Mousepox and bioweapons
by Danger Mouse Dave
Australian scientists tried to create a strain of the mousepox virus that they thought would cause sterility in female mice. What they created was a killer virus that vaccination could only partially control. The research has prompted fears that the strategy could be used for the development of biological weapons.
Reference
Expression of Mouse Interleukin-4 by a Recombinant Ectromelia Virus Suppresses Cytolytic Lymphocyte Responses and Overcomes Genetic Resistance to Mousepox
Ronald J. Jackson et al.
Journal of Virology, February 2001, p. 1205-1210, Vol. 75
Periodic mouse plagues that occur in rural areas can eat their way through thousands of hectares of crops. Combating these plagues usually involves air drops of poisoned baits. Whilst this is sometimes effective, a more reliable and specific method of controlling mouse populations is needed.
So scientists at Canberra’s Co-operative Research Centre (CRC) for the Biological Control of Pest Animals teamed up with the John Curtin School of Medical Research at the Australian National University in Canberra to create a strain of the mousepox virus that would cause sterility in female mice.
Left: The vaccinia (cowpox) virus
Mousepox is a virus that belongs to a family of viruses known as the Poxviridae. They are double-stranded DNA viruses. The pox viruses take their name from the pustules (pocks) that erupt on the skin of infected organisms. These pustules contain fluid teaming with newly made virus particles. Physical contact is the primary mode of transmission of the virus. They are a significant family of viruses in that there is a pox virus for just about every mammal you can think of - foulpox, camelpox, mousepox, cowpox, monkeypox, chickenpox, swinepox, foxpox, sealpox, dolphinpox kangaroopox and many many more. There are also pox viruses for invertebrates. Most pox viruses are specific for their host, although there are examples of pox viruses that can infect multiple species. There are around 20 pox viruses that can infect humans. The most common and well known is chickenpox. The name is misleading in that it is not a chicken virus, it is a human virus known as the Varicella Zoster virus.
Left: Pharaoh Ramses V
For the most part, pox viruses cause relatively minor infections that leave the organism immune to future infections. There are some notable exceptions to this generalisation. The Varicella Zoster virus can cause chickenpox (mostly in childhood) but can recur later in life and cause the painful condition known as Shingles. The biggest exception is a virus known as the Variola virus. This causes an infection known as smallpox, one of history’s deadliest diseases. Smallpox was one of the most contagious and virulent diseases ever known. It killed countless millions across the world, especially in Europe, India and China. The Pharaoh Ramses V died of smallpox in 1157 BC. The disease reached Europe in 710 AD and was transferred to America by Hernando Cortez in 1520. 3.5 million Aztecs died in the next 2 years. In the cities of 18th century Europe, smallpox reached plague proportions and was a feared scourge. Five reigning European monarchs died from smallpox during the 18th century. In Europe, nearly everyone caught it at some stage in their lives. About 10-20% of infected people died as a result. Of the survivors, around 15% were permanently disfigured by the scars left from the pustules that covered the body. The English physician Edward Jenner developed the first smallpox vaccine 1798 by discovering that inoculating people with cowpox stimulated immunity to both cowpox and smallpox. Jenner coined the terms "vaccination" and "vaccine" from the name for the cowpox virus (the Vaccinia virus), which in turn comes from the Latin for cow (vacca). Since then better smallpox vaccines have been developed. The World Health Organization’s world-wide smallpox vaccination campaign has resulted in the eradication of the disease. The last recorded case was in Somalia in 1977. Samples of the smallpox virus are kept in various high-security laboratories in the USA and Russia.
The scientists genetically engineered a mousepox virus to carry the mouse egg shell protein ZP3, or zona pellucida 3, as a mouse contraceptive. The reasoning was that by infecting mice with the engineered virus, the mice would contract a mild mousepox infection and launch an immune response to the virus. But the virus expresses the mouse egg protein, so infected mice would not only make antibodies against the virus, but also against the egg protein. In females, these anti-ZP3 antibodies would then attack their own eggs in their ovaries and cause sterility whilst leaving the mice healthy (once they recovered from the mild mousepox disease) and free to mate with males. Thus, these sterile females would "dilute out" the effectiveness of breeding males in a population. This was the theory and it seemed to work quite well in one laboratory strain of mouse, but in other strains it was ineffective. If the virus is to be useful in the wild then it will have to be able to work across different strains of mice.
In order to try and make the engineered virus more effective across different strains, the scientists inserted another gene into the virus - the interleukin-4 gene (IL-4). This gene codes for a cytokine (or hormone) that is one of the many that regulate the functioning of the immune system. There was previous work that suggested a virus expressing IL-4 would increase the antibody-producing response in mice and tone down the effectiveness of virus-clearing cells of the immune system (called "killer T-cells"). It was hoped that this would increase the immune response to the ZP3 protein and make the virus an effective contraceptive across multiple strains of mice.
The results from the newly engineered mousepox virus were very unexpected. They expected only to strengthen the antibody response in resistant strains, but instead the virus overwhelmed the mice, proliferating out of control and destroying their livers. Even mice that had been vaccinated against mousepox (which is normally extremely effective at conferring resistance) fared poorly, with half dying immediately and the remainder developing a chronic abscess at the site of infection.
On a virology and immunology level, this is a very interesting result. But the implications go far beyond that. The goal of the research was certainly benign, but the study provided the first evidence to suggest that all it takes to transform an innocuous virus into a deadly virus is the insertion of a single gene. Something that was thought to be hard - increasing the pathogenicity of a virus - appears, in this case, to be easy. This has some alarming implications for the development of biological weapons. Up until now the concerns regarding biological weapons centred on the use of existing pathogens. A terrorist’s ultimate aim would be to obtain a sample of smallpox. It has been 23 years since anyone’s immune system has seen the smallpox virus. Smallpox vaccination is no longer included in the standard course of childhood vaccinations, and stocks of the vaccine are low. But smallpox is very difficult to obtain, so the next best options are bacterial pathogens - Bacillus anthracis (which causes Anthrax) and Yersinia pestis (which causes The Plague). However, when it comes to the large scale production of these pathogens, biological weapons inspectors know what to look for. The specific facilities and reagents needed are a dead give-away. The mousepox result, however, may indicate that commonly used technology found in any biotechnology laboratory in the world could be used to create new viruses that overwhelm resistance and render vaccination useless. The close relationship between the pox viruses raises the question as to whether it would be possible to transform other members of the family, including those that infect humans. By inserting IL-4 into chickenpox, would it be possible to transform chickenpox into a virus that is more deadly than smallpox?
The team spent 18 months confirming the data and debating whether to go public with them. In the end, disclosure won out over concerns about educating future bioterrorists and alarming the public. In an interesting twist, on publication of the results it was not environmentalists or media commentators that were sounding the warning, but the scientists themselves. On 16 January the CRC issued a press release timed to accompany the article that pleaded for stronger measures to combat the threat of biowarfare arising from such good intentions. Not surprisingly, the press release triggered sensational warnings in the Australian media and elsewhere. The scientists said that it should serve as a warning to the community to be more aware of the potentially harmful consequences of their work. "We need the public to trust us if we are going to seek their approval to release pest-control viruses down the track," says CRC director Bob Seamark, who led the research. But any intentional release, he hastens to add, won't involve viruses carrying the IL-4 gene. "These are confined to the high-security lab."
Despite the warnings, it's not clear whether the unexpected result, which turned a vector into a potent killer, could be duplicated in viruses that affect humans. Such fears may be overrated, says Ron Jackson, the CRC virologist who carried out much of the work. Jackson suspects that the findings may be peculiar to the mousepox virus, which naturally carries other proteins that weaken the antiviral response. He notes that the same result did not occur with the vaccinia virus (cowpox) in other experiments.
Deterring bioweapons development
The implications of this finding are of intense interest to organizations such as the Federation of American Scientists, which has formed a working group to develop a protocol that would add verification powers to the currently toothless international convention on biological weapons - the 1975 Biological and Toxin Weapons Convention (BTWC).
The emerging fields of genomics and proteomics holds great potential for the development of new useful biological reagents, but many of these will also have utility as biological weapons, or will suggest ways of creating such weapons. The treaty does not prohibit research, but it does prohibit the development, production, or stockpiling of biological or toxic agents and of devices to deliver such agents for other than peaceful purposes. However, with no provisions for verification, the treaty has proved to be a weak deterrent to nations committed to biological weapons development.
For this reason, an addendum to the BTWC has been negotiated over the past five years. The addendum calls for….
- annual declarations of facilities with the potential for use in a biological weapons program.
- random visits to such facilities by teams of international inspectors.
- establishing a mechanism for investigation of suspicions of violation of the BTWC.
Its adoption would significantly improve international security and reduce the risk of bioterrorism by inhibiting bioweapons development. The United States, however, has consistently delayed progress and pressed for a weakening of the new provisions, and now might completely derail the negotiations by stalling past the deadline imposed for completion of the addendum.
Synthetic viruses just around the corner (Nature Science Update)
Killer virus (New Scientist)
Bugs of war (New Scientist)