Influenza vaccination reveals sex dimorphic imprints of prior mild COVID-19.

Acute viral infections can have durable functional impacts on the immune system long after recovery, but how they affect homeostatic immune states and responses to future perturbations remain poorly understood1-4. Here we use systems immunology approaches, including longitudinal multimodal single-cell analysis (surface proteins, transcriptome and V(D)J sequences) to comparatively assess baseline immune statuses and responses to influenza vaccination in 33 healthy individuals after recovery from mild, non-hospitalized COVID-19 (mean, 151 days after diagnosis) and 40 age- and sex-matched control individuals who had never had COVID-19. At the baseline and independent of time after COVID-19, recoverees had elevated T cell activation signatures and lower expression of innate immune genes including Toll-like receptors in monocytes. Male individuals who had recovered from COVID-19 had coordinately higher innate, influenza-specific plasmablast, and antibody responses after vaccination compared with healthy male individuals and female individuals who had recovered from COVID-19, in part because male recoverees had monocytes with higher IL-15 responses early after vaccination coupled with elevated prevaccination frequencies of 'virtual memory'-like CD8+ T cells poised to produce more IFNγ after IL-15 stimulation. Moreover, the expression of the repressed innate immune genes in monocytes increased by day 1 to day 28 after vaccination in recoverees, therefore moving towards the prevaccination baseline of the healthy control individuals. By contrast, these genes decreased on day 1 and returned to the baseline by day 28 in the control individuals. Our study reveals sex-dimorphic effects of previous mild COVID-19 and suggests that viral infections in humans can establish new immunological set-points that affect future immune responses in an antigen-agnostic manner.

Preconfiguration of the antigen-binding site during affinity maturation of a broadly neutralizing influenza virus antibody.

Affinity maturation refines a naive B-cell response by selecting mutations in antibody variable domains that enhance antigen binding. We describe a B-cell lineage expressing broadly neutralizing influenza virus antibodies derived from a subject immunized with the 2007 trivalent vaccine. The lineage comprises three mature antibodies, the unmutated common ancestor, and a common intermediate. Their heavy-chain complementarity determining region inserts into the conserved receptor-binding pocket of influenza HA. We show by analysis of structures, binding kinetics and long time-scale molecular dynamics simulations that antibody evolution in this lineage has rigidified the initially flexible heavy-chain complementarity determining region by two nearly independent pathways and that this preconfiguration accounts for most of the affinity gain. The results advance our understanding of strategies for developing more broadly effective influenza vaccines.

A live attenuated influenza A(H5N1) vaccine induces long-term immunity in the absence of a primary antibody response.

BACKGROUND: Highly pathogenic avian influenza A(H5N1) causes severe infections in humans. We generated 2 influenza A(H5N1) live attenuated influenza vaccines for pandemic use (pLAIVs), but they failed to elicit a primary immune response. Our objective was to determine whether the vaccines primed or established long-lasting immunity that could be detected by administration of inactivated subvirion influenza A(H5N1) vaccine (ISIV). METHODS: The following groups were invited to participate in the study: persons who previously received influenza A(H5N1) pLAIV; persons who previously received an irrelevant influenza A(H7N3) pLAIV; and community members who were naive to influenza A(H5N1) and LAIV. LAIV-experienced subjects received a single 45-µg dose of influenza A(H5N1) ISIV. Influenza A(H5N1)- and LAIV-naive subjects received either 1 or 2 doses of ISIV. RESULTS: In subjects who had previously received antigenically matched influenza A(H5N1) pLAIV followed by 1 dose of ISIV compared with those who were naive to influenza A(H5N1) and LAIV and received 2 doses of ISIV, we observed an increased frequency of antibody response (82% vs 50%, by the hemagglutination inhibition assay) and a significantly higher antibody titer (112 vs 76; P = .04). The affinity of antibody and breadth of cross-clade neutralization was also enhanced in influenza A(H5N1) pLAIV-primed subjects. CONCLUSIONS: ISIV administration unmasked long-lasting immunity in influenza A(H5N1) pLAIV recipients, with a rapid, high-titer, high-quality antibody response that was broadly cross-reactive across several influenza A(H5N1) clades. CLINICAL TRIALS REGISTRATION: NCT01109329.

Effects of postchallenge administration of ST-246 on dissemination of IHD-J-Luc vaccinia virus in normal mice and in immune-deficient mice reconstituted with T cells.

Whole-body bioimaging was used to study dissemination of vaccinia virus (VACV) in normal and in immune deficient (nu(-)/nu(-)) mice protected from lethality by postchallenge administration of ST-246. Total fluxes were recorded in the liver, spleen, lungs, and nasal cavities of live mice after intranasal infection with a recombinant IHD-J-Luc VACV expressing luciferase. Areas under the flux curve were calculated for individual mice to assess viral loads. Treatment for 2 to 5 days of normal BALB/c mice with ST-246 at 100 mg/kg starting 24 h postchallenge conferred 100% protection and reduced viral loads in four organs compared to control mice. Mice also survived after 5 days of treatment with ST-246 at 30 mg/kg, and yet the viral loads and poxes were higher in these mice compared to 100-mg/kg treatment group. Nude mice were not protected by ST-246 alone or by 10 million adoptively transferred T cells. In contrast, nude mice that received T cells and 7-day treatment with ST-246 survived infection and exhibited reduced viral loads compared to nonreconstituted and ST-246-treated mice after ST-246 was stopped. Similar protection of nude mice was achieved using adoptively transferred 1.0 and 0.1 million, but not 0.01 million, purified T cells or CD4(+) or CD8(+) T cells in conjunction with ST-246 treatment. These data suggest that ST-246 protects immunocompetent mice from lethality and reduces viral dissemination in internal organs and poxvirus lesions. Furthermore, immune-deficient animals with partial T cell reconstitution can control virus replication after a course of ST-246 and survive lethal vaccinia virus challenge.

Humans with chronic granulomatous disease maintain humoral immunologic memory despite low frequencies of circulating memory B cells.

CD27(+) memory B cells are reduced in the blood of patients with chronic granulomatous disease (CGD) for reasons and consequences that remain unclear. Here we confirm not only decreased CD27(+) but also IgG(+) B cells in the blood of CGD patients compared with healthy donors (HDs). However, among IgG(+) B cells, the ratio of CD27(-) to CD27(+) was significantly higher in CGD patients compared with HDs. Similar to conventional memory B cells, CD27(-)IgG(+) B cells of CGD patients expressed activation markers and had undergone somatic hypermutation, albeit at levels lower than their CD27(+) counterparts. Functional analyses revealed slight reductions in frequencies of total IgG but not influenza-specific memory B-cell responses, as measured by Elispot in CGD patients compared with HDs. Serum IgG levels and influenza-specific antibodies were also normal in these CGD patients. Finally, we provide evidence that influenza-specific memory B cells can be present within the CD27(-)IgG(+) B-cell compartment. Together, these findings show that, despite reduced circulating CD27(+) memory B cells, CGD patients maintain an intact humoral immunologic memory, with potential contribution from CD27(-) B cells.

Broadly neutralizing human antibody that recognizes the receptor-binding pocket of influenza virus hemagglutinin.

Seasonal antigenic drift of circulating influenza virus leads to a requirement for frequent changes in vaccine composition, because exposure or vaccination elicits human antibodies with limited cross-neutralization of drifted strains. We describe a human monoclonal antibody, CH65, obtained by isolating rearranged heavy- and light-chain genes from sorted single plasma cells, coming from a subject immunized with the 2007 trivalent influenza vaccine. The crystal structure of a complex of the hemagglutinin (HA) from H1N1 strain A/Solomon Islands/3/2006 with the Fab of CH65 shows that the tip of the CH65 heavy-chain complementarity determining region 3 (CDR3) inserts into the receptor binding pocket on HA1, mimicking in many respects the interaction of the physiological receptor, sialic acid. CH65 neutralizes infectivity of 30 out of 36 H1N1 strains tested. The resistant strains have a single-residue insertion near the rim of the sialic-acid pocket. We conclude that broad neutralization of influenza virus can be achieved by antibodies with contacts that mimic those of the receptor.

DNA priming prior to inactivated influenza A(H5N1) vaccination expands the antibody epitope repertoire and increases affinity maturation in a boost-interval-dependent manner in adults.

DNA priming improves the response to inactivated influenza A(H5N1) vaccination. We compared the immunogenicity of an H5 DNA prime (using strain A/Indonesia/5/2005) followed by an H5N1 monovalent inactivated vaccine boost at 4, 8, 12, 16, or 24 weeks to that of 2 doses of H5N1 monovalent inactivated vaccine in adults. Antibody epitope repertoires were elucidated by genome-fragment phage-display library analysis, and antibody avidities for HA1 and HA2 domains were measured by surface plasmon resonance. H5 DNA priming expanded the H5-specific antibody epitope repertoire and enhanced antibody avidity to the HA1 (but not the HA2) domain in an interval-dependent manner. Enhanced HA1 binding and avidity after an interval of ≥12 weeks between prime and boost correlated with improved neutralization of homologous and heterologous H5N1 strains. Clinical trials registration NCT01086657.

Bacterial HA1 vaccine against pandemic H5N1 influenza virus: evidence of oligomerization, hemagglutination, and cross-protective immunity in ferrets.

The impending influenza virus pandemic requires global vaccination to prevent large-scale mortality and morbidity, but traditional influenza virus vaccine production is too slow for rapid responses. We have developed bacterial systems for expression and purification of properly folded functional hemagglutinin as a rapid response to emerging pandemic strains. A recombinant H5N1 (A/Vietnam/1203/2004) hemagglutinin globular domain (HA1) was produced in Escherichia coli under controlled redox refolding conditions. Importantly, the properly folded HA1(1-320), i.e., HA1 lacking amino acids 321 to 330, contained ≥75% functional oligomers without addition of foreign oligomerization sequence. Site-directed mutagenesis mapped the oligomerization signal to the HA1 N-terminal Ile-Cys-Ile residues at positions 3 to 5. The purified HA1 oligomers (but not monomers) bound fetuin and agglutinated red blood cells. Upon immunization of rabbits, the oligomeric HA1(1-320) elicited potent neutralizing antibodies against homologous and heterologous H5N1 viruses more rapidly than HA1(28-320) containing only monomers. Ferrets vaccinated with oligomeric HA1 (but not monomeric HA1 with the N terminus deleted) at 15 and 3 µg/dose were fully protected from lethality and weight loss after challenge with homologous H5N1 (A/Vietnam/1203/2004, clade 1) virus, as well as heterologous clade 2.2 H5N1 (A/WooperSwan/Mongolia/244/2005) virus. Protection was associated with a significant reduction in viral loads in the nasal washes of homologous and heterologous virus challenged ferrets. This is the first study that describes the presence of an N-terminal oligomerization sequence in the globular domain of influenza virus hemagglutinin. Our findings suggest that functional oligomeric rHA1-based vaccines can be produced efficiently in bacterial systems and can be easily upscaled in response to a pandemic influenza virus threat.

Passive immunotherapies protect WRvFire and IHD-J-Luc vaccinia virus-infected mice from lethality by reducing viral loads in the upper respiratory tract and internal organs.

Whole-body bioimaging was employed to study the effects of passive immunotherapies on lethality and viral dissemination in BALB/c mice challenged with recombinant vaccinia viruses expressing luciferase. WRvFire and IHD-J-Luc vaccinia viruses induced lethality with similar times to death following intranasal infection, but WRvFire replicated at higher levels than IHD-J-Luc in the upper and lower respiratory tracts. Three types of therapies were tested: licensed human anti-vaccinia virus immunoglobulin intravenous (VIGIV); recombinant anti-vaccinia virus immunoglobulin (rVIG; Symphogen, Denmark), an investigational product containing a mixture of 26 human monoclonal antibodies (HuMAbs) against mature virion (MV) and enveloped virion (EV); and HuMAb compositions targeting subsets of MV or EV proteins. Bioluminescence recorded daily showed that pretreatment with VIGIV (30 mg) or with rVIG (100 µg) on day -2 protected mice from death but did not prevent viral replication at the site of inoculation and dissemination to internal organs. Compositions containing HuMAbs against MV or EV proteins were protective in both infection models at 100 µg per animal, but at 30 µg, only anti-EV antibodies conferred protection. Importantly, the t statistic of the mean total fluxes revealed that viral loads in surviving mice were significantly reduced in at least 3 sites for 3 consecutive days (days 3 to 5) postchallenge, while significant reduction for 1 or 2 days in any individual site did not confer protection. Our data suggest that reduction of viral replication at multiple sites, including respiratory tract, spleen, and liver, as monitored by whole-body bioluminescence can be used to predict the effectiveness of passive immunotherapies in mouse models.

H5N1 virus-like particle vaccine elicits cross-reactive neutralizing antibodies that preferentially bind to the oligomeric form of influenza virus hemagglutinin in humans.

Transmission of pathogenic avian influenza viruses (AIV) from wild birds to domestic poultry and humans is continuing in multiple countries around the world. In preparation for a potential AIV pandemic, multiple vaccine candidates are under development. In the case of H5N1 AIV, a clear shift in transmission from clade 1 to clade 2 viruses occurred in recent years. The virus-like particle (VLP) represents an economical approach to pandemic vaccine development. In the current study, we evaluated the humoral immune response in humans vaccinated with H5N1 A/Indonesia/05/2005 (clade 2.1) VLP vaccine manufactured in Sf9 insect cells. The VLPs were comprised of the influenza virus hemagglutinin (HA), neuraminidase (NA), and matrix 1 (M1) proteins. In an FDA-approved phase I/II human clinical study, two doses of H5N1 VLPs at 15, 45, or 90 µg HA/dose resulted in seroconversion and production of functional antibodies. Moreover, cross-reactivity against other clade 2 subtypes was demonstrated using virus neutralization assays. H5N1 whole-genome fragment phage display libraries (GFPDL) were used to elucidate the antibody epitope repertoire in postvaccination human sera. Diverse epitopes in HA1/HA2 and NA were recognized by postvaccination sera from the two high-dose groups, including large segments spanning the HA1 receptor binding domain. Importantly, the vaccine elicited sera that preferentially bound to an oligomeric form of recombinant HA1 compared with monomeric HA1. The oligomeric/monomeric HA1 binding ratios of the sera correlated with the virus neutralizing titers. Additionally, the two high-dose VLP vaccine groups generated NA-inhibiting antibodies that were associated with binding to a C-terminal epitope close to the sialic acid binding site. These findings represent the first report describing the quality of the antibody responses in humans following AIV VLP immunization and support further development of such vaccines against emerging influenza virus strains.

Smallpox vaccine-induced antibodies are necessary and sufficient for protection against monkeypox virus.

Vaccination with live vaccinia virus affords long-lasting protection against variola virus, the agent of smallpox. Its mode of protection in humans, however, has not been clearly defined. Here we report that vaccinia-specific B-cell responses are essential for protection of macaques from monkeypox virus, a variola virus ortholog. Antibody-mediated depletion of B cells, but not CD4+ or CD8+ T cells, abrogated vaccine-induced protection from a lethal intravenous challenge with monkeypox virus. In addition, passive transfer of human vaccinia-neutralizing antibodies protected nonimmunized macaques from severe disease. Thus, vaccines able to induce long-lasting protective antibody responses may constitute realistic alternatives to the currently available smallpox vaccine (Dryvax).

Influenza vaccination and single cell multiomics reveal sex dimorphic immune imprints of prior mild COVID-19.

Viral infections can have profound and durable functional impacts on the immune system. There is an urgent need to characterize the long-term immune effects of SARS-CoV-2 infection given the persistence of symptoms in some individuals and the continued threat of novel variants. Here we use systems immunology, including longitudinal multimodal single cell analysis (surface proteins, transcriptome, and V(D)J sequences) from 33 previously healthy individuals after recovery from mild, non-hospitalized COVID-19 and 40 age- and sex-matched healthy controls with no history of COVID-19 to comparatively assess the post-infection immune status (mean: 151 days after diagnosis) and subsequent innate and adaptive responses to seasonal influenza vaccination. Identification of both sex-specific and -independent temporally stable changes, including signatures of T-cell activation and repression of innate defense/immune receptor genes (e.g., Toll-like receptors) in monocytes, suggest that mild COVID-19 can establish new post-recovery immunological set-points. COVID-19-recovered males had higher innate, influenza-specific plasmablast, and antibody responses after vaccination compared to healthy males and COVID-19-recovered females, partly attributable to elevated pre-vaccination frequencies of a GPR56 expressing CD8+ T-cell subset in male recoverees that are "poised" to produce higher levels of IFNγ upon inflammatory stimulation. Intriguingly, by day 1 post-vaccination in COVID-19-recovered subjects, the expression of the repressed genes in monocytes increased and moved towards the pre-vaccination baseline of healthy controls, suggesting that the acute inflammation induced by vaccination could partly reset the immune states established by mild COVID-19. Our study reveals sex-dimorphic immune imprints and in vivo functional impacts of mild COVID-19 in humans, suggesting that prior COVID-19, and possibly respiratory viral infections in general, could change future responses to vaccination and in turn, vaccines could help reset the immune system after COVID-19, both in an antigen-agnostic manner.

Application of bioluminescence imaging to the prediction of lethality in vaccinia virus-infected mice.

To find an alternative endpoint for the efficacy of antismallpox treatments, bioluminescence was measured in live BALB/c mice following lethal challenge with a recombinant WR vaccinia virus expressing luciferase. Intravenous vaccinia immunoglobulin treatments were used to confer protection on a proportion of animals. Using known lethality outcomes in 200 animals and total fluxes recorded daily in live animals, we performed univariate receiver operating characteristic (ROC) curve analysis to assess whether lethality can be predicted based on bioluminescence. Total fluxes in the spleens on day 3 and in the livers on day 5 generated accurate predictive models; the area under the ROC curve (AUC) was 0.91. Multiple logistic regression analysis utilizing a linear combination of six measurements: total flux in the liver on days 2, 3, and 5; in the spleen on days 1 and 3; and in the nasal cavity on day 4 generated the most accurate predictions (AUC = 0.96). This model predicted lethality in 90% of animals with only 10% of nonsurviving animals incorrectly predicted to survive. Compared with bioluminescence, ROC analysis with 25% and 30% weight loss as thresholds accurately predicted survival on day 5, but lethality predictions were low until day 9. Collectively, our data support the use of bioimaging for lethality prediction following vaccinia virus challenge and for gaining insight into protective mechanisms conferred by vaccines and therapeutics.

Electroporation of synthetic DNA antigens offers protection in nonhuman primates challenged with highly pathogenic avian influenza virus.

Avian influenza highlights the need for novel vaccination techniques that would allow for the rapid design and production of safe and effective vaccines. An ideal platform would be capable of inducing both protective antibodies and potent cellular immune responses. These potential advantages of DNA vaccines remain unrealized due to a lack of efficacy in large animal studies and in human trials. Questions remain regarding the potential utility of cellular immune responses against influenza virus in primates. In this study, by construct optimization and in vivo electroporation of synthetic DNA-encoded antigens, we observed the induction of cross-reactive cellular and humoral immune responses individually capable of providing protection from influenza virus infection in the rhesus macaque. These studies advance the DNA vaccine field and provide a novel, more tolerable vaccine with broad immunogenicity to avian influenza virus. This approach appears important for further investigation, including studies with humans.

Antigenic fingerprinting of H5N1 avian influenza using convalescent sera and monoclonal antibodies reveals potential vaccine and diagnostic targets.

BACKGROUND: Transmission of highly pathogenic avian H5N1 viruses from poultry to humans have raised fears of an impending influenza pandemic. Concerted efforts are underway to prepare effective vaccines and therapies including polyclonal or monoclonal antibodies against H5N1. Current efforts are hampered by the paucity of information on protective immune responses against avian influenza. Characterizing the B cell responses in convalescent individuals could help in the design of future vaccines and therapeutics. METHODS AND FINDINGS: To address this need, we generated whole-genome-fragment phage display libraries (GFPDL) expressing fragments of 15-350 amino acids covering all the proteins of A/Vietnam/1203/2004 (H5N1). These GFPDL were used to analyze neutralizing human monoclonal antibodies and sera of five individuals who had recovered from H5N1 infection. This approach led to the mapping of two broadly neutralizing human monoclonal antibodies with conformation-dependent epitopes. In H5N1 convalescent sera, we have identified several potentially protective H5N1-specific human antibody epitopes in H5 HA[(-10)-223], neuraminidase catalytic site, and M2 ectodomain. In addition, for the first time to our knowledge in humans, we identified strong reactivity against PB1-F2, a putative virulence factor, following H5N1 infection. Importantly, novel epitopes were identified, which were recognized by H5N1-convalescent sera but did not react with sera from control individuals (H5N1 naïve, H1N1 or H3N2 seropositive). CONCLUSION: This is the first study, to our knowledge, describing the complete antibody repertoire following H5N1 infection. Collectively, these data will contribute to rational vaccine design and new H5N1-specific serodiagnostic surveillance tools.

Repeat vaccination reduces antibody affinity maturation across different influenza vaccine platforms in humans.

Several vaccines are approved in the United States for seasonal influenza vaccination every year. Here we compare the impact of repeat influenza vaccination on hemagglutination inhibition (HI) titers, antibody binding and affinity maturation to individual hemagglutinin (HA) domains, HA1 and HA2, across vaccine platforms. Fold change in HI and antibody binding to HA1 trends higher for H1N1pdm09 and H3N2 but not against B strains in groups vaccinated with FluBlok compared with FluCelvax and Fluzone. Antibody-affinity maturation occurs against HA1 domain of H1N1pdm09, H3N2 and B following vaccination with all vaccine platforms, but not against H1N1pdm09-HA2. Importantly, prior year vaccination of subjects receiving repeat vaccinations demonstrated reduced antibody-affinity maturation to HA1 of all three influenza virus strains irrespective of the vaccine platform. This study identifies an important impact of repeat vaccination on antibody-affinity maturation following vaccination, which may contribute to lower vaccine effectiveness of seasonal influenza vaccines in humans.

AS03-adjuvanted H5N1 vaccine promotes antibody diversity and affinity maturation, NAI titers, cross-clade H5N1 neutralization, but not H1N1 cross-subtype neutralization.

Immune responses to inactivated vaccines against avian influenza are poor due in part to lack of immune memory. Adjuvants significantly increased virus neutralizing titers. We performed comprehensive analyses of polyclonal antibody responses following FDA-approved adjuvanted H5N1-A/Indonesia vaccine, administered in presence or absence of AS03. Using Whole Genome Fragment Phage Display Libraries, we observed that AS03 induced antibody epitope diversity to viral hemagglutinin (HA) and neuraminidase compared with unadjuvanted vaccine. Furthermore, AS03 promoted significant antibody affinity maturation to properly folded H5-HA1 (but not to HA2) domain, which correlated with neutralization titers against both vaccine and heterologous H5N1 strains. However, no increase in heterosubtypic cross-neutralization of Group1-H1N1 seasonal strains was observed. AS03-H5N1 vaccine also induced higher neuraminidase inhibition antibody titers. This study provides insight into the differential impacts of AS03 adjuvant on H5N1 vaccine-induced antibody responses that may help optimize vaccine platforms for future vaccines with improved protection against seasonal and pandemic influenza strains.

Durable neutralizing antibodies after remote smallpox vaccination among adults with and without HIV infection.

The only US licensed vaccine with established efficacy against smallpox, Dryvax, is contraindicated for HIV patients. Detectable smallpox-neutralizing antibodies are still present among US adults. This study compared vaccinia-neutralizing antibody titers between 20 HIV-infected and 20 uninfected veterans matched for age and military entry. Vaccinia-neutralizing antibodies were detected in 95% HIV-infected and 100% uninfected veterans; 40% HIV-infected and 70% uninfected adults had protective titers. Therefore, after robust vaccination, neutralizing antibodies are maintained for prolonged times despite CD4 cell depletion.

A Potent Germline-like Human Monoclonal Antibody Targets a pH-Sensitive Epitope on H7N9 Influenza Hemagglutinin.

The H7N9 influenza virus causes high-mortality disease in humans but no effective therapeutics are available. Here we report a human monoclonal antibody, m826, that binds to H7 hemagglutinin (HA) and protects against H7N9 infection. m826 binds to H7N9 HA with subnanomolar affinity at acidic pH and 10-fold lower affinity at neutral pH. The high-resolution (1.9 Å) crystal structure of m826 complexed with H7N9 HA indicates that m826 binds an epitope that may be fully exposed upon pH-induced conformational changes in HA. m826 fully protects mice against lethal challenge with H7N9 virus through mechanisms likely involving antibody-dependent cell-mediated cytotoxicity. Interestingly, immunogenetic analysis indicates that m826 is a germline antibody, and m826-like sequences can be identified in H7N9-infected patients, healthy adults, and newborn babies. These m826 properties offer a template for H7N9 vaccine immunogens, a promising candidate therapeutic, and a tool for exploring mechanisms of virus infection inhibition by antibodies.

Clinical and immunological comparison of smallpox vaccine administered to the outer versus the inner upper arms of vaccinia-naive adults.

BACKGROUND: Current recommendations direct health care providers to administer smallpox vaccine to the upper outer arm. However, concerns about cosmetically bothersome scarring, accidental contact transmission, interference by body tattoos, and even malignant transformation suggest evaluation of alternate vaccination sites is warranted. METHODS: We randomized 20 vaccinia-naive adults to undergo smallpox vaccination on the outer (n = 10) or inner (n = 10) upper arm. Evaluations included major reaction ("take") rates and vaccination site cultures on postvaccination day 7, determination of serum vaccinia-specific neutralizing antibody titers on days 0 (prevaccination) and 21, and determination of adverse events. RESULTS: On postvaccination day 7, a total of 18 participants (9 per group) had major reactions, 17 of whom had culture evidence of viable vaccinia. The inner and outer arm groups had similar major reaction mean sizes (P = .17), but the amount of erythema (in square centimeters) was smaller in the inner arm group (P = .05). At day 21, all participants had higher titers of vaccinia-specific neutralizing antibodies, compared with at day 0, and the geometric mean titer values of the 2 vaccine groups were similar (P = .45). Adverse event rates were similar. CONCLUSION: The comparable clinical, immunological, and tolerability outcomes between smallpox vaccine applied to the conventional upper outer arm site versus the upper inner arm, coupled with modestly less vaccine-site erythema on the inner arm, indicate that the inner arm may be a suitable alternate vaccination site.