VARIOLA VIRUS (VARV) (from the Latin varius (spotted) or varus (pimple))

Table of contents :

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Epidemiology Genomics Proteomics Transmission Pathogenesis Symptoms & signs Therapy Experimental animal models Web resources

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Epidemiology : smallpox originated in Northeastern Africa around 10000 BC (mummified remains of the 18^th Egyptian dinasty (1580 to 1350 BC) and of the better known Pharaoh Ramses V (1157 BC)). An epidemic occurred in Athens in 430 BC and is described by Thucydides. In 1096-1291 crusaders bring smallpox back to Europe and between 1400 and 1800 fatalities routinely exceed 500,000 / yr. In 1520 smallpox caused collapse of naive Aztec and Incan Empires after arrival of Spanish conquistadors and in 1763 it was first used as a bioweapon against native American Indians.

• last USA case occurred in 1949

In 1966 the WHO undertake a vaccination campaign to eradicate variola virus by using anti-Orthopoxvirus attenuated vaccine

• last India case occurred in 1975 (Saiban Bibi)
• last worldwide natural infection : Ali Maow Maalin, a cook at the hospital in Merca, Somalia, and sometime vaccinator, diagnosed as infectd by variola minor virus on September 11, 1977, and declared recovered on October 26, 1977
• last worldwide artificial infection : on August 1978, Janet Parker, a 40-years old British photographer, became accidentally infected while taking picture in a Birmingham University Medical School lab where some samples were stored and used by the virologist Henry Bedson : the operation of the air handling systems within the facility sucked the air in from the laboratory to the floor above. The virologist committed suicide before she died (she had been vaccinated 12 years before) and she infected (secondary transmission) her father, who died probably not of smallpox but of a heart attack, and her mother, the last person that we know of to have been infected with variola virus, that survived. None of 290 other contacts, all of whom were preventively vaccinated, became ill.

Below is the document signed on December 9, 1979, by members of the WHO Global Commission, certifying that smallpox had been eradicated : the document became public at the World Health Assembly on May 8, 1980.

Eradication was obtained thanks to :

• absence of animal reservoir
• absence of latent or persistent infections in humans (no healthy carriers)
• easily and early recognition of symptoms, so that infected individuals could be quarantined and their contacts vaccinated.
• rare antigenic variation due to high replication fidelity of poxvirus DNA polymerase

Genomics :

• Complete genome (1993)
• PubMed citations

Proteomics :

• smallpox inhibitor of complement enzyme (SPICE) is a co-factor for factor I-mediated degradation of both C3b and C4b.
• IFN-g receptor-like
• G4R/G2R ORF codes for a TNF decoy similar to TNFRII

Environmental resistance :

• historical books written about smallpox virus and the vaccine virus, vaccinia virus, suggests that these viruses could remain viable at room temperature or its equivalent for 2-3 weeks only outside of infected individuals. Variola virus inoculation by scabs stuffed into the nose was first started in the Far East, and by inference, the scab was the result of crusting of acute smallpox pustules. Inoculation of smallpox from an infected person to one who had never been exposed to smallpox was started in the Middle East with fresh "matter" taken directly from true smallpox lesions. There are no reports of inoculations from material older than new scabs or crusts
• in the late 1960s, a study published by the World Health Organization used scabs stored in envelopes to show that the virus stayed viable that way for at least 13 years, likely longer.
• smallpox scabs stored in cotton at room temperature, a relative humidity of 55-75% (the virus does not survive for long periods at customary ambient temperatures in aerosol, inversely proportional to both temperature and humidity), and in indirect sunlight still were viable after 18 months. Thus the virus does not appear to survive, even under optimal conditions, for > 2 years.
• vaccinia virus could be stored in glycerol for long periods of time.
• however, there are no reported experiments or observations that directly focus on how long smallpox material can remain infectious outside of the body; it could, however, only infect humans
• one of the reasons smallpox was eventually eradicated was that there is no human carrier state, meaning that spread was almost always by aerosol during epidemics. Smallpox infection has not been contracted from old mass grave sites of persons who died from smallpox, or potentially infected environmental surfaces known to be exposed to smallpox material that remained for extended periods of time. There was no evidence for the virus lurking in graves, although linen and scabs might survive for a time in vaults above ground : after a cemetery in Quebec which harbored victims of the epidemics of 1650 and 1708 was disturbed in 1854 in order to build sewers, smallpox broke out in its neighborhood, but this was probably just a coincidence. In 1985 it was suggested that smallpox could live as long as a century in the crypts of victims interred in cool, dry climates^{ref1, ref2, ref3, ref4}. In view of the ability of poxviruses to retain infectivity for long periods (possibly decades) in desiccated scabs, it has been suggested that viable smallpox virus or fragments of its intact genomic nucleic acid, might be recoverable from centuries-old cadavers or preserved medical specimens, or alternatively from frozen corpses in the Arctic : so far these predictions have not been fulfilled. Examination of skin lesions from the mummified body of a 16th century Neapolitan child clearly revealed distinctive smallpox-like lesions and poxvirus-like particles by electron microscopy and so it was inferred that this child had died of a severe form of smallpox : however, neither infectious virus, specific antigen, nor recognizable genomic material could be demonstrated^ref. Several years ago in Kentucky a construction crew unearthed a metal coffin containing the mummified corpse of an apparent smallpox victim that researchers traced to the mid-1800s : the CDC checked the tissue for live virus and came up empty
• on 31 March 2003 the 23-year-old librarian Susanne Caro in the College of Santa Fe's Fogelson Library found a small, yellowed envelope tucked between the pages of a  1888 book on Civil War medicine, with the inscription "scabs from vaccination of W.B. Yarrington's children" in the corner, with the signature "Dr. W.D. Kelly," the book's author. He had done work on childhood vaccinations in the late 1800s and during the late 1800s, pus or bits of scabs from smallpox patients with mild cases were implanted in the skins of healthy people to generate a mild illness that bestowed lifetime immunity. It will be interesting to learn in due course whether the contents of the envelope are indeed smallpox scabs, and whether any infectivity has survived.

Transmission : inhalatory (saliva droplets) or percutaneous route (rarely from scabs^ref due to tightly binding in its fibrin matrix); this is an anthroponosis with no known reservoir
Pathogenesis : typically enters the host through the oropharynx, invades the mucosal epithelium, and migrates to regional lymph nodes => asymptomatic viremia => multiplication of virus in the spleen, the bone marrow and other lymph nodes, where viral replication occurs => secondary viremia^ref. After an incubation period of 12–14 days, virus enters the blood within leukocytes, which seed the skin and produce the characteristic skin lesions (pox), whereas most other tissues are spared. The fact that virus seems to travel in leukocytes that specifically exit blood vessels in the papillary dermis indicates that variola virus preferentially associates with leukocytes that can home to skin; alternatively, it might be that only skin tissues can support the subsequent replication steps that are required for lesion formation.
Symptoms & signs : ordinary smallpox / Kaffir pox (a.k.a. vaiolo in Italy; poc or pocca = a bag or pouch; as opposed to great pox or syphilis) => 7-17 days incubation => 2-5 days with high fever, malaise, prostration, headache and backache => infected macrophage in small vessels of the dermis migrate into and infect basal layers of the epidermis => necrosis and edema => maculopapular exanthema on mouth and oropharynx (=> ulcer => saliva => contagion source, expecially in the first week of illness, after onset of rash), face, forearms => trunk and legs, all at the same stage of development => vesicular within 48 hrs => round and tense pustule deeply embedded in the dermis => about 8 days after onset of the rash, the pustules dry up and become crusts by day 14 => by the end of the third week crusts detach leaving pigment-free pockmark (a depressed scar). 33% lethality within 2 weeks due to :

• overwhelming viremia (=> confluent, flat, or malignant smallpox : severe toxemia and delayed appearance of the skin lesions, which were commonly flat and solid; 70-80% lethality)
• type III hypersensitivity (=> MOF)
• encephalitis
• internal (mainly gastrointestinal) hemorrhages in immunocompromised hosts (fulminant or hemorrhagic smallpox from strain Harvey : bleeding from the conjunctiva and mucous membranes, very severe toxemia, and early 100% lethality
• before onset of the rash (early type)
• after the rash appears (late type)

Blindness was a rare complication, usually occurring in cases where there was malnutrition and/or a secondary bacterial infection. Cases of smallpox among pregnant women often resulted in spontaneous abortion of the fetus or a stillborn infant with evidence of lesions on the skin.
Whitepox is the name given to 4 isolates alleged to derive from kidneys of African monkeys and Rodentia in the early 1970s, which are distinguishable by Variola virus only for inducing white pocks in chorioallantoic membrane : no taxonomy allocation as been purposed.
A milder form of disease was seen among those who had partial immunity :

• those who had been vaccinated many years earlier (varioloid / modified smallpox : skin lesions evolve quickly and are more variable in size)
• individuals affected by Variola minor virus (alastrim / Cuban itch)

Depending on whether the rash is present and its density, this variety is divided into 4 types :

• variola sine eruptione : no rash occurs in individuals with good immunity
• confluent smallpox
• discrete smallpox
• semiconfluent smallpox

Variola virus did not persist in the body after recovery. Upon recovery, neutralizing antibodies were long-lasting, whereas HA-inhibiting antibodies declined to low levels within 5 years, and complement-fixing antibodies rarely persisted for longer than 6 months. Little is known about the development of CMI.
Probability of infection for unvaccinated close contacts = 40-90%.
Complications :

• skin overinfection
• acute laryngitis and bronchopneumonia
• heart failure
• encephalomyelitis => coma vigil / "typhoid status" and psychosis
• neuritis
• keratitis
• uveitis
• panophthalmitis
• acute glomerulonephritis
• septic arthritis
• osteomyelitis
• otitis
• arteritis
• phlebitis
• orchitis

Therapy :

• attenuated vaccination within 4 days (as antibody response after primary vaccination usually occurs 4 to 8 days earlier than the response after naturally acquired smallpox infection). In the second half of incubation period IFNs-mediated interference by the already established variola virus prevent the less virulent vaccinia virus from establishing itself.
• IV cidofovir or PO HDP-cidofovir
• methisazone was once used with poor results
• the Poxviridae family members vaccinia and variola virus enter mammalian cells, replicate outside the nucleus and produce virions that travel to the cell surface along microtubules, fuse with the plasma membrane and egress from infected cells toward apposing cells on actin-filled membranous protrusions. Cell-associated enveloped virions (CEV) use Abl- and Src-family tyrosine kinases for actin motility, and that these kinases act in a redundant fashion, perhaps permitting motility in a greater range of cell types. Additionally, release of CEV from the cell requires Abl- but not Src-family tyrosine kinases, and is blocked by STI-571, an Abl-family kinase inhibitor used to treat chronic myelogenous leukemia in humans. STI-571 reduces viral dissemination by 5 orders of magnitude and promotes survival in infected mice, suggesting possible use for this drug in treating smallpox or complications associated with vaccination. This therapeutic approach may prove generally efficacious in treating microbial infections that rely on host tyrosine kinases, and, because the drug targets host but not viral molecules, this strategy is much less likely to engender resistance compared to conventional antimicrobial therapies^ref.

Although the preventive use of smallpox vaccination has been well documented, little is known about its efficacy when used after exposure to the virus. Here we compare the effectiveness of (1) post-exposure smallpox vaccination and (2) antiviral treatment with either cidofovir or with a related acyclic nucleoside phosphonate analogue (HPMPO–DAPy) after lethal intratracheal infection of cynomolgus monkeys (Macaca fascicularis) with monkeypox virus (MPXV). MPXV causes a disease similar to human smallpox and this animal model can be used to measure differences in the protective efficacies of classical and new-generation candidate smallpox vaccines. Initiation of antiviral treatment 24 h after lethal intratracheal MPXV infection, using either of the antiviral agents and applying various systemic treatment regimens, resulted in significantly reduced mortality and reduced numbers of cutaneous monkeypox lesions. In contrast, when monkeys were vaccinated 24 h after MPXV infection, using a standard human dose of a currently recommended smallpox vaccine (Elstree-RIVM), no significant reduction in mortality was observed. When antiviral therapy was terminated 13 days after infection, all surviving animals had virus-specific serum antibodies and antiviral T lymphocytes. These data show that adequate preparedness for a biological threat involving smallpox should include the possibility of treating exposed individuals with antiviral compounds such as cidofovir or other selective anti-poxvirus drugs^ref

At a meeting in Geneva on 4-5 Nov 2004, the Advisory Committee on Variola Virus Research voted to recommend approval for genetically altering the smallpox virus with GFP to speed up the screening of anti-smallpox drugs and help to use up the last remaining stocks of the virus. In practice the proposal will first be seen by the WHO executive board and director general before it is considered by the World Health Assembly in May 2005. the marker would accelerate screening by factors of 10 or 100. It would also reduce the amount of time that lab workers would need to be in BSL-4 labs handling live variola virus. The committee's first proviso for its recommendation is that the research must only be done in the 2 laboratories currently handling live variola, at the CDC in the USA and at Novosibirsk in Russia. The second is that the proposal must also be put to the relevant institutional and national safety committees and submitted to the scientific subcommittee of the Advisory Committee on Variola Virus Research. According to WHO, the committee requested more detailed research proposals and a full safety assessment. There is some concern about the stability of the large virus and a small risk of increasing virulence. The committee also made at least 6 other research recommendations that would allow labs around the world to work with fragments of the variola virus as large as 20% of the whole genome : some consider this a problem because different labs with different chunks "would only have to link them together, and it would be a lot easier" to recreate the entire smallpox genome than it would be to synthesize it. They believe a 10–15% threshold would be preferable because the 20% threshold corresponds to about 37,000 bp, a too large a number. The proposals would also permit the 2 smallpox repository labs—one at the CDC and the other at Novosibirsk in Russia—to insert variola genes one at a time into other viruses in the orthopox family, like monkey pox and cowpox, to create a better model for drug screening, as they can be worked on at lower containment levels than the BSL-4 that smallpox requires, but anyway at one higher BSL than each orthopox virus normally requires. 2 other recommendations would allow the Russian and American teams to share their smallpox samples with one another for the first time and to perform experiments on variola and other orthopox viruses simultaneously, provided the work is performed at BSL-4. Another 2 propose that other researchers elsewhere be permitted to synthesize smallpox fragments up to 500 bp in their labs but prohibited from synthesizing longer ones and, with microarrays, permitted to include the whole variola genome, just as long as individual array fragments are no longer than 80 bp. The 6 recommendations the committee approved last week were all proposed by its technical subcommittee in 2003^ref.
The World Health Organization (WHO) announced on Fri 18 May 2007 that it is postponing for at least 4 years any decision on when to destroy the world's last known stockpiles of smallpox virus, a deadly virus eradicated nearly 30 years go. There is no treatment for the viral disease that killed millions of people a year as recently as the 1960s and left many more blind and scarred. In 1979, it became the 1st disease officially stamped out after a worldwide vaccination campaign. The United States and Russia, however, which hold the only known stockpiles of the virus in high-security laboratories, have long resisted calls to destroy them in case smallpox is found to exist elsewhere. The 60th annual World Health Assembly, the top decision-making body of the United Nations agency, reaffirmed a previous commitment to destroy the remaining stockpiles, but agreed to postpone any decision on when this should happen until its 2011 meeting. In 2010, the WHO secretariat will carry out a review of all research undertaken and still planned, in order that the 64th World Health Assembly may reach global consensus on the timing of the destruction of existing variola smallpox virus stocks. A previous 2002 deadline for destroying smallpox virus was delayed by WHO until new vaccines or treatments for smallpox were found, after the United States said it would keep stocks on hand to combat any re-emergence of the
disease. That decision was made in the wake of the "September 11" suicide plane attacks on the United States, which stirred fears that deadly viruses could fall into the hands of terrorist groups^ref.
Experimental animal models : none except IV inoculation in monkeys.
It is considered by CDC as a category A biological weapon.