Smallpox must be distinguished from other vesicular exanthems, such as chickenpox, erythema multiforme with bullae, or allergic contact dermatitis. In a confirmed outbreak, smallpox would likely be a clinical diagnosis. Particularly problematic to the necessary infection control measures would be the failure to recognize relatively mild cases of smallpox in persons with partial immunity, or extremely severe cases in patients without classical disease. Therefore, isolation of suspected cases, quarantine of potential exposures, and initiation of medical countermeasures should be promptly followed by an accurate laboratory diagnosis. Providers who collect or process specimens should be vaccinated and should exercise contact and airborne precautions. Specimens should be collected only under the direction of public health officials. Typical variola specimens might include scrapings of skin lesions, lesion fluid, crusts, blood, or pharyngeal swabs.

A method of presumptive diagnosis is demonstration of characteristic poxvirus virions on electron microscopy of vesicular scrapings. Under light microscopy, aggregations of variola virus particles, called Guarnieri bodies, can be found. Another rapid but relatively insensitive test for Guarnieri bodies in vesicular scrapings is Gispen's modified silver stain, in which cytoplasmic inclusions appear black. However, none of the above laboratory tests is capable of discriminating variola from vaccinia, monkeypox, or cowpox.

Definitive diagnosis of variola has classically required isolation of the virus and characterization of its growth on chicken egg chorioallantoic membrane or in cell culture. Culture of variola is presently available only at Laboratory Response Network (LRN) national laboratories (CDC and USAMRIID) under BSL-4 conditions. Several nucleic acid techniques have been developed for specific poxvirus identification, with PCR becoming more widely available.

Neutralizing antibodies to variola form in the first week of illness and may be present for many years. Hemaglutination-inhibition antibodies are detectible by the 16th day of infection and complement fixation antibodies by the 18th, but both begin to decrease after 1 year.

Associated laboratory findings, including the complete blood counts (CBC) of patients with ordinary smallpox, often exhibited a neutropenia and lymphocytosis during the eruptive stage. Neutrophils could become elevated during the late pustular stage when secondary bacterial infections would occur. Mild thrombocytopenia was common. In hemorrhagic smallpox, thrombocytopenia was progressive and severe as DIC (disseminated intravascular coagulation) developed.

Differential Diagnosis

Other rash illnesses, outlined in the table below, are included in the differential diagnosis of smallpox.

Differential Diagnosis for Smallpox



Distinguishing features

Ordinary and Flat-Type Smallpoxa



See Distinguishing Features Between Smallpox, Monkeypox, and Chickenpox below

Human monkeypox

Monkeypox virus

See Monkeypox below

Disseminated herpes zoster


—Usually occurs in immunocompromised hosts
—History of chickenpox


Staphylococcus aureus
Streptococcus pyogenes

—Lesions often pruritic and not painful
—Lesions focal and not usually disseminated
—Lesions not "shotty"
—Gold-colored crusted plaques are classic
—Lesions superficial and not embedded into dermis
—Constitutional symptoms generally absent or minimal
—Usually occurs in children

Hand, foot, and mouth disease


—Usually occurs in children <10 yr of age
—Has autumn seasonal pattern
—Lesions may be confined to hands and feet (although dissemination may occur)

Disseminated herpes simplex

Herpes simplex virus

—Usually occurs in immunocompromised hosts
—Lesions are vesicular and do not progress to pustules

Secondary syphilis

Treponema pallidum

—Rash generally does not include vesicular phase
—Lesions not "shotty"
—Constitutional symptoms relatively mild
—Lesions generally evolve slowly from macules to papules to pustules (often over several weeks)

Molluscum contagiosum


—Usually occurs in healthy children or HIV-positive adults
—In healthy adults, lesions generally occur in genital area
—Lesions are painless
—Constitutional symptoms generally are absent
—Lesions may persist for several months (or longer in immunocompromised patients)

Erythema multiforme major (including Stevens-Johnson syndrome)

Associated with various infectious and noninfectious processes

—Constitutional symptoms and rash usually appear at same time
—Rash evolves rapidly
—Bullae or "bull's-eye" lesions common
—Extensive mucous membrane involvement, including conjunctivitis, common

Drug eruptions


—Lesions generally not pustular
—History of drug exposure
—Fever may be present, but severe toxemia usually absent

Bullous pemphigoid


—Tense bullae characteristic
—Occurs most commonly in elderly
—Intense pruritus may be present
—Constitutional symptoms usually absent
—Peripheral eosinophilia may be noted

Other skin conditions


—Insect bites
—Contact dermatitis

Hemorrhagic Smallpox


Neisseria meningitidis

Rapid progression to shock and often death

Hemorrhagic varicella


Usually occurs in immunocompromised children

Rocky Mountain spotted fever

Rickettsia rickettsii

—Tick exposure history may be obtained
—Occurs April through May
—Most US cases occur in southeastern and south-central states


Ehrlichia chaffeensis
Erhlichia phagocytophilia

—Tick exposure history may be obtained
—Petechial rash uncommon
—Peripheral blood smear may show morulae in neutrophils of patients with human granulocytic ehrlichiosis

Septicemia caused by gram-negative bacteria

Various bacterial agents

—Underlying illness usually present

Abbreviation: VZV, varicella-zoster virus.

aOther rash illnesses (eg, measles, rubella, scabies, scarlet fever) also may be considered in the differential diagnosis, although the rashes caused by these conditions generally are not characteristic of smallpox.

Adapted from CDC 2007: Acute, generalized vesicular or pustular rash illness testing protocol in the United States, Fenner 1988, Moore 2006.


Historical perspective in Africa

Human monkeypox is caused by monkeypox virus, which, like variola virus, is in the Orthopoxvirus genus. Monkeypox is similar to smallpox, but illness is generally milder. Recognized cases have occurred predominantly in western and central Africa. Pertinent historic information about monkeypox in Africa is as follows:

  • The illness in humans is similar to discrete or semiconfluent ordinary smallpox (Jezek 1987). A prodrome (fever, headache, backache) lasting 1 to 3 days occurs, followed by eruption of a smallpox-like rash that lasts 2 to 4 weeks.
  • Monkeypox cases tend to have prominent lymphadenopathy, which generally is not a feature of either chickenpox or smallpox (Arita 1985, Breman 1980, Jezek 1987). This can be an important distinguishing characteristic between the three conditions.
  • The first human case was recognized in 1970; since then, sporadic cases and outbreaks have been recognized in Africa, although the illness appears to be relatively uncommon.
  • Natural animal reservoirs in Africa include several squirrel species and forest-dwelling primates (Khodakevich 1988). Lagomorphs (rabbits, hares, etc) and other rodents in addition to squirrels, including prairie dogs, also appear to be susceptible to infection. The ecological requirements and geographic distributions have been identified, and these may support further field studies and guide public health intervention strategies (Levine 2007).
  • The case-fatality rate was 11% in one series of 282 patients (Jezek 1987) and was 3% in one outbreak involving 71 cases (CDC 1997: Human monkeypox—Kasai Oriental, Zaire, 1996-1997), suggesting that the illness is less severe than smallpox. In both investigations, all deaths occurred in children younger than 10 years old (who had not received earlier smallpox vaccination).
  • Person-to-person transmission has been demonstrated (Arita 1985, Breman 1980, CDC 1997: Human monkeypox—Kasai Oriental, Zaire, 1996-1997, Jezek 1986, Jezek 1988). Secondary attack rates of 7.2%, 7.5%, and 15% have been reported among household contacts who had not received prior smallpox vaccination (Arita 1985, Jezek 1986, Jezek 1988). These secondary attack rates are lower than those observed for smallpox and reflect the lower propensity for spread of monkeypox compared with smallpox.

US 2003 outbreak

In June 2003, an outbreak of monkeypox was recognized in the midwestern United States (CDC 2003: Multistate outbreak of monkeypox—Illinois, Indiana, Kansas, Missouri, Ohio, and Wisconsin, Reed 2004). Key findings from the outbreak are as follows:

  • Seventy-one cases were reported to the CDC; 18 (26%) patients were hospitalized but no fatalities occurred.
  • No exposed healthcare workers developed monkeypox symptoms, although one worker had serologic evidence of recent orthopoxvirus infection; that person had received smallpox vaccine during the previous year (Fleischauer 2005).
  • Thirty exposed persons received smallpox vaccine to prevent monkeypox; three reported rash within 2 weeks after vaccination, and one of these persons was confirmed as having monkeypox.
  • In one report, three exposed persons were identified who had no symptoms but had serologic evidence of recent monkeypox infection. All three had received smallpox vaccine in the past (13, 29, and 48 years earlier). These findings suggest long-term persistence of cross-protective immunity to orthopoxvirus infection following smallpox vaccination in these individuals (Hammarlund 2005).
  • The outbreak was traced to contact with infected prairie dogs; the prairie dogs became infected through contact with six species of African rodents (including Gambian giant rats, rope squirrels, tree squirrels, brushtail porcupines, striped mice, and dormice).
  • In November 2003, the Food and Drug Administration (FDA) and the CDC issued an interim final rule to prevent further introduction and transmission of the monkeypox virus (HHS 2003). This rule included one set of regulations administered by the CDC that prohibited importation of African rodents. A second set of regulations, administered by the FDA, placed restrictions on the interstate movement of several species of African rodents and on prairie dogs. In September 2008, the FDA removed its restrictions because they were deemed no longer necessary; the CDC ban on importation of African rodents remains in effect (FDA 2008).
  • The CDC has published guidance for autopsy and safe handling of human remains of monkeypox patients (CDC 2003: Interim guidance for autopsy and safe handling of human remains of monkeypox patients).

Distinguishing Features Between Smallpox, Monkeypox, and Chickenpox

Smallpox and monkeypox are generally quite similar, although smallpox often is more severe and the case-fatality rate is higher. Early in the clinical course, smallpox or monkeypox may be mistaken for chickenpox if the clinical suspicion for orthopoxvirus infection is low. Also, smallpox in partially immune patients may be mild and may resemble chickenpox. An assessment of suspected smallpox cases referred to the CDC between 2002 and 2004 found that chickenpox accounted for more than half of the cases (Seward 2004). Distinguishing features of the three illnesses are outlined in the table below.

Distinguishing Features Between Smallpox, Chickenpox, and Monkeypox


Smallpox (Variola Major)a




Lasts 2-4 days, with high fever, headache, backache, and severe prostration; vomiting and severe abdominal pain may occur

Prodrome often absent; if present, it is mild and brief (ie, about 1 day)

Lasts about 2 days and is similar to that seen with smallpox

Distribution of rash

Begins on oral mucosa, spreads to face, then expands in centrifugal pattern (ie, most dense on face and distal extremities)

Begins on trunk and expands in centripetal pattern (ie, most dense on trunk)

Often begins on face and spreads in centrifugal pattern (although cases have been reported with centripetal pattern of spread)

Lesions on palms and soles


Almost never occur





Common (up to 90%)

Timing for occurrence of lesions

Generally emerge over 1-2 days and then progress at same rate

Occur in "crops" and may be at different stages of maturation at any given point in time

Lesions usually progress at same rate but may occur in crops (in about 20% of patients)

Evolution of lesions

Progress over several days from macules (day 1), to papules (day 2), to vesicles (days 3-5), to pustules (days 7 to about 14), to scabs (day 14 to about 20)

Progress quickly over about 24 hr from macules to papules to vesicles, then to crusted lesions

Progress in pattern similar to smallpox

Sensation associated with lesions

May be painful and only become pruritic during scabbing stage

Often intensely pruritic; not usually painful unless superimposed bacterial infection occurs

May be painful (although often milder than smallpox)

Depth of lesions

Extend deep into dermis and often cause pitted scarring

Superficial and generally do not cause scarring

Generally superficial (although pitted scarring can occur)

Duration of illness

14-21 days

4-7 days

14-21 days


Patients often appear toxic and case-fatality rate may be as high as 50%

Patients often do not appear severely ill and illness is rarely fatal

Illness can vary in severity but often is mild and self-limited


Cases can be expected to occur in all age-groups; illness may be somewhat milder in adults over age 30 who were vaccinated as young children

Most cases occur in children; adults likely to be immune

Cases can be expected to occur in all age-groups; illness may be milder in people who have received smallpox vaccination

aIllness may be milder in patients with partial immunity; fever may be less common and fewer lesions may occur with more rapid healing.
bAlthough the prodrome may be milder in patients with chickenpox, a review of 932 chickenpox cases demonstrated that 7% to 17% of unvaccinated patients with chickenpox may meet the smallpox febrile prodrome criteria as put forth by the CDC (Moore 2004).

Adapted from CDC 2007: Acute, generalized vesicular or pustular rash illness testing protocol in the United States, Di Giulio 2004.

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