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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., Pub. in PLoS one, v5 n10, p1-11, Dec 2010.

Monkeypox virus (MPXV) infection in humans results in clinical symptoms very similar to ordinary smallpox. Aerosol is a route of secondary transmission for monkeypox, and a primary route of smallpox transmission in humans. Therefore, an animal model for aerosol exposure to MPXV is needed to test medical countermeasures. To characterize the pathogenesis in cynomolgus macaques (Macaca fascicularis), groups of macaques were exposed to four different doses of aerosolized MPXV. Blood was collected the day before, and every other day after exposure and assessed for complete blood count (CBC), clinical chemistry analysis, and quantitative PCR. Macaques showed mild anorexia, depression, and fever on day 6 post-exposure. Lymphadenopathy, which differentiates monkeypox from smallpox, was observed in exposed macaques around day 6 post-exposure. CBC and clinical chemistries showed abnormalities similar to human monkeypox cases. Whole blood and throat swab viral loads peaked around day 10, and in survivors, gradually decreased until day 28 post-exposure. Survival was not dose dependent. As such, doses of 4x10,000 PFU, 1x100,000 PFU, or 1x1,000,000 PFU resulted in lethality for 70% of the animals, whereas a dose of 4x100,000 PFU resulted in 85% lethality. Overall, cynomolgus macaques exposed to aerosolized MPXV develop a clinical disease that resembles that of human monkeypox. These findings provide a strong foundation for the use of aerosolized MPXV exposure of cynomolgus macaques as an animal model to test medical countermeasures against orthopoxviruses., Published in PLoS ONE, v5 n9 e12880, Sep 2010.

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(Registered) (Wyeth Laboratories, Inc.), was highly effective, but the supply was insufficient to vaccinate the entire current US population. Additionally Dryvax(R) 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(Trademarked) (Acambis, Inc.(TM)), a single plaque-purified vaccinia virus derivative of Dryvax(R), aseptically propagated in cell culture. Preclinical and clinical trials reported in 2008 demonstrated that ACAM2000(TM) has comparable immunogenicity to that of Dryvax(R) and causes a similar frequency of adverse events. Furthermore, like Dryvax(R), ACAM2000(TM) vaccination has been shown by careful cardiac screening to result in an unexpectedly high rate of myocarditis and pericarditis. ACAM2000(TM) received US Food and Drug Administration (FDA) approval in August 2007, and replaced Dryvax(R) for all smallpox vaccinations in February 2008. Currently, over 200 million doses of ACAM2000(TM) have been produced for the US Strategic National Stockpile. This review of ACAM2000(TM) addresses the production, characterization clinical trials, and adverse events associated with this new smallpox vaccine., Published in Drug Design, Development and Therapy v3687 p71-79, 2010. An open access article.

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, vacc

The genus Orthopoxvirus contains several species of related viruses, including the causative agent of smallpox (Variola virus). Despite the eradication of smallpox, several other members of the genus are capable of causing human infection, including monkeypox, cowpox, and other zoonotic rodent-borne poxviruses. Therefore, a single assay that can accurately identify all orthopoxviruses could provide a valuable tool for rapid detection and surveillance. In this study, we developed a pan-Orthopoxvirus assay based on PCR/electrospray ionization-mass spectrometry (PCR/ESI-MS) on the Ibis T5000 system. The assay employs four PCR reactions targeting the orthopoxvirus DNA and RNA and helicase and polymerase genes. In this study, we evaluated this assay for its ability to correctly identify a panel of known orthopoxviruses and to determine the assay's ability to detect and identify virus from the blood of rabbitpox virus-infected rabbits. We demonstrate that the assay can broadly detect and identify a diverse collection of orthopoxviruses and the assay is sensitive down to a few plaque-forming units per milliliter of blood and the stochastic limits of PCR., The original document contains color images. Pub. in PLoS One, v4 n7, pe6432-e6342, Jul 2009 Sponsored in part by Defense Threat Reduction Agency and Office Biodefense Research Affairs, National Institute of Allergy and Infectious Diseases. USAMRIID Research Plan no. 114538.

Infection of rabbits with aerosolized rabbitpox virus (RPXV) produces a disease similar to monkeypox and smallpox in humans and provides a valuable, informative model system to test medical countermeasures against orthopoxviruses. Due to the eradication of smallpox, efficacy evaluation of new-generation smallpox vaccines depends on relevant well-developed animal efficacy studies for vaccine licensure. In this study, we tested the efficacy of IMVAMUNE [Modified Vaccinia Virus Ankara-Bavarian Nordic (MVA-BN )] for protecting rabbits against aerosolized RPXV. Rabbits were vaccinated with either phosphate-buffered saline (PBS), Dryvax , a single low dose of IMVAMUNE , a single high dose of IMVAMUNE , or twice with a high dose of IMVAMUNE . Aerosol challenge with a lethal dose of RPXV was performed 4 weeks after the last vaccination. All PBS control animals succumbed to the disease or were euthanized because of the disease within 7 days postchallenge. The rabbits vaccinated with Dryvax , a low dose of IMVAMUNE , or a single high dose of IMVAMUNE showed minimal to moderate clinical signs of the disease, and all of them recovered. The only clinical sign displayed by rabbits that had been vaccinated twice with a high dose of IMVAMUNE was mild transient anorexia in just two animals. This study shows that IMVAMUNE can be a very effective vaccine against aerosolized RPXV., The original document contains color images. Pub. in Vaccine, v27 p5496-5504, 2009.

Orthopoxviruses, such as variola and monkeypox viruses, can cause severe disease in humans when delivered by the aerosol route, and thus represent significant threats to both military and civilian populations. Currently, there are no antiviral therapies approved by the U.S. Food and Drug Administration (FDA) to treat smallpox or monkeypox infection. In this study, we showed that administration of the antiviral compound ST-246 to rabbits by oral gavage, once daily for 14 days beginning 1h postexposure (p.e.), resulted in 100% survival in a lethal aerosolized rabbitpox model used as a surrogate for smallpox. Furthermore, efficacy of delayed treatment with ST-246 was evaluated by beginning treatment on days 1, 2, 3, and 4 p.e. Although a limited number of rabbits showed less severe signs of the rabbitpox disease from the day 1 and day 2 p.e. treatment groups, their illness resolved very quickly, and the survival rates for these group of rabbits were 88% and 100%, respectively. But when the treatment was started on days 3 or 4 p.e., survival was 67% and 33%, respectively. This work suggests that ST-246 is a very potent antiviral compound against aerosolized rabbitpox in rabbits and should be investigated for further development for all orthopoxvirus diseases., The original document contains color images. Pub. in Antiviral Research v79, p121-127, 2008. Prepared in cooperation with STGA Technologies, Inc., Corvallis, OR.

Arthropod-borne viruses ("arboviruses") cause significant human illness ranging from mild, asymptomatic infection to fatal encephalitis or hemorrhagic fever. The most significant arboviruses causing human illness belong to genera in three viral families, Togaviridae, Flaviviridae, and Bunyaviridae. These viruses represent a significant public health threat to many parts of the world, and, as evidenced by the recent introduction of the West Nile virus (WNV) to the Western Hemisphere, they can no longer be considered specific to any one country or region of the world. Like most viral diseases, there are no specific therapies for the arboviral encephalitides; therefore, effective vaccines remain the front line of defense for these diseases. With this in mind, the development of new, more effective vaccines and the appropriate animal models in which to test them become paramount. In fact, for many important arboviruses (e.g. California serogroup and St. Louis encephalitis viruses), there are currently no approved vaccines available for human use. For others, such as the alphaviruses, human vaccines are available only as Investigational New Drugs, and thus are not in widespread use. On the other hand, safe and effective vaccines against tick-borne encephalitis virus (TBEV) and Japanese encephalitis virus (JEV) have been in use for decades. New challenges in vaccine development have been met with new technologies in vaccine research. Many of the newer vaccines are now being developed by recombinant DNA technology. For example, chimeric virus vaccines have been developed using infectious clone technology for many of the arboviruses including, WNV, JEV, and TBEV. Other successful approaches have involved the use of naked DNA encoding and subsequently expressing the desired protective epitopes. Naked DNA vaccines have been used for TBEV and JEV and are currently under development for use against WNV., Pub. in Antiviral Research, v60, p153-174, 2003.