B; Poland, Gregory A

    2016-09-01

    Smallpox has shaped human history, from the earliest human civilizations well into the 20th century. With high mortality rates, rapid transmission, and serious long-term effects on survivors, smallpox was a much-feared disease. The eradication of smallpox represents an unprecedented medical victory for the lasting benefit of human health and prosperity. Concerns remain, however, about the development and use of the smallpox virus as a biological weapon, which necessitates the need for continued vaccine development. Smallpox vaccine development is thus a much-reviewed topic of high interest. This review focuses on the current state of smallpox vaccines and their context in biodefense efforts.

  13. Traditional Smallpox Vaccines and Atopic Dermatitis

    Science.gov (United States)

    ... Trials 2016 Research Grant Request DONATE Traditional Smallpox Vaccines and Atopic Dermatitis Frequently Asked Questions Eczema Living ... Involved Eczema Products News Research Donate Traditional Smallpox Vaccines and Atopic Dermatitis Frequently Asked Questions What is ...

  14. Preventing the return of smallpox: molecular modeling studies on thymidylate kinase from Variola virus.

    Science.gov (United States)

    Guimarães, Ana Paula; Ramalho, Teodorico Castro; França, Tanos Celmar Costa

    2014-01-01

    Smallpox was one of the most devastating diseases in the human history and still represents a serious menace today due to its potential use by bioterrorists. Considering this threat and the non-existence of effective chemotherapy, we propose the enzyme thymidylate kinase from Variola virus (VarTMPK) as a potential target to the drug design against smallpox. We first built a homology model for VarTMPK and performed molecular docking studies on it in order to investigate the interactions with inhibitors of Vaccinia virus TMPK (VacTMPK). Subsequently, molecular dynamics (MD) simulations of these compounds inside VarTMPK and human TMPK (HssTMPK) were carried out in order to select the most promising and selective compounds as leads for the design of potential VarTMPK inhibitors. Results of the docking and MD simulations corroborated to each other, suggesting selectivity towards VarTMPK and, also, a good correlation with the experimental data.

  15. TLR3 and TLR9 agonists improve postexposure vaccination efficacy of live smallpox vaccines.

    Science.gov (United States)

    Israely, Tomer; Melamed, Sharon; Achdout, Hagit; Erez, Noam; Politi, Boaz; Waner, Trevor; Lustig, Shlomo; Paran, Nir

    2014-01-01

    Eradication of smallpox and discontinuation of the vaccination campaign resulted in an increase in the percentage of unvaccinated individuals, highlighting the need for postexposure efficient countermeasures in case of accidental or deliberate viral release. Intranasal infection of mice with ectromelia virus (ECTV), a model for human smallpox, is curable by vaccination with a high vaccine dose given up to 3 days postexposure. To further extend this protective window and to reduce morbidity, mice were vaccinated postexposure with Vaccinia-Lister, the conventional smallpox vaccine or Modified Vaccinia Ankara, a highly attenuated vaccine in conjunction with TLR3 or TLR9 agonists. We show that co-administration of the TLR3 agonist poly(I:C) even 5 days postexposure conferred protection, avoiding the need to increase the vaccination dose. Efficacious treatments prevented death, ameliorated disease symptoms, reduced viral load and maintained tissue integrity of target organs. Protection was associated with significant elevation of serum IFNα and anti-vaccinia IgM antibodies, modulation of IFNγ response, and balanced activation of NK and T cells. TLR9 agonists (CpG ODNs) were less protective than the TLR3 agonist poly(I:C). We show that activation of type 1 IFN by poly(I:C) and protection is achievable even without co-vaccination, requiring sufficient amount of the viral antigens of the infective agent or the vaccine. This study demonstrated the therapeutic potential of postexposure immune modulation by TLR activation, allowing to alleviate the disease symptoms and to further extend the protective window of postexposure vaccination.

  16. TLR3 and TLR9 agonists improve postexposure vaccination efficacy of live smallpox vaccines.

    Directory of Open Access Journals (Sweden)

    Tomer Israely

    Full Text Available Eradication of smallpox and discontinuation of the vaccination campaign resulted in an increase in the percentage of unvaccinated individuals, highlighting the need for postexposure efficient countermeasures in case of accidental or deliberate viral release. Intranasal infection of mice with ectromelia virus (ECTV, a model for human smallpox, is curable by vaccination with a high vaccine dose given up to 3 days postexposure. To further extend this protective window and to reduce morbidity, mice were vaccinated postexposure with Vaccinia-Lister, the conventional smallpox vaccine or Modified Vaccinia Ankara, a highly attenuated vaccine in conjunction with TLR3 or TLR9 agonists. We show that co-administration of the TLR3 agonist poly(I:C even 5 days postexposure conferred protection, avoiding the need to increase the vaccination dose. Efficacious treatments prevented death, ameliorated disease symptoms, reduced viral load and maintained tissue integrity of target organs. Protection was associated with significant elevation of serum IFNα and anti-vaccinia IgM antibodies, modulation of IFNγ response, and balanced activation of NK and T cells. TLR9 agonists (CpG ODNs were less protective than the TLR3 agonist poly(I:C. We show that activation of type 1 IFN by poly(I:C and protection is achievable even without co-vaccination, requiring sufficient amount of the viral antigens of the infective agent or the vaccine. This study demonstrated the therapeutic potential of postexposure immune modulation by TLR activation, allowing to alleviate the disease symptoms and to further extend the protective window of postexposure vaccination.

  17. Protective effect of Toll-like receptor 4 in pulmonary vaccinia infection.

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    Martha A Hutchens

    Full Text Available Innate immune responses are essential for controlling poxvirus infection. The threat of a bioterrorist attack using Variola major, the smallpox virus, or zoonotic transmission of other poxviruses has renewed interest in understanding interactions between these viruses and their hosts. We recently determined that TLR3 regulates a detrimental innate immune response that enhances replication, morbidity, and mortality in mice in response to vaccinia virus, a model pathogen for studies of poxviruses. To further investigate Toll-like receptor signaling in vaccinia infection, we first focused on TRIF, the only known adapter protein for TLR3. Unexpectedly, bioluminescence imaging showed that mice lacking TRIF are more susceptible to vaccinia infection than wild-type mice. We then focused on TLR4, the other Toll-like receptor that signals through TRIF. Following respiratory infection with vaccinia, mice lacking TLR4 signaling had greater viral replication, hypothermia, and mortality than control animals. The mechanism of TLR4-mediated protection was not due to increased release of proinflammatory cytokines or changes in total numbers of immune cells recruited to the lung. Challenge of primary bone marrow macrophages isolated from TLR4 mutant and control mice suggested that TLR4 recognizes a viral ligand rather than an endogenous ligand. These data establish that TLR4 mediates a protective innate immune response against vaccinia virus, which informs development of new vaccines and therapeutic agents targeted against poxviruses.

  18. Incongruencies in Vaccinia Virus Phylogenetic Trees

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    Chad Smithson

    2014-10-01

    Full Text Available Over the years, as more complete poxvirus genomes have been sequenced, phylogenetic studies of these viruses have become more prevalent. In general, the results show similar relationships between the poxvirus species; however, some inconsistencies are notable. Previous analyses of the viral genomes contained within the vaccinia virus (VACV-Dryvax vaccine revealed that their phylogenetic relationships were sometimes clouded by low bootstrapping confidence. To analyze the VACV-Dryvax genomes in detail, a new tool-set was developed and integrated into the Base-By-Base bioinformatics software package. Analyses showed that fewer unique positions were present in each VACV-Dryvax genome than expected. A series of patterns, each containing several single nucleotide polymorphisms (SNPs were identified that were counter to the results of the phylogenetic analysis. The VACV genomes were found to contain short DNA sequence blocks that matched more distantly related clades. Additionally, similar non-conforming SNP patterns were observed in (1 the variola virus clade; (2 some cowpox clades; and (3 VACV-CVA, the direct ancestor of VACV-MVA. Thus, traces of past recombination events are common in the various orthopoxvirus clades, including those associated with smallpox and cowpox viruses.

  19. Modified Vaccinia Virus Ankara: History, Value in Basic Research, and Current Perspectives for Vaccine Development.

    Science.gov (United States)

    Volz, A; Sutter, G

    2017-01-01

    Safety tested Modified Vaccinia virus Ankara (MVA) is licensed as third-generation vaccine against smallpox and serves as a potent vector system for development of new candidate vaccines against infectious diseases and cancer. Historically, MVA was developed by serial tissue culture passage in primary chicken cells of vaccinia virus strain Ankara, and clinically used to avoid the undesirable side effects of conventional smallpox vaccination. Adapted to growth in avian cells MVA lost the ability to replicate in mammalian hosts and lacks many of the genes orthopoxviruses use to conquer their host (cell) environment. As a biologically well-characterized mutant virus, MVA facilitates fundamental research to elucidate the functions of poxvirus host-interaction factors. As extremely safe viral vectors MVA vaccines have been found immunogenic and protective in various preclinical infection models. Multiple recombinant MVA currently undergo clinical testing for vaccination against human immunodeficiency viruses, Mycobacterium tuberculosis or Plasmodium falciparum. The versatility of the MVA vector vaccine platform is readily demonstrated by the swift development of experimental vaccines for immunization against emerging infections such as the Middle East Respiratory Syndrome. Recent advances include promising results from the clinical testing of recombinant MVA-producing antigens of highly pathogenic avian influenza virus H5N1 or Ebola virus. This review summarizes our current knowledge about MVA as a unique strain of vaccinia virus, and discusses the prospects of exploiting this virus as research tool in poxvirus biology or as safe viral vector vaccine to challenge existing and future bottlenecks in vaccinology.

  20. Attitudes of healthcare workers in U.S. hospitals regarding smallpox vaccination

    Directory of Open Access Journals (Sweden)

    O'Brien Megan A

    2003-06-01

    Full Text Available Abstract Background The United States is implementing plans to immunize 500,000 hospital-based healthcare workers against smallpox. Vaccination is voluntary, and it is unknown what factors drive vaccine acceptance. This study's aims were to estimate the proportion of workers willing to accept vaccination and to identify factors likely to influence their decisions. Methods The survey was conducted among physicians, nurses, and others working primarily in emergency departments or intensive care units at 21 acute-care hospitals in 10 states during the two weeks before the U.S. national immunization program for healthcare workers was announced in December 2002. Of the questionnaires distributed, 1,165 were returned, for a response rate of 81%. The data were analyzed by logistic regression and were adjusted for clustering within hospital and for different number of responses per hospital, using generalized linear mixed models and SAS's NLMIXED procedure. Results Sixty-one percent of respondents said they would definitely or probably be vaccinated, while 39% were undecided or inclined against it. Fifty-three percent rated the risk of a bioterrorist attack using smallpox in the United States in the next two years as either intermediate or high. Forty-seven percent did not feel well-informed about the risks and benefits of vaccination. Principal concerns were adverse reactions and the risk of transmitting vaccinia. In multivariate analysis, four variables were associated with willingness to be vaccinated: perceived risk of an attack, self-assessed knowledge about smallpox vaccination, self-assessed previous smallpox vaccination status, and gender. Conclusions The success of smallpox vaccination efforts will ultimately depend on the relative weight in people's minds of the risk of vaccine adverse events compared with the risk of being exposed to the disease. Although more than half of the respondents thought the likelihood of a bioterrorist smallpox

  1. A novel high-throughput vaccinia virus neutralization assay and preexisting immunity in populations from different geographic regions in China.

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    Qiang Liu

    Full Text Available 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

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

    Science.gov (United States)

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

    2012-01-01

    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

  3. A novel high-throughput vaccinia virus neutralization assay and preexisting immunity in populations from different geographic regions in China.

    Science.gov (United States)

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

    2012-01-01

    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. 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. 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.

  4. Cytokine production associated with smallpox vaccine responses.

    Science.gov (United States)

    Simon, Whitney L; Salk, Hannah M; Ovsyannikova, Inna G; Kennedy, Richard B; Poland, Gregory A

    2014-01-01

    Smallpox was eradicated 34 years ago due to the success of the smallpox vaccine; yet, the vaccine continues to be studied because of its importance in responding to potential biological warfare and the adverse events associated with current smallpox vaccines. Interindividual variations in vaccine response are observed and are, in part, due to genetic variation. In some cases, these varying responses lead to adverse events, which occur at a relatively high rate for the smallpox vaccine compared with other vaccines. Here, we aim to summarize the cytokine responses associated with smallpox vaccine response to date. Along with a description of each of these cytokines, we describe the genetic and adverse event data associated with cytokine responses to smallpox vaccination.

  5. Smallpox - Multiple Languages: MedlinePlus

    Science.gov (United States)

    ... Hmong) PDF Minnesota Department of Health Khmer (Khmer) Smallpox Vaccine English វ៉ាក់សាំង smallpox - Khmer (Khmer) PDF ... Centers for Disease Control and Prevention Russian (Русский) Smallpox Vaccine English Вакцина против оспы - Русский (Russian) PDF Immunization ...

  6. Logistics in smallpox: the legacy.

    Science.gov (United States)

    Wickett, John; Carrasco, Peter

    2011-12-30

    Logistics, defined as "the time-related positioning of resources" was critical to the implementation of the smallpox eradication strategy of surveillance and containment. Logistical challenges in the smallpox programme included vaccine delivery, supplies, staffing, vehicle maintenance, and financing. Ensuring mobility was essential as health workers had to travel to outbreaks to contain them. Three examples illustrate a range of logistic challenges which required imagination and innovation. Standard price lists were developed to expedite vehicle maintenance and repair in Bihar, India. Innovative staffing ensured an adequate infrastructure for vehicle maintenance in Bangladesh. The use of disaster relief mechanisms in Somalia provided airlifts, vehicles and funding within 27 days of their initiation. In contrast the Expanded Programme on Immunization (EPI) faces more complex logistical challenges. Copyright © 2011 Elsevier Ltd. All rights reserved.

  7. Isolation and characterization of cidofovir resistant vaccinia viruses

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    Prichard Mark N

    2008-05-01

    Full Text Available Abstract Background The emergence of drug resistant viruses, together with the possibility of increased virulence, is an important concern in the development of new antiviral compounds. Cidofovir (CDV is a phosphonate nucleotide that is approved for use against cytomegalovirus retinitis and for the emergency treatment of smallpox or complications following vaccination. One mode of action for CDV has been demonstrated to be the inhibition of the viral DNA polymerase. Results We have isolated several CDV resistant (CDVR vaccinia viruses through a one step process, two of which have unique single mutations within the DNA polymerase. An additional resistant virus isolate provides evidence of a second site mutation within the genome involved in CDV resistance. The CDVR viruses were 3–7 fold more resistant to the drug than the parental viruses. The virulence of the CDVR viruses was tested in mice inoculated intranasally and all were found to be attenuated. Conclusion Resistance to CDV in vaccinia virus can be conferred individually by at least two different mutations within the DNA polymerase gene. Additional genes may be involved. This one step approach for isolating resistant viruses without serial passage and in the presence of low doses of drug minimizes unintended secondary mutations and is applicable to other potential antiviral agents.

  8. Emergence and reemergence of smallpox: the need for development of a new generation smallpox vaccine.

    Science.gov (United States)

    Shchelkunov, Sergei N

    2011-12-30

    The review summarizes the archive data on smallpox, history of ancient civilizations, and the most recent data on the genome organization of orthopoxviruses, their evolutionary relationships, and the time points of smallpox emergence. The performed analysis provides the grounds for the hypothesis that smallpox could have emerged several times as a result of evolutionary changes in the zoonotic ancestor virus and disappeared due to insufficient population size of ancient civilizations. Smallpox reemerged in the Indian subcontinent approximately 2500-3000 years before present, which resulted in endemization of this anthroponotic infection, which had been preserved until the smallpox eradication in the 20th century AD. The conclusion suggests a potential possibility of future variola virus reemergence, presenting a great menace for mankind, as well as the need for development of new safe smallpox vaccines, design of anti-smallpox drugs, and activation of the control of zoonotic human orthopoxvirus infections.

  9. The De Novo Synthesis of Horsepox Virus: Implications for Biosecurity and Recommendations for Preventing the Reemergence of Smallpox.

    Science.gov (United States)

    Koblentz, Gregory D

    2017-08-24

    In March 2017, the American biotech company Tonix announced that a Canadian scientist had synthesized horsepox virus as part of a project to develop a safer vaccine against smallpox. The first de novo synthesis of an orthopoxvirus, a closely related group of viruses that includes horsepox and the variola virus that causes smallpox, crosses an important Rubicon in the field of biosecurity. The synthesis of horsepox virus takes the world one step closer to the reemergence of smallpox as a threat to global health security. That threat has been held at bay for the past 40 years by the extreme difficulty of obtaining variola virus and the availability of effective medical countermeasures. The techniques demonstrated by the synthesis of horsepox have the potential to erase both of these barriers. The primary risk posed by this research is that it will open the door to the routine and widespread synthesis of other orthopoxviruses, such as vaccinia, for use in research, public health, and medicine. The normalization and globalization of orthopoxvirus synthesis for these beneficial applications will create a cadre of laboratories and scientists that will also have the capability and expertise to create infectious variola virus from synthetic DNA. Unless the safeguards against the synthesis of variola virus are strengthened, the capability to reintroduce smallpox into the human population will be globally distributed and either loosely or completely unregulated, providing the foundation for a disgruntled or radicalized scientist, sophisticated terrorist group, unscrupulous company, or rogue state to recreate one of humanity's most feared microbial enemies. The reemergence of smallpox-because of a laboratory accident or an intentional release-would be a global health disaster. International organizations, national governments, the DNA synthesis industry, and the synthetic biology community all have a role to play in devising new approaches to preventing the reemergence of

  10. Smallpox: A Review for Health Educators

    Science.gov (United States)

    Bungum, Timothy J.

    2003-01-01

    Since the declaration of the eradication of smallpox in May of 1980 concern about this virus has ebbed. However, recent world events, including the destabilization of governments, have raised concerns that smallpox could fall into the hands of nefarious individuals or groups who might attempt to use the virus as a weapon. In Centers for Disease…

  11. The Smallpox Threat: The School Nurse's Role

    Science.gov (United States)

    Martin, Mary E.; Didion, Judy

    2003-01-01

    Today, with the threat of bioterrorism and war, there is a new dimension to the traditional role of the school nurse. The smallpox threat to public health will invoke the school nurse's role as an educator, liaison, and consultant in the community. This article discusses smallpox, the vaccination process, adverse effects, and postvaccination care.…

  12. Smallpox: a disease of the past? Consideration for midwives.

    Science.gov (United States)

    Constantin, Carolyn M; Martinelli, Angela M; Foster, Stanley O; Bonney, Elizabeth A; Strickland, Ora L

    2003-01-01

    Smallpox infection was often more severe in pregnant women than in non-pregnant women or in men, regardless of vaccination status. Women with smallpox infection during pregnancy have higher rates of abortions, stillbirths, and preterm deliveries than women without the disease. Pregnant women have high incidences of hemorrhagic-type and flat-type smallpox, which are associated with extremely high fatality rates. Although smallpox was eradicated in the late 1970s, current international concern exists regarding the potential use of smallpox virus as an agent for bioterrorism. This manuscript reviews clinical aspects of smallpox, smallpox immunization, and outcomes in pregnant women.

  13. 17(th) Century Variola Virus Reveals the Recent History of Smallpox.

    Science.gov (United States)

    Duggan, Ana T; Perdomo, Maria F; Piombino-Mascali, Dario; Marciniak, Stephanie; Poinar, Debi; Emery, Matthew V; Buchmann, Jan P; Duchêne, Sebastian; Jankauskas, Rimantas; Humphreys, Margaret; Golding, G Brian; Southon, John; Devault, Alison; Rouillard, Jean-Marie; Sahl, Jason W; Dutour, Olivier; Hedman, Klaus; Sajantila, Antti; Smith, Geoffrey L; Holmes, Edward C; Poinar, Hendrik N

    2016-12-19

    Smallpox holds a unique position in the history of medicine. It was the first disease for which a vaccine was developed and remains the only human disease eradicated by vaccination. Although there have been claims of smallpox in Egypt, India, and China dating back millennia [1-4], the timescale of emergence of the causative agent, variola virus (VARV), and how it evolved in the context of increasingly widespread immunization, have proven controversial [4-9]. In particular, some molecular-clock-based studies have suggested that key events in VARV evolution only occurred during the last two centuries [4-6] and hence in apparent conflict with anecdotal historical reports, although it is difficult to distinguish smallpox from other pustular rashes by description alone. To address these issues, we captured, sequenced, and reconstructed a draft genome of an ancient strain of VARV, sampled from a Lithuanian child mummy dating between 1643 and 1665 and close to the time of several documented European epidemics [1, 2, 10]. When compared to vaccinia virus, this archival strain contained the same pattern of gene degradation as 20(th) century VARVs, indicating that such loss of gene function had occurred before ca. 1650. Strikingly, the mummy sequence fell basal to all currently sequenced strains of VARV on phylogenetic trees. Molecular-clock analyses revealed a strong clock-like structure and that the timescale of smallpox evolution is more recent than often supposed, with the diversification of major viral lineages only occurring within the 18(th) and 19(th) centuries, concomitant with the development of modern vaccination. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

  14. STAINING OF VACCINIA ANTIGEN BY IMMUNOURANIUM TECHNIQUE,

    Science.gov (United States)

    An attempt to follow morphologically the development of vaccinia antigen in helium-lanthanum ( HeLa ) cells is reported. The conversion of rabbit...antisera to vaccinia virus and the preparation of vaccinia-infected HeLa cells for electron microscopy are described. With specific staining, viral

  15. [Smallpox is a dormant volcano].

    Science.gov (United States)

    L'vov, D K; Zverev, V V; Gintsburg, A L; Marennikova, S S; Pal'tsev, M A

    2008-01-01

    The presence of rodent-associated natural foci containing at least 6 of the known 11 viruses belonging to the genus Orthopoxvirus (Poxviridae, Chordopoxvirinae) within the equatorial, tropical, subtropical, temperate, and subarctic climatic zones; the increasing aggravation of the monkey pox epidemic situation in equatorial Africa with an increase in human mortality by an average of 9.8% with a possibility of 2 to 8 passages in 30-70% of patients; the possible persistence of a virus in the human cadavers buried in the permafrost of Eurasia and America; bioterrorism threat due to the unaccounted viral reserves persisting somewhere or somebody; no postvaccinal human immunity since vaccination and vaccine manufacture stopped 30 years ago as recommended by the WHO, make the risk of the deteriorating epidemic situation with disastrous effects greater now and in the foreseeable future than it was 20-30 years ago. Health care academic circles and bodies do not know methods for rapid diagnosis in the field conditions of species-specific identification smallpox virus or preventive (low-reactogenic, effective vaccines, and those accessible for mass production) and therapeutic (nontoxic drugs, those satisfactory for mass production, inexpedient, effective ones when orally used) agents. Basic studies of biodiversity, functional properties of viral DNA and proteins, pathogenesis, and evolution are required. Live smallpox virus should be used at certain and particularly final stages for these studies that are of scientific and applied significance.

  16. Chronic inhibition of cyclooxygenase-2 attenuates antibody responses against vaccinia infection.

    Science.gov (United States)

    Bernard, Matthew P; Bancos, Simona; Chapman, Timothy J; Ryan, Elizabeth P; Treanor, John J; Rose, Robert C; Topham, David J; Phipps, Richard P

    2010-02-01

    Generation of optimal humoral immunity to vaccination is essential to protect against devastating infectious agents such as the variola virus that causes smallpox. Vaccinia virus (VV), employed as a vaccine against smallpox, provides an important model of infection. Herein, we evaluated the importance cyclooxygenase-2 (Cox-2) in immunity to VV using Cox-2 deficient mice and Cox-2 selective inhibitory drugs. The effects of Cox-2 inhibition on antibody responses to live viruses such as vaccinia have not been previously described. Here, we used VV infection in Cox-2 deficient mice and in mice chronically treated with Cox-2 selective inhibitors and show that the frequency of VV-specific B cells was reduced, as well as the production of neutralizing IgG. VV titers were approximately 70 times higher in mice treated with a Cox-2 selective inhibitor. Interestingly, Cox-2 inhibition also reduced the frequency of IFN-gamma producing CD4(+) T helper cells, important for class switching. The significance of these results is that the chronic use of non-steroidal anti-inflammatory drugs (NSAIDs), and other drugs that inhibit Cox-2 activity or expression, blunt the ability of B cells to produce anti-viral antibodies, thereby making vaccines less effective and possibly increasing susceptibility to viral infection. These new findings support an essential role for Cox-2 in regulating humoral immunity.

  17. Engineering the vaccinia virus L1 protein for increased neutralizing antibody response after DNA immunization

    Directory of Open Access Journals (Sweden)

    Moss Bernard

    2009-03-01

    Full Text Available Abstract Background The licensed smallpox vaccine, comprised of infectious vaccinia virus, has associated adverse effects, particularly for immunocompromised individuals. Therefore, safer DNA and protein vaccines are being investigated. The L1 protein, a component of the mature virion membrane that is conserved in all sequenced poxviruses, is required for vaccinia virus entry into host cells and is a target for neutralizing antibody. When expressed by vaccinia virus, the unglycosylated, myristoylated L1 protein attaches to the viral membrane via a C-terminal transmembrane anchor without traversing the secretory pathway. The purpose of the present study was to investigate modifications of the gene expressing the L1 protein that would increase immunogenicity in mice when delivered by a gene gun. Results The L1 gene was codon modified for optimal expression in mammalian cells and potential N-glycosylation sites removed. Addition of a signal sequence to the N-terminus of L1 increased cell surface expression as shown by confocal microscopy and flow cytometry of transfected cells. Removal of the transmembrane domain led to secretion of L1 into the medium. Induction of binding and neutralizing antibodies in mice was enhanced by gene gun delivery of L1 containing the signal sequence with or without the transmembrane domain. Each L1 construct partially protected mice against weight loss caused by intranasal administration of vaccinia virus. Conclusion Modifications of the vaccinia virus L1 gene including codon optimization and addition of a signal sequence with or without deletion of the transmembrane domain can enhance the neutralizing antibody response of a DNA vaccine.

  18. Recommendations for using smallpox vaccine in a pre-event vaccination program. Supplemental recommendations of the Advisory Committee on Immunization Practices (ACIP) and the Healthcare Infection Control Practices Advisory Committee (HICPAC).

    Science.gov (United States)

    Wharton, Melinda; Strikas, Raymond A; Harpaz, Rafael; Rotz, Lisa D; Schwartz, Benjamin; Casey, Christine G; Pearson, Michele L; Anderson, Larry J

    2003-04-01

    This report supplements the 2001 statement by the Advisory Committee on Immunization Practices (ACIP) (CDC. Vaccinia [smallpox] vaccine: recommendations of the Advisory Committee on Immunization Practices [ACIP], 2001. MMWR 2001;50[No. RR-10]:1-25). This supplemental report provides recommendations for using smallpox vaccine in the pre-event vaccination program in the United States. To facilitate preparedness and response, smallpox vaccination is recommended for persons designated by public health authorities to conduct investigation and follow-up of initial smallpox cases that might necessitate direct patient contact. ACIP recommends that each state and territory establish and maintain > or = 1 smallpox response team. ACIP and the Healthcare Infection Control Practices Advisory Committee (HICPAC) recommend that each acute-care hospital identify health-care workers who can be vaccinated and trained to provide direct medical care for the first smallpox patients requiring hospital admission and to evaluate and manage patients who are suspected as having smallpox. When feasible, the first-stage vaccination program should include previously vaccinated health-care personnel to decrease the potential for adverse events. Additionally persons administering smallpox vaccine in this pre-event vaccination program should be vaccinated. Smallpox vaccine is administered by using the multiple-puncture technique with a bifurcated needle, packaged with the vaccine and diluent. According to the product labeling, 2-3 punctures are recommended for primary vaccination and 15 punctures for revaccination. A trace of blood should appear at the vaccination site after 15-20 seconds; if no trace of blood is visible, an additional 3 insertions should be made by using the same bifurcated needle without reinserting the needle into the vaccine vial. If no evidence of vaccine take is apparent after 7 days, the person can be vaccinated again. Optimal infection-control practices and appropriate

  19. Poxviruses: smallpox vaccine, its complications and chemotherapy

    Directory of Open Access Journals (Sweden)

    Mimi Remichkova

    2010-04-01

    Full Text Available Mimi RemichkovaDepartment of Pathogenic Bacteria, The Stephan Angeloff Institute of Microbiology, Bulgarian Academy of Sciences, Sofia, BulgariaAbstract: The threat of bioterrorism in the recent years has once again posed to mankind the unresolved problems of contagious diseases, well forgotten in the past. Smallpox (variola is among the most dangerous and highly contagious viral infections affecting humans. The last natural case in Somalia marked the end of a successful World Health Organization campaign for smallpox eradication by vaccination on worldwide scale. Smallpox virus still exists today in some laboratories, specially designated for that purpose. The contemporary response in the treatment of the post-vaccine complications, which would occur upon enforcing new programs for mass-scale smallpox immunization, includes application of effective chemotherapeutics and their combinations. The goals are to provide the highest possible level of protection and safety of the population in case of eventual terrorist attack. This review describes the characteristic features of the poxviruses, smallpox vaccination, its adverse reactions, and poxvirus chemotherapy.Keywords: poxvirus, smallpox vaccine, post vaccine complications, inhibitors

  20. Smallpox

    Science.gov (United States)

    ... samples saved for government research. Researchers continue to debate whether or not to kill the last remaining ... biological weapon It is unknown how long past vaccinations stay effective. People who received the vaccine many ...

  1. Smallpox

    Science.gov (United States)

    ... of Award Clinical Terms of Award Restriction for China Clinical Terms Guidance Compliance Sample Letter Inclusion Codes ... Division of AIDS Division of Allergy, Immunology, and Transplantation Division of Microbiology and Infectious Diseases Division of ...

  2. Evaluating anti-Orthopoxvirus antibodies in individuals from Brazilian rural areas prior to the bovine vaccinia era

    Directory of Open Access Journals (Sweden)

    Poliana de Oliveira Figueiredo

    2015-09-01

    Full Text Available Vaccinia virus naturally circulates in Brazil and is the causative agent of a zoonotic disease known as bovine vaccinia (BV. We retrospectively evaluated two populations from the Amazon and Southeast Regions. BV outbreaks had not been reported in these regions before sample collection. Neutralising antibodies were found in 13 individuals (n = 132 with titres ranging from 100 ≥ 6,400 neutralising units/mL. Univariate analysis identified age and vaccination as statistically significant risk factors in individuals from the Southeast Region. The absence of detectable antibodies in vaccinated individuals raises questions about the protection of smallpox vaccine years after vaccination and reinforces the need for surveillance of Orthopoxvirus in Brazilian populations without evidence of previous outbreaks.

  3. Potent T cell Responses Induced by Single DNA Vaccine Boosted with Recombinant Vaccinia Vaccine

    Institute of Scientific and Technical Information of China (English)

    Lianxing Liu; Chao Qiu; Yang Huang; Jianqing Xu; Yiming Shao

    2013-01-01

    Plasmid DNA,an effective vaccine vector,can induce both cellular and humoral immune responses.However,plasmid DNA raises issues concerning potential genomic integration after injection.This issue should be considered in preclinical studies.Tiantan vaccinia virus (TV) has been most widely utilized in eradicating smallpox in China.This virus has also been considered as a successful vaccine vector against a few infectious diseases.Potent T cell responses through T-cell receptor (TCR) could be induced by three injections of the DNA prime vaccine followed by a single injection of recombinant vaccinia vaccine.To develop a safer immunization strategy,a single DNA prime followed by a single recombinant Tiantan vaccinia (rTV) AIDS vaccine was used to immunize mice.Our data demonstrated that one DNA prime/rTV boost regimen induced mature TCR activation with high functional avidity,preferential T cell Vβ receptor usage and high sensitivity to anti-CD3 antibody stimulation.No differences in T cell responses were observed among one,two or three DNA prime/rTV boost regimens.This study shows that one DNA prime/rTV boost regimen is sufficient to induce potent T cell responses against HIV.

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

    Directory of Open Access Journals (Sweden)

    Susan A Holechek

    Full Text Available 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-γ 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-γ in the serum. These results suggest that protection provided by IFN-γ expressed by VACV may be mediated by its immunoregulatory activities rather than its antiviral effects. These results highlight the importance of IFN-γ 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.

  5. Real-Time PCR Assay To Detect Smallpox Virus

    Science.gov (United States)

    Sofi Ibrahim, M.; Kulesh, David A.; Saleh, Sharron S.; Damon, Inger K.; Esposito, Joseph J.; Schmaljohn, Alan L.; Jahrling, Peter B.

    2003-01-01

    We developed a highly sensitive and specific assay for the rapid detection of smallpox virus DNA on both the Smart Cycler and LightCycler platforms. The assay is based on TaqMan chemistry with the orthopoxvirus hemagglutinin gene used as the target sequence. With genomic DNA purified from variola virus Bangladesh 1975, the limit of detection was estimated to be approximately 25 copies on both machines. The assay was evaluated in a blinded study with 322 coded samples that included genomic DNA from 48 different isolates of variola virus; 25 different strains and isolates of camelpox, cowpox, ectromelia, gerbilpox, herpes, monkeypox, myxoma, rabbitpox, raccoonpox, skunkpox, vaccinia, and varicella-zoster viruses; and two rickettsial species at concentrations mostly ranging from 100 fg/μl to 1 ng/μl. Contained within those 322 samples were variola virus DNA, obtained from purified viral preparations, at concentrations of 1 fg/μl to 1 ng/μl. On the Smart Cycler platform, 2 samples with false-positive results were detected among the 116 samples not containing variola virus tested; i.e., the overall specificity of the assay was 98.3%. On the LightCycler platform, five samples with false-positive results were detected (overall specificity, 95.7%). Of the 206 samples that contained variola virus DNA ranging in concentrations from 100 fg/μl to 1 ng/μl, 8 samples were considered negative on the Smart Cycler platform and 1 sample was considered negative on the LightCycler platform. Thus, the clinical sensitivities were 96.1% for the Smart Cycler instrument and 99.5% for the LightCycler instrument. The vast majority of these samples were derived from virus-infected cell cultures and variola virus-infected tissues; thus, the DNA material contained both viral DNA and cellular DNA. Of the 43 samples that contained purified variola virus DNA ranging in concentration from 1 fg/μl to 1 ng/μl, the assay correctly detected the virus in all 43 samples on both the Smart Cycler

  6. Analysis of the complete genome of smallpox variola major virus strain Bangladesh-1975.

    Science.gov (United States)

    Massung, R F; Liu, L I; Qi, J; Knight, J C; Yuran, T E; Kerlavage, A R; Parsons, J M; Venter, J C; Esposito, J J

    1994-06-01

    We analyzed the 186,102 base pairs (bp) that constitute the entire DNA genome of a highly virulent variola virus isolated from Bangladesh in 1975. The linear, double-stranded molecule has relatively small (725 bp) inverted terminal repeat (ITR) sequences containing three 69-bp direct repeat elements, a 54-bp partial repeat element, and a 105-base telomeric end-loop that can be maximally base-paired to contain 17 mismatches. Proximal to the right-end ITR sequences are another seven 69-bp elements and a 53- and a 27-bp partial element. Sequence analysis showed 187 closely spaced open reading frames specifying putative major proteins containing > or = 65 amino acids. Most of the virus proteins correspond to proteins in current databases, including 150 proteins that have > 90% identity to major gene products encoded by vaccinia virus, the smallpox vaccine. Variola virus has a group of proteins that are truncated compared with vaccinia virus counterparts and a smaller group of proteins that are elongated. The terminal regions encode several novel proteins and variants of other poxvirus proteins that potentially augment variola virus transmissibility and virulence for its only natural host, humans.

  7. An Attenuated LC16m8 Smallpox Vaccine: Analysis of Full-Genome Sequence and Induction of Immune Protection§

    Science.gov (United States)

    Morikawa, Shigeru; Sakiyama, Tokuki; Hasegawa, Hideki; Saijo, Masayuki; Maeda, Akihiko; Kurane, Ichiro; Maeno, Go; Kimura, Junko; Hirama, Chie; Yoshida, Teruhiko; Asahi-Ozaki, Yasuko; Sata, Tetsutaro; Kurata, Takeshi; Kojima, Asato

    2005-01-01

    The potential threat of smallpox bioterrorism has made urgent the development of lower-virulence vaccinia virus vaccines. An attenuated LC16m8 (m8) vaccine was developed in 1975 from the Lister strain used in the World Health Organization smallpox eradication program but was not used against endemic smallpox. Today, no vaccines can be tested with variola virus for efficacy in humans, and the mechanisms of immune protection against the major intracellular mature virion (IMV) and minor extracellular enveloped virion (EEV) populations of poxviruses are poorly understood. Here, we determined the full-genome sequences of the m8, parental LC16mO (mO), and grandparental Lister (LO) strains and analyzed their evolutionary relationships. Sequence data and PCR analysis indicated that m8 was a progeny of LO and that m8 preserved almost all of the open reading frames of vaccinia virus except for the disrupted EEV envelope gene B5R. In accordance with this genomic background, m8 induced 100% protection against a highly pathogenic vaccinia WR virus in mice by a single vaccination, despite the lack of anti-B5R and anti-EEV antibodies. The immunogenicity and priming efficacy with the m8 vaccine consisting mainly of IMV were as high as those with the intact-EEV parental mO and grandparental LO vaccines. Thus, mice vaccinated with 107 PFU of m8 produced low levels of anti-B5R antibodies after WR challenge, probably because of quick clearance of B5R-expressing WR EEV by strong immunity induced by the vaccination. These results suggest that priming with m8 IMV provides efficient protection despite undetectable levels of immunity against EEV. PMID:16140764

  8. One time intranasal vaccination with a modified vaccinia Tiantan strain MVTT(ZCI) protects animals against pathogenic viral challenge.

    Science.gov (United States)

    Yu, Wenbo; Fang, Qing; Zhu, Weijun; Wang, Haibo; Tien, Po; Zhang, Linqi; Chen, Zhiwei

    2010-02-25

    To combat variola virus in bioterrorist attacks, it is desirable to develop a noninvasive vaccine. Based on the vaccinia Tiantan (VTT) strain, which was historically used to eradicate the smallpox in China, we generated a modified VTT (MVTT(ZCI)) by removing the hemagglutinin gene and an 11,944bp genomic region from HindIII fragment C2L to F3L. MVTT(ZCI) was characterized for its host cell range in vitro and preclinical safety and efficacy profiles in mice. Despite replication-competency in some cell lines, unlike VTT, MVTT(ZCI) did not cause death after intracranial injection or body weight loss after intranasal inoculation. MVTT(ZCI) did not replicate in mouse brain and was safe in immunodeficient mice. MVTT(ZCI) induced neutralizing antibodies via the intranasal route of immunization. One time intranasal immunization protected animals from the challenge of the pathogenic vaccinia WR strain. This study established proof-of-concept that the attenuated replicating MVTT(ZCI) may serve as a safe noninvasive smallpox vaccine candidate.

  9. Comparison of the replication characteristics of vaccinia virus strains Guang 9 and Tian Tan in vivo and in vitro.

    Science.gov (United States)

    Zhu, Rong; Liu, Qiang; Huang, Weijin; Yu, Yongxin; Wang, Youchun

    2014-10-01

    Vaccinia virus is widely used as a vector in the development of recombinant vaccines. Vaccinia virus strain Guang 9 (VG9), which was derived from vaccinia virus strain Tian Tan (VTT) by successive plaque-cloning purification, was more attenuated than VTT. In this study, the host cell range and the growth and replication of VG9 were compared with those of VTT. The results showed that both VG9 and VTT could infect permissive cells (Vero, TK-143 and CEF) and semipermissive cells PK (15) and induced a visible cytopathic effect (CPE). Both strains could infect nonpermissive CHO-K1 cells but neither was able to reproduce. The replicative ability of VG9 was a little lower than that of VTT. Additionally, recombinant vaccinia viruses containing a firefly luciferase gene (VG9-L and VTT-L) were constructed, and their expression in vitro and replication and spread in vivo were compared. The expression ability of VG9-L was lower than that of VTT-L. Whole-animal imaging data indicated that VG9-L could reproduce quickly and express the exogenous protein at the site of inoculation, regardless of whether the intramuscular, intracutaneous, subcutaneous or celiac inoculation route was used. VG9-L was better in its ability to express a foreign protein than VTT-L, but the time during which expression occurred was shorter. There was no dissemination of virus in mice inoculated with either strain. In summary, this study demonstrates the possibility of using VG9 for the production of smallpox vaccines or the construction of recombinant vaccinia virus vaccines.

  10. Frequently Asked Questions and Answers on Smallpox

    Science.gov (United States)

    ... protect anyone else judged to have a high risk of exposure to smallpox. The vaccine cannot be used in ... only to those persons who have a high risk of coming into contact with the virus which ... health workers and hospital personnel around the world have been trained to ...

  11. Demographic and clinical factors associated with response to smallpox vaccine in preimmunized volunteers.

    Directory of Open Access Journals (Sweden)

    Philippe Bossi

    Full Text Available CONTEXT: In March 2003, the French Ministry of Health implemented a program on preparedness and response to a biological attack using smallpox as weapon. This program included the establishment of a preoutbreak national team that could be revaccinated against smallpox. OBJECTIVE: To identify demographic and clinical factors associated with vaccination success defined as the presence of a pustule at the inoculation site at day 8 (days 7-9, with an undiluted vaccinia virus derived from a Lister strain among preimmunized volunteers. VOLUNTEERS AND METHODS: From March 2003 to November 2006, we have studied prospectively 226 eligible volunteers. Demographic data were recorded for each volunteer (age, sex, number of previously smallpox vaccinations and date of the last vaccination. Smallpox vaccine adverse reactions were diagnosed on the basis of clinical examination performed at days 0, 7, 14, 21 and 28 after revaccination. RESULTS: A total of 226 volunteers (sex ratio H/F = 2.7 were revaccinated. Median age was 45 years (range: 27-63 yrs. All volunteers completed follow-up. Median number of vaccinations before revaccination was 2 (range: 1-8. The median delay between time of the study and the last vaccination was 29 years (range; 18-60 yrs. Sixty-one volunteers (27% experienced one (n = 40 or more (n = 21 minor side effects during the 2-14 days after revaccination. Successful vaccination was noted in 216/226 volunteers (95.6% at day 8 and the median of the pustule diameter was 5 mm (range: 1-20 mm. Size of the pustule at day 8 was correlated with age (p = 0.03 and with the presence of axillary adenopathy after revaccination (p = 0.007. Sex, number of prior vaccinations, delay between the last vaccination and revaccination, and local or systemic side effects with the exception of axillary adenopathy, were not correlated with the size of the pustule at day 8. CONCLUSIONS: Previously vaccinated volunteers can be successfully revaccinated with the

  12. Medical Surveillance Monthly Report (MSMR). Volume 21, Number 8, August 2014

    Science.gov (United States)

    2014-08-01

    Greenberg M. Complications of vaccination against smallpox. Am J Dis Child . 1948;76:492– 502. 2. Leake JP. Question and answers on smallpox vaccination ...documented.12,13 Th e only smallpox vaccine currently available for use in the U.S. is ACAM2000®, which was approved by the Food and Drug...Force train- ing installation, select technical training students receive the smallpox vaccination in accordance with current Department of Defense

  13. Smallpox vaccination and all-cause infectious disease hospitalization

    DEFF Research Database (Denmark)

    Sørup, Signe; Villumsen, Marie; Ravn, Henrik

    2011-01-01

    There is growing evidence from observational studies and randomized trials in low-income countries that vaccinations have non-specific effects. Administration of live vaccines reduces overall child morbidity and mortality, presumably due to protection against non-targeted infections. In Denmark......, the live vaccine against smallpox was phased out in the 1970s due to the eradication of smallpox. We used the phasing-out period to investigate the effect of smallpox vaccination on the risk of hospitalization for infections....

  14. Comparative Proteomics of Human Monkeypox and Vaccinia Intracellular Mature and Extracellular Enveloped Virions

    Energy Technology Data Exchange (ETDEWEB)

    Manes, Nathan P.; Estep, Ryan D.; Mottaz, Heather M.; Moore, Ronald J.; Clauss, Therese RW; Monroe, Matthew E.; Du, Xiuxia; Adkins, Joshua N.; Wong, Scott; Smith, Richard D.

    2008-03-07

    Orthopoxviruses are the largest and most complex of the animal viruses. In response to the recent emergence of monkeypox in Africa and the threat of smallpox bioterrorism, virulent (monkeypox virus) and benign (vaccinia virus) orthopoxviruses were proteomically compared with the goal of identifying proteins required for pathogenesis. Orthopoxviruses were grown in HeLa cells to two different viral forms (intracellular mature virus and extracellular enveloped virus), purified by sucrose gradient ultracentrifugation, denatured using RapiGest™ surfactant, and digested with trypsin. Unfractionated samples and strong cation exchange HPLC fractions were analyzed by reversed-phase LC-MS/MS, and analyses of the MS/MS spectra using SEQUEST® and X! Tandem resulted in the identification of hundreds of monkeypox, vaccinia, and copurified host proteins. The unfractionated samples were additionally analyzed by LC-MS on an LTQ-Orbitrap™, and the accurate mass and elution time tag approach was used to perform quantitative comparisons. Possible pathophysiological roles of differentially expressed orthopoxvirus genes are discussed.

  15. 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.

    Directory of Open Access Journals (Sweden)

    Clement A Meseda

    Full Text Available 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.

  16. How long ago did smallpox virus emerge?

    Science.gov (United States)

    Shchelkunov, Sergei N

    2009-01-01

    Unlike vertebrates, for which paleontological data are available, and RNA viruses, which display a high rate of genetic variation, an objective estimate of time parameters for the molecular evolution of DNA viruses, which display a low rate of accumulation of mutations, is a complex problem. Genomic studies of a set of smallpox (variola) virus (VARV) isolates demonstrated the patterns of phylogenetic relationships between geographic variants of this virus. Using archival data on smallpox outbreaks and the results of phylogenetic analyses of poxvirus genomes, different research teams have obtained contradictory data on the possible time point of VARV origin. I discuss the approaches used for dating of VARV evolution and adduce the arguments favoring its historically recent origin.

  17. Smallpox: Is the Department of Defense Prepared?

    Science.gov (United States)

    2003-01-01

    medications ( cidofovir , adefovir dipivoxil, cyclic cidofivir, and ribavirin) show in vitro activity against variola virus and are undergoing further testing...further treatment including VIG or cidofovir . Dose Related Effects of Smallpox Vaccine (Frey 2002) provides data from a dilution study of Dryvax...antiviral medications. Thus far, in vitro testing has shown the drug cidofovir to have activity against variola virus and makes it the leading antiviral

  18. Smallpox: emergence, global spread, and eradication.

    Science.gov (United States)

    Fenner, F

    1993-01-01

    Speculatively, it is suggested that variola virus, the cause of smallpox, evolved from an orthopoxvirus of animals of the central African rain forests (possibly now represented by Tatera poxvirus), some thousands of years ago, and first became established as a virus specific for human beings in the dense populations of the Nile valley perhaps five thousand years ago. By the end of the first millennium of the Christian era, it had spread to all the densely populated parts of the Eurasian continent and along the Mediterranean fringe of north Africa. It became established in Europe during the times of the Crusades. The great voyages of European colonization carried smallpox to the Americas and to Africa south of the Sahara. Transported across the Atlantic by Europeans and their African slaves, it played a major role in the conquest of Mexico and Peru and the European settlement of north America. Variolation, an effective preventive inoculation, was devised as early as the tenth century. In 1798 this practice was supplanted by Jenner's cowpox vaccine. In 1967, when the disease was still endemic in 31 countries and caused ten to fifteen million cases and about two million deaths annually, the World Health Organization embarked on a programme that was to see the disease eradicated globally just over ten years later, and the world was formally declared to be free of smallpox in May 1980. Smallpox is unique--a specifically human disease that emerged from some animal reservoir, spread to become a worldwide, severe and almost universal affliction, and finally underwent the reverse process to emergence, namely global eradication.

  19. Vaccinia Virus LC16m8∆ as a Vaccine Vector for Clinical Applications

    Directory of Open Access Journals (Sweden)

    Minoru Kidokoro

    2014-10-01

    Full Text Available The LC16m8 strain of vaccinia virus, the active ingredient in the Japanese smallpox vaccine, was derived from the Lister/Elstree strain. LC16m8 is replication-competent and has been administered to over 100,000 infants and 3,000 adults with no serious adverse reactions. Despite this outstanding safety profile, the occurrence of spontaneously-generated large plaque-forming virulent LC16m8 revertants following passage in cell culture is a major drawback. We identified the gene responsible for the reversion and deleted the gene (B5R from LC16m8 to derive LC16m8Δ. LC16m8∆ is non-pathogenic in immunodeficient severe combined immunodeficiency (SCID mice, genetically-stable and does not reverse to a large-plaque phenotype upon passage in cell culture, even under conditions in which most LC16m8 populations are replaced by revertants. Moreover, LC16m8∆ is >500-fold more effective than the non-replicating vaccinia virus (VV, Modified Vaccinia Ankara (MVA, at inducing murine immune responses against pathogenic VV. LC16m8∆, which expresses the SIV gag gene, also induced anti-Gag CD8+ T-cells more efficiently than MVA and another non-replicating VV, Dairen I minute-pock variants (DIs. Moreover, LC16m8∆ expressing HIV-1 Env in combination with a Sendai virus vector induced the production of anti-Env antibodies and CD8+ T-cells. Thus, the safety and efficacy of LC16m8∆ mean that it represents an outstanding platform for the development of human vaccine vectors.

  20. Frequency of Adverse Events after Vaccination with Different Vaccinia Strains.

    NARCIS (Netherlands)

    Kretzschmar, Mirjam; Wallinga, Jacco; Teunis, Peter F M; Xing, Shuqin; Mikolajczyk, Rafael

    2006-01-01

    BACKGROUND: Large quantities of smallpox vaccine have been stockpiled to protect entire nations against a possible reintroduction of smallpox. Planning for an appropriate use of these stockpiled vaccines in response to a smallpox outbreak requires a rational assessment of the risks of vaccination-re

  1. Frequency of Adverse Events after Vaccination with Different Vaccinia Strains.

    NARCIS (Netherlands)

    Kretzschmar, Mirjam; Wallinga, Jacco; Teunis, Peter F M; Xing, Shuqin; Mikolajczyk, Rafael

    2006-01-01

    BACKGROUND: Large quantities of smallpox vaccine have been stockpiled to protect entire nations against a possible reintroduction of smallpox. Planning for an appropriate use of these stockpiled vaccines in response to a smallpox outbreak requires a rational assessment of the risks of vaccination-re

  2. Smallpox epidemic in a Brazilian community.

    Science.gov (United States)

    de Azeredo Costa, E; Morris, L

    1975-06-01

    The most severe outbreak of smallpox in Brazil in 1969 took place in the municipio of Utinga, Bahia. Of 507 cases, 246 occurred in residents of the urban area of the municipio. This paper describes the findings of the detailed study of the urban area outbreak. An inverse relationship between age and susceptibility was evident; fully 91% of the children less than 5 years of age as well as 75% of the school-age children had no history of smallpox and had never been vaccinated. The overall attack rate was 11% with nearly three-fourths of the cases in children under 15 years of age. Only 4 of the 246 patients had a vaccination scar, and the estimate of vaccine efficacy was 94%. The continued occurrence of cases after the initial vaccination containment measures prompted detailed studies during 2 follow-up visits to the community which demonstrated that 1) transmission of smallpox may continue in a pocket of susceptibles although the overall immune status of a community is high following containment measures, and 2) clinical illness may have been aborted in those affected individuals who were only in their second to fourth day of incubation at the time of vaccination.

  3. Prospective surveillance for cardiac adverse events in healthy adults receiving modified vaccinia Ankara vaccines: a systematic review.

    Directory of Open Access Journals (Sweden)

    Marnie L Elizaga

    Full Text Available 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

  4. Smallpox-Related Knowledge and Beliefs among Recent College Graduates

    Science.gov (United States)

    Bungum, Timothy; Day, Charlene

    2006-01-01

    Recent world events have increased concern and preparations for possible bioterror events. Despite worldwide efforts to limit access to bio-weapons, smallpox is still considered a potential bioterror threat. Americans' understanding of smallpox could prevent panic and enhance the willingness of citizens to receive vaccinations. Objective: The…

  5. Critical role of perforin-dependent CD8+ T cell immunity for rapid protective vaccination in a murine model for human smallpox.

    Directory of Open Access Journals (Sweden)

    Melanie Kremer

    Full Text Available Vaccination is highly effective in preventing various infectious diseases, whereas the constant threat of new emerging pathogens necessitates the development of innovative vaccination principles that also confer rapid protection in a case of emergency. Although increasing evidence points to T cell immunity playing a critical role in vaccination against viral diseases, vaccine efficacy is mostly associated with the induction of antibody responses. Here we analyze the immunological mechanism(s of rapidly protective vaccinia virus immunization using mousepox as surrogate model for human smallpox. We found that fast protection against lethal systemic poxvirus disease solely depended on CD4 and CD8 T cell responses induced by vaccination with highly attenuated modified vaccinia virus Ankara (MVA or conventional vaccinia virus. Of note, CD4 T cells were critically required to allow for MVA induced CD8 T cell expansion and perforin-mediated cytotoxicity was a key mechanism of MVA induced protection. In contrast, selected components of the innate immune system and B cell-mediated responses were fully dispensable for prevention of fatal disease by immunization given two days before challenge. In conclusion, our data clearly demonstrate that perforin-dependent CD8 T cell immunity plays a key role in MVA conferred short term protection against lethal mousepox. Rapid induction of T cell immunity might serve as a new paradigm for treatments that need to fit into a scenario of protective emergency vaccination.

  6. One more piece in the VACV ecological puzzle: could peridomestic rodents be the link between wildlife and bovine vaccinia outbreaks in Brazil?

    Directory of Open Access Journals (Sweden)

    Jônatas S Abrahão

    Full Text Available BACKGROUND: Despite 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 milkers. Little is known about VACV's natural hosts and its epidemiological and ecological characteristics. Although VACV was isolated and/or serologically detected in Brazilian wild animals, the link between wildlife and farms has not yet been elucidated. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we describe for the first time, to our knowledge, the isolation of a VACV (Mariana virus - MARV from a mouse during a BV outbreak. Genetic data, in association with biological assays, showed that this isolate was the same etiological agent causing exanthematic lesions observed in the cattle and human inhabitants of a particular BV-affected area. Phylogenetic analysis grouped MARV with other VACV isolated during BV outbreaks. CONCLUSION/SIGNIFICANCE: These data provide new biological and epidemiological information on VACV and lead to an interesting question: could peridomestic rodents be the link between wildlife and BV outbreaks?

  7. The effects of post-exposure smallpox vaccination on clinical disease presentation: addressing the data gaps between historical epidemiology and modern surrogate model data.

    Science.gov (United States)

    Keckler, M Shannon; Reynolds, Mary G; Damon, Inger K; Karem, Kevin L

    2013-10-25

    Decades after public health interventions - including pre- and post-exposure vaccination - were used to eradicate smallpox, zoonotic orthopoxvirus outbreaks and the potential threat of a release of variola virus remain public health concerns. Routine prophylactic smallpox vaccination of the public ceased worldwide in 1980, and the adverse event rate associated with the currently licensed live vaccinia virus vaccine makes reinstatement of policies recommending routine pre-exposure vaccination unlikely in the absence of an orthopoxvirus outbreak. Consequently, licensing of safer vaccines and therapeutics that can be used post-orthopoxvirus exposure is necessary to protect the global population from these threats. Variola virus is a solely human pathogen that does not naturally infect any other known animal species. Therefore, the use of surrogate viruses in animal models of orthopoxvirus infection is important for the development of novel vaccines and therapeutics. Major complications involved with the use of surrogate models include both the absence of a model that accurately mimics all aspects of human smallpox disease and a lack of reproducibility across model species. These complications limit our ability to model post-exposure vaccination with newer vaccines for application to human orthopoxvirus outbreaks. This review seeks to (1) summarize conclusions about the efficacy of post-exposure smallpox vaccination from historic epidemiological reports and modern animal studies; (2) identify data gaps in these studies; and (3) summarize the clinical features of orthopoxvirus-associated infections in various animal models to identify those models that are most useful for post-exposure vaccination studies. The ultimate purpose of this review is to provide observations and comments regarding available model systems and data gaps for use in improving post-exposure medical countermeasures against orthopoxviruses.

  8. A Multicenter, Open-Label, Controlled Phase II Study to Evaluate Safety and Immunogenicity of MVA Smallpox Vaccine (IMVAMUNE in 18-40 Year Old Subjects with Diagnosed Atopic Dermatitis.

    Directory of Open Access Journals (Sweden)

    Richard N Greenberg

    Full Text Available Replicating smallpox vaccines can cause severe complications in individuals with atopic dermatitis (AD. Prior studies evaluating Modified Vaccinia Ankara virus (MVA, a non-replicating vaccine in humans, showed a favorable safety and immunogenicity profile in healthy volunteers.This Phase II study compared the safety and immunogenicity of MVA enrolling groups of 350 subjects with AD (SCORAD ≤ 30 and 282 healthy subjects.Subjects were vaccinated twice with MVA, each dose given subcutaneously 4 weeks apart. Adverse events, cardiac parameters, and the development of vaccinia virus humoral immune responses were monitored.The overall safety of the vaccine was similar in both groups. Adverse events affecting skin were experienced significantly more often in subjects with AD, but the majority of these events were mild to moderate in intensity. Seroconversion rates and geometric mean titers for total and neutralizing vaccinia-specific antibodies in the AD group were non-inferior compared to the healthy subjects.The size of the study population limited the detection of serious adverse events occurring at a frequency less than 1%.MVA has a favorable safety profile and the ability to elicit vaccinia-specific immune responses in subjects with AD.ClinicalTrials.gov NCT00316602.

  9. A Multicenter, Open-Label, Controlled Phase II Study to Evaluate Safety and Immunogenicity of MVA Smallpox Vaccine (IMVAMUNE) in 18–40 Year Old Subjects with Diagnosed Atopic Dermatitis

    Science.gov (United States)

    Greenberg, Richard N; Hurley, Yadira; Dinh, Dinh V.; Mraz, Serena; Vera, Javier Gomez; von Bredow, Dorothea; von Krempelhuber, Alfred; Roesch, Siegfried; Virgin, Garth; Arndtz-Wiedemann, Nathaly; Meyer, Thomas Peter; Schmidt, Darja; Nichols, Richard; Young, Philip; Chaplin, Paul

    2015-01-01

    Background Replicating smallpox vaccines can cause severe complications in individuals with atopic dermatitis (AD). Prior studies evaluating Modified Vaccinia Ankara virus (MVA), a non-replicating vaccine in humans, showed a favorable safety and immunogenicity profile in healthy volunteers. Objective This Phase II study compared the safety and immunogenicity of MVA enrolling groups of 350 subjects with AD (SCORAD ≤ 30) and 282 healthy subjects. Methods Subjects were vaccinated twice with MVA, each dose given subcutaneously 4 weeks apart. Adverse events, cardiac parameters, and the development of vaccinia virus humoral immune responses were monitored. Results The overall safety of the vaccine was similar in both groups. Adverse events affecting skin were experienced significantly more often in subjects with AD, but the majority of these events were mild to moderate in intensity. Seroconversion rates and geometric mean titers for total and neutralizing vaccinia-specific antibodies in the AD group were non-inferior compared to the healthy subjects. Limitations The size of the study population limited the detection of serious adverse events occurring at a frequency less than 1%. Conclusion MVA has a favorable safety profile and the ability to elicit vaccinia-specific immune responses in subjects with AD. Trial Registration ClinicalTrials.gov NCT00316602 PMID:26439129

  10. Activity, specificity, and probe design for the smallpox virus protease K7L.

    Science.gov (United States)

    Aleshin, Alexander E; Drag, Marcin; Gombosuren, Naran; Wei, Ge; Mikolajczyk, Jowita; Satterthwait, Arnold C; Strongin, Alex Y; Liddington, Robert C; Salvesen, Guy S

    2012-11-16

    The K7L gene product of the smallpox virus is a protease implicated in the maturation of viral proteins. K7L belongs to protease Clan CE, which includes distantly related cysteine proteases from eukaryotes, pathogenic bacteria, and viruses. Here, we describe its recombinant high level expression, biochemical mechanism, substrate preference, and regulation. Earlier studies inferred that the orthologous I7L vaccinia protease cleaves at an AG-X motif in six viral proteins. Our data for K7L suggest that the AG-X motif is necessary but not sufficient for optimal cleavage activity. Thus, K7L requires peptides extended into the P7 and P8 positions for efficient substrate cleavage. Catalytic activity of K7L is substantially enhanced by homodimerization, by the substrate protein P25K as well as by glycerol. RNA and DNA also enhance cleavage of the P25K protein but not of synthetic peptides, suggesting that nucleic acids augment the interaction of K7L with its protein substrate. Library-based peptide preference analyses enabled us to design an activity-based probe that covalently and selectively labels K7L in lysates of transfected and infected cells. Our study thus provides proof-of-concept for the design of inhibitors and probes that may contribute both to a better understanding of the role of K7L in the virus life cycle and the design of novel anti-virals.

  11. An update on the vaccinia virus genome.

    Science.gov (United States)

    Johnson, G P; Goebel, S J; Paoletti, E

    1993-10-01

    This communication is intended as a single source update of the initial report (Goebel et al., 1990a,b) which described the complete DNA sequence of the vaccinia virus genome. We have integrated published information as well as unpublished data. Our understanding of the complexities of the genetic functional organization of poxviruses is increasing at a remarkable rate. While some previously unknown identities have since been elucidated, the fact that the majority of vaccinia-encoded gene products still lack assigned functions lends excitement to the immediate future of poxvirus research.

  12. On the origin of smallpox: correlating variola phylogenics with historical smallpox records.

    Science.gov (United States)

    Li, Yu; Carroll, Darin S; Gardner, Shea N; Walsh, Matthew C; Vitalis, Elizabeth A; Damon, Inger K

    2007-10-02

    Human disease likely attributable to variola virus (VARV), the etiologic agent of smallpox, has been reported in human populations for >2,000 years. VARV is unique among orthopoxviruses in that it is an exclusively human pathogen. Because VARV has a large, slowly evolving DNA genome, we were able to construct a robust phylogeny of VARV by analyzing concatenated single nucleotide polymorphisms (SNPs) from genome sequences of 47 VARV isolates with broad geographic distributions. Our results show two primary VARV clades, which likely diverged from an ancestral African rodent-borne variola-like virus either approximately 16,000 or approximately 68,000 years before present (YBP), depending on which historical records (East Asian or African) are used to calibrate the molecular clock. One primary clade was represented by the Asian VARV major strains, the more clinically severe form of smallpox, which spread from Asia either 400 or 1,600 YBP. Another primary clade included both alastrim minor, a phenotypically mild smallpox described from the American continents, and isolates from West Africa. This clade diverged from an ancestral VARV either 1,400 or 6,300 YBP, and then further diverged into two subclades at least 800 YBP. All of these analyses indicate that the divergence of alastrim and variola major occurred earlier than previously believed.

  13. Pathogenicity and immunogenicity of recombinant Tiantan Vaccinia Virus with deleted C12L and A53R genes.

    Science.gov (United States)

    Dai, Kaifan; Liu, Ying; Liu, Mingjie; Xu, Jianqing; Huang, Wei; Huang, Xianggang; Liu, Lianxing; Wan, Yanmin; Hao, Yanling; Shao, Yiming

    2008-09-15

    Interest is increasing regarding replicating poxvirus as HIV vaccine vector. In China, the Tiantan Vaccinia Virus (TV) has been used most extensively in the battle of eradicating smallpox. Recently, TV was developing as vaccine vector to fight against infectious diseases such as human immunodeficiency virus (HIV). However, replicating vaccinia virus sometimes may pose serious post-vaccination complications, especially in immunosuppressed individuals. To develop a safer and more effective TV-based vector, we constructed C12L (vIL-18 binding protein) and A53R (vTNF receptor homolog) gene-deleted mutants which are based on parental TV and VTKgpe (TV expressing HIV gagpol and env gene), respectively. The pathogenicity and immunogenicity were also evaluated. Deleting these two immunomodulatory genes lessened the virulence of the parental virus in both mice and rabbit models. Notably, C12L deletion mutant attenuated the skin virulence of parental virus by as high as approximate 2 logs. Furthermore, VTKgpe with A53R and C12L gene deletion retains the high immunogenicity of the parental virus to elicit strong humoral and cellular responses to the HIV target genes despite the remarkable attenuation. These data suggest that deletion of the cytokine viroceptor gene is feasible to obtain a safer and replication-competent TV vector for vaccination and immunotherapy.

  14. Protective Effect of Surfactant Protein D in Pulmonary Vaccinia Virus Infection: Implication of A27 Viral Protein

    Directory of Open Access Journals (Sweden)

    Julien Perino

    2013-03-01

    Full Text Available Vaccinia virus (VACV was used as a surrogate of variola virus (VARV (genus Orthopoxvirus, the causative agent of smallpox, to study Orthopoxvirus infection. VARV is principally transmitted between humans by aerosol droplets. Once inhaled, VARV first infects the respiratory tract where it could encounter surfactant components, such as soluble pattern recognition receptors. Surfactant protein D (SP-D, constitutively present in the lining fluids of the respiratory tract, plays important roles in innate host defense against virus infection. We investigated the role of SP-D in VACV infection and studied the A27 viral protein involvement in the interaction with SP-D. Interaction between SP-D and VACV caused viral inhibition in a lung cell model. Interaction of SP-D with VACV was mediated by the A27 viral protein. Binding required Ca2+ and interactions were blocked in the presence of excess of SP-D saccharide ligands. A27, which lacks glycosylation, directly interacted with SP-D. The interaction between SP-D and the viral particle was also observed using electron microscopy. Infection of mice lacking SP-D (SP-D-/- resulted in increased mortality compared to SP-D+/+ mice. Altogether, our data show that SP-D participates in host defense against the vaccinia virus infection and that the interaction occurs with the viral surface protein A27.

  15. Proteomic basis of the antibody response to monkeypox virus infection examined in cynomolgus macaques and a comparison to human smallpox vaccination.

    Directory of Open Access Journals (Sweden)

    Sarah Keasey

    Full Text Available 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 effective for monkeypox due to a high degree of sequence conservation. Yet, the identity of antigens within the monkeypox virus proteome contributing to immune responses has not been described in detail. We compared antibody responses to monkeypox virus infection and human smallpox vaccination by using a protein microarray covering 92-95% (166-192 proteins of representative proteomes from monkeypox viral clades of Central and West Africa, including 92% coverage (250 proteins of the vaccinia virus proteome as a reference orthopox vaccine. All viral gene clones were verified by sequencing and purified recombinant proteins were used to construct the microarray. Serum IgG of cynomolgus macaques that recovered from monkeypox recognized at least 23 separate proteins within the orthopox proteome, while only 14 of these proteins were recognized by IgG from vaccinated humans. There were 12 of 14 antigens detected by sera of human vaccinees that were also recognized by IgG from convalescent macaques. The greatest level of IgG binding for macaques occurred with the structural proteins F13L and A33R, and the membrane scaffold protein D13L. Significant IgM responses directed towards A44R, F13L and A33R of monkeypox virus were detected before onset of clinical symptoms in macaques. Thus, antibodies from vaccination recognized a small number of proteins shared with pathogenic virus strains, while recovery from infection also involved humoral responses to antigens uniquely recognized within the monkeypox virus proteome.

  16. In vitro transcription of a cloned vaccinia virus gene by a soluble extract prepared from vaccinia virus-infected HeLa cells.

    OpenAIRE

    Foglesong, P D

    1985-01-01

    Faithful transcription of a vaccinia virus gene was accomplished in vitro by using a soluble extract prepared from vaccinia virus-infected HeLa cells. Specific transcription of the cloned vaccinia virus gene was detected by using template DNA restricted within the transcribed region. The vaccinia virus gene was not transcribed by extracts prepared from uninfected HeLa cells even with supplementation by purified vaccinia virus RNA polymerase, nor was a clone of adenovirus 2 DNA bearing the maj...

  17. Susceptibility of Vaccinia Virus to Chemical Disinfectants

    Science.gov (United States)

    de Oliveira, Tércia Moreira Ludolfo; Rehfeld, Izabelle Silva; Coelho Guedes, Maria Isabel Maldonado; Ferreira, Jaqueline Maria Siqueira; Kroon, Erna Geessien; Lobato, Zélia Inês Portela

    2011-01-01

    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. PMID:21734141

  18. The complete DNA sequence of vaccinia virus.

    Science.gov (United States)

    Goebel, S J; Johnson, G P; Perkus, M E; Davis, S W; Winslow, J P; Paoletti, E

    1990-11-01

    The complete DNA sequence of the genome of vaccinia virus has been determined. The genome consisted of 191,636 bp with a base composition of 66.6% A + T. We have identified 198 "major" protein-coding regions and 65 overlapping "minor" regions, for a total of 263 potential genes. Genes encoded by the virus were located by examination of DNA sequence characteristics and compared with existing vaccinia virus mapping analyses, sequence data, and transcription data. These genes were found to be compactly organized along the genome with relatively few regions of noncoding sequences. Whereas several similarities to proteins of known function were discerned, the function of the majority of proteins encoded by these open reading frames is as yet undetermined.

  19. A New Vaccinia Virus Intermediate Transcription Factor

    OpenAIRE

    Sanz, Patrick; Moss, Bernard

    1998-01-01

    Transcription of the vaccinia virus genome is mediated by a virus-encoded multisubunit DNA-dependent RNA polymerase in conjunction with early-, intermediate-, and late-stage-specific factors. Previous studies indicated that two virus-encoded proteins (capping enzyme and VITF-1) and one unidentified cellular protein (VITF-2) are required for specific transcription of an intermediate promoter template in vitro. We have now extensively purified an additional virus-induced intermediate transcript...

  20. CD8 T cells are essential for recovery from a respiratory vaccinia virus infection.

    Science.gov (United States)

    Goulding, John; Bogue, Rebecka; Tahiliani, Vikas; Croft, Michael; Salek-Ardakani, Shahram

    2012-09-01

    The precise immune components required for protection against a respiratory Orthopoxvirus infection, such as human smallpox or monkeypox, remain to be fully identified. In this study, we used the virulent Western Reserve strain of vaccinia virus (VACV-WR) to model a primary respiratory Orthopoxvirus infection. Naive mice infected with VACV-WR mounted an early CD8 T cell response directed against dominant and subdominant VACV-WR Ags, followed by a CD4 T cell and Ig response. In contrast to other VACV-WR infection models that highlight the critical requirement for CD4 T cells and Ig, we found that only mice deficient in CD8 T cells presented with severe cachexia, pulmonary inflammation, viral dissemination, and 100% mortality. Depletion of CD8 T cells at specified times throughout infection highlighted that they perform their critical function between days 4 and 6 postinfection and that their protective requirement is critically dictated by initial viral load and virulence. Finally, the ability of adoptively transferred naive CD8 T cells to protect RAG⁻/⁻ mice against a lethal VACV-WR infection demonstrated that they are both necessary and sufficient in protecting against a primary VACV-WR infection of the respiratory tract.

  1. Genomic identification of human vaccinia virus keratoconjunctivitis and its importance as a laboratory-acquired infection

    Directory of Open Access Journals (Sweden)

    Zahra Movahedi Motlagh

    2016-01-01

    Full Text Available Context: Vaccinia virus (VACV is a member of orthopoxvirus genus of the family Poxviridae. VACVs are enveloped, double-stranded DNA viruses. Several species of this family, for example, molluscum contagiosum, smallpox, deerpox, horsepox, rabbitpox, and VACVs may cause conjunctivitis. Aims: Given the high incidence of keratoconjunctivitis in Iran (approximately 3.6%-53.9% and insufficient clinical diagnostic measures, laboratory tests for detection of its causes and determination of accurate keratoconjunctivitis/conjunctivitis prevalence due to different pathogens are essential. Settings and Design: In this research, conjunctival samples collected from 100 patients with keratoconjunctivitis signs were referred to an eye hospital of Iran. Subjects and Methods: After DNA extraction, polymerase chain reaction (PCR was carried out for detection of VACV. PCR-positive products were further subjected to DNA sequencing. Statistical Analysis Used: The results were analyzed using Chi-square test. Results: In this study, 28% of the samples were positive and a statistically significant relationship obtained between working in medical or research laboratories and VACV prevalence (P < 0.05. Conclusions: This study showed a high rate of VACV keratoconjunctivitis, and therefore, further studies for its prevention and control are necessary.

  2. 75 FR 3244 - Prospective Grant of Exclusive License: Monoclonal Antibodies Against Smallpox/Orthopoxviruses

    Science.gov (United States)

    2010-01-20

    ... Antibodies Against Smallpox/Orthopoxviruses AGENCY: National Institutes of Health, Public Health Service...: Concerns that variola (smallpox) virus might be used as a biological weapon have led to the recommendation... prevention of smallpox, it is well documented that various adverse reactions in individuals have been caused...

  3. The Migration of Smallpox and Its Indelible Footprint on Latin American History. Junior Division Winner.

    Science.gov (United States)

    Thomson, Mark

    1998-01-01

    Addresses the migration of smallpox into the New World where it caused the extinction of entire indigenous civilizations and altered the survivors' cultures. Discusses the historical origins of smallpox and highlights the migration and consequences of smallpox in Central and South America. Includes an annotated bibliography, research description,…

  4. Epidemiologic Assessment of the Protective Effects of Smallpox Vaccination

    Directory of Open Access Journals (Sweden)

    Hiroshi Nishiura

    2006-01-01

    Full Text Available Despite smallpox eradication, the widely discussed possibility of a bioterrorist attack using a variola virus makes it necessary to review the epidemiology of smallpox and the effects of various vaccination schemes. This paper provides a literature review concerning the epidemiologic assessment of the protective effects of smallpox vaccination, with particular emphasis on the statistical and theoretical points of view. Although smallpox vaccination has the longest history of all vaccinations, we lack precise epidemiologic estimates of its effectiveness. Vaccination practice continually evolved and many places experienced vaccinations with various strains over time. Despite the weak statistical evidence, it can be stated with certainty that smallpox vaccines prevented infection for a few decades after primary vaccination and that vaccinated individuals had the benefit of a longer lasting partial protection when they contracted the disease. Confronted with the huge uncertainties and with the necessity to rely on laboratory evidence, appropriate preparedness plans for countermeasures using vaccination must be based on the best available evidence.

  5. A prospective study of the incidence of myocarditis/pericarditis and new onset cardiac symptoms following smallpox and influenza vaccination.

    Directory of Open Access Journals (Sweden)

    Renata J M Engler

    Full Text Available Although myocarditis/pericarditis (MP has been identified as an adverse event following smallpox vaccine (SPX, the prospective incidence of this reaction and new onset cardiac symptoms, including possible subclinical injury, has not been prospectively defined.The study's primary objective was to determine the prospective incidence of new onset cardiac symptoms, clinical and possible subclinical MP in temporal association with immunization.New onset cardiac symptoms, clinical MP and cardiac specific troponin T (cTnT elevations following SPX (above individual baseline values were measured in a multi-center prospective, active surveillance cohort study of healthy subjects receiving either smallpox vaccine or trivalent influenza vaccine (TIV.New onset chest pain, dyspnea, and/or palpitations occurred in 10.6% of SPX-vaccinees and 2.6% of TIV-vaccinees within 30 days of immunization (relative risk (RR 4.0, 95% CI: 1.7-9.3. Among the 1081 SPX-vaccinees with complete follow-up, 4 Caucasian males were diagnosed with probable myocarditis and 1 female with suspected pericarditis. This indicates a post-SPX incidence rate more than 200-times higher than the pre-SPX background population surveillance rate of myocarditis/pericarditis (RR 214, 95% CI 65-558. Additionally, 31 SPX-vaccinees without specific cardiac symptoms were found to have over 2-fold increases in cTnT (>99th percentile from baseline (pre-SPX during the window of risk for clinical myocarditis/pericarditis and meeting a proposed case definition for possible subclinical myocarditis. This rate is 60-times higher than the incidence rate of overt clinical cases. No clinical or possible subclinical myocarditis cases were identified in the TIV-vaccinated group.Passive surveillance significantly underestimates the true incidence of myocarditis/pericarditis after smallpox immunization. Evidence of subclinical transient cardiac muscle injury post-vaccinia immunization is a finding that requires further

  6. Smallpox virus destruction and the implications of a new vaccine.

    Science.gov (United States)

    Henderson, D A

    2011-06-01

    The World Health Assembly is scheduled to decide in May 2011 whether the 2 known remaining stockpiles of smallpox virus are to be destroyed or retained. In preparation for this, a WHO-appointed committee undertook a comprehensive review of the status of smallpox virus research from 1999 to 2010. It concluded that, considering the nature of the studies already completed with respect to vaccine, drugs, and diagnostics, there was no reason to retain live smallpox virus except to satisfy restrictive regulatory requirements. The committee advised that researchers and regulators define alternative models for testing the vaccines and drugs. Apart from other considerations, the costs of new products are significant and important. These include prospective expenditures required for the development, manufacture, testing, and storage of new products. This commentary provides approximations of these costs and the incremental contribution that a newly developed vaccine might make in terms of public health security.

  7. A novel replication-competent vaccinia vector MVTT is superior to MVA for inducing high levels of neutralizing antibody via mucosal vaccination.

    Science.gov (United States)

    Huang, Xiaoxing; Lu, Bin; Yu, Wenbo; Fang, Qing; Liu, Li; Zhuang, Ke; Shen, Tingting; Wang, Haibo; Tian, Po; Zhang, Linqi; Chen, Zhiwei

    2009-01-01

    Mucosal vaccination offers great advantage for inducing protective immune response to prevent viral transmission and dissemination. Here, we report our findings of a head-to-head comparison of two viral vectors modified vaccinia Ankara (MVA) and a novel replication-competent modified vaccinia Tian Tan (MVTT) for inducing neutralizing antibodies (Nabs) via intramuscular and mucosal vaccinations in mice. MVTT is an attenuated variant of the wild-type VTT, which was historically used as a smallpox vaccine for millions of Chinese people. The spike glycoprotein (S) of SARS-CoV was used as the test antigen after the S gene was constructed in the identical genomic location of two vectors to generate vaccine candidates MVTT-S and MVA-S. Using identical doses, MVTT-S induced lower levels ( approximately 2-3-fold) of anti- SARS-CoV neutralizing antibodies (Nabs) than MVA-S through intramuscular inoculation. MVTT-S, however, was capable of inducing consistently 20-to-100-fold higher levels of Nabs than MVA-S when inoculated via either intranasal or intraoral routes. These levels of MVTT-S-induced Nab responses were substantially (approximately 10-fold) higher than that induced via the intramuscular route in the same experiments. Moreover, pre-exposure to the wild-type VTT via intranasal or intraoral route impaired the Nab response via the same routes of MVTT-S vaccination probably due to the pre-existing anti-VTT Nab response. The efficacy of intranasal or intraoral vaccination, however, was still 20-to-50-fold better than intramuscular inoculation despite the subcutaneous pre-exposure to wild-type VTT. Our data have implications for people who maintain low levels of anti-VTT Nabs after historical smallpox vaccination. MVTT is therefore an attractive live viral vector for mucosal vaccination.

  8. A novel replication-competent vaccinia vector MVTT is superior to MVA for inducing high levels of neutralizing antibody via mucosal vaccination.

    Directory of Open Access Journals (Sweden)

    Xiaoxing Huang

    Full Text Available Mucosal vaccination offers great advantage for inducing protective immune response to prevent viral transmission and dissemination. Here, we report our findings of a head-to-head comparison of two viral vectors modified vaccinia Ankara (MVA and a novel replication-competent modified vaccinia Tian Tan (MVTT for inducing neutralizing antibodies (Nabs via intramuscular and mucosal vaccinations in mice. MVTT is an attenuated variant of the wild-type VTT, which was historically used as a smallpox vaccine for millions of Chinese people. The spike glycoprotein (S of SARS-CoV was used as the test antigen after the S gene was constructed in the identical genomic location of two vectors to generate vaccine candidates MVTT-S and MVA-S. Using identical doses, MVTT-S induced lower levels ( approximately 2-3-fold of anti- SARS-CoV neutralizing antibodies (Nabs than MVA-S through intramuscular inoculation. MVTT-S, however, was capable of inducing consistently 20-to-100-fold higher levels of Nabs than MVA-S when inoculated via either intranasal or intraoral routes. These levels of MVTT-S-induced Nab responses were substantially (approximately 10-fold higher than that induced via the intramuscular route in the same experiments. Moreover, pre-exposure to the wild-type VTT via intranasal or intraoral route impaired the Nab response via the same routes of MVTT-S vaccination probably due to the pre-existing anti-VTT Nab response. The efficacy of intranasal or intraoral vaccination, however, was still 20-to-50-fold better than intramuscular inoculation despite the subcutaneous pre-exposure to wild-type VTT. Our data have implications for people who maintain low levels of anti-VTT Nabs after historical smallpox vaccination. MVTT is therefore an attractive live viral vector for mucosal vaccination.

  9. Smallpox Vaccination of Laboratory Workers at US Variola Testing Sites.

    Science.gov (United States)

    Medcalf, Sharon; Bilek, Laura; Hartman, Teresa; Iwen, Peter C; Leuschen, Patricia; Miller, Hannah; O'Keefe, Anne; Sayles, Harlan; Smith, Philip W

    2015-08-01

    To evaluate the need to revaccinate laboratory workers against smallpox, we assessed regular revaccination at the US Laboratory Response Network's variola testing sites by examining barriers to revaccination and the potential for persistence of immunity. Our data do not provide evidence to suggest prolonging the recommended interval for revaccination.

  10. Bioterrorism and Smallpox: Policies, Practices, and Implications for Social Work

    Science.gov (United States)

    Mackelprang, Romel W.; Mackelprang, Romel D.; Thirkill, Ashley D.

    2005-01-01

    Terrorist acts and the fear of terrorism have become a part of everyday life in the early 21st century. Among the threats most feared is bioterrorism, including the intentional release of smallpox. With the invasion of Iraq and toppling of the Saddam Hussein regime, acute bioterrorism fears have abated; however, an ongoing threat remains. This…

  11. Bioterrorism and Smallpox: Policies, Practices, and Implications for Social Work

    Science.gov (United States)

    Mackelprang, Romel W.; Mackelprang, Romel D.; Thirkill, Ashley D.

    2005-01-01

    Terrorist acts and the fear of terrorism have become a part of everyday life in the early 21st century. Among the threats most feared is bioterrorism, including the intentional release of smallpox. With the invasion of Iraq and toppling of the Saddam Hussein regime, acute bioterrorism fears have abated; however, an ongoing threat remains. This…

  12. The Emergency campaign for smallpox eradication from Somalia (1977-1979)--revisited.

    Science.gov (United States)

    Deria, Abdullahi

    2011-12-30

    The historical significance of smallpox eradication from Somalia lies in the fact that the country was the last to record the last endemic smallpox case in the world. Before 1977 the programme was mismanaged. In the mid-1970s, the programme was plagued with concealment. Confirmation of smallpox outbreak in Mogadishu in September 1976 delayed global smallpox eradication. The Government maintained that there was no ongoing smallpox transmission in the country after the Mogadishu outbreak and frustrated independent attempts to verify its claim. In February 1977 the Government allowed World Health Organization (WHO) epidemiologists to search, unhindered, for smallpox outside Mogadishu. Soon widespread smallpox transmission was detected. The Government appealed for international support. The strategy to stop the smallpox transmission was based on surveillance and containment. The WHO took the leading role of the campaign which, in spite of the Somalia/Ethiopia war of 1977/78, culminated in the eradication of smallpox from the country. Somalia was certified smallpox-free on 19 October 1979.

  13. GeneChip Resequencing of the Smallpox Virus Genome Can Identify Novel Strains: a Biodefense Application▿

    Science.gov (United States)

    Sulaiman, Irshad M.; Tang, Kevin; Osborne, John; Sammons, Scott; Wohlhueter, Robert M.

    2007-01-01

    We developed a set of seven resequencing GeneChips, based on the complete genome sequences of 24 strains of smallpox virus (variola virus), for rapid characterization of this human-pathogenic virus. Each GeneChip was designed to analyze a divergent segment of approximately 30,000 bases of the smallpox virus genome. This study includes the hybridization results of 14 smallpox virus strains. Of the 14 smallpox virus strains hybridized, only 7 had sequence information included in the design of the smallpox virus resequencing GeneChips; similar information for the remaining strains was not tiled as a reference in these GeneChips. By use of variola virus-specific primers and long-range PCR, 22 overlapping amplicons were amplified to cover nearly the complete genome and hybridized with the smallpox virus resequencing GeneChip set. These GeneChips were successful in generating nucleotide sequences for all 14 of the smallpox virus strains hybridized. Analysis of the data indicated that the GeneChip resequencing by hybridization was fast and reproducible and that the smallpox virus resequencing GeneChips could differentiate the 14 smallpox virus strains characterized. This study also suggests that high-density resequencing GeneChips have potential biodefense applications and may be used as an alternate tool for rapid identification of smallpox virus in the future. PMID:17182757

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

    Science.gov (United States)

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

    2013-01-02

    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.

  15. Smallpox DNA Vaccine Protects Nonhuman Primates Against Lethal Monkeypox

    Science.gov (United States)

    2004-05-01

    threatening (e.g., eczema vac- cinatum, encephalitis, and progressive vaccinia) (6). Serious adverse events that are not necessarily causally associated...This outbreak (71 suspected and 35 laboratory-confirmed cases as of 8 July 2003) may have been transmitted from prairie dogs that were infected after

  16. Vaccinia Virus Infections in a Martial Arts Gym

    Centers for Disease Control (CDC) Podcasts

    2011-04-04

    This podcast discusses an outbreak of vaccinia virus in Maryland in 2008. Christine Hughes, a health scientist with the Poxvirus and Rabies Branch at CDC, and co-author of a paper in the April 2011 issue of CDC's journal, discusses vaccinia virus infections in a martial arts gym.  Created: 4/4/2011 by National Center for Emerging Zoonotic and Infectious Diseases (NCEZID).   Date Released: 4/5/2011.

  17. [The history of smallpox vaccination in the Imperial Moscow foster house].

    Science.gov (United States)

    Sher, S A

    2011-01-01

    The article deals with the history of vaccination against natural smallpox which is directly connected to the Imperial Moscow foster house which became one of smallpox vaccination centers in Russia of XIX century. In 1801, when variolations were substituted by more safe cowpox vaccinations, in Russia the first vaccination using the method of Jenner was made exactly in in the Imperial Moscow foster house. From 1805, the smallpox vaccination received the status of force of law, the Imperial Moscow foster house began to produce and to distribute the smallpox vaccine all over the country and apply the smallpox vaccination not only to its foster children but to all turned to and, besides that, to train the smallpox vaccination. In 1857, the Imperial Moscow foster house became the first establishment in Russia where the revaccination was applied. In 1980, the WHO proclaimed that the implementation of the global program of smallpox irradiation resulted in the natural smallpox elimination on Earth. The smallpox became the first communicable disease defeated due to mass vaccination. One third of Earth population was vaccinated by the Soviet vaccine, which originated mainly because of the activities of physicians of the Imperial Moscow foster house.

  18. Comparative Pathology of Smallpox and Monkeypox in Man and Macaques

    Science.gov (United States)

    Cann, J. A.; Jahrling, P. B.; Hensley, L. E.; Wahl-Jensen, V.

    2012-01-01

    Summary In the three decades since the eradication of smallpox and cessation of routine vaccination, the collective memory of the devastating epidemics caused by this orthopoxvirus has waned, and the human population has become increasingly susceptible to a disease that remains high on the list of possible bioterrorism agents. Research using surrogate orthopoxviruses in their natural hosts, as well as limited variola virus research in animal models, continues worldwide; however, interpretation of findings is often limited by our relative lack of knowledge about the naturally occurring disease. For modern comparative pathologists, many of whom have no first-hand knowledge of naturally occurring smallpox, this work provides a contemporary review of this historical disease, as well as discussion of how it compares with human monkeypox and the corresponding diseases in macaques. PMID:22884034

  19. [Immunological characteristics of the 2-stage method of smallpox vaccination].

    Science.gov (United States)

    Marennikova, S S; Matsevich, G R; Sokolova, A F; Shul'ga, L G; Manenkova, G M

    1977-05-01

    As a result of observations carried out on children the authors present immunological characteristics of two-stage smallpox vaccination at different intervals (1 to 60 days) between the injection of inactivated and live vaccine. There proved to be acceleration and intensification of antibody formation after two-stage immunization in comparison with the rutine vaccination. A seven-day interval between the injection of the inactivated and live preparations was recommended on the basis of the data obtained.

  20. Smallpox as a Bioweapon: Should We Be Concerned?

    Science.gov (United States)

    2012-03-01

    Story of Eradicating a Worldwide Killer (New York: Prometheus Books, 2009), 32. 15 Bill Frist, When Every Moment Counts: What You Need to Know...Hammond, and Leonard Cole, eds., Essentials of Terror Medicine (New York, NY: Springer, 2009), 203. 46 Henderson and Borio, “Smallpox and Monkeypox,” 629...Shapira, Hammond, and Cole, eds., Essentials of Terror Medicine , 204; Centers for Disease Control and Prevention, Division of Bioterrorism

  1. In vitro efficacy of ST246 against smallpox and monkeypox.

    Science.gov (United States)

    Smith, Scott K; Olson, Victoria A; Karem, Kevin L; Jordan, Robert; Hruby, Dennis E; Damon, Inger K

    2009-03-01

    Since the eradication of smallpox and the cessation of routine childhood vaccination for smallpox, the proportion of the world's population susceptible to infection with orthopoxviruses, such as variola virus (the causative agent of smallpox) and monkeypox virus, has grown substantially. In the United States, the only vaccines for smallpox licensed by the Food and Drug Administration (FDA) have been live virus vaccines. Unfortunately, a substantial number of people cannot receive live virus vaccines due to contraindications. Furthermore, no antiviral drugs have been fully approved by the FDA for the prevention or treatment of orthopoxvirus infection. Here, we show the inhibitory effect of one new antiviral compound, ST-246, on the in vitro growth properties of six variola virus strains and seven monkeypox virus strains. We performed multiple assays to monitor the cytopathic effect and to evaluate the reduction of viral progeny production and release in the presence of the compound. ST-246 had 50% effective concentrations of

  2. Safety, Immunogenicity and Efficacy of Modified Vaccinia Ankara (MVA) Against Dryvax® Challenge in Vaccinia-Naïve and Vaccinia-Immune Individuals

    Science.gov (United States)

    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

    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. PMID:17126963

  3. Characterization of Two Historic Smallpox Specimens from a Czech Museum.

    Science.gov (United States)

    Pajer, Petr; Dresler, Jiri; Kabíckova, Hana; Písa, Libor; Aganov, Pavel; Fucik, Karel; Elleder, Daniel; Hron, Tomas; Kuzelka, Vitezslav; Velemínsky, Petr; Klimentova, Jana; Fucikova, Alena; Pejchal, Jaroslav; Hrabakova, Rita; Benes, Vladimir; Rausch, Tobias; Dundr, Pavel; Pilin, Alexander; Cabala, Radomir; Hubalek, Martin; Stríbrny, Jan; Antwerpen, Markus H; Meyer, Hermann

    2017-07-27

    Although smallpox has been known for centuries, the oldest available variola virus strains were isolated in the early 1940s. At that time, large regions of the world were already smallpox-free. Therefore, genetic information of these strains can represent only the very last fraction of a long evolutionary process. Based on the genomes of 48 strains, two clades are differentiated: Clade 1 includes variants of variola major, and clade 2 includes West African and variola minor (Alastrim) strains. Recently, the genome of an almost 400-year-old Lithuanian mummy was determined, which fell basal to all currently sequenced strains of variola virus on phylogenetic trees. Here, we determined two complete variola virus genomes from human tissues kept in a museum in Prague dating back 60 and 160 years, respectively. Moreover, mass spectrometry-based proteomic, chemical, and microscopic examinations were performed. The 60-year-old specimen was most likely an importation from India, a country with endemic smallpox at that time. The genome of the 160-year-old specimen is related to clade 2 West African and variola minor strains. This sequence likely represents a new endemic European variant of variola virus circulating in the midst of the 19th century in Europe.

  4. New insights about host response to smallpox using microarray data

    Directory of Open Access Journals (Sweden)

    Dias Rodrigo A

    2007-08-01

    Full Text Available Abstract Background Smallpox is a lethal disease that was endemic in many parts of the world until eradicated by massive immunization. Due to its lethality, there are serious concerns about its use as a bioweapon. Here we analyze publicly available microarray data to further understand survival of smallpox infected macaques, using systems biology approaches. Our goal is to improve the knowledge about the progression of this disease. Results We used KEGG pathways annotations to define groups of genes (or modules, and subsequently compared them to macaque survival times. This technique provided additional insights about the host response to this disease, such as increased expression of the cytokines and ECM receptors in the individuals with higher survival times. These results could indicate that these gene groups could influence an effective response from the host to smallpox. Conclusion Macaques with higher survival times clearly express some specific pathways previously unidentified using regular gene-by-gene approaches. Our work also shows how third party analysis of public datasets can be important to support new hypotheses to relevant biological problems.

  5. Should smallpox vaccine be made available to the general public?

    Science.gov (United States)

    May, Thomas; Silverman, Ross D

    2003-06-01

    In June 2002, the Advisory Committee on Immunization Practices (ACIP) approved draft recommendations concerning preparation for potential biological terror attacks that utilize the smallpox virus. ACIP recommends against both mandatory and voluntary vaccination of the general public. The present paper examines the moral and political considerations both for and against each of the general public vaccination options considered by the ACIP in the context of the state's authority over vaccination for the purposes of protecting public health. Although it is clear that compulsory mass vaccination is not justified at this time, the issues surrounding voluntary vaccination are more complex. Should smallpox vaccination prior to an outbreak be made available to the general public? The paper concludes that the vaccine should not be made available at this time. This conclusion, however, is based upon contingent features of current circumstances, which would change once an outbreak occurred. In the even of a terror-related outbreak of smallpox, the general public's access to voluntary vaccination would become justified, even in areas beyond where the outbreak has occurred.

  6. Vaccination scars in HIV infected patients – does vaccinia vaccination confer protection against HIV?

    DEFF Research Database (Denmark)

    Jespersen, Sanne; Hønge, Bo Langhoff; Medina, Candida

    Vaccination scars in HIV infected patients – does vaccinia vaccination confer protection against HIV?......Vaccination scars in HIV infected patients – does vaccinia vaccination confer protection against HIV?...

  7. Safety and immunogenicity of IMVAMUNE® smallpox vaccine using different strategies for a post event scenario.

    Science.gov (United States)

    Frey, Sharon E; Winokur, Patricia L; Salata, Robert A; El-Kamary, Samer S; Turley, Christine B; Walter, Emmanuel B; Hay, Christine Mhorag; Newman, Frances K; Hill, Heather R; Zhang, Ying; Chaplin, Paul; Tary-Lehmann, Magdalena; Belshe, Robert B

    2013-06-24

    Reintroduction of Variola major as an agent of bioterrorism remains a concern. A shortened dosing schedule of Bavarian Nordic's (BN) IMVAMUNE(®) (modified vaccinia Ankara vaccine against smallpox) was compared to the currently recommended 0- and 28-day schedule for non-inferiority by evaluating the magnitude and kinetics of the immune responses. Subjects were assigned to receive IMVAMUNE or placebo administered subcutaneously on Days 0 and 7, Days 0 and 28, or Day 0. Blood was collected for antibody and cell-mediated immune assays. Subjects were followed for safety for 12 months after last vaccination. The primary endpoint of this study was the geometric mean antibody titers (GMT) at 14 days post last vaccination. Of 208 subjects enrolled, 191 received vaccine (Group: 0+7, Group: 0+28 and Group: 0) and 17 received placebo. Moderate/severe systemic reactogenicity after any vaccination were reported by 31.1%, 25.4%, and 28.6% of the subjects for Group: 0+7, Group: 0+28, and Group: 0, respectively (Chi-square test, P=0.77). Based on BN's Plaque Reduction Assay GMTs, Group: 0+7 was non-inferior to Group: 0+28 at Day 4, 180, and 365 after the second vaccination. On Day 14, Group: 0+7 and Group: 0+28 GMT were 10.8 (CI: 9.0, 12.9) and 30.2 (CI: 22.1, 41.1), respectively. Based on BN's Enzyme-linked immunosorbent assay, the proportion of subjects with positive titers for Group: 0+28 was significantly greater than that for Group: 0+7 after second vaccination at Days 4 and 180. By Day 14 after the second dose, the IFN-γ enzyme-linked immunosorbent spot (ELISPOT) responses were similar for Group: 0+28 and Group: 0+7. Overall, a standard dose of IMVAMUNE (0.5 mL of 1 x 10(8) TCID/mL) administered subcutaneously was safe and well tolerated. A second dose of IMVAMUNE at Day 28 compared to Day 7 provided greater antibody responses and the maximal number of responders. By Day 14 after the second dose, IFN-γ ELISPOT responses were similar for Group: 0+28 and Group: 0

  8. 42 CFR 102.51 - Documentation a smallpox vaccine recipient must submit to be deemed eligible by the Secretary.

    Science.gov (United States)

    2010-10-01

    ... 42 Public Health 1 2010-10-01 2010-10-01 false Documentation a smallpox vaccine recipient must..., DEPARTMENT OF HEALTH AND HUMAN SERVICES VACCINES SMALLPOX COMPENSATION PROGRAM Required Documentation To Be Deemed Eligible § 102.51 Documentation a smallpox vaccine recipient must submit to be deemed eligible...

  9. Infectious vaccinia virus recombinants that express hepatitis B virus surface antigen

    Science.gov (United States)

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

    1983-04-01

    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.

  10. Lessons for Implementation from the World's Most Successful Programme: The Global Eradication of Smallpox.

    Science.gov (United States)

    Pratt, David

    1999-01-01

    Focuses on lessons educators might learn from the Intensified Campaign for the Global Eradication of Smallpox. Outlines the history of smallpox eradication. Discusses the eradication effort's obstacles, campaign, and costs and benefits. Considers five factors relevant to the successful implementation of educational programs. (CMK)

  11. A recombinant vaccinia virus expressing human carcinoembryonic antigen (CEA).

    Science.gov (United States)

    Kaufman, H; Schlom, J; Kantor, J

    1991-07-30

    Carcinoembryonic antigen (CEA) is a 180-kDa glycoprotein expressed on most gastrointestinal carcinomas. A 2.4-kb cDNA clone, containing the complete coding sequence, was isolated from a human colon tumor cell library and inserted into a vaccinia virus genome. This newly developed construct was characterized by Southern blotting, DNA hybridization studies, and polymerase chain reaction analysis. The CEA gene was stably integrated into the vaccinia virus thymidine kinase gene. The recombinant was efficiently replicated upon serial passages in cell cultures and in animals. The recombinant virus expresses on the surface of infected cells a protein product recognized by a monoclonal antibody (COL-I) directed against CEA. Immunization of mice with the vaccinia construct elicited a humoral immune response against CEA. Pilot studies also showed that administration of the recombinant CEA vaccinia construct was able to greatly reduce the growth in mice of a syngeneic murine colon adenocarcinoma which had been transduced with the human CEA gene. The use of this new recombinant CEA vaccinia construct may thus provide an approach in the specific active immunotherapy of human GI cancer and other CEA expressing carcinoma types.

  12. Cryo-electron tomography of vaccinia virus

    Science.gov (United States)

    Cyrklaff, Marek; Risco, Cristina; Fernández, Jose Jesús; Jiménez, Maria Victoria; Estéban, Mariano; Baumeister, Wolfgang; Carrascosa, José L.

    2005-01-01

    The combination of cryo-microscopy and electron tomographic reconstruction has allowed us to determine the structure of one of the more complex viruses, intracellular mature vaccinia virus, at a resolution of 4–6 nm. The tomographic reconstruction allows us to dissect the different structural components of the viral particle, avoiding projection artifacts derived from previous microscopic observations. A surface-rendering representation revealed brick-shaped viral particles with slightly rounded edges and dimensions of ≈360 × 270 × 250 nm. The outer layer was consistent with a lipid membrane (5–6 nm thick), below which usually two lateral bodies were found, built up by a heterogeneous material without apparent ordering or repetitive features. The internal core presented an inner cavity with electron dense coils of presumptive DNA–protein complexes, together with areas of very low density. The core was surrounded by two layers comprising an overall thickness of ≈18–19 nm; the inner layer was consistent with a lipid membrane. The outer layer was discontinuous, formed by a periodic palisade built by the side interaction of T-shaped protein spikes that were anchored in the lower membrane and were arranged into small hexagonal crystallites. It was possible to detect a few pore-like structures that communicated the inner side of the core with the region outside the layer built by the T-shaped spike palisade. PMID:15699328

  13. Unraveling the structure of the variola topoisomerase IB-DNA complex: a possible new twist on smallpox therapy.

    Science.gov (United States)

    Osheroff, Neil

    2006-10-01

    Smallpox is a serious and highly contagious disease that is caused by the variola virus. It is one of the most severe infectious human diseases known, with mortality rates as high as 30%. A successful worldwide vaccination program led to the eradication of smallpox in 1980. However, the high transmission rate of variola virus, coupled with the deadly nature of smallpox, makes this virus a potentially devastating weapon for bioterrorism. Currently, there is no specific treatment for smallpox. However, a recent article on the structure of a variola topoisomerase IB-DNA complex provides an intriguing starting point for the rational design of drugs with potential activity against smallpox.

  14. Group interest versus self-interest in smallpox vaccination policy.

    Science.gov (United States)

    Bauch, Chris T; Galvani, Alison P; Earn, David J D

    2003-09-02

    The recent threat of bioterrorism has fueled debate on smallpox vaccination policy for the United States. Certain policy proposals call for voluntary mass vaccination; however, if individuals decide whether to vaccinate according to self-interest, the level of herd immunity achieved may differ from what is best for the population as a whole. We present a synthesis of game theory and epidemic modeling that formalizes this conflict between self-interest and group interest and shows that voluntary vaccination is unlikely to reach the group-optimal level. This shortfall results in a substantial increase in expected mortality after an attack.

  15. Outbreaks of human monkeypox after cessation of smallpox vaccination.

    Science.gov (United States)

    Reynolds, Mary G; Damon, Inger K

    2012-02-01

    The recent observation of a surge in human monkeypox in the Democratic Republic of the Congo (DRC) prompts the question of whether cessation of smallpox vaccination is driving the phenomenon, and if so, why is re-emergence not universal throughout the historic geographic range of the virus? Research addressing the virus's mechanisms for immune evasion and induction, as well as that directed at elucidating the genes involved in pathogenesis in different viral lineages (West African vs Congo Basin), provide insights to help explain why emergence appears to be geographically limited. Novel vaccines offer one solution to curtail the spread of this disease.

  16. Immunodomination during peripheral vaccinia virus infection.

    Directory of Open Access Journals (Sweden)

    Leon C W Lin

    Full Text Available Immunodominance is a fundamental property of CD8(+ T cell responses to viruses and vaccines. It had been observed that route of administration alters immunodominance after vaccinia virus (VACV infection, but only a few epitopes were examined and no mechanism was provided. We re-visited this issue, examining a panel of 15 VACV epitopes and four routes, namely intradermal (i.d., subcutaneous (s.c., intraperitoneal (i.p. and intravenous (i.v. injection. We found that immunodominance is sharpened following peripheral routes of infection (i.d. and s.c. compared with those that allow systemic virus dissemination (i.p. and i.v.. This increased immunodominance was demonstrated with native epitopes of VACV and with herpes simplex virus glycoprotein B when expressed from VACV. Responses to some subdominant epitopes were altered by as much as fourfold. Tracking of virus, examination of priming sites, and experiments restricting virus spread showed that priming of CD8(+ T cells in the spleen was necessary, but not sufficient to broaden responses. Further, we directly demonstrated that immunodomination occurs more readily when priming is mainly in lymph nodes. Finally, we were able to reduce immunodominance after i.d., but not i.p. infection, using a VACV expressing the costimulators CD80 (B7-1 and CD86 (B7-2, which is notable because VACV-based vaccines incorporating these molecules are in clinical trials. Taken together, our data indicate that resources for CD8(+ T cell priming are limiting in local draining lymph nodes, leading to greater immunodomination. Further, we provide evidence that costimulation can be a limiting factor that contributes to immunodomination. These results shed light on a possible mechanism of immunodomination and highlight the need to consider multiple epitopes across the spectrum of immunogenicities in studies aimed at understanding CD8(+ T cell immunity to viruses.

  17. Vaccinations against smallpox and tuberculosis are associated with better long-term survival

    DEFF Research Database (Denmark)

    Rieckmann, Andreas; Villumsen, Marie; Sørup, Signe;

    2016-01-01

    -up), and 401 deaths due to natural causes (841 deaths in total) occurred in the full cohort. Compared with individuals who had not received vaccinia or BCG, those who had received both vaccinia and BCG had an adjusted hazard ratio (aHR) of 0.54 [95% confidence interval (CI): 0.36-0.81] for mortality due...

  18. Smallpox and its eradication in the Democratic Republic of Congo: lessons learned.

    Science.gov (United States)

    Muyembe-Tamfum, Jean-Jacques; Mulembakani, Prime; Lekie, René Botee; Szczeniowski, Mark; Ježek, Zdeněk; Doshi, Reena; Hoff, Nicole; Rimoin, Anne W

    2011-12-30

    Smallpox eradication is considered to be one of the most remarkable accomplishments of the 20th century. Lessons learned from the campaign during the 1960s and 1970s in the Democratic Republic of Congo (DRC) can provide important information for the development of other eradication programs including polio. The DRC is the third largest country in Africa; the population suffers from extreme poverty, deteriorating infrastructure and health systems, and long periods of civil strife. Despite these challenges, DRC's smallpox eradication campaign was successful, eradicating smallpox only 41 months after initiation. DRC had been polio free since 2001; however, in 2006, imported cases were identified in the country. Polio transmission has since been re-established and DRC now has the second greatest number of reported polio cases in the world. Challenges which existed during the smallpox campaign in DRC are still present today; additionally, the polio vaccine itself poses unique challenges which include requiring multiple doses to confer immunity. In the fight against polio in DRC, it will be important to draw from the smallpox eradication experience. A number of important themes emerged during the campaign that could be beneficial to eradicating polio and future eradication programs that may follow. During the smallpox campaign, a standard vaccination program was implemented, surveillance was intensified, and there were strong collaborative programs with community involvement. These successful elements of the smallpox campaign should be adapted and applied in DRC in polio eradication programs. Copyright © 2011 Elsevier Ltd. All rights reserved.

  19. Development of the small-molecule antiviral ST-246® as a smallpox therapeutic

    Science.gov (United States)

    Grosenbach, Douglas W; Jordan, Robert; Hruby, Dennis E

    2011-01-01

    Naturally occurring smallpox has been eradicated, yet it remains as one of the highest priority pathogens due to its potential as a biological weapon. The majority of the US population would be vulnerable in a smallpox outbreak. SIGA Technologies, Inc. has responded to the call of the US government to develop and supply to the Strategic National Stockpile a smallpox antiviral to be deployed in the event of a smallpox outbreak. ST-246® (tecovirimat) was initially identified via a high-throughput screen in 2002, and in the ensuing years, our drug-development activities have spanned in vitro analysis, preclinical safety, pharmacokinetics and efficacy testing (all according to the ‘animal rule’). Additionally, SIGA has conducted Phase I and II clinical trials to evaluate the safety, tolerability and pharmacokinetics of ST-246, bringing us to our current late stage of clinical development. This article reviews the need for a smallpox therapeutic and our experience in developing ST-246, and provides perspective on the role of a smallpox antiviral during a smallpox public health emergency. PMID:21837250

  20. Antigen profiling analysis of vaccinia virus injected canine tumors

    Science.gov (United States)

    Cecil, Alexander; Gentschev, Ivaylo; Adelfinger, Marion; Nolte, Ingo; Dandekar, Thomas; Szalay, Aladar A

    2014-01-01

    Virotherapy on the basis of oncolytic vaccinia virus (VACV) strains is a novel approach for cancer therapy. In this study we describe for the first time the use of dynamic boolean modeling for tumor growth prediction of vaccinia virus GLV-1h68-injected canine tumors including canine mammary adenoma (ZMTH3), canine mammary carcinoma (MTH52c), canine prostate carcinoma (CT1258), and canine soft tissue sarcoma (STSA-1). Additionally, the STSA-1 xenografted mice were injected with either LIVP 1.1.1 or LIVP 5.1.1 vaccinia virus strains.   Antigen profiling data of the four different vaccinia virus-injected canine tumors were obtained, analyzed and used to calculate differences in the tumor growth signaling network by type and tumor type. Our model combines networks for apoptosis, MAPK, p53, WNT, Hedgehog, TK cell, Interferon, and Interleukin signaling networks. The in silico findings conform with in vivo findings of tumor growth. Boolean modeling describes tumor growth and remission semi-quantitatively with a good fit to the data obtained for all cancer type variants. At the same time it monitors all signaling activities as a basis for treatment planning according to antigen levels. Mitigation and elimination of VACV- susceptible tumor types as well as effects on the non-susceptible type CT1258 are predicted correctly. Thus the combination of Antigen profiling and semi-quantitative modeling optimizes the therapy already before its start. PMID:25482233

  1. A new inhibitor of apoptosis from vaccinia virus and eukaryotes.

    NARCIS (Netherlands)

    Gubser, C.; Bergamaschi, D.; Hollinshead, M.; Lu, X.; Kuppeveld, F.J.M. van; Smith, G.L.

    2007-01-01

    A new apoptosis inhibitor is described from vaccinia virus, camelpox virus, and eukaryotic cells. The inhibitor is a hydrophobic, multiple transmembrane protein that is resident in the Golgi and is named GAAP (Golgi anti-apoptotic protein). Stable expression of both viral GAAP (v-GAAP) and human GAA

  2. Smallpox, risks of terrorist attacks, and the Nash equilibrium: an introduction to game theory and an examination of the smallpox vaccination program.

    Science.gov (United States)

    Hamilton, Richard; McCain, Roger

    2009-01-01

    The smallpox vaccination emergency preparedness program has been unsuccessful in enrolling sufficient numbers of healthcare workers. The objective of this study was to use game theory to analyze a pre-event vaccination versus post-event vaccination program using the example of a terrorist considering an attack with smallpox or a hoax. A three-person game (normal and extensive form), and an in-person game are played for pre-event and post-event vaccinations of healthcare workers facing the possibility of a smallpox attack or hoax. Full pre-event vaccinations of all targeted healthcare workers are not necessary to deter a terrorist attack. In addition, coordinating vaccinations among healthcare workers, individual healthcare worker risk aversion, and the degree to which terrorists make the last move based on specific information on the status of pre-event vaccination all greatly impact strategy selection for both sides. A Nash Equilibrium of pre- and post-event vaccination strategies among a large number of healthcare professionals will tend to eliminate the advantage (of the terrorists) of a smallpox attack over a hoax, but may not eliminate some probability of a smallpox attack. Emergency preparedness would benefit from game theory analysis of the costs and payoffs of specific terrorism/counter-terrorism strategies.

  3. A brief history of vaccines: smallpox to the present.

    Science.gov (United States)

    Hsu, Jennifer L

    2013-01-01

    Modern vaccine history began in the late 18th century with the discovery of smallpox immunization by Edward Jenner. This pivotal step led to substantial progress in prevention of infectious diseases with inactivated vaccines for multiple infectious diseases, including typhoid, plague and cholera. Each advance produced significant decreases in infection-associated morbidity and mortality, thus shaping our modem cultures. As knowledge of microbiology and immunology grew through the 20th century, techniques were developed for cell culture of viruses. This allowed for rapid advances in prevention of polio, varicella, influenza and others. Finally, recent research has led to development of alternative vaccine strategies through use of vectored antigens, pathogen subunits (purified proteins or polysaccharides) or genetically engineered antigens. As the science of vaccinology continues to rapidly evolve, knowledge of the past creates added emphasis on the importance of developing safe and effective strategies for infectious disease prevention in the 21st century.

  4. [Should the human smallpox virus (variola) be destroyed?].

    Science.gov (United States)

    Tryland, Morten

    2004-10-21

    Smallpox, caused by variola virus, was a terror for civilizations around the world for more than 3000 years. Although the disease is eradicated, hundreds of variola virus isolates are kept in two WHO-collaborating facilities, one in USA and one in Russia. In spite of several agreements on destruction, it is now doubtful that these virus isolates will be destroyed. Variola virus may exist in other places and may be used as a biological weapon in war or for terror. Further research on variola virus is thus essential in order to achieve a better understanding of the pathogenicity of the virus and to develop new anti-variola virus vaccines and antiviral drugs.

  5. Drought, smallpox, and emergence of Leishmania braziliensis in northeastern Brazil.

    Science.gov (United States)

    Sousa, Anastácio Q; Pearson, Richard

    2009-06-01

    Cutaneous leishmaniasis caused by Leishmania (Vianna) braziliensis is a major health problem in the state of Ceará in northeastern Brazil. We propose that the disease emerged as a consequence of the displacement of persons from Ceará to the Amazon region following the Great Drought and smallpox epidemic of 1877-1879. As the economic and social situation in Ceará deteriorated, approximately 55,000 residents migrated to the Amazon region to find work, many on rubber plantations. Those that returned likely introduced L. (V.) brazilensis into Ceará, where the first cases of cutaneous leishmaniasis were reported early in the 20th century. The absence of an animal reservoir in Ceará, apart from dogs, supports the hypothesis. The spread of HIV/AIDS into the region and the possibility of concurrent cutaneous leishmaniasis raise the possibility of future problems.

  6. Public health and bioterrorism: renewed threat of anthrax and smallpox.

    Science.gov (United States)

    Wallin, Arūne; Luksiene, Zivile; Zagminas, Kestutis; Surkiene, Gene

    2007-01-01

    Bioterrorism is one of the main public health categorical domains. According to sociological analytics, in postmodern society terrorism is one of the real threats of the 21st century. While rare, the use of biological weapons has a long history. Recently, anthrax has been evaluated as one of the most dangerous biological weapons. Naturally occurring anthrax in humans is a disease acquired from contact with anthrax-infected animals or anthrax-contaminated animal products. Usually anthrax infection occurs in humans by three major routes: inhalational, cutaneous, and gastrointestinal. Inhalational anthrax is expected to account for most serious morbidity and most mortality. The clinical presentation of inhalation anthrax has been described as a two-stage illness. Many factors contribute to the pathogenesis of Bacillus anthracis. Antibiotics, anthrax globulin, corticosteroids, mechanical ventilation, vaccine are possible tools of therapy. Smallpox existed in two forms: variola major, which accounted for most morbidity and mortality, and a milder form, variola minor. Smallpox spreads from person to person primarily by droplet nuclei or aerosols expelled from the oropharynx of infected persons and by direct contact. In the event of limited outbreak with few cases, patients should be admitted to the hospital and confined to rooms that are under negative pressure and equipped with high-efficiency particulate air filtration. In larger outbreaks, home isolation and care should be the objective for most patients. Progress in detection, suitable vaccines, postexposure prophylaxis, infection control, and decontamination might be serious tools in fight against the most powerful biological weapon. To assure that the public health and healthcare system can respond to emergencies, the government should direct resources to strengthen the emergency-response system, create medication stockpiles, and improve the public health infrastructure.

  7. UNIT 14A.4 Generation of Recombinant Vaccinia Viruses

    Science.gov (United States)

    Earl, Patricia L.; Moss, Bernard; Wyatt, Linda S.

    2016-01-01

    This unit describes how to infect cells with vaccinia virus and then transfect them with a plasmid-transfer vector or PCR fragment to generate a recombinant virus. Selection and screening methods used to isolate recombinant viruses and a method for the amplification of recombinant viruses are described. Finally, a method for live immunostaining that has been used primarily for detection of recombinant modified vaccinia virus Ankara (MVA) is presented. This unit first describes how to infect cells with vaccinia virus and then transfect them with a plasmid-transfer vector or PCR fragment to generate a recombinant virus (see Basic Protocol 1). Also presented are selection and screening methods used to isolate recombinant viruses (see Basic Protocol 2) and a method for the amplification of recombinant viruses (see Basic Protocol 3). Finally, a method for live immunostaining that has been used primarily for detection of recombinant modified vaccinia virus Ankara (MVA) is presented (see Basic Protocol 4). HeLa S3 cells are recommended for large-scale growth of vaccinia virus. BS-C-1 cells may be used for xanthine-guanine phosphoribosyltransferase (XGPRT) and plaque size selection, fluorescent protein screening, transfection and determination of virus titer (UNIT 14A.3). For thymidine kinase (TK) selection, HuTK− 143B cells are used. With MVA, all steps are carried out in CEF or BHK-21 cells (UNIT 14A.3). CAUTION Proceed carefully and follow biosafety level 2 (BL-2) practices when working with standard vaccinia virus (see UNIT 14A.3 for safety precautions). [*Copy Editor: The original CPMB unit referenced CPMB Unit 16.15 for safety. The chapter editor asked that the authors include some of the safety information in the revised units – CPMB 16.16 and 16.17 – which are now CPMC Unit 14A.3 and 14A.4. As a result, the authors changed the safety citation here to “Unit 14A.3”, which doesn’t have nearly as much information as the original CPMB Unit 16.15. Perhaps the

  8. Pharmacodynamics of cidofovir for vaccinia virus infection in an in vitro hollow-fiber infection model system.

    Science.gov (United States)

    McSharry, James J; Deziel, Mark R; Zager, Kris; Weng, Qingmei; Drusano, George L

    2009-01-01

    Variola major virus remains a potent weapon of bioterror. There is currently an investigational-new-drug application for cidofovir for the therapy of variola major virus infections. Stittelaar and colleagues compared the levels of effectiveness of postexposure smallpox vaccination (Elstree-RIVM) and antiviral treatment with cidofovir or an acyclic nucleoside phosphonate analogue 6-[2-(phosphonomethoxy)alkoxy]-2,4-diaminopyrimidine (HPMPO-DAPy) after lethal intratracheal infection of cynomolgus monkeys with monkeypox virus, a variola virus surrogate. Their results demonstrated that either compound was more effective than vaccination with the Ellstree vaccine (K. J. Stittelaar et al., Nature 439:745-748, 2006). An unanswered question is how to translate this information into therapy for poxvirus infections in people. In a proof-of-principle study, we used a novel in vitro hollow-fiber infection model system to determine the pharmacodynamics of vaccinia virus infection of HeLa-S3 cells treated with cidofovir. Our results demonstrate that the currently licensed dose of cidofovir of 5 mg/kg of body weight weekly with probenecid (which ameliorates nephrotoxicity) is unlikely to provide protection for patients intentionally exposed to Variola major virus. We further demonstrate that the antiviral effect is independent of the schedule of drug administration. Exposures (area under the concentration-time curve) to cidofovir that will have a robust protective effect will require doses that are 5 to 10 times that currently administered to humans. Such doses may cause nephrotoxicity, and therefore, approaches that include probenecid administration as well as schedules of administration that will help ameliorate the uptake of cidofovir into renal tubular epithelial cells need to be considered when addressing such treatment for people.

  9. Myxoma and Vaccinia Viruses Bind Differentially to Human Leukocytes

    OpenAIRE

    Chan, Winnie M.; Bartee, Eric C.; Moreb, Jan S.; Dower, Ken; Connor, John H.; McFadden, Grant

    2013-01-01

    Myxoma virus (MYXV) and vaccinia virus (VACV), two distinct members of the family Poxviridae, are both currently being developed as oncolytic virotherapeutic agents. Recent studies have demonstrated that ex vivo treatment with MYXV can selectively recognize and kill contaminating cancerous cells from autologous bone marrow transplants without perturbing the engraftment of normal CD34+ hematopoietic stem and progenitor cells. However, the mechanism(s) by which MYXV specifically recognizes and ...

  10. Inhibition of Translation Initiation by Protein 169: A Vaccinia Virus Strategy to Suppress Innate and Adaptive Immunity and Alter Virus Virulence.

    Directory of Open Access Journals (Sweden)

    Pavla Strnadova

    2015-09-01

    Full Text Available Vaccinia virus (VACV is the prototypic orthopoxvirus and the vaccine used to eradicate smallpox. Here we show that VACV strain Western Reserve protein 169 is a cytoplasmic polypeptide expressed early during infection that is excluded from virus factories and inhibits the initiation of cap-dependent and cap-independent translation. Ectopic expression of protein 169 causes the accumulation of 80S ribosomes, a reduction of polysomes, and inhibition of protein expression deriving from activation of multiple innate immune signaling pathways. A virus lacking 169 (vΔ169 replicates and spreads normally in cell culture but is more virulent than parental and revertant control viruses in intranasal and intradermal murine models of infection. Intranasal infection by vΔ169 caused increased pro-inflammatory cytokines and chemokines, infiltration of pulmonary leukocytes, and lung weight. These alterations in innate immunity resulted in a stronger CD8+ T-cell memory response and better protection against virus challenge. This work illustrates how inhibition of host protein synthesis can be a strategy for virus suppression of innate and adaptive immunity.

  11. GMCSF-armed vaccinia virus induces an antitumor immune response.

    Science.gov (United States)

    Parviainen, Suvi; Ahonen, Marko; Diaconu, Iulia; Kipar, Anja; Siurala, Mikko; Vähä-Koskela, Markus; Kanerva, Anna; Cerullo, Vincenzo; Hemminki, Akseli

    2015-03-01

    Oncolytic Western Reserve strain vaccinia virus selective for epidermal growth factor receptor pathway mutations and tumor-associated hypermetabolism was armed with human granulocyte-macrophage colony-stimulating factor (GMCSF) and a tdTomato fluorophore. As the assessment of immunological responses to human transgenes is challenging in the most commonly used animal models, we used immunocompetent Syrian golden hamsters, known to be sensitive to human GMCSF and semipermissive to vaccinia virus. Efficacy was initially tested in vitro on various human and hamster cell lines and oncolytic potency of transgene-carrying viruses was similar to unarmed virus. The hGMCSF-encoding virus was able to completely eradicate subcutaneous pancreatic tumors in hamsters, and to fully protect the animals from subsequent rechallenge with the same tumor. Induction of specific antitumor immunity was also shown by ex vivo co-culture experiments with hamster splenocytes. In addition, histological examination revealed increased infiltration of neutrophils and macrophages in GMCSF-virus-treated tumors. These findings help clarify the mechanism of action of GMCSF-armed vaccinia viruses undergoing clinical trials.

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

    Science.gov (United States)

    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

    2014-01-01

    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

  13. Logistics of community smallpox control through contact tracing and ring vaccination: a stochastic network model

    Directory of Open Access Journals (Sweden)

    Portnoy Diane L

    2004-08-01

    Full Text Available Abstract Background Previous smallpox ring vaccination models based on contact tracing over a network suggest that ring vaccination would be effective, but have not explicitly included response logistics and limited numbers of vaccinators. Methods We developed a continuous-time stochastic simulation of smallpox transmission, including network structure, post-exposure vaccination, vaccination of contacts of contacts, limited response capacity, heterogeneity in symptoms and infectiousness, vaccination prior to the discontinuation of routine vaccination, more rapid diagnosis due to public awareness, surveillance of asymptomatic contacts, and isolation of cases. Results We found that even in cases of very rapidly spreading smallpox, ring vaccination (when coupled with surveillance is sufficient in most cases to eliminate smallpox quickly, assuming that 95% of household contacts are traced, 80% of workplace or social contacts are traced, and no casual contacts are traced, and that in most cases the ability to trace 1–5 individuals per day per index case is sufficient. If smallpox is assumed to be transmitted very quickly to contacts, it may at times escape containment by ring vaccination, but could be controlled in these circumstances by mass vaccination. Conclusions Small introductions of smallpox are likely to be easily contained by ring vaccination, provided contact tracing is feasible. Uncertainties in the nature of bioterrorist smallpox (infectiousness, vaccine efficacy support continued planning for ring vaccination as well as mass vaccination. If initiated, ring vaccination should be conducted without delays in vaccination, should include contacts of contacts (whenever there is sufficient capacity and should be accompanied by increased public awareness and surveillance.

  14. The H4 subunit of vaccinia virus RNA polymerase is not required for transcription initiation at a viral late promoter.

    OpenAIRE

    Wright, C F; Coroneos, A M

    1995-01-01

    Chromatography of RNA polymerase purified from vaccinia virions and from vaccinia virus-infected HeLa cells resulted in the separation of populations active for early and late transcription. An RNA polymerase population immunodepleted for the vaccinia virus H4 gene peptide could support late transcription.

  15. Progress in the research of smallpox vaccines%天花疫苗研究进展

    Institute of Scientific and Technical Information of China (English)

    孟昕; 阮力

    2010-01-01

    In the last decade, the research on smallpox vaccine has been paid much more attention because of the special position of variola virus in bioterrorism and biological warfare. To date, 4 generations of smallpox vaccines have been developed, and of which 3 generations can be used. Among them, the first generation smallpox vaccines play an important role in the achievement of global smallpox eradication, but their adverse reactions are strong. The second generation smallpox vaccines possess the theoretical advantages conferred by improved manufacturing process and lowered risk of contamination by adventitious agents. The third generation smallpox vaccines have reduced virulence and improved safety, but the immunological efficacy has also descended. The fourth generation smallpox vaccines are developed by techniques for genetic engineering, and the efficacy remains to be evaluated. Owing to global smallpox eradication, the evaluation of the immunological efficacy of new smallpox vaccines becomes the key procedure which can hinder the development of new vaccines.%鉴于天花病毒在生物恐怖和生物战中的特殊地位,近十年来,天花疫苗的研究重新得到重视.到目前为止,天花疫苗研究已进入第四代,已有三代天花疫苗可供使用.其中,第一代天花疫苗在全球消灭天花中起到重要和关键的作用,但不良反应较强.第二代疫苗改进了生产工艺,降低了外源因子污染的可能.第三代疫苗有效地降低了疫苗的毒力,增强了安全性,但免疫效力有所下降.第四代疫苗系使用基因工程技术研制的疫苗,效果尚待评价.由于天花已经消灭,在新疫苗的研究中,如何评价其免疫效果成为妨碍疫苗研究进展的关键环节.

  16. Efficacy of tecovirimat (ST-246) in nonhuman primates infected with variola virus (Smallpox).

    Science.gov (United States)

    Mucker, Eric M; Goff, Arthur J; Shamblin, Joshua D; Grosenbach, Douglas W; Damon, Inger K; Mehal, Jason M; Holman, Robert C; Carroll, Darin; Gallardo, Nadia; Olson, Victoria A; Clemmons, Cody J; Hudson, Paul; Hruby, Dennis E

    2013-12-01

    Naturally occurring smallpox has been eradicated but remains a considerable threat as a biowarfare/bioterrorist weapon (F. Fleck, Bull. World Health Organ. 81:917-918, 2003). While effective, the smallpox vaccine is currently not recommended for routine use in the general public due to safety concerns (http://www.bt.cdc.gov/agent/smallpox/vaccination). Safe and effective countermeasures, particularly those effective after exposure to smallpox, are needed. Currently, SIGA Technologies is developing the small-molecule oral drug, tecovirimat (previously known as ST-246), as a postexposure therapeutic treatment of orthopoxvirus disease, including smallpox. Tecovirimat has been shown to be efficacious in preventing lethal orthopoxviral disease in numerous animal models (G. Yang, D. C. Pevear, M. H. Davies, M. S. Collett, T. Bailey, et al., J. Virol. 79:13139-13149, 2005; D. C. Quenelle, R. M. Buller, S. Parker, K. A. Keith, D. E. Hruby, et al., Antimicrob. Agents Chemother., 51:689-695, 2007; E. Sbrana, R. Jordan, D. E. Hruby, R. I. Mateo, S. Y. Xiao, et al., Am. J. Trop. Med. Hyg. 76:768-773, 2007). Furthermore, in clinical trials thus far, the drug appears to be safe, with a good pharmacokinetic profile. In this study, the efficacy of tecovirimat was evaluated in both a prelesional and postlesional setting in nonhuman primates challenged intravenously with 1 × 10(8) PFU of Variola virus (VARV; the causative agent of smallpox), a model for smallpox disease in humans. Following challenge, 50% of placebo-treated controls succumbed to infection, while all tecovirimat-treated animals survived regardless of whether treatment was started at 2 or 4 days postinfection. In addition, tecovirimat treatment resulted in dramatic reductions in dermal lesion counts, oropharyngeal virus shedding, and viral DNA circulating in the blood. Although clinical disease was evident in tecovirimat-treated animals, it was generally very mild and appeared to resolve earlier than in placebo

  17. [Lessons learnt from the German smallpox outbreaks after World War II].

    Science.gov (United States)

    Sasse, Julia; Gelderblom, Hans R

    2015-07-01

    Even though smallpox was declared eradicated by WHO in 1980, it cannot be ruled out that the etiological variola virus could be used as a biological weapon. Undestroyed viruses from biowarfare programmes, virus strains left undetected in a freezer or dangerous recombinant poxvirus constructs could cause dangerous outbreaks in a relatively unprotected population. Despite an abundance of studies performed during the eradication of smallpox, epidemiological data for preparedness planning and outbreak control in modern, industrialized countries are scarce. Full-text hand search for the period from 1945 to 1975 in the main German public health journals. After World War II 12 smallpox outbreaks occurred in Germany. They were studied with the focus on the period of contagiousness, the protective effect of vaccination, booster-effect of revaccination and the place of infection. A total of 95 individuals contracted smallpox, including 10 fatalities. Despite having been previously vaccinated, 81 vaccinated persons came down with smallpox, yet 91% of them developed only mild symptoms. These patients presented a high risk for spreading the infection to contact persons due to misinterpretation of symptoms and the continuing social contacts. Basically, the risk of transmission in the first 2 to 3 days after onset of symptoms was low, thus facilitating antiepidemic measures. The importance of hospital preparedness is emphasized by the fact that most infections occurred in hospitals. The data analyzed provide valuable information for today's outbreak response planning and counter bioterrorism preparedness.

  18. Tecovirimat, a p37 envelope protein inhibitor for the treatment of smallpox infection.

    Science.gov (United States)

    Duraffour, Sophie; Andrei, Graciela; Snoeck, Robert

    2010-03-01

    Since the eradication of naturally occurring smallpox in 1980, the fear that variola virus could be used as a biological weapon has become real. Over the last 10 years, emergency preparedness programs have been launched to protect populations against a smallpox outbreak or the possible emergence in humans of other orthopoxvirus infections, such as monkeypox. Vaccination against smallpox was responsible for its eradication, but was linked with high rates of adverse events and contraindications. In this context, intensive research in the poxvirus field has led to the development of safer vaccines and to an increase in the number of anti-poxvirus agents in the pipeline. SIGA Technologies Inc, under license from ViroPharma Inc, is developing tecovirimat (ST-246). Tecovirimat is a novel antiviral that inhibits the egress of orthopoxviruses by targeting viral p37 protein orthologs. The development of tecovirimat during the last 5 years for the treatment of smallpox and for its potential use as adjunct to smallpox vaccine is reviewed here.

  19. A chemokine-binding domain in the tumor necrosis factor receptor from variola (smallpox) virus.

    Science.gov (United States)

    Alejo, Alí; Ruiz-Argüello, M Begoña; Ho, Yin; Smith, Vincent P; Saraiva, Margarida; Alcami, Antonio

    2006-04-11

    Variola virus (VaV) is the causative agent of smallpox, one of the most devastating diseases encountered by man, that was eradicated in 1980. The deliberate release of VaV would have catastrophic consequences on global public health. However, the mechanisms that contribute to smallpox pathogenesis are poorly understood at the molecular level. The ability of viruses to evade the host defense mechanisms is an important determinant of viral pathogenesis. Here we show that the tumor necrosis factor receptor (TNFR) homologue CrmB encoded by VaV functions not only as a soluble decoy TNFR but also as a highly specific binding protein for several chemokines that mediate recruitment of immune cells to mucosal surfaces and the skin, sites of virus entry and viral replication at late stages of smallpox. CrmB binds chemokines through its C-terminal domain, which is unrelated to TNFRs, was named smallpox virus-encoded chemokine receptor (SECRET) domain and uncovers a family of poxvirus chemokine inhibitors. An active SECRET domain was found in another viral TNFR (CrmD) and three secreted proteins encoded by orthopoxviruses. These findings identify a previously undescribed chemokine-binding and inhibitory domain unrelated to host chemokine receptors and a mechanism of immune modulation in VaV that may influence smallpox pathogenesis.

  20. SIMULTANEOUS BCG AND SMALLPOX VACCINATION ON NEWBORN INFANTS

    Directory of Open Access Journals (Sweden)

    Abdul Rivai

    2012-09-01

    Full Text Available Telah dikemukakan anggapan-anggapan yang terdapat dewasa ini tentang vaksinasi BCG dan cacar secara simultan. Telah dilakukan vaksinasi BCG dan cacar secara simultan pada 729 neonati dengan freeze dried Smallpox vaccine buatan dari Bio Farma dan freeze dried BCG vaccine Tokyo. Pencacaran dilakukan secara multiple puncture dan bifurcated needle dengan suntikan BCG dengan jarum dan spuit khusus intracutan dengan dosis 0,1 ml. Tuberkulin test dilakukan dengan PPD dari Kopenhagen dengan kekuatan 2 TU 9 minggu setelah vaksinasi. Dari 741 bayi yang diikut sertakan dalam survey, 12 menolak, 3 bayi tidak dapat dilakukan pemeriksaan pertama, 35 bayi belum diperiksa, pemeriksaan pertama telah dilakukan pada 691 bayi. Dari 406 bayi yang seharusnya sudah diperiksa untuk pemeriksaan kedua, 23 dapat dilakukan karena tidak dapat dijumpai atau meninggal. Telah dikemukakan bahwa pencatatan alamat yang jelas dan lengkap serta kesungguhan dalam melakukan home visits sangat penting untuk berhasilnya penyelidikan semacam ini. Dari hasil-hasil yang didapatkan sampai sekarang telah dapat diambil kesimpulan sementara, bahwa vaksinasi BCG dan cacar secara simultan memberikan hasil yang memuaskan, juga bila dibandingkan dengan hasil-hasil penyelidikan diluar negeri take pada pencacaran 99.4 percent, test tuberkulin dengan PPD 2 TU 9 minggu setelah vaksinasi memberikan indurasi lebih dari 5 mm pada 99.75 percent dan tidak menimbulkan komplikasi-komplikasi. Pelaksanaan vaksinasi BCG dan cacar dapat dilakukan oleh tenaga paramedis yang telah mendapat latihan khusus dan diawasi oleh dokter yang kompeten. Dianjurkan untuk melakukan follow up pada bayi-bayi yang diikut sertakan dalam survey ini.

  1. SIMULTANEOUS BCG AND SMALLPOX VACCINATION ON NEWBORN INFANTS

    Directory of Open Access Journals (Sweden)

    Abdul Rivai

    2012-09-01

    Full Text Available Telah dikemukakan anggapan-anggapan yang terdapat dewasa ini tentang vaksinasi BCG dan cacar secara simultan. Telah dilakukan vaksinasi BCG dan cacar secara simultan pada 729 neonati dengan freeze dried Smallpox vaccine buatan dari Bio Farma dan freeze dried BCG vaccine Tokyo. Pencacaran dilakukan secara multiple puncture dan bifurcated needle dengan suntikan BCG dengan jarum dan spuit khusus intracutan dengan dosis 0,1 ml. Tuberkulin test dilakukan dengan PPD dari Kopenhagen dengan kekuatan 2 TU 9 minggu setelah vaksinasi. Dari 741 bayi yang diikut sertakan dalam survey, 12 menolak, 3 bayi tidak dapat dilakukan pemeriksaan pertama, 35 bayi belum diperiksa, pemeriksaan pertama telah dilakukan pada 691 bayi. Dari 406 bayi yang seharusnya sudah diperiksa untuk pemeriksaan kedua, 23 dapat dilakukan karena tidak dapat dijumpai atau meninggal. Telah dikemukakan bahwa pencatatan alamat yang jelas dan lengkap serta kesungguhan dalam melakukan home visits sangat penting untuk berhasilnya penyelidikan semacam ini. Dari hasil-hasil yang didapatkan sampai sekarang telah dapat diambil kesimpulan sementara, bahwa vaksinasi BCG dan cacar secara simultan memberikan hasil yang memuaskan, juga bila dibandingkan dengan hasil-hasil penyelidikan diluar negeri take pada pencacaran 99.4 percent, test tuberkulin dengan PPD 2 TU 9 minggu setelah vaksinasi memberikan indurasi lebih dari 5 mm pada 99.75 percent dan tidak menimbulkan komplikasi-komplikasi. Pelaksanaan vaksinasi BCG dan cacar dapat dilakukan oleh tenaga paramedis yang telah mendapat latihan khusus dan diawasi oleh dokter yang kompeten. Dianjurkan untuk melakukan follow up pada bayi-bayi yang diikut sertakan dalam survey ini.

  2. Multifaceted contributions: health workers and smallpox eradication in India.

    Science.gov (United States)

    Bhattacharya, Sanjoy

    2008-01-01

    Smallpox eradication in South Asia was a result of the efforts of many grades of health-workers. Working from within the confines of international organisations and government structures, the role of the field officials, who were of various nationalities and also drawn from the cities and rural enclaves of the countries in these regions, was crucial to the development and deployment of policies. However, the role of these personnel is often downplayed in official histories and academic histories, which highlight instead the roles played by a handful of senior officials within the World Health Organization and the federal governments in the sub-continent. This article attempts to provide a more rounded assessment of the complex situation in the field. In this regard, an effort is made to underline the great usefulness of the operational flexibility displayed by field officers, wherein lessons learnt in the field were made an integral part of deploying local campaigns; careful engagement with the communities being targeted, as well as the employment of short term workers from amongst them, was an important feature of this work.

  3. Transmission patterns of smallpox: Systematic review of natural outbreaks in Europe and North America since World War II

    NARCIS (Netherlands)

    V. Bhatnagar (Vibha); M.A. Stoto (Michael); S.C. Morton (Sally); R. Boer (Rob); S.A. Bozzette (Samuel)

    2006-01-01

    textabstractBackground: Because smallpox (variola major) may be used as a biological weapon, we reviewed outbreaks in post-World War II Europe and North America in order to understand smallpox transmission patterns. Methods: A systematic review was used to identify papers from the National Library o

  4. The efficacy and pharmacokinetics of brincidofovir for the treatment of lethal rabbitpox virus infection: a model of smallpox disease.

    Science.gov (United States)

    Trost, Lawrence C; Rose, Michelle L; Khouri, Jody; Keilholz, Laurie; Long, James; Godin, Stephen J; Foster, Scott A

    2015-05-01

    Brincidofovir (BCV) has broad-spectrum in vitro activity against dsDNA viruses, including smallpox, and is being developed as a treatment for smallpox as well as infections caused by other dsDNA viruses. BCV has previously been shown to be active in multiple animal models of smallpox. Here we present the results of a randomized, blinded, placebo-controlled study of the efficacy and pharmacokinetics of a novel, "humanized" regimen of BCV for treatment of New Zealand White rabbits infected with a highly lethal inoculum of rabbitpox virus, a well characterized model of smallpox. Compared with placebo, a dose-dependent increase in survival was observed in all BCV-treatment groups. Concentrations of cidofovir diphosphate (CDV-PP), the active antiviral, in rabbit peripheral blood mononuclear cells (PBMCs) were determined for comparison to those produced in humans at the dose proposed for treatment of smallpox. CDV-PP exposure in PBMCs from rabbits given BCV scaled to human exposures at the dose proposed for treatment of smallpox, which is also currently under evaluation for other indications. The results of this study demonstrate the activity of BCV in the rabbitpox model of smallpox and the feasibility of scaling doses efficacious in the model to a proposed human dose and regimen for treatment of smallpox. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

  5. Transmission patterns of smallpox: Systematic review of natural outbreaks in Europe and North America since World War II

    NARCIS (Netherlands)

    V. Bhatnagar (Vibha); M.A. Stoto (Michael); S.C. Morton (Sally); R. Boer (Rob); S.A. Bozzette (Samuel)

    2006-01-01

    textabstractBackground: Because smallpox (variola major) may be used as a biological weapon, we reviewed outbreaks in post-World War II Europe and North America in order to understand smallpox transmission patterns. Methods: A systematic review was used to identify papers from the National Library o

  6. The discontinuation of routine smallpox vaccination in the United States, 1960-1976: an unlikely affirmation of biomedical hegemony.

    Science.gov (United States)

    Rich, Miriam

    2011-02-01

    This article seeks to understand the discursive context of the cessation of routine smallpox vaccination in the United States in the early 1970s. The United States has a long tradition of opposition to compulsory smallpox vaccination, usually expressed in terms of concerns about personal liberties, the extent of state authority, and challenges to the hegemony of orthodox biomedicine. The practice of routine smallpox vaccination continued in the United States until its termination in the 1970s, following a 1971 recommendation against the practice issued by the United States Public Health Service. This history investigates the ways in which opposition to compulsory smallpox vaccination in the 1960s and 70s was articulated and understood by contemporaries through an analysis of the rhetoric used in leading medical journals and popular newspapers. It finds that this ultimately successful movement to end routine smallpox vaccination drew upon the language of biomedical authority rather than political protest.

  7. Safety of attenuated smallpox vaccine LC16m8 in immunodeficient mice.

    Science.gov (United States)

    Yokote, Hiroyuki; Shinmura, Yasuhiko; Kanehara, Tomomi; Maruno, Shinichi; Kuranaga, Masahiko; Matsui, Hajime; Hashizume, So

    2014-09-01

    Freeze-dried live attenuated smallpox vaccine LC16m8 prepared in cell culture has been the sole smallpox vaccine licensed in Japan since 1975 and was recently recommended as a WHO stockpile vaccine. We evaluated the safety of recently remanufactured lots of LC16m8 using a series of immunodeficient mouse models. These models included suckling mice, severe combined immunodeficiency disease (SCID) mice, and wild-type mice treated with cyclosporine. LC16m8 showed extremely low virulence in each of the three mouse models compared with that of its parental strains, Lister and LC16mO. These results provide further evidence that LC16m8 is one of the safest replication-competent smallpox vaccines in the world and may be considered for use in immunodeficient patients.

  8. Smallpox as an actual biothreat: lessons learned from its outbreak in ex-Yugoslavia in 1972.

    Science.gov (United States)

    Ristanovic, Elizabeta; Gligic, Ana; Atanasievska, Sonja; Protic-Djokic, Vesna; Jovanovic, Dragutin; Radunovic, Miodrag

    2016-01-01

    Variola (smallpox) virus is classified as class A of potential biological weapons, due to its microbiological, genetic, antigenic and epidemiological characteristics. The potential danger is more real because vaccination against smallpox has stopped since disease eradication in 1979. That is why we want to share our unique, rich experience and acquired knowledge in the fight against this highly contagious and deadly disease during the smallpox outbreak in ex-Yugoslavia in 1972. It was the largest postwar outbreak in Europe when there were officially registered 175 ill patients, 35 of them with lethal outcome. This outbreak was specific by the time of its occurrence, the affected territory, dimensions and some epidemiological characteristics, but also by the well-organized, synchronized and efficient reaction of the competent state services in the fight against it.

  9. The eradication of smallpox: organizational learning and innovation in international health administration.

    Science.gov (United States)

    Hopkins, J W

    1988-04-01

    The WHO smallpox eradication campaign represents perhaps the best example of a successful international health administration. In the 1st year of the campaign (1967), the guiding strategy was to vaccinate people en masse over a 2-3 year period in countries where smallpox was epidemic thereby conquering the disease. In Western Nigeria where 90% of the population had been vaccinated, a smallpox outbreak occurred in a religious sect resisting vaccinations and a delay in delivery of supplies forced a change in strategy. Campaign staff learned to rapidly isolate infected persons and swiftly vaccinate the uninfected in an outbreak area in order to break the transmission of smallpox, even where 1/2 the population had been vaccinated. Technological advancements also contributed to the campaign's success. For example, the jet injector vaccinated 1000 people/hour with efficient, reliable, mass produced potent, stable freeze dried vaccines (often produced in target countries) or the less costly and virtually maintenance free bifurcated needle was used. The most significant contribution to the success of the campaign, however, was the flexible mode of management adopted by the campaign staff at WHO which provided an appropriate environment for organizational learning and innovation. Although management was open and flexible, the campaign did depend on careful planning and setting of goals, continual assessment, and rapid response to field requests for assistance or advice. Trends in the incidence of smallpox was chosen as the indicator of success as opposed to the number of vaccinations. The campaign demonstrated the need for cultural adaptations as it operated in each country and region. This evaluation of the success of the smallpox campaign presents conclusions that serve as guidelines to the organization and administration of international programs designed to solve other health problems.

  10. Modeling the Effect of Herd Immunity and Contagiousness in Mitigating a Smallpox Outbreak.

    Science.gov (United States)

    Graeden, Ellie; Fielding, Russel; Steinhouse, Kyle E; Rubin, Ilan N

    2015-07-01

    The smallpox antiviral tecovirimat has recently been purchased by the U.S. Strategic National Stockpile. Given significant uncertainty regarding both the contagiousness of smallpox in a contemporary outbreak and the efficiency of a mass vaccination campaign, vaccine prophylaxis alone may be unable to control a smallpox outbreak following a bioterror attack. Here, we present the results of a compartmental epidemiological model that identifies conditions under which tecovirimat is required to curtail the epidemic by exploring how the interaction between contagiousness and prophylaxis coverage of the affected population affects the ability of the public health response to control a large-scale smallpox outbreak. Each parameter value in the model is based on published empirical data. We describe contagiousness parametrically using a novel method of distributing an assumed R-value over the disease course based on the relative rates of daily viral shedding from human and animal studies of cognate orthopoxvirus infections. Our results suggest that vaccination prophylaxis is sufficient to control the outbreak when caused either by a minimally contagious virus or when a very high percentage of the population receives prophylaxis. As vaccination coverage of the affected population decreases below 70%, vaccine prophylaxis alone is progressively less capable of controlling outbreaks, even those caused by a less contagious virus (R0 less than 4). In these scenarios, tecovirimat treatment is required to control the outbreak (total number of cases under an order of magnitude more than the number of initial infections). The first study to determine the relative importance of smallpox prophylaxis and treatment under a range of highly uncertain epidemiological parameters, this work provides public health decision-makers with an evidence-based guide for responding to a large-scale smallpox outbreak.

  11. Protein Primary Structure of the Vaccinia Virion at Increased Resolution

    Science.gov (United States)

    Ngo, Tuan; Mirzakhanyan, Yeva; Moussatche, Nissin; Gershon, Paul David

    2016-11-01

    Here we examine the protein covalent structure of the vaccinia virus virion. Within two virion preparations, >88% of the theoretical vaccinia virus-encoded proteome was detected with high confidence, including the first detection of products from 27 open reading frames (ORFs) previously designated "predicted," "uncharacterized," "inferred," or "hypothetical" polypeptides containing as few as 39 amino acids (aa) and six proteins whose detection required nontryptic proteolysis. We also detected the expression of four short ORFs, each of which was located within an ORF ("ORF-within-ORF"), including one not previously recognized or known to be expressed. Using quantitative mass spectrometry (MS), between 58 and 74 proteins were determined to be packaged. A total of 63 host proteins were also identified as candidates for packaging. Evidence is provided that some portion of virion proteins are "nicked" via a combination of endoproteolysis and concerted exoproteolysis in a manner, and at sites, independent of virus origin or laboratory procedures. The size of the characterized virion phosphoproteome was doubled from 189 (J. Matson, W. Chou, T. Ngo, and P. D. Gershon, Virology 452-453:310-323, 2014, doi:http://dx.doi.org/10.1016/j.virol.2014.01.012) to 396 confident, unique phosphorylation sites, 268 of which were within the packaged proteome. This included the unambiguous identification of phosphorylation "hot spots" within virion proteins. Using isotopically enriched ATP, 23 sites of intravirion kinase phosphorylation were detected within nine virion proteins, all at sites already partially occupied within the virion preparations. The clear phosphorylation of proteins RAP94 and RP19 was consistent with the roles of these proteins in intravirion early gene transcription. In a blind search for protein modifications, cysteine glutathionylation and O-linked glycosylation featured prominently. We provide evidence for the phosphoglycosylation of vaccinia virus proteins.

  12. [Study of allergic reactions in the body after experimental smallpox vaccination].

    Science.gov (United States)

    Vorob'ev, A A; Afanas'ev, S S; Patrikeev, G T; Prigoda, A S; Podkuiĭko, V N

    1977-09-01

    Experiments on guinea pigs demonstrated a mixed cellular-humoral character of allergy in smallpox vaccination. The ratio of cellular and humoral components depended on the site of the vaccina application. Skin manifestations in smallpox vaccination (hyperemia, infiltration, papula) were due to the multiplication of the virus proper, increased sensitivity of the surrounding tissues to the live vibrio, and specific immunological reconstruction. The skin of the sensitized animal after the anaphylactic shock in response to the intracardiac injection of the challenging dose of the vaccine virus acquired areactivity to its subsequent application.

  13. New effective chemically synthesized anti-smallpox compound NIOCH-14.

    Science.gov (United States)

    Mazurkov, Oleg Yu; Kabanov, Alexey S; Shishkina, Larisa N; Sergeev, Alexander A; Skarnovich, Maksim O; Bormotov, Nikolay I; Skarnovich, Maria A; Ovchinnikova, Alena S; Titova, Ksenya A; Galahova, Darya O; Bulychev, Leonid E; Sergeev, Artemiy A; Taranov, Oleg S; Selivanov, Boris A; Tikhonov, Alexey Ya; Zavjalov, Evgenii L; Agafonov, Alexander P; Sergeev, Alexander N

    2016-05-01

    Antiviral activity of the new chemically synthesized compound NIOCH-14 (a derivative of tricyclodicarboxylic acid) in comparison with ST-246 (the condensed derivative of pyrroledione) was observed in experiments in vitro and in vivo using orthopoxviruses including highly pathogenic ones. After oral administration of NIOCH-14 to outbred ICR mice infected intranasally with 100 % lethal dose of ectromelia virus, it was shown that 50 % effective doses of NIOCH-14 and ST-246 did not significantly differ. The 'therapeutic window' varied from 1 day before infection to 6 days post-infection (p.i.) to achieve 100-60 % survival rate. The administration of NIOCH-14 and ST-246 to mice resulted in a significant reduction of ectromelia virus titres in organs examined as compared with the control and also reduced pathological changes in the lungs 6 days p.i. Oral administration of NIOCH-14 and ST-246 to ICR mice and marmots challenged with monkeypox virus as compared with the control resulted in a significant reduction of virus production in the lungs and the proportion of infected mice 7 days p.i. as well as the absence of disease in marmots. Significantly lower proportions of infected mice and virus production levels in the lungs as compared with the control were demonstrated in experiments after oral administration of NIOCH-14 and ST-246 to ICR mice and immunodeficient SCID mice challenged with variola virus 3 and 4 days p.i., respectively. The results obtained suggest good prospects for further study of the chemical compound NIOCH-14 to create a new smallpox drug on its basis.

  14. A new inhibitor of apoptosis from vaccinia virus and eukaryotes.

    Directory of Open Access Journals (Sweden)

    Caroline Gubser

    2007-02-01

    Full Text Available A new apoptosis inhibitor is described from vaccinia virus, camelpox virus, and eukaryotic cells. The inhibitor is a hydrophobic, multiple transmembrane protein that is resident in the Golgi and is named GAAP (Golgi anti-apoptotic protein. Stable expression of both viral GAAP (v-GAAP and human GAAP (h-GAAP, which is expressed in all human tissues tested, inhibited apoptosis induced by intrinsic and extrinsic apoptotic stimuli. Conversely, knockout of h-GAAP by siRNA induced cell death by apoptosis. v-GAAP and h-GAAP display overlapping functions as shown by the ability of v-GAAP to complement for the loss of h-GAAP. Lastly, deletion of the v-GAAP gene from vaccinia virus did not affect virus replication in cell culture, but affected virus virulence in a murine infection model. This study identifies a new regulator of cell death that is highly conserved in evolution from plants to insects, amphibians, mammals, and poxviruses.

  15. Successive site translocating inoculation potentiates DNA/recombinant vaccinia vaccination.

    Science.gov (United States)

    Ren, Yanqin; Wang, Na; Hu, Weiguo; Zhang, Xiaoyan; Xu, Jianqing; Wan, Yanmin

    2015-12-15

    DNA vaccines have advantages over traditional vaccine modalities; however the relatively low immunogenicity restrains its translation into clinical use. Further optimizations are needed to get the immunogenicity of DNA vaccine closer to the level required for human use. Here we show that intramuscularly inoculating into a different limb each time significantly improves the immunogenicities of both DNA and recombinant vaccinia vaccines during multiple vaccinations, compared to repeated vaccination on the same limb. We term this strategy successive site translocating inoculation (SSTI). SSTI could work in synergy with genetic adjuvant and DNA prime-recombinant vaccinia boost regimen. By comparing in vivo antigen expression, we found that SSTI avoided the specific inhibition of in vivo antigen expression, which was observed in the limbs being repeatedly inoculated. Employing in vivo T cell depletion and passive IgG transfer, we delineated that the inhibition was not mediated by CD8(+) T cells but by specific antibodies. Finally, by using C3(-/-) mouse model and in vivo NK cells depletion, we identified that specific antibodies negatively regulated the in vivo antigen expression primarily in a complement depended way.

  16. In silico-accelerated identification of conserved and immunogenic variola/vaccinia T-cell epitopes

    DEFF Research Database (Denmark)

    Moise, Leonard; McMurry, Julie A; Buus, Søren

    2009-01-01

    Epitopes shared by the vaccinia and variola viruses underlie the protective effect of vaccinia immunization against variola infection. We set out to identify a subset of cross-reactive epitopes using bioinformatics and immunological methods. Putative T-cell epitopes were computationally predicted...... from highly conserved open reading frames from seven complete vaccinia and variola genomes using EpiMatrix. Over 100 epitopes bearing low human sequence homology were selected and assessed in HLA binding assays and in T-cell antigenicity assays using PBMCs isolated from Dryvax-immunized subjects...

  17. Non-coding RNAs and heme oxygenase-1 in vaccinia virus infection

    Science.gov (United States)

    Meseda, Clement A.; Srinivasan, Kumar; Wise, Jasen; Catalano, Jennifer; Yamada, Kenneth M.; Dhawan, Subhash

    2017-01-01

    Small nuclear RNAs (snRNAs) are undetermined, a population of snRNAs is produced during the early phase of infection of cells by vaccinia virus. In the present study, we demonstrate a direct correlation between expression of the cytoprotective enzyme heme oxygenase-1 (HO-1), suppression of selective snRNA expression, and inhibition of vaccinia virus infection of macrophages. Hemin induced HO-1 expression, completely reversed virus-induced host snRNA expression, and suppressed vaccinia virus infection. This involvement of specific virus-induced snRNAs and associated gene clusters suggests a novel HO-1-dependent host-defense pathway in poxvirus infection. PMID:25450361

  18. Smallpox inoculation (variolation) in East Africa with special reference to the practice among the Boran and Gabra of Northern Kenya.

    Science.gov (United States)

    Imperato, Pascal James; Imperato, Gavin H

    2014-12-01

    Smallpox inoculation (variolation) was widely reported in sub-Sahara Africa before, during, and after the colonial era. The infective smallpox materials and techniques used, as well as the anatomical sites for inoculation, varied widely among different ethnic groups. The practice among the Boran and Gabra pastoralists of northern Kenya resembled that which was prevalent in a number of areas of Ethiopia. This is not surprising as the Boran also live in southern Ethiopia, and Gabra herdsmen frequently cross the border into this region. The Boran and Gabra technique for smallpox inoculation consisted of taking infective material from the vesicles or pustules of those with active smallpox, and scraping it into the skin on the dorsum of the lower forearm. Although the intent was to cause a local reaction and at most a mild form of smallpox, severe cases of the disease not infrequently resulted. Also, variolated individuals were capable of infecting others with smallpox, thereby augmenting outbreaks and sustaining them. The limited known reports of smallpox inoculation among the Boran and Gabra are presented in this communication. The expansion of vaccination with effective heat stable vaccines, the development of medical and public health infrastructures, and educational programs all contributed to the eventual disappearance of the practice among the Boran and Gabra.

  19. The role of vaccinia termination factor and cis-acting elements in vaccinia virus early gene transcription termination.

    Science.gov (United States)

    Tate, Jessica; Gollnick, Paul

    2015-11-01

    Vaccinia virus early gene transcription termination requires the virion form of the viral RNA polymerase (vRNAP), Nucleoside Triphosphate Phosphohydrolase I (NPHI), ATP, the vaccinia termination factor (VTF), and a U5NU termination signal in the nascent transcript. VTF, also the viral mRNA capping enzyme, binds U5NU, and NPHI hydrolyzes ATP to release the transcript. NPHI can release transcripts independent of VTF and U5NU if vRNAP is not actively elongating. However, VTF and U5NU are required for transcript release from an elongating vRNAP, suggesting that the function of VTF and U5NU may be to stall the polymerase. Here we demonstrate that VTF inhibits transcription elongation by enhancing vRNAP pausing. Hence VTF provides the connection between the termination signal in the RNA transcript and viral RNA polymerase to initiate transcription termination. We also provide evidence that a second cis-acting element downstream of U5NU influences the location and efficiency of early gene transcription termination.

  20. Les hasards de la variole The hazards of smallpox

    Directory of Open Access Journals (Sweden)

    Jean-Marc Rohrbasser

    2011-12-01

    Full Text Available La nécessité d’un calcul ayant pour fin d’estimer un risque peut être révoquée en doute lorsqu’il s’agit de prendre une décision en situation d’incertitude, a fortiori lorsqu’il s’agit d’une question de vie ou de mort. Dans la controverse engagée sur l’opportunité d’inoculer la variole, la position de D’Alembert constitue un cas exemplaire de scepticisme portant sur l’application des mathématiques, et en l’occurrence du calcul des probabilités, à des décisions relatives à la vie humaine. D’Alembert, en effet, conteste aux mathématiques sociales le pouvoir de rendre compte de phénomènes humains en y cherchant des régularités et des formalisations sans dissocier les dimensions mathématiques et probabilistes des dimensions philosophiques et éthiques. En suivant le débat qui, au milieu du xviiie siècle, se déroule entre le mathématicien français et son homologue suisse Daniel Bernoulli, on assiste à l’un des épisodes de la lente gestation des notions de prise de risque, de décision et de rationalité.The necessity of a calculation aiming to evaluate a risk can be revoked in doubt when the question is to make a decision in a situation of uncertainty, all the more when the question is about life or death. In the controversy opened on the opportunity to inoculate the smallpox, D'Alembert’s position constitutes an exemplary case of scepticism concerning the application of the mathematics, and in this particular case the probability theory, to decisions relative to the human life. D’Alembert, indeed, disputes in the social mathematics the power to describe human phenomena by looking for regularities and for formalizations without separating the mathematical and probability dimensions of the philosophic and ethical ones. By following the debate which, in the middle of the 18th century, takes place between the French mathematician and his Swiss counterpart Daniel Bernoulli, we attend one of the

  1. 痘苗病毒天坛株载体研究与应用概述%Research and application of vaccinia virus Tiantan strain vector

    Institute of Scientific and Technical Information of China (English)

    阮力

    2013-01-01

    1982年,文献首次报道外源基因成功在痘苗病毒WR株(小鼠嗜神经毒株)中获得表达,引起了中国科学家的极大关注.1984年,中国医学科学院病毒学研究所在朱既明院士的组织下成立了痘苗病毒基因表达载体研究协作组,提出使用痘苗病毒天坛株开发可用于人体的痘苗病毒基因表达载体的新思路.该研究历时10余年,成功构建了痘苗病毒天坛株高效表达载体,并广泛用于外源基因表达、基因工程疫苗研究、单克隆抗体研制和诊断试剂开发.本文简单介绍了该载体的研究及应用,并对载体疫苗存在的问题及发展前景进行了讨论.%In 1982, a foreign gene successfully expressed in the vaccinia virus WR strain (mouse neurotropic strain) aroused a great concern of Chinese scientists. In 1984, Dr. ZHU Ji-Ming, academician of Chinese Academy of Science organized a collaborative research group in the Institute of Virology, Chinese Academy of Medical Sciences to develop a novel gene expression system based on vaccinia virus Tiantan strain which was used to prepare smallpox vaccine for human use more than 50 years in China. After more than ten years of effort, a highly efficient vector system for foreign gene expression was successfully established and widely used for the expression of foreign genes, recombinant vaccines, and the development of monoclonal antibodies and diagnostic reagents. This paper briefly described the research and application of the gene expression system and discussed the problems and prospects of vector vaccine development.

  2. [From dreaded epidemic to rare disease - smallpox in Sweden 1750-1900].

    Science.gov (United States)

    Sköld, P

    1994-01-01

    Smallpox contributed to the death of 300,000 people in Sweden between 1750 and 1800. It was one of the most feared diseases of the time. Most victims were children under the age of 10 years. It is estimated that one out of every ten children died from smallpox. The mortality rate was between 10% and 20% but those who survived faced severe complications. Most common were the disfiguring pockmarks among those previously infected. It turned out that women who had contracted smallpox married much later in life than the healthy. Also, both female and male fertility was lowered by smallpox infection. By the mid 1750s inoculation was introduced in Sweden. This was a preventive method where a mild infection was given by putting smallpox matter into an incision in the skin. Success was restricted to the wealthy, even if the doctors tried to reach the common people. The main reasons were lack of confidence for doctors, medical and epidemiological risks, costs and an ineffective organization. In 1801 Eberhard Zacharias Munch of Rosenschöld performed the first vaccination with cowpox matter in Sweden. In a few years an extensive practise was stabilized, which made the Swedish population one of the best vaccinated in the world. Moreover, a unique registration was compiled which, together with the parish records, makes the country one of the best documented. The main reasons behind the success was previous experiences of inoculation, international influences, abolition of the physicians monopoly, an effective organization, better opportunities for financing and rewards, and the compulsory law of 1816.

  3. Vaccinia scars associated with better survival for adults. An observational study from Guinea-Bissau

    DEFF Research Database (Denmark)

    Aaby, Peter; Gustafson, Per; Roth, Adam Anders Edvin

    2006-01-01

    Live vaccines including BCG and measles may have non-targeted beneficial effects on childhood survival in areas with high mortality. The authors therefore undertook a survey of vaccinia scars to evaluate subsequent mortality.......Live vaccines including BCG and measles may have non-targeted beneficial effects on childhood survival in areas with high mortality. The authors therefore undertook a survey of vaccinia scars to evaluate subsequent mortality....

  4. Down Regulation of Gene Expression by the Vaccinia Virus D10 Protein

    OpenAIRE

    Shors, Teri; Keck, James G.; Moss, Bernard

    1999-01-01

    Vaccinia virus genes are expressed in a sequential fashion, suggesting a role for negative as well as positive regulatory mechanisms. A potential down regulator of gene expression was mapped by transfection assays to vaccinia virus open reading frame D10, which encodes a protein with no previously known function. Inhibition was independent of the promoter type used for the reporter gene, indicating that the mechanism did not involve promoter sequence recognition. The inhibition was overcome, ...

  5. Early death after feline infectious peritonitis virus challenge due to recombinant vaccinia virus immunization.

    OpenAIRE

    Vennema, H.; de Groot, R J; Harbour, D. A.; Dalderup, M.; Gruffydd-Jones, T.; Horzinek, M.C.; Spaan, W J

    1990-01-01

    The gene encoding the fusogenic spike protein of the coronavirus causing feline infectious peritonitis was recombined into the genome of vaccinia virus. The recombinant induced spike-protein-specific, in vitro neutralizing antibodies in mice. When kittens were immunized with the recombinant, low titers of neutralizing antibodies were obtained. After challenge with feline infectious peritonitis virus, these animals succumbed earlier than did the control group immunized with wild-type vaccinia ...

  6. Paramunity-inducing effects of vaccinia strain MVA.

    Science.gov (United States)

    Vilsmeier, B

    1999-09-01

    Vaccinia virus MVA is harmless for humans and animals both locally and parenterally. It offers paraspecific activities similar to those of comparable attenuated viruses of other pox genera, e.g. avipox or parapox. At the systemic level, MVA protects baby mice against lethal challenge with vesicular stomatitis virus (dose-response curve). MVA raises phagocytosis and NK-cell activity in humans and animals, whilst encouraging the induction of interferon alpha, interleukin-2 and -12 and colony-stimulating activity at the same time. The paramunity-inducing properties of MVA make it an ideal vector for the insertion of foreign genes. It is superior to other virus vectors because of its complex function. Inactivated MVA is also suitable as an inducer of paramunity.

  7. The Spanish royal philanthropic expedition to bring smallpox vaccination to the New World and Asia in the 19th century.

    Science.gov (United States)

    Franco-Paredes, Carlos; Lammoglia, Lorena; Santos-Preciado, José Ignacio

    2005-11-01

    The New World was ravaged by smallpox for several centuries after the Spanish conquest. Jenner's discovery of the smallpox vaccine made possible the prevention and control of smallpox epidemics. In response to a large outbreak of smallpox in the Spanish colonies, King Charles IV appointed Francisco Xavier de Balmis to lead an expedition that would introduce Jenner's vaccine to these colonies. During the journey, the vaccine was kept viable by passing it from arm to arm in orphaned children, who were brought along expressly for that purpose and remained under the care of the orphanage's director. This expedition was the first large scale mass vaccination of its kind. The historic legacy of this pioneering event in international health should be revisited in the current era of persistent inequalities in global health.

  8. Susceptibility of Marmosets (Callithrix jacchus) to Monkeypox Virus: A Low Dose Prospective Model for Monkeypox and Smallpox Disease.

    Science.gov (United States)

    Mucker, Eric M; Chapman, Jennifer; Huzella, Louis M; Huggins, John W; Shamblin, Joshua; Robinson, Camenzind G; Hensley, Lisa E

    2015-01-01

    Although current nonhuman primate models of monkeypox and smallpox diseases provide some insight into disease pathogenesis, they require a high titer inoculum, use an unnatural route of infection, and/or do not accurately represent the entire disease course. This is a concern when developing smallpox and/or monkeypox countermeasures or trying to understand host pathogen relationships. In our studies, we altered half of the test system by using a New World nonhuman primate host, the common marmoset. Based on dose finding studies, we found that marmosets are susceptible to monkeypox virus infection, produce a high viremia, and have pathological features consistent with smallpox and monkeypox in humans. The low dose (48 plaque forming units) required to elicit a uniformly lethal disease and the extended incubation (preclinical signs) are unique features among nonhuman primate models utilizing monkeypox virus. The uniform lethality, hemorrhagic rash, high viremia, decrease in platelets, pathology, and abbreviated acute phase are reflective of early-type hemorrhagic smallpox.

  9. Following in the footsteps of smallpox: can we achieve the global eradication of measles?

    Directory of Open Access Journals (Sweden)

    Morgan Oliver WC

    2004-03-01

    Full Text Available Abstract Background Although an effective measles vaccine has been available for almost 40 years, in 2000 there were about 30 million measles infections worldwide and 777,000 measles-related deaths. The history of smallpox suggests that achieving measles eradication depends on several factors; the biological characteristics of the organism; vaccine technology; surveillance and laboratory identification; effective delivery of vaccination programmes and international commitment to eradication. Discussion Like smallpox, measles virus has several biological characteristics that favour eradication. Humans are the only reservoir for the virus, which causes a visible illness and infection leading to life-long immunity. As the measles virus has only one genetic serotype which is relatively stable over time, the same basic vaccine can be used world-wide. Vaccination provides protection against measles infection for at least 15 years, although efficacy may be reduced due to host factors such as nutritional status. Measles vaccination may also confer other non-specific health benefits leading to reduced mortality. Accurate laboratory identification of measles cases enables enhanced surveillance to support elimination programmes. The "catch-up, keep-up, follow-up" vaccination programme implemented in the Americas has shown that measles elimination is possible using existing technologies. On 17th October 2003 the "Cape Town Measles Declaration" by the World Health Organisation and the United Nations Childrens Fund called on governments to intensify efforts to reduce measles mortality by supporting universal vaccination coverage and the development of more effective vaccination. Summary Although more difficult than for smallpox, recent experience in the Americas suggests that measles eradication is technically feasible. Growing international support to deliver these programmes means that measles, like smallpox, may very well become a curiosity of history.

  10. Risk of Inflammatory Bowel Disease following Bacille Calmette-Guérin and Smallpox Vaccination

    DEFF Research Database (Denmark)

    Villumsen, Anne Marie; Jess, Tine; Sørup, Signe

    2013-01-01

    Childhood immunology has been suggested to play a role in development of inflammatory bowel disease (IBD) based on the studies of childhood vaccinations, infections, and treatment with antibiotics. Bacille Calmette-Guérin (BCG) and smallpox vaccinations were gradually phased-out in Denmark...... for children born between 1965 and 1976, hence allowing the study of subsequent risk of Crohn's disease and ulcerative colitis in a unique prospective design....

  11. Extracting key information from historical data to quantify the transmission dynamics of smallpox

    Directory of Open Access Journals (Sweden)

    Brockmann Stefan O

    2008-08-01

    Full Text Available Abstract Background Quantification of the transmission dynamics of smallpox is crucial for optimizing intervention strategies in the event of a bioterrorist attack. This article reviews basic methods and findings in mathematical and statistical studies of smallpox which estimate key transmission parameters from historical data. Main findings First, critically important aspects in extracting key information from historical data are briefly summarized. We mention different sources of heterogeneity and potential pitfalls in utilizing historical records. Second, we discuss how smallpox spreads in the absence of interventions and how the optimal timing of quarantine and isolation measures can be determined. Case studies demonstrate the following. (1 The upper confidence limit of the 99th percentile of the incubation period is 22.2 days, suggesting that quarantine should last 23 days. (2 The highest frequency (61.8% of secondary transmissions occurs 3–5 days after onset of fever so that infected individuals should be isolated before the appearance of rash. (3 The U-shaped age-specific case fatality implies a vulnerability of infants and elderly among non-immune individuals. Estimates of the transmission potential are subsequently reviewed, followed by an assessment of vaccination effects and of the expected effectiveness of interventions. Conclusion Current debates on bio-terrorism preparedness indicate that public health decision making must account for the complex interplay and balance between vaccination strategies and other public health measures (e.g. case isolation and contact tracing taking into account the frequency of adverse events to vaccination. In this review, we summarize what has already been clarified and point out needs to analyze previous smallpox outbreaks systematically.

  12. The First Smallpox Epidemic on the Canadian Plains: In the Fur-traders' Words

    Directory of Open Access Journals (Sweden)

    C Stuart Houston

    2000-01-01

    Full Text Available William Tomison, in charge of the Hudson's Bay Company's Cumberland House on the Saskatchewan River, described the devastating smallpox epidemic of 1781 and 1782. He understood contagion, practised isolation and disinfection, and provided mortality statistics during a 'virgin soil' epidemic. Above all, he showed remarkable compassion. He and his men took dying Indians into their already crowded quarters, and provided them with food, shelter and 24 h care. This article describes the epidemic and its aftermath.

  13. [Treatment and remedies against smallpox outbreaks in Ferrara in the late nineteenth century].

    Science.gov (United States)

    Vicentini, Chiara Beatrice; Manfredini, Stefano; Altieri, Lorenzo; Lupi, Silvia; Guidi, Enrica; Contini, Carlo

    2013-09-01

    Health interventions against smallpox during the two epidemics in the second half of the 19th century are outlined. The 1871 hospital health report and the medical report on smallpox patients treated at the hospital and poorhouse of Ferrara between January 1891 and January 1892, drawn up by Alessandro Bennati, provide both interesting data and insights into the treatments and remedies of the time. The treatment of this illness was - and indeed could be - nothing other than symptomatic, there being no real means to halt the spread of the disease. Rather, other remedies were found by alleviating pain and regaining energy during the various stages of the disease. A close relationship between vaccination and the incidence and gravity of the illness is underlined. When the practice of vaccination started to be widely employed at the end of the century, there were almost no cases of death due to smallpox. The pharmacopoeias of the time, Antonio Campana's Farmacopea ferrarese in particular, proved an essential guide in the analysis of each document.

  14. SWOT analysis: strengths, weaknesses, opportunities and threats of the Israeli Smallpox Revaccination Program.

    Science.gov (United States)

    Huerta, Michael; Balicer, Ran D; Leventhal, Alex

    2003-01-01

    During September 2002, Israel began its current revaccination program against smallpox, targeting previously vaccinated "first responders" among medical and emergency workers. In order to identify the potential strengths and weaknesses of this program and the conditions under which critical decisions were reached, we conducted a SWOT analysis of the current Israeli revaccination program, designed to identify its intrinsic strengths and weaknesses, as well as opportunities for its success and threats against it. SWOT analysis--a practical tool for the study of public health policy decisions and the social and political contexts in which they are reached--revealed clear and substantial strengths and weaknesses of the current smallpox revaccination program, intrinsic to the vaccine itself. A number of threats were identified that may jeopardize the success of the current program, chief among them the appearance of severe complications of vaccination. Our finding of a lack of a generation of knowledge on smallpox vaccination highlights the need for improved physician education and dissipation of misconceptions that are prevalent in the public today.

  15. Using ICR and SCID mice as animal models for smallpox to assess antiviral drug efficacy.

    Science.gov (United States)

    Titova, Ksenya A; Sergeev, Alexander A; Zamedyanskaya, Alena S; Galahova, Darya O; Kabanov, Alexey S; Morozova, Anastasia A; Bulychev, Leonid E; Sergeev, Artemiy A; Glotova, Tanyana I; Shishkina, Larisa N; Taranov, Oleg S; Omigov, Vladimir V; Zavjalov, Evgenii L; Agafonov, Alexander P; Sergeev, Alexander N

    2015-09-01

    The possibility of using immunocompetent ICR mice and immunodeficient SCID mice as model animals for smallpox to assess antiviral drug efficacy was investigated. Clinical signs of the disease did not appear following intranasal (i.n.) challenge of mice with strain Ind-3a of variola virus (VARV), even when using the highest possible dose of the virus (5.2 log10 p.f.u.). The 50 % infective doses (ID50) of VARV, estimated by the virus presence or absence in the lungs 3 and 4 days post-infection, were 2.7 ± 0.4 log10 p.f.u. for ICR mice and 3.5 ± 0.7 log10 p.f.u. for SCID mice. After i.n. challenge of ICR and SCID mice with VARV 30 and 50 ID50, respectively, steady reproduction of the virus occurred only in the respiratory tract (lungs and nose). Pathological inflammatory destructive changes were revealed in the respiratory tract and the primary target cells for VARV (macrophages and epithelial cells) in mice, similar to those in humans and cynomolgus macaques. The use of mice to assess antiviral efficacies of NIOCH-14 and ST-246 demonstrated the compliance of results with those described in scientific literature, which opens up the prospect of their use as an animal model for smallpox to develop anti-smallpox drugs intended for humans.

  16. Non-coding RNAs and heme oxygenase-1 in vaccinia virus infection

    Energy Technology Data Exchange (ETDEWEB)

    Meseda, Clement A. [Division of Viral Products, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD (United States); Srinivasan, Kumar [Division of Transfusion Transmitted Diseases, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD (United States); Wise, Jasen [Qiagen, Frederick, MD (United States); Catalano, Jennifer [Center for Tobacco Products, Food and Drug Administration, Bethesda, MD (United States); Yamada, Kenneth M. [National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD (United States); Dhawan, Subhash, E-mail: subhash.dhawan@fda.hhs.gov [Division of Transfusion Transmitted Diseases, Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD (United States)

    2014-11-07

    Highlights: • Heme oxygenase-1 (HO-1) induction inhibited vaccinia virus infection of macrophages. • Reduced infectivity inversely correlated with increased expression of non-coding RNAs. • The regulation of HO-1 and ncRNAs suggests a novel host defense response against vaccinia virus infection. - Abstract: Small nuclear RNAs (snRNAs) are <200 nucleotide non-coding uridylate-rich RNAs. Although the functions of many snRNAs remain undetermined, a population of snRNAs is produced during the early phase of infection of cells by vaccinia virus. In the present study, we demonstrate a direct correlation between expression of the cytoprotective enzyme heme oxygenase-1 (HO-1), suppression of selective snRNA expression, and inhibition of vaccinia virus infection of macrophages. Hemin induced HO-1 expression, completely reversed virus-induced host snRNA expression, and suppressed vaccinia virus infection. This involvement of specific virus-induced snRNAs and associated gene clusters suggests a novel HO-1-dependent host-defense pathway in poxvirus infection.

  17. SUBCELLULAR LOCALIZATION OF ANTIGENS BY IMMUNE AND CYTOCHEMICAL TECHNIQUES IN ELECTRON MICROSCOPY. MITOCHONDRIA, A SITE OF VACCINIA PROTEIN.

    Science.gov (United States)

    in monocytic leukemic blood, and vaccinia antigen in infected HeLa cells . The results of this study suggest the following conclusions. (1) Platelets...synthesis in mitochondrial fractions of vaccinia-infected HeLa cells was found to be greater than in uninfected cells and increased as infection progressed

  18. 天花病名流变考%A Study on the Chinese Name of Smallpox

    Institute of Scientific and Technical Information of China (English)

    刘思媛

    2015-01-01

    现知最早记载天花的中医古籍《肘后备急方》中称天花为“虏疮”,后来又出现了“圣疮”“天疮”“豌豆疮”“麸豆疮”“痘疹”等多种称谓。历史上天花病名的繁杂更替,是中医关于天花的理论认识逐渐演变的显著标志。天花病名与中医相关诊断治疗、民间相关社会习俗均有联系。民国时期,“天花”最终替代“痘疹”成为这种病毒性传染病的专有名称,是牛痘接种术传入之后,中医在天花防治中的重要地位被预防接种所取代的结果。%The Zhou hou fang ( Handbook of Formulae) is the earliest known ancient medical book carrying the title of smallpox, called“lu chuan” ( captive sore) .Since then, various names had been ap-plied to this disorder, including “sheng chuang” (holy sore),“tian chuang” (celestial sore), “wan dou chuang” (pea sore),“fu dou chuang” (bran sore), and“dou zhen” (pox-rash).This frequent changes of disease titles in its history is a significant mark of the evolution of TCM recognition on smallpox related to its treatment and prevention, and folk-social customs.It was not until the Republican period that the title of“pox-rash” was substituted by smallpox, a specific title for this viral infection.This is also the outcome showing the important position of TCM treatment and prevention for smallpox replaced by the preventive method of modern smallpox vaccination.

  19. Redundancy complicates the definition of essential genes for vaccinia virus.

    Science.gov (United States)

    Dobson, Bianca M; Tscharke, David C

    2015-11-01

    Vaccinia virus (VACV) genes are characterized as either essential or non-essential for growth in culture. It seems intuitively obvious that if a gene can be deleted without imparting a growth defect in vitro it does not have a function related to basic replication or spread. However, this interpretation relies on the untested assumption that there is no redundancy across the genes that have roles in growth in cell culture. First, we provide a comprehensive summary of the literature that describes the essential genes of VACV. Next, we looked for interactions between large blocks of non-essential genes located at the ends of the genome by investigating sets of VACVs with large deletions at the genomic termini. Viruses with deletions at either end of the genome behaved as expected, exhibiting only mild or host-range defects. In contrast, combining deletions at both ends of the genome for the VACV Western Reserve (WR) strain caused a devastating growth defect on all cell lines tested. Unexpectedly, we found that the well-studied VACV growth factor homologue encoded by C11R has a role in growth in vitro that is exposed when 42 genes are absent from the left end of the VACV WR genome. These results demonstrate that some non-essential genes contribute to basic viral growth, but redundancy means these functions are not revealed by single-gene-deletion mutants.

  20. Vaccinia virus recombinants expressing an 11-kilodalton beta-galactosidase fusion protein incorporate active beta-galactosidase in virus particles.

    Science.gov (United States)

    Huang, C; Samsonoff, W A; Grzelecki, A

    1988-10-01

    Recombinant plasmids in which vaccinia virus transcriptional regulatory sequences were fused to the Escherichia coli lacZ gene were constructed for insertion of the lacZ gene into the vaccinia virus genome. beta-Galactosidase (beta-gal) was found in some purified recombinant vaccinia virions. By enzyme activity, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and microscopic techniques, the evidence suggested that beta-gal accounted for 5% of the total protein in the virion. These recombinant viruses were constructed so that a portion of the coding sequences of a late vaccinia virus structural polypeptide was fused to the amino terminus of beta-gal to produce the fusion protein. Removal of the coding sequences resulted in the complete loss of beta-gal activity. This demonstrated that a vaccinia virus DNA segment from a late structural gene is responsible for the incorporation of beta-gal into the virion.

  1. Effects of poliovirus 2A(pro) on vaccinia virus gene expression.

    Science.gov (United States)

    Feduchi, E; Aldabe, R; Novoa, I; Carrasco, L

    1995-12-15

    The effects of transient expression of poliovirus 2A(pro) on p220 cleavage in COS cells have been analyzed. When 2A(pro) was cloned in plasmid pTM1 and transiently expressed in COS cells, efficient cleavage of p220 occurred after infection of these cells with a recombinant vaccinia virus bearing phage T7 RNA polymerase. High numbers of COS cells were transfected with pTM1-2A, as judged by p220 cleavage, thereby allowing an analysis of the effects of poliovirus 2A(pro) on vaccinia virus gene expression. A 40-50% cleavage of p220 by transfected poliovirus 2A(pro) was observed ten hours post infection and cleavage was almost complete (80-90%) 20-25 hours post infection with vaccinia virus. Profound inhibition of vaccinia virus protein synthesis was detectable ten hours post infection and was maximal 20-25 hours post infection. This inhibition resulted from neither a blockade of transcription of vaccinia virus nor a lack of translatability of the mRNAs present in cells that synthesize poliovirus 2A(pro). Addition of ara-C inhibited the replication of vaccinia virus and allowed the continued synthesis of cellular proteins. Under these conditions, 2A(pro) is expressed and blocks cellular translation. Finally, p220 cleavage by 2A(pro) did not inhibit the translation of a mRNA encoding poliovirus protein 2C, as directed by the 5' leader sequences of encephalomiocarditis virus. Therefore, these findings show a correlation between p220 cleavage and inhibition of translation from newly made mRNAs. Our results are discussed in the light of present knowledge of p220 function, and new approaches are considered that might provide further insights into the function(s) of initiation factor eIF-4F.

  2. Vaccinia virus infection attenuates innate immune responses and antigen presentation by epidermal dendritic cells.

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

    2006-10-01

    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 this infection displays an atypical response to anti-poxvirus agents. Whereas adenosine N1-oxide blocked virus production and viral protein synthesis during a synchronous infection, cytosine arabinoside had no effect at concentrations sufficient to prevent virus replication in BSC40 monkey kidney cells. Vaccinia virus infection of XS52 cells not only failed to elicit the production of various cytokines, including tumor necrosis factor alpha (TNF-alpha), interleukin-1beta (IL-1beta), IL-6, IL-10, IL-12 p40, alpha interferon (IFN-alpha), and IFN-gamma, it actively inhibited the production of proinflammatory cytokines TNF-alpha and IL-6 by XS52 cells in response to exogenous lipopolysaccharide (LPS) or poly(I:C). Infection with a vaccinia virus mutant lacking the E3L gene resulted in TNF-alpha secretion in the absence of applied stimuli. Infection of XS52 cells or BSC40 cells with the DeltaE3L virus, but not wild-type vaccinia virus, triggered proteolytic decay of IkappaBalpha. These results suggest a novel role for the E3L protein as an antagonist of the NF-kappaB signaling pathway. DeltaE3L-infected XS52 cells secreted higher levels of TNF-alpha and IL-6 in response to LPS and poly(I:C) than did cells infected with the wild-type virus. XS52 cells were productively infected by a vaccinia virus mutant lacking the K1L gene. DeltaK1L-infected cells secreted higher levels of TNF-alpha and IL-6 in response to LPS than wild-type virus-infected cells. Vaccinia virus infection of primary LCs harvested from mouse epidermis was nonpermissive, although a viral reporter protein was

  3. Preparation of national standard for determination of vaccinia virus titer%痘苗病毒滴度测定用国家标准品的制备

    Institute of Scientific and Technical Information of China (English)

    黄维金; 周艳; 孟淑芳; 俞永新; 王佑春; 朱蓉; 赵晨燕; 徐静; 万延民; 齐影; 刘强; 王文波; 温智恒

    2012-01-01

    Objective To prepare the national standard for determination of titers of virus vector-based vaccines. Methods Vaccinia virus was prepared with chick embryo fibroblasts (CEFs), purified by density gradient ultracentrifuge, added with preservative for smallpox vaccine, then filled and stored in frozen, and distributed to four laboratories for cooperative calibration by plaque assay with crystal violet staining. The samples were diluted serially, and two dilutions were used for each test. Five parallel tests were performed on each dilution. The geometric mean of determination results in various laboratories was calculated, based on which the vaccinia virus titer in standard was defined. The virus standard after storage at various temperatures for various time durations were evaluated for stability, and subjected to overall control tests. Results A total of 44 tests were performed in four laboratories, and the results, with a geometric mean of 3. 05 × 106 PFU/ml, a 95% CI of 2. 00 × 106 ~ 4. 65 × 106 PFU/ml, a standard deviation of 0. 09 and a total CV value of 1. 41%, showed no significant difference (P > 0. 05). The prepared vaccinia virus standard was stable after storage at 4℃ for 10 d, while was less stable at 37℃, and was qualified in filling quantity (1 ml per container) and sterility test, of which the pH value was 6. 9 and the precision for filling was 0. 95% based on weight. Conclusion The prepared vaccinia virus standard may be used for determination of titers of vaccinia virus vector-based vaccines.%目的 制备病毒载体疫苗滴度测定用国家标准品.方法 以鸡胚成纤维细胞制备痘苗病毒,经密度梯度超速离心纯化后,加入天花疫苗保护剂,分装冻存,分发至我国4个不同实验室,按照统一的结晶紫染色蚀斑法实验方案进行协作标定,每次实验稀释后取2个稀释度,每个稀释度设5个平行孔,计算各实验室检测结果的几何均数,对标准品中痘苗病毒滴度进行赋值.

  4. Myxoma and vaccinia viruses bind differentially to human leukocytes.

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    Chan, Winnie M; Bartee, Eric C; Moreb, Jan S; Dower, Ken; Connor, John H; McFadden, Grant

    2013-04-01

    Myxoma virus (MYXV) and vaccinia virus (VACV), two distinct members of the family Poxviridae, are both currently being developed as oncolytic virotherapeutic agents. Recent studies have demonstrated that ex vivo treatment with MYXV can selectively recognize and kill contaminating cancerous cells from autologous bone marrow transplants without perturbing the engraftment of normal CD34(+) hematopoietic stem and progenitor cells. However, the mechanism(s) by which MYXV specifically recognizes and eliminates the cancer cells in the autografts is not understood. While little is known about the cellular attachment factor(s) exploited by MYXV for entry into any target cells, VACV has been shown to utilize cell surface glycosaminoglycans such as heparan sulfate (HS), the extracellular matrix protein laminin, and/or integrin β1. We have constructed MYXV and VACV virions tagged with the Venus fluorescent protein and compared their characteristics of binding to various human cancer cell lines as well as to primary human leukocytes. We report that the binding of MYXV or VACV to some adherent cell lines could be partially inhibited by heparin, but laminin blocked only VACV binding. In contrast to cultured fibroblasts, the binding of MYXV and VACV to a wide spectrum of primary human leukocytes could not be competed by either HS or laminin. Additionally, MYXV and VACV exhibited very different binding characteristics against certain select human leukocytes, suggesting that the two poxviruses utilize different cell surface determinants for the attachment to these cells. These results indicate that VACV and MYXV can exhibit very different oncolytic tropisms against some cancerous human leukocytes.

  5. Live-Cell Imaging of Vaccinia Virus Recombination.

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    Patrick Paszkowski

    2016-08-01

    Full Text Available Recombination between co-infecting poxviruses provides an important mechanism for generating the genetic diversity that underpins evolution. However, poxviruses replicate in membrane-bound cytoplasmic structures known as factories or virosomes. These are enclosed structures that could impede DNA mixing between co-infecting viruses, and mixing would seem to be essential for this process. We hypothesize that virosome fusion events would be a prerequisite for recombination between co-infecting poxviruses, and this requirement could delay or limit viral recombination. We have engineered vaccinia virus (VACV to express overlapping portions of mCherry fluorescent protein fused to a cro DNA-binding element. In cells also expressing an EGFP-cro fusion protein, this permits live tracking of virus DNA and genetic recombination using confocal microscopy. Our studies show that different types of recombination events exhibit different timing patterns, depending upon the relative locations of the recombining elements. Recombination between partly duplicated sequences is detected soon after post-replicative genes are expressed, as long as the reporter gene sequences are located in cis within an infecting genome. The same kinetics are also observed when the recombining elements are divided between VACV and transfected DNA. In contrast, recombination is delayed when the recombining sequences are located on different co-infecting viruses, and mature recombinants aren't detected until well after late gene expression is well established. The delay supports the hypothesis that factories impede inter-viral recombination, but even after factories merge there remain further constraints limiting virus DNA mixing and recombinant gene assembly. This delay could be related to the continued presence of ER-derived membranes within the fused virosomes, membranes that may once have wrapped individual factories.

  6. Oncolytic vaccinia virus synergizes with irinotecan in colorectal cancer.

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    Ottolino-Perry, Kathryn; Acuna, Sergio A; Angarita, Fernando A; Sellers, Clara; Zerhouni, Siham; Tang, Nan; McCart, J Andrea

    2015-10-01

    Metastatic colorectal cancer (CRC) is complex clinical challenge for which there are limited treatment options. Chemotherapy with or without surgery provides moderate improvements in overall survival and quality of life; nevertheless the 5-year survival remains below 30%. Oncolytic vaccinia virus (VV) shows strong anti-tumour activity in models of CRC, however transient delays in disease progression are insufficient to lead to long-term survival. Here we examined the efficacy of VV with oxaliplatin or SN-38 (active metabolite of irinotecan) in CRC cell lines in vitro and VV with irinotecan in an orthotopic model of metastatic CRC. Synergistic improvements in in vitro cell killing were observed in multiple cell lines. Combination therapy was well tolerated in tumour-bearing mice and the median survival was significantly increased relative to monotherapy despite a drug-dependent decrease in the mean tumour titer. Increased apoptosis following in vitro and in vivo combination therapy was observed. In vitro cell cycle analysis showed increases in S-phase cells following infection occurred in both infected and uninfected cell populations. This corresponded to a 4-fold greater increase in apoptosis in the uninfected compared to infected cells following combination therapy. Combination treatment strategies are among the best options for patients with advanced cancers. VV is currently under clinical investigation in patients with CRC and the data presented here suggest that its combination with irinotecan may provide benefit to a subset of CRC patients. Further, investigation of this combination is necessary to determine the tumour characteristics responsible for mediating synergy. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  7. Transmission patterns of smallpox: systematic review of natural outbreaks in Europe and North America since World War II

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    Boer Rob

    2006-05-01

    Full Text Available Abstract Background Because smallpox (variola major may be used as a biological weapon, we reviewed outbreaks in post-World War II Europe and North America in order to understand smallpox transmission patterns. Methods A systematic review was used to identify papers from the National Library of Medicine, Embase, Biosis, Cochrane Library, Defense Technical Information Center, WorldCat, and reference lists of included publications. Two authors reviewed selected papers for smallpox outbreaks. Results 51 relevant outbreaks were identified from 1,389 publications. The median for the effective first generation reproduction rate (initial R was 2 (range 0–38. The majority outbreaks were small (less than 5 cases and contained within one generation. Outbreaks with few hospitalized patients had low initial R values (median of 1 and were prolonged if not initially recognized (median of 3 generations; outbreaks with mostly hospitalized patients had higher initial R values (median 12 and were shorter (median of 3 generations. Index cases with an atypical presentation of smallpox were less likely to have been diagnosed with smallpox; outbreaks in which the index case was not correctly diagnosed were larger (median of 27.5 cases and longer (median of 3 generations compared to outbreaks in which the index case was correctly diagnosed (median of 3 cases and 1 generation. Conclusion Patterns of spread during Smallpox outbreaks varied with circumstances, but early detection and implementation of control measures is a most important influence on the magnitude of outbreaks. The majority of outbreaks studied in Europe and North America were controlled within a few generations if detected early.

  8. Vaccination policy against smallpox, 1835-1914: a comparison of England with Prussia and Imperial Germany.

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    Hennock, E P

    1998-04-01

    There are three identifiable phases in comparing vaccination policy in England, Prussia and Imperial Germany. (1) Prior to the 1870's the tradition of medical police in Prussia resulted in the vaccination of the population being treated as a State responsibility earlier than in England and provided an appropriate administrative framework. The administrative pressure that could be exerted persuaded the Prussian authorities that legislation to make vaccination compulsory was unnecessary. In contrast, England and Wales lacked both the tradition and administrative structures of a medical police. Legislation (1840, 1853) for free and universal infant vaccination was followed by radical ideological and administrative innovation. (2) From 1875 to 1889 both countries provided free and compulsory vaccination for all. In England this was limited to infants; in Germany including Prussia, it included the re-vaccination of children. (3) After 1889 England and Germany began to diverge more sharply. In England vaccination rates fell and after 1898 conscientious objectors were excused from having to have their children vaccinated. Germany retained compulsory vaccination and rates in the two countries increasingly diverged. England came to rely on the local public health administration for the surveillance and containment of smallpox, including selective vaccination of contacts. Despite these differences smallpox mortality dropped sharply in both countries, although in Germany somewhat earlier. The English reliance on surveillance and containment prefigures that of the WHO in the eradication of smallpox in the Third World. It suggests that the emphasis on the importance of high levels of mass vaccination in the German literature should perhaps be revised.

  9. Population-level differences in disease transmission: a Bayesian analysis of multiple smallpox epidemics.

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    Elderd, Bret D; Dwyer, Greg; Dukic, Vanja

    2013-09-01

    Estimates of a disease's basic reproductive rate R0 play a central role in understanding outbreaks and planning intervention strategies. In many calculations of R0, a simplifying assumption is that different host populations have effectively identical transmission rates. This assumption can lead to an underestimate of the overall uncertainty associated with R0, which, due to the non-linearity of epidemic processes, may result in a mis-estimate of epidemic intensity and miscalculated expenditures associated with public-health interventions. In this paper, we utilize a Bayesian method for quantifying the overall uncertainty arising from differences in population-specific basic reproductive rates. Using this method, we fit spatial and non-spatial susceptible-exposed-infected-recovered (SEIR) models to a series of 13 smallpox outbreaks. Five outbreaks occurred in populations that had been previously exposed to smallpox, while the remaining eight occurred in Native-American populations that were naïve to the disease at the time. The Native-American outbreaks were close in a spatial and temporal sense. Using Bayesian Information Criterion (BIC), we show that the best model includes population-specific R0 values. These differences in R0 values may, in part, be due to differences in genetic background, social structure, or food and water availability. As a result of these inter-population differences, the overall uncertainty associated with the "population average" value of smallpox R0 is larger, a finding that can have important consequences for controlling epidemics. In general, Bayesian hierarchical models are able to properly account for the uncertainty associated with multiple epidemics, provide a clearer understanding of variability in epidemic dynamics, and yield a better assessment of the range of potential risks and consequences that decision makers face.

  10. Genome sequence diversity and clues to the evolution of variola (smallpox) virus.

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    Esposito, Joseph J; Sammons, Scott A; Frace, A Michael; Osborne, John D; Olsen-Rasmussen, Melissa; Zhang, Ming; Govil, Dhwani; Damon, Inger K; Kline, Richard; Laker, Miriam; Li, Yu; Smith, Geoffrey L; Meyer, Hermann; Leduc, James W; Wohlhueter, Robert M

    2006-08-11

    Comparative genomics of 45 epidemiologically varied variola virus isolates from the past 30 years of the smallpox era indicate low sequence diversity, suggesting that there is probably little difference in the isolates' functional gene content. Phylogenetic clustering inferred three clades coincident with their geographical origin and case-fatality rate; the latter implicated putative proteins that mediate viral virulence differences. Analysis of the viral linear DNA genome suggests that its evolution involved direct descent and DNA end-region recombination events. Knowing the sequences will help understand the viral proteome and improve diagnostic test precision, therapeutics, and systems for their assessment.

  11. Antigen profiling analysis of vaccinia virus injected canine tumors: oncolytic virus efficiency predicted by boolean models.

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    Cecil, Alexander; Gentschev, Ivaylo; Adelfinger, Marion; Nolte, Ingo; Dandekar, Thomas; Szalay, Aladar A

    2014-01-01

    Virotherapy on the basis of oncolytic vaccinia virus (VACV) strains is a novel approach for cancer therapy. In this study we describe for the first time the use of dynamic boolean modeling for tumor growth prediction of vaccinia virus GLV-1h68-injected canine tumors including canine mammary adenoma (ZMTH3), canine mammary carcinoma (MTH52c), canine prostate carcinoma (CT1258), and canine soft tissue sarcoma (STSA-1). Additionally, the STSA-1 xenografted mice were injected with either LIVP 1.1.1 or LIVP 5.1.1 vaccinia virus strains.   Antigen profiling data of the four different vaccinia virus-injected canine tumors were obtained, analyzed and used to calculate differences in the tumor growth signaling network by type and tumor type. Our model combines networks for apoptosis, MAPK, p53, WNT, Hedgehog, TK cell, Interferon, and Interleukin signaling networks. The in silico findings conform with in vivo findings of tumor growth. Boolean modeling describes tumor growth and remission semi-quantitatively with a good fit to the data obtained for all cancer type variants. At the same time it monitors all signaling activities as a basis for treatment planning according to antigen levels. Mitigation and elimination of VACV- susceptible tumor types as well as effects on the non-susceptible type CT1258 are predicted correctly. Thus the combination of Antigen profiling and semi-quantitative modeling optimizes the therapy already before its start.

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

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    Stöcklein Walter

    2007-08-01

    Full Text Available Abstract 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.

  13. Production and characterisation of a monoclonal antibody to human papillomavirus type 16 using recombinant vaccinia virus.

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    McLean, C S; Churcher, M J; Meinke, J; Smith, G L; Higgins, G; Stanley, M; Minson, A C

    1990-06-01

    A monoclonal antibody was raised against the major capsid protein L1 of human papillomavirus type 16, using a recombinant vaccinia virus that expresses the L1 protein, as a target for screening. This antibody, designated CAMVIR-1, reacted with a 56 kilodalton protein in cells infected with L1-vaccinia virus, and the protein was present in a predominantly nuclear location. The antibody also detects the HPV-16 L1 antigen in formalin fixed, paraffin wax embedded biopsy specimens and on routine cervical smears. The antibody reacts strongly and consistently with biopsy specimens containing HPV-16 or HPV-33, but very weak reactions were occasionally observed with biopsy specimens or smears containing HPV-6 or HPV-11. The potential advantages of using a vaccinia recombinant are (i) the target protein is synthesised in a eukoryotic cell so that its "processing" and location are normal; (ii) cells infected with vaccinia recombinants can be subjected to various fixing procedures similar to those used for routine clinical material. This greatly increases the probability that an identified antibody will be useful in a clinical setting.

  14. Proteome analysis of vaccinia virus IHD-W-infected HEK 293 cells with 2-dimensional gel electrophoresis and MALDI-PSD-TOF MS of on solid phase support N-terminally sulfonated peptides

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    Bartel Sebastian

    2011-08-01

    Full Text Available Abstract Background Despite the successful eradication of smallpox by the WHO-led vaccination programme, pox virus infections remain a considerable health threat. The possible use of smallpox as a bioterrorism agent as well as the continuous occurrence of zoonotic pox virus infections document the relevance to deepen the understanding for virus host interactions. Since the permissiveness of pox infections is independent of hosts surface receptors, but correlates with the ability of the virus to infiltrate the antiviral host response, it directly depends on the hosts proteome set. In this report the proteome of HEK293 cells infected with Vaccinia Virus strain IHD-W was analyzed by 2-dimensional gel electrophoresis and MALDI-PSD-TOF MS in a bottom-up approach. Results The cellular and viral proteomes of VACV IHD-W infected HEK293 cells, UV-inactivated VACV IHD-W-treated as well as non-infected cells were compared. Derivatization of peptides with 4-sulfophenyl isothiocyanate (SPITC carried out on ZipTipμ-C18 columns enabled protein identification via the peptides' primary sequence, providing improved s/n ratios as well as signal intensities of the PSD spectra. The expression of more than 24 human proteins was modulated by the viral infection. Effects of UV-inactivated and infectious viruses on the hosts' proteome concerning energy metabolism and proteins associated with gene expression and protein-biosynthesis were quite similar. These effects might therefore be attributed to virus entry and virion proteins. However, the modulation of proteins involved in apoptosis was clearly correlated to infectious viruses. Conclusions The proteome analysis of infected cells provides insight into apoptosis modulation, regulation of cellular gene expression and the regulation of energy metabolism. The confidence of protein identifications was clearly improved by the peptides' derivatization with SPITC on a solid phase support. Some of the identified proteins

  15. The complete genomic sequence of the modified vaccinia Ankara strain: comparison with other orthopoxviruses.

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    Antoine, G; Scheiflinger, F; Dorner, F; Falkner, F G

    1998-05-10

    The complete genomic DNA sequence of the highly attenuated vaccinia strain modified vaccinia Ankara (MVA) was determined. The genome of MVA is 178 kb in length, significantly smaller than that of the vaccinia Copenhagen genome, which is 192 kb. The 193 open reading frames (ORFs) mapped in the MVA genome probably correspond to 177 genes, 25 of which are split and/or have suffered mutations resulting in truncated proteins. The left terminal genomic region of MVA contains four large deletions and one large insertion relative to the Copenhagen strain. In addition, many ORFs in this region are fragmented, leaving only eight genes structurally intact and therefore presumably functional. The inserted DNA codes for a cluster of genes that is also found in the vaccinia WR strain and in cowpox virus and includes a highly fragmented gene homologous to the cowpox virus host range gene, providing further evidence that a cowpox-like virus was the ancestor of vaccinia. Surprisingly, the central conserved region of the genome also contains some fragmented genes, including ORF F5L, encoding a major membrane protein, and ORFs F11L and O1L, encoding proteins of 39.7 and 77.6 kDa, respectively. The right terminal genomic region carries three large deletions all classical poxviral immune evasion genes and all ankyrin-like genes located in this region are fragmented except for those encoding the interleukin-1 beta receptor and the 68-kDa ankyrin-like protein B18R. Thus, the attenuated phenotype of MVA is the result of numerous mutations, particularly affecting the host interactive proteins, including the ankyrin-like genes, but also involving some structural proteins.

  16. Reflections on New York City's 1947 Smallpox Vaccination Program and Its 1976 Swine Influenza Immunization Program.

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    Imperato, Pascal James

    2015-06-01

    In 1947, a smallpox outbreak occurred in New York City with a total of twelve cases and two deaths. In order to contain this outbreak, the New York City Department of Health launched a mass immunization campaign that over a period of some 60 days vaccinated 6.35 million people. This article examines in detail the epidemiology of this outbreak and the measures employed to contain it. In 1976, a swine influenza strain was isolated among a few recruits at a US Army training camp at Fort Dix, New Jersey. It was concluded at the time that this virus possibly represented a re-appearance of the 1918 influenza pandemic influenza strain. As a result, a mass national immunization program was launched by the federal government. From its inception, the program encountered a myriad of challenges ranging from doubts that it was even necessary to the development of Guillain-Barré paralysis among some vaccine recipients. This paper examines the planning for and implementation of the swine flu immunization program in New York City. It also compares it to the smallpox vaccination program of 1947. Despite equivalent financial and personnel resources, leadership and organizational skills, the 1976 program only immunized approximately a tenth of the number of New York City residents vaccinated in 1947. The reasons for these marked differences in outcomes are discussed in detail.

  17. US military service members vaccinated against smallpox in 2003 and 2004 experience a slightly higher risk of hospitalization postvaccination.

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    Jacobson, Isabel G; Smith, Tyler C; Smith, Besa; Wells, Timothy S; Reed, Robert J; Ryan, Margaret A K

    2008-07-29

    This study explores adverse events severe enough to warrant hospitalization that may have been associated with receiving the smallpox vaccine in conjunction with military service. Cox proportional hazards modeling was used to identify the risk of hospitalization among US active-duty military personnel during a 1-year period following receipt of the smallpox vaccine. The reference group consisted of active-duty military personnel who also received the smallpox vaccine after the conclusion of their health care observation period, allowing for comparison to a temporally and demographically similar population. The risk of hospitalization was slightly elevated among the postvaccine group for any-cause hospitalization and for hospitalization in several broad diagnostic categories. Hospitalizations for asthma, autoimmune diseases, and myopericarditis, were more likely in the postvaccine group. The increased risk of hospitalization for varied outcomes does not necessarily imply a cause-effect relationship, but it does offer areas for more focused study, using longitudinal data to explore the long-term impact of smallpox vaccination on the health of young adults.

  18. Susceptibility of Marmosets (Callithrix jacchus to Monkeypox Virus: A Low Dose Prospective Model for Monkeypox and Smallpox Disease.

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    Eric M Mucker

    Full Text Available Although current nonhuman primate models of monkeypox and smallpox diseases provide some insight into disease pathogenesis, they require a high titer inoculum, use an unnatural route of infection, and/or do not accurately represent the entire disease course. This is a concern when developing smallpox and/or monkeypox countermeasures or trying to understand host pathogen relationships. In our studies, we altered half of the test system by using a New World nonhuman primate host, the common marmoset. Based on dose finding studies, we found that marmosets are susceptible to monkeypox virus infection, produce a high viremia, and have pathological features consistent with smallpox and monkeypox in humans. The low dose (48 plaque forming units required to elicit a uniformly lethal disease and the extended incubation (preclinical signs are unique features among nonhuman primate models utilizing monkeypox virus. The uniform lethality, hemorrhagic rash, high viremia, decrease in platelets, pathology, and abbreviated acute phase are reflective of early-type hemorrhagic smallpox.

  19. The prevention and eradication of smallpox: a commentary on Sloane (1755) ‘An account of inoculation’

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    Weiss, Robin A.; Esparza, José

    2015-01-01

    Sir Hans Sloane's account of inoculation as a means to protect against smallpox followed several earlier articles published in Philosophical Transactions on this procedure. Inoculation (also called ‘variolation’) involved the introduction of small amounts of infectious material from smallpox vesicles into the skin of healthy subjects, with the goal of inducing mild symptoms that would result in protection against the more severe naturally acquired disease. It began to be practised in England in 1721 thanks to the efforts of Lady Mary Wortley Montagu who influenced Sloane to promote its use, including the inoculation of the royal family's children. When Edward Jenner's inoculation with the cow pox (‘vaccination’) followed 75 years later as a safer yet equally effective procedure, the scene was set for the eventual control of smallpox epidemics culminating in the worldwide eradication of smallpox in 1977, officially proclaimed by WHO in 1980. Here, we discuss the significance of variolation and vaccination with respect to scientific, public health and ethical controversies concerning these ‘weapons of mass protection’. This commentary was written to celebrate the 350th anniversary of the journal Philosophical Transactions of the Royal Society. PMID:25750241

  20. Characterization of a new Vaccinia virus isolate reveals the C23L gene as a putative genetic marker for autochthonous Group 1 Brazilian Vaccinia virus.

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    Felipe L Assis

    Full Text Available 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.

  1. Smallpox virus resequencing GeneChips can also rapidly ascertain species status for some zoonotic non-variola orthopoxviruses.

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    Sulaiman, Irshad M; Sammons, Scott A; Wohlhueter, Robert M

    2008-04-01

    We recently developed a set of seven resequencing GeneChips for the rapid sequencing of Variola virus strains in the WHO Repository of the Centers for Disease Control and Prevention. In this study, we attempted to hybridize these GeneChips with some known non-Variola orthopoxvirus isolates, including monkeypox, cowpox, and vaccinia viruses, for rapid detection.

  2. Design of inhibitors of thymidylate kinase from Variola virus as new selective drugs against smallpox.

    Science.gov (United States)

    Guimarães, Ana P; de Souza, Felipe R; Oliveira, Aline A; Gonçalves, Arlan S; de Alencastro, Ricardo B; Ramalho, Teodorico C; França, Tanos C C

    2015-02-16

    Recently we constructed a homology model of the enzyme thymidylate kinase from Variola virus (VarTMPK) and proposed it as a new target to the drug design against smallpox. In the present work, we used the antivirals cidofovir and acyclovir as reference compounds to choose eleven compounds as leads to the drug design of inhibitors for VarTMPK. Docking and molecular dynamics (MD) studies of the interactions of these compounds inside VarTMPK and human TMPK (HssTMPK) suggest that they compete for the binding region of the substrate and were used to propose the structures of ten new inhibitors for VarTMPK. Further docking and MD simulations of these compounds, inside VarTMPK and HssTMPK, suggest that nine among ten are potential selective inhibitors of VarTMPK. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  3. p53 Downregulates Its Activating Vaccinia-Related Kinase 1, Forming a New Autoregulatory Loop

    OpenAIRE

    2006-01-01

    The stable accumulation of p53 is detrimental to the cell because it blocks cell growth and division. Therefore, increases in p53 levels are tightly regulated, mainly by its transcriptional target, mdm2, that downregulates p53. Elucidation of new signaling pathways requires the characterization of the members and the nature of their connection. Vaccinia-related kinase 1 (VRK1) contributes to p53 stabilization by partly interfering with its mdm2-mediated degradation, among other mechanisms; th...

  4. Modulation of the Myxoma Virus Plaque Phenotype by Vaccinia Virus Protein F11

    OpenAIRE

    Irwin, Chad R.; Evans, David H.

    2012-01-01

    Vaccinia virus (VACV) produces large plaques consisting of a rapidly expanding ring of infected cells surrounding a lytic core, whereas myxoma virus (MYXV) produces small plaques that resemble a focus of transformed cells. This is odd, because bioinformatics suggests that MYXV carries homologs of nearly all of the genes regulating Orthopoxvirus attachment, entry, and exit. So why does MYXV produce foci? One notable difference is that MYXV-infected cells produce few of the actin microfilaments...

  5. The Florey lecture, 1983. Biological control, as exemplified by smallpox eradication and myxomatosis.

    Science.gov (United States)

    Fenner, F

    1983-06-22

    Biological control is an important method of dealing with plant and insect pests. The control of rabbits by myxomatosis and the eradication of smallpox by vaccination are unusual examples of biological control, in that they involve a vertebrate and a viral pest respectively. Myxomatosis is a benign disease in Sylvilagus rabbits in South America which is transmitted mechanically by mosquitoes. In the European rabbit, Oryctolagus, which is a pest in Australia and England, the virus from Sylvilagus produces a generalized disease that is almost always lethal. Myxomatosis was deliberately introduced into Australia in 1950 and into Europe in 1952. It was at first spectacularly successful in controlling the rabbit pest, but biological adjustments occurred in the virulence of the virus and the genetic resistances of rabbits. After 30 years of interaction, natural selection has resulted in a balance at a fairly high level of viral virulence. Smallpox has been a major scourge of mankind for over 1500 years. It spread from Asia to Europe in the Middle ages and from Europe to Africa and the Americas in the 15th and 16th centuries. Jenner's cowpox vaccine provided a method of control that reduced the severity of the disease during the 19th century but failed to eliminate the disease from many countries before the 1930s. Thereafter it was eradicated from Europe and North America, but remained endemic in South America, Africa and Asia. In 1967 it was still endemic in 33 countries and W.H.O. established a programme for global eradication within 10 years. The goal was achieved in 1977. Problems of the eradication programme and reasons for its success will be described.

  6. The vaccinia virus E6 protein influences virion protein localization during virus assembly

    Energy Technology Data Exchange (ETDEWEB)

    Condit, Richard C., E-mail: condit@mgm.ufl.edu; Moussatche, Nissin

    2015-08-15

    Vaccinia virus mutants in which expression of the virion core protein gene E6R is repressed are defective in virion morphogenesis. E6 deficient infections fail to properly package viroplasm into viral membranes, resulting in an accumulation of empty immature virions and large aggregates of viroplasm. We have used immunogold electron microscopy and immunofluorescence confocal microscopy to assess the intracellular localization of several virion structural proteins and enzymes during E6R mutant infections. We find that during E6R mutant infections virion membrane proteins and virion transcription enzymes maintain a normal localization within viral factories while several major core and lateral body proteins accumulate in aggregated virosomes. The results support a model in which vaccinia virions are assembled from at least three substructures, the membrane, the viroplasm and a “pre-nucleocapsid”, and that the E6 protein is essential for maintaining proper localization of the seven-protein complex and the viroplasm during assembly. - Highlights: • Mutation of E6 disrupts association of viral membranes with viral core proteins • Mutation of E6 does not perturb viral membrane biosynthesis • Mutation of E6 does not perturb localization of viral transcription enzymes • Mutation of E6 causes mis-localization and aggregation of viral core proteins • Vaccinia assembly uses three subassemblies: membranes, viroplasm, prenucleocapsid.

  7. Initial characterization of Vaccinia Virus B4 suggests a role in virus spread

    Energy Technology Data Exchange (ETDEWEB)

    Burles, Kristin; Irwin, Chad R.; Burton, Robyn-Lee [Li Ka Shing Institute of Virology, Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada T6G 2S2 (Canada); Schriewer, Jill [Department of Molecular Microbiology and Immunology, Saint Louis University School of Medicine, St. Louis, MO (United States); Evans, David H. [Li Ka Shing Institute of Virology, Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada T6G 2S2 (Canada); Buller, R. Mark [Department of Molecular Microbiology and Immunology, Saint Louis University School of Medicine, St. Louis, MO (United States); Barry, Michele, E-mail: michele.barry@ualberta.ca [Li Ka Shing Institute of Virology, Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada T6G 2S2 (Canada)

    2014-05-15

    Currently, little is known about the ankyrin/F-box protein B4. Here, we report that B4R-null viruses exhibited reduced plaque size in tissue culture, and decreased ability to spread, as assessed by multiple-step growth analysis. Electron microscopy indicated that B4R-null viruses still formed mature and extracellular virions; however, there was a slight decrease of virions released into the media following deletion of B4R. Deletion of B4R did not affect the ability of the virus to rearrange actin; however, VACV811, a large vaccinia virus deletion mutant missing 55 open reading frames, had decreased ability to produce actin tails. Using ectromelia virus, a natural mouse pathogen, we demonstrated that virus devoid of EVM154, the B4R homolog, showed decreased spread to organs and was attenuated during infection. This initial characterization suggests that B4 may play a role in virus spread, and that other unidentified mediators of actin tail formation may exist in vaccinia virus. - Highlights: • B4R-null viruses show reduced plaque size, and decreased ability to spread. • B4R-null viruses formed mature and extracellular virions; and rearranged actin. • Virus devoid of EVM154, the B4R homolog, was attenuated during infection. • Initial characterization suggests that B4 may play a role in virus spread. • Unidentified mediators of actin tail formation may exist in vaccinia virus.

  8. Viral glycoprotein-mediated cell fusion assays using vaccinia virus vectors.

    Science.gov (United States)

    Bossart, Katharine N; Broder, Christopher C

    2004-01-01

    The vaccinia virus-based expression of viral envelope glycoprotein genes-derived from enveloped viruses that infect their respective host cells through a pH-independent mechanism of membrane fusion-has been a powerful tool in helping to characterize these important attachment and fusion proteins. The cellular expression of these viral envelope glycoproteins has allowed for the measurement of membrane fusion events using cell-cell fusion or syncytia formation. This method has been enhanced by the addition of a reporter-gene system to the vaccinia virus-based cell-cell fusion assay. This improvement has provided a high-throughput and quantitative aspect to this assay, which can serve as a surrogate for virus entry and is therefore ideally suited in the characterization of numerous enveloped viruses, including biological safety level-4 (BSL-4) agents. This chapter will detail the methods of the vaccinia virus-based reporter-gene fusion assay and how it may be used to characterize the fusion mediated by the BSL-4-classified Hendra and Nipah viruses.

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

    Directory of Open Access Journals (Sweden)

    Bedognetti Davide

    2011-10-01

    Full Text Available Abstract 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.

  10. Escherichia coli gpt gene provides dominant selection for vaccinia virus open reading frame expression vectors.

    Science.gov (United States)

    Falkner, F G; Moss, B

    1988-06-01

    Mycophenolic acid, an inhibitor of purine metabolism, was shown to block the replication of vaccinia virus in normal cell lines. This observation led to the development of a dominant one-step plaque selection system, based on expression of the Escherichia coli gpt gene, for the isolation of recombinant vaccinia viruses. Synthesis of xanthine-guanine phosphoribosyltransferase enabled only the recombinant viruses to form large plaques in a selective medium containing mycophenolic acid, xanthine, and hypoxanthine. To utilize the selection system efficiently, we constructed a series of plasmids that contain the E. coli gpt gene and allow insertion of foreign genes into multiple unique restriction endonuclease sites in all three reading frames between the translation initiation codon of a strong late promoter and synthetic translation termination sequences. The selection-expression cassette is flanked by vaccinia virus DNA that directs homologous recombination into the virus genome. The new vectors allow high-level expression of complete or partial open reading frames and rapid construction of recombinant viruses by facilitating the cloning steps and by simplifying their isolation. The system was tested by cloning the E. coli beta-galactosidase gene; in 24 h, this enzyme accounted for approximately 3.5% of the total infected-cell protein.

  11. Determination of the promoter region of an early vaccinia virus gene encoding thymidine kinase.

    Science.gov (United States)

    Weir, J P; Moss, B

    1987-05-01

    Nine recombinant vaccinia viruses that contain overlapping segments of the putative promoter region of the vaccinia virus thymidine kinase (TK) gene linked to DNA coding for the prokaryotic enzyme chloramphenicol acetyltransferase (CAT) were constructed. In each case, the RNA start site and 5 bp of DNA downstream were retained. No significant difference in CAT expression occurred as the deletion was extended from 352 to 32 bp before the RNA start site. Deletion of a further 10 bp, however, led to complete cessation of early promoter activity. Primer extension analysis of the 5' ends of the transcripts verified that the natural TK RNA start site was still used when only 32 bp of upstream DNA remained. Loss of early promoter activity was previously found when deletions were extended from 31 to 24 bp before the RNA start site of another vaccinia gene that is expressed constitutively throughout infection (M.A. Cochran, C. Puckett, and B. Moss, 1985, Proc. Natl. Acad. Sci. USA 82, 19-23). Sequence similarities in the promoter regions of these two genes were noted.

  12. Annual Report to Congress

    Science.gov (United States)

    2008-05-01

    8429) Vaccinia Immune Globulin, Intravenous (IND • #9141) Vaccinia Immune Globulin, Intravenous • (IND#10351, emergency use protocol) Cidofovir ...Smallpox Therapeutic, Intravenous • (S&T) Cidofovir Smallpox Therapeutic, Oral (S&T)• SIGA-246 Smallpox Therapeutic, Oral (fast track • status (S&T

  13. Incomplete but infectious vaccinia virions are produced in the absence of oncolysis in feline SCCF1 cells.

    Science.gov (United States)

    Parviainen, Suvi; Autio, Karoliina; Vähä-Koskela, Markus; Guse, Kilian; Pesonen, Sari; Rosol, Thomas J; Zhao, Fang; Hemminki, Akseli

    2015-01-01

    Vaccinia virus is a large, enveloped virus of the poxvirus family. It has broad tropism and typically virus replication culminates in accumulation and lytic release of intracellular mature virus (IMV), the most abundant form of infectious virus, as well as release by budding of extracellular enveloped virus (EEV). Vaccinia viruses have been modified to replicate selectively in cancer cells and clinically tested as oncolytic agents. During preclinical screening of relevant cancer targets for a recombinant Western Reserve strain deleted for both copies of the thymidine kinase and vaccinia growth factor genes, we noticed that confluent monolayers of SCCF1 cat squamous carcinoma cells were not destroyed even after prolonged infection. Interestingly, although SCCF1 cells were not killed, they continuously secreted virus into the cell culture supernatant. To investigate this finding further, we performed detailed studies by electron microscopy. Both intracellular and secreted virions showed morphological abnormalities on ultrastructural inspection, suggesting compromised maturation and morphogenesis of vaccinia virus in SCCF1 cells. Our data suggest that SCCF1 cells produce a morphologically abnormal virus which is nevertheless infective, providing new information on the virus-host cell interactions and intracellular biology of vaccinia virus.

  14. Incomplete but infectious vaccinia virions are produced in the absence of oncolysis in feline SCCF1 cells.

    Directory of Open Access Journals (Sweden)

    Suvi Parviainen

    Full Text Available Vaccinia virus is a large, enveloped virus of the poxvirus family. It has broad tropism and typically virus replication culminates in accumulation and lytic release of intracellular mature virus (IMV, the most abundant form of infectious virus, as well as release by budding of extracellular enveloped virus (EEV. Vaccinia viruses have been modified to replicate selectively in cancer cells and clinically tested as oncolytic agents. During preclinical screening of relevant cancer targets for a recombinant Western Reserve strain deleted for both copies of the thymidine kinase and vaccinia growth factor genes, we noticed that confluent monolayers of SCCF1 cat squamous carcinoma cells were not destroyed even after prolonged infection. Interestingly, although SCCF1 cells were not killed, they continuously secreted virus into the cell culture supernatant. To investigate this finding further, we performed detailed studies by electron microscopy. Both intracellular and secreted virions showed morphological abnormalities on ultrastructural inspection, suggesting compromised maturation and morphogenesis of vaccinia virus in SCCF1 cells. Our data suggest that SCCF1 cells produce a morphologically abnormal virus which is nevertheless infective, providing new information on the virus-host cell interactions and intracellular biology of vaccinia virus.

  15. Oncolytic and immunologic cancer therapy with GM-CSF-armed vaccinia virus of Tian Tan strain Guang9.

    Science.gov (United States)

    Deng, Lili; Fan, Jun; Guo, Mingming; Huang, Biao

    2016-03-28

    Targeted oncolytic vaccinia viruses are being developed as a novel strategy in cancer therapy. Arming vaccinia viruses with immunostimulatory cytokines can enhance antitumor efficacy. Such engineered oncolytic viruses, like JX-594, a Wyeth strain vaccinia virus modified with human granulocyte-macrophage colony-stimulating factor (GM-CSF), have shown promising results and have proceeded rapidly in clinical trials. However, the oncolytic potential of the Chinese vaccine strain Tian Tan (VTT) has not been explored. In this study, we constructed a targeted oncolytic vaccinia virus of Tian Tan strain Guang9 (VG9) expressing murine GM-CSF (VG9-GMCSF) and evaluated the antitumor effect of this recombinant vaccinia virus in a murine melanoma model. In vitro, viral replication and cytotoxicity of VG9-GMCSF was as potent as VG9; in vivo, VG9-GMCSF significantly inhibited the growth of subcutaneously implanted melanoma tumors, prolonged the survival of tumor-bearing mice, and produced an antitumor cytotoxic response. Such antitumor effect may be due to the lytic nature of virus as well as the stimulation of immune activity by GM-CSF production. Our results indicate that VG9-GMCSF induces strong tumoricidal activity, providing a potential therapeutic strategy for combating cancer.

  16. "A wild and wondrous ride": CDC field epidemiologists in the east Pakistan smallpox and cholera epidemics of 1958.

    Science.gov (United States)

    Greenough, Paul

    2011-02-01

    In mid-April of 1958 the Government of Pakistan summoned the press to announce a grave need for international aid to cope with smallpox and cholera epidemics in East Pakistan. In response, and with the backing of the US State Department, Dr. Alexander D. Langmuir, chief epidemiologist of the CDC, led a team of epidemiologists to assist authorities in Dacca strengthen their immunization programs. Langmuir's superiors hoped for a Cold War advantage, but he saw an opportunity for trainees in the Epidemic Intelligence Service to learn about public health in a developing country. Langmuir later described the episode as a "wild and wondrous ride," but it had been more like a nightmare: the East Pakistan health department had collapsed; a popular movement had taken over vaccination and squandered vaccine supplies; hostile journalists had questioned the Americans' deeper motives; and a professional rivalry opened between the Americans and a British epidemiologist named Aidan Cockburn. By the time the epidemic subsided in July 1958, 30 million Bengalis had been vaccinated for smallpox but another 20,000 had succumbed to the disease. This episode was CDC's first sustained foreign intervention, a precursor to its extensive role in the 1970s helping WHO eradicate smallpox from Bangladesh.

  17. Non-essential genes in the vaccinia virus HindIII K fragment: a gene related to serine protease inhibitors and a gene related to the 37K vaccinia virus major envelope antigen.

    Science.gov (United States)

    Boursnell, M E; Foulds, I J; Campbell, J I; Binns, M M

    1988-12-01

    The complete nucleotide sequence of a cloned copy of the HindIII K fragment of the WR strain of vaccinia virus has been determined. Eight open reading frames (ORFs) have been identified, on the basis of size and codon usage. The predicted amino acid sequences of the putative genes have been compared to the Protein Identification Resource and to published vaccinia virus sequences. One gene, predicted to encode a 42.2K protein, is highly related to the family of serine protease inhibitors. It shows approximately 25% identity to human antithrombin III and 19% identity to the cowpox virus 38K protein gene which is also related to serine protease inhibitors. The product of another gene shows a similar high level of identity to the 37K vaccinia virus major envelope antigen. The existence of viable deletion mutants and recombinants containing foreign DNA inserted into both these genes indicates that they are non-essential.

  18. Efficient cleavage of p220 by poliovirus 2Apro expression in mammalian cells: effects on vaccinia virus.

    Science.gov (United States)

    Aldabe, R; Feduchi, E; Novoa, I; Carrasco, L

    1995-10-24

    Poliovirus protease 2A cleaves p220, a component of initiation factor eIF-4F. Polyclonal antibodies that recognize p220 and the cleaved products from different species have been raised. Transfection of several cell lines with poliovirus 2Apro cloned in different plasmids leads to efficient cleavage of p220 upon infection with VT7, a recombinant vaccinia virus that expresses the T7 RNA polymerase. Under these conditions vaccinia virus protein synthesis is severely inhibited, while expression of poliovirus protein 2C from a similar plasmid has no effect. These results show by the first time the effects of p220 cleavage on vaccinia virus translation in the infected cells.

  19. Host range restriction of vaccinia virus in Chinese hamster ovary cells: relationship to shutoff of protein synthesis.

    Science.gov (United States)

    Drillien, R; Spehner, D; Kirn, A

    1978-12-01

    Chinese hamster ovary cells were found to be nonpermissive for vaccinia virus. Although early virus-induced events occurred in these cells (RNA and polypeptide synthesis), subsequent events appeared to be prevented by a very rapid and nonselective shutoff of protein synthesis. Within less than 2 h after infection, both host and viral protein syntheses were arrested. At low multiplicities of infection, inhibition of RNA synthesis with cordycepin resulted in failure of the virus to block protein synthesis. Moreover, infection of the cells in the presence of cycloheximide prevented the immediate onset of shutoff after reversal of cycloheximide. Inactivation of virus particles by UV irradiation also impaired the capacity of the virus to inhibit protein synthesis. These results suggested that an early vaccinia virus-coded product was implicated in the shutoff of protein synthesis. Either the nonpermissive Chinese hamster ovary cells were more sensitive to this inhibition than permissive cells, or a regulatory control of the vaccinia shutoff function was defective.

  20. A varíola, uma antiga inimiga Smallpox, an old foe

    Directory of Open Access Journals (Sweden)

    Hermann G. Schatzmayr

    2001-12-01

    Full Text Available A varíola acompanhou o homem por muitos séculos, causando mortes e lesões graves e irreversíveis. Usada como arma biológica em situações de guerra, volta a ser tema de discussão no mundo exatamente por essa possibilidade, apesar de ter sido erradicada das Américas em 1971, e do mundo em 1977. Os dados acumulados durante as Campanhas de Erradicação, mostram que a infecção se disseminava com relativa lentidão, através de contato muito próximo do receptor com o paciente. Infecções sub-clínicas em não-vacinados eram raras e vacinações de bloqueio em torno de novos casos, desde que estes fossem identificados e confirmados com rapidez, eram capazes de impedir a disseminação da infecção. As transmissões indiretas através de aerossóis eram menos comuns. Vacinados mesmo uma única vez, raramente apresentavam doença grave, no caso de reinfecção. A possibilidade de uso do vírus da varíola como arma biológica deve ser considerada como real, apesar de, com base nos conhecimentos atuais, serem necessárias suspensões virais de alta potência, lançadas muito próximo das pessoas a serem atingidas em grande número.Smallpox has accompanied mankind for centuries, causing deaths and permanent lesions. Used in the past as a biological weapon during wars, it has come into focus again precisely because of this renewed possibility, although the disease has been eradicated in the Americas since 1971 and worldwide since 1977. Data gathered during the eradication campaigns show that the disease spread relatively slowly through close contacts between patients and susceptibles. Sub-clinical infection in non-vaccinated individuals was a rare event, and blockade vaccination surrounding new cases (as long as these cases were confirmed early was able to prevent the disease from spreading in the community. Even with only one dose, vaccinated individuals rarely developed a serious case of the disease upon reinfection. The use of smallpox as

  1. Patent landscape of countermeasures against smallpox and estimation of grant attraction capability through patent landscape data.

    Science.gov (United States)

    Mayburd, Anatoly L; Kedia, Govind; Evans, Haydn W; Kaslival, Pritesh C

    2010-11-01

    The study was concerned with countermeasures against a possible smallpox outbreak. In the process of assessment 18 landscaping sectors were defined and described, the advantages and drawbacks of the corresponding countermeasures being reviewed. The data of the previously published influenza landscape were revisited. The current economic climate of deficit cutting (austerity) also puts emphasis on the optimization of capital investment. We used the materials of the landscape to define and analyze metrics of capital placement optimization. Value score was obtained by fitting patent landscape internals to the sale price of individual patents. Success score was obtained as a product of a-priori parameters that measure likelihood of emergence of a marketable product in a technological sector. Both scores were combined in a qualitative metric. Our methodology defined weight as a product of the sector size by the success score. We hypothesized - based on the material of two landscapes- that a life cycle of a technology begins in IP space with a high patent quality low volume "bud" of low weight, reaches maximum weight and then weight falls again when the technology becomes outdated. The weight and the annual dynamic of weight can serve a measure of investment risk and return. In this report we modeled investment by issue of government grants or purchase of patents by government. In the smallpox landscape the number of patents purchased by government agencies was the highest in the sectors with the highest weight and the trend was confirmed by the count of NIH grants issued in support of the technological sectors. In the influenza landscape only grant issue count was statistically meaningful and the trend was also confirmed. To better fit the grant support levels, the weight expression was optimized by using training coefficients. We propose to use value scores for evaluation of individual patent publications/company portfolios and to use weights for assessment of

  2. Cellular DNA ligase I is recruited to cytoplasmic vaccinia virus factories and masks the role of the vaccinia ligase in viral DNA replication.

    Science.gov (United States)

    Paran, Nir; De Silva, Frank S; Senkevich, Tatiana G; Moss, Bernard

    2009-12-17

    Vaccinia virus (VACV) encodes DNA polymerase and additional proteins that enable cytoplasmic replication. We confirmed the ability of VACV DNA ligase mutants to replicate and tested the hypothesis that cellular ligases compensate for loss of viral gene expression. RNA silencing of human DNA ligase I expression and a small molecule inhibitor of human DNA ligase I [corrected] severely reduced replication of viral DNA in cells infected with VACV ligase-deficient mutants, indicating that the cellular enzyme plays a complementary role. Replication of ligase-deficient VACV was greatly reduced and delayed in resting primary cells, correlating with initial low levels of ligase I and subsequent viral induction and localization of ligase I in virus factories. These studies indicate that DNA ligation is essential for poxvirus replication and explain the ability of ligase deletion mutants to replicate in dividing cells but exhibit decreased pathogenicity in mice. Encoding its own ligase might allow VACV to "jump-start" DNA synthesis.

  3. Ligation of double-stranded and single-stranded [Oligo(dT)] DNA by vaccinia virus DNA ligase

    OpenAIRE

    1996-01-01

    Vaccinia virus DNA ligase has been expressed in Escherichia coli, purified, and biochemically characterized. The enzyme ligates double-stranded (ds) DNA substrates with either cohesive or blunt-end termini and the latter reaction is stimulated by PEG. Vaccinia virus DNA ligase can also ligate oligo(dT) when annealed to either a poly(dA) or a poly(rA) backbone and, remarkably, free oligo(dT). This ligation of a single-stranded (ss) substrate is unique among eukaryotic DNA ligases. The enzyme r...

  4. Could hantavirus circulation superpose areas of highly endemic vaccinia virus outbreaks? A retrospective seroepidemiological study in State of Minas Gerais

    Directory of Open Access Journals (Sweden)

    Giliane de Souza Trindade

    2014-12-01

    Full Text Available Introduction Hantavirus infections have been described in several regions in Brazil through seroepidemiological studies. Usually, populations are associated with rural and wild environment mainly due to close contact to species of Sigmodontinae rodents, considered hantavirus reservoirs. Methods A retrospective serosurvey was conducted to access the hantavirus seroprevalence in people living in regions affected by bovine vaccinia outbreaks. Results Sera from 53 patients were analyzed and none of them presented anti-hantavirus IgG antibodies. Conclusions This study presents an opportunity to analyze seronegativity despite close and recurrent contact with known hantavirus reservoirs. Aspects of hantavirus and bovine vaccinia emergence are also discussed.

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

    Directory of Open Access Journals (Sweden)

    Rapp Ingrid

    2010-06-01

    Full Text Available Abstract 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.

  6. Construction of Recombinant Modified Vaccinia Ankara (MVA) Expressing Hepatitis B Virus Surface Antigen

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The T lymphocyte response has been shown to be the determinant in the clearance of many viral infections.Hence, therapeutic vaccine candidates against HBV are designed to enhance this response of the immune system.Vaccinia virus vector-based vaccines have been proposed as excellent candidates to elicit long-term and strong T lymphocyte mediated immune responses. In this study, the recombinant MVA expressing HBV surface antigen has been constructed, which can elicit a potent T cell mediated response. The ELISA results for the surface protein in the medium of the recombinant MVA, strongly indicate that the recombinant virus has been successfully obtained.

  7. Purification and Characterization of Recombinant Vaccinia L1R Protein from Escherichia coli

    Science.gov (United States)

    2016-08-01

    genomic Vaccinia DNA using high-fidelity PfuTurbo DNA polymerase (Agilent Technologies, Santa Clara, CA). Subsequently, the ORF was subcloned into...performing protein extraction . Harvested cells were resuspended in lysis buffer consisting of 100 mM Tris HCl, pH 8.0, 150 mM NaCl, 1% Triton X-100, 5 mM...showed sufficient cell lysis to extract L1R, although the yields were slightly lower. 2.3 Inclusion Body Enrichment and Solubilization After lysis

  8. Enhanced expression of the Epstein-Barr virus latent membrane protein by a recombinant vaccinia virus.

    Science.gov (United States)

    Stewart, J P; Hampson, I N; Heinrich, H W; Mackett, M; Arrand, J R

    1989-05-01

    The complete coding sequence of the Epstein-Barr virus strain B95-8 latent membrane protein (LMP) was cloned using a Raji cell cDNA library and genomic B95-8 DNA. The clone was characterized by sequencing and then used to make a recombinant vaccinia virus. This virus (VLMP) was shown to express a relatively high level of LMP in an authentic fashion. Antisera raised in rabbits against VLMP were shown to react with B95-8 LMP as well as cross-reacting with a 50K cellular protein.

  9. Analysis of a large cluster of nonessential genes deleted from a vaccinia virus terminal transposition mutant.

    Science.gov (United States)

    Kotwal, G J; Moss, B

    1988-12-01

    The principal objectives of this study were to analyze the structure and coding potential of a long segment of DNA missing from a previously isolated (B. Moss, E. Winters, and J. A. Cooper (1981) J. Virol. 40, 387-395) attenuated variant of vaccinia virus strain WR and to examine the precise changes in the genome accompanying the deletion. The sequences of a 14.5-kbp region located at the left end of the standard vaccinia virus genome, extending from within the inverted terminal repetition (ITR) of the HindIII C fragment to the end of the HindIII N fragment, and of a 3-kbp segment from a corresponding region of the variant genome were determined. A comparison of these sequences revealed that the variant contained a deletion of 12 kbp and an insertion of 2.1 kbp. The origin of the inserted DNA was traced to the HindIII B region by using oligonucleotide probes indicating that a transposition of unique DNA located adjacent to the right ITR had occurred. Structural analysis indicated no extensive homologies, nucleotide substitutions, additions, or deletions at the boundaries of the transposed DNA. Examination of the right end of the variant genome indicated that a copy of the transposed DNA was still present and, therefore, the length of the ITR had been increased by 2.1 kbp. The variant genome could have formed by a mechanism that resulted in the replacement of a 22-kbp left-terminal fragment with a 12-kbp right-terminal fragment. The DNA missing from the variant and contained within the standard vaccinia virus WR genome contains 17 contiguous open reading frames (ORFs), all of which are directed leftward and apparently not required for replication in cultured cells. One deleted ORF has a 60% sequence similarity to another gene encoding a 42,000-Da protein present within the ITR suggesting that duplications have previously occurred during the evolution of vaccinia virus. Another deleted ORF has a 39% sequence similarity to a complement 4b binding protein. The

  10. Vectores recombinantes basados en el virus Vaccinia modificado de Ankara (MVA) como vacunas contra la leishmaniasis

    OpenAIRE

    Pérez Jiménez, Eva; Larraga, Vicente; Esteban, Mariano

    2005-01-01

    Vectores recombinantes basados en el virus vaccinia modificado de Ankara (MVA) como vacunas contra la leishmaniasis. Los vectores de la invención contienen secuencias codificantes de la proteína LACK, preferentemente insertadas en el locus de hemaglutinina del virus y bajo el control de un promotor que permite su expresión a lo largo del ciclo de infección del virus. Son vectores seguros, estables, que dan lugar a una potente respuesta inmune que confiere protección frente a la leishmaniasis,...

  11. Serological studies on an outbreak of smallpox in the State of Bahia - Brazil in 1969

    Directory of Open Access Journals (Sweden)

    Eduardo de Azeredo Costa

    1972-01-01

    Full Text Available Four weeks after Containment Vaccination undertaken against the largest outbreak of smallpox occured in Brazil in 1969, that of the municipality of Utinga, Bahia, 99 samples of serum were collected from the local population. These samples were classified in four groups: a - Individuals with a history of variola prior to the beginning of present outbreak in town (15 sera; "Previous smallpox group"; b - Individuals with primary vaccination, with no record variola, at the time of containment measures (15 sera. "Primary vaccinated group"; c - Individuals with no previous record of variola revaccinated with "take" at the time of containment (15 sera0, "Revaccinated group"; d - Individuals who contracted variola in present outbreak (54 sera these were subdivided in four sub-groups, according to dates on which cases ocurred, "Variola in outbreak group". Serological study of samples was done by tests of hemagglutination inhibition, neutralization, and complement fixation. It was observed that HI titers were significantly lower in cases of previous smallpox than in other groups. Although they were slightly higher on revaccinated individuals than on primary vaccinated group and than in the group of variola in outbreak, this difference was not significant. Those same antibodies were present in all cases of variola in outbreak, and it was found that titers decreased in direct proportion to time elapsed from occurrence of cases. Neutralizing antibodies proved to be significantly higher on the revaccinated group than on variola in outbreak group, and higher on these than on primary vaccinated and on the previous smallpox groups. In cases from the variola in outbreak it was verified that neutralizing antibodies remained stable, although with great variation in titers. Tests of complement fixation could not be undertaken on all samples, because many of them proved to have anticomplementarity. However, it was found that complement fixing antibodies diminished

  12. Game theory of pre-emptive vaccination before bioterrorism or accidental release of smallpox.

    Science.gov (United States)

    Molina, Chai; Earn, David J D

    2015-06-06

    Smallpox was eradicated in the 1970s, but new outbreaks could be seeded by bioterrorism or accidental release. Substantial vaccine-induced morbidity and mortality make pre-emptive mass vaccination controversial, and if vaccination is voluntary, then there is a conflict between self- and group interests. This conflict can be framed as a tragedy of the commons, in which herd immunity plays the role of the commons, and free-riding (i.e. not vaccinating pre-emptively) is analogous to exploiting the commons. This game has been analysed previously for a particular post-outbreak vaccination scenario. We consider several post-outbreak vaccination scenarios and compare the expected increase in mortality that results from voluntary versus imposed vaccination. Below a threshold level of post-outbreak vaccination effort, expected mortality is independent of the level of response effort. A lag between an outbreak starting and a response being initiated increases the post-outbreak vaccination effort necessary to reduce mortality. For some post-outbreak vaccination scenarios, even modest response lags make it impractical to reduce mortality by increasing post-outbreak vaccination effort. In such situations, if decreasing the response lag is impossible, the only practical way to reduce mortality is to make the vaccine safer (greater post-outbreak vaccination effort leads only to fewer people vaccinating pre-emptively).

  13. Taking advantage of the positive side-effects of smallpox vaccination.

    Science.gov (United States)

    Mayr, A

    2004-06-01

    From the introduction of smallpox vaccination approximately 200 years ago right up to its discontinuation (1980), reports by physicians and scientists about positive side-effects such as healing of chronic skin rashes, reduced susceptibility to various infectious diseases, e.g. measles, scarlet fever and whooping cough, and even the prophylactic use of the vaccination, e.g. against syphilis, were published again and again. Comparison with the period after cessation of vaccination confirms the experiences of the above vaccinators. As early as 1956, targeted research on these observations led to evidence of the 'ring-zone phenomenon', i.e. the production of soluble antiviral substances in infected chicken embryos and cell cultures. With the help of modern immunological and bioengineering methods, it was later possible to demonstrate that these effects are based on the activation of lymphoreticular cells and the regulatory effect of certain cytokines within the context of the non-specific immune system. These findings led to the development of paramunization with paraspecific vaccines from highly attenuated animal pox viruses. During attenuation, deletions in the virus DNA occur. Attenuated animal pox strains are therefore suited for the production of vector vaccines. The fact that these vector vaccines demonstrate an especially high level of paraspecific efficacy and lack harmful effects is likewise the result of the attenuated animal pox viruses. Optimum regulation of the entire immune system leads to increased paramunity already in the first few days after vaccination and to enhanced antigen recognition and thus accelerated commencement of specific immunity.

  14. 痘苗病毒天坛株的减毒与其作为疫苗载体研究进展%Attenuation of the smallpox vaccine vaccinia Tiantan strain and its use as a safe vaccine live vector

    Institute of Scientific and Technical Information of China (English)

    余文博; 刘利; 陈志伟

    2011-01-01

    1 痘苗病毒天坛株的历史天花是人类历史上最可怕的传染病之一.天花由天花病毒(variola virus)引起,人是该病毒的惟一宿主.人感染天花病毒后的死亡率达到30%~40%[1].世界上公认的对天花最早的准确记录来自中国,晋代药学家葛洪于公元430年左右,在《肘后备急方》中对天花有清楚的描述[1].

  15. Oral Immunization with Recombinant Vaccinia Virus Prime and Intramuscular Protein Boost Provides Protection against Intrarectal Simian-Human Immunodeficiency Virus Challenge in Macaques.

    Science.gov (United States)

    Thippeshappa, Rajesh; Tian, Baoping; Cleveland, Brad; Guo, Wenjin; Polacino, Patricia; Hu, Shiu-Lok

    2015-12-30

    Human immunodeficiency virus type 1 (HIV-1) acquisition occurs predominantly through mucosal transmission. We hypothesized that greater mucosal immune responses and protective efficacy against mucosal HIV-1 infection may be achieved by prime-boost immunization at mucosal sites. We used a macaque model to determine the safety, immunogenicity, and protective efficacy of orally delivered, replication-competent but attenuated recombinant vaccinia viruses expressing full-length HIV-1 SF162 envelope (Env) or simian immunodeficiency virus (SIV) Gag-Pol proteins. We examined the dose and route that are suitable for oral immunization with recombinant vaccinia viruses. We showed that sublingual inoculation of two vaccinia virus-naive pigtailed macaques with 5 × 10(8) PFU of recombinant vaccinia viruses was safe. However, sublingual inoculation with a higher dose or tonsillar inoculation resulted in secondary oral lesions, indicating the need to optimize the dose and route for oral immunization with replication-competent vaccinia virus vectors. Oral priming alone elicited antibody responses to vaccinia virus and to the SF162 Env protein. Intramuscular immunization with the SF162 gp120 protein at either 20 or 21 weeks postpriming resulted in a significant boost in antibody responses in both systemic and mucosal compartments. Furthermore, we showed that immune responses induced by recombinant vaccinia virus priming and intramuscular protein boosting provided protection against intrarectal challenge with the simian-human immunodeficiency virus SHIV-SF162-P4.

  16. [Seeds against smallpox: Joaquim Vás and the scientific translation of bananeira brava seeds in Goa, India (1894-1930)].

    Science.gov (United States)

    Roque, Ricardo

    2004-01-01

    In 1914, in the former Portuguese colony of Goa, India, the physician António Joaquim Vás announced the discovery of a wonderful treatment for smallpox, entailing clinical application of seeds from the bananeira brava (Heliconia biabi Sw.m.), a plant remedy allegedly derived from Indian medical practices. The present article explores the circumstances surrounding the successes and failures of this discovery. The concept of scientific translation is used to interpret the transformation of bananeira brava seeds into an early twentieth-century remedy for smallpox. This transfer from indigenous use to scientific therapeutic constitutes the creation of a quasi-medicine, that is, a case of 'medium translation'. Although these seeds occupy a problematic place within the program of scientific translation, they enjoyed active circulation within science and remained a part of medical practices for combating smallpox.

  17. Analysis of pregnancy and infant health outcomes among women in the National Smallpox Vaccine in Pregnancy Registry who received Anthrax Vaccine Adsorbed.

    Science.gov (United States)

    Conlin, Ava Marie S; Bukowinski, Anna T; Gumbs, Gia R

    2015-08-26

    The National Smallpox Vaccine in Pregnancy Registry (NSVIPR) actively follows women inadvertently vaccinated with smallpox vaccine during or shortly before pregnancy to evaluate their reproductive health outcomes. Approximately 65% of NSVIPR participants also inadvertently received Anthrax Vaccine Adsorbed (AVA) while pregnant, providing a ready opportunity to evaluate pregnancy and infant health outcomes among these women. AVA-exposed pregnancies were ascertained using NSVIPR and electronic healthcare data. Rates of pregnancy loss and infant health outcomes, including major birth defects, were compared between AVA-exposed and AVA-unexposed pregnancies. Analyses included AVA-exposed and AVA-unexposed pregnant women who also received smallpox vaccine 28 days prior to or during pregnancy. Rates of adverse outcomes among the AVA-exposed group were similar to or lower than expected when compared with published reference rates and the AVA-unexposed population. The findings provide reassurance of the safety of AVA when inadvertently received by a relatively young and healthy population during pregnancy.

  18. Risk of lymphoma and leukaemia after bacille Calmette-Guérin and smallpox vaccination: a Danish case-cohort study

    DEFF Research Database (Denmark)

    Villumsen, Marie; Sørup, Signe; Jess, Tine

    2009-01-01

    Vaccines may have non-specific effects as suggested mainly in mortality studies from low-income countries. The objective was to examine the effects of BCG and smallpox vaccinations on subsequent risk of lymphoma and leukaemia in a Danish population experiencing rapid out-phasing of these vaccines...... cohort and analysed in a case-cohort design. BCG vaccination reduced the risk of lymphomas (HR=0.49 (95% CI: 0.26-0.93)), whereas smallpox vaccination did not (HR=1.32 (0.56-3.08)). With the small number of leukaemia cases, the analysis of leukaemia had limited power (BCG vaccination HR=0.81 (0.......31-2.16); smallpox vaccination HR=1.32 (0.49-3.53)). The present study with very reliable vaccine history information indicates a beneficial effect of BCG vaccination on the risk of lymphomas....

  19. Fine structure of the vaccinia virion determined by controlled degradation and immunolocalization

    Energy Technology Data Exchange (ETDEWEB)

    Moussatche, Nissin, E-mail: nissin@ufl.edu; Condit, Richard C.

    2015-01-15

    The vaccinia virion is a membraned, slightly flattened, barrel-shaped particle, with a complex internal structure featuring a biconcave core flanked by lateral bodies. Although the architecture of the purified mature virion has been intensely characterized by electron microscopy, the distribution of the proteins within the virion has been examined primarily using biochemical procedures. Thus, it has been shown that non-ionic and ionic detergents combined or not with a sulfhydryl reagent can be used to disrupt virions and, to a limited degree, separate the constituent proteins in different fractions. Applying a controlled degradation technique to virions adsorbed on EM grids, we were able to immuno-localize viral proteins within the virion particle. Our results show after NP40 and DTT treatment, membrane proteins are removed from the virion surface revealing proteins that are associated with the lateral bodies and the outer layer of the core wall. Combined treatment using high salt and high DTT removed lateral body proteins and exposed proteins of the internal core wall. Cores treated with proteases could be disrupted and the internal components were exposed. Cts8, a mutant in the A3 protein, produces aberrant virus that, when treated with NP-40 and DTT, releases to the exterior the virus DNA associated with other internal core proteins. With these results, we are able to propose a model for the structure the vaccinia virion.

  20. Modified vaccinia virus Ankara exerts potent immune modulatory activities in a murine model.

    Directory of Open Access Journals (Sweden)

    Miriam Nörder

    Full Text Available BACKGROUND: Modified vaccinia virus Ankara (MVA, a highly attenuated strain of vaccinia virus, has been used as vaccine delivery vector in preclinical and clinical studies against infectious diseases and malignancies. Here, we investigated whether an MVA which does not encode any antigen (Ag could be exploited as adjuvant per se. METHODOLOGY/PRINCIPAL FINDINGS: We showed that dendritic cells infected in vitro with non-recombinant (nr MVA expressed maturation and activation markers and were able to efficiently present exogenously pulsed Ag to T cells. In contrast to the dominant T helper (Th 1 biased responses elicited against Ags produced by recombinant MVA vectors, the use of nrMVA as adjuvant for the co-administered soluble Ags resulted in a long lasting mixed Th1/Th2 responses. CONCLUSIONS/SIGNIFICANCE: These findings open new ways to potentiate and modulate the immune responses to vaccine Ags depending on whether they are co-administered with MVA or encoded by recombinant viruses.

  1. Microbiota is an essential element for mice to initiate a protective immunity against Vaccinia virus.

    Science.gov (United States)

    Lima, Maurício T; Andrade, Ana C S P; Oliveira, Graziele P; Calixto, Rafael S; Oliveira, Danilo B; Souza, Éricka L S; Trindade, Giliane S; Nicoli, Jacques R; Kroon, Erna G; Martins, Flaviano S; Abrahão, Jônatas S

    2016-02-01

    The gastrointestinal tract of vertebrates harbors one of the most complex ecosystems known in microbial ecology and this indigenous microbiota almost always has a profound influence on host-parasite relationships, which can enhance or reduce the pathology of the infection. In this context, the impact of the microbiota during the infection of several viral groups remains poorly studied, including the family Poxviridae. Vaccinia virus (VACV) is a member of this family and is the causative agent of bovine vaccinia, responsible for outbreaks that affect bovines and humans. To determine the influence of the microbiota in the development of the disease caused by VACV, a comparative study using a murine model was performed. Germ-free and conventional, 6- to 7-week-old Swiss NIH mice were infected by tail scarification and intranasally with VACV. Moreover, immunosuppression and microbiota reposition were performed, to establish the interactions among the host's immune system, microbiota and VACV. The data demonstrate that the microbiota is essential for the effective immune response of mice against VACV in intranasal inoculation and to control the virus at the primary site of infection. Furthermore, this study is the first to show that Swiss conventional mice are refractory to the intranasal infection of VACV.

  2. Elements in the Development of a Production Process for Modified Vaccinia Virus Ankara

    Directory of Open Access Journals (Sweden)

    Volker Sandig

    2013-11-01

    Full Text Available The production of several viral vaccines depends on chicken embryo fibroblasts or embryonated chicken eggs. To replace this logistically demanding substrate, we created continuous anatine suspension cell lines (CR and CR.pIX, developed chemically-defined media, and established production processes for different vaccine viruses. One of the processes investigated in greater detail was developed for modified vaccinia virus Ankara (MVA. MVA is highly attenuated for human recipients and an efficient vector for reactogenic expression of foreign genes. Because direct cell-to-cell spread is one important mechanism for vaccinia virus replication, cultivation of MVA in bioreactors is facilitated if cell aggregates are induced after infection. This dependency may be the mechanism behind our observation that a novel viral genotype (MVA-CR accumulates with serial passage in suspension cultures. Sequencing of a major part of the genomic DNA of the new strain revealed point mutations in three genes. We hypothesize that these changes confer an advantage because they may allow a greater fraction of MVA-CR viruses to escape the host cells for infection of distant targets. Production and purification of MVA-based vaccines may be simplified by this combination of designed avian cell line, chemically defined media and the novel virus strain.

  3. Treatment of Vaccinia and Cowpox Virus Infections in Mice with CMX001 and ST-246

    Directory of Open Access Journals (Sweden)

    Earl R. Kern

    2010-12-01

    Full Text Available Although a large number of compounds have been identified with antiviral activity against orthopoxviruses in tissue culture systems, it is highly preferred that these compounds have activity in vivo before they can be seriously considered for further development. One of the most commonly used animal models for the confirmation of this activity has been the use of mice infected with either vaccinia or cowpox viruses. These model systems have the advantage that they are relatively inexpensive, readily available and do not require any special containment facilities; therefore, relatively large numbers of compounds can be evaluated in vivo for their activity. The two antiviral agents that have progressed from preclinical studies to human safety trials for the treatment of orthopoxvirus infections are the cidofovir analog, CMX001, and an inhibitor of extracellular virus formation, ST-246. These compounds are the ones most likely to be used in the event of a bioterror attack. The purpose of this communication is to review the advantages and disadvantages of using mice infected with vaccinia and cowpox virus as surrogate models for human orthopoxvirus infections and to summarize the activity of CMX001 and ST-246 in these model infections.

  4. Preclinical evaluation of oncolytic vaccinia virus for therapy of canine soft tissue sarcoma.

    Directory of Open Access Journals (Sweden)

    Ivaylo Gentschev

    Full Text Available Virotherapy using oncolytic vaccinia virus (VACV strains is one promising new strategy for canine cancer therapy. In this study we describe the establishment of an in vivo model of canine soft tissue sarcoma (CSTS using the new isolated cell line STSA-1 and the analysis of the virus-mediated oncolytic and immunological effects of two different Lister VACV LIVP1.1.1 and GLV-1h68 strains against CSTS. Cell culture data demonstrated that both tested VACV strains efficiently infected and destroyed cells of the canine soft tissue sarcoma line STSA-1. In addition, in our new canine sarcoma tumor xenograft mouse model, systemic administration of LIVP1.1.1 or GLV-1h68 viruses led to significant inhibition of tumor growth compared to control mice. Furthermore, LIVP1.1.1 mediated therapy resulted in almost complete tumor regression and resulted in long-term survival of sarcoma-bearing mice. The replication of the tested VACV strains in tumor tissues led to strong oncolytic effects accompanied by an intense intratumoral infiltration of host immune cells, mainly neutrophils. These findings suggest that the direct viral oncolysis of tumor cells and the virus-dependent activation of tumor-associated host immune cells could be crucial parts of anti-tumor mechanism in STSA-1 xenografts. In summary, the data showed that both tested vaccinia virus strains and especially LIVP1.1.1 have great potential for effective treatment of CSTS.

  5. Early transcription factor subunits are encoded by vaccinia virus late genes.

    Science.gov (United States)

    Gershon, P D; Moss, B

    1990-06-01

    The vaccinia virus early transcription factor (VETF) was shown to be a virus-encoded heterodimer. The gene for the 82-kDa subunit was identified as open reading frame (ORF) A8L, based on the N-terminal sequence of factor purified by using DNA-affinity magnetic beads. The 70-kDa subunit of VETF was refractory to N-terminal analysis, and so N-terminal sequences were obtained for three internal tryptic peptides. All three peptides matched sequences within ORF D6R. ORFs A8L and D6R are located within the central region of the vaccinia virus genome and are separated by about 13,600 base pairs. Proteins corresponding to the 3' ends of ORFs A8L and D6R were overexpressed in Escherichia coli and used to prepare antisera that bound to the larger and smaller subunits, respectively, of affinity-purified VETF. Immunoblot analysis of proteins from infected cells indicated that both subunits are expressed exclusively in the late phase of infection, just prior to their packaging in virus particles. The two subunits of VETF have no significant local or overall amino acid sequence homology to one another, to other entries in biological sequence data bases including bacterial sigma factors, or to recently determined sequences of some eukaryotic transcription factors. The 70-kDa subunit, however, has motifs in common with a super-family of established and putative DNA and RNA helicases.

  6. DNA sequence analysis of the Hind III M fragment from Chinese vaccine strain of vaccinia virus.

    Science.gov (United States)

    Liu, V J; Jin, Q; Jin, D Y; Hou, Y D

    1989-01-01

    The complete DNA sequence of the Hind III M fragment of vaccinia virus (VV) Tian Tan strain genome was determined by the dideoxynucleotide chain termination method. Three open reading frames (ORFs) were identified in the complementary strand of the sequence, comprised of 2218bp. Among them, ORF K1 initiates its transcription at -45 of the Hind III K fragment. The deduced peptide encoded by K1 contains 284 amino acids with a calculated molecular weight of 32.48 KDa. Its sequence is homologous to the host range protein of VV Copenhagen strain; the variation is only 2.46% at the amino acid level. ORF M2 could encode a peptide of 21.94 KDa with 196 amino acids. This gene was shown to be homologous to that of the 23 KDa peptide of herpes simplex virus type I. A non-coding region of 204bp located between K1 and M2 is rich in palindromic structures. ORF M1 extends its 3' terminus into the Hind III N fragment. Within the M fragment, M1 can only encode 212 amino acids. The major part of ORF M1 is very similar to the M portion of a possible alpha-amanitin resistance gene isolated from VV-WR strain. This work provides a molecular foundation in the construction of a new insertion vector for the preparation of a recombinant vaccinia virus to be used as a polyvalent live vaccine.

  7. Middle east respiratory syndrome coronavirus spike protein delivered by modified vaccinia virus ankara efficiently induces virus-neutralizing antibodies

    NARCIS (Netherlands)

    F. Song (Fei); R. Fux (Robert); L.B.V. Provacia (Lisette); A. Volz (Asisa); M. Eickmann; S. Becker (Stephan); A.D.M.E. Osterhaus (Albert); B.L. Haagmans (Bart); G. Suttera (Gerd)

    2013-01-01

    textabstractMiddle 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 sta

  8. Immunogenicity and protective efficacy of recombinant Modified Vaccinia virus Ankara candidate vaccines delivering West Nile virus envelope antigens

    NARCIS (Netherlands)

    Volz, Asisa; Lim, Stephanie; Kaserer, Martina; Pijlman, Gorben P.

    2016-01-01

    West Nile virus (WNV) cycles between insects and wild birds, and is transmitted via mosquito vectors to horses and humans, potentially causing severe neuroinvasive disease. Modified Vaccinia virus Ankara (MVA) is an advanced viral vector for developing new recombinant vaccines against infectious

  9. CD40 ligand and tdTomato-armed vaccinia virus for induction of antitumor immune response and tumor imaging.

    Science.gov (United States)

    Parviainen, S; Ahonen, M; Diaconu, I; Hirvinen, M; Karttunen, Å; Vähä-Koskela, M; Hemminki, A; Cerullo, V

    2014-02-01

    Oncolytic vaccinia virus is an attractive platform for immunotherapy. Oncolysis releases tumor antigens and provides co-stimulatory danger signals. However, arming the virus can improve efficacy further. CD40 ligand (CD40L, CD154) can induce apoptosis of tumor cells and it also triggers several immune mechanisms. One of these is a T-helper type 1 (Th1) response that leads to activation of cytotoxic T-cells and reduction of immune suppression. Therefore, we constructed an oncolytic vaccinia virus expressing hCD40L (vvdd-hCD40L-tdTomato), which in addition features a cDNA expressing the tdTomato fluorochrome for detection of virus, potentially important for biosafety evaluation. We show effective expression of functional CD40L both in vitro and in vivo. In a xenograft model of bladder carcinoma sensitive to CD40L treatment, we show that growth of tumors was significantly inhibited by the oncolysis and apoptosis following both intravenous and intratumoral administration. In a CD40-negative model, CD40L expression did not add potency to vaccinia oncolysis. Tumors treated with vvdd-mCD40L-tdtomato showed enhanced efficacy in a syngenic mouse model and induced recruitment of antigen-presenting cells and lymphocytes at the tumor site. In summary, oncolytic vaccinia virus coding for CD40L mediates multiple antitumor effects including oncolysis, apoptosis and induction of Th1 type T-cell responses.

  10. Immunogenicity and protective efficacy of recombinant Modified Vaccinia virus Ankara candidate vaccines delivering West Nile virus envelope antigens

    NARCIS (Netherlands)

    Volz, Asisa; Lim, Stephanie; Kaserer, Martina; Pijlman, Gorben P.

    2016-01-01

    West Nile virus (WNV) cycles between insects and wild birds, and is transmitted via mosquito vectors to horses and humans, potentially causing severe neuroinvasive disease. Modified Vaccinia virus Ankara (MVA) is an advanced viral vector for developing new recombinant vaccines against infectious

  11. Dogs and Opossums Positive for Vaccinia Virus during Outbreak Affecting Cattle and Humans, São Paulo State, Brazil.

    Science.gov (United States)

    Peres, Marina G; Barros, Claudenice B; Appolinário, Camila M; Antunes, João M A P; Mioni, Mateus S R; Bacchiega, Thais S; Allendorf, Susan D; Vicente, Acácia F; Fonseca, Clóvis R; Megid, Jane

    2016-02-01

    During a vaccinia virus (VACV) outbreak in São Paulo State, Brazil, blood samples were collected from cows, humans, other domestic animals, and wild mammals. Samples from 3 dogs and 3 opossums were positive for VACV by PCR. Results of gene sequencing yielded major questions regarding other mammalian species acting as reservoirs of VACV.

  12. Chemical inactivation of recombinant vaccinia viruses and the effects on antigenicity and immunogenicity of recombinant simian immunodeficiency virus envelope glycoproteins.

    NARCIS (Netherlands)

    E.G.J. Hulskotte (Ellen); M.E.M. Dings (Marlinda); S.G. Norley (Stephen); A.D.M.E. Osterhaus (Albert)

    1997-01-01

    textabstractThe efficiency of paraformaldehyde (PFA) and binary ethylenimine (BEI) in inactivating recombinant vaccinia virus (rVV), present in baby hamster kidney cells expressing simian immunodeficiency virus envelope glycoproteins (SIV-Env), was measured in a series of inactivation studies. Both

  13. Concanavalin A-mediated cell agglutinability induced by Vaccinia virions. [Uv radiation, /sup 125/I tracer technique

    Energy Technology Data Exchange (ETDEWEB)

    Mbuy, G.; Bubel, H.C.

    1978-12-01

    The induction of enhanced concanavalin A (Con A)-mediated cellular agglutinability by purified vaccinia virus was examined quantitatively. Increased HEp-2 cell agglutinability by the lectin occurred within the first hour of infection and persisted without further change throughout the virus infectious cycle. Ultraviolet, but not heat-inactivated, virus was as effective as infectious virus in causing increased Con A agglutinability. Inhibition of viral and host cell protein synthesis by Streptovitacin A failed to alter the lectin response to vaccinia virus infection. Fluorescein-labeled Con A was observed to form clusters and large fluorescent patches on the infected cell surface during the earliest stage of infection. Studies with /sup 125/I-labeled Con A revealed an early but minimal increase in lectin binding to infected cells. After the first hour of infection, no further increase in Con A binding was observed. When cells were exposed to purified vaccinia virus surface tubules increased Con A agglutinability comparable to that obtained with native virus was demonstrated. Con A-mediated agglutinability of cells was temperature-dependent and displayed a higher temperature transition in infected cells. These data suggest that upon contact with the host cell, vaccinia virions or surface tubules induce alterations in the plasma membrane which are reflected in an enhanced agglutinability by Con A.

  14. Combined immunity of DNA vector and recombinant vaccinia virus expressing Gag proteins of equine infectious anemia virus

    Institute of Scientific and Technical Information of China (English)

    DAI Chunming; ZHANG Xiaoyan; WANG Shuhui; LIU Ying; DUAN Danli; SHEN Rongxian; SHAO Yiming

    2004-01-01

    In order to develop a new vaccine candidate for equine infectious anemia virus (EIAV), gag gene of Chinese donkey leukocyte attenuated strain (EIAV DLV) and its parental virulent strain (EIAV LN) were inserted respectively into the TK region of the Tiantan strain (VV) of vaccinia virus by homologous recombination and the positive clone was confirmed by blue plaque assay. Protein expression was examined by Western blot. Prime and prime-boost procedures were used to immunize mice with two DNA vectors and two recombinant vaccinia viruses expressing EIAV Gag proteins. The results showed that the specific lysis of CTL responses in the DNA+rVV groups was stronger than those in the DNA groups, amounting to 31%. Although the levels of specific antibodies were not significantly different, we could conclude that the recombinant vaccinia virus could boost the cellular responses following DNA vector priming. There was no detectable difference between the immune responses induced by DLV and LN Gag proteins. This data demonstrates that the combined immunity of DNA vector and recombinant vaccinia virus expressing EIAV gag proteins, utilizing the prime-boost procedure, can drive immunized mice to produce powerful cellular responses. These results lay an important foundation for the development of a new EIAV genetic engineering vaccine.

  15. Analysis of the L1 gene product of human papillomavirus type 16 by expression in a vaccinia virus recombinant.

    Science.gov (United States)

    Browne, H M; Churcher, M J; Stanley, M A; Smith, G L; Minson, A C

    1988-06-01

    The L1 open reading frame of human papillomavirus type 16 (HPV16) has been expressed in vaccinia virus under the control of both the 7.5K early and late promoter, and the 4b major late promoter. Antibodies to a beta-galactosidase fusion protein containing a C-terminal portion of the HPV16 L1 gene product were used to compare the levels of L1 expression in the two recombinants, and showed that greater levels of expression were obtained when the gene was placed under the control of the 4b late promoter. Immunofluorescence studies revealed a nuclear location of the L1 gene product when expressed in vaccinia virus. Antibodies to the beta-galactosidase fusion protein detected a major polypeptide species of 57K and a minor species of 64K in Western blots of recombinant-infected cell lysates. The 64K species was not detected when cells were infected in the presence of tunicamycin, indicating that the primary translation product of the HPV16 L1 open reading frame is modified by N-linked glycosylation when expressed in vaccinia virus. Whereas antibodies to HPV16 L1 fusion proteins and to a peptide containing amino acids from the C terminus of HPV16 L1 reacted well in Western blots with the HPV16 L1 target expressed in vaccinia virus, no reactivity was observed with antibodies to bovine papillomavirus type 1 particles or to a HPV6b fusion protein.

  16. [The journey of the vaccine against smallpox: one expedition, two oceans, three continents, and thousands of children].

    Science.gov (United States)

    Tuells, José; Duro-Torrijos, José Luis

    2015-01-01

    Spain encouraged, during the Bourbon dynasty, the formation of scientific expeditions, among which was the Royal Philanthropic Vaccine Expedition, an example of biopolitics applied by the state in order to protect health. The expedition went all over the world, using children as a reservoir to transport the vaccine fluid. Francisco Xavier Balmis established a human chain that arm-to-arm materialized the success of the mission. The characteristics and difficulties which children had to pass through and their contribution to the spread of the smallpox vaccine are analyzed.

  17. US Military Service Members Vaccinated Against Smallpox in 2003 and 2004 Experience a Slightly Higher Risk of Hospitalization Postvaccination

    Science.gov (United States)

    2008-01-01

    conditions 18 0.0 7 0.0 1.92 0.79, 4.69 280 Iron deficiency anemias 16 0.0 7 0.0 1.52 0.60, 3.84 288 Diseases of white blood cells 11 0.0 6 0.0 1.43...public release: distribution is unlimited. Naval Health Research Center 140 Sylvester Road San Diego, California 92106 This article appeared...Naval Health Research Center US Military Service Members Vaccinated Against Smallpox in 2003 and 2004 Experience a Slightly Higher Risk of

  18. Large-scale use of freeze-dried smallpox vaccine prepared in primary cultures of rabbit kidney cells

    Science.gov (United States)

    Hekker, A. C.; Bos, J. M.; Rai, N. Kumara; Keja, J.; Cuboni, G.; Emmet, B.; Djalins, J.

    1976-01-01

    A lyophilized smallpox vaccine made from infected monolayer cultures of primary rabbit kidney cells was used together with a calf lymph vaccine in a field trial in Lombok, Indonesia, in 1973. About 60 000 children below 15 years of age were vaccinated: some 50 000 with the tissue culture vaccine and about 10 000 with calf lymph vaccine. Similar results were obtained with both vaccines in primary vaccinees and in revaccinees as regards the take rate, pock reactions, and serious secondary reactions. PMID:1088108

  19. Infecções humanas causadas por poxvirus relacionados ao vírus vaccinia no Brasil Human infections caused by vaccinia-like poxviruses in Brazil

    Directory of Open Access Journals (Sweden)

    Hermann G. Schatzmayr

    2009-12-01

    Full Text Available A partir de 1999, infecções humanas por Orthopoxvirus vem sendo observadas em pelo menos oito estados no país, com a formação de vesículas as quais evoluem para pústulas e crostas, principalmente nos membros superiores e face, após contacto com bovinos apresentando lesões semelhantes no úbere. Alem das lesões na pele, foram descritas nos pacientes reações ganglionares axilares por vezes dolorosas, febre, cefaléia, fadiga, desidratação, anorexia, sudorese, artralgia e mialgia, evoluindo o quadro por três a quatro semanas. Lesão vulvar bem como transmissão intrafamiliar foram igualmente descritas. Estudos moleculares demonstraram que os poxvirus identificados são geneticamente relacionados a amostras do vírus vaccinia utilizadas no passado, nas campanhas de vacinação. Especimens clínicos de 80 infecções humanas foram estudados no laboratório e a infecção por orthopoxvirus confirmada em 68 casos. São apresentadas lesões observadas em pacientes bem como discutidas as implicações desta zoonose no Brasil.Since 1999, human infection caused by Orthopoxvirus has been observed in at least eight Brazilian states, with the presence of vesicles that evolve to pustules and crusts, especially on the hands, arms and face, after contact with cows showing comparable lesions on the udder. In addition to the skin lesions, there have been descriptions of patients with axillary ganglionic reactions that are sometimes painful, along with fever, headache, fatigue, dehydration, anorexia, sudoresis, arthralgia and muscle pain. The condition evolves over a three to four-week period. Vulvar lesions and transmission within families have also been described. Molecular studies have shown that the poxviruses identified are genetically related to vaccinia virus samples that were used in vaccination campaigns in the past. Clinical specimens from 80 human infections were studied in the laboratory, and orthopoxvirus infections were confirmed in 68

  20. 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

    Science.gov (United States)

    Baxby, Derrick

    1972-01-01

    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 PMID:4624399

  1. [EVALUATION OF THE HUMAN SENSITIVITY TO SMALLPOX VIRUS BY THE PRIMARY CULTURES OF THE MONOCYTE-MACROPHAGES].

    Science.gov (United States)

    Zamedyanskaya, A S; Titova, K A; Sergeev, Al A; Kabanov, A S; Bulychev, L E; Sergeev, Ar A; Galakhova, D O; Nesterov, A E; Nosareva, O V; Shishkina, L N; Taranov, O S; Omigov, V V; Agafonov, A P; Sergeev, A N

    2016-01-01

    Studies of the primary cultures of granulocytes, mononuclear, and monocyte-macrophage cells derived from human blood were performed using variola virus (VARV) in the doses of 0.001-0.021 PFU/cell (plaques-forming units per cell). Positive dynamics of the virus accumulation was observed only in the monocyte-macrophages with maximum values of virus concentration (5.0-5.5 Ig PFU/ml) mainly within six days after the infection. The fact of VARV replication in the monocyte-macrophages was confirmed by the data of electron microscopy. At the same time, virus vaccines when tested in doses 3.3 and 4.2 Ig PFU/ml did not show the ability to reproduce in these human cells. The people sensitivity to VARV as assessed from the data obtained on human monocyte-macrophages corresponded to -1 PFU (taking into account the smooth interaction of the virus in the body to the cells of this type), which is consistent to previously found theoretical data on the virus sensitivity. The human susceptibility to VARV assessed experimentally can be used to predict the adequacy of developed smallpox models (in vivo) based on susceptible animals. This is necessary for reliable assessment of the efficiency of development of drugs for treatment and prophylaxis of the smallpox.

  2. Vaccinia DNA ligase complements Saccharomyces cerevisiae cdc9, localizes in cytoplasmic factories and affects virulence and virus sensitivity to DNA damaging agents.

    OpenAIRE

    Kerr, S M; Johnston, L H; Odell, M; Duncan, S A; Law, K M; Smith, G L

    1991-01-01

    The functional compatibility of vaccinia virus DNA ligase with eukaryotic counterparts was demonstrated by its ability to complement Saccharomyces cerevisiae cdc9. The vaccinia DNA ligase is a 63 kDa protein expressed early during infection that is non-essential for virus DNA replication and recombination in cultured cells. This implies complementation by a mammalian DNA ligase, yet no obvious recruitment of host DNA ligase I from the nucleus to the cytoplasm was observed during infection. An...

  3. Heterogeneity in the A33 Protein Impacts the Cross-Protective Efficacy of a Candidate Smallpox DNA Vaccine

    Science.gov (United States)

    2008-01-01

    1997). By 24-h post-transfection, COS cells were labeled with 35S-methionine/ cystine for 4 h and lysed using 4% zwit- tergent buffer (4% zwittergent...3753–3762. Roper, R.L., Wolffe, E.J., Weisberg, A., Moss, B., 1998. The envelope protein encoded by the A33R gene is required for formation of actin...Smith, G.L., Way, M., 1999. Interactions be- tween vaccinia virus IEV membrane proteins and their roles in IEV assembly and actin tail formation . J

  4. The Effect of Smallpox and Bacillus Calmette-Guérin Vaccination on the Risk of Human Immunodeficiency Virus-1 Infection in Guinea-Bissau and Denmark

    DEFF Research Database (Denmark)

    Rieckmann, Andreas; Villumsen, Marie; Jensen, Mette Lundsby

    2017-01-01

    BACKGROUND: The live smallpox and Bacillus Calmette-Guérin (BCG) vaccinations have been associated with better adult survival in both Guinea-Bissau and Denmark. In Guinea-Bissau, human immunodeficiency virus (HIV)-1 became an important cause of death after smallpox vaccination was phased out...... globally in 1980. We hypothesised that smallpox and BCG vaccinations were associated with a lower prevalence of HIV-1 infection, and we tested this hypothesis in both Guinea-Bissau and Denmark. METHODS: We conducted 2 studies: (1) a cross-sectional study of HIV infection and vaccination scars in Guinea...... Calmette-Guérin and/or smallpox vaccination compared with neither of these vaccines was associated with an adjusted odds ratio (aOR) for HIV-1 of 0.62 (95% confidence interval [CI], 0.36-1.07) in Guinea-Bissau and 0.70 (95% CI, 0.43-1.15) in Denmark. We combined the results from both settings in a meta...

  5. Sensitivity to ultraviolet radiation of Lassa, vaccinia, and Ebola viruses dried on surfaces.

    Science.gov (United States)

    Sagripanti, Jose-Luis; Lytle, C David

    2011-03-01

    Germicidal UV (also known as UVC) provides a means to decontaminate infected environments as well as a measure of viral sensitivity to sunlight. The present study determined UVC inactivation slopes (and derived D(37) values) of viruses dried onto nonporous (glass) surfaces. The data obtained indicate that the UV resistance of Lassa virus is higher than that of Ebola virus. The UV sensitivity of vaccinia virus (a surrogate for variola virus) appeared intermediate between that of the two virulent viruses studied. In addition, the three viruses dried on surfaces showed a relatively small but significant population of virions (from 3 to 10 % of virus in the inoculum) that appeared substantially more protected by their environment from the effect of UV than the majority of virions tested. The findings reported in this study should assist in estimating the threat posed by the persistence of virus in environments contaminated during epidemics or after an accidental or intentional release.

  6. Myxoma and vaccinia viruses exploit different mechanisms to enter and infect human cancer cells.

    Science.gov (United States)

    Villa, Nancy Y; Bartee, Eric; Mohamed, Mohamed R; Rahman, Masmudur M; Barrett, John W; McFadden, Grant

    2010-06-05

    Myxoma (MYXV) and vaccinia (VACV) viruses have recently emerged as potential oncolytic agents that can infect and kill different human cancer cells. Although both are structurally similar, it is unknown whether the pathway(s) used by these poxviruses to enter and cause oncolysis in cancer cells are mechanistically similar. Here, we compared the entry of MYXV and VACV-WR into various human cancer cells and observed significant differences: 1--low-pH treatment accelerates fusion-mediated entry of VACV but not MYXV, 2--the tyrosine kinase inhibitor genistein inhibits entry of VACV, but not MYXV, 3--knockdown of PAK1 revealed that it is required for a late stage event downstream of MYXV entry into cancer cells, whereas PAK1 is required for VACV entry into the same target cells. These results suggest that VACV and MYXV exploit different mechanisms to enter into human cancer cells, thus providing some rationale for their divergent cancer cell tropisms.

  7. Dynamic monitoring of membrane nanotubes formation induced by vaccinia virus on a high throughput microfluidic chip

    Science.gov (United States)

    Xiao, Min; Xu, Na; Wang, Cheng; Pang, Dai-Wen; Zhang, Zhi-Ling

    2017-03-01

    Membrane nanotubes (MNTs) are physical connections for intercellular communication and induced by various viruses. However, the formation of vaccinia virus (VACV)-induced MNTs has never been studied. In this report, VACV-induced MNTs formation process was monitored on a microfluidic chip equipped with a series of side chambers, which protected MNTs from fluidic shear stress. MNTs were formed between susceptible cells and be facilitated by VACV infection through three patterns. The formed MNTs varied with cell migration and virus concentration. The length of MNTs was positively correlated with the distance of cell migration. With increasing virus titer, the peak value of the ratio of MNT-carried cell appeared earlier. The immunofluorescence assay indicated that the rearrangement of actin fibers induced by VACV infection may lead to the formation of MNTs. This study presents evidence for the formation of MNTs induced by virus and helps us to understand the relationship between pathogens and MNTs.

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

    Science.gov (United States)

    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

    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. Copyright © 2012 Elsevier B.V. All rights reserved.

  9. Vaccinia complement control protein: Multi-functional protein and a potential wonder drug

    Indian Academy of Sciences (India)

    Purushottam Jha; Girish J Kotwal

    2003-04-01

    Vaccinia virus complement control protein (VCP) was one of the first viral molecules demonstrated to have a role in blocking complement and hence in the evasion of host defense. Structurally it is very similar to the human C4b-BP and the other members of complement control protein. Functionally it is most similar to the CR1 protein. VCP blocks both major pathways of complement activation. The crystal structure of VCP was determined a little over a year ago and it is the only known structure of an intact and complete complement control protein. In addition to binding complement, VCP also binds to heparin. These two binding abilities can take place simultaneously and contribute to its many function and to its potential use in several inflammatory diseases, e.g. Alzheimer’s disease (AD), CNS injury, xenotransplantation, etc. making it a truly fascinating molecule and potential drug.

  10. Vaccinia virus Transmission through Experimentally Contaminated Milk Using a Murine Model.

    Science.gov (United States)

    Rehfeld, Izabelle Silva; Guedes, Maria Isabel Maldonado Coelho; Fraiha, Ana Luiza Soares; Costa, Aristóteles Gomes; Matos, Ana Carolina Diniz; Fiúza, Aparecida Tatiane Lino; Lobato, Zélia Inês Portela

    2015-01-01

    Bovine vaccinia (BV) is a zoonosis caused by Vaccinia virus (VACV), which affects dairy cattle and humans. Previous studies have detected the presence of viable virus particles in bovine milk samples naturally and experimentally contaminated with VACV. However, it is not known whether milk contaminated with VACV could be a route of viral transmission. However, anti-Orthopoxvirus antibodies were detected in humans from BV endemic areas, whom had no contact with affected cows, which suggest that other VACV transmission routes are possible, such as consumption of contaminated milk and dairy products. Therefore, it is important to study the possibility of VACV transmission by contaminated milk. This study aimed to examine VACV transmission, pathogenesis and shedding in mice orally inoculated with experimentally contaminated milk. Thirty mice were orally inoculated with milk containing 107 PFU/ml of VACV, and ten mice were orally inoculated with uncontaminated milk. Clinical examinations were performed for 30 consecutive days, and fecal samples and oral swabs (OSs) were collected every other day. Mice were euthanized on predetermined days, and tissue and blood samples were collected. Nested-PCR, plaque reduction neutralization test (PRNT), viral isolation, histopathology, and immunohistochemistry (IHC) methods were performed on the collected samples. No clinical changes were observed in the animals. Viral DNA was detected in feces, blood, OSs and tissues, at least in one of the times tested. The lungs displayed moderate to severe interstitial lymphohistiocytic infiltrates, and only the heart, tonsils, tongue, and stomach did not show immunostaining at the IHC analysis. Neutralizing antibodies were detected at the 20th and 30th days post infection in 50% of infected mice. The results revealed that VACV contaminated milk could be a route of viral transmission in mice experimentally infected, showing systemic distribution and shedding through feces and oral mucosa, albeit

  11. Modulation of gene expression in a human cell line caused by poliovirus, vaccinia virus and interferon

    Directory of Open Access Journals (Sweden)

    Hoddevik Gunnar

    2007-03-01

    Full Text Available Abstract Background The project was initiated to describe the response of a human embryonic fibroblast cell line to the replication of two different viruses, and, more specifically, to look for candidate genes involved in viral defense. For this purpose, the cells were synchronously infected with poliovirus in the absence or presence of interferon-alpha, or with vaccinia virus, a virus that is not inhibited by interferon. By comparing the changes in transcriptosome due to these different challenges, it should be possible to suggest genes that might be involved in defense. Results The viral titers were sufficient to yield productive infection in a majority of the cells. The cells were harvested in triplicate at various time-points, and the transcriptosome compared with mock infected cells using oligo-based, global 35 k microarrays. While there was very limited similarities in the response to the different viruses, a large proportion of the genes up-regulated by interferon-alpha were also up-regulated by poliovirus. Interferon-alpha inhibited poliovirus replication, but there were no signs of any interferons being induced by poliovirus. The observations suggest that the cells do launch an antiviral response to poliovirus in the absence of interferon. Analyses of the data led to a list of candidate antiviral genes. Functional information was limited, or absent, for most of the candidate genes. Conclusion The data are relevant for our understanding of how the cells respond to poliovirus and vaccinia virus infection. More annotations, and more microarray studies with related viruses, are required in order to narrow the list of putative defence-related genes.

  12. Deletion of Fifteen Open Reading Frames from Modified Vaccinia Virus Ankara Fails to Improve Immunogenicity.

    Directory of Open Access Journals (Sweden)

    Naif Khalaf Alharbi

    Full Text Available Modified vaccinia virus Ankara (MVA is a highly attenuated strain of vaccinia virus, which has been used as a recombinant vaccine vector in many vaccine development programmes. The loss of many immunosuppressive and host-range genes resulted in a safe and immunogenic vaccine vector. However it still retains some immunomodulatory genes that may reduce MVA immunogenicity. Earlier reports demonstrated that the deletion of the A41L, B15R, C6L, or C12L open reading frames (ORFs enhanced cellular immune responses in recombinant MVA (rMVA by up to 2-fold. However, previously, we showed that deletion of the C12L, A44L, A46R, B7R, or B15R ORFs from rMVA, using MVA-BAC recombineering technology, did not enhance rMVA immunogenicity at either peak or memory cellular immune responses. Here, we extend our previous study to examine the effect of deleting clusters of genes on rMVA cellular immunogenicity. Two clusters of fifteen genes were deleted in one rMVA mutant that encodes either the 85A antigen of Mycobacterium tuberculosis or an immunodominant H2-Kd-restricted murine malaria epitope (pb9. The deletion mutants were tested in prime only or prime and boost vaccination regimens. The responses showed no improved peak or memory CD8+ T cell frequencies. Our results suggest that the reported small increases in MVA deletion mutants could not be replicated with different antigens, or epitopes. Therefore, the gene deletion strategy may not be taken as a generic approach for improving the immunogenicity of MVA-based vaccines, and should be carefully assessed for every individual recombinant antigen.

  13. Modulation of the myxoma virus plaque phenotype by vaccinia virus protein F11.

    Science.gov (United States)

    Irwin, Chad R; Evans, David H

    2012-07-01

    Vaccinia virus (VACV) produces large plaques consisting of a rapidly expanding ring of infected cells surrounding a lytic core, whereas myxoma virus (MYXV) produces small plaques that resemble a focus of transformed cells. This is odd, because bioinformatics suggests that MYXV carries homologs of nearly all of the genes regulating Orthopoxvirus attachment, entry, and exit. So why does MYXV produce foci? One notable difference is that MYXV-infected cells produce few of the actin microfilaments that promote VACV exit and spread. This suggested that although MYXV carries homologs of the required genes (A33R, A34R, A36R, and B5R), they are dysfunctional. To test this, we produced MYXV recombinants expressing these genes, but we could not enhance actin projectile formation even in cells expressing all four VACV proteins. Another notable difference between these viruses is that MYXV lacks a homolog of the F11L gene. F11 inhibits the RhoA-mDia signaling that maintains the integrity of the cortical actin layer. We constructed an MYXV strain encoding F11L and observed that, unlike wild-type MYXV, the recombinant virus disrupted actin stress fibers and produced plaques up to 4-fold larger than those of controls, and these plaques expanded ∼6-fold faster. These viruses also grew to higher titers in multistep growth conditions, produced higher levels of actin projectiles, and promoted infected cell movement, although neither process was to the extent of that observed in VACV-infected cells. Thus, one reason for why MYXV produces small plaques is that it cannot spread via actin filaments, although the reason for this deficiency remains obscure. A second reason is that leporipoxviruses lack vaccinia's capacity to disrupt cortical actin.

  14. Recombination-mediated genetic engineering of a bacterial artificial chromosome clone of modified vaccinia virus Ankara (MVA)

    DEFF Research Database (Denmark)

    Cottingham, Matthew G; Andersen, Rikke F; Spencer, Alexandra J

    2008-01-01

    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, PNAS99 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......-2006). In addition, we found a higher frequency of triple-positive IFN-gamma, TNF-alpha 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 Gal...

  15. Vaccinia virus-mediated melanin production allows MR and optoacoustic deep tissue imaging and laser-induced thermotherapy of cancer.

    Science.gov (United States)

    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

    2013-02-26

    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.

  16. Gel-eletroforese no diagnóstico da varíola Gel-electrophoresis in the smallpox diagnosis

    Directory of Open Access Journals (Sweden)

    Julio A. Mesquita

    1972-01-01

    Full Text Available O emprego de gel-eletroforese no diagnóstico da varíola, demonstrou ser ao menos trinta vezes (30X mais sensível que o teste de agar-gel, nas condições descritas (tabela I. Doze (12 espécimes, cujos testes convencionais de inoculação em ovos embrionados e de agar-gel resultaram positivos, foram testados em suas diluições originais congeladas por mais de um ano, sendo seis deles revelados por gel-eletroforese enquanto nenhum o foi por agar-gel (tabela II. Trinta e três (33 amostras isoladas no laboratório, foram testadas com material colhido de membrana cório-alantóica da primeira inoculação para o diagnóstico, conservado em glicerina 50%, resultando 15 positivas em gel-eletroforese e apenas 3 em agar-gel (tabela II. Os últimos 60 espécimes recebidos para diagnóstico, através a Campanha de Erradicação da Varíola, também resultaram negativos em gel-eletroforese, que não mostrou falsos-positivos nas condições descritas.The test of gel-electrophoresis applied to the pox virus group showed to be at least thirth times (30X more sensitive than agar-gel test on the described conditions (Table I. Twelve specimens, which were positives form Smallpox in the conventional tests of egg inoculation and agar-gel difusion test, have been screened in their original dilutions frozen for more than 1 year and six of them were still detectable by gel-eletrophoresis, while by agar-gel test any of them was positive (Table II. Thirty three Smallpox isolates have been tested with material from first egg inoculation (chorioallantoic membranes which have been stored in glycerin 50%, at - 15ºC. Fifteen of them were still positive by gel-electrophoresis and only 3 by agar-gel (Table II. The last 60 specimens received for diagnosis from Smallpox Erradication Campaign (CEV, were negatives by both tests. The gel-electrophoresis, did not show false-positives on described conditions.

  17. Cross-protective immunity against multiple influenza virus subtypes by a novel modified vaccinia Ankara (MVA) vectored vaccine in mice.

    Science.gov (United States)

    Brewoo, Joseph N; Powell, Tim D; Jones, Jeremy C; Gundlach, Nancy A; Young, Ginger R; Chu, Haiyan; Das, Subash C; Partidos, Charalambos D; Stinchcomb, Dan T; Osorio, Jorge E

    2013-04-03

    Development of an influenza vaccine that provides cross-protective immunity remains a challenge. Candidate vaccines based on a recombinant modified vaccinia Ankara (MVA) viral vector expressing antigens from influenza (MVA/Flu) viruses were constructed. A vaccine candidate, designated MVA/HA1/C13L/NP, that expresses the hemagglutinin from pandemic H1N1 (A/California/04/09) and the nucleoprotein (NP) from highly pathogenic H5N1 (A/Vietnam/1203/04) fused to a secretory signal sequence from vaccinia virus was highly protective. The vaccine elicited strong antibody titers to homologous H1N1 viruses while cross-reactive antibodies to heterologous viruses were not detectable. In mice, this MVA/HA1/C13L/NP vaccine conferred complete protection against lethal challenge with A/Vietnam/1203/04 (H5N1), A/Norway/3487-2/09 (pandemic H1N1) or A/Influenza/Puerto Rico/8/34 (seasonal H1N1) and partial protection (57.1%) against challenge with seasonal H3N2 virus (A/Aichi/68). The protective efficacy of the vaccine was not affected by pre-existing immunity to vaccinia. Our findings highlight MVA as suitable vector to express multiple influenza antigens that could afford broad cross-protective immunity against multiple subtypes of influenza virus.

  18. [Modified vaccinia virus ankara (MVA)--development as recombinant vaccine and prospects for use in veterinary medicine].

    Science.gov (United States)

    Volz, Asisa; Fux, Robert; Langenmayer, Martin C; Sutter, Gerd

    2015-01-01

    Poxviruses as expression vectors are widely used in medical research for the development of recombinant vaccines and molecular therapies. Here we review recent accomplishments in vaccine research using recombinant modified vaccinia virus ankara (MVA). MVA is a highly attenuated vaccinia virus strain that originated from serial tissue culture passage in chicken embryo fibroblasts more than 40 years ago. Growth adaptation to avian host cells caused deletions and mutations in the viral genome affecting about 15% of the original genetic information. In consequence, MVA is replication-deficient in cells of mammalian origin and fails to produce many of the virulence factors encoded by conventional vaccinia virus. Because of its safety for the general environment MVA can be handled under conditions of biosafety level one. Non-replicating MVA can enter any target cell and activate its molecular life cycle to express all classes of viral and recombinant genes. Therefore, recombinant MVA have been established as an extremely safe and efficient vector system for vaccine development in medical research. By now, various recombinant MVA vaccines have been found safe and immunogenic when used for phase I/II clinical testing in humans, and suitable for industrial scale production following good practice of manufacturing. Thus, there is an obvious usefulness of recombinant MVA vaccines for novel prophylactic and therapeutic approaches also in veterinary medicine. Results from first studies in companion and farm animals are highly promising.

  19. Extracts from rabbit skin inflamed by the vaccinia virus attenuate bupivacaine-induced spinal neurotoxicity in pregnant rats

    Institute of Scientific and Technical Information of China (English)

    Rui Cui; Shiyuan Xu; Liang Wang; Hongyi Lei; Qingxiang Cai; Hongfei Zhang; Dongmei Wang

    2013-01-01

    Extracts from rabbit skin inflamed by the vaccinia virus can relieve pain and promote repair of nerve injury. The present study intraperitoneally injected extracts from rabbit skin inflamed by the vaccinia virus for 3 and 4 days prior to and following intrathecal injection of bupivacaine into pregnant rats. The pain threshold test after bupivacaine injection showed that the maximum possible effect of tail-flick latency peaked 1 day after intrathecal injection of bupivacaine in the extract-pretreatment group, and gradually decreased, while the maximum possible effect in the bupivacaine group continued to increase after intrathecal injection of bupivacaine. Histological observation showed that after 4 days of intrathecal injection of bupivacaine, the number of shrunken, vacuolated, apoptotic and caspase-9-positive cells in the dorsal root ganglion in the extract-pretreatment group was significantly reduced compared with the bupivacaine group. These findings indicate that extracts from rabbit skin inflamed by the vaccinia virus can attenuate neurotoxicity induced by intrathecal injection of bupivacaine in pregnant rats, possibly by inhibiting caspase-9 protein expression and suppressing nerve cell apoptosis.

  20. Evaluation of a new recombinant oncolytic vaccinia virus strain GLV-5b451 for feline mammary carcinoma therapy.

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    Marion Adelfinger

    Full Text Available Virotherapy on the basis of oncolytic vaccinia virus (VACV infection is a promising approach for cancer therapy. In this study we describe the establishment of a new preclinical model of feline mammary carcinoma (FMC using a recently established cancer cell line, DT09/06. In addition, we evaluated a recombinant vaccinia virus strain, GLV-5b451, expressing the anti-vascular endothelial growth factor (VEGF single-chain antibody (scAb GLAF-2 as an oncolytic agent against FMC. Cell culture data demonstrate that GLV-5b451 virus efficiently infected, replicated in and destroyed DT09/06 cancer cells. In the selected xenografts of FMC, a single systemic administration of GLV-5b451 led to significant inhibition of tumor growth in comparison to untreated tumor-bearing mice. Furthermore, tumor-specific virus infection led to overproduction of functional scAb GLAF-2, which caused drastic reduction of intratumoral VEGF levels and inhibition of angiogenesis. In summary, here we have shown, for the first time, that the vaccinia virus strains and especially GLV-5b451 have great potential for effective treatment of FMC in animal model.

  1. Efficient colonization and therapy of human hepatocellular carcinoma (HCC using the oncolytic vaccinia virus strain GLV-1h68.

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    Ivaylo Gentschev

    Full Text Available Virotherapy using oncolytic vaccinia virus strains is one of the most promising new strategies for cancer therapy. In this study, we analyzed for the first time the therapeutic efficacy of the oncolytic vaccinia virus GLV-1h68 in two human hepatocellular carcinoma cell lines HuH7 and PLC/PRF/5 (PLC in cell culture and in tumor xenograft models. By viral proliferation assays and cell survival tests, we demonstrated that GLV-1h68 efficiently colonized, replicated in, and did lyse these cancer cells in culture. Experiments with HuH7 and PLC xenografts have revealed that a single intravenous injection (i.v. of mice with GLV-1h68 resulted in a significant reduction of primary tumor sizes compared to uninjected controls. In addition, replication of GLV-1h68 in tumor cells led to strong inflammatory and oncolytic effects resulting in intense infiltration of MHC class II-positive cells like neutrophils, macrophages, B cells and dendritic cells and in up-regulation of 13 pro-inflammatory cytokines. Furthermore, GLV-1h68 infection of PLC tumors inhibited the formation of hemorrhagic structures which occur naturally in PLC tumors. Interestingly, we found a strongly reduced vascular density in infected PLC tumors only, but not in the non-hemorrhagic HuH7 tumor model. These data demonstrate that the GLV-1h68 vaccinia virus may have an enormous potential for treatment of human hepatocellular carcinoma in man.

  2. Induction of neutralizing antibodies by varicella-zoster virus gpII glycoprotein expressed from recombinant vaccinia virus.

    Science.gov (United States)

    Massaer, M; Haumont, M; Place, M; Bollen, A; Jacobs, P

    1993-03-01

    The gpII glycoprotein of varicella-zoster virus (VZV) was produced in CV1 cells via vaccinia virus recombinants. Two different DNA constructs were expressed: the first one encodes the complete gpII protein (gpII s+a+) and the second a truncated species lacking the membrane anchorage domain (gpII s+a-). To achieve expression both coding sequences had to be engineered at the 5' end by substituting the unusually short (24 bp) natural signal sequence by a more conventional one encoding 29 amino acids. Recombinant gpII proteins were detected in vaccinia virus-infected cells by ELISA and immunoprecipitation. Both forms of recombinant gpII proteins were produced as glycosylated single-chain molecules of respectively 110K and 90K. Upon reduction these were only partially converted into subunits. A rabbit infected with the vaccinia virus recombinant expressing the complete gpII produced antibodies which recognized VZV antigens and neutralized VZV infectivity in vitro, independent of complement.

  3. A PCR-based method for manipulation of the vaccinia virus genome that eliminates the need for cloning.

    Science.gov (United States)

    Turner, P C; Moyer, R W

    1992-11-01

    A general method is described for altering specific genes of vaccinia virus (VV). We demonstrate and evaluate the procedure by gene inactivation, using a dominant selectable marker in conjunction with recombinant polymerase chain reaction (PCR). Primers based on the sequence of the target gene enable amplification of flanking arms and their subsequent attachment to the gpt cassette that confers resistance to mycophenolic acid. Linear PCR constructs are transfected into cells infected with wild-type vaccinia virus. Mutant viruses with gpt inserted into the target gene by homologous recombination are then selected by growth in the presence of MPA. This technique was applied to the vaccinia virus thymidine kinase gene and compared to the traditional method of constructing gpt-containing plasmids by cloning. The PCR scheme was found to be highly efficient and could theoretically be used to insert any foreign DNA element into any nonessential target gene for which partial or complete sequence information is available. The procedure can potentially be used for a wide variety of genetic modifications, including the insertion of foreign genes, with poxviruses and other DNA viruses. Genomes of microorganisms, such as bacteria and yeast that can be transformed with linear DNA, are also candidates for manipulation by this methodology.

  4. Vergleich von rekombinanten Vaccinia- und DNA-Vektoren zur Tumorimmuntherapie im C57BL/6-Mausmodell

    Science.gov (United States)

    Johnen, Heiko

    2002-10-01

    In der vorliegenden Arbeit wurden Tumorimpfstoffe auf der Basis des Plasmid-Vektors pCI, modified vaccinia virus Ankara (MVA) und MVA-infizierten dendritischen Zellen entwickelt und durch Sequenzierung, Western blotting und durchflußzytometrische Analyse überprüft. Die in vivo Wirksamkeit der Vakzinen wurde in verschiedenen Tumormodellen in C57BL/6 Mäusen verglichen. Die auf dem eukaryotischen Expressionsvektor pCI basierende DNA-Vakzinierung induzierte einen sehr wirksamen, antigenspezifischen und langfristigen Schutz vor Muzin, CEA oder beta-Galactosidase exprimierenden Tumoren. Eine MVA-Vakzinierung bietet in den in dieser Arbeit durchgeführten Tumormodellen keinen signifikanten Schutz vor Muzin oder beta-Galactosidase exprimierenden Tumoren. Sowohl humane, als auch murine in vitro generierte dendritische Zellen lassen sich mit MVA – im Vergleich zu anderen viralen Vektoren – sehr gut infizieren. Die Expressionsrate der eingefügten Gene ist aber gering im Vergleich zur Expression in permissiven Wirtszellen des Virus (embryonale Hühnerfibroblasten). Es konnte gezeigt werden, daß eine MVA-Infektion dendritischer Zellen ähnliche Auswirkungen auf den Reifezustand humaner und muriner dendritischer Zellen hat, wie eine Infektion mit replikationskompetenten Vakzinia-Stämmen, und außerdem die Hochregulation von CD40 während der terminalen Reifung von murinen dendritischen Zellen inhibiert wird. Die während der langfristigen in vitro Kultur auf CEF-Zellen entstandenen Deletionen im MVA Genom führten zu einer starken Attenuierung und dem Verlust einiger Gene, die immunmodulatorische Proteine kodieren, jedoch nicht zu einer Verminderung des zytopathischen Effekts in dendritischen Zellen. Die geringe Expressionsrate und die beobachtete Inhibition der Expression kostimulatorischer Moleküle auf dendritischen Zellen kann für eine wenig effektive Induktion einer Immunantwort in MVA vakzinierten Tieren durch cross priming oder die direkte Infektion antigenpr

  5. Pharmacokinetic and pharmacodynamic modeling to determine the dose of ST-246 to protect against smallpox in humans.

    Science.gov (United States)

    Leeds, Janet M; Fenneteau, Frederique; Gosselin, Nathalie H; Mouksassi, Mohamad-Samer; Kassir, Nastya; Marier, J F; Chen, Yali; Grosenbach, Doug; Frimm, Annie E; Honeychurch, Kady M; Chinsangaram, Jarasvech; Tyavanagimatt, Shanthakumar R; Hruby, Dennis E; Jordan, Robert

    2013-03-01

    Although smallpox has been eradicated, the United States government considers it a "material threat" and has funded the discovery and development of potential therapeutic compounds. As reported here, the human efficacious dose for one of these compounds, ST-246, was determined using efficacy studies in nonhuman primates (NHPs), together with pharmacokinetic and pharmacodynamic analysis that predicted the appropriate dose and exposure levels to provide therapeutic benefit in humans. The efficacy analysis combined the data from studies conducted at three separate facilities that evaluated treatment following infection with a closely related virus, monkeypox virus (MPXV), in a total of 96 NHPs. The effect of infection on ST-246 pharmacokinetics in NHPs was applied to humans using population pharmacokinetic models. Exposure at the selected human dose of 600 mg is more than 4-fold higher than the lowest efficacious dose in NHPs and is predicted to provide protection to more than 95% of the population.

  6. From foe to friend: geographical and environmental factors and the control and eradication of smallpox in India.

    Science.gov (United States)

    Bhattacharya, Sanjoy

    2003-01-01

    Due to the highly visible nature of the disease, smallpox received a lot of attention from the colonial and independent Indian governments. An assessment of the changing official views about the impact of geographical and environmental factors on modes of variola causation and control presents insights into themes that are generally ignored in the existing historiography. Rather than being synchronised efforts, imposed top-down, provincial level officials in charge of running vaccination programmes were able to retain a great degree of autonomy and shape the nature of local immunisation drive. As a result, vaccination work in the country was often disjointed, marked by official disagreements about the usefulness of certain strategies and technological inputs, with these trends being most noticeable in rural areas. Thus, this study seeks to highlight the importance of recognising--and studying--the fractured nature of medical and public health administration in the South Asian sub-continent.

  7. Reverse genetics of SARS-related coronavirus using vaccinia virus-based recombination.

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    Sjoerd H E van den Worm

    Full Text Available 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.

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

    Science.gov (United States)

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

    2012-01-01

    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. PMID:22412934

  9. 42 CFR 102.50 - Medical records necessary to establish that a covered injury was sustained.

    Science.gov (United States)

    2010-10-01

    ... date of the smallpox vaccination or exposure to vaccinia; and (2) All inpatient hospital medical records, including the admission history and physical examination, the discharge summary, all physician... of the smallpox vaccination or exposure to vaccinia. (b) A requester may submit additional...

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

    Energy Technology Data Exchange (ETDEWEB)

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

    1986-07-15

    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.

  11. Study of Vaccinia and Cowpox viruses' replication in Rac1-N17 dominant-negative cells

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    Ana Paula Carneiro Salgado

    2013-08-01

    Full Text Available Interfering with cellular signal transduction pathways is a common strategy used by many viruses to create a propitious intracellular environment for an efficient replication. Our group has been studying cellular signalling pathways activated by the orthopoxviruses Vaccinia (VACV and Cowpox (CPXV and their significance to viral replication. In the present study our aim was to investigate whether the GTPase Rac1 was an upstream signal that led to the activation of MEK/ERK1/2, JNK1/2 or Akt pathways upon VACV or CPXV' infections. Therefore, we generated stable murine fibroblasts exhibiting negative dominance to Rac1-N17 to evaluate viral growth and the phosphorylation status of ERK1/2, JNK1/2 and Akt. Our results demonstrated that VACV replication, but not CPXV, was affected in dominant-negative (DN Rac1-N17 cell lines in which viral yield was reduced in about 10-fold. Viral late gene expression, but not early, was also reduced. Furthermore, our data showed that Akt phosphorylation was diminished upon VACV infection in DN Rac1-N17 cells, suggesting that Rac1 participates in the phosphoinositide-3 kinase pathway leading to the activation of Akt. In conclusion, our results indicate that while Rac1 indeed plays a role in VACV biology, perhaps another GTPase may be involved in CPXV replication.

  12. Potential effect of prior raccoonpox virus infection in raccoons on vaccinia-based rabies immunization

    Science.gov (United States)

    Root, J Jeffrey; McLean, Robert G; Slate, Dennis; MacCarthy, Kathleen A; Osorio, Jorge E

    2008-01-01

    Background The USDA, Wildlife Services cooperative oral rabies vaccination (ORV) program uses a live vaccinia virus-vectored (genus Orthopoxvirus) vaccine, Raboral V-RG® (V-RG), to vaccinate specific wildlife species against rabies virus in several regions of the U.S. Several naturally occurring orthopoxviruses have been found in North America, including one isolated from asymptomatic raccoons (Procyon lotor). The effect of naturally occurring antibodies to orthopoxviruses on successful V-RG vaccination in raccoons is the focus of this study. Results Overall, raccoons pre-immunized (n = 10) with a recombinant raccoonpox virus vaccine (RCN-F1) responded to vaccination with V-RG with lower rabies virus neutralizing antibody (VNA) titers than those which were not pre-immunized (n = 10) and some failed to seroconvert for rabies VNA to detectable levels. Conclusion These results suggest that the success of some ORV campaigns may be hindered where raccoonpox virus or possibly other orthopoxvirus antibodies are common in wildlife species targeted for ORV. If these areas are identified, different vaccination strategies may be warranted. PMID:18834520

  13. Intradermal delivery of recombinant vaccinia virus vector DIs induces gut-mucosal immunity.

    Science.gov (United States)

    Yoshino, N; Kanekiyo, M; Hagiwara, Y; Okamura, T; Someya, K; Matsuo, K; Ami, Y; Sato, S; Yamamoto, N; Honda, M

    2010-08-01

    Antigen-specific mucosal immunity is generally induced by the stimulation of inductive mucosal sites. In this study, we found that the replication-deficient vaccinia virus vector, DIs, generates antigen-specific mucosal immunity and systemic responses. Following intradermal injection of recombinant DIs expressing simian immunodeficiency virus gag (rDIsSIVgag), we observed increased levels of SIV p27-specific IgA and IgG antibodies in faecal extracts and plasma samples, and antibody-forming cells in the intestinal mucosa and spleen of C57BL/6 mice. Antibodies against p27 were not detected in nasal washes, saliva, and vaginal washes. The enhanced mucosal and systemic immunity persisted for 1 year of observation. Induction of Gag-specific IFN-gamma spot-forming CD8(+) T cells in the spleen, small intestinal intraepithelial lymphocytes, and submandibular lymph nodes was observed in the intradermally injected mice. Heat-inactivated rDIsSIVgag rarely induced antigen-specific humoral and T-helper immunity. Moreover, rDIsSIVgag was detected in MHC class II IA antigen-positive (IA(+)) cells at the injection site. Consequently, intradermal delivery of rDIs effectively induces antigen-specific humoral and cellular immunity in gut-mucosal tissues of mice. Our data suggest that intradermal injection of an rDIs vaccine may be useful against mucosally transmitted pathogens.

  14. Delivery of Echinococcus granulosus antigen EG95 to mice and sheep using recombinant vaccinia virus.

    Science.gov (United States)

    Dutton, S; Fleming, S B; Ueda, N; Heath, D D; Hibma, M H; Mercer, A A

    2012-06-01

    The tapeworm Echinococcus granulosus is the causative agent of hydatid disease and affects sheep, cattle, dogs and humans worldwide. It has a two-stage life cycle existing as worms in the gut of infected dogs (definitive host) and as cysts in herbivores and humans (intermediate host). The disease is debilitating and can be life threatening where the cysts interfere with organ function. Interruption of the hydatid life cycle in the intermediate host by vaccination may be a way to control the disease, and a protective oncosphere antigen EG95 has been shown to protect animals against challenge with E. granulosus eggs. We explored the use of recombinant vaccinia virus as a delivery vehicle for EG95. Mice and sheep were immunized with the recombinant vector, and the result monitored at the circulating antibody level. In addition, sera from immunized mice were assayed for the ability to kill E. granulosus oncospheres in vitro. Mice immunized once intranasally developed effective oncosphere-killing antibody by day 42 post-infection. Antibody responses and oncosphere killing were correlated and were significantly enhanced by boosting mice with either EG95 protein or recombinant vector. Sheep antibody responses to the recombinant vector or to EG95 protein mirrored those in mice.

  15. Antigen Gene Transfer to Human Plasmacytoid Dendritic Cells Using Recombinant Adenovirus and Vaccinia Virus Vectors

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    Hetty J. Bontkes

    2005-01-01

    Full Text Available Recombinant adenoviruses (RAd and recombinant vaccinia viruses (RVV expressing tumour-associated antigens (TAA are used as anti-tumour vaccines. It is important that these vaccines deliver the TAA to dendritic cells (DC for the induction of a strong immune response. Infection of myeloid DC (MDC with RAd alone is relatively inefficient but CD40 retargeting significantly increases transduction efficiency and DC maturation. Infection with RVV is efficient without DC maturation. Plasmacytoid dendritic cells (PDC play a role in the innate immune response to viral infections through the secretion of IFNα but may also play a role in specific T-cell induction. The aim of our study was to investigate whether PDC are better targets for RAd and RVV based vaccines. RAd alone hardly infected PDC (2% while CD40 retargeting did not improve transduction efficiency, but it did increase PDC maturation (25% CD83 positive cells. Accordingly, specific CTL activation by RAd infected PDC was limited (the number of IFNγ producing CTL was reduced by 75% compared to stimulation with peptide loaded PDC. RVV infected PDC specifically stimulated CTL but PDC were not activated. These Results indicate that PDC are not ideal targets for RAd and RVV based vaccines. However, PDC induced specific CTL activation after pulsing with recombinant protein, indicating that PDC can also cross-present antigens released from surrounding infected cells.

  16. DNA strand transfer catalyzed by vaccinia topoisomerase: ligation of DNAs containing a 3' mononucleotide overhang.

    Science.gov (United States)

    Cheng, C; Shuman, S

    2000-05-01

    The specificity of vaccinia topoisomerase for transesterification to DNA at the sequence 5'-CCCTT and its versatility in strand transfer have illuminated the recombinogenic properties of type IB topoisomerases and spawned topoisomerase-based strategies for DNA cloning. Here we characterize a pathway of topoisomerase-mediated DNA ligation in which enzyme bound covalently to a CCCTT end with an unpaired +1T nucleotide rapidly and efficiently joins the CCCTT strand to a duplex DNA containing a 3' A overhang. The joining reaction occurs with high efficiency, albeit slowly, to duplex DNAs containing 3' G, T or C overhangs. Strand transfer can be restricted to the correctly paired 3' A overhang by including 0.5 M NaCl in the ligation reaction mixture. The effects of base mismatches and increased ionic strength on the rates of 3' overhang ligation provide a quantitative picture of the relative contributions of +1 T:A base pairing and electrostatic interactions downstream of the scissile phosphate to the productive binding of an unlinked acceptor DNA to the active site. The results clarify the biochemistry underlying topoisomerase-cloning of PCR products with non-templated 3' overhangs.

  17. Increased ATP generation in the host cell is required for efficient vaccinia virus production

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    Hsu Che-Fang

    2009-09-01

    Full Text Available Abstract 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 μM 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.

  18. RAB1A promotes Vaccinia virus replication by facilitating the production of intracellular enveloped virions

    Energy Technology Data Exchange (ETDEWEB)

    Pechenick Jowers, Tali; Featherstone, Rebecca J.; Reynolds, Danielle K.; Brown, Helen K. [The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Roslin, Midlothian EH25 9RG, Scotland (United Kingdom); James, John; Prescott, Alan [Division of Cell Signalling and Immunology, College of Life Sciences, University of Dundee, Dundee DD1 5EH, Scotland (United Kingdom); Haga, Ismar R. [The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Roslin, Midlothian EH25 9RG, Scotland (United Kingdom); Beard, Philippa M., E-mail: pip.beard@roslin.ed.ac.uk [The Roslin Institute and Royal (Dick) School of Veterinary Studies, University of Edinburgh, Roslin, Midlothian EH25 9RG, Scotland (United Kingdom)

    2015-01-15

    Vaccinia virus (VACV) is a large double-stranded DNA virus with a complex cytoplasmic replication cycle that exploits numerous cellular proteins. This work characterises the role of a proviral cellular protein, the small GTPase RAB1A, in VACV replication. Using siRNA, we identified RAB1A as required for the production of extracellular enveloped virions (EEVs), but not intracellular mature virions (IMVs). Immunofluorescence and electron microscopy further refined the role of RAB1A as facilitating the wrapping of IMVs to become intracellular enveloped virions (IEVs). This is consistent with the known function of RAB1A in maintenance of ER to Golgi transport. VACV can therefore be added to the growing list of viruses which require RAB1A for optimal replication, highlighting this protein as a broadly proviral host factor. - Highlights: • Characterisation of the role of the small GTPase RAB1A in VACV replication. • RAB1A is not required for production of the primary virion form (IMV). • RAB1A is required for production of processed virion forms (IEVs, CEVs and EEVs). • Consistent with known role of RAB1A in ER to Golgi transport.

  19. Mapping vaccinia virus DNA replication origins at nucleotide level by deep sequencing.

    Science.gov (United States)

    Senkevich, Tatiana G; Bruno, Daniel; Martens, Craig; Porcella, Stephen F; Wolf, Yuri I; Moss, Bernard

    2015-09-01

    Poxviruses reproduce in the host cytoplasm and encode most or all of the enzymes and factors needed for expression and synthesis of their double-stranded DNA genomes. Nevertheless, the mode of poxvirus DNA replication and the nature and location of the replication origins remain unknown. A current but unsubstantiated model posits only leading strand synthesis starting at a nick near one covalently closed end of the genome and continuing around the other end to generate a concatemer that is subsequently resolved into unit genomes. The existence of specific origins has been questioned because any plasmid can replicate in cells infected by vaccinia virus (VACV), the prototype poxvirus. We applied directional deep sequencing of short single-stranded DNA fragments enriched for RNA-primed nascent strands isolated from the cytoplasm of VACV-infected cells to pinpoint replication origins. The origins were identified as the switching points of the fragment directions, which correspond to the transition from continuous to discontinuous DNA synthesis. Origins containing a prominent initiation point mapped to a sequence within the hairpin loop at one end of the VACV genome and to the same sequence within the concatemeric junction of replication intermediates. These findings support a model for poxvirus genome replication that involves leading and lagging strand synthesis and is consistent with the requirements for primase and ligase activities as well as earlier electron microscopic and biochemical studies implicating a replication origin at the end of the VACV genome.

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

    Science.gov (United States)

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

    2013-09-17

    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-γ (IFN-γ) 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-γ. 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-γ under engineered tetracycline-inducible promoters. Immunodeficient SCID mice inoculated with VACVs not expressing IFN-γ demonstrated severe weight loss, whereas those given VACVs expressing IFN-γ under constitutive VACV promoters showed no signs of infection. Most importantly, mice inoculated with a VACV expressing the IFN-γ 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-γ under a tightly controlled tetracycline-inducible VACV promoter for use in vaccines and oncolytic cancer therapies.

  1. High cytokine production and effective antitumor activity of a recombinant vaccinia virus encoding murine interleukin 12.

    Science.gov (United States)

    Meko, J B; Yim, J H; Tsung, K; Norton, J A

    1995-11-01

    We have constructed a recombinant vaccinia virus (recVV), vKT0334 mIL-12, containing the genes encoding the p35 and p40 subunits of murine interleukin-12 (mIL-12). In vitro experiments demonstrated that vKT0334 mIL-12 efficiently infected a variety of murine and human tumor cell lines and produced very high amounts (1.5 micrograms/10(6) cells/24 h) of biologically active mIL-12. Mice injected s.c. with 10(6) MCA 105 sarcoma cells, followed by injection at the same site with saline or a control recVV, vKT033, containing no mIL-12 genes, all developed progressively growing tumor, whereas 60% of animals injected with vKT0334 mIL-12 remained tumor free (P < 0.0005). Furthermore, tumor growth was significantly reduced in the remaining mice treated with vKT0334 mIL-12 that did develop tumor compared with mice treated with vKT033 (P < 0.03) or saline (P < 0.0001). We conclude that recVV expressing high levels of mIL-12 offers an effective in vivo method of cytokine gene delivery and expression in tumors with subsequent antitumor effect.

  2. Brazilian vaccinia virus strains show a classical orthopoxvirus in-fection course and cross-protection

    Institute of Scientific and Technical Information of China (English)

    Betania Paiva Drumond; Jonatas Santos Abraho; Zlia Ins Portela Lobato; Cludio Antonio Bonjardim; Paulo Csar Peregrino Ferreira; Erna Geessien Kroon

    2009-01-01

    Objectives:The purpose of this work was to study the infection course and cross-protection in mice after intra-dermal injection of Vaccinia virus (VACV ) strain Western Reserve and three Brazilian VACV strains:Araatuba,Muriaéand BeAn58058 isolated from cow,human and rodent,respectively.Methods:Balb /c mice were inoculated by footpad and back scarification and daily monitored regarding lesion development and weight loss.To check cross protection after intradermal VACV inoculation,mice were subsequently infected with different VACV strains and monitored to check lesion development.Serum neutralization assays were per-formed to check for the presence of antibodies against Orthopoxvirus.Results:After VACV intradermal inocu-lation the lesion development pattern was similar in mice infected with the different virus strains.By using the footpad scarification model,cross-protection among VACV strains was observed.Moreover,neutralizing anti-bodies against Orthopoxvirus were detected in sera from mice infected with all VACV strains.Conclusion:Al-though it was not possible to observe virulence differences among VACV strains isolated from cow,rodent and human using the murine model,this inoculation route showed to be an appropriated model to study lesions de-velopment since it mimics natural infections by VACV in nature.

  3. Generation of an attenuated Tiantan vaccinia virus by deletion of the ribonucleotide reductase large subunit.

    Science.gov (United States)

    Kan, Shifu; Jia, Peng; Sun, Lili; Hu, Ningning; Li, Chang; Lu, Huijun; Tian, Mingyao; Qi, Yanxin; Jin, Ningyi; Li, Xiao

    2014-09-01

    Attenuation of the virulence of vaccinia Tiantan virus (VTT) underlies the strategy adopted for mass vaccination campaigns. This strategy provides advantages of safety and efficacy over traditional vaccines and is aimed at minimization of adverse health effects. In this study, a mutant form of the virus, MVTT was derived from VTT by deletion of the ribonucleotide reductase large subunit (R1) (TI4L). Compared to wild-type parental (VTT) and revertant (VTT-rev) viruses, virulence of the mutant MVTT was reduced by 100-fold based on body weight reduction and by 3,200-fold based on determination of the intracranial 50% lethal infectious dose. However, the immunogenicity of MVTT was equivalent to that of the parental VTT. We also demonstrated that the TI4L gene is not required for efficient replication. These data support the conclusion that MVTT can be used as a replicating virus vector or as a platform for the development of vaccines against infectious diseases and for cancer therapy.

  4. Potential effect of prior raccoonpox virus infection in raccoons on vaccinia-based rabies immunization

    Directory of Open Access Journals (Sweden)

    MacCarthy Kathleen A

    2008-10-01

    Full Text Available Abstract Background The USDA, Wildlife Services cooperative oral rabies vaccination (ORV program uses a live vaccinia virus-vectored (genus Orthopoxvirus vaccine, Raboral V-RG® (V-RG, to vaccinate specific wildlife species against rabies virus in several regions of the U.S. Several naturally occurring orthopoxviruses have been found in North America, including one isolated from asymptomatic raccoons (Procyon lotor. The effect of naturally occurring antibodies to orthopoxviruses on successful V-RG vaccination in raccoons is the focus of this study. Results Overall, raccoons pre-immunized (n = 10 with a recombinant raccoonpox virus vaccine (RCN-F1 responded to vaccination with V-RG with lower rabies virus neutralizing antibody (VNA titers than those which were not pre-immunized (n = 10 and some failed to seroconvert for rabies VNA to detectable levels. Conclusion These results suggest that the success of some ORV campaigns may be hindered where raccoonpox virus or possibly other orthopoxvirus antibodies are common in wildlife species targeted for ORV. If these areas are identified, different vaccination strategies may be warranted.

  5. Vaccinia Virus Inhibits T Cell Receptor–Dependent Responses by Human γδ T Cells

    Science.gov (United States)

    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

    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 γδ T cells because of the role they play in immune control of this virus. Exposure to VV failed to trigger proliferative responses in γδ T cells from unprimed individuals, but it was an unexpected finding that VV blocked responses to model antigens by the Vγ2Vδ2 T cell subset. Infectious or ultraviolet light–inactivated VV inhibited proliferative Vγ2Vδ2 T cell responses to phosphoantigens and tumor cells, prevented cytolysis of Daudi B cells, and reduced cytokine production. Inhibiting Vγ2Vδ2 T cells may be a mechanism for evading host immunity and increasing VV virulence. Increased VV replication or expression in the absence of γδ T cell responses might contribute to its potency as a vaccine against poxvirus and recombinant antigens. PMID:17152007

  6. Vaccinia virus inhibits T cell receptor-dependent responses by human gammadelta T cells.

    Science.gov (United States)

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

    2007-01-01

    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 gammadelta T cells because of the role they play in immune control of this virus. Exposure to VV failed to trigger proliferative responses in gammadelta T cells from unprimed individuals, but it was an unexpected finding that VV blocked responses to model antigens by the Vgamma2Vdelta2 T cell subset. Infectious or ultraviolet light-inactivated VV inhibited proliferative Vgamma2Vdelta2 T cell responses to phosphoantigens and tumor cells, prevented cytolysis of Daudi B cells, and reduced cytokine production. Inhibiting Vgamma2Vdelta2 T cells may be a mechanism for evading host immunity and increasing VV virulence. Increased VV replication or expression in the absence of gammadelta T cell responses might contribute to its potency as a vaccine against poxvirus and recombinant antigens.

  7. Research progress in response strategies to smallpox bioterrorism%生物恐怖视角下的天花应对策略研究进展

    Institute of Scientific and Technical Information of China (English)

    祖正虎; 许晴; 张文斗; 徐致靖; 黄培堂; 郑涛

    2013-01-01

    Emergency prevention and control of smallpox transmission in the context of bioterrorism require effective response strategies. This paper began by describing the general process of computational experiments on smallpox response strategies. Then,the current achievement of this method was summarized in terms of the experimental environment, the computational model and intervention strategies. Finally, problems with smallpox response strategy simulation and directions of further research were discussed.%生物恐怖视角下天花传播的应急防控需要科学的应对策略.本文首先介绍天花应对策略计算实验的一般过程,然后从实验环境、计算模型及干预策略3个方面总结当前的研究及取得的进展,并就天花应对策略模拟存在的问题及进一步研究方向进行讨论.

  8. Variola minor in coalfield areas of England and Wales, 1921-34: Geographical determinants of a national smallpox epidemic that spread out of effective control.

    Science.gov (United States)

    Smallman-Raynor, Matthew R; Rafferty, Sarah; Cliff, Andrew D

    2017-05-01

    This paper uses techniques of binary logistic regression to identify the spatial determinants of the last national epidemic of smallpox to spread in England and Wales, the variola minor epidemic of 1921-34. Adjusting for age and county-level variations in vaccination coverage in infancy, the analysis identifies a dose-response gradient with increasing odds of elevated smallpox rates in local government areas with (i) medium (odds ratio [OR] = 5.32, 95% Confidence Interval [95% CI] 1.96-14.41) and high (OR = 11.32, 95% CI 4.20-31.59) coal mining occupation rates and (ii) medium (OR = 16.74, 95% CI 2.24-125.21) and high (OR = 63.43, 95% CI 7.82-497.21) levels of residential density. The results imply that the spatial transmission of variola virus was facilitated by the close spatial packing of individuals, with a heightened transmission risk in coal mining areas of the country. A syndemic interaction between common respiratory conditions arising from exposure to coal dust and smallpox virus transmission is postulated to have contributed to the findings. We suggest that further studies of the geographical intersection of coal mining and acute infections that are transmitted via respiratory secretions are warranted. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Small(pox success? Erradicação da varíola: medida do sucesso?

    Directory of Open Access Journals (Sweden)

    Anne-Emanuelle Birn

    2011-02-01

    Full Text Available The 30th anniversary of the World Health Organization's (WHO official certification of smallpox eradication was marked by a slew of events hailing the campaign's dramatic tale of technological and organizational triumph against an ancient scourge. Yet commemorations also serve as moments of critical reflection. This article questions the acclaim showered upon smallpox eradication as the single greatest public health success in history. It examines how and why smallpox eradication and WHO's concurrent social justice-oriented primary health care approach (following from the Declaration of Alma-Ata became competing paradigms. It synthesizes critiques of eradication's shortcomings and debunks some of the myths surrounding the global eradication campaign as a public health priority and necessity, and as a Cold War victory of cooperation. The article concludes with thoughts on integrating technical and social-political aspects of health within the context of welfare states as the means to achieving widespread and enduring global public health success.O 30º aniversário da certificação oficial da Organização Mundial da Saúde (OMS da erradicação da varíola foi marcado por uma série de eventos que saudaram a dramática história do triunfo tecnológico e organizacional contra este antigo flagelo. Todavia, as comemorações também servem como momentos para uma reflexão crítica. Este artigo questiona os aplausos excessivos para a erradicação da varíola como o único e absoluto sucesso na história da saúde pública. Examina como e por que a erradicação da varíola e a abordagem da atenção básica de saúde orientada para a justiça social (de acordo com a Declaração de Alma-Ata se tornaram paradigmas concorrentes. Sintetiza críticas sobre as deficiências da erradicação e busca desvelar alguns dos mitos que envolvem a campanha de erradicação global como uma prioridade e necessidade da saúde pública, e como uma vitória da

  10. Deletion of C7L and K1L Genes Leads to Significantly Decreased Virulence of Recombinant Vaccinia Virus TianTan

    OpenAIRE

    Zheng Liu; Shuhui Wang; Qicheng Zhang; Meijuan Tian; Jue Hou; Rongmin Wang; Chang Liu; Xu Ji; Ying Liu; Yiming Shao

    2013-01-01

    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 o...

  11. Extrachromosomal recombination in vaccinia-infected cells requires a functional DNA polymerase participating at a level other than DNA replication.

    Science.gov (United States)

    Colinas, R J; Condit, R C; Paoletti, E

    1990-12-01

    Homologous recombination was measured in vaccinia-infected cells cotransfected with two plasmid recombination substrates. One plasmid contains a vaccinia protein lacZ coding region bearing a 1.1 kb 3' terminal deletion while the other plasmid contains a non-promoted lacZ coding region bearing a 1.1 kb 5' terminal deletion. Homologous recombination occurring between the 825 bp of lacZ common to both plasmids regenerates a functional lacZ gene from which B-galactosidase expression was measured. The entire 3 kb lacZ gene was used as a positive control. A panel of thermosensitive mutants was screened in cells either transfected with the positive control plasmid or cotransfected with the recombination substrates. A DNA - mutant, ts42, known to map to the viral DNA polymerase gene was found to be defective in recombination. Significantly, other DNA - mutants, ts17 or ts25, or other DNA polymerase mutants did not exhibit a defect in recombination similar to ts42. Inhibitors of viral DNA synthesis did not uniformly affect recombination. Cytosine arabinoside and aphidicolin inhibited B-galactosidase expression from the recombination substrates but not from the positive control plasmid, whereas hydroxyurea enhanced expression from both. Marker rescue with the cloned wildtype DNA polymerase gene repaired the defect in ts42. Southern and western analyses demonstrated that B-galactosidase activity was consistent with a recombined lacZ gene and unit size 116 kDa protein. Measurement of plasmid and viral DNA replication in cells infected with the different DNA - mutants indicated that recombination was independent of plasmid and viral DNA replication. Together these results suggest that the vaccinia DNA polymerase participates in homologous recombination at a level other than that of DNA replication.

  12. Involvement of the Cellular Phosphatase DUSP1 in Vaccinia Virus Infection

    Science.gov (United States)

    Cáceres, Ana; Perdiguero, Beatriz; Gómez, Carmen E.; Cepeda, Maria Victoria; Caelles, Carme; Sorzano, Carlos Oscar; Esteban, Mariano

    2013-01-01

    Poxviruses encode a large variety of proteins that mimic, block or enhance host cell signaling pathways on their own benefit. It has been reported that mitogen-activated protein kinases (MAPKs) are specifically upregulated during vaccinia virus (VACV) infection. Here, we have evaluated the role of the MAPK negative regulator dual specificity phosphatase 1 (DUSP1) in the infection of VACV. We demonstrated that DUSP1 expression is enhanced upon infection with the replicative WR virus and with the attenuated VACV viruses MVA and NYVAC. This upregulation is dependent on early viral gene expression. In the absence of DUSP1 in cultured cells, there is an increased activation of its molecular targets JNK and ERK and an enhanced WR replication. Moreover, DUSP1 knock-out (KO) mice are more susceptible to WR infection as a result of enhanced virus replication in the lungs. Significantly, MVA, which is known to produce non-permissive infections in most mammalian cell lines, is able to grow in DUSP1 KO immortalized murine embryo fibroblasts (MEFs). By confocal and electron microscopy assays, we showed that in the absence of DUSP1 MVA morphogenesis is similar as in permissive cell lines and demonstrated that DUSP1 is involved at the stage of transition between IVN and MV in VACV morphogenesis. In addition, we have observed that the secretion of pro-inflammatory cytokines at early times post-infection in KO mice infected with MVA and NYVAC is increased and that the adaptive immune response is enhanced in comparison with WT-infected mice. Altogether, these findings reveal that DUSP1 is involved in the replication and host range of VACV and in the regulation of host immune responses through the modulation of MAPKs. Thus, in this study we demonstrate that DUSP1 is actively involved in the antiviral host defense mechanism against a poxvirus infection. PMID:24244156

  13. The changes of vaccinia related kinase 1 in grafted heart after rat heart transplantation

    Science.gov (United States)

    Qian, Shiguo; Yang, Xuechao; Wu, Kunpeng; Lv, Qiangsheng; Zhang, Yuanyuan; Dai, Jiahong; Chen, Cheng

    2014-01-01

    Objective To assess the expression and significance of vaccinia-related kinase 1 (VRK1) after rat heart transplantation. Materials and methods Lewis and Wistar rats weighing 250 to 300 g were used as donors and recipients. Allografts were from Wistar transplanted into Lewis, and isografts were transplanted from Lewis into Lewis. Grafts were harvested at 1, 3, 5, and 7 days after transplantation. We performed Western Blot of heart tissues after cardiac transplantation. To analyze VRK1 express between the isografts and allografts for immunohistochemical staining. At 5th day after heart transplantation use related cytokines VRK1 for immunohistochemical. We used double immunofluorescent staining on transverse cryosections of graft tissues by co-labeling with different markers, including those for VRK1, activate caspase-3, α-actinin, VCAM-1, CD4. Results Compared with rare expression in syngeneic Lewis rat hearts, VRK1 protein level in allogeneic hearts were detected at various survival times after heterotopic heart transplantation, which observably expressed on day 5 postoperative. In addition, we examined the expression of activate caspase-3 in allogeneic hearts, which has a similar expression with VRK1. Immunohistochemical and immunofluorescent method displayed that VRK1 was widely expressed in cytoplasm of cardiac tissue and activate caspase-3 was also expressed in cardiomyocytes. However, the VRK1 wasn’t express in inflammation. Conclusions The VRK1 expression has increased after heart transplantation in allograft and isograft, and VRK1 may play a significant role in myocardial apoptosis after heterotopic heart transplantation in rats. PMID:25589968

  14. Single-particle characterization of oncolytic vaccinia virus by flow virometry.

    Science.gov (United States)

    Tang, Vera A; Renner, Tyler M; Varette, Oliver; Le Boeuf, Fabrice; Wang, Jiahu; Diallo, Jean-Simon; Bell, John C; Langlois, Marc-André

    2016-09-30

    Vaccinia virus (VV) is an oncolytic virus that is currently being evaluated as a promising cancer vaccine in several phase I, II and III clinical trials. Although several quality control tests are performed on each new batch of virus, these do not routinely include a systematic characterization of virus particle homogeneity, or relate the infectious titer to the total number of submicron sized particles (SSPs) present in the sample. SSPs are comprised of infectious virus and non-infectious viral particles, but also cell contaminants derived from the virus isolation procedures, such as cellular vesicles and debris. Here we have employed flow virometry (FV) analysis and sorting to isolate and characterize distinct SSP populations in therapeutic oncolytic VV preparations. We show that VV preparations contain SSPs heterogeneous in size and include large numbers of non-infectious VV particles. Furthermore, we used FV to illustrate how VV has a propensity to aggregate over time and under various handling and storage procedures. Accordingly, we find that together the infectious titer, the total number of SSPs, the number of viral genomes and the level of particle aggregation in a sample constitute useful parameters that greatly facilitate inter-sample assessment of physical quality, and also provides a means to monitor sample deterioration over time. Additionally, we have successfully employed FV sorting to further isolate virus from other particles by identifying a lipophilic dye that preferentially stains VV over other SSPs in the sample. Overall, we demonstrate that FV is a fast and effective tool that can be used to perform quality, and consistency control assessments of oncolytic VV vaccine preparations.

  15. Seven major genomic deletions of vaccinia virus Tiantan strain are sufficient to decrease pathogenicity.

    Science.gov (United States)

    Li, Yiquan; Sheng, Yuan; Chu, Yunjie; Ji, Huifan; Jiang, Shuang; Lan, Tian; Li, Min; Chen, Shuang; Fan, Yuanyuan; Li, Wenjie; Li, Xiao; Sun, Lili; Jin, Ningyi

    2016-05-01

    Attenuated strain TTVAC7, as a multi-gene-deleted vaccinia virus Tiantan strain (VTT), was constructed by knocking out parts of non-essential genes related to virulence, host range and immunomodulation of VTT, and by combining double marker screening with exogenous selectable marker knockout techniques. In this study, shuttle vector plasmids pTC-EGFP, pTA35-EGFP, pTA66-EGFP, pTE-EGFP, pTB-EGFP, pTI-EGFP and pTJ-EGFP were constructed, which contained seven pairs of recombinant arms linked to the early and late strong promoter pE/L, as well as to enhanced green fluorescent protein (EGFP) as an exogenous selectable marker. BHK cells were co-transfected/infected successively with the above plasmids and VTT or gene-deleted VTT, and homologous recombination and fluorescence plaque screening methods were used to knock out the gene fragments (TC: TC7L ∼ TK2L; TA35: TA35L; TA66: TA66R; TE: TE3L ∼ TE4L; TB: TB13R; TI: TI4L; TJ: TJ2R). The Cre/LoxP system was then applied to knock out the exogenous selectable marker, and ultimately the gene-deleted attenuated strain TTVAC7 was obtained. A series of in vivo and in vitro experiments demonstrated that not only the host range of TTVAC7 could be narrowed and its toxicity weakened significantly, but its high immunogenicity was maintained at the same time. These results support the potential of TTVAC7 to be developed as a safe viral vector or vaccine.

  16. Apoptosis and necrosis in vaccinia virus-infected HeLa G and BSC-40 cells.

    Science.gov (United States)

    Liskova, Jana; Knitlova, Jarmila; Honner, Richard; Melkova, Zora

    2011-09-01

    In most cells, vaccinia virus (VACV) infection is considered to cause a lytic cell death, an equivalent of necrosis. However, upon infection of the epithelial cell lines HeLa G and BSC-40 with VACV strain Western Reserve (WR), we have previously observed an increased activation of and activity attributable to caspases, a typical sign of apoptosis. In this paper, we have further analyzed the type of cell death in VACV-infected cells HeLa G and BSC-40. In a cell-based flow cytometric assay, we showed a specific activation of caspase-2 and 4 in HeLa G and BSC-40 cells infected with VACV, strain WR, while we did not find any effects of inhibitors of calpain and cathepsin D and E. The actual activity of the two caspases, but also of caspase-3, was then confirmed in lysates of infected HeLa G, but not in BSC-40 cells. Accordingly, poly(ADP)-ribose polymerase (PARP) cleavage was found increased only in infected HeLa G cells. Consequently, we have determined morphological features of apoptosis and/or activity of the executioner caspase-3 in infected HeLa G cells in situ, while only a background apoptosis was observed in infected BSC-40 cells. Finally, vaccination strains Dryvax and Praha were found to induce apoptosis in both HeLa G and BSC-40 cells, as characterized morphologically and by PARP cleavage. These findings may be important for understanding the differences in VACV-host interactions and post-vaccination complications in different individuals.

  17. Complete nucleotide sequences of two adjacent early vaccinia virus genes located within the inverted terminal repetition.

    Science.gov (United States)

    Venkatesan, S; Gershowitz, A; Moss, B

    1982-11-01

    The proximal part of the 10,000-base pair (bp) inverted terminal repetition of vaccinia virus DNA encodes at least three early mRNAs. A 2,236-bp segment of the repetition was sequenced to characterize two of the genes. This task was facilitated by constructing a series of recombinants containing overlapping deletions; oligonucleotide linkers with synthetic restriction sites provided points for radioactive labeling before sequencing by the chemical degradation method of Maxam and Gilbert (Methods Enzymol. 65:499-560, 1980). The ends of the transcripts were mapped by hybridizing labeled DNA fragments to early viral RNA and resolving nuclease S1-protected fragments in sequencing gels, by sequencing cDNA clones, and from the lengths of the RNAs. The nucleotide sequences for at least 60 bp upstream of both transcriptional initiation sites are more than 80% adenine . thymine rich and contain long runs of adenines and thymines with some homology to procaryotic and eucaryotic consensus sequences. The gene transcribed in the rightward direction encodes an RNA of approximately 530 nucleotides with a single open reading frame of 420 nucleotides. Preceding the first AUG, there is a heptanucleotide that can hybridize to the 3' end of 18S rRNA with only one mismatch. The derived amino acid sequence of the protein indicated a molecular weight of 15,500. The gene transcribed in the leftward direction encodes an RNA 1,000 to 1,100 nucleotides long with an open reading frame of 996 nucleotides and a leader sequence of only 5 to 6 nucleotides. The derived amino acid sequence of this protein indicated a molecular weight of 38,500. The 3' ends of the two transcripts were located within 100 bp of each other. Although there are adenine . thymine-rich clusters near the putative transcriptional termination sites, specific AATAAA polyadenylic acid signal sequences are absent.

  18. 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

    Science.gov (United States)

    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

    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. PMID:22398243

  19. Parallel simulation of smallpox spreading in large populations based on social contact networks%基于社会接触网络的大规模人群天花传播并行仿真

    Institute of Scientific and Technical Information of China (English)

    许晴; 祖正虎; 张文斗; 徐致靖; 黄培堂; 郑涛

    2012-01-01

    天花生物恐怖是世界各国面临的重大潜在威胁.本文利用并行仿真的技术手段,研究天花在大规模城市人群中的传播规律并对环形接种和大规模疫苗接种这两种主要的天花疫苗接种策略进行有效性分析,结果表明环形接种措施能够有效控制天花疫情的扩散,在天花疫苗储备不足的情况下可以优先考虑采用该措施.%Smallpox bioterrorism is a potentially significant threat to the world. This paper used the parallel simulation method to study the spread of smallpox in large urban populations and analyzed the effectiveness of two main smallpox vaccination strategies,which are ring vaccination and mass vaccination. The result showed that ring vaccination strategy can effectively control smallpox epidemic and should be given priority in case of shortage of smallpox vaccine reserves.

  20. Political epidemiology: strengthening socio-political analysis for mass immunisation - lessons from the smallpox and polio programmes.

    Science.gov (United States)

    Taylor, S

    2009-01-01

    Control and reduction of infectious diseases is a key to attaining the Millennium Development Goals. An important element of this work is the successful immunisation, especially in resource-poor countries. Mass immunisation, most intensively in the case of eradication, depends on a combination of reliable demand (e.g. public willingness to comply with the vaccine protocol) and effective supply (e.g. robust, generally state-led, vaccine delivery). This balance of compliance and enforceability is, quintessentially, socio-political in nature - conditioned by popular perceptions of disease and risk, wider conditions of economic development and poverty, technical aspects of vaccine delivery, and the prevailing international norms regarding power relations between states and peoples. In the past 100 years, three out of six disease eradication programmes have failed. The explanations for failure have focused on biotechnical and managerial or financial issues. Less attention is paid to socio-political aspects. Yet socio-political explanations are key. Eradication is neither inherently prone to failure, nor necessarily doomed in the case of polio. However, eradication, and similar mass immunisation initiatives, which fail to address social and political realities of intervention may be. A comparison of the smallpox and polio eradication programmes illustrates the importance of disease-specific socio-political analysis in programme conceptualisation, design, and management.

  1. A case of exemplarity: C. F. Rottböll's history of smallpox inoculation in Denmark-Norway, 1766.

    Science.gov (United States)

    Eriksen, Anne

    2010-01-01

    Smallpox inoculation was one of the great discoveries of the 18th century and has been written into the grand narrative of medical progress, describing the taming of epidemic disease. Setting the perspective of progress aside, the article explores how this medical innovation was situated in 18th-century society and culture. The aim is to investigate how medical practice was intertwined with social structure and cultural patterns. The article takes its case from a book published in Copenhagen in 1766 by Professor C. F. Rottböll, former Head Physician of the Royal Inoculation House in Copenhagen. Being the first medical treatise on inoculation in Denmark-Norway, the book also has a historical section followed by a collection of reports and letters written by a number of other authors from various parts of the kingdom. Through close reading, the article explores how the introduction of the new technique was described in the texts. The reports were written to present practice and discuss cases. In doing so, they also presented a variety of other concerns so that a diversity of aims and intentions are added to the medical ones. The social and rhetorical strategies employed illuminate social ambition and systems of patronage, as well as understandings of history and of truth.

  2. Smallpox vaccination in the early 19th century using live carriers: the travels of Francisco Xavier de Balmis.

    Science.gov (United States)

    Aldrete, J Antonio

    2004-04-01

    Realizing that the Spanish colonies were being devastated by epidemics of smallpox resulting in thousands of deaths, Charles IV, King of Spain, sent one of his court's physicians to apply the recently discovered vaccine. Without refrigeration, the vaccine was passed from one child to another (boys taken out of orphanages). Francisco Xavier de Balmis and a team that included three assistants, two surgeons, and three nurses sailed from Spain on November 30. 1803. They vaccinated more than 100,000 people from the Caribbean Islands and South, Central, and North America, reaching up to San Antonio, Texas, and then traveled to the Philippines, Macao, Canton, and Santa Elena Island, landing back in Cadiz on September 7, 1806. During his journey, Balmis instructed local physicians on how to prepare, preserve, and apply the vaccine, while collecting rare biologic specimens. On the 200th anniversary of their sailing, recognition is given to this group for conducting what was the first global vaccination campaign that reached Texas and California.

  3. [The historical development of immunization in Germany. From compulsory smallpox vaccination to a National Action Plan on Immunization].

    Science.gov (United States)

    Klein, S; Schöneberg, I; Krause, G

    2012-11-01

    In the German Reich, smallpox vaccinations were organized by the state. A mandatory vaccination throughout the empire was introduced in 1874, which was continued in the Federal Republic of Germany (FRG) and the German Democratic Republic (GDR) until 1982/1983. From 1935, health departments were responsible for vaccinations. In the GDR, immunization was tightly organized: The state made great efforts to achieve high vaccination rates. Responsibilities were clearly defined at all levels and for all ages. While vaccination was initially mandatory only at the regional level, the legally mandated immunization schedule later contained compulsory vaccinations, e.g., against measles. In the beginning there were mandatory vaccinations in the FRG at the Länder level. Since 1961, the Federal Epidemics Act has impeded obligatory vaccinations. Instead, voluntary vaccinations based on recommendations were stressed. Since the 1980s, vaccinations have been shifted from the public health service sector to office-based physicians. Today, public health authorities offer mainly supplementary vaccinations. In 2007, protective immunizations were introduced as compulsory benefits of the statutory health insurance (SHI). Recently, the German federal states developed a National Vaccination Plan to support immunization strategies.

  4. The sequence of camelpox virus shows it is most closely related to variola virus, the cause of smallpox.

    Science.gov (United States)

    Gubser, Caroline; Smith, Geoffrey L

    2002-04-01

    Camelpox virus (CMPV) and variola virus (VAR) are orthopoxviruses (OPVs) that share several biological features and cause high mortality and morbidity in their single host species. The sequence of a virulent CMPV strain was determined; it is 202182 bp long, with inverted terminal repeats (ITRs) of 6045 bp and has 206 predicted open reading frames (ORFs). As for other poxviruses, the genes are tightly packed with little non-coding sequence. Most genes within 25 kb of each terminus are transcribed outwards towards the terminus, whereas genes within the centre of the genome are transcribed from either DNA strand. The central region of the genome contains genes that are highly conserved in other OPVs and 87 of these are conserved in all sequenced chordopoxviruses. In contrast, genes towards either terminus are more variable and encode proteins involved in host range, virulence or immunomodulation. In some cases, these are broken versions of genes found in other OPVs. The relationship of CMPV to other OPVs was analysed by comparisons of DNA and predicted protein sequences, repeats within the ITRs and arrangement of ORFs within the terminal regions. Each comparison gave the same conclusion: CMPV is the closest known virus to variola virus, the cause of smallpox.

  5. Ectromelia Virus Disease Characterization in the BALB/c Mouse: A Surrogate Model for Assessment of Smallpox Medical Countermeasures

    Directory of Open Access Journals (Sweden)

    Jennifer Garver

    2016-07-01

    Full Text Available In 2007, the United States– Food and Drug Administration (FDA issued guidance concerning animal models for testing the efficacy of medical countermeasures against variola virus (VARV, the etiologic agent for smallpox. Ectromelia virus (ECTV is naturally-occurring and responsible for severe mortality and morbidity as a result of mousepox disease in the murine model, displaying similarities to variola infection in humans. Due to the increased need of acceptable surrogate animal models for poxvirus disease, we have characterized ECTV infection in the BALB/c mouse. Mice were inoculated intranasally with a high lethal dose (125 PFU of ECTV, resulting in complete mortality 10 days after infection. Decreases in weight and temperature from baseline were observed eight to nine days following infection. Viral titers via quantitative polymerase chain reaction (qPCR and plaque assay were first observed in the blood at 4.5 days post-infection and in tissue (spleen and liver at 3.5 days post-infection. Adverse clinical signs of disease were first observed four and five days post-infection, with severe signs occurring on day 7. Pathological changes consistent with ECTV infection were first observed five days after infection. Examination of data obtained from these parameters suggests the ECTV BALB/c model is suitable for potential use in medical countermeasures (MCMs development and efficacy testing.

  6. Ectromelia Virus Disease Characterization in the BALB/c Mouse: A Surrogate Model for Assessment of Smallpox Medical Countermeasures.

    Science.gov (United States)

    Garver, Jennifer; Weber, Lauren; Vela, Eric M; Anderson, Mike; Warren, Richard; Merchlinsky, Michael; Houchens, Christopher; Rogers, James V

    2016-07-22

    In 2007, the United States- Food and Drug Administration (FDA) issued guidance concerning animal models for testing the efficacy of medical countermeasures against variola virus (VARV), the etiologic agent for smallpox. Ectromelia virus (ECTV) is naturally-occurring and responsible for severe mortality and morbidity as a result of mousepox disease in the murine model, displaying similarities to variola infection in humans. Due to the increased need of acceptable surrogate animal models for poxvirus disease, we have characterized ECTV infection in the BALB/c mouse. Mice were inoculated intranasally with a high lethal dose (125 PFU) of ECTV, resulting in complete mortality 10 days after infection. Decreases in weight and temperature from baseline were observed eight to nine days following infection. Viral titers via quantitative polymerase chain reaction (qPCR) and plaque assay were first observed in the blood at 4.5 days post-infection and in tissue (spleen and liver) at 3.5 days post-infection. Adverse clinical signs of disease were first observed four and five days post-infection, with severe signs occurring on day 7. Pathological changes consistent with ECTV infection were first observed five days after infection. Examination of data obtained from these parameters suggests the ECTV BALB/c model is suitable for potential use in medical countermeasures (MCMs) development and efficacy testing.

  7. Bob H. Reinhardt: The End of A Global Pox. America and the Eradication of Smallpox in the Cold War Era : The University of North Carolina Press, Chapel Hill, 2015, 268 pp., Hard cover, Illustrated, Figure, Maps, Table, Notes, Bibliography, Index., $39.95.

    Science.gov (United States)

    Imperato, Pascal James

    2017-04-01

    This review examines in detail Bob H. Reinhardt's meticulous analyses of smallpox eradication within the broad context of American liberalism, Cold War politics, and the exercise of technological, medical, and political power on the part of the United States. As a result, his book provides a unique perspective on the eradication of smallpox.

  8. Safety and biodistribution of a double-deleted oncolytic vaccinia virus encoding CD40 ligand in laboratory Beagles.

    Science.gov (United States)

    Autio, Karoliina; Knuuttila, Anna; Kipar, Anja; Pesonen, Sari; Guse, Kilian; Parviainen, Suvi; Rajamäki, Minna; Laitinen-Vapaavuori, Outi; Vähä-Koskela, Markus; Kanerva, Anna; Hemminki, Akseli

    2014-01-01

    We evaluated adverse events, biodistribution and shedding of oncolytic vaccinia virus encoding CD40 ligand in two Beagles, in preparation for a phase 1 trial in canine cancer patients. Dog 1 received one dose of vaccinia virus and was euthanized 24 hours afterwards, while dog 2 received virus four times once weekly and was euthanized 7 days after that. Dogs were monitored for adverse events and underwent a detailed postmortem examination. Blood, saliva, urine, feces, and organs were collected for virus detection. Dog 1 had mild fever and lethargy while dog 2 experienced a possible seizure 5.5 hours after first virus administration. Viral DNA declined quickly in the blood after virus administration in both dogs but was still detectable 1 week later by quantitative polymerase chain reaction. Only samples taken directly after virus infusion contained infectious virus. Small amounts of viral DNA, but no infectious virus, were detected in a few saliva and urine samples. Necropsies did not reveal any relevant pathological changes and virus DNA was detected mainly in the spleen. The dogs in the study did not have cancer, and thus adverse events could be more common and viral load higher in dogs with tumors which allow viral amplification.

  9. Protective immunity in macaques vaccinated with a modified vaccinia virus Ankara-based measles virus vaccine in the presence of passively acquired antibodies

    NARCIS (Netherlands)

    K.J. Stittelaar (Koert); L.S. Wyatt (Linda); R.L. de Swart (Rik); H.W. Vos (Helma); J. Groen (Jan); G. van Amerongen (Geert); R.S. van Binnendijk (Rob); S. Rozenblatt (Shmuel); B. Moss (Bernard); A.D.M.E. Osterhaus (Albert)

    2000-01-01

    textabstractRecombinant modified vaccinia virus Ankara (MVA), encoding the measles virus (MV) fusion (F) and hemagglutinin (H) (MVA-FH) glycoproteins, was evaluated in an MV vaccination-challenge model with macaques. Animals were vaccinated twice in the absence or presence of passively transferred M

  10. Protective immunity in macaques vaccinated with a modified vaccinia virus Ankara-based measles virus vaccine in the presence of passively acquired antibodies

    NARCIS (Netherlands)

    K.J. Stittelaar (Koert); L.S. Wyatt (Linda); R.L. de Swart (Rik); H.W. Vos (Helma); J. Groen (Jan); R.S. van Binnendijk (Rob); S. Rozenblatt (Shmuel); B. Moss (Bernard); G. van Amerongen (Geert); A.D.M.E. Osterhaus (Albert)

    2000-01-01

    textabstractRecombinant modified vaccinia virus Ankara (MVA), encoding the measles virus (MV) fusion (F) and hemagglutinin (H) (MVA-FH) glycoproteins, was evaluated in an MV vaccination-challenge model with macaques. Animals were vaccinated twice in the absence or presence of passi

  11. High-affinity human leucocyte antigen class I binding variola-derived peptides induce CD4(+) T cell responses more than 30 years post-vaccinia virus vaccination

    DEFF Research Database (Denmark)

    Wang, M.; Tang, Sheila Tuyet; Lund, Ole;

    2009-01-01

    Interferon-gamma secreting T lymphocytes against pox virus-derived synthetic 9-mer peptides were tested by enzyme-linked immunospot in peripheral blood of individuals vaccinated with vaccinia virus more than 30 years ago. The peptides were characterized biochemically as high-affinity human...

  12. Molecular network, pathway, and functional analysis of time-dependent gene changes associated with pancreatic cancer susceptibility to oncolytic vaccinia virotherapy

    Directory of Open Access Journals (Sweden)

    Dana Haddad

    2016-01-01

    Conclusions: Our study reveals the ability to assess time-dependent changes in gene expression patterns in pancreatic cancer cells associated with infection and susceptibility to vaccinia viruses. This suggests that molecular assays may be useful to develop safer and more efficacious oncolyticvirotherapies and support the idea that these treatments may target pathways implicated in pancreatic cancer resistance to conventional therapies.

  13. Biophysical analysis of bacterial and viral systems. A shock tube study of bio-aerosols and a correlated AFM/nanosims investigation of vaccinia virus

    Energy Technology Data Exchange (ETDEWEB)

    Gates, Sean Damien [Stanford Univ., CA (United States)

    2013-05-01

    The work presented herein is concerned with the development of biophysical methodology designed to address pertinent questions regarding the behavior and structure of select pathogenic agents. Two distinct studies are documented: a shock tube analysis of endospore-laden bio-aerosols and a correlated AFM/NanoSIMS study of the structure of vaccinia virus.

  14. Increased antibody responses to human papillomavirus type 16 L1 protein expressed by recombinant vaccinia virus lacking serine protease inhibitor genes.

    Science.gov (United States)

    Zhou, J; Crawford, L; McLean, L; Sun, X Y; Stanley, M; Almond, N; Smith, G L

    1990-09-01

    The L1 gene of human papillomavirus type 16 (HPV-16) driven by the vaccinia virus major late 4b gene promoter has been inserted into three different sites of the vaccinia virus genome. Insertion into the thymidine kinase (TK) gene was achieved by selection of TK- mutants in BUdR on TK- cells. Insertion into two vaccinia virus serine protease inhibitor (serpin) genes was achieved by co-insertion of the Escherichia coli xanthine guanine phosphoribosyltransferase gene linked to the vaccinia virus 7.5K promoter and selection of mycophenolic acid-resistant recombinant viruses. Each recombinant virus expressed a 57K L1 protein at similar levels and with similar kinetics. However, immunization of mice with these recombinant viruses induced different levels of antibody to the L1 protein. Viruses lacking serpin genes B13R and B24R induced significantly higher antibody levels than did viruses lacking the TK gene. The presence of functional B13R and B24R gene products is therefore somehow immunosuppressive at least for antibody responses to the L1 protein of HPV-16.

  15. Virotherapy of canine tumors with oncolytic vaccinia virus GLV-1h109 expressing an anti-VEGF single-chain antibody.

    Directory of Open Access Journals (Sweden)

    Sandeep S Patil

    Full Text Available Virotherapy using oncolytic vaccinia virus (VACV strains is one promising new strategy for cancer therapy. We have previously reported that oncolytic vaccinia virus strains expressing an anti-VEGF (Vascular Endothelial Growth Factor single-chain antibody (scAb GLAF-1 exhibited significant therapeutic efficacy for treatment of human tumor xenografts. Here, we describe the use of oncolytic vaccinia virus GLV-1h109 encoding GLAF-1 for canine cancer therapy. In this study we analyzed the virus-mediated delivery and production of scAb GLAF-1 and the oncolytic and immunological effects of the GLV-1h109 vaccinia virus strain against canine soft tissue sarcoma and canine prostate carcinoma in xenograft models. Cell culture data demonstrated that the GLV-1h109 virus efficiently infect, replicate in and destroy both tested canine cancer cell lines. In addition, successful expression of GLAF-1 was demonstrated in virus-infected canine cancer cells and the antibody specifically recognized canine VEGF. In two different xenograft models, the systemic administration of the GLV-1h109 virus was found to be safe and led to anti-tumor and immunological effects resulting in the significant reduction of tumor growth in comparison to untreated control mice. Furthermore, tumor-specific virus infection led to a continued production of functional scAb GLAF-1, resulting in inhibition of angiogenesis. Overall, the GLV-1h109-mediated cancer therapy and production of immunotherapeutic anti-VEGF scAb may open the way for combination therapy concept i.e. vaccinia virus mediated oncolysis and intratumoral production of therapeutic drugs in canine cancer patients.

  16. 天花生物恐怖风险下最优疫苗接种比例%Optimal proportion of immunized populations under different smallpox bioterrorist attack risks

    Institute of Scientific and Technical Information of China (English)

    许晴; 祖正虎; 张文斗; 徐致靖; 黄培堂; 郑涛

    2012-01-01

    天花是最危险的生物剂之一,目前全世界已经消除天花疾病,但发生天花生物恐怖袭击的可能性仍然存在.大规模接种天花疫苗是最有效的天花生物恐怖预防措施,但有可能会导致大量接种者产生严重的不良反应甚至死亡.因此,在存在天花恐怖事件可能性但尚未发生的当前,保有一定比例的天花免疫人群会减少真正发生时的整体损失.本文从最小化质量调整生命年损失的角度,探讨国家在不同天花生物恐怖袭击风险下应保有的最佳免疫人群比例.%Smallpox is one of the most dangerous biological agents. Although smallbox has been eliminated globally, the possibility of smallpox being used during a bioterrorist attack still exists. Large-scale smallpox vaccination is the most effective measure against smallpox bioterrorism, but may lead to serious side effects and even death. Therefore, retaining a certain proportion of immunized populations will reduce the overall losses in case of smallpox bioterrorist incidents. Based on minimized quality-adjusted life years losses, this article explored the optimal proportion of immunized populations that a country shoulol retain under the different smallpox bioterrorist attack risks.

  17. Enfrentando el bioterrorismo: aspectos epidemiológicos, clínicos y preventivos de la viruela Confronting bioterrorism: epidemiologic, clinical, and preventive aspects of smallpox

    Directory of Open Access Journals (Sweden)

    Carlos Franco-Paredes

    2003-08-01

    Full Text Available Uno de los grandes logros de la salud pública mundial, la erradicación de la viruela, puede verse mermado por el posible riesgo de bioterrorismo. El debate acerca de la destrucción de los restos del virus en los dos laboratorios de referencia de la Organización Mundial de la Salud ha cambiado diametralmente debido a los eventos terroristas y a la dispersión intencional de Bacillus anthracis ocurridos en poblaciones civiles en Estados Unidos de América en el año 2001. La liberación del virus Variola con fines terroristas constituye un riesgo mínimo no cuantificable, pero desafortunadamente real. El impacto podría ser devastador debido a la elevada morbimortalidad de la enfermedad aunada al pánico y a la desestabilización social que podría ocasionar. Es por ello que el establecimiento de un plan de respuesta, sumado a disponibilidad de vacuna para ser utilizada pos-exposición, es importante dentro de los planes de contingencia contra el bioterrorismo. El reiniciar un programa limitado de vacunación contra la viruela, como parte de dicho plan, ha sido recientemente recomendado por el Comité Asesor de Vacunación, del Centro para el Control de las Enfermedades, pero la vacuna disponible puede causar complicaciones graves e incluso la muerte, por lo que dicha recomendación no ha sido universalmente aceptada. No obstante, el personal médico y de salud pública requiere de información actualizada sobre la viruela y su prevención, ya que ellos son la primera línea de defensa en caso de un posible brote a consecuencia de un ataque bioterrorista. El presente artículo presenta una revisión dirigida a proporcionar al personal de salud un enfoque clínico, epidemiológico y preventivo sobre la viruela.The worldwide eradication of smallpox, a major achievement in public health, is currently threatened by the risk of bioterrorism. The debate on the destruction of the Variola virus in the two reference laboratories of the World Health

  18. Reactions of Cre with methylphosphonate DNA: similarities and contrasts with Flp and vaccinia topoisomerase.

    Directory of Open Access Journals (Sweden)

    Chien-Hui Ma

    Full Text Available BACKGROUND: Reactions of vaccinia topoisomerase and the tyrosine site-specific recombinase Flp with methylphosphonate (MeP substituted DNA substrates, have provided important insights into the electrostatic features of the strand cleavage and strand joining steps catalyzed by them. A conserved arginine residue in the catalytic pentad, Arg-223 in topoisomerase and Arg-308 in Flp, is not essential for stabilizing the MeP transition state. Topoisomerase or its R223A variant promotes cleavage of the MeP bond by the active site nucleophile Tyr-274, followed by the rapid hydrolysis of the MeP-tyrosyl intermediate. Flp(R308A, but not wild type Flp, mediates direct hydrolysis of the activated MeP bond. These findings are consistent with a potential role for phosphate electrostatics and active site electrostatics in protecting DNA relaxation and site-specific recombination, respectively, against abortive hydrolysis. METHODOLOGY/PRINCIPAL FINDINGS: We have examined the effects of DNA containing MeP substitution in the Flp related Cre recombination system. Neutralizing the negative charge at the scissile position does not render the tyrosyl intermediate formed by Cre susceptible to rapid hydrolysis. Furthermore, combining the active site R292A mutation in Cre (equivalent to the R223A and R308A mutations in topoisomerase and Flp, respectively with MeP substitution does not lead to direct hydrolysis of the scissile MeP bond in DNA. Whereas Cre follows the topoisomerase paradigm during the strand cleavage step, it follows the Flp paradigm during the strand joining step. CONCLUSIONS/SIGNIFICANCE: Collectively, the Cre, Flp and topoisomerase results highlight the contribution of conserved electrostatic complementarity between substrate and active site towards transition state stabilization during site-specific recombination and DNA relaxation. They have potential implications for how transesterification reactions in nucleic acids are protected against

  19. Nucleotide sequence of XhoI O fragment of ectromelia virus DNA reveals significant differences from vaccinia virus.

    Science.gov (United States)

    Senkevich, T G; Muravnik, G L; Pozdnyakov, S G; Chizhikov, V E; Ryazankina, O I; Shchelkunov, S N; Koonin, E V; Chernos, V I

    1993-10-01

    The nucleotide sequence of the 3913 base pair XhoI O fragment located in an evolutionary variable region adjacent to the right end of the genome of ectromelia virus (EMV) was determined. The sequence contains two long open reading frames coding for putative proteins of 559 amino acid residues (p65) and 344 amino acid residues (p39). Amino acid database searches showed that p39 is closely related to vaccinia virus (VV), strain WR, B22R gene product (C12L gene product of strain Copenhagen), which belongs to the family of serine protease inhibitors (serpins). Despite the overall high conservation, differences were observed in the sequences of p39, B22R, and C12L in the site known to interact with proteases in other serpins, suggesting that the serpins of EMV and two strains of VV may all inhibit proteases with different specificities. The gene coding for the ortholog of p65 is lacking in the Copenhagen strain of vaccinia virus; the WR strain contains a truncated variant of this gene (B21R) potentially coding for a small protein (p16) corresponding to the C-terminal region of p65. p65 is a new member of the family of poxvirus proteins including vaccinia virus proteins A55R, C2L and F3L, and a group of related proteins of leporipoxviruses, Shope fibroma and myxoma viruses (T6, T8, T9, M9). These proteins are homologous to the Drosophila protein Kelch involved in egg development. Both Kelch protein and the related poxvirus proteins contain two distinct domains. The N-terminal domain is related to the similarly located domains of transcription factors Ttk, Br-C (Drosophila), and KUP (human), and GCL protein involved in early development in Drosophila. The C-terminal domain consists of an array of four to five imperfect repeats and is related to human placental protein MIPP. Phylogenetic analysis of the family of poxvirus proteins showed that their genes have undergone a complex succession of duplications, and complete or partial deletions.

  20. Evaluation of radiation effects against C6 glioma in combination with vaccinia virus-p53 gene therapy

    Science.gov (United States)

    Gridley, D. S.; Andres, M. L.; Li, J.; Timiryasova, T.; Chen, B.; Fodor, I.; Nelson, G. A. (Principal Investigator)

    1998-01-01

    The primary objective of this study was to evaluate the antitumor effects of recombinant vaccinia virus-p53 (rVV-p53) in combination with radiation therapy against the C6 rat glioma, a p53 deficient tumor that is relatively radioresistant. VV-LIVP, the parental virus (Lister strain), was used as a control. Localized treatment of subcutaneous C6 tumors in athymic mice with either rVV-p53 or VV-LIVP together with tumor irradiation resulted in low tumor incidence and significantly slower tumor progression compared to the agents given as single modalities. Assays of blood and spleen indicated that immune system activation may account, at least partly, for the enhance tumor inhibition seen with combined treatment. No overt signs of treatment-related toxicity were noted.

  1. Absence of vaccinia virus detection in a remote region of the Northern Amazon forests, 2005-2015.

    Science.gov (United States)

    Costa, Galileu Barbosa; Lavergne, Anne; Darcissac, Edith; Lacoste, Vincent; Drumond, Betânia Paiva; Abrahão, Jônatas Santos; Kroon, Erna Geessien; de Thoisy, Benoît; de Souza Trindade, Giliane

    2017-08-01

    Vaccinia virus (VACV) circulates in Brazil and other South America countries and is responsible for a zoonotic disease that usually affects dairy cattle and humans, causing economic losses and impacting animal and human health. Furthermore, it has been detected in wild areas in the Brazilian Amazon. To better understand the natural history of VACV, we investigated its circulation in wildlife from French Guiana, a remote region in the Northern Amazon forest. ELISA and plaque reduction neutralization tests were performed to detect anti-orthopoxvirus antibodies. Real-time and standard PCR targeting C11R, A56R and A26L were applied to detect VACV DNA in serum, saliva and tissue samples. No evidence of VACV infection was found in any of the samples tested. These findings provide additional information on the VACV epidemiological puzzle. The virus could nevertheless be circulating at low levels that were not detected in areas where no humans or cattle are present.

  2. Detection of Vaccinia Virus in Milk: Evidence of a Systemic and Persistent Infection in Experimentally Infected Cows.

    Science.gov (United States)

    de Oliveira, Tércia Moreira Ludolfo; Guedes, Maria Isabel Maldonado Coelho; Rehfeld, Izabelle Silva; Matos, Ana Carolina Diniz; Rivetti, Anselmo Vasconcelos; Alves, Pedro Augusto; Galinari, Grazielle Cossenzo Florentino; Cerqueira, Mônica Maria Oliveira Pinho; Abrahão, Jônatas Santos; Lobato, Zélia Inês Portela

    2015-11-01

    Bovine vaccinia (BV) is a zoonosis caused by Vaccinia virus (VACV), which affects lactating cows and milkers. VACV DNA and infectious particles have been detected in milk of naturally infected cows. However, the period and pattern of VACV shedding in milk is unknown, as is whether the presence of VACV in milk is due to a localized or a systemic infection. To address those questions, eight lactating cows were inoculated with VACV in previously scarified teats. The experiment was divided in two phases. In Phase 1, milk samples were collected daily for 33 days, and in Phase 2, four animals from the first phase were immunosuppressed. In both phases, milk was collected with a sterile catheter on even days and by hand milking on odd days. All animals showed typical BV lesions in the inoculated teats. All milk samples were subjected to nested polymerase chain reaction (PCR) and real-time quantitative PCR to detect VACV DNA. PCR-positive samples were subjected to virus isolation. VACV DNA was intermittently detected in milk in both phases and infectious viral particles could be detected only in phase 2, on the 69th, 73rd, 74th, 77th, 79th, and 81st days postinfection. Despite the possibility of propagation of VACV through milk, it is known that milk continues to be drawn and marketed normally during outbreaks of the disease. The detection of both VACV DNA and infectious particles in milk samples draws attention to the potential public health risk associated with the consumption of milk from BV outbreaks. Detection of VACV in the milk from noninfected teats demonstrated that VACV shedding in milk might be related to a systemic infection. Moreover, it was shown that VACV DNA and viral infectious particles could be detected in milk even after healing of the lesions, demonstrating that VACV may cause a persistent infection in cattle.

  3. A human multi-epitope recombinant vaccinia virus as a universal T cell vaccine candidate against influenza virus.

    Directory of Open Access Journals (Sweden)

    Alan G Goodman

    Full Text Available There is a need to develop a universal vaccine against influenza virus infection to avoid developing new formulations of a seasonal vaccine each year. Many of the vaccine strategies for a universal vaccine target strain-conserved influenza virus proteins, such as the matrix, polymerase, and nucleoproteins, rather than the surface hemagglutinin and neuraminidase proteins. In addition, non-disease-causing viral vectors are a popular choice as a delivery system for the influenza virus antigens. As a proof-of-concept, we have designed a novel influenza virus immunogen based on the NP backbone containing human T cell epitopes for M1, NS1, NP, PB1 and PA proteins (referred as NPmix as well as a construct containing the conserved regions of influenza virus neuraminidase (N-terminal and hemagglutinin (C-terminal (referred as NA-HA. DNA vectors and vaccinia virus recombinants expressing NPmix (WR-NP or both NPmix plus NA-HA (WR-flu in the cytosol were tested in a heterologous DNA-prime/vaccinia virus-boost vaccine regimen in mice. We observed an increase in the number of influenza virus-specific IFNγ-secreting splenocytes, composed of populations marked by CD4(+ and CD8(+ T cells producing IFNγ or TNFα. Upon challenge with influenza virus, the vaccinated mice exhibited decreased viral load in the lungs and a delay in mortality. These findings suggest that DNA prime/poxvirus boost with human multi-epitope recombinant influenza virus proteins is a valid approach for a general T-cell vaccine to protect against influenza virus infection.

  4. Induction of antibody responses to African horse sickness virus (AHSV in ponies after vaccination with recombinant modified vaccinia Ankara (MVA.

    Directory of Open Access Journals (Sweden)

    Rachael Chiam

    Full Text Available 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.

  5. HIV-1 vaccines based on replication-competent Tiantan vaccinia protected Chinese rhesus macaques from simian HIV infection.

    Science.gov (United States)

    Liu, Qiang; Li, Yue; Luo, Zhenwu; Yang, Guibo; Liu, Yong; Liu, Ying; Sun, Maosheng; Dai, Jiejie; Li, Qihan; Qin, Chuan; Shao, Yiming

    2015-03-27

    To assess the efficacy of HIV vaccines constructed from replication-competent Tiantan vaccinia virus (rTV) alone or combined with DNA in protecting Chinese rhesus macaques from homologous Simian/Human Immunodeficiency Virus (SHIV)-CN97001 challenge. The nef, gag, pol, and gp140 genes from strain CRF07_BC HIV-1 CN54 were selected to construct an HIV vaccine using the rTV or rTV/DNA vaccine. After vaccination, the vaccine and control groups were intravenously challenged with SHIV-CN97001 (32 MID50). HIV-specific antibodies and neutralizing antibodies, gp70 V1V2 binding antibodies, and cytotoxic T-lymphocyte responses were measured prospectively after vaccination with an ELISA, a virus infectivity assay in TZM-bl cells, and ELISPOT assays, respectively. Viral RNA was quantified after challenge with real-time reverse transcriptase-PCR (RT-PCR), and protection efficacy was determined with an analysis of CD8 lymphocyte depletion in vivo. Both rTV and DNA/rTV vaccine groups developed strong cellular and humoral responses against HIV-1 CN54 antigens, including Gag and Env, and also developed significant and persistent anti-Env antibodies and neutralizing antibodies after immunization. Both the rTV and DNA/rTV groups were significantly protected against SHIV-CN97001 or displayed lower viremia than the controls. After CD8 lymphocyte depletion, no viremia was detectable in the vaccinated monkeys, but rebounded rapidly in the control animals. Protection against infection correlated with vaccine-elicited neutralizing antibodies specific for homologous HIV-1 viruses. An rTV-based HIV-1 vaccine, with or without a DNA primer, provided protection from SHIV challenge in a macaque model. Replication-competent Tiantan vaccinia is a promising vector and should enable advances in HIV-1 vaccine development.

  6. A role for the H4 subunit of vaccinia RNA polymerase in transcription initiation at a viral early promoter.

    Science.gov (United States)

    Deng, L; Shuman, S

    1994-05-13

    The vaccinia virus H4 gene encodes an essential subunit of the DNA-dependent RNA polymerase holoenzyme encapsidated within virus particles (Ahn, B., and Moss, B. (1992) Proc. Natl. Acad. Sci. U.S.A. 89, 3536-3540; Kane, E. M., and Shuman, S. (1992) J. Virol. 66, 5752-5762). The role of this protein in transcription of viral early genes was revealed by the effects of affinity-purified anti-H4 antibody on discrete phases of the early transcription reaction in vitro. Anti-H4 specifically prevented the synthesis of a 21-nucleotide nascent RNA chain but had no impact on elongation of the 21-mer RNA by preassembled ternary complexes. Inhibition of initiation but not elongation was also observed with affinity-purified anti-D6 antibody directed against the 70-kDa subunit of the vaccinia early transcription initiation factor (ETF). Native gel mobility-shift assays showed that anti-H4 prevented the NTP-dependent recruitment of RNA polymerase to the preinitiation complex of ETF bound at the early promoter. Two species of ternary complexes could be resolved by native gel electrophoresis. Addition of anti-H4 to preformed complexes elicited a supershift of both ternary species but not of the preinitiation complex. Supeshift by anti-D6 revealed that the more rapidly migrating species of ternary complex did not contain immunoreactive ETF. Loss of ETF from the ternary complex was time-dependent. Thus, whereas the H4 protein was a stable constituent of the elongation complex, ETF was dissociable. We suggest that H4 functions as a molecular bridge to ETF and thereby allows specific recognition of early promoters by the core RNA polymerase. H4 is unlike bacterial sigma factor in that it remains bound to polymerase after the elongation complex is established.

  7. The Integration of Epistasis Network and Functional Interactions in a GWAS Implicates RXR Pathway Genes in the Immune Response to Smallpox Vaccine

    Science.gov (United States)

    McKinney, Brett A.; Lareau, Caleb; Oberg, Ann L.; Kennedy, Richard B.; Ovsyannikova, Inna G.; Poland, Gregory A.

    2016-01-01

    Although many diseases and traits show large heritability, few genetic variants have been found to strongly separate phenotype groups by genotype. Complex regulatory networks of variants and expression of multiple genes lead to small individual-variant effects and difficulty replicating the effect of any single variant in an affected pathway. Interaction network modeling of GWAS identifies effects ignored by univariate models, but population differences may still cause specific genes to not replicate. Integrative network models may help detect indirect effects of variants in the underlying biological pathway. In this study, we used gene-level functional interaction information from the Integrative Multi-species Prediction (IMP) tool to reveal important genes associated with a complex phenotype through evidence from epistasis networks and pathway enrichment. We test this method for augmenting variant-based network analyses with functional interactions by applying it to a smallpox vaccine immune response GWAS. The integrative analysis spotlights the role of genes related to retinoid X receptor alpha (RXRA), which has been implicated in a previous epistasis network analysis of smallpox vaccine. PMID:27513748

  8. One-step duplex polymerase chain reaction for the detection of swinepox and vaccinia viruses in skin lesions of swine with poxvirus-related disease.

    Science.gov (United States)

    Medaglia, Maria Luiza G; Sá, Natália M B; Correa, Isadora A; Costa, Luciana J; Damaso, Clarissa R

    2015-07-01

    Infection of pigs with swinepox virus (SWPV) was reported in Brazil in 2011. SWPV causes a systemic pustular disease in pigs and the symptoms are clinically indistinguishable from those caused by vaccinia virus (VACV) infection. Pigs infected with VACV have been reported in various countries; however, VACV is endemic in Brazil, India and other countries, where it affects mainly dairy cows, dairy buffaloes and dairy workers causing localized pustules. The transmission of VACV to other susceptible hosts has also been detected in Brazil. Therefore, VACV should be investigated as a possible etiologic agent of pustular skin disorders in pigs. This work describes the development of a one-step duplex assay to detect swinepox and vaccinia viruses simultaneously in skin lesions of pigs with generalized pustular disease. The investigation of VACV infection in pigs is important in countries where this zoonosis is endemic and should be differentiated from SWPV infection. Copyright © 2015 Elsevier B.V. All rights reserved.

  9. Regression of Human Prostate Tumors and Metastases in Nude Mice following Treatment with the Recombinant Oncolytic Vaccinia Virus GLV-1h68

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    Ivaylo Gentschev

    2010-01-01

    Full Text Available Virotherapy using oncolytic vaccinia virus strains is one of the most promising new strategies for cancer therapy. In the current study, we analyzed the therapeutic efficacy of the oncolytic vaccinia virus GLV-1h68 against two human prostate cancer cell lines DU-145 and PC-3 in cell culture and in tumor xenograft models. By viral proliferation assays and cell survival tests, we demonstrated that GLV-1h68 was able to infect, replicate in, and lyse these prostate cancer cells in culture. In DU-145 and PC-3 tumor xenograft models, a single intravenous injection with GLV-1h68 resulted in a significant reduction of primary tumor size. In addition, the GLV-1h68-infection led to strong inflammatory and oncolytic effects resulting in drastic reduction of regional lymph nodes with PC-3 metastases. Our data documented that the GLV-1h68 virus has a great potential for treatment of human prostate carcinoma.

  10. The epidemic of smallpox in Guangdong Province and its prevention and treatment in the Republican period%民国时期广东的天花流行与防治

    Institute of Scientific and Technical Information of China (English)

    陈丽楠; 刘玲娣

    2014-01-01

    民国时期,广东几乎每年都有天花疫情发生.为了应对传染病,广东省建立了卫生行政及传染病防治机构,并自制牛痘疫苗;相关机构按周、月、年对疫情进行统计,并定期上报,以了解疫情进展.同时,还成立了传染病医院收容天花患者.港口检疫则将天花列为重点检查对象.为了推广牛痘接种,政府与社会合作,组织大规模的“种痘运动”,以儿童、学生和年轻人作为主要接种对象,并借助媒体积极宣传天花防治知识.%During the Republican period,the smallpox broke out in high frequency almost annually in Guangdong.In response to infectious diseases,the Guangdong government established the Health Administration and institutions for infectious disease's prevention and treatment,and prepared smallpox vaccine by themselves.In order to grasp the situation of the epidenmic,related institutions collected epidemic data weekly,monthly,and annually with the statistics reported regularly.Meanwhile,infectious diseasc hospital was established for smallpox patients.Harbor quarantine put smallpox as a key target of inspection.With the joint effort of the government and social organizations,massive "vaccination campaigns" was organized to promote vaccination,in which children,students and young people were the main subjects for inoculations.Prevention knowledge and anti-epidemic concept towards smallpox have been actively publicized and improved by media.

  11. Suitability of vaccinia virus and bovine viral diarrhea virus (BVDV for determining activities of three commonly-used alcohol-based hand rubs against enveloped viruses

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    Steinmann Jochen

    2007-02-01

    Full Text Available Abstract Background A procedure for including activity against enveloped viruses in the post-contamination treatment of hands has been recommended, but so far no European standard is available to implement it. In 2004, the German Robert Koch-Institute (RKI and the German Association for the Control of Virus Disease (DVV suggested that vaccinia virus and bovine viral diarrhea virus (BVDV should be used as test viruses in a quantitative suspension test to determine the activity of a disinfectant against all enveloped viruses. Methods We have studied the activities of three commonly-used alcohol-based hand rubs (hand rub A, based on 45% propan-2-ol, 30% propan-1-ol and 0.2% mecetronium etilsulfate; hand rub B, based on 80% ethanol; hand rub C, based on 95% ethanol against vaccinia virus and BVDV, and in addition against four other clinically relevant enveloped viruses: herpes simplex virus (HSV types 1 and 2, and human and avian influenza A virus. The hand rubs were challenged with different organic loads at exposure time of 15, 30 and 60 s. According to the guidelines of both BGA/RKI and DVV, and EN 14476:2005, the reduction of infectivity of each test virus was measured on appropriate cell lines using a quantitative suspension test. Results All three alcohol-based hand rubs reduced the infectivity of vaccinia virus and BVDV by ≥ 4 log10-steps within 15 s, irrespective of the type of organic load. Similar reductions of infectivity were seen against the other four enveloped viruses within 15 s in the presence of different types of organic load. Conclusion Commonly used alcohol-based hand rubs with a total alcohol concentration ≥ 75% can be assumed to be active against clinically relevant enveloped viruses if they effectively reduce the infectivities of vaccinia virus and BVDV in a quantitative suspension test.

  12. Immunogenicity in mice and rhesus monkeys vaccinated with recombinant vaccinia virus expressing bivalent E7E6 fusion proteins from human papillomavirus types 16 and 18

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    Tian Houwen

    2011-06-01

    Full Text Available Abstract Background Persistent infection with high-risk human papillomavirus (HPV is a predominant cause of cervical cancer, and HPV16 and HPV18 occur in 50% and 20% of cervical cancer cases, respectively. The viral oncogenes E6 and E7 are constitutively expressed by HPV-associated tumour cells and can therefore be used as target antigens for immunotherapy. In this study, we constructed a recombinant vaccinia virus co-expressing the HPV16/18 E7E6 fusion proteins (rVVJ16/18E7E6 for use as a therapeutic vaccine for the treatment of HPV16+ and HPV18+ cancers. Methods We constructed a bivalent recombinant vaccinia virus expressing modified E7E6 fusion proteins of HPV type 16 and 18 (rVVJ16/18E7E6 based on the vaccinia virus Tiantan strain. We then defined the cellular immune responses to the virus in mice and rhesus monkeys and assessed antitumour efficacy of these responses in mice using the TC-1 tumour challenge model. Results Our data demonstrated that rVVJ16/18E7E6 was able to elicit varying levels of CD8+ T cell immune responses and lysis of target cells in mice in response to peptides HPV16E749-57 and HPV18E667-75. Furthermore, the virus was also able to induce anti-tumour responses in the HPV16+ TC-1 tumour challenge model, including partial protection (30-40% and delayed tumour appearance. In addition, the virus was able to induce immune responses in rhesus monkeys. Conclusions The recombinant vaccinia virus rVVJ16/18E7E6 can generate clear and significant cellular immunity in both mice and rhesus monkeys. These data provide a basis for the use of this recombinant virus as a potential vaccine candidate for further study.

  13. Recombinant Vaccinia Virus is an Effective and Non-perturbing Vector for Human Dendritic Cells Transfected with Epstein-Barr Virus Latent Membrane Protein 2A

    Institute of Scientific and Technical Information of China (English)

    许继军; 姚堃; 彭光勇; 谢芳艺; 丁传林; 朱建中; 秦健

    2002-01-01

    ObjectiveTo study the effects of dendritic cells (DC) transfected with recombinant vaccinia virus encoding Epstein-Barr virus (EBV) latent membrane protein 2A(LMP2A) gene,and to provide evidence for further investigation on the therapeutic vaccines against EBV-associated malignancies.MethodsMature DC were transfected with EBV-LMP2A recombinant vaccinia virus (rVV-LMP2A).Before and after the transfection,the expression of surface antigens on mature DC including CD1a,CD83,CD40,CD80,HLA-DR was measured by fluorescence activated cell sorter (FACS) and the function of DC to stimulate allogeneic T cells proliferation was measured by mixed leukocyte reactions (MLR).ResultsLMP2A protein was highly expressed (66.1%) in DC after the transfection of rVV-LMP2A.No significant changes in the primary surface antigens expression and in the MLR were detected during the transfection.Transfected DC still had strong potential in stimulating the proliferation of allogeneic T cells.ConclusionRecombinant vaccinia virus was an effective and non-perturbing vector to mediate the transfection of LMP2A into DC.The functions of mature DC were not affected significantly by the transfection of Vac-LMP2A.This study could provide evidence for the further immunotherapy of EBV-associated malignancies,e.g.nasopharyngeal carcinoma (NPC).``

  14. Recombinant Vaccinia Virus is an Effective and Non—perturbing Vector for Human Dendritic Cells Transfected with Epstein—Barr Virus Latent Membrane Protein 2A

    Institute of Scientific and Technical Information of China (English)

    许继军; 姚Kun; 等

    2002-01-01

    Objective To study the effects of dendritic cells(DC) transfected with recombinant vaccinia virus encoding Epstein-Barr virus(EBV) latent membrane protein 2A(LMP2A) gene,and to provide evidence for further investigation on the therapeutic uaccines against EBV-associated malignancies.Methods Mature DC were transfected with EVB-LMP2A recombinant vaccinia virus(rVV-LMP2A).Before and after the transfection,the expression of surface antigens on mature DC including CD1a,CD83,CD40,CD80,HLA-DR was measured by fluorescence activated cell sorter(FACS) and the function of DC to stimulate allogeneic T cells proliferation was measured by mixed leukocyte reactions(MLR).Results LMP2A protein was highly expressed (66.1%) in DC after the transfection of rVV-LMP2A.No significant changes in the primary surface antigens expression and in the MLR were detected during the transfection.Transfected DC still had strong potential in stimulating the proliferation of allogeneic T cells.Conclusion Recombinant vaccinia virus was an effective and non-perturbing vector to mediate the transfection of LMP2A into DC.The functions of mature DC were not affected significantly by the transfection of Vac-LMP2A.This study could provide evidence for the further immunotherapy of EBV-associated malignancies,e.g.nasopharyngeal carcinoma(NPC).

  15. The hemagglutinin-neuraminidase (HN) gene of Newcastle disease virus strain Italien (ndv Italien): comparison with HNs of other strains and expression by a vaccinia recombinant.

    Science.gov (United States)

    Wemers, C D; de Henau, S; Neyt, C; Espion, D; Letellier, C; Meulemans, G; Burny, A

    1987-01-01

    A cDNA library was constructed with poly(A+) mRNA from cells infected with the virulent Italien NDV strain. A clone that hybridized to the HN gene mRNA was sequenced. A long open reading-frame encodes for a protein of 571 amino acids, with a calculated molecular weight of 61,900, including 13 cysteine residues and six potential glycosylation sites. To define the sequence changes that occurred in the avian paramyxovirus hemagglutinin-neuraminidase (HN) during the evolution of virulence, we have studied the HNs of the virulent Italien NDV strain, the mesovirulent Beaudette strain and the nonvirulent Hitchner strain. The majority of amino acid variations are conservative changes but they cluster at 4 preferential sites in the putative head of HN. The clusters of amino acid substitutions are intimately associated or overlap with regions of HN rich in charged amino acid residues and in cysteines. The latter are conserved not only between HNs from all 3 NDV strains but also between HNs of 4 different paramyxoviruses, NDV, SV 5, Sendai and PI 3. The HN coding sequence was inserted into the genome of vaccinia virus under the control of vaccinia P 7.5 K transcriptional regulatory sequences. Expression of native HN proteins at the surface of recombinant HN vaccinia-infected cells was demonstrated by indirect immunofluorescence with 2 anti-HN monoclonals.

  16. Comparison on virulence and immunogenicity of two recombinant vaccinia vaccines, Tian Tan and Guang9 strains, expressing the HIV-1 envelope gene.

    Science.gov (United States)

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

    2012-01-01

    The vaccinia virus Guang9 strain (VG9), derived from the vaccinia virus Tian Tan strain (VTT) has been found to be less virulent than VTT. 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-γ response which represented cellular immune response induced by VG9-E was significantly higher than that did by VTT-E. 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.

  17. Recombination-mediated genetic engineering of a bacterial artificial chromosome clone of modified vaccinia virus Ankara (MVA.

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    Matthew G Cottingham

    Full Text Available 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, PNAS99 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-gamma, TNF-alpha 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

  18. Vaccinia virus-mediated intra-tumoral expression of matrix metalloproteinase 9 enhances oncolysis of PC-3 xenograft tumors

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    Schäfer Simon

    2012-08-01

    Full Text Available Abstract Background Oncolytic viruses, including vaccinia virus (VACV, are a promising alternative to classical mono-cancer treatment methods such as surgery, chemo- or radiotherapy. However, combined therapeutic modalities may be more effective than mono-therapies. In this study, we enhanced the effectiveness of oncolytic virotherapy by matrix metalloproteinase (MMP-9-mediated degradation of proteins of the tumoral extracellular matrix (ECM, leading to increased viral distribution within the tumors. Methods For this study, the oncolytic vaccinia virus GLV-1h255, containing the mmp-9 gene, was constructed and used to treat PC-3 tumor-bearing mice, achieving an intra-tumoral over-expression of MMP-9. The intra-tumoral MMP-9 content was quantified by immunohistochemistry in tumor sections. Therapeutic efficacy of GLV-1h255 was evaluated by monitoring tumor growth kinetics and intra-tumoral virus titers. Microenvironmental changes mediated by the intra-tumoral MMP-9 over-expression were investigated by microscopic quantification of the collagen IV content, the blood vessel density (BVD and the analysis of lymph node metastasis formation. Results GLV-1h255-treatment of PC-3 tumors led to a significant over-expression of intra-tumoral MMP-9, accompanied by a marked decrease in collagen IV content in infected tumor areas, when compared to GLV-1h68-infected tumor areas. This led to considerably elevated virus titers in GLV-1h255 infected tumors, and to enhanced tumor regression. The analysis of the BVD, as well as the lumbar and renal lymph node volumes, revealed lower BVD and significantly smaller lymph nodes in both GLV-1h68- and GLV-1h255- injected mice compared to those injected with PBS, indicating that MMP-9 over-expression does not alter the metastasis-reducing effect of oncolytic VACV. Conclusions Taken together, these results indicate that a GLV-1h255-mediated intra-tumoral over-expression of MMP-9 leads to a degradation of collagen IV

  19. [Strategies, actors, promises and fears in the smallpox vaccinations campaigns in Mexico: from the Porfiriato to the Post-revolution (1880-1940)].

    Science.gov (United States)

    Agostoni, Claudia

    2011-02-01

    The article examines some of the strategies employed by the Mexican health authorities that led to the organization of massive and obligatory smallpox vaccination campaigns from the late 1880s to the 1940s, a period of Mexican history that corresponds to the Porfirio Díaz regime (1877-1911), to the armed phase of the Mexican Revolution (1910-1920), and to the first two decades of the Post-revolutionary governments (1920-1940). Attention will be placed of the vaccination programs in the main urban settings, notably in Mexico City, as well as the gradual but decisive organization and regulation of vaccination campaigns in the heterogeneous rural milieu. Furthermore, the importance that hygienic education acquired will be explored, as well as the divergent and contested responses that emerged due to the obligatory vaccination campaigns, responses that included resistance, fear, uncertainty and widespread acceptance.

  20. Immunogenicity of Attenuated Vaccinia Virus Guang 9 Strain%痘苗病毒弱毒株广9株的免疫原性分析

    Institute of Scientific and Technical Information of China (English)

    朱蓉; 黄维金; 王佑春; 俞永新

    2011-01-01

    目的 分析痘苗病毒弱毒株广9株(Vaccinia virus Guang 9 strain,VG9)对BALB/c小鼠的免疫原性.方法 用鸡胚成纤维细胞培养痘苗病毒VG9株和天坛株(Vaccinia virus Tian Tan strain,VTT),离心纯化病毒,经小鼠背部皮内免疫后,每周采血,共4次,采用ELISA法检测小鼠血清中抗痘苗病毒特异性抗体效价;中和试验检测免疫后3周和4周小鼠血清中和抗体效价.结果 两株痘苗病毒免疫后1周,小鼠血清中即产生抗痘苗病毒特异性抗体,效价均达1∶4050;至第3周,抗体效价上升至1∶12 150;第4周时,VG9株抗体效价维持在1∶12 150,而VTT株效价可达1:36 450.两株病毒诱导的中和抗体水平均在免疫后3周达峰值,且具有交叉中和作用,均对VTT株中和效果更好,VG9株诱导产生中和抗体的能力较VTT株略低.结论 痘苗病毒VG9株与VTT株一样,具有良好的免疫原性,皮内免疫均能刺激小鼠产生良好的抗痘菌病毒特异性抗体和中和抗体.%Objective To analyze the immunogenicity of attenuated vaccinia virus Guang 9 (VG9) strain. Methods VG9 and Vaccinia virus Tian Tan (VTT) strains were cultured in chick embryo fibroblasts (CEFs), purified by centrifugation and inoculated I.d. Into BALB /c mice. Serum samples were collected once a week for 4 times, and determined for specific antibody liter against vaccinia virus by ELJSA. The neutralizing antibody liters in sera 3 and 4 weeks after immunization were determined by neutralization test. Results Specific antibody liters against vaccinia virus in sera of mice were 1 : 4 050 one week after immunization with both the strains and increased to 1 : 12 150 on week 3. However, the antibody titers against VG9 and VTT strains on week 4 after immunization were 1 : 12 150 and 1 : 36 450 respectively. Both the neutralizing antibody levels induced by the two strains reached peak values 3 weeks after immunization, and cross neutralization was observed. The neutralizing effect on

  1. Smallpox Antiviral Drug

    Science.gov (United States)

    2007-01-01

    strategies is essential. ACKNOWLEDGMENTS We thank Robert Jordan, Sean Amberg, and Tove’ Bolken for critical reviews of the manuscript. REFERENCES...part II. Adverse events. Clin Infect Dis 37:251–271. Ghosn J, Chaix ML, Peytavin G, Rey E, Bresson JL, Goujard C, Katlama C, Viard JP, Treluyer JM

  2. Smallpox Vaccine Overview

    Science.gov (United States)

    ... Agent and Organophosphate Pesticide Poisoning Sarin Soman Tabun VX Nicotine Case Definition: Nicotine Poisoning Nitrogen mustard (HN- ... Poisoning Unidentified chemical Vesicants/blister agents Vomiting agents VX Facts About VX Case Definition: Nerve Agents or ...

  3. Live Virus Smallpox Vaccine

    Science.gov (United States)

    ... Agent and Organophosphate Pesticide Poisoning Sarin Soman Tabun VX Nicotine Case Definition: Nicotine Poisoning Nitrogen mustard (HN- ... Poisoning Unidentified chemical Vesicants/blister agents Vomiting agents VX Facts About VX Case Definition: Nerve Agents or ...

  4. Vaccinia-related kinase 1 is required for the maintenance of undifferentiated spermatogonia in mouse male germ cells.

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    Yoon Ha Choi

    Full Text Available Vaccinia-related kinase 1 (VRK1 is a crucial protein kinase for mitotic regulation. VRK1 is known to play a role in germ cell development, and its deficiency results in sterility. Here we describe that VRK1 is essential for the maintenance of spermatogonial stem cells. To determine whether VRK1 plays a role in these cells, we assessed the population size of undifferentiated spermatogonia. Flow cytometry analyses showed that the number of undifferentiated spermatogonia was markedly reduced in VRK1-deficient testes. VRK1 was highly expressed in spermatogonial populations, and approximately 66% of undifferentiated spermatogonia that were sorted as an Ep-CAM+/c-kit-/alpha-6-integrin+ population showed a positive signal for VRK1. Undifferentiated stem cells expressing Plzf and Oct4 but not c-kit also expressed VRK1, suggesting that VRK1 is an intrinsic factor for the maintenance of spermatogonial stem cells. Microarray analyses of the global testicular transcriptome and quantitative RT-PCR of VRK1-deficient testes revealed significantly reduced expression levels of undifferentiated spermatogonial marker genes in early postnatal mice. Together, these results suggest that VRK1 is required for the proliferation and differentiation of undifferentiated spermatogonia, which are essential for spermatogenic cell maintenance.

  5. Recombinant modified vaccinia virus Ankara expressing glycoprotein E2 of Chikungunya virus protects AG129 mice against lethal challenge.

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    Petra van den Doel

    2014-09-01

    Full Text Available Chikungunya virus (CHIKV infection is characterized by rash, acute high fever, chills, headache, nausea, photophobia, vomiting, and severe polyarthralgia. There is evidence that arthralgia can persist for years and result in long-term discomfort. Neurologic disease with fatal outcome has been documented, although at low incidences. The CHIKV RNA genome encodes five structural proteins (C, E1, E2, E3 and 6K. The E1 spike protein drives the fusion process within the cytoplasm, while the E2 protein is believed to interact with cellular receptors and therefore most probably constitutes the target of neutralizing antibodies. We have constructed recombinant Modified Vaccinia Ankara (MVA expressing E3E2, 6KE1, or the entire CHIKV envelope polyprotein cassette E3E26KE1. MVA is an appropriate platform because of its demonstrated clinical safety and its suitability for expression of various heterologous proteins. After completing the immunization scheme, animals were challenged with CHIV-S27. Immunization of AG129 mice with MVAs expressing E2 or E3E26KE1 elicited neutralizing antibodies in all animals and provided 100% protection against lethal disease. In contrast, 75% of the animals immunized with 6KE1 were protected against lethal infection. In conclusion, MVA expressing the glycoprotein E2 of CHIKV represents as an immunogenic and effective candidate vaccine against CHIKV infections.

  6. Immunogenicity and protective efficacy of recombinant Modified Vaccinia virus Ankara candidate vaccines delivering West Nile virus envelope antigens.

    Science.gov (United States)

    Volz, Asisa; Lim, Stephanie; Kaserer, Martina; Lülf, Anna; Marr, Lisa; Jany, Sylvia; Deeg, Cornelia A; Pijlman, Gorben P; Koraka, Penelope; Osterhaus, Albert D M E; Martina, Byron E; Sutter, Gerd

    2016-04-07

    West Nile virus (WNV) cycles between insects and wild birds, and is transmitted via mosquito vectors to horses and humans, potentially causing severe neuroinvasive disease. Modified Vaccinia virus Ankara (MVA) is an advanced viral vector for developing new recombinant vaccines against infectious diseases and cancer. Here, we generated and evaluated recombinant MVA candidate vaccines that deliver WNV envelope (E) antigens and fulfil all the requirements to proceed to clinical testing in humans. Infections of human and equine cell cultures with recombinant MVA demonstrated efficient synthesis and secretion of WNV envelope proteins in mammalian cells non-permissive for MVA replication. Prime-boost immunizations in BALB/c mice readily induced circulating serum antibodies binding to recombinant WNV E protein and neutralizing WNV in tissue culture infections. Vaccinations in HLA-A2.1-/HLA-DR1-transgenic H-2 class I-/class II-knockout mice elicited WNV E-specific CD8+ T cell responses. Moreover, the MVA-WNV candidate vaccines protected C57BL/6 mice against lineage 1 and lineage 2 WNV infection and induced heterologous neutralizing antibodies. Thus, further studies are warranted to evaluate these recombinant MVA-WNV vaccines in other preclinical models and use them as candidate vaccine in humans.

  7. Human and animal infections by vaccinia-like viruses in the state of Rio de Janeiro: a novel expanding zoonosis.

    Science.gov (United States)

    Schatzmayr, H G; Costa, R V C; Gonçalves, M C R; D'Andréa, P S; Barth, O M

    2011-12-30

    Since 1999, vesicular infections caused by Orthopoxvirus in humans and animals, mainly in dairy cattle, have been identified in 20 municipalities in the Rio de Janeiro state of Brazil. This paper describes studies conducted in counties of the northwestern, middle-Paraíba Valley and southern regions of the Rio de Janeiro state where 77 human, 346 bovine and 78 rodent samples were collected over the past ten years. Laboratory investigations using virus isolation, electron microscopy, molecular biology (PCR) and serological analysis confirmed Orthopoxvirus infections in 77.9% of human, 49.2% of dairy cattle and 17.9% of rodent samples. The characterisation of the Cantagalo/IOC strain reconfirmed that this virus was a vaccinia-like virus. In other regions of the Rio de Janeiro state, vesicular/pustular infections in animals and humans are suspected but these have not yet been confirmed. A continuous surveillance system has been established to monitor these regions in addition to several other states of the Brazilian Federation.

  8. Serro 2 Virus Highlights the Fundamental Genomic and Biological Features of a Natural Vaccinia Virus Infecting Humans

    Science.gov (United States)

    Trindade, Giliane de Souza; Emerson, Ginny L.; Sammons, Scott; Frace, Michael; Govil, Dhwani; Fernandes Mota, Bruno Eduardo; Abrahão, Jônatas Santos; de Assis, Felipe Lopes; Olsen-Rasmussen, Melissa; Goldsmith, Cynthia S.; Li, Yu; Carroll, Darin; Guimarães da Fonseca, Flavio; Kroon, Erna; Damon, Inger K.

    2016-01-01

    Vaccinia virus (VACV) has been implicated in infections of dairy cattle and humans, and outbreaks have substantially impacted local economies and public health in Brazil. During a 2005 outbreak, a VACV strain designated Serro 2 virus (S2V) was collected from a 30-year old male milker. Our aim was to phenotypically and genetically characterize this VACV Brazilian isolate. S2V produced small round plaques without associated comets when grown in BSC40 cells. Furthermore, S2V was less virulent than the prototype strain VACV-Western Reserve (WR) in a murine model of intradermal infection, producing a tiny lesion with virtually no surrounding inflammation. The genome of S2V was sequenced by primer walking. The coding region spans 184,572 bp and contains 211 predicted genes. Mutations in envelope genes specifically associated with small plaque phenotypes were not found in S2V; however, other alterations in amino acid sequences within these genes were identified. In addition, some immunomodulatory genes were truncated in S2V. Phylogenetic analysis using immune regulatory-related genes, besides the hemagglutinin gene, segregated the Brazilian viruses into two clusters, grouping the S2V into Brazilian VACV group 1. S2V is the first naturally-circulating human-associated VACV, with a low passage history, to be extensively genetically and phenotypically characterized. PMID:27973399

  9. Efficacy and safety/toxicity study of recombinant vaccinia virus JX-594 in two immunocompetent animal models of glioma.

    Science.gov (United States)

    Lun, XueQing; Chan, Jennifer; Zhou, Hongyuan; Sun, Beichen; Kelly, John J P; Stechishin, Owen Owen; Bell, John C; Parato, Kelley; Hu, Kang; Vaillant, Dominique; Wang, Jiahu; Liu, Ta-Chiang; Breitbach, Caroline; Kirn, David; Senger, Donna L; Forsyth, Peter A

    2010-11-01

    The purpose of this study was to investigate the oncolytic potential of the recombinant, granulocyte macrophage colony-stimulating factor (GM-CSF)-expressing vaccinia virus (VV) JX-594 in experimental malignant glioma (MGs) in vitro and in immunocompetent rodent models. We have found that JX-594 killed all MG cell lines tested in vitro. Intratumoral (i.t.) administration of JX-594 significantly inhibited tumor growth and prolonged survival in rats-bearing RG2 intracranial (i.c.) tumors and mice-bearing GL261 brain tumors. Combination therapy with JX-594 and rapamycin significantly increased viral replication and further prolonged survival in both immunocompetent i.c. MG models with several animals considered "cured" (three out of seven rats >120 days, terminated experiment). JX-594 infected and killed brain tumor-initiating cells (BTICs) from patient samples grown ex vivo, and did so more efficiently than other oncolytic viruses MYXV, Reovirus type-3, and VSV(ΔM51). Additional safety/toxicity studies in nontumor-bearing rodents treated with a supratherapeutic dose of JX-594 demonstrated GM-CSF-dependent inflammation and necrosis. These results suggest that i.c. administered JX-594 triggers a predictable GM-CSF-mediated inflammation in murine models. Before proceeding to clinical trials, JX-594 should be evaluated in the brains of nonhuman primates and optimized for the viral doses, delivery routes as well as the combination agents (e.g., mTOR inhibitors).

  10. Azathioprine inhibits vaccinia virus replication in both BSC-40 and RAG cell lines acting on different stages of virus cycle.

    Science.gov (United States)

    Damaso, Clarissa R A; Oliveira, Marcus F; Massarani, Susana M; Moussatché, Nissin

    2002-08-15

    In the present study we demonstrate that azathioprine (AZA) inhibits vaccinia virus (VV) replication in both BSC-40 and RAG cell lines, acting on different stages of virus cycle. In BSC-40 cells, early protein synthesis was not significantly affected, but late gene expression was severely impaired. In RAG cells all stages of gene expression were completed during synchronous infection in the presence of the drug. The onset of DNA replication was not affected in RAG cells, but a severe inhibition was observed in BSC-40 cells. Electron microscopic analysis of VV-infected RAG cells treated with AZA revealed brick-shaped particles presenting abnormal definition of the internal structure. Purified virions from AZA-treated RAG cells presented several modifications of the protein content, a lesser amount of DNA, and a lower PFU:particle ratio. Our results suggest that in VV-infected RAG cells AZA interfered with virus morphogenesis, whereas in BSC-40 cells the replicative cycle was inhibited at the DNA replication stage.

  11. Immunogenicity analysis following human immunodeficiency virus recombinant DNA and recombinant vaccinia virus Tian Tan prime-boost immunization.

    Science.gov (United States)

    Liu, Cunxia; Du, Shouwen; Li, Chang; Wang, Yuhang; Wang, Maopeng; Li, Yi; Yin, Ronglan; Li, Xiao; Ren, Dayong; Qin, Yanqing; Ren, Jingqiang; Jin, Ningyi

    2013-06-01

    This study assessed and compared the immunogenicity of various immunization strategies in mice using combinations of recombinant DNA (pCCMp24) and recombinant attenuated vaccinia virus Tian Tan (rddVTT-CCMp24). Intramuscular immunization was performed on days 0 (prime) and 21 (boost). The immunogenicity of the vaccine schedules was determined by measuring human immunodeficiency virus (HIV)-specific binding antibody levels and cytokine (interleukin-2 and interleukin-4) concentrations in peripheral blood, analyzing lymphocyte proliferation capacity against HIV epitopes and CD4(+)/CD8(+) cell ratio, and monitoring interferon-gamma levels at different times post-immunization. The results showed that pCCMp24, rddVTT-CCMp24 and their prime-boost immunization induced humoral and cellular immune responses. The pCCMp24/rddVTT-CCMp24 immunization strategy increased CD8(+) T cells and induced more IFN-γ-secreting cells compared with single-shot rDNA. The prime-boost immunization strategy also induced the generation of cellular immunological memory to HIV epitope peptides. These results demonstrated that prime-boost immunization with rDNA and rddVTT-CCMp24 had a tendency to induce greater cellular immune response than single-shot vaccinations, especially IFN-γ response, providing a basis for further studies.

  12. Transcription and translation mapping of the 13 genes in the vaccinia virus HindIII D fragment.

    Science.gov (United States)

    Lee-Chen, G J; Niles, E G

    1988-03-01

    The vaccinia virus HindIII D fragment is 160,060 bp in length and encodes 13 complete open reading frames [Niles et al. (1986) Virology 153, 96-112; S. L. Weinrich and D. E. Hruby (1986). Nucleic Acids Res. 14, 3003-3016]. We have employed a two-step Northern hybridization protocol using single-stranded DNA probes from M13 recombinants in order to identify the mRNA products from the 13 genes. Six of these genes are expressed only at early times after infection; six are transcribed only at late times; one gene is expressed at both early and late times after virus infection. The D11 gene is transcribed into two late mRNA species, one full-length and the other derived from the 3' one-third of the coding sequence. Translation of hybrid-selected mRNA was carried out in an attempt to identify the protein products encoded by each mRNA. Protein products were found for each early gene but translation was successful for only two of the eight late mRNAs. With the completion of the physical map it is apparent that the early and late genes in the HindIII D fragment are arranged in order to minimize potential interference caused by the expression of closely packed viral genes.

  13. A genotype of modified vaccinia Ankara (MVA) that facilitates replication in suspension cultures in chemically defined medium.

    Science.gov (United States)

    Jordan, Ingo; Horn, Deborah; John, Katrin; Sandig, Volker

    2013-01-21

    While vectored vaccines, based on hyperattenuated viruses, may lead to new treatment options against infectious diseases and certain cancers, they are also complex products and sometimes difficult to provide in sufficient amount and purity. To facilitate vaccine programs utilizing host-restricted poxviruses, we established avian suspension cell lines (CR and CR.pIX) and developed a robust, chemically defined, culturing process for production of this class of vectors. For one prominent member, modified vaccinia Ankara (MVA), we now describe a new strain that appears to replicate to greater yields of infectious units, especially in the cell-free supernatant of cultures in chemically defined media. The new strain was obtained by repeated passaging in CR suspension cultures and, consistent with reports on the exceptional genetic stability of MVA, sequencing of 135 kb of the viral genomic DNA revealed that only three structural proteins (A3L, A9L and A34R) each carry a single amino acid exchange (H639Y, K75E and D86Y, respectively). Host restriction in a plaque-purified isolate of the new genotype appears to be maintained in cell culture. Processing towards an injectable vaccine preparation may be simplified with this strain as a complete lysate, containing the main burden of host cell contaminants, may not be required anymore to obtain adequate yields.

  14. Systemic treatment of xenografts with vaccinia virus GLV-1h68 reveals the immunologic facet of oncolytic therapy

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    Liu Hui

    2009-07-01

    Full Text Available Abstract Background GLV-1h68 is an attenuated recombinant vaccinia virus (VACV that selectively colonizes established human xenografts inducing their complete regression. Results Here, we explored xenograft/VACV/host interactions in vivo adopting organism-specific expression arrays and tumor cell/VACV in vitro comparing VACV replication patterns. There were no clear-cut differences in vitro among responding and non-responding tumors, however, tumor rejection was associated in vivo with activation of interferon-stimulated genes (ISGs and innate immune host's effector functions (IEFs correlating with VACV colonization of the xenografts. These signatures precisely reproduce those observed in humans during immune-mediated tissue-specific destruction (TSD that causes tumor or allograft rejection, autoimmunity or clearance of pathogens. We recently defined these common pathways in the "immunologic constant of rejection" hypothesis (ICR. Conclusion This study provides the first prospective validation of a universal mechanism associated with TSD. Thus, xenograft infection by oncolytic VACV, beyond offering a promising therapy of established cancers, may represent a reliable pre-clinical model to test therapeutic strategies aimed at modulating the central pathways leading to TSD; this information may lead to the identification of principles that could refine the treatment of cancer and chronic infection by immune stimulation or autoimmunity and allograft rejection through immune tolerance.

  15. Human vaccinia-like virus outbreaks in São Paulo and Goiás States, Brazil: virus detection, isolation and identification Surtos de vírus Vaccinia-like nos Estados de São Paulo e Goiás, Brasil: detecção, isolamento e identificação viral

    Directory of Open Access Journals (Sweden)

    Teresa Keico Nagasse-Sugahara

    2004-04-01

    Full Text Available Since October 2001, the Adolfo Lutz Institute has been receiving vesicular fluids and scab specimens of patients from Paraíba Valley region in the São Paulo and Minas Gerais States and from São Patricio Valley, in the Goiás State. Epidemiological data suggested that the outbreaks were caused by Cowpox virus or Vaccinia virus. Most of the patients are dairy milkers that had vesiculo-pustular lesions on the hands, arms, forearms, and some of them, on the face. Virus particles with orthopoxvirus morphology were detected by direct electron microscopy (DEM in samples of 49 (66.21% patients of a total of 74 analyzed. Viruses were isolated in Vero cell culture and on chorioallantoic membrane (CAM of embryonated chicken eggs. Among 21 samples submitted to PCR using primers for hemagglutinin (HA gene, 19 were positive. Restriction digestion with TaqI resulted in four characteristic Vaccinia virus fragments. HA nucleotide sequences showed 99.9% similarity with Cantagalo virus, described as a strain of Vaccinia virus. The only difference observed was the substitution of one nucleotide in the position 616 leading to change in one amino acid of the protein in the position 206. The phylogenetic analysis showed that the isolates clustered together with Cantagalo virus, other Vaccinia strains and Rabbitpox virus.A partir de outubro de 2001, o Instituto Adolfo Lutz tem recebido amostras de líquido vesicular e crostas de lesões de pele de pacientes das regiões do Vale do Paraíba, Estado de São Paulo e do Vale do São Patricio, Estado de Goiás. Os dados clínicos e epidemiológicos sugeriam que os surtos poderiam ser causados por Cowpox virus ou Vaccinia virus. A maioria dos pacientes era ordenhadores que tinham lesões vesicopustulares nas mãos, braços, antebraços e alguns na face. A análise por microscopia eletrônica direta (MED detectou partículas com morfologia de vírus do gênero Orthopoxvirus em amostras de 49 (66,21% pacientes dos 74

  16. Deletion of C7L and K1L genes leads to significantly decreased virulence of recombinant vaccinia virus TianTan.

    Science.gov (United States)

    Liu, Zheng; Wang, Shuhui; Zhang, Qicheng; Tian, Meijuan; Hou, Jue; Wang, Rongmin; Liu, Chang; Ji, Xu; Liu, Ying; Shao, Yiming

    2013-01-01

    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.

  17. Deletion of C7L and K1L genes leads to significantly decreased virulence of recombinant vaccinia virus TianTan.

    Directory of Open Access Journals (Sweden)

    Zheng Liu

    Full Text Available 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.

  18. A selective Seoul-Fluor-based bioprobe, SfBP, for vaccinia H1-related phosphatase--a dual-specific protein tyrosine phosphatase.

    Science.gov (United States)

    Jeong, Myeong Seon; Kim, Eunha; Kang, Hyo Jin; Choi, Eun Joung; Cho, Alvin R; Chung, Sang J; Park, Seung Bum

    2012-07-04

    We report a Seoul-Fluor-based bioprobe, SfBP, for selective monitoring of protein tyrosine phosphatases (PTPs). A rational design based on the structures at the active site of dual-specific PTPs can enable SfBP to selectively monitor the activity of these PTPs with a 93-fold change in brightness. Moreover, screening results of SfBP against 30 classical PTPs and 35 dual-specific PTPs show that it is selective toward vaccinia H1-related (VHR) phosphatase, a dual-specific PTP (DUSP-3).

  19. HIVIS-DNA or HIVISopt-DNA priming followed by CMDR vaccinia-based boosts induce both humoral and cellular murine immune responses to HIV

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    J Hinkula

    2017-06-01

    Conclusions: HIVIS-DNA was modified to obtain HIVISopt-DNA that had fewer plasmids, and additional epitopes. Even with one DNA prime followed by two MVA-CMDR boosts, humoral and cell-mediated immune responses were readily induced by priming with either DNA construct composition. Priming by HIV-DNA augmented neutralizing antibody responses revealed by boosting with the vaccinia-based heterologous sequences. Cellular and antibody responses covered selected strains representing HIV-1 subtypes A, B, C and CRF01_AE. We assume this is related to the inclusion of heterologous full genes in the vaccine schedule.

  20. A Candidate HIV/AIDS Vaccine (MVA-B) Lacking Vaccinia Virus Gene C6L Enhances Memory HIV-1-Specific T-Cell Responses

    OpenAIRE

    Juan García-Arriaza; José Luis Nájera; Carmen E Gómez; Nolawit Tewabe; Sorzano, Carlos Oscar S.; Thierry Calandra; Thierry Roger; Mariano Esteban

    2011-01-01

    The vaccinia virus (VACV) C6 protein has sequence similarities with the poxvirus family Pox_A46, involved in regulation of host immune responses, but its role is unknown. Here, we have characterized the C6 protein and its effects in virus replication, innate immune sensing and immunogenicity in vivo. C6 is a 18.2 kDa protein, which is expressed early during virus infection and localizes to the cytoplasm of infected cells. Deletion of the C6L gene from the poxvirus vector MVA-B expressing HIV-...

  1. The D10 Decapping Enzyme of Vaccinia Virus Contributes to Decay of Cellular and Viral mRNAs and to Virulence in Mice

    OpenAIRE

    2014-01-01

    Posttranscriptional mechanisms are important for regulation of cellular and viral gene expression. The presence of the 5′ cap structure m7G(5′)ppp(5′)Nm is a general feature of mRNAs that provides protection from exoribonuclease digestion and enhances translation. Vaccinia virus and other poxviruses encode enzymes for both cap synthesis and decapping. Decapping is mediated by two related enzymes, D9 and D10, which are synthesized before and after viral DNA replication, respectively. The timin...

  2. Quantitative Analysis of MicroRNAs in Vaccinia virus Infection Reveals Diversity in Their Susceptibility to Modification and Suppression.

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    Amy H Buck

    Full Text Available Vaccinia virus (VACV is a large cytoplasmic DNA virus that causes dramatic alterations to many cellular pathways including microRNA biogenesis. The virus encodes a poly(A polymerase which was previously shown to add poly(A tails to the 3' end of cellular miRNAs, resulting in their degradation by 24 hours post infection (hpi. Here we used small RNA sequencing to quantify the impact of VACV infection on cellular miRNAs in human cells at both early (6 h and late (24 h times post infection. A detailed quantitative analysis of individual miRNAs revealed marked diversity in the extent of their modification and relative change in abundance during infection. Some miRNAs became highly modified (e.g. miR-29a-3p, miR-27b-3p whereas others appeared resistant (e.g. miR-16-5p. Furthermore, miRNAs that were highly tailed at 6 hpi were not necessarily among the most reduced at 24 hpi. These results suggest that intrinsic features of human cellular miRNAs cause them to be differentially polyadenylated and altered in abundance during VACV infection. We also demonstrate that intermediate and late VACV gene expression are required for optimal repression of some miRNAs including miR-27-3p. Overall this work reveals complex and varied consequences of VACV infection on host miRNAs and identifies miRNAs which are largely resistant to VACV-induced polyadenylation and are therefore present at functional levels during the initial stages of infection and replication.

  3. Intrarectal vaccination with recombinant vaccinia virus expressing carcinoembronic antigen induces mucosal and systemic immunity and prevents progression of colorectal cancer.

    Science.gov (United States)

    Kim-Schulze, Seunghee; Kim, Hong Sung; Wainstein, Alberto; Kim, Dae Won; Yang, Wein Cui; Moroziewicz, Dorota; Mong, Phyllus Y; Bereta, Michal; Taback, Bret; Wang, Qin; Kaufman, Howard L

    2008-12-01

    The gastrointestinal mucosa contains an intact immune system that protects the host from pathogens and communicates with the systemic immune system. Absorptive epithelial cells in the mucosa give rise to malignant tumors although the interaction between tumor cells and the mucosal immune system is not well defined. The pathophysiology of colorectal cancer has been elucidated through studies of hereditary syndromes, such as familial adenomatous polyposis, a cancer predisposition syndrome caused by germline mutations in the adenomatous polyposis coli tumor suppressor gene. Patients with FAP develop adenomas and inevitably progress to invasive carcinomas by the age of 40. To better delineate the role of mucosal immunity in colorectal cancer, we evaluated the efficacy of intrarectal recombinant vaccinia virus expressing the human carcinoembryonic Ag (CEA) in a murine FAP model in which mice are predisposed to colorectal cancer and also express human CEA in the gut. Mucosal vaccination reduced the incidence of spontaneous adenomas and completely prevented progression to invasive carcinoma. The therapeutic effects were associated with induction of mucosal CEA-specific IgA Ab titers and CD8(+) CTLs. Mucosal vaccination was also associated with an increase in systemic CEA-specific IgG Ab titers, CD4(+) and CD8(+) T cell responses and resulted in growth inhibition of s.c. implanted CEA-expressing tumors suggesting communication between mucosal and systemic immune compartments. Thus, intrarectal vaccination induces mucosal and systemic antitumor immunity and prevents progression of spontaneous colorectal cancer. These results have implications for the prevention of colorectal cancer in high-risk individuals.

  4. Direct TLR2 signaling is critical for NK cell activation and function in response to vaccinia viral infection.

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    Jennifer Martinez

    2010-03-01

    Full Text Available Natural killer (NK cells play an essential role in innate immune control of poxviral infections in vivo. However, the mechanism(s underlying NK cell activation and function in response to poxviruses remains poorly understood. In a mouse model of infection with vaccinia virus (VV, the most studied member of the poxvirus family, we identified that the Toll-like receptor (TLR 2-myeloid differentiating factor 88 (MyD88 pathway was critical for the activation of NK cells and the control of VV infection in vivo. We further showed that TLR2 signaling on NK cells, but not on accessory cells such as dendritic cells (DCs, was necessary for NK cell activation and that this intrinsic TLR2-MyD88 signaling pathway was required for NK cell activation and played a critical role in the control of VV infection in vivo. In addition, we showed that the activating receptor NKG2D was also important for efficient NK activation and function, as well as recognition of VV-infected targets. We further demonstrated that VV could directly activate NK cells via TLR2 in the presence of cytokines in vitro and TLR2-MyD88-dependent activation of NK cells by VV was mediated through the phosphatidylinositol 3-kinase (PI3K-extracellular signal-regulated kinase (ERK pathway. Taken together, these results represent the first evidence that intrinsic TLR signaling is critical for NK cell activation and function in the control of a viral infection in vivo, indicate that multiple pathways are required for efficient NK cell activation and function in response to VV infection, and may provide important insights into the design of effective strategies to combat poxviral infections.

  5. The membrane fusion step of vaccinia virus entry is cooperatively mediated by multiple viral proteins and host cell components.

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    Jason P Laliberte

    2011-12-01

    Full Text Available For many viruses, one or two proteins allow cell attachment and entry, which occurs through the plasma membrane or following endocytosis at low pH. In contrast, vaccinia virus (VACV enters cells by both neutral and low pH routes; four proteins mediate cell attachment and twelve that are associated in a membrane complex and conserved in all poxviruses are dedicated to entry. The aim of the present study was to determine the roles of cellular and viral proteins in initial stages of entry, specifically fusion of the membranes of the mature virion and cell. For analysis of the role of cellular components, we used well characterized inhibitors and measured binding of a recombinant VACV virion containing Gaussia luciferase fused to a core protein; viral and cellular membrane lipid mixing with a self-quenching fluorescent probe in the virion membrane; and core entry with a recombinant VACV expressing firefly luciferase and electron microscopy. We determined that inhibitors of tyrosine protein kinases, dynamin GTPase and actin dynamics had little effect on binding of virions to cells but impaired membrane fusion, whereas partial cholesterol depletion and inhibitors of endosomal acidification and membrane blebbing had a severe effect at the later stage of core entry. To determine the role of viral proteins, virions lacking individual membrane components were purified from cells infected with members of a panel of ten conditional-lethal inducible mutants. Each of the entry protein-deficient virions had severely reduced infectivity and except for A28, L1 and L5 greatly impaired membrane fusion. In addition, a potent neutralizing L1 monoclonal antibody blocked entry at a post-membrane lipid-mixing step. Taken together, these results suggested a 2-step entry model and implicated an unprecedented number of viral proteins and cellular components involved in signaling and actin rearrangement for initiation of virus-cell membrane fusion during poxvirus entry.

  6. Vaccinia Virus Immunomodulator A46: A Lipid and Protein-Binding Scaffold for Sequestering Host TIR-Domain Proteins.

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    Sofiya Fedosyuk

    2016-12-01

    Full Text Available Vaccinia virus interferes with early events of the activation pathway of the transcriptional factor NF-kB by binding to numerous host TIR-domain containing adaptor proteins. We have previously determined the X-ray structure of the A46 C-terminal domain; however, the structure and function of the A46 N-terminal domain and its relationship to the C-terminal domain have remained unclear. Here, we biophysically characterize residues 1-83 of the N-terminal domain of A46 and present the X-ray structure at 1.55 Å. Crystallographic phases were obtained by a recently developed ab initio method entitled ARCIMBOLDO_BORGES that employs tertiary structure libraries extracted from the Protein Data Bank; data analysis revealed an all β-sheet structure. This is the first such structure solved by this method which should be applicable to any protein composed entirely of β-sheets. The A46(1-83 structure itself is a β-sandwich containing a co-purified molecule of myristic acid inside a hydrophobic pocket and represents a previously unknown lipid-binding fold. Mass spectrometry analysis confirmed the presence of long-chain fatty acids in both N-terminal and full-length A46; mutation of the hydrophobic pocket reduced the lipid content. Using a combination of high resolution X-ray structures of the N- and C-terminal domains and SAXS analysis of full-length protein A46(1-240, we present here a structural model of A46 in a tetrameric assembly. Integrating affinity measurements and structural data, we propose how A46 simultaneously interferes with several TIR-domain containing proteins to inhibit NF-κB activation and postulate that A46 employs a bipartite binding arrangement to sequester the host immune adaptors TRAM and MyD88.

  7. Identification from diverse mammalian poxviruses of host-range regulatory genes functioning equivalently to vaccinia virus C7L.

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    Meng, Xiangzhi; Chao, Jie; Xiang, Yan

    2008-03-15

    Vaccinia virus (VACV) C7L is a host-range gene that regulates cellular tropism of VACV. Distantly related C7L homologues are encoded by nearly all mammalian poxviruses, but whether they are host-range genes functioning similar to VACV C7L has not been determined. Here, we used VACV as a model system to analyze five different C7L homologues from diverse mammalian poxviruses for their abilities to regulate poxvirus cellular tropism. Three C7L homologues (myxoma virus M63R, M64R and cowpox virus 020), when expressed with an epitope tag and from a VACV mutant lacking the host-range genes K1L and C7L (vK1L-C7L-), failed to support productive viral replication in human and murine cells. In nonpermissive cells, these viruses did not synthesize viral late proteins, expressed a reduced level of the early protein E3L, and were defective at suppressing cellular PKR activation. In contrast, two other C7L homologues, myxoma virus (MYXV) M62R and yaba-like disease virus (YLDV) 67R, when expressed with an epitope tag and from vK1L(-)C7L(-), supported normal viral replication in human and murine cells and restored the ability of the virus to suppress PKR activation. Furthermore, M62R rescued the defect of vK1L(-)C7L(-) at replicating and disseminating in mice following intranasal inoculation. These results show that MYXV M62R and YLDV 67R function equivalently to C7L at supporting VACV replication in mammalian hosts and suggest that a C7L-like host-range gene is essential for the replication of many mammalian poxviruses in mammalian hosts.

  8. A novel naturally occurring tandem promoter in modified vaccinia virus ankara drives very early gene expression and potent immune responses.

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    Sonia T Wennier

    Full Text Available Modified vaccinia virus Ankara (MVA has been shown to be suitable for the generation of experimental vaccines against cancer and infectious diseases, eliciting strong humoral and cellular immune responses. In viral vectored vaccines, strong recombinant antigen expression and timing of expression influence the quantity and quality of the immune response. Screening of synthetic and native poxvirus promoters for strong protein expression in vitro and potent immune responses in vivo led to the identification of the MVA13.5L promoter, a unique and novel naturally occurring tandem promoter in MVA composed of two 44 nucleotide long repeated motifs, each containing an early promoter element. The MVA13.5L gene is highly conserved across orthopoxviruses, yet its function is unknown. The unique structure of its promoter is not found for any other gene in the MVA genome and is also conserved in other orthopoxviruses. Comparison of the MVA13.5L promoter activity with synthetic poxviral promoters revealed that the MVA13.5L promoter produced higher levels of protein early during infection in HeLa cells and particularly in MDBK cells, a cell line in which MVA replication stops at an early stage before the expression of late genes. Finally, a recombinant antigen expressed under the control of this novel promoter induced high antibody titers and increased CD8 T cell responses in homologous prime-boost immunization compared to commonly used promoters. In particular, the recombinant antigen specific CD8 T cell responses dominated over the immunodominant B8R vector-specific responses after three vaccinations and even more during the memory phase. These results have identified the native MVA13.5L promoter as a new potent promoter for use in MVA vectored preventive and therapeutic vaccines.

  9. Transforming growth factor alpha, Shope fibroma growth factor, and vaccinia growth factor can replace myxoma growth factor in the induction of myxomatosis in rabbits.

    Science.gov (United States)

    Opgenorth, A; Nation, N; Graham, K; McFadden, G

    1993-02-01

    The epidermal growth factor (EGF) homologues encoded by vaccinia virus, myxoma virus, and malignant rabbit fibroma virus have been shown to contribute to the pathogenicity of virus infection upon inoculation of susceptible hosts. However, since the primary structures of these growth factors and the disease profiles induced by different poxvirus genera vary substantially, the degree to which the various EGF homologues perform similar roles in viral pathogenesis remains unclear. In order to determine whether different EGF-like growth factors can perform qualitatively similar functions in the induction of myxomatosis in rabbits, we created recombinant myxoma virus variants in which the native growth factor, myxoma growth factor (MGF), was disrupted and replaced with either vaccinia virus growth factor, Shope fibroma growth factor, or rat transforming growth factor alpha. Unlike the control virus containing an inactivated MGF gene, which caused marked attenuation of the disease syndrome and substantially less proliferation of the epithelial cell layers in the conjunctiva and respiratory tract, the recombinant myxoma virus strains expressing heterologous growth factors produced infections which were both clinically and histopathologically indistinguishable from wild-type myxomatosis. We conclude that these poxviral and cellular EGF-like growth factors, which are diverse with respect to primary structure and origin, have similar biological functions in the context of myxoma virus pathogenesis and are mitogenic for the same target cells.

  10. Role of metalloproteases in vaccinia virus epitope processing for transporter associated with antigen processing (TAP)-independent human leukocyte antigen (HLA)-B7 class I antigen presentation.

    Science.gov (United States)

    Lorente, Elena; García, Ruth; Mir, Carmen; Barriga, Alejandro; Lemonnier, François A; Ramos, Manuel; López, Daniel

    2012-03-23

    The transporter associated with antigen processing (TAP) translocates the viral proteolytic peptides generated by the proteasome and other proteases in the cytosol to the endoplasmic reticulum lumen. There, they complex with nascent human leukocyte antigen (HLA) class I molecules, which are subsequently recognized by the CD8(+) lymphocyte cellular response. However, individuals with nonfunctional TAP complexes or tumor or infected cells with blocked TAP molecules are able to present HLA class I ligands generated by TAP-independent processing pathways. Herein, using a TAP-independent polyclonal vaccinia virus-polyspecific CD8(+) T cell line, two conserved vaccinia-derived TAP-independent HLA-B*0702 epitopes were identified. The presentation of these epitopes in normal cells occurs via complex antigen-processing pathways involving the proteasome and/or different subsets of metalloproteinases (amino-, carboxy-, and endoproteases), which were blocked in infected cells with specific chemical inhibitors. These data support the hypothesis that the abundant cellular proteolytic systems contribute to the supply of peptides recognized by the antiviral cellular immune response, thereby facilitating immunosurveillance. These data may explain why TAP-deficient individuals live normal life spans without any increased susceptibility to viral infections.

  11. The attenuation of vaccinia Tian Tan strain by the removal of the viral M1L-K2L genes.

    Science.gov (United States)

    Zhu, Weijun; Fang, Qing; Zhuang, Ke; Wang, Haibo; Yu, Wenbo; Zhou, Jingying; Liu, Li; Tien, Po; Zhang, Linqi; Chen, Zhiwei

    2007-09-01

    To generate a safe vaccinia Tian Tan (VTT)-based vaccine vector, it is necessary to develop a method to attenuate the virus. A modified VTT (MVTT(2-GFP)) was constructed by replacing the viral M1L-K2L genes with a GFP gene. In comparison to the parental VTT, MVTT(2-GFP) lost its replication capacity in rabbit RK13 and human HeLa cell lines. The life cycle of viral replication was blocked at different stages in these two cell lines as determined by electron microscope examination. MVTT(2-GFP) was less virulent than VTT for 100-fold by measuring mouse body weight loss after intranasal viral inoculation and for 340-fold by determining the intracranial LD(50) value in mice. The foreign GFP gene was stable genetically after 10 rounds of passage in Vero cells. Importantly, MVTT(2-GFP) elicited both humoral and cell-mediated immune responses to the GFP gene in mice. With two intramuscular inoculations of 10(5)PFU virus, the anti-GFP antibody reciprocal endpoint titer reached over 700 as determined by an ELISA. The number of IFN-gamma secreting T cells reached over 350SFU per million splenocytes against a CD8+ T cell-specific epitope of GFP. Collectively, the removal of the M1L-K2L genes is a useful method to generate an attenuated vaccinia Tian Tan vaccine vector.

  12. The acidic C-terminus of vaccinia virus I3 single-strand binding protein promotes proper assembly of DNA-protein complexes.

    Science.gov (United States)

    Harrison, Melissa L; Desaulniers, Megan A; Noyce, Ryan S; Evans, David H

    2016-02-01

    The vaccinia virus I3L gene encodes a single-stranded DNA binding protein (SSB) that is essential for virus DNA replication and is conserved in all Chordopoxviruses. The I3 protein contains a negatively charged C-terminal tail that is a common feature of SSBs. Such acidic tails are critical for SSB-dependent replication, recombination and repair. We cloned and purified variants of the I3 protein, along with a homolog from molluscum contagiosum virus, and tested how the acidic tail affected DNA-protein interactions. Deleting the C terminus of I3 enhanced the affinity for single-stranded DNA cellulose and gel shift analyses showed that it also altered the migration of I3-DNA complexes in agarose gels. Microinjecting an antibody against I3 into vaccinia-infected cells also selectively inhibited virus replication. We suggest that this domain promotes cooperative binding of I3 to DNA in a way that would maintain an open DNA configuration around a replication site.

  13. Functional hyper-IL-6 from vaccinia virus-colonized tumors triggers platelet formation and helps to alleviate toxicity of mitomycin C enhanced virus therapy

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    Sturm Julia B

    2012-01-01

    Full Text Available Abstract Background Combination of oncolytic vaccinia virus therapy with conventional chemotherapy has shown promise for tumor therapy. However, side effects of chemotherapy including thrombocytopenia, still remain problematic. Methods Here, we describe a novel approach to optimize combination therapy of oncolytic virus and chemotherapy utilizing virus-encoding hyper-IL-6, GLV-1h90, to reduce chemotherapy-associated side effects. Results We showed that the hyper-IL-6 cytokine was successfully produced by GLV-1h90 and was functional both in cell culture as well as in tumor-bearing animals, in which the cytokine-producing vaccinia virus strain was well tolerated. When combined with the chemotherapeutic mitomycin C, the anti-tumor effect of the oncolytic virotherapy was significantly enhanced. Moreover, hyper-IL-6 expression greatly reduced the time interval during which the mice suffered from chemotherapy-induced thrombocytopenia. Conclusion Therefore, future clinical application would benefit from careful investigation of additional cytokine treatment to reduce chemotherapy-induced side effects.

  14. On the falsifications in history: Juan José Heydeck (n. 1755 and his «amazing» discovery of a vaccine against the smallpox

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    Olagüe de Ros, Guillermo

    2007-06-01

    Full Text Available Juan Jose Heydeck (b. 1755 is a turbid figure that in the last years he is acquiring certain historical relief. His false find of a vaccine has been overvalued, in excess, against the smallpox in goats of Madrid. In this note, which is a response to an article appeared in Asclepio in 2006, this personage is checked from an historical point of view and there is demonstrated the falsehood of his find, fruit of his excessive ambition.

    Juan José Heydeck (n. 1755 es una turbia figura que en los últimos años está adquiriendo cierto relieve historiográfico. Se ha sobrevalorado, en exceso, su falso hallazgo de una vacuna contra la
    viruela en cabras de Madrid. En esta nota, que es respuesta a un artículo aparecido en esta revista en 2006, se revisa historiográficamente a este personaje y se demuestra la falsedad de su hallazgo, fruto de su ambiciosa y desmedida ambición.

  15. Effects of nasal or pulmonary delivered treatments with an adenovirus vectored interferon (mDEF201 on respiratory and systemic infections in mice caused by cowpox and vaccinia viruses.

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    Donald F Smee

    Full Text Available An adenovirus 5 vector encoding for mouse interferon alpha, subtype 5 (mDEF201 was evaluated for efficacy against lethal cowpox (Brighton strain and vaccinia (WR strain virus respiratory and systemic infections in mice. Two routes of mDEF201 administration were used, nasal sinus (5-µl and pulmonary (50-µl, to compare differences in efficacy, since the preferred treatment of humans would be in a relatively small volume delivered intranasally. Lower respiratory infections (LRI, upper respiratory infections (URI, and systemic infections were induced by 50-µl intranasal, 10-µl intranasal, and 100-µl intraperitoneal virus challenges, respectively. mDEF201 treatments were given prophylactically either 24 h (short term or 56d (long-term prior to virus challenge. Single nasal sinus treatments of 10(6 and 10(7 PFU/mouse of mDEF201 protected all mice from vaccinia-induced LRI mortality (comparable to published studies with pulmonary delivered mDEF201. Systemic vaccinia infections responded significantly better to nasal sinus delivered mDEF201 than to pulmonary treatments. Cowpox LRI infections responded to 10(7 mDEF201 treatments, but a 10(6 dose was only weakly protective. Cowpox URI infections were equally treatable by nasal sinus and pulmonary delivered mDEF201 at 10(7 PFU/mouse. Dose-responsive prophylaxis with mDEF201, given one time only 56 d prior to initiating a vaccinia virus LRI infection, was 100% protective from 10(5 to 10(7 PFU/mouse. Improvements in lung hemorrhage score and lung weight were evident, as were decreases in liver, lung, and spleen virus titers. Thus, mDEF201 was able to treat different vaccinia and cowpox virus infections using both nasal sinus and pulmonary treatment regimens, supporting its development for humans.

  16. Vaccinia virus K1 ankyrin repeat protein inhibits NF-κB activation by preventing RelA acetylation.

    Science.gov (United States)

    Bravo Cruz, Ariana G; Shisler, Joanna L

    2016-10-01

    The vaccinia virus (VACV) K1 protein inhibits dsRNA-dependent protein kinase (PKR) activation. A consequence of this function is that K1 inhibits PKR-induced NF-κB activation during VACV infection. However, transient expression of K1 also inhibits Toll-like receptor (TLR)-induced NF-κB activation. This suggests that K1 has a second NF-κB inhibitory mechanism that is PKR-independent. This possibility was explored by expressing K1 independently of infection and stimulating NF-κB under conditions that minimized or excluded PKR activation. K1 inhibited both TNF- and phorbol 12-myristate 13-acetate (PMA)-induced NF-κB activation, as detected by transcription of synthetic (e.g. luciferase) and natural (e.g. CXCL8) genes controlled by NF-κB. K1 also inhibited NF-κB activity in PKRkd cells, cells that have greatly decreased amounts of PKR. K1 no longer prevented IκBα degradation or NF-κB nuclear translocation in the absence of PKR, suggesting that K1 acted on a nuclear event. Indeed, K1 was present in the nucleus and cytoplasm of stimulated and unstimulated cells. K1 inhibited acetylation of the RelA (p65) subunit of NF-κB, a nuclear event known to be required for NF-κB activation. Moreover, p65-CBP (CREB-binding protein) interactions were blocked in the presence of K1. However, K1 did not preclude NF-κB binding to oligonucleotides containing κB-binding sites. The current interpretation of these data is that NF-κB-promoter interactions still occur in the presence of K1, but NF-κB cannot properly trigger transcriptional activation because K1 antagonizes acetylation of RelA. Thus, in comparison to all known VACV NF-κB inhibitory proteins, K1 acts at one of the most downstream events of NF-κB activation.

  17. Imaging characteristics, tissue distribution, and spread of a novel oncolytic vaccinia virus carrying the human sodium iodide symporter.

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    Dana Haddad

    Full Text Available INTRODUCTION: Oncolytic viruses show promise for treating cancer. However, to assess therapy and potential toxicity, a noninvasive imaging modality is needed. This study aims to determine the in vivo biodistribution, and imaging and timing characteristics of a vaccinia virus, GLV-1h153, encoding the human sodium iodide symporter (hNIS. METHODS: GLV-1h153 was modified from GLV-1h68 to encode the hNIS gene. Timing of cellular uptake of radioiodide (131I in human pancreatic carcinoma cells PANC-1 was assessed using radiouptake assays. Viral biodistribution was determined in nude mice bearing PANC-1 xenografts, and infection in tumors confirmed histologically and optically via Green Fluorescent Protein (GFP and bioluminescence. Timing characteristics of enhanced radiouptake in xenografts were assessed via (124I-positron emission tomography (PET. Detection of systemic administration of virus was investigated with both (124I-PET and 99m-technecium gamma-scintigraphy. RESULTS: GLV-1h153 successfully facilitated time-dependent intracellular uptake of (131I in PANC-1 cells with a maximum uptake at 24 hours postinfection (P<0.05. In vivo, biodistribution profiles revealed persistence of virus in tumors 5 weeks postinjection at 10(9 plaque-forming unit (PFU/gm tissue, with the virus mainly cleared from all other major organs. Tumor infection by GLV-1h153 was confirmed via optical imaging and histology. GLV-1h153 facilitated imaging virus replication in tumors via PET even at 8 hours post radiotracer injection, with a mean %ID/gm of 3.82 ± 0.46 (P<0.05 2 days after intratumoral administration of virus, confirmed via tissue radiouptake assays. One week post systemic administration, GLV-1h153-infected tumors were detected via (124I-PET and 99m-technecium-scintigraphy. CONCLUSION: GLV-1h153 is a promising oncolytic agent against pancreatic cancer with a promising biosafety profile. GLV-1h153 facilitated time-dependent hNIS-specific radiouptake in pancreatic

  18. Vectors based on modified vaccinia Ankara expressing influenza H5N1 hemagglutinin induce substantial cross-clade protective immunity.

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    Annett Hessel

    Full Text Available BACKGROUND: New highly pathogenic H5N1 influenza viruses are continuing to evolve with a potential threat for an influenza pandemic. So far, the H5N1 influenza viruses have not widely circulated in humans and therefore constitute a high risk for the non immune population. The aim of this study was to evaluate the cross-protective potential of the hemagglutinins of five H5N1 strains of divergent clades using a live attenuated modified vaccinia Ankara (MVA vector vaccine. METHODOLOGY/PRINCIPAL FINDINGS: The replication-deficient MVA virus was used to express influenza hemagglutinin (HA proteins. Specifically, recombinant MVA viruses expressing the HA genes of the clade 1 virus A/Vietnam/1203/2004 (VN/1203, the clade 2.1.3 virus A/Indonesia/5/2005 (IN5/05, the clade 2.2 viruses A/turkey/Turkey/1/2005 (TT01/05 and A/chicken/Egypt/3/2006 (CE/06, and the clade 2.3.4 virus A/Anhui/1/2005 (AH1/05 were constructed. These experimental live vaccines were assessed in a lethal mouse model. Mice vaccinated with the VN/1203 hemagglutinin-expressing MVA induced excellent protection against all the above mentioned clades. Also mice vaccinated with the IN5/05 HA expressing MVA induced substantial protection against homologous and heterologous AH1/05 challenge. After vaccination with the CE/06 HA expressing MVA, mice were fully protected against clade 2.2 challenge and partially protected against challenge of other clades. Mice vaccinated with AH1/05 HA expressing MVA vectors were only partially protected against homologous and heterologous challenge. The live vaccines induced substantial amounts of neutralizing antibodies, mainly directed against the homologous challenge virus, and high levels of HA-specific IFN-γ secreting CD4 and CD8 T-cells against epitopes conserved among the H5 clades and subclades. CONCLUSIONS/SIGNIFICANCE: The highest level of cross-protection was induced by the HA derived from the VN/1203 strain, suggesting that pandemic H5 vaccines

  19. 表达HPV16L1、L2和E7蛋白的非复制重组痘苗病毒的构建%Construction of non-replicating recombinant vaccinia virus expressing HPV16 L1, L2E7 proteins

    Institute of Scientific and Technical Information of China (English)

    Jiangtao Fan; Xinqiu Chen; Wei Huang; Houwen Tian

    2009-01-01

    Objective:To construct a non-replicating vaccinia virus expressing human papitlomavirus 16 (HPV16) L1, L2E7 proteins as a candidate vaccine for cervical cancer. Methods:Using vaccinia virus vector, we generated a strain of non-repli-cating recombinant vaccinia virus vaccine expressing HPV16 L1, L2E7 proteins by homologous recombination and identified by PCR and Western-bloting. Results:We demonstrated that the L1, L2E7 gene of HPV16 were integrated into vaccinia genosome and could express L1, L2E7 protein stably when infected the CEF using PCR and Western-blot assay. Conclu-sion:NTVJL1/L2E7 can express L1, L2E7 protein of HPV16 and can be taken as a candidate vaccine for HPV16-associated diseases.

  20. Vacinação, varíola e uma cultura da imunização no Brasil Vaccination, smallpox, and a culture of immunization in Brazil

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    Gilberto Hochman

    2011-02-01

    Full Text Available O objetivo deste artigo é discutir a emergência e o estabelecimento de uma "cultura da imunização" no Brasil contemporâneo a partir da erradicação da varíola. Essa cultura está associada a um longo processo de introdução de vacinas, de campanhas de vacinação e de vacinação em massa empreendidas pelo Estado brasileiro desde o final do século XIX. Particular importância é atribuída à campanha de erradicação da varíola no Brasil (1966-1973. A experiência da vacinação em massa da população contra a varíola é contrastada com episódios de resistência como a "Revolta da Vacina", e considerada como crucial na configuração de novas políticas e novas compreensões sobre o papel da imunização na saúde pública e seu lugar na sociedade brasileira.The aim of this paper is to discuss the emergence and establishment of a "culture of immunization" in the contemporary Brazil from the eradication of smallpox. This culture is associated with a long process of introduction of vaccines, vaccination campaigns and mass vaccination undertaken by the Brazilian government since the late nineteenth century. Particular importance is attributed to the campaign to eradicate smallpox in Brazil (1966-1973. The experience of mass vaccination of the population against smallpox is contrasted with episodes of resistance as the "revolt against the vaccine", and regarded as crucial in shaping new policies and new understandings about the role of immunization in public health and its place in society Brazil.

  1. Disease, religion and medicine: smallpox in nineteenth-century Benin Doenças, religião e medicina: a varíola no Benim, século XIX

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    Elisée Soumonni

    2012-12-01

    Full Text Available The essay examines, with special reference to smallpox, the perception and interpretation of disease in pre-colonial Dahomey, present-day Republic of Benin. Because disease is seen primarily as a punishment from the gods and not just as a medical problem or a bodily disorder, traditional cult priests play a leading role in making diagnoses and prescribing remedies, mostly based on medicinal plants. The prominence of Sakpata, god of smallpox, coupled with the influence of its priests is evaluated within the context of Dahomey's political history and the spread of the disease. This pivotal position was to constitute a challenge to the French colonial campaign to vaccinate against smallpox.O ensaio examina - com especial atenção à varíola - as percepções e interpretações das doenças no Daomé pré-colonial, atual República do Benim. Uma vez que as doenças eram vistas antes de tudo como punição divina, e não como problema ou distúrbio do corpo, os sacerdotes tradicionais exerciam papel central no seu diagnóstico e na prescrição de remédios, com base principalmente em plantas medicinais. A importância do culto a Sakpata, deus da varíola, juntamente com a influência dos sacerdotes é avaliada dentro do contexto da história política do Daomé e da disseminação das doenças. A posição crucial desse culto constituiu-se como um desafio para a campanha colonial francesa de vacinação contra a varíola.

  2. Marcados en la piel: vacunación y viruela en Argentina (1870-1910 Marked on the skin: vaccination and smallpox in Argentina (1870-1910

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    Maria Silvia Di Liscia

    2011-02-01

    Full Text Available Este artículo estudia la vacunación antivariólica en Argentina desde 1870, cuando se inician los debates al respecto, hasta la década de 1910, cuando se amplía al resto del país. Se analizan las prácticas de inmunización puestas en marcha con anterioridad a la Ley de vacunación obligatoria, aprobada en 1886 para la Capital y en 1904 para el resto del país. Tal medida fue resistida desde diferentes sectores. Su aprobación dependió de las consecuencias del proceso de modernización y urbanización acelerada, del peso de los higienistas en el escenario político y su extensión dependió de una concepción administrativa diferente, que incorporaba nuevas áreas y sectores al escenario nacional.This paper studies the smallpox vaccination in Argentina since 1870, when these discussions were inittiaded until the 1910s, when they were extended to the rest of the country. We analyze immunization practices implemented prior to the compulsory vaccination law, passed in 1886 for the Capital and in 1904 for the rest of the country. Such a move found resistance from different sectors. Its approval depended on the consequences of modernization and urbanization, the weight of hygienists in the political arena, and its extension depended on a different administrative conception, incorporating new areas and sectors to the national scenario.

  3. Attenuation of vaccinia Tian Tan strain by removal of viral TC7L-TK2L and TA35R genes.

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    Shifu Kan

    Full Text Available Vaccinia Tian Tan (VTT was attenuated by deletion of the TC7L-TK2L and TA35R genes to generate MVTT3. The mutant was generated by replacing the open reading frames by a gene encoding enhanced green fluorescent protein (EGFP flanked by loxP sites. Viruses expressing EGFP were then screened for and purified by serial plaque formation. In a second step the marker EGFP gene was removed by transfecting cells with a plasmid encoding cre recombinase and selecting for viruses that had lost the EGFP phenotype. The MVTT3 mutant was shown to be avirulent and immunogenic. These results support the conclusion that TC7L-TK2L and TA35R deletion mutants can be used as safe viral vectors or as platform for vaccines.

  4. Vaccinia virus protein A3 is required for the production of normal immature virions and for the encapsidation of the nucleocapsid protein L4.

    Science.gov (United States)

    Jesus, Desyree Murta; Moussatche, Nissin; McFadden, Baron B D; Nielsen, Casey Paulasue; D'Costa, Susan M; Condit, Richard C

    2015-07-01

    Maturation of the vaccinia virion is an intricate process that results in the organization of the viroplasm contained in immature virions into the lateral bodies, core wall and nucleocapsid observed in the mature particles. It is unclear how this organization takes place and studies with mutants are indispensable in understanding this process. By characterizing an inducible mutant in the A3L gene, we revealed that A3, an inner core wall protein, is important for formation of normal immature viruses and also for the correct localization of L4, a nucleocapsid protein. L4 did not accumulate in the viral factories in the absence of A3 and was not encapsidated in the particles that do not contain A3. These data strengthen our previously suggested hypothesis that A3 and L4 interact and that this interaction is critical for proper formation of the core wall and nucleocapsid.

  5. Protective effects of a Modified Vaccinia Ankara-based vaccine candidate against Crimean-Congo Haemorrhagic Fever virus require both cellular and humoral responses.

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    Stuart D Dowall

    Full Text Available Crimean-Congo Haemorrhagic Fever (CCHF is a severe tick-borne disease, endemic in many countries in Africa, the Middle East, Eastern Europe and Asia. There is no approved vaccine currently available against CCHF. The most promising candidate, which has previously been shown to confer protection in the small animal model, is a modified Vaccinia Ankara virus vector expressing the CCHF viral glycoprotein (MVA-GP. It has been shown that MVA-GP induces both humoral and cellular immunogenicity. In the present study, sera and T-lymphocytes were passively and adoptively transferred into recipient mice prior to challenge with CCHF virus. Results demonstrated that mediators from both arms of the immune system were required to demonstrate protective effects against lethal challenge.

  6. Protective effects of a Modified Vaccinia Ankara-based vaccine candidate against Crimean-Congo Haemorrhagic Fever virus require both cellular and humoral responses.

    Science.gov (United States)

    Dowall, Stuart D; Graham, Victoria A; Rayner, Emma; Hunter, Laura; Watson, Robert; Taylor, Irene; Rule, Antony; Carroll, Miles W; Hewson, Roger

    2016-01-01

    Crimean-Congo Haemorrhagic Fever (CCHF) is a severe tick-borne disease, endemic in many countries in Africa, the Middle East, Eastern Europe and Asia. There is no approved vaccine currently available against CCHF. The most promising candidate, which has previously been shown to confer protection in the small animal model, is a modified Vaccinia Ankara virus vector expressing the CCHF viral glycoprotein (MVA-GP). It has been shown that MVA-GP induces both humoral and cellular immunogenicity. In the present study, sera and T-lymphocytes were passively and adoptively transferred into recipient mice prior to challenge with CCHF virus. Results demonstrated that mediators from both arms of the immune system were required to demonstrate protective effects against lethal challenge.

  7. Attenuation of Vaccinia Tian Tan Strain by Removal of Viral TC7L-TK2L and TA35R Genes

    Science.gov (United States)

    Kan, Shifu; Wang, Yuhang; Sun, Lili; Jia, Peng; Qi, Yanxin; Su, Jiaqiang; Liu, Lei; Yang, Guohua; Liu, Liming; Wang, Zhuoyue; Wang, Jinhui; Liu, Guangchen; Jin, Ningyi; Li, Xiao; Ding, Zhuang

    2012-01-01

    Vaccinia Tian Tan (VTT) was attenuated by deletion of the TC7L-TK2L and TA35R genes to generate MVTT3. The mutant was generated by replacing the open reading frames by a gene encoding enhanced green fluorescent protein (EGFP) flanked by loxP sites. Viruses expressing EGFP were then screened for and purified by serial plaque formation. In a second step the marker EGFP gene was removed by transfecting cells with a plasmid encoding cre recombinase and selecting for viruses that had lost the EGFP phenotype. The MVTT3 mutant was shown to be avirulent and immunogenic. These results support the conclusion that TC7L-TK2L and TA35R deletion mutants can be used as safe viral vectors or as platform for vaccines. PMID:22363781

  8. Enhanced immunogenicity for CD8+ T cell induction and complete protective efficacy of malaria DNA vaccination by boosting with modified vaccinia virus Ankara.

    Science.gov (United States)

    Schneider, J; Gilbert, S C; Blanchard, T J; Hanke, T; Robson, K J; Hannan, C M; Becker, M; Sinden, R; Smith, G L; Hill, A V

    1998-04-01

    Immunization with irradiated sporozoites can protect against malaria infection and intensive efforts are aimed at reproducing this effect with subunit vaccines. A particular sequence of subunit immunization with pre-erythrocytic antigens of Plasmodium berghei, consisting of single dose priming with plasmid DNA followed by a single boost with a recombinant modified vaccinia virus Ankara (MVA) expressing the same antigen, induced unprecedented complete protection against P. berghei sporozoite challenge in two strains of mice. Protection was associated with very high levels of splenic peptide-specific interferon-gamma-secreting CD8+ T cells and was abrogated when the order of immunization was reversed. DNA priming followed by MVA boosting may provide a general immunization regime for induction of high levels of CD8+ T cells.

  9. The nucleotide sequence of the chicken thymidine kinase gene and the relationship of its predicted polypeptide to that of the vaccinia virus thymidine kinase.

    Science.gov (United States)

    Kwoh, T J; Engler, J A

    1984-05-11

    The entire DNA nucleotide sequence of a 3.0 kilobase pair Hind III fragment containing the chicken cytoplasmic thymidine kinase gene was determined. Oligonucleotide linker insertion mutations distributed throughout this gene and having known effects upon gene activity ( Kwoh , T.J., Zipser , D., and Wigler , M. 1983. J. Mol. Appl. Genet. 2, 191-200), were used to access regions of the Hind III fragment for sequencing reactions. The complete nucleotide sequence, together with the positions of the linker insertion mutations within the sequence, allows us to propose a structure for the chicken thymidine kinase gene. The protein coding sequence of the gene is divided into seven small segments (each less than 160 base pairs) by six small introns (each less than 230 base pairs). The proposed 244 amino acid polypeptide encoded by this gene bears strong homology to the vaccinia virus thymidine kinase. No homology with the thymidine kinases of the herpes simplex viruses was found.

  10. Characterization of a 7-kilodalton subunit of vaccinia virus DNA-dependent RNA polymerase with structural similarities to the smallest subunit of eukaryotic RNA polymerase II.

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    Amegadzie, B Y; Ahn, B Y; Moss, B

    1992-05-01

    A previously unrecognized 7-kDa polypeptide copurified with the DNA-dependent RNA polymerase of vaccinia virus virions. Internal amino acid sequences of the small protein matched a viral genomic open reading frame of 63 codons. Antipeptide antiserum was used to confirm the specific and complete association of the 7-kDa protein with RNA polymerase. The amino acid sequence predicted from the viral gene, named rpo7, was 23% identical to that of the smallest subunit of Saccharomyces cerevisiae RNA polymerase II, and a metal-binding motif, Cys-X-X-Cys-Gly, was located at precisely the same location near the N terminus in the two proteins. RNA analyses demonstrated early transcriptional initiation and termination signals in the rpo7 gene sequence. The viral RNA polymerase subunit was synthesized during the early phase of infection and continued to accumulate during the late phase.

  11. Vaccinia mature virus fusion regulator A26 protein binds to A16 and G9 proteins of the viral entry fusion complex and dissociates from mature virions at low pH.

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    Chang, Shu-Jung; Shih, Ao-Chun; Tang, Yin-Liang; Chang, Wen

    2012-04-01

    Vaccinia mature virus enters cells through either endocytosis or plasma membrane fusion, depending on virus strain and cell type. Our previous results showed that vaccinia virus mature virions containing viral A26 protein enter HeLa cells preferentially through endocytosis, whereas mature virions lacking A26 protein enter through plasma membrane fusion, leading us to propose that A26 acts as an acid-sensitive fusion suppressor for mature virus (S. J. Chang, Y. X. Chang, R. Izmailyan R, Y. L. Tang, and W. Chang, J. Virol. 84:8422-8432, 2010). In the present study, we investigated the fusion suppression mechanism of A26 protein. We found that A26 protein was coimmunoprecipitated with multiple components of the viral entry-fusion complex (EFC) in infected HeLa cells. Transient expression of viral EFC components in HeLa cells revealed that vaccinia virus A26 protein interacted directly with A16 and G9 but not with G3, L5 and H2 proteins of the EFC components. Consistently, a glutathione S-transferase (GST)-A26 fusion protein, but not GST, pulled down A16 and G9 proteins individually in vitro. Together, our results supported the idea that A26 protein binds to A16 and G9 protein at neutral pH contributing to suppression of vaccinia virus-triggered membrane fusion from without. Since vaccinia virus extracellular envelope proteins A56/K2 were recently shown to bind to the A16/G9 subcomplex to suppress virus-induced fusion from within, our results also highlight an evolutionary convergence in which vaccinia viral fusion suppressor proteins regulate membrane fusion by targeting the A16 and G9 components of the viral EFC complex. Finally, we provide evidence that acid (pH 4.7) treatment induced A26 protein and A26-A27 protein complexes of 70 kDa and 90 kDa to dissociate from mature virions, suggesting that the structure of A26 protein is acid sensitive.

  12. Modified vaccinia virus Ankara triggers type I IFN production in murine conventional dendritic cells via a cGAS/STING-mediated cytosolic DNA-sensing pathway.

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    Peihong Dai

    2014-04-01

    Full Text Available 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.

  13. Insertion of the human sodium iodide symporter to facilitate deep tissue imaging does not alter oncolytic or replication capability of a novel vaccinia virus

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    Mittra Arjun

    2011-03-01

    Full Text Available Abstract Introduction Oncolytic viruses show promise for treating cancer. However, to assess therapeutic efficacy and potential toxicity, a noninvasive imaging modality is needed. This study aimed to determine if insertion of the human sodium iodide symporter (hNIS cDNA as a marker for non-invasive imaging of virotherapy alters the replication and oncolytic capability of a novel vaccinia virus, GLV-1h153. Methods GLV-1h153 was modified from parental vaccinia virus GLV-1h68 to carry hNIS via homologous recombination. GLV-1h153 was tested against human pancreatic cancer cell line PANC-1 for replication via viral plaque assays and flow cytometry. Expression and transportation of hNIS in infected cells was evaluated using Westernblot and immunofluorescence. Intracellular uptake of radioiodide was assessed using radiouptake assays. Viral cytotoxicity and tumor regression of treated PANC-1tumor xenografts in nude mice was also determined. Finally, tumor radiouptake in xenografts was assessed via positron emission tomography (PET utilizing carrier-free 124I radiotracer. Results GLV-1h153 infected, replicated within, and killed PANC-1 cells as efficiently as GLV-1h68. GLV-1h153 provided dose-dependent levels of hNIS expression in infected cells. Immunofluorescence detected transport of the protein to the cell membrane prior to cell lysis, enhancing hNIS-specific radiouptake (P In vivo, GLV-1h153 was as safe and effective as GLV-1h68 in regressing pancreatic cancer xenografts (P 124I-PET. Conclusion Insertion of the hNIS gene does not hinder replication or oncolytic capability of GLV-1h153, rendering this novel virus a promising new candidate for the noninvasive imaging and tracking of oncolytic viral therapy.

  14. A pilot study comparing the development of EIAV Env-specific antibodies induced by DNA/recombinant vaccinia-vectored vaccines and an attenuated Chinese EIAV vaccine.

    Science.gov (United States)

    Meng, Qinglai; Lin, Yuezhi; Ma, Jian; Ma, Yan; Zhao, Liping; Li, Shenwei; Yang, Kai; Zhou, Jianhua; Shen, Rongxian; Zhang, Xiaoyan; Shao, Yiming

    2012-12-01

    Data from successful attenuated lentiviral vaccine studies indicate that fully mature Env-specific antibodies characterized by high titer, high avidity, and the predominant recognition of conformational epitopes are associated with protective efficacy. Although vaccination with a DNA prime/recombinant vaccinia-vectored vaccine boost strategy has been found to be effective in some trials with non-human primate/simian/human immunodeficiency virus (SHIV) models, it remains unclear whether this vaccination strategy could elicit mature equine infectious anemia virus (EIAV) Env-specific antibodies, thus protecting vaccinated horses against EIAV infection. Therefore, in this pilot study we vaccinated horses using a strategy based on DNA prime/recombinant Tiantan vaccinia (rTTV)-vectored vaccines encoding EIAV env and gag genes, and observed the development of Env-specific antibodies, neutralizing antibodies, and p26-specific antibodies. Vaccination with DNA induced low titer, low avidity, and the predominant recognition of linear epitopes by Env-specific antibodies, which was enhanced by boosting vaccinations with rTTV vaccines. However, the maturation levels of Env-specific antibodies induced by the DNA/rTTV vaccines were significantly lower than those induced by the attenuated vaccine EIAV(FDDV). Additionally, DNA/rTTV vaccines did not elicit broadly neutralizing antibodies. After challenge with a virulent EIAV strain, all of the vaccinees and control horses died from EIAV disease. These data indicate that the regimen of DNA prime/rTTV vaccine boost did not induce mature Env-specific antibodies, which might have contributed to immune protection failure.

  15. Vaccination of prostate cancer patients with modified vaccinia ankara delivering the tumor antigen 5T4 (TroVax): a phase 2 trial.

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    Amato, Robert J; Drury, Noel; Naylor, Stuart; Jac, Jaroslaw; Saxena, Somya; Cao, Amy; Hernandez-McClain, Joan; Harrop, Richard

    2008-01-01

    The attenuated vaccinia virus, modified vaccinia Ankara, has been engineered to deliver the tumor antigen 5T4 (TroVax). TroVax has been evaluated in an open-label phase 2 trial in hormone refractory prostate cancer patients in which the vaccine was administered either alone or in combination with granulocyte macrophage-colony stimulating factor (GM-CSF). The comparative safety and immunologic and clinical efficacy of TroVax alone or in combination with GM-CSF was determined. Twenty-seven patients with metastatic hormone refractory prostate cancer were treated with TroVax alone (n=14) or TroVax+GM-CSF (n=13). 5T4-specific cellular and humoral responses were monitored throughout the study. Clinical responses were assessed by quantifying prostate-specific antigen concentrations and measuring changes in tumor burden by computer-assisted tomography scan. TroVax was well tolerated in all patients with no serious adverse events attributed to vaccination. Of 24 immunologically evaluable patients, all mounted 5T4-specific antibody responses. Periods of disease stabilization from 2 to >10 months were observed. Time to progression was significantly greater in patients who mounted 5T4-specific cellular responses compared with those who did not (5.6 vs. 2.3 mo, respectively). There were no objective clinical responses seen in this study. In this study, the combination of GM-CSF with TroVax showed similar clinical and immunologic responses to TroVax alone. The high frequency of 5T4-specific immune responses and relationship with enhanced time to progression is encouraging and warrants further investigation.

  16. Efficacy of vaccination with recombinant vaccinia and fowlpox vectors expressing NY-ESO-1 antigen in ovarian cancer and melanoma patients.

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    Odunsi, Kunle; Matsuzaki, Junko; Karbach, Julia; Neumann, Antje; Mhawech-Fauceglia, Paulette; Miller, Austin; Beck, Amy; Morrison, Carl D; Ritter, Gerd; Godoy, Heidi; Lele, Shashikant; duPont, Nefertiti; Edwards, Robert; Shrikant, Protul; Old, Lloyd J; Gnjatic, Sacha; Jäger, Elke

    2012-04-10

    Recombinant poxviruses (vaccinia and fowlpox) expressing tumor-associated antigens are currently being evaluated in clinical trials as cancer vaccines to induce tumor-specific immune responses that will improve clinical outcome. To test whether a diversified prime and boost regimen targeting NY-ESO-1 will result in clinical benefit, we conducted two parallel phase II clinical trials of recombinant vaccinia-NY-ESO-1 (rV-NY-ESO-1), followed by booster vaccinations with recombinant fowlpox-NY-ESO-1 (rF-NY-ESO-1) in 25 melanoma and 22 epithelial ovarian cancer (EOC) patients with advanced disease who were at high risk for recurrence/progression. Integrated NY-ESO-1-specific antibody and CD4(+) and CD8(+) T cells were induced in a high proportion of melanoma and EOC patients. In melanoma patients, objective response rate [complete and partial response (CR+PR)] was 14%, mixed response was 5%, and disease stabilization was 52%, amounting to a clinical benefit rate (CBR) of 72% in melanoma patients. The median PFS in the melanoma patients was 9 mo (range, 0-84 mo) and the median OS was 48 mo (range, 3-106 mo). In EOC patients, the median PFS was 21 mo (95% CI, 16-29 mo), and median OS was 48 mo (CI, not estimable). CD8(+) T cells derived from vaccinated patients were shown to lyse NY-ESO-1-expressing tumor targets. These data provide preliminary evidence of clinically meaningful benefit for diversified prime and boost recombinant pox-viral-based vaccines in melanoma and ovarian cancer and support further evaluation of this approach in these patient populations.

  17. A vaccinia virus recombinant transcribing an alphavirus replicon and expressing alphavirus structural proteins leads to packaging of alphavirus infectious single cycle particles.

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    Juana M Sánchez-Puig

    Full Text Available Poxviruses and Alphaviruses constitute two promising viral vectors that have been used extensively as expression systems, or as vehicles for vaccine purposes. Poxviruses, like vaccinia virus (VV are well-established vaccine vectors having large insertion capacity, excellent stability, and ease of administration. In turn, replicons derived from Alphaviruses like Semliki Forest virus (SFV are potent protein expression and immunization vectors but stocks are difficult to produce and maintain. In an attempt to demonstrate the use of a Poxvirus as a means for the delivery of small vaccine vectors, we have constructed and characterized VV/SFV hybrid vectors. A SFV replicon cDNA was inserted in the VV genome and placed under the control of a VV early promoter. The replicon, transcribed from the VV genome as an early transcript, was functional, and thus capable of initiating its own replication and transcription. Further, we constructed a VV recombinant additionally expressing the SFV structural proteins under the control of a vaccinia synthetic early/late promoter. Infection with this recombinant produced concurrent transcription of the replicon and expression of SFV structural proteins, and led to the generation of replicon-containing SFV particles that were released to the medium and were able to infect additional cells. This combined VV/SFV system in a single virus allows the use of VV as a SFV delivery vehicle in vivo. The combination of two vectors, and the possibility of generating in vivo single-cycle, replicon containing alphavirus particles, may open new strategies in vaccine development or in the design of oncolytic viruses.

  18. A vaccinia virus recombinant transcribing an alphavirus replicon and expressing alphavirus structural proteins leads to packaging of alphavirus infectious single cycle particles.

    Science.gov (United States)

    Sánchez-Puig, Juana M; Lorenzo, María M; Blasco, Rafael

    2013-01-01

    Poxviruses and Alphaviruses constitute two promising viral vectors that have been used extensively as expression systems, or as vehicles for vaccine purposes. Poxviruses, like vaccinia virus (VV) are well-established vaccine vectors having large insertion capacity, excellent stability, and ease of administration. In turn, replicons derived from Alphaviruses like Semliki Forest virus (SFV) are potent protein expression and immunization vectors but stocks are difficult to produce and maintain. In an attempt to demonstrate the use of a Poxvirus as a means for the delivery of small vaccine vectors, we have constructed and characterized VV/SFV hybrid vectors. A SFV replicon cDNA was inserted in the VV genome and placed under the control of a VV early promoter. The replicon, transcribed from the VV genome as an early transcript, was functional, and thus capable of initiating its own replication and transcription. Further, we constructed a VV recombinant additionally expressing the SFV structural proteins under the control of a vaccinia synthetic early/late promoter. Infection with this recombinant produced concurrent transcription of the replicon and expression of SFV structural proteins, and led to the generation of replicon-containing SFV particles that were released to the medium and were able to infect additional cells. This combined VV/SFV system in a single virus allows the use of VV as a SFV delivery vehicle in vivo. The combination of two vectors, and the possibility of generating in vivo single-cycle, replicon containing alphavirus particles, may open new strategies in vaccine development or in the design of oncolytic viruses.

  19. Modified vaccinia virus Ankara triggers type I IFN production in murine conventional dendritic cells via a cGAS/STING-mediated cytosolic DNA-sensing pathway.

    Science.gov (United States)

    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

    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.

  20. Preferential colonization of metastases by oncolytic vaccinia virus strain GLV-1h68 in a human PC-3 prostate cancer model in nude mice.

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    Ulrike Donat

    Full Text Available Recently, we showed that the oncolytic vaccinia virus GLV-1h68 has a significant therapeutic potential in treating lymph node metastases of human PC-3 prostate carcinoma in tumor xenografts. In this study, underlying mechanisms of the virus-mediated metastases reduction were analyzed. Immunohistochemistry demonstrated that virus-treatment resulted in a drastically decrease of blood and lymph vessels, representing essential routes for PC-3 cell migration, in both tumors and metastases. Thus, GLV-1h68 drastically reduced essential routes for the metastatic spread of PC-3 cells. Furthermore, analysis of viral distribution in GLV-1h68-injected tumor-bearing mice by plaque assays, revealed significantly higher virus titers in metastases compared to solid tumors. To elucidate conditions potentially mediating the preferential viral colonization and eradication of metastases, microenvironmental components of uninfected tumors and metastases were compared by microscopic studies. These analyses revealed that PC-3 lymph node metastases showed increased vascular permeability, higher proliferation status of tumor cells as determined by BrdU- and Ki-67 assays and lesser necrosis of PC-3 cells than solid tumors. Moreover, an increased number of immune cells (MHCII(+/CD68(+ macrophages, MHCII(+/CD19(+ B lymphocytes combined with an up-regulated expression of pro-inflammatory cytokines was observed in metastases in comparison to primary PC-3 tumors. We propose that these microenvironmental components mediated the metastatic tropism of GLV-1h68. Therefore, vaccinia virus-based oncolytic virotherapy might offer a novel treatment of metastatic prostate carcinomas in humans.