Smallpox: Could a vanquished foe return?

Smallpox, one of the most devastating diseases in human history, has ancient roots. Detected in mummified remains dating to the Egyptian dynasties, the disease would eventually claim hundreds of millions of victims in the 20th century alone. As recently as 1967, smallpox was still killing around 15 million people annually.

In 1980, however, the World Health Assembly declared smallpox officially eradicated, thanks to aggressive global vaccination efforts. This triumph is often cited as one of the greatest success stories in modern medicine.

But while naturally-occurring smallpox may be a thing of the past, humanity could nevertheless experience a new outbreak of this dreaded disease. Samples of variola, the virus responsible for causing smallpox, still lurk in laboratory freezers in the U.S. and Russia. Despite elaborate security measures, some threat remains of the virus escaping to wreak havoc.

More ominously, technical advances in synthetic biology could permit variola or related sequences to be duplicated from scratch in a laboratory, raising the possibility of accidental spread or use of the pathogen as a bioweapon.

Monitoring a menace

In new research, PLuS Alliance student Dillon Adam, supervised by Arizona State University Associate Professor Matthew Scotch and Raina MacIntyre of UNSW’s Kirby Institute, investigates methods for rapidly identifying the incidence and global migration routes of variola, using techniques known as phylogenetics and phylogeography.

The methods described offer powerful tools that could be used in the field to pinpoint and arrest a future outbreak of smallpox at the earliest phase of an epidemic.

Further, the new study uses phylogenetics and phylogeography to chart historic migration patterns of smallpox.

Matthew Scotch, assistant director of the Biodesign Center for Environmental Health Engineering.

Scotch is the assistant director of ASU's Biodesign Center for Environmental Health Engineering. He and MacIntyre co-authored the new paper, led by Adam. Adam and MacIntyre are researchers at UNSW’s Kirby Institute, where most of the work was carried out.

The group’s research findings recently appeared in the Oxford journal Molecular Biology and Evolution.

“Phylogeography is a discipline that can inform public health surveillance,” Scotch said. “We show that in this work by focusing on select smallpox genes. This virus still represents a serious concern among epidemiologists and hopefully, this work will help us in a response, if ever needed.”

Preparing for the worst

A follow-up simulation conducted by the researchers underscored the urgency of developing reliable, rapid response strategies for dealing with a future bioterror attack using smallpox or a related pathogen.

Researchers joined forces with stakeholders in health, foreign affairs and trade, defense, police and key nongovernmental agencies to conduct the bioterror exercise. Their aim was to model the effects of a smallpox attack that begins in Fiji, followed shortly by a much larger event occurring in a bigger country in Asia.

The exercise was conducted by The NHMRC Centre for Research Excellence, Integrated Systems for Epidemic Response (ISER), led by UNSW, which conducts applied systems research on epidemic control.

The results are alarming. The simulation suggests that an epidemic of smallpox could — within weeks — spread like a wildfire across the globe. Stores of vaccine would quickly be depleted. The patient load would overwhelm hospitals and health care systems, and severe impacts on travel and trade would occur. Further, as health care workers, military and police were affected by the disease, they would be left unable to cope with increasingly dire conditions. Organized society would begin to unravel.  

A variety of factors conspire to make this scenario particularly destructive. These include a world population that has either never been vaccinated or has waning immunity to smallpox; delayed diagnosis due to unfamiliarity with symptoms of smallpox by most of today’s physicians and rapid transit between all points on the globe, permitting brisk dissemination of the virus.

The simulation suggests that a lightning-fast response by health care officials would be critical for avoiding catastrophe. Aggressive, coordinated action against a smallpox outbreak will have to occur within one week of the first infectious case or the epidemic will spiral out of control.

Time is the enemy

Until now, however, positive laboratory diagnosis for variola has been cumbersome, time-consuming and costly. In the new study, researchers use just three genes from the virus, instead of the entire genome, to identify it. Studying sequence alterations in different isolates of variola can help establish where an epidemic began, track its rate of spread and establish its routes of migration. Sequencing minimal genetic material to identify a future re-emergence of smallpox could radically accelerate the mobilization of health care resources, improving outcomes.

The study results show that examination of 3 critical diagnostic genes, knows as hemagglutinin (HA), cytokine response modifier B (CrmB) and A-type inclusion protein (ATI) can not only identify variola, but distinguish between the two leading strains of the virus: Variola Minor and the significantly more lethal Variola Major.

Once the diagnostic viral genes have been sequenced, researchers apply sophisticated statistical methods, known as Bayesian analysis, in order to evaluate the most likely disease epicenter, rates of viral spread and geographical distribution over time. Health care workers can apply this information to formulate the best approach to containing and eradicating the disease, before an epidemic rages out of control.

Phylogenetics uses the appearance of a pathogen (or its morphology), combined with molecular data — typically DNA sequence. Through careful analysis of these factors, phylogenetic researchers can build family histories of organisms that reveal their degree of relatedness.

The technique depends in part on the fact that pathogens develop mutations at different rates over time. RNA viruses mutate quickly, DNA viruses (including smallpox), less rapidly and bacteria, comparatively slowly. Molecular clocks of mutation rate have been established for most known pathogens and are used to build and calibrate phylogenetic trees.

The integration of evolutionary frameworks with epidemiological data from a given outbreak can be used to produce phylogeographic models tracking disease source, speed of diffusion and extent of spread. The combined techniques of phylogenetics and phylogeography offer potent tools for health agencies confronted with a contagious outbreak.

Phylogeography has already been used to investigate viral dispersal for diseases including dengue fever, rabies, influenza and HIV. It is also poised to significantly improve viral mapping of origin and spread for avian influenza, though it has yet to be applied in the case of an active disease epidemic. The increasing availability of rapid, low-cost sequencing technologies will lead to more widespread application of phylogenetics and phylogeography for infectious disease monitoring.

Profile of a killer

In addition to providing techniques to pinpoint a future outbreak of smallpox, the study uses the most complete dataset currently available of whole genome and partially sequenced variola isolates to show how waves of destruction in past outbreaks of the disease spread across the continents.

Using reduced genetic datasets of variola sequence information, sampled from 1654 to 1977, and Bayesian phylogeogeaphy, the group was able to map, for the first time, 10 statistically likely routes of historic smallpox transmission between 8 discrete regions: Latin America, Europe, Western Africa, Eastern Africa and the Middle East, Southern Africa, Southern Asia, Asia Pacific and East Asia.

The methods outlined are the first to empirically describe global transmission of historic variola isolates through the use of phylogeography, revealing two discrete lineages of Variola Major circulating between East Asia and Europe following WWII.

Renewed threat

Could smallpox really rise from the dead to cause a new and terrifying pandemic? In addition to 571 known samples of variola hibernating in laboratories, an additional threat exists. Advances in the manipulation of the molecules of life through techniques of synthetic biology have enabled the construction of smallpox or a similarly deadly pathogen.

In 2017, Canadian researchers synthesized another form of poxvirus extinct in nature, establishing the frightening potential of fast-paced genetic innovations.

According to MacIntyre, who leads the Biosecurity Program at the Kirby Institute: “Biowarfare has been used since recorded history. Certain terror groups have openly called for biological attacks in July this year. The capability for attacks is increased by publicly available methods for making synthetic viruses, so we have both intent and capability. This means such threats must be taken seriously.”

In addition to heightened vigilance, techniques like those described in the current research offer the best hope of averting a large-scale smallpox catastrophe in the future and fulfilling the World Health Organization’s mandate for improved smallpox preparedness.

MacIntyre is also an adjunct professor at ASU and a fellow (along with Scotch) in the PLuS Alliance, a collaborative effort by ASU, King’s College London and UNSW Sydney aimed at addressing global challenges in health, social justice, sustainability, technology and innovation.