We'd Better Think Twice Before Eradicating All Smallpox Virus Stocks

The global eradication of smallpox as a threat to human health is one of the milestone achievements of modern medical science. It was accomplished through a unique international collaboration sponsored by the World Health Organization (WHO) and directed by virologist Donald A. Henderson, who now serves as deputy assistant secretary in the United States Department of Health and Human Services. The principal research centers supporting the eradication program were in Moscow and in Atlanta. While this collaboration was proceeding, the Cold War also nourished a high degree of mutual suspicion between the U.S. and the then-USSR about the possible abuse of such research for potential application in biological weaponry. While there are many technical reasons for giving smallpox very low priority as a weapon threat, there was great pressure to conclude an agreement to destroy all research stocks of the smallpox (variola) virus, as part of confidence-building between the superpowers. Meanwhile, the world has experienced enormous changes. Russia is no longer a military threat, and it is clear that its civilian research establishment shares the same medical and scientific values as the rest of the world's scientific community. Any lingering questions about secret military work would not be resolved by the proposed destruction of research stocks in civilian hands. We have also become sensitized to the fate of many other species, and now dedicate considerable political and economic resources to the conservation of natural biodiversity. Thus, a reexamination of the policies about smallpox is very much in order. The following appeal to reexamine the proposed destruction of smallpox virus stocks is made by Russian biologist Lev Sandakhchiev, who has played a notable part in the scientific study of the smallpox and other viruses. Joshua Lederberg University Professor Rockefeller University New York In 1980, the World Health Assembly announced the complete eradication of smallpox disease throughout the world--an achievement brought about by an international program overseen by the World Health Organization (WHO) for more than 20 years. Since 1984, the only collections of the variola virus (VAR)--the cause of smallpox--have been stored at two WHO Collaborating Centers on Smallpox and Related Infections: the Centers for Disease Control and Prevention (CDC) in Atlanta and the Institute for Viral Preparations in Moscow. In March 1986, an ad hoc WHO panel--the Committee on Orthopoxvirus Infections--recommended that, since smallpox had been officially eradicated, all remaining collections of variola virus be destroyed. And in May 1990, Louis W. Sullivan, then the United States Health and Human Services secretary, asked WHO to see that the VAR collections were indeed eradicated as soon as the complete sequencing of the VAR genome had been achieved. Later that same year, in December 1990, the ad hoc WHO committee- -the same panel that in 1986 recommended the destruction of all remaining variola virus collections--approved projects by American and Russian scientists aimed at sequencing the genomes `of various VAR strains. To accommodate this process, the committee recommended the destruction of all existing VAR stocks and recombinant plasmids containing VAR DNA fragments by Dec. 31, 1993, given that sufficient sequence information was ascertained by that time. However, Russian investigators (working under the aegis of the National Program on Conservation of Genetic Material of the Russian Collection of Variola Virus Strains) believe that the recommended eradication of VAR stocks by the end of this year is not justified from the scientific point of view and is not supported by the available data concerning the structural and functional organization of the variola virus genome. So far, we have accomplished the sequencing of the complete coding region of variola major virus strain India-1967, excluding the non-coding short inverted terminal DNA fragments. Currently, the Russian researchers, together with American scientists based at CDC, are performing the DNA sequencing of the variola minor virus strain Garcia-1966. The CDC scientists are also carrying out the sequencing of the complete genome of the variola major virus strain Bangladesh-1975. Moreover, genomic fragments of several other VAR strains are being sequenced simultaneously by British scientists at Oxford University. To date, the following information concerning VAR genomic structure has been revealed: The organization of VAR terminal inverted sequences is unique. It is not yet understood, however, whether and how it affects the manifestation of biological features of this virus. Evidently, there is no possibility of re-creating the terminal structures of the VAR genome through recombination of VAR DNA fragments cloned in the molecular vectors, and the genomes of some other orthopoxviruses. A major part of VAR genes is highly conserved with respect to vaccinia virus. A number of VAR genes have revealed code for the truncated versions of the analogous vaccinia proteins. Several VAR long coding sequences have the truncated analogs in the genome of vaccinia virus. We suggest that VAR and vaccinia virus originated from a common ancestor and that cowpox virus (CPV), persisting now, is likely to be the analog of the ancestor. The ultimate conclusion on the origin of VAR and vaccinia viruses demands the genome sequencing of at least one CPV strain. A variety of genes were revealed in the genome of VAR code for the proteins analogous to TNF-receptor, gamma-IFN-receptor, and complement-binding proteins, as well as the sets of proteins belonging to the family of serine protease inhibitors, ankyrin- like proteins, proteins of the T6/8/9 family of tumorigenic Shope fibroma virus, and others. Several revealed VAR genes encode the proteins of unique structure, which have no analogs among the other poxviruses. However, to affirm the unique character of these VAR genes and their genomic structure on the whole, it is necessary to sequence the genomes of monkeypox (MPV) and cowpox viruses, both being pathogenic for humans. Basing our opinion on the above results and on our understanding of the ecology and evolution of orthopoxviruses, we consider the following conclusions essential: There is no sense in the destruction of collections of recombinant plasmids containing VAR genome fragments. These plasmids can be handled in complete safety. There is no way to reconstruct any variola-like virus with the help of these plasmids. On the contrary, they can be quite useful. Using the polymerase chain reaction (PCR), one can amplify the chosen coding sequences without viral promoters and transcription terminators, insert these sequences into the expression plasmids, and synthesize the VAR proteins with subsequent study of their characteristics and comparison with analogous proteins of the other poxviruses. To our minds, a number of VAR proteins, especially those possessing pronounced immunomodulating features, can initiate the development of unique medical preparations. There is no sense in the eradication of VAR stocks until the sequencing of genomes of human pathogenic monkeypox and cowpox viruses is completed. MPV and CPV are zoonotic and persisting at present in nature. The present level of knowledge does not allow one to predict with validity the possibility (or impossibility) of the formation of a variola-like virus in nature through recombinational and evolutionary changes of monkeypox and/or cowpox viruses. Thus, along with studying VAR, it is essential to pay more attention to investigating the structural and functional organization of MPV and CPV. The WHO Technical Committee has recommended carrying out the complete genome sequencing of the variola minor virus strain Garcia-1966, along with the complete sequencing of the variola major virus strains India-1967 and Bangladesh-1975. This work is not likely to be completed before 1994, if we consider the previous experience within the frames of the variola genome sequencing program. Authenticity of the determined structure of the viral genome-- that is, its correspondence to the native structure of original natural isolate--presents a real problem. All viral strains under sequencing by now have undergone several passages on chorioallantoic membranes of developing chick embryos and/or cell cultures. Nobody knows what changes in VAR genome can possibly have arisen as a result of these passages. On the other hand, the process of cloning via plaques involves the selection of accidental variants, which can differ considerably from the dominant structure of the genome to the complete loss of a number of genes. Only the comparative study of several isolates that have not undergone the laboratory passages and a strain that has been cultivated for a long time (such as Harvey-44) can clarify the situation. After the comparison of the genomic sequences of various VAR strains, it obviously will be necessary to PCR-amplify and subsequently sequence at least some of the essential VAR DNA regions from stored unpassaged human material. It is evident that such work could not be completed by the end of 1993. No information is available now concerning the rate of mutation accumulation in the VAR genome on the level of both amino acid substitutions in proteins and "silent" nucleotide changes. The molecular mechanism of this process on the population level is also unknown. It should be noted that comparison of constant genomic regions rather than the variable ones would give the most important results as well as the comparison of silent substitutions in the coding regions that are not subjected to the direct selection pressure of antibodies and tissue-specificity changes. Data concerning the inner microheterogeneity of natural populations of this virus are almost completely lacking at this point. Clarification of this question requires the sequencing of a considerable number of clones of the same genomic regions obtained from several viruses that have not been adapted to laboratory cultivation. Destruction of VAR collections at the WHO Collaborating Centers on Smallpox and Related Infections does not ensure the complete eradication of VAR samples from the Earth. The possibility of VAR stocks being preserved in some laboratory not under WHO's control cannot be ruled out; nor can preservation of the virus in the tissues of patients who have died of smallpox. If the world community considers that the existing WHO Collaborating Centers do not provide a guarantee against the use of VAR as a potential biological weapon, we suggest the organization of an International Laboratory on Smallpox. This laboratory would house scientists from different countries; would have codirectors from, for example, the U.S. and Russia; would work under constant control of WHO (or the United Nations); and would provide both the preservation of VAR strains and the advanced studies of genomic organization and evolutionary changes of VAR and other orthopoxviruses. In the permafrost region of Russia, there are burial grounds of smallpox victims. It is not impossible that viable viruses are still stored in such places, which is of great danger to mankind. During summer periods since 1991, such a possibility has been tested. Study of the tissue samples taken from corpses with skin lesions characteristic of smallpox revealed the orthopoxviral antigens but failed to reveal the viable virus. However, these studies have not yet been completed--and there is no point in eradication of VAR collections before the completion of this work. To sum up, the modern level of knowledge and laboratory methodology about viruses is limited. The complete sequence of the viral genome does not make available the complete knowledge of viral features. It is not possible to predict what methods will be developed in the future and in what way they could be used to characterize VAR stocks. The untimely destruction of viral collections could lead to the irrevocable loss of information concerning the pathways and mechanisms of variola virus evolution, especially its population aspects, and would as well eliminate the possibility of understanding the molecular epidemiological regularities of population interactions of virus and host. Lev S. Sandakhchiev is director of the Vector Laboratories of the All Union Molecular Biology Research Institute Koltsovo, Novosibirsk, Russia.

We'd Better Think Twice Before Eradicating All Smallpox Virus Stocks

Interested in reading more?

Become a Member of