Improved assay to detect neutralizing antibody following vaccination with diluted or undiluted vaccinia (Dryvax) vaccine

The assessment of immunogenicity of a diluted vaccinia vaccine for possible widespread use of a diluted vaccine in the event of a bioterrorist attack prompted us to focus on the development of a sensitive and specific plaque reduction neutralization (PRN) assay to assess the antibody response of volunteers to a vaccinia (Dryvax) vaccine. Two incubation times, 1 h or overnight (approximately 15 h), were explored for the neutralization step of the assay. In addition, serum samples were evaluated using both sonicated and nonsonicated virus in PRN assays with 1 and 15 h of incubation. The use of the overnight incubation method resulted in the detection of antibody in two vaccinated individuals who exhibited a take, i.e., a major reaction indicative of successive vaccination as defined by the Centers for Disease Control and Prevention, but did not have a fourfold increase in antibody to vaccinia virus by the 1-h-incubation method and increased the sensitivity from 94 to 100%. In addition to the increased sensitivity of the assay, we noted a significant increase (approximately 40-fold) in the PRN titer of serum samples tested with the 15-h-incubation method. The use of sonicated virus increased the reproducibility of the virus titers and PRN titers. Forty-two percent of the samples tested using sonicated virus had a PRN titer that was fourfold higher or greater than that of nonsonicated virus in the assay. A PRN titer that was threefold higher or greater was observed in more than half (58%) of the samples using sonicated virus. Therefore, the more sensitive, specific, and reproducible plaque neutralization assay for the detection of antibody to vaccinia virus is the method using a 15-h-incubation time and freshly sonicated vaccinia virus.     

Race and sex-based differences in cytokine immune responses to smallpox vaccine in healthy individuals

We assessed the effects of sex, race and ethnicity on smallpox vaccine-induced immune responses in 1071 armed forces members after primary Dryvax® smallpox vaccination, including 790 males and 281 females; 580 Caucasians, 217 African–Americans, and 217 Hispanics. Analysis of vaccinia-specific cytokine responses revealed that Caucasians had higher total IFNγ ELISPOT responses (median 57 spot-forming units/SFUs per 200,000 cells, p = 0.01) and CD8⁺IFNγ ELISPOT responses (12 SFUs, p < 0.001) than African–Americans (51 and 4 SFUs, respectively) and Hispanics (47 and 8 SFUs, respectively). Similarly, Caucasians secreted higher levels of vaccinia-specific IL-2 (p = 0.003) and IFNα (p < 0.001) compared to other racial/ethnic groups. Males had higher total IFNγ ELISPOT responses (median 55 SFUs) compared to females (41 SFUs, p < 0.001). We observed statistically significant sex-related differences in the secretion of IL-2 (p < 0.001), IL-1β (p < 0.001) and IL-10 (p = 0.017). These data suggest that vaccinia-specific cytokine responses following primary smallpox vaccination are significantly influenced by race and sex of vaccinees.     

Cell-mediated immune responses to smallpox vaccination

We report that vaccine dilution (1:1 or 1:10) and previous vaccinia virus vaccination status had no significant effect on cell-mediated immune responses (i.e., the immediate vaccinia virus-specific gamma interferon-producing T-cell response measured by enzyme-linked immunospot assay) 1 month after smallpox vaccination (Lancy-Vaxina; Berna Biotech, Switzerland).     

Smallpox, vaccination and adverse reactions to smallpox vaccine

PURPOSE OF REVIEW: Public fear of the reintroduction of smallpox as a biological weapon or agent of bioterrorism has led to a renaissance of interest in smallpox, and a military and public health vaccination programme in the USA. Clinical experience from the last century together with novel immunobiological findings is the basis for current knowledge on smallpox as a disease. Pre-existing knowledge on smallpox vaccination, plus recent vaccination campaign-derived data, is the basis of current risk-benefit assessments. This article summarizes, from a dermatologist's point of view, current aspects of smallpox, smallpox vaccination and adverse reactions to vaccinia, the live virus smallpox vaccine. RECENT FINDINGS: The smallpox vaccination campaign in the USA has involved over 600,000 vaccinees, and has largely confirmed incidence data on complications. An increased rate of myopericarditis is the new finding in the current vaccination campaign. Immunodeficiencies, manifest atopic dermatitis lesions and a history of atopic dermatitis remain contraindications to vaccination. Plasmacytoid dendritic cells are a key regulator of the human antiviral immune response and are recruited to inflamed skin in many skin diseases, but are depleted in atopic dermatitis lesions. The lack of plasmacytoid dendritic cell recruitment, together with the missing upregulation of antiviral peptides such as cathelicidin LL37 in atopic dermatitis lesions, is considered relevant for an atopic dermatitis patient's susceptibility to eczema vaccinatum. SUMMARY: Recent experience from the US smallpox vaccination campaign has largely confirmed what was known in the 1960s. Current immunobiological research will enhance our understanding of the interaction between poxviruses and the skin's immune system.     

Bacteriophages isolated from smallpox vaccine

A number of lots of dermal and tissue culture smallpox vaccine was studied for the presence of bacteriophage. The presence of bacteriophage was established in lots of tissue culture vaccine but not in the dermal vaccine. Bacteriophage was detected by a modified method for phage isolation. The concentration and activity of the isolated bacteriophage were low. Its activity was increased by passages. The bacteriophage was stable and lysed the main serotypes of enteropathogenic E. coli as well as strains of Shigella sonnei and Salmonella of typhoid A.     

200 years of the smallpox vaccine

The expression "inoculation of smallpox" was first employed by Emanuele Timone, native of Chios island and graduated from the Universities of Padua and Oxford. He learned about this procedure in Constantinople. This method was introduced in North America, during the great epidemic outbreak of 1721, by two Bostonian citizens: Cotton Mather and Zabdiel Boylston. The French physician Henri Etienne Morel introduced the procedure into New Spain during the smallpox epidemic of 1779. Nevertheless only in 1798 the English physician Edward Jenner published the results of his observations and experience concerning the "vaccination" in his book "Inquiry into the cause and effects of the variolae vaccinae." After some initial oppositions, this method rapidly spreaded to the rest of Europe. It arrived to Spain in 1801 and thence was transferred to Spanish America and Philippine Islands with the expedition leaded by Francisco Xavier Balmis. Along the XIX century the methods for obtaining and keeping the vaccine were notably improved. Both Jenner and Balmis are worthy of remembrance as great humanity benefactors.     

Clinical and immunological study of percutaneous revaccination in children who originally received smallpox vaccine subcutaneously.

In a large multicenter smallpox vaccination study carried out from 1970 to 1973, it was found that 39% of children who were initially immunized by the subcutaneous route and then challenged percutaneously with a standard vaccination did not have measurable neutralizing antibody upon follow-up. Because of this finding, a percutaneous revaccination study was conducted at the St. Louis study center in 1975 and 1976. There were four study groups, which were composed on the basis of route of primary immunization (subcutaneous or percutaneous) and whether neutralizing antibody was detectable following the original percutaneous challenge immunization. Of 52 children revaccinated, all but four had accelerated reactions. There was no difference in size of lesions or day of maximum erythema among the four study groups. Only 66% of children originally vaccinated subcutaneously who did not have postchallenge neutralizing antibody had measurable neutralizing antibody following revaccination. Transformation studies with vaccinia viral antigen before and after revaccination were performed on lymphocytes from 50 children. There was no appreciable differences in responses either before or after revaccination when the four groups were compared. However, the mean stimulation ratio for the total group increased from 2.4 before revaccination to 4.6 3 weeks later. In primary subcutaneous vaccine recipients without pre-revaccination neutralizing antibody, lymphocyte transformation correlated directly with the neutralizing antibody response.     

The new cell culture smallpox vaccine should be offered to the general population

A series of major factors must be weighed in deciding whether or not, and to what extent, a particular country should consider pre-exposure vaccination for smallpox. These include the risk of a bioterrorist attack using smallpox, the risk of secondary spread from another country, the risks and benefits of vaccination, the effectivenes s of vaccination pre- and post-exposure, the prevalence of immunocompromised persons, the capacity of the medical care delivery system and the wealth of a nation. We review here the issues and variables relevant for policy making, propose a framework for country-specific decision making and suggest the World Health Organization has a key role to play, particularly with regard to lower-income countries. In doing so, we support the proposition.     

Cannabinoids lead to enhanced virulence of the smallpox vaccine (vaccinia) virus

Indian hemp is used since thousands of years as herbal drug. We found that a single dose of cannabis resin was equally active as Δ9-tetrahydrocannabinol (THC) enhancing severity and duration of symptoms in vaccinia virus infected mice. Cowpox virus did not cause symptomatic disease, but some reduction of specific antibody production was observed in drug treated animals. In vitro cannabis was superior to THC alone at inhibiting mitogen stimulated proliferation of human and mouse spleen cells and peripheral blood mononuclear cells. Also resin sub-fractions other than THC, cannabidiol and cannabinol, recovered also from cigarette smoke, were found inhibitory, suggesting additional involvement of constituents other than psychoactive THC. The immunoregulatory effects must be differentiated from apoptotic effects on spleen cells and lymphocytic mouse cell lines, which were observed with resin and THC but not with cannabidiol or cannabinol. A significant contribution of cytotoxic effects seems unlikely as drug treated lymphocytes were still capable of producing cytokines after T-cell receptor-specific stimulation. Considering a recent case of unusually severe cowpox virus infection in a young drug taker these data confirm a risk of “soft drugs” for acquiring poxvirus infection or enhancing side effects of the smallpox vaccine and perhaps also other live vaccines.