Elicit Antibody Responses Research Articles

Temperature-regulated protein synthesis by Leptospira interrogans.

Leptospira interrogans is an important mammalian pathogen. Transmission from an environmental source requires adaptations to a range of new environmental conditions in the organs and tissues of the infected host. Since many pathogenic bacteria utilize temperature to discern their environment and regulate the synthesis of appropriate proteins, we investigated the effects of temperature on protein synthesis in L. interrogans. Bacteria were grown for several days after culture temperatures were shifted from 30 to 37 degrees C. Triton X-114 cellular fractionation identified several proteins of the cytoplasm, periplasm, and outer membrane for which synthesis was dependent on the culture temperature. Synthesis of a cytoplasmic protein of 20 kDa was switched off at 37 degrees C, whereas synthesis of a 66-kDa periplasmic protein was increased at the higher temperature. Increased synthesis of a 25-kDa outer membrane protein was observed when the organisms were shifted from 30 to 37 degrees C. A 36-kDa protein synthesized at 30 but not at 37 degrees C was identified as LipL36, an outer membrane lipoprotein. In contrast, expression of another lipoprotein, LipL41, was the same at either temperature. Immunoblotting with convalescent equine sera revealed that some proteins exhibiting thermoregulation of synthesis elicited antibody responses during infection. Our results show that sera from horses which aborted as a result of naturally acquired infection with L. interrogans serovar pomona type kennewicki recognize periplasmic and outer membrane proteins which are differentially synthesized in response to temperature and which therefore may be important in the host-pathogen interaction during infection.

Strategies for Mimicking Neisserial Saccharide Epitopes as Vaccines

Monoclonal antibody (mAb) 2C7 recognizes a conserved and widely expressed oligosaccharide (OS) epitope on Neisseria gonorrhoeae. This OS epitope evokes a significant bactericidal and opsonic immune response after natural infection and vaccination. The OS epitope structure represents an excellent target for a potential protective gonococcal vaccine. Because carbohydrate antigens are T-cell independent, inducing weak antibody responses, OS molecules are not useful immunogens. We developed and examined two different strategies to mimic the 2C7 OS epitope: (i) an anti-idiotope (mAb CAI); and (ii) a peptide (PEP-1). These surrogate immunogens elicited antibody responses in mice (CAI and PEP-1) and rabbits (CAI) that were bactericidal in vitro against gonococci. Both CAI and PEP-1 arc true immunologic mimics of OS and may form a basis for the development of vaccine candidates for human immunization against N. gonorrhoeae.

Glycosylphosphatidylinositol anchors of Plasmodium falciparum: molecular characterization and naturally elicited antibody response that may provide immunity to malaria pathogenesis.

Induction of proinflammatory cytokine responses by glycosylphosphatidylinositols (GPIs) of intraerythrocytic Plasmodium falciparum is believed to contribute to malaria pathogenesis. In this study, we purified the GPIs of P. falciparum to homogeneity and determined their structures by biochemical degradations and mass spectrometry. The parasite GPIs differ from those of the host in that they contain palmitic (major) and myristic (minor) acids at C-2 of inositol, predominantly C18:0 and C18:1 at sn-1 and sn-2, respectively, and do not contain additional phosphoethanolamine substitution in their core glycan structures. The purified parasite GPIs can induce tumor necrosis factor alpha release from macrophages. We also report a new finding that adults who have resistance to clinical malaria contain high levels of persistent anti-GPI antibodies, whereas susceptible children lack or have low levels of short-lived antibody response. Individuals who were not exposed to the malaria parasite completely lack anti-GPI antibodies. Absence of a persistent anti-GPI antibody response correlated with malaria-specific anemia and fever, suggesting that anti-GPI antibodies provide protection against clinical malaria. The antibodies are mainly directed against the acylated phosphoinositol portion of GPIs. These results are likely to be valuable in studies aimed at the evaluation of chemically defined structures for toxicity versus immunogenicity with implications for the development of GPI-based therapies or vaccines.

Contribution of defined amino acid residues to the immunogenicity of recombinant Escherichia coli heat-stable enterotoxin fusion proteins

We investigated whether the toxicity-associated receptor-binding domain of the non-immunogenic Escherichia coli heat-stable enterotoxin (STh) as a fusion with a carrier protein and the inclusion of an appropriate spacer are critical factors for eliciting antibody responses against the native toxin. The immunological properties of three toxic and one non-toxic fusion proteins, consisting of STh N-terminally joined to the C-terminus of the major subunit ClpG of E. coli CS31A fimbriae, were compared. In contrast to the non-toxic hybrid STh with glycine and leucine simultaneously substituted for the receptor-interacting Pro 13 and Ala 14 amino acids, the toxic chimeras responded by producing high serum levels of anti-STh antibodies in immunized animals. On the other hand, only the toxic ClpG-STh construct with the natural peptide 47KSGPESM 53 of Pro-STh as spacer stimulated STh-neutralizing responses against both native toxin and enterotoxigenic live E. coli cells. Altogether, these findings suggest a close relationship between conformational similarity to the native structure of STh and the ability to elicit specific antibody responses against STh.

Safety and immunogenicity of NYVAC-JEV and ALVAC-JEV attenuated recombinant Japanese encephalitis virus — poxvirus vaccines in vaccinia-nonimmune and vaccinia-immune humans

A controlled, randomized, double-blind clinical trial evaluated whether two attenuated recombinant poxviruses with identical Japanese encephalitis virus (JEV) gene insertions, NYVAC-JEV and ALVAC-JEV, were safe and immunogenic in volunteers. Groups of 10 volunteers distinguished by vaccinia immune status received two doses of each vaccine. The vaccines appeared to be equally safe and well tolerated in volunteers, but more reactogenic than licensed formalin-inactivated JE and placebo vaccines given as controls. NYVAC-JEV and ALVAC-JEV vaccine recipients had frequent occurrence of local warmth, erythema, tenderness, and/or arm pain after vaccination. There was no apparent effect of vaccinia immune status on frequency or magnitude of local and systemic reactions. NYVAC-JEV elicited antibody responses to JEV antigens in recipients but ALVAC-JEV vaccine poorly induced antibody responses. However, NYVAC-JEV vaccine induced neutralizing antibody responses only in vaccinia-nonimmune recipients while vaccinia-immune volunteers failed to develop protective antibodies (5/5 vs. 0/5 seroconversion, p<0.01). These data suggest that preexisting immunity to poxvirus vector may suppress antibody responses to recombinant gene products.

Accessory cell defect in unresponsiveness of neonates and aged to polysaccharide vaccines

T independent antigens elicit antibody responses in the absence of carrier specific T helper cells but require signals from accessory cells (macrophages and dendritic cells) or specific cytokines. They are further subdivided into TI-1 and TI-2 categories based on the ability of TI-1 but not TI-2 antigens to elicit immune responses from neonates. Most bacterial polysaccharides including the pneumococcal polysaccharide vaccines belong to the TI-2 class. It is hypothesized that defects in accessory cell function play a critical role in the failure of neonates to respond to such TI-2 antigens. Immune responses to these TI-2 stimuli are also reduced in the aged, also due to a quantitative deficiency in accessory cells. Agents that can stimulate accessory cell function may provide an alternative strategy to improve the immunogenicity of the polysaccharide vaccines in the neonates and the aged.

Avaliação da resposta de antitoxinas beta e épsilon de Clostridium perfringens induzidas em bovinos e coelhos por seis vacinas comerciais no Brasil

Avaliou-se a resposta de antitoxinas beta e épsilon de Clostridium perfringens em bovinos vacinados contra clostridioses com seis vacinas disponíveis no mercado. Quarenta e oito bezerros de seis a sete meses de idade foram divididos em oito grupos (T1 a T8) de seis animais cada. Os grupos de número 2 a 7 receberam as vacinas T2 a T7 nos dias 0 e 42 com a dose e via recomendadas pelos fabricantes. Solução salina e toxóide padrão foram usados nos mesmos dias nos grupos 1 e 8 (T1 e T8), respectivamente, como controles negativo e positivo. Amostras de sangue foram coletadas nos dias 0, 42 e 56 pós-vacinação (PV), para titulação de anticorpos no soro. As vacinas e os controles foram também testados em oito coelhos cada, inoculados nos dias 0 e 21 com metade da dose indicada para bovinos. Os coelhos foram sangrados no dia 35 e os soros foram misturados em partes iguais para cada vacina para a titulação de anticorpos. Os soros dos bovinos foram titulados individualmente contra as toxinas beta e épsilon de C. perfringens pelo método de soroneutralização em camundongos. A vacina T2 apresentou títulos de anticorpos de 22,6 e 5,6 UI/ml e a vacina T4 11,2 e 7,0 UI/ml, respectivamente, contra toxinas beta e épsilon em coelhos. Os títulos do toxóide padrão (T8) foram 45,2 UI/ml contra ambas as toxinas. Em bovinos, as médias dos títulos de anticorpos contra a toxina beta nos dias 42 e 56 PV com a vacina T2 (1,15 UI/ml e 8,0 UI/ml) foram similares ao toxóide padrão (2,02 e 10,03 UI/ml). A vacina T4 (0,73 e 4,54 UI/ml) teve títulos menores (P&lt;0,05) que o toxóide padrão e similares a T2. Contra a toxina épsilon, o toxóide padrão teve média de título (0,97 UI/ml) no dia 42 que foi significativamente maior (P&lt;0,05) do que T4 (0,15 UI/ml) e similar a T2 (0,42 UI/ml). No dia 56, T2 (4,27 UI/ml) teve títulos significativamente maiores (P&lt;0,05) do que T4 (0,68 UI/ml) e similares ao toxóide padrão (4,98 UI/ml). Em cada tratamento, a resposta aos 56 dias foi superior (P&lt;0,05) em relação aos 42 dias após a primeira vacinação. As outras vacinas e a solução salina não induziram respostas de antitoxinas beta e épsilon de C. perfringens detectáveis nos soros dos coelhos e dos bovinos. A vacina T2 induziu altos títulos de anticorpos, maiores que aqueles induzidos por T4 e similares ao T8.

Complement Depletion Facilitates the Infection of Multiple Brain Tumors by an Intravascular, Replication-Conditional Herpes Simplex Virus Mutant

Intravascular routes of administration can provide a means to target gene- and virus-based therapies to multiple tumor foci located within an organ, such as the brain. However, we demonstrate here that rodent plasma inhibits cell transduction by replication-conditional (oncolytic) herpes simplex viruses (HSV), replication-defective HSV, and adenovirus vectors. In vitro depletion of complement with mild heat treatment or in vivo depletion by treatment of athymic rats with cobra venom factor (CVF) partially reverses this effect. Without CVF, inhibition of cell infection by HSV is observed at plasma dilution as high as 1:32, while plasma from CVF-treated animals displays anti-HSV activity at lower dilutions (1:8). When applied to the therapy of intracerebral brain tumors, in vivo complement depletion facilitates the initial infection (assayed at the 2-day time point) by an intra-arterial replication-conditional HSV of tumor cells, located within three separate and distinct human glioma masses. However, at the 4-day time point, no propagation of HSV from initially infected tumor cells could be observed. Previously, we have shown that the immunosuppressive agent, cyclophosphamide (CPA), facilitates the in vivo propagation of an oncolytic HSV, delivered intravascularly, within infected multiple intracerebral masses, by inhibition of both innate and elicited anti-HSV neutralizing antibody response (K. Ikeda et al., Nat. Med. 5:881-889, 1999). In this study, we thus show that the addition of CPA to the CVF treatment results in a significant increase in viral propagation within infected tumors, measured at the 4-day time period. The concerted action of CVF and CPA significantly increases the life span of athymic rodents harboring three separate and large glioma xenografts after treatment with intravascular, oncolytic HSV. Southern analysis of viral genomes analyzed by PCR reveals the presence of the oncolytic virus in the brains, livers, spleens, kidneys, and intestine of treated animals, although none of these tissues displays evidence of HSV-mediated gene expression. In light of clinical trials of oncolytic HSV for malignant brain tumors, these findings suggest that antitumor efficacy may be limited by the host innate and elicited humoral responses.

Introduction

Atherosclerosis, and its complications cerebral and myocardial infarction and gangrene of the extremities, is the leading cause of death in industrialized societies. Large epidemiological studies have revealed the main risk factors associated with an increased risk for atherosclerotic disease to be smoking, hypertension, diabetes and dyslipidaemias. However, these classical risk factors explain only a part of the epidemiological features of atherosclerotic vascular disease. During the last two decades, immunological mechanisms have been increasingly implicated. Postulates for the involvement of innate and adaptive immunity with cellular as well as humoral components have met with considerable experimental support. It is now well recognized that atherosclerotic lesions contain activated, immunocompetent cells, including T cells reactive towards oxidized LDL lipoproteins, heat shock proteins and possibly also other autoantigens. Systemic humoral immune responses can also be detected against such antigens and have been correlated to disease progression in some prospective clinical studies. Immune complexes can appear both in the circulation and in the artery wall, where they can activate complement and thus cause tissue destruction. Immune mechanisms appear to be particularly active when plaques are activated and rupture and they may therefore cause acute coronary syndromes. At such occasions, acute phase reactants, immune cytokines and activated T cells can be detected in the systemic circulation in amounts that correlate to the clinical course of disease. In addition to autoantigens, microbes have been implicated as potential pathogenic factors in atherosclerosis in several recent studies. In particular, Chlamydia pneumoniae can be detected in plaques, can elicit antibody responses that are associated with ischaemic heart disease and has been reported to accelerate experimental atherosclerosis. Although still not completely understood, the possibility of a microbial pathogenic component in atherosclerosis is an exciting challenge for further research and could provide new opportunities for therapy and prevention. Recent research in several laboratories throughout the world have provided important new insights into immunopathogenic and infection mechanisms in atherosclerosis. The Journal of Internal Medicine sponsored a conference with leading investigators in this field in April 1999. The present volume contains state-of-the-art reviews on this controversial topic by the speakers at the conference. It is our hope that it will serve as an introduction and an overview of this important aspect of current research in internal medicine.

Foreign gene expression in Corynebacterium pseudotuberculosis: development of a live vaccine vector

A defined phospholipase D mutant of Corynebacterium pseudotuberculosis, designated Toxminus, was used as a live vector to express and deliver a range of candidate vaccine antigens to sheep. Expression levels of the foreign genes in Toxminus were evaluated when directed from a number of different promoters, both constitutively expressed and inducible, as fusions with expressed genes including a signal sequence, and from chromosomal and episomal loci. In general expression levels were low and it appeared that some of the recombinant proteins were tolerated by C. pseudotuberculosis Toxminus better than others. Gene expression was however sufficiently high for three of the genes to elicit antibody responses specific to the recombinant protein following a single dose of the live Toxminus vector vaccine. This work suggests that C. pseudotuberculosis Toxminus has potential for development as a live veterinary vaccine vector.

Engineering parvovirus-like particles for the induction of B-cell, CD4 + and CTL responses

An antigen delivery system based on hybrid recombinant parvovirus-like particles (VLPs) formed by the self-assembly of the capsid VP2 protein of porcine (PPV) or canine parvovirus (CPV) expressed in insect cells with the baculovirus system has been developed. PPV:VLPs containing a CD8 + epitope from the LCMV nucleoprotein evoked a potent CTL response and were able to protect mice against a lethal infection with the virus. Also, PPV:VLPs containing the C3:T epitope from poliovirus elicited a CD4 + response in mice. These T-cell epitopes were located in the N-terminus of the VP2 protein. Unfortunately, the N-terminus is not adequate for antibody responses because it is inside the particle. Previous attempts to modify the surface-exposed regions of the capsid were unsuccessful. Deletions in the four loops of the surface resulted in the complete disruption of the particles except those in loop 2. However, preliminary insertions of the C3:B epitope of poliovirus in VP2 residue 225 of the loop 2 were not able to elicit antibody responses. To optimise the insertion site, the epitope was inserted consecutively in positions 226, 227 and 228, which seem to be more accessible on the capsid. Mice immunised with these chimeric C3:B CPV:VLPs were able to elicit a strong neutralising antibody response (>3 log 10 units) against poliovirus. The possibility of combining different types of epitopes in different positions of a single particle to stimulate different branches of the immune system paves the way to the production of more potent vaccines in a simple and cheap way.

Parvovirus-Like Particles as Vaccine Vectors

A wide array of systems have been developed to improve “classic” vaccines. The use of small polypeptides able to elicit potent antibody and cytotoxic responses seems to have enormous potential in the design of safer vaccines. While peptide coupling to large soluble proteins such as keyhole limpet hemocyanin is the current method of choice for eliciting antibody responses and insertion in live viruses for cytotoxic T-lymphocyte responses, alternative cheaper and/or safer methods will clearly be required in the future. Virus-like particles constitute very immunogenic molecules that allow for covalent coupling of the epitopes of interest in a simple way. In this article, we detail the methodology employed for the preparation of efficient virus vectors as delivery systems. We used parvovirus as the model for the design of new vaccine vectors. Recently parvovirus-like particles have been engineered to express foreign polypeptides in certain positions, resulting in the production of large quantities of highly immunogenic peptides, and to induce strong antibody, helper-T-cell, and cytotoxic T-lymphocyte responses. We discuss the different alternatives and the necessary steps to carry out this process, placing special emphasis on the flow of decisions that need to be made during the project.

Vaccination with carbohydrate peptide mimotopes promotes anti-tumor responses.

Tumor-associated carbohydrate (TAC) antigens are important targets in cancer vaccine efforts. Carbohydrates are, however, frequently poor immunogens, in that they are T-cell-independent antigens. Molecular mimicry of TAC by peptides is an alternative approach to generating anti-carbohydrate immune responses. Here we demonstrate that peptide mimotopes can elicit antibody responses that cross-react with representative human TAC antigens. Primary immunization with such a multiple antigenic peptide, along with QS-21 as adjuvant, elicits cytotoxic antibodies reactive with naturally occurring forms of TAC expressed on tumor cells, and vaccination of mice with peptide mimotopes reduced tumor growth and prolonged host survival in a murine tumor model.

Antigenic and immunogenic properties of totally synthetic peptide-based anti-fertility vaccines.

In this study we describe the results of experiments in which a variety of totally synthetic luteinizing hormone releasing hormone (LHRH) vaccines were assembled and examined for their abilities to elicit antibody responses and induce sterility in mice. It is shown that totally synthetic vaccines consisting of a 15 residue-defined T cell epitope and the 10 residue LHRH epitope not only induced high titers of antibody but also induced sterility. This effect did not appear to correlate with antibody titer, antibody isotype or comparative antibody affinity, but may be related to the length of time for which antibodies are present to exert their influence.

Detection of anti-VacA antibody responses in serum and gastric juice samples using type s1/m1 and s2/m2 Helicobacter pylori VacA antigens.

Several different families of vacuolating toxin (vacA) alleles are present in Helicobacter pylori, and they encode products with differing functional activities. H. pylori strains containing certain types of vacA alleles have been associated with an increased risk for peptic ulcer disease. In this study, we tested serum samples and gastric juice from 19 H. pylori-negative and 39 H. pylori-positive patients for enzyme-linked immunosorbent assay reactivity with two different types of VacA antigens (types s1/m1 and s2/m2), which were purified from H. pylori 60190 and 86-338, respectively. Both antigens were recognized better by serum immunoglobulin G (IgG) from H. pylori-positive persons than by serum IgG from H. pylori-negative persons (P < 0.01). The s1/m1 VacA antigen was better recognized by sera from patients carrying vacA type s1/m1 strains than by sera from patients carrying vacA type s2/m2 or s1/m2 strains (P < 0.01). Conversely, the s2/m2 VacA antigen was better recognized by sera from patients carrying type s2/m2 or s1/m2 strains (P = 0.03). Serum IgG anti-VacA antibodies were present more frequently in patients carrying type s1/m1 strains than in other H. pylori-positive patients (P = 0.0002). In addition, the highest levels of IgA anti-VacA antibodies were detected in the gastric juice of patients carrying type s1/m1 strains. These data indicate that different VacA isoforms have distinct antigenic properties and that multiple forms of VacA elicit antibody responses in H. pylori-positive humans.

Xenogeneic and allogeneic anti-MHC immune responses induced by plasmid DNA immunization

Major histocompatibility complex (MHC) proteins are known to be incorporated into the human immunodeficiency virus (HIV-1) envelope as the virion buds from the host cell surface. Studies using simian immunodeficiency virus (SIV) infection of macaques have demonstrated that immunization with uninfected human cells or purified HLA proteins can provide protection from challenge with live SIV when it is grown in human cells expressing the same MHC alleles. Thus the induction of anti-MHC immune responses represents an important option to consider with respect to vaccine design for SIV and HIV. Here we examine plasmid DNA immunization strategies as an alternative to cellular or protein immunogens for the induction of xenogeneic and allogeneic immune responses in C57BL/6 mice and in an HLA transgenic mouse model system, respectively. We compared the immunogenicity of HLA-A2- and HLA-B27-expressing splenocytes with the corresponding plasmid DNA immunogens. Results from the transgenic mouse experiments indicate that plasmid DNA immunization with both class I and class II MHC-encoding vectors can elicit antibody responses recognizing conformationally intact MHC molecules. Our data also show that immunization with class I MHC-encoding DNA immunogens can elicit cytotoxic T-lymphocyte responses, demonstrating the potential to mobilize both antibody and cell-mediated anti-MHC immune responses in the context of this approach to HIV-1 vaccine design.

The major Epstein-Barr virus (EBV) envelope glycoprotein gp340 when incorporated into Iscoms primes cytotoxic T-cell responses directed against EBV lymphoblastoid cell lines

A recombinant form of the EBV envelope glycoprotein and vaccine candidate gp340, lacking its hydrophobic transmembrane region, was incorporated into Iscoms after coupling to phosphatidyl ethanolamine via carbohydrate residues. Coupling by partial oxidation of gp340 carbohydrate with sodium periodate partly denatured the incorporated gp340 as indicated by its reduced reactivity with monoclonal antibodies that recognise the major neutralising epitope. Immunisation of cottontop tamarins with these Iscoms elicited antibody responses to gp340, but these antibodies only poorly recognised the major neutralising epitope in a competition ELISA and were unable to neutralise EBV in vitro. Despite the lack of neutralising antibody, immunisation with these Iscoms primed significant in vitro proliferative responses to soluble gp340 in lymphocytes from the draining lymph nodes and spleen. T-cell lines were raised from both immunised and control animals by in vitro stimulation of peripheral blood lymphocytes or spleen cells with autologous EBV-transformed lymphoblastoid cell lines. The T-cell lines from control animals had higher numbers of CD4+ T-cells than CD8+ T-cells and were not cytotoxic for autologous lymphoblastoid cell lines (LCL). In contrast the lines from immunised animals contained more CD8+ T-cells than CD4+ T-cells and had marked cytotoxicity for autologous LCL.

Epitopes of human chorionic gonadotropin and their relationship to immunogenicity and cross-reactivity of beta-chain mutants.

Human chrionic gonadotropin (hCG) is a placental glycoprotein hormone, a heterodimeric molecule, consisting of alpha and beta chains. It induces the synthesis of progesterone, which is essential for the maintenance of the fertilized egg. Antibodies directed against hCG can, therefore, prevent pregnancy and serve as a vaccine. hCG belongs to the glycoprotein hormone family and shares the alpha chain with the other members. The beta chain is a hormone-specific subunit that is unique to hCG, but still possesses 85% amino acid homology with the beta chain of luteinizing hormone (LH), which means that prolonged immunization with hCG produces antibodies that cross-react with LH. We have taken an approach involving the mutation of beta hCG to eliminate cross-reactive epitopes without affecting the natural folding of the polypeptide chain and thus the unique beta hCG-specific epitopes. Several mutants have been constructed that have maintained the binding to hCG-specific monoclonal antibodies (mAbs) but have lost the ability to bind to a panel of LH cross-reactive mAbs. To investigate the immunogenicity of selected mutants, mice were immunized with expression plasmid DNA, containing the gene for wild-type beta hCG and two mutants: mutant 3, with four amino acid substitutions (68 Arg-->Glu; 74 Arg-->Ser; 75 Gly-->His; 79 Val-->His), and mutant 7, with a single amino acid substitution (68 Arg-->Glu). Although both mutants were able to elicit antibody responses in at least some animals, the levels were less than those seen with the wild-type beta hCG DNA, and there seems still to be a residual cross-reactivity with LH. Attempts to improve the immunogenicity of the mutants and to further modify the sequence to remove the cross-reactivity are currently underway.

Differential kinetics and distribution of antibodies in serum and nasal and vaginal secretions after nasal and oral vaccination of humans.

Although nasal vaccination has emerged as an interesting alternative to systemic or oral vaccination, knowledge is scarce about the immune responses after such immunization in humans. In the present study, we have compared the kinetics and organ distribution of the antibody responses after nasal and oral vaccination. We immunized female volunteers nasally or orally with cholera toxin B subunit (CTB) and determined the specific antibody levels in serum and nasal and vaginal secretions, as well as the number of circulating antibody-secreting cells, before immunization and 1, 2, 3, 6, and 26 weeks thereafter. Nasal vaccination induced 9-fold CTB-specific immunoglobulin A (IgA) and 56-fold specific IgG antibody increases in nasal secretions, whereas no significant IgA increase was seen after oral vaccination. Both oral and nasal vaccination resulted in 5- to 6-fold CTB-specific IgA and 20- to 30-fold specific IgG increases in vaginal secretions. Strong serum responses to CTB were also induced by both routes of vaccination. A notable difference between nasal and oral vaccination was that the nasal route elicited a specific antibody response with a later onset but of much longer duration than did the oral route. We conclude from this study that the nasal route is superior to the oral route for administering at least nonliving vaccines against infections in the upper respiratory tract, whereas either oral or nasal vaccination might be used for eliciting antibody responses in the female genital tract.

DNA Vaccines Expressing either the GP or NP Genes of Ebola Virus Protect Mice from Lethal Challenge

DNA vaccines expressing the envelope glycoprotein (GP) or nucleocapsid protein (NP) genes of Ebola virus were evaluated in adult, immunocompetent mice. The vaccines were delivered into the skin by particle bombardment of DNA-coated gold beads with thePowderJect-XRgene gun. Both vaccines elicited antibody responses as measured by ELISA and elicited cytotoxic T cell responses as measured by chromium release assays. From one to four vaccinations with 0.5 μg of the GP DNA vaccine resulted in a dose-dependent protection from Ebola virus challenge. Maximal protection (78% survival) was achieved after four vaccinations. Mice were completely protected with a priming dose of 0.5 μg of GP DNA followed by three or four subsequent vaccinations with 1.5 μg of DNA. Partial protection could be observed for at least 9 months after three immunizations with 0.5 μg of the GP DNA vaccine. Comparing the GP and NP vaccines indicated that approximately the same level of protection could be achieved with either vaccine.