MicrobiologyBytes

It seems like only yesterday we were on the verge of international agreement to destroy all remaing laboratory stocks of smallpox (variola) virus. So it comes as something of a surprise to find people infecting monkeys with smallpox to study the pathogenesis of the disease. Such studies significantly advance our understanding of variola pathogenesis in primates and could help development of new antiviral drugs, improved bioterrorism countermeasures, and suggest new potential targets for therapeutic intervention in humans.

But is it a good idea?

Progression of Pathogenic Events in Cynomolgus Macaques Infected with Variola Virus. (2011) PLoS ONE 6(10): e24832. doi:10.1371/journal.pone.0024832
Smallpox, caused by variola virus (VARV), is a devastating human disease that affected millions worldwide until the virus was eradicated in the 1970 s. Subsequent cessation of vaccination has resulted in an immunologically naive human population that would be at risk should VARV be used as an agent of bioterrorism. The development of antivirals and improved vaccines to counter this threat would be facilitated by the development of animal models using authentic VARV. Towards this end, cynomolgus macaques were identified as adequate hosts for VARV, developing ordinary or hemorrhagic smallpox in a dose-dependent fashion. To further refine this model, we performed a serial sampling study on macaques exposed to doses of VARV strain Harper calibrated to induce ordinary or hemorrhagic disease. Several key differences were noted between these models. In the ordinary smallpox model, lymphoid and myeloid hyperplasias were consistently found whereas lymphocytolysis and hematopoietic necrosis developed in hemorrhagic smallpox. Viral antigen accumulation, as assessed immunohistochemically, was mild and transient in the ordinary smallpox model. In contrast, in the hemorrhagic model antigen distribution was widespread and included tissues and cells not involved in the ordinary model. Hemorrhagic smallpox developed only in the presence of secondary bacterial infections – an observation also commonly noted in historical reports of human smallpox. Together, our results support the macaque model as an excellent surrogate for human smallpox in terms of disease onset, acute disease course, and gross and histopathological lesions.

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In 2011 the World Health Organization (WHO) plans to announce its recommendation regarding the final destruction of all known remaining smallpox virus stockpiles. Smallpox, an ancient human scourge of unparalleled destructive importance throughout most of recorded human history, is believed to have emerged in the Middle East some 6,000–10,000 years ago from either camelpox or the gerbil-specific taterapox. It holds a status as one of the great killers in all human history, having produced the horrific deaths of up to 500 million persons in just the 20th century alone. At first glance, the answer to this conundrum – whether or not smallpox should be forever relegated to the autoclave of extinction – might seem an easy one. Beaten back by the Jenner vaccine first proposed in 1796, smallpox was finally declared eradicated in 1980, in one of the most profound public health achievements in human history. Since that time, WHO has made it generally known that they would like to see the elimination of all remaining variola stockpiles and made the United States and Russia the repository for all remaining stocks. At the 60th Annual World Health Assembly in 2007, the organization postponed the final decision for any recommended destruction deadline until their next meeting in 2011.

Should Remaining Stockpiles of Smallpox Virus (Variola) Be Destroyed? Emerging Infectious Diseases 17(4) April 2011

“Twenty miles south-east of Novosibirsk, in Siberia, several dozen concrete buildings have been erected outside the town of Koltsovo. The settlement is ringed with triple rows of barbed wire fences. Video cameras and motion sensors monitor any activity near the wires while soldiers from an elite Russian army unit patrol its perimeter.
This is Russia’s State Research Centre of Virology and Biotechnology – or Vector, as it is usually known. Frozen in winter, when temperatures plunge below -30^oC, and then scorched in summer, when the heat routinely rises above 30^oC, the place is as unwelcoming as you could imagine. Given its name, location and a high-security protection, Vector would make an ideal setting for a James Bond film.
This would be a fitting accolade, for Vector contains a number of unsettling scientific secrets, with the most sinister being housed in bio-containment laboratory P-4, in Building 6. Here a small storage plant, chilled by liquid nitrogen, holds phials of one of the deadliest pathogens known to medical science: the smallpox virus…”

Read more: Smallpox virus: crunch time for the fate of a global killer | The Observer

Should we destroy the remaining laboratory stocks of smallpox virus?

Now that the 20th century has passed into the domain of history books, we can retrospectively begin to assess the relative contributions that the many advances in the realm of infectious disease have actually made to public health in general. At the top of this virtuous list will surely be the discovery of antibiotics in the 1930s and the use of vaccination to eradicate smallpox as an extant human disease in the 1960s and 1970s. As clearly pointed out in a recent book by D. A. Henderson, one of the leaders of the global smallpox eradication program, this task of ridding Homo sapiens from the curse of this ancestral disease was neither easy nor without controversy. In fact, the history of the many consequences of smallpox on humankind reads like a long litany of human misery and calamitous events, but is juxtaposed with the more noble accomplishments that began with the discovery of vaccination by Jenner in 1798 and culminated with the World Health Organization (WHO) certifying the world free of smallpox in 1980. With this singular accomplishment, as many as 60–100 million individuals who would have been predicted to die of smallpox have been spared from a truly gruesome death. Nevertheless, the narrative of smallpox did not stop with its eradication as a pandemic human disease. Instead, we find ourselves still wrestling with an issue that intermingles public health policy, philosophy, national security, and bioterrorism, and affects our perceptions of research ethics with extreme pathogens in general. It boils down to a not-so-simple question: What exactly should the Victor do with the Vanquished?

Killing a Killer: What Next for Smallpox? 2010 PLoS Pathog 6(1): e1000727. doi:10.1371/journal.ppat.1000727

Related:

• W.H.O. ducks smallpox decision (again)

Melioidosis, which can present as an acute septic illness or as a chronic low-grade infection, was first described in Burma more than 100 years ago. Largely due to its recognition as a biological threat agent, current knowledge on melioidosis, caused by the Gram-negative bacterium Burkholderia pseudomallei, has increased tremendously over recent years as more and more research groups are working on this disease. The global distribution boundaries of melioidosis continue to expand well beyond the traditionally recognized endemic region of southeast-Asia and northern-Australia. For instance, new cases have been described in Brazil and southeast-Africa, although it has to be clarified whether melioidosis is indeed endemic in these regions. Of interest, phylogenetic studies have recently hypothesized that B. pseudomallei has originated in tropical Australia with subsequent spread to southeast-Asia (dubbed as the Gondwana hypothesis). Pneumonia with bacterial dissemination to distant sites and abscess formation is a common presentation. Current treatment recommendations on the basis of clinical trial evidence are parental ceftazidim or carbapenem for 10–14 days or longer as clinically indicated, followed by oral trimethoprim–sulfamethoxazole ± doxycycline for at least 12–20 weeks. This review summarizes present knowledge on the molecular characterization of B. pseudomallei and the immunology of melioidosis with a special emphasis on its potential therapeutic implications.

Immunity to Burkholderia pseudomallei. Curr Opin Infect Dis. 2009 22(2): 102-108
Largely due to its recognition as a biological threat agent, current knowledge on melioidosis, caused by the Gram-negative bacterium Burkholderia pseudomallei, has increased tremendously over the last years. This review summarizes current understanding on the molecular characterization of B. pseudomallei and the immunology of melioidosis. The genome of B. pseudomallei is composed of two chromosomes of which the largest part represents the B. pseudomallei core genome, whereas the remaining accessory genome has been associated with bacterial virulence. Virulence factors, most notably quorum sensing, type III secretion system, lipopolysaccharide and other surface polysaccharides, flagella and various factors essential for the intracellular life cycle of B. pseudomallei, have been further characterized. The neutrophils play a critical in host defense, which is initiated by the Toll-like receptors. The proinflammatory immune response – including the activation of coagulation – and its regulation have been further dissected. Severe melioidosis can probably be seen as the clinical manifestation of a pathogen recognition receptor mediated dysregulation of the immune response to invading B. pseudomallei. B. pseudomallei employs numerous tactics to evade the immune response. Studies on host-pathogen interactions in melioidosis have identified a whole range of potential new treatment targets.

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• Glanders and Melioidosis – coming soon to a crowded shopping centre near you?
• Pathogenic soil bacterium is influenced by land management

Releasing highly pathogenic organisms into an urban population is a act of bioterrorism that could result in a large number of casualties. The first indication that a covert open-air release has occurred is quite likely to be individuals reporting for medical attention. If such an attack is suspected, then public health authorities would attempt to identify those individuals who have been infected in order to provide rapid treatment with the aim of reducing the possibility of disease and potential death. Aiming treatment at too small an area might miss individuals infected further down and/or up wind, whereas issues surrounding both treatment resources and serious side effects may rule out mass treatment campaigns of large sections of the population.

A new paper describes a statistical method that can estimate the origin and time of an aerosolized release of anthrax following detection of the first few cases. The method predicts where the most critically affected areas will be following the release of this highly pathogenic agent, which may enable preventative treatment of individuals at risk and protection from the disease. Previously published methods can estimate the date and scale of anthrax release but not the source location or geographic extent of human exposure. The new method uses information about the first people infected, including when they started to experience symptoms of infection and where they live and work, combined with recent weather information, such as wind direction. Anthrax has the potential to cause a large number of deaths in the event of a covert, open air release. If such a release were to occur, it is critical for public health decision makers to evaluate its extent and the potential impact on the population and then to identify the people most at risk of infection as soon as possible. It is critical to treat people as soon as possible after exposure to anthrax. While forecasts based on small numbers of early cases are less reliable than those obtained later in an outbreak, treating individuals based on early estimates is still likely to save lives overall.

Estimating the Location and Spatial Extent of a Covert Anthrax Release. 2009 PLoS Comput Biol 5(1): e1000356
Rapidly identifying the features of a covert release of an agent such as anthrax could help to inform the planning of public health mitigation strategies. Previous studies have sought to estimate the time and size of a bioterror attack based on the symptomatic onset dates of early cases. We extend the scope of these methods by proposing a method for characterizing the time, strength, and also the location of an aerosolized pathogen release. A back-calculation method is developed allowing the characterization of the release based on the data on the first few observed cases of the subsequent outbreak, meteorological data, population densities, and data on population travel patterns. We evaluate this method on small simulated anthrax outbreaks (about 25–35 cases) and show that it could date and localize a release after a few cases have been observed, although misspecifications of the spore dispersion model, or the within-host dynamics model, on which the method relies can bias the estimates. Our method could also provide an estimate of the outbreak’s geographical extent and, as a consequence, could help to identify populations at risk and, therefore, requiring prophylactic treatment. Our analysis demonstrates that while estimates based on the first ten or 15 observed cases were more accurate and less sensitive to model misspecifications than those based on five cases, overall mortality is minimized by targeting prophylactic treatment early on the basis of estimates made using data on the first five cases. The method we propose could provide early estimates of the time, strength, and location of an aerosolized anthrax release and the geographical extent of the subsequent outbreak. In addition, estimates of release features could be used to parameterize more detailed models allowing the simulation of control strategies and intervention logistics.

There is a pressing need for antiviral agents that are effective against multiple classes of viruses. Broad specificity might be achieved by targeting phospholipids that are widely expressed on infected host cells or viral envelopes. We reasoned that events occurring during virus replication (for example, cell activation or preapoptotic changes) would trigger the exposure of normally intracellular anionic phospholipids on the outer surface of virus-infected cells. A chimeric antibody, bavituximab, was used to identify and target the exposed anionic phospholipids. Infection of cells with Pichinde virus (a model for Lassa fever virus, a potential bioterrorism agent) led to the exposure of anionic phospholipids. Bavituximab treatment cured overt disease in guinea pigs lethally infected with Pichinde virus. Direct clearance of infectious virus from the blood and antibody-dependent cellular cytotoxicity of virus-infected cells seemed to be the major antiviral mechanisms. Combination therapy with bavituximab and ribavirin was more effective than either drug alone. Bavituximab also bound to cells infected with multiple other viruses and rescued mice with lethal mouse cytomegalovirus infections. Targeting exposed anionic phospholipids with bavituximab seems to be safe and effective. Our study demonstrates that anionic phospholipids on infected host cells and virions may provide a new target for the generation of antiviral agents.

Targeting inside-out phosphatidylserine as a therapeutic strategy for viral diseases. 2008 Nature Medicine 14: 1357-1362

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In the years during and after World War II, the Microbiological Research Establishment (MRE) at Porton Down was the UK Government’s centre for germ warfare research (or defence, depending on who you believe). There were two parts to the MRE, the part “outside the wire”, which contained among other things, the Common Cold Unit, and the section “inside the wire”, which was top secret and has been the subject of much rumour and speculation.

I recently came across an article by Bill Parker, who worked at the MRE for two years and which lifts some of the curtain of secrecy. Makes interesting reading.

Radionuclides in the environment are one of the major concerns to human health and ecotoxicology. The explosion at the Chernobyl nuclear power plant renewed interest in the role played by fungi in mediating radionuclide movement in ecosystems. As a result of these studies, our knowledge of the importance of fungi, especially in their mycorrhizal habit, in long-term accumulation of radionuclides, transfer up the food chain and regulation of accumulation by their host plants was increased. Micro-fungi have been found to be highly resilient to exposure to ionizing radiation, with fungi having been isolated from within and around the Chernobyl plant. Radioresistance of some fungal species has been linked to the presence of melanin, which has been shown to have emerging properties of acting as an energy transporter for metabolism and has been implicated in enhancing hyphal growth and directed growth of sensitized hyphae towards sources of radiation. Using this recently acquired knowledge, we may be in a better position to suggest the use of fungi in bioremediation of radioactively contaminated sites and cleanup of industrial effluent.

Fungi appear to be very resistant to radionuclides in the environment. Part of this resistance may be the smaller amount of DNA per nucleus than mammalian cells, but evidence from comparative studies between pigment and nonpigmented fungi suggest there may be other factors that confer radioresistance. Melanin pigment may provide some protection against ionizing radiation and be integral to the absorption and retention of radionuclides. Owing to the long-lived and extensive hyphal network and biomass in upper soil horizons of forest ecosystems, fungi are very efficient in absorbing radionuclides, and are an important component of long term accumulation of radionuclides. This may be why radionuclide adsorption/desorption models do not conform to observed patterns, as mycorrhizal fungi mediate soil-to-plant transfer rates. Internal translocation of radionuclides to and hyper-accumulation into harvestable fruit bodies of macro-fungi has potential for environmental remediation but needs further evaluation.