Approach Considerations
Suitably vaccinated and trained personnel should obtain viral swabs of the patient's pharynx and/or open skin lesions (eg, pustule contents, material from the base of the scab).
Under biosafety level 4 (BSL4) laboratory conditions, these samples can be examined for the presence of virions by using electron microscopy, PCR assay, or immunohistochemical analysis or by growing the virus on live cell cultures. [13, 14]
Serologic testing can be performed to detect neutralizing antibodies, but the results cannot be used to differentiate Orthopoxvirus species.
Although smallpox and all other viruses in the Orthopoxvirus genus exhibit identically appearing brick-shaped virions, the clinical aspects of these diseases generally suffice for distinguishing cowpox and vaccinia from smallpox.
Monkeypox virions may also be indistinguishable from smallpox virions, but naturally occurring monkeypox is typically limited to tropical rain forest areas of Africa.
Smallpox infection may be confirmed based on the presence of brick-shaped virions viewed with electron microscopy examination of vesicular or pustular fluid or scabs. PCR assay and electron microscopy can be used to examine inactivated samples and therefore do not require such high levels of isolation and can be performed in local laboratories.
However, although electron microscopy can help to identify the virus as a member of the Orthopoxvirus genus, it cannot help to determine the exact species.
PCR assay can be used to identify the species and can even distinguish minor genetic variations in the different strains. PCR assay has been used to identify variola only twice previously, and never in a clinical situation. PCR assay can amplify small and specific lengths of DNA and can accurately differentiate variola virus DNA from other species in the genus. The sensitivity is 5-10 copies of DNA. PCR assay can be useful in distinguishing between chickenpox and smallpox.
Cell culture is seldom used, because it is not as effective as the other methods and because it requires the use of live virus, which, in turn, requires the use of a BSL-4 laboratory.
Depending on the presenting clinical symptoms, other diseases, such as meningococcemia, leukemia, herpes viruses, and drug eruptions, must be ruled out. A meticulous drug history should be obtained. Tests likely to be performed include the following:
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Tzanck preparations
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Direct fluorescent antibody testing for herpes viruses
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Blood tests
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Skin biopsy
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Lumbar puncture
Specimen Collection and Handling
A smallpox skin specimen should be collected with precautions in place. Gloves should be worn during collection; fluid from lesions can be harvested on a cotton swab. Prior to shipping specimens, state and local health department laboratories should be contacted for specific instructions.
The CDC recommends the following procedures for handling specimens obtained from a patient thought to be infected with the smallpox virus:
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Specimens should be collected by someone who has recently been vaccinated (or who has been vaccinated that day) and who is wearing gloves and a mask
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To obtain vesicular or pustular fluid, the lesions may need to be opened with the blunt edge of a scalpel; the fluid can then be harvested on a cotton swab; scabs can be picked off with forceps
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Specimens should be deposited in a Vacutainer tube; the tube should be sealed with adhesive tape at the juncture of the stopper and the tube, and this tube, in turn, should be enclosed in a second durable and watertight container
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State or local health department laboratories should be contacted immediately for proper specimen shipping protocols.
Laboratory examination should be performed only in designated BSL-4 laboratories. Once it has been established that an epidemic is being caused by the smallpox virus, clinically similar cases do not require further laboratory testing.
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Smallpox virion. Courtesy of US Centers for Disease Control and Prevention.
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After exposure to the smallpox virus, a symptom-free incubation period follows. It normally lasts 10-12 days but may vary from 7-17 days. Smallpox begins with fever, headache, and severe backache. A rash appears after 2-4 days and progresses through characteristic stages of papules, vesicles, pustules, and, finally, scabs. The scabs desquamate at the end of the third or fourth week. Courtesy of the World Health Organization.
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Smallpox rash at days 3, 5, and 7 of evolution. Lesions are denser on the face and extremities than on the trunk. They also appear on the palms of the hand and have a similar appearance. Courtesy of the World Health Organization.
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Flat-type smallpox on day 6 of the rash. Courtesy of the US Centers for Disease Control and Prevention.
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This patient with smallpox survived toxemia to succumb to secondary tissue damage days after this photo was taken. Courtesy of the US Centers for Disease Control and Prevention.
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Smallpox vaccination with bifurcated needle. Reconstituted vaccine is held between the prongs of the needle and injected subcutaneously by multiple punctures; 15 rapid strokes, at right angles to the skin over the deltoid muscle, are made within a 5-mm area. Courtesy of the World Health Organization.
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Smallpox vaccination. Evolving primary vaccination appearance. Courtesy of the US Centers for Disease Control and Prevention.
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Typical temperature chart of a patient with smallpox infection (from Henderson, 1999).
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Characteristic skin lesion of variola viral infection on the arms and the legs of an adolescent. Photo used with the permission of the World Health Organization (WHO).
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Small child with pustular lesions due to variola viral infection. Photo used with the permission of the World Health Organization (WHO).
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Infant with advanced lesions due to variola viral infection. Photo used with the permission of the World Health Organization (WHO).
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Unvaccinated infant with the ordinary form of the variola major strain of smallpox has centrifugally distributed umbilicated pustules on day 3 in the course of the disease. Reprinted with permission from the World Health Organization (WHO).
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Unvaccinated infant with the ordinary form of the variola major strain of smallpox has centrifugally distributed umbilicated pustules on day 5 in the course of the disease. Reprinted with permission from the World Health Organization (WHO).
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Unvaccinated infant with the ordinary form of the variola major strain of smallpox has centrifugally distributed umbilicated pustules on day 7 in the course of the disease. Reprinted with permission from the World Health Organization (WHO).
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The ordinary form of the variola minor strain of smallpox (alastrim) in an unvaccinated woman 12 days after the onset of skin lesions. The facial lesions are sparser and evolved more rapidly than the extremity lesions. Reprinted with permission from the World Health Organization (WHO).
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The ordinary form of the variola minor strain of smallpox (alastrim) in an unvaccinated woman 12 days after the onset of skin lesions. The facial lesions are sparser and evolved more rapidly than the extremity lesions. Reprinted with permission from the World Health Organization (WHO).
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The ordinary form of the variola minor strain of smallpox (alastrim) in an unvaccinated woman 12 days after the onset of skin lesions. The facial lesions are sparser and evolved more rapidly than the extremity lesions. Reprinted with permission from the World Health Organization (WHO).
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Adult with variola major with hundreds of pustular lesions centrifugally distributed. Photo from Fitzsimmons Army Medical Center slide file.
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Hemorrhagic-type variola major lesions. Death usually ensued before typical pustules developed. Reprinted with permission from the World Health Organization (WHO). 1988; 10-14, 35-36.
