Viral eradication and the evolution of smallpox

The control and eventual eradication of the smallpox virus from the wild is one of the most heralded success stories in all of public health. Indeed, smallpox has played a central role in the history of vaccination. Even prior to Edward Jenner's use of the related cowpox virus to protect against smallpox disease, it was known that inoculation with materials from an infectious smallpox pustule or scab (dubbed "variolation") could protect an individual from death due to smallpox, generally resulting instead in a mild form of the illness. Jenner's observation that milkmaids seemed to be protected from the disease--and his use of material from cowpox pustules instead of smallpox--resulted in the development of the science of vaccination. World-wide use of the smallpox vaccine, along with a mass vaccination campaign led by the World Health Orgainzation, resulted in the end of naturally-occurring smallpox on the planet, with the exception of stores of the virus held in the United States and Russia.

This feat is being attempted currently with measles and polio viruses, but it's been much more difficult, and eradication of these viruses may never be attainable. Below I discuss some aspects that are critical to a campaign that seeks to eradicate a disease, and a new paper on the evolution of smallpox viruses.

A number of conditions must first be met to even consider an organism for eradication.

First, the organism must be a pathogen only of humans, and can't have a non-human reservoir. Imagine the difficulty of trying to eradicate something like E. coli or Salmonella, which have a practically endless reservoir in animals and the environment.

Second, the infection must induce long-lasting immunity. An organism that can re-infect a person multiple times, or has a number of different serotypes (such as influenza) is not a good candidate for eradication. Additionally, a vaccine that requires only a single dose for long-lasting protection is also ideal.

Third, to go along with #2, there must be a tool or intervention that stops the chain of transmission between individuals. Though this is often assumed to be a vaccine, it could be an antimicrobial drug, or even a physical quarantine, depending on the organism and its current distribution.

Fourth, there has to be a committment by multiple agencies and countries to organism eradication. This includes not only funds and time but a willingness to work with people in different countries and cultures to bring the eradication plan to fruition.

Fifth, and probably most critical, the disease must be considered important enough to justify the expenditure of time and money. A worldwide effort to eradicate, say, athelete's foot just ain't gonna cut it--it needs to be a disease that's serious in terms of morbidity and/or mortality.

Smallpox met all of these criteria. Notably, it was a scourge of the ages, so the critical point five was easily met. Though related to a number of poxviruses of animals, smallpox is an exclusively human pathogen with no zoonotic or environmental reservoir. The methods of stopping viral transmission--vaccination, isolation of cases, and quarantine of those who were potentially infected--had worked successfully for many years. Natural infection induced life-long immunity, and the vaccination also produced long-lasting immunity. There was an internationally-supported effort to eradicate the disease, and even war-torn countries negotiated cease-fires in order to allow their people to be vaccinated. Additionally, there weren't multiple serotypes of the virus--the related vaccinia virus, used as a vaccine, was able to provide enough cross-protective immunity to protect the vaccinated individual from smallpox infection.

However, the eradication of smallpox took place before the golden age of molecular biology, prior to the polymerase chain reaction (PCR) and certainly before large-scale genetic sequencing. Therefore, smallpox viruses haven't been extensively examined using these methods, until now. A new Science paper has examined the genetics of 45 isolates of smallpox virus, taken from stocks at the CDC and VECTOR (Russia's State Research Center of Virology and Biotechnology) and isolated from patients during the eradication campaign in the 1960s and 1970s, taking a comparative genomics approach in order to examine the evolution and diversity of the strains. They also included a cowpox, vaccinia, and gerbilpox virus.

They found that, overall, there was a relatively small amount of sequence diversity among the isolates, with many of the differences grouping in one region of the genome that had already been suggested to play a role in virulence. This has implications for bioterrorism; if a virus happened to be released, an examination of this region could identify the viral strain relatively quickly (or, alternatively, rule out that it's one of these strains). Additionally, when they examined the proteins produced (the proteome), they found that even for the most divergent proteins, only a few amino acid differences were found between isolates.

Going along with a topic discussed on this blog recently here and here regarding misuse of scientific information (such as genomic sequences) for nefarious purposes, the authors also examined approximately how many mutations would be necessary in order to take one of the animal poxviruses (camel, monkey, or gerbil) and end up with a variola-like virus. The number was fairly substantial (a few thousand base pairs), but they note:

The results indicate that just a few thousand mutations by one of several rapid methods could convert such orthopoxvirus DNAs into VARV (variola virus) DNA. Recently, infectious VACV (vaccinia virus) was recovered by rescuing its genome from a bacterial artificial chromosome containing its DNA. In theory, one could use sequences to synthesize long oligonucleotides to reconstruct VARV DNA. Such rapidly advancing technology makes it important to understand sequence diversity and the proteome so as to develop and maintain countermeasures against malefic-use-created pathogens.

Looking at the phylogeny of the viruses, they grouped generally into three clusters corresponding to geographic areas (Asia, West Africa, and South America), and that they could see some rough differences between isolates, supporting a previous observation that certain viral strains were generally associated with high, low, or intermediate mortality. They also note that the variola viruses were most closely related to the gerbilpox virus, of the ones they sequenced, and speculate on the zoonotic transmission of this virus into the human population. They also note that a large problem with this is that the gerbilpox virus is also contemporary, and of course we lack any isolates from closer to the time when these virus lineages split apart. Another limitation they touched on but didn't explicity discuss is that they only included a very small number of animal poxviruses. A more complete collection, including not only other poxvirus species but also multiple samples of each, may have yielded different results.

Finally, again, these isolates are all from patients, and convenience isolates at that. Of course, with a pathogen as unique as smallpox the investigators have to take what they can get, but that also places a limitation on their conclusions. Still, a fascinating study adding additonal insight into this ancient--and potentially future--scourge.

Reference

Esposito et al. 2006. Genome Sequence Diversity and Clues to the Evolution of Variola (Smallpox) Virus. Science. 313:807 - 812. Link.

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