Immunopotentiation of local and systemic humoral immune responses by ISCOMs, liposomes and FCA: role in protection against influenza A in mice

The immunogenicity and protective efficacy of an influenza A subunit vaccine preparation administered to mice in an aqueous form, or presented as immunostimulatory complexes (ISCOMs), liposomes or with Freund's complete adjuvant (FCA), were assessed in comparative studies with live infectious virus. Both intranasal and parenteral routes of administration were assessed. An enzyme-linked immunosorbent assay (ELISA) was used to measure nasal wash and serum antibody responses in groups of unprimed mice, while protection was determined by the recovery of homologous influenza virus from mouse nasal washes and lung homogenates following challenge infection by the intranasal route. The results showed that parenteral administration of the influenza antigen preparations induced variable levels of both local and systemic antibodies at weeks 3, 7 and 22 postimmunization. Although the overall greatest levels of antibody and protection were elicited in mice following live virus infection, formulation of influenza surface haemagglutinin (HA) and neuraminidase (NA) proteins into ISCOMs elicited high and persistent antibody responses and provided relatively good protection of the upper and lower respiratory tracts of these animals. The results also show a relatively poor effect of the subunit antigen preparations in promoting humoral immune responses and protection irrespective of the nature of their presentation, when given by the intranasal route.     

Pathogenicity of Mycoplasma synoviae in broiler chickens

Six isolates of Mycoplasma synoviae, identified as WVU 1853, K1968, K1858, 92D8034, F10-2AS, and FMT, were compared for pathogenicity in broiler chickens. Specific-pathogen-free chickens were inoculated, in two groups of 20, with each isolate by footpad or eyedrop inoculation at 1 day of age and were examined at necropsy 7, 14, 28, and 42 days postinoculation. Specimens were taken for histopathology, culture, polymerase chain reaction assay, and hemagglutination-inhibition serology. Isolates were grouped according to pathogenicity on the basis of differences in lesion development and tissue distribution in the respiratory system, other viscera, and the skeletal system. K1968 (pathogenic) induced lesions in all sites examined in both the footpad and eyedrop inoculation groups. It was detected in all sites following footpad inoculation and in all sites except viscera following eyedrop inoculation. WVU 1853, K1858, and 92D8034 (moderately pathogenic) induced lesions and were detected in all sites following footpad inoculation. With eyedrop inoculation, lesions were identified only in upper and lower respiratory sites, and organisms were detected only in upper respiratory sites. F10-2AS (moderately pathogenic) was similar; however, footpad inoculation failed to induce visceral lesions or permit organism detection in any site. F10-2AS was detected in upper and lower respiratory tissues following eyedrop inoculation. FMT (mildly pathogenic) induced only upper respiratory lesions when either footpad or eyedrop inoculation was used, and detection was restricted to upper respiratory sites following eyedrop inoculation. These results are useful in comparative evaluations of the virulence of other M. synoviae isolates and form a basis for characterization of virulence factors of M. synoviae.     

Protective immune response of chickens against Newcastle disease, induced by the intranasal vaccination with inactivated virus

Intranasal vaccination of chickens with inactivated Newcastle disease virus (NDV) induced both local and systemic antibody responses, resulting in protection against intranasal challenge with a lethal dose of a virulent NDV strain. The immune response was enhanced by the use of cholera toxin B subunit (CTB) as an adjuvant and only small amounts of the challenge virus were recovered from the birds vaccinated together with CTB. On the other hand, subcutaneous vaccination with the same antigen induced only a serum antibody response in chickens, allowing the challenge virus to replicate in the sinus. The present results indicate that secretory antibodies induced on the respiratory mucosal surface by intranasal vaccination with inactivated NDV protected chickens from lethal infection by inhibiting virus replication at the portal of entry for the virus.     

Effect of mu-opioids morphine and buprenorphine on the development of adjuvant arthritis in rats

OBJECTIVE AND DESIGN: On the basis that endogenous opioids play a role in the physiological response to inflammation, this study tests the anti-arthritic effects of a mu-opioid agonist, morphine and the partial mu-agonist, buprenorphine. MATERIAL: Male Lewis rats were used. TREATMENT: Rats were inoculated subcutaneously with 0.05 ml of Freund's complete adjuvant (5 mg/ml) into the right hind paw to produce adjuvant arthritis. Morphine (either 10 to 60 mg/kg/day s.c. bolus or 60 mg/kg/day s.c. infusion) and buprenorphine (0.65 +/- 0.06 mg/kg/day, orally), respectively, were administered for 3 days during the primary inflammatory phase of adjuvant arthritis. METHODS: The progression of adjuvant arthritis was monitored every three days by body weight change and hind limb oedema (ipsilateral and contralateral). On day 21 the animals were sacrificed and histology and radiography of the contralateral limb were performed. In rats receiving Freund's adjuvant and no drug treatment, the incidence of arthritis was 89%. Effect was expressed as the pooled severity index (PSI) derived from the arithmetic average of the volume, histology and radiography scores in the contralateral hind limb. RESULTS: Buprenorphine had no effect on experimental arthritis (PSI control vs treated: 242 +/- 28 vs 253 +/- 28%). In contrast, morphine by subcutaneous injection twice daily (10 to 60 mg/kg/day) but not by subcutaneous infusion (60 mg/kg/day) was found to attenuate the progression of adjuvant arthritis in a dose-dependent manner. This indicates that the anti-arthritic effects of morphine are opioid receptor mediated (ED50, 58 +/- 9 mg/kg) and suggests that the local concentration reached effective levels only after subcutaneous injection. It is also possible that the high doses of morphine were anti-inflammatory through effects at the kappa receptor. However, these high doses of morphine produced death in one third of the rats, the calculated lethal dose (LD50, 63 +/- 2 mg/kg) being close to the effective dose. CONCLUSION: Anti-arthritic effects of morphine are opioid receptor mediated but morphine use for this indication is restricted by its adverse effects.     

Effects of radiolysis on yttrium-90-labeled Lym-1 antibody preparations

An Australian field isolate of Mycoplasma synoviae (MS), 89079/7NS, was exposed to the mutagen N-nitro-N'-methyl-N-nitrosoguanidine. Fifteen clones from the exposed culture were characterized for temperature sensitivity. Four clones labelled B, D, G, and H were temperature sensitive and were further characterized for their ability to colonize chickens and elicit an immune response. Serum antibody responses to MS were detected 3 wk after infection, by eyedrop, in 10 of 10 birds inoculated with 86079/7NS and clones B and G and in 9 of 10 birds inoculated with clone H. No MS antibody response was observed in any bird inoculated with clone D. MS was recovered from the upper trachea of 10 of 10 birds inoculated with clones B, G, and H at 2 wk after infection. No MS was isolated from birds inoculated with clone D. Clone H, designated MS-H, was selected as a potential vaccine candidate.     

Macrophage inflammatory protein-1alpha (MIP-1alpha) expression plasmid enhances DNA vaccine-induced immune response against HIV-1

CD8+ cell-secreted CC-chemokines, MIP-1alpha, and MIP-beta have recently been identified as factors which suppress HIV. In this study we co-inoculated MIP-1alpha expression plasmid with a DNA vaccine constructed from HIV-1 pCMV160IIIB and pcREV, and evaluated the effect of the adjuvant on HIV-specific immune responses following intramuscular and intranasal immunization. The levels of both cytotoxic T lymphocyte (CTL) activity and DTH showed that HIV-specific cell-mediated immunity (CMI) was significantly enhanced by co-inoculation of the MIP-1alpha expression plasmid with the DNA vaccine compared with inoculation of the DNA vaccine alone. The HIV-specific serum IgG1/IgG2a ratio was significantly lowered when the plasmid was co-inoculated in both intramuscular and intranasal routes, suggesting a strong elicitation of the T helper (Th) 1-type response. When the MIP-1alpha expression plasmid was inoculated intramuscularly with the DNA vaccine, an infiltration of mononuclear cells was observed at the injection site. After intranasal administration, the level of mucosal secretory IgA antibody was markedly enhanced. These findings demonstrate that MIP-1alpha expression plasmid inoculated together with DNA vaccine acts as a strong adjuvant for eliciting Th1-derived immunity.     

Protection of chickens with or without maternal antibodies against both Marek's and Newcastle diseases by one-time vaccination with recombinant vaccine of Marek's disease virus type 1

We constructed a stable recombinant Marek's disease virus type 1 (rMDV1) expressing the fusion protein (F) of Newcastle disease virus (NDV) by inserting the coding sequence within the US10 gene of MDV1 by homologous recombination and designated this as rMDV1-US10L(F). The NDV-F protein was significantly expressed under control of the SV40 late promoter in cultured cells infected with the rMDV1. To examine the protective efficacy of the rMDV1, specific pathogen-free (SPF) chickens were vaccinated with rMDV1 at one-day-old. Almost all birds (> 95%) were protected from NDV challenge via intramuscular, ocular, intranasal and intratrachial routes at 4 weeks after vaccination. The rMDV1 showed 100% protection against virulent MDV1 challenge in SPF chickens. Antibody responses against NDV-F and MDV1 antigens were observed at least up to 11 weeks after immunization. When the sera from chickens vaccinated with the rMDV1 were examined for the presence of anti-NDV-F antibody on the day of NDV challenge, the vaccinated bird group which did not survive from NDV challenge were found to show lower antibody titers than the surviving group. The rMDV1 also provided sufficient protection against NDV and MDV1 challenges in commercial chickens with maternal antibodies against NDV-F and MDV1 antigens.     

Focal epithelial hyperplasia in an HIV positive man. An illustrated case and review of the literature

Chickens 14 days old were experimentally inoculated with Mycoplasma gallisepticum (MG) R-P10 strain. After development of respiratory symptoms, birds were left unmedicated or medicated for 5 consecutive days with Difloxacin 5, 7.5 or 10 mg/kg body weight per day or Enrofloxacin at the dose level of 10 mg/kg body weight per day. Evaluation of efficacy was based on body weight, symptoms, post-mortem findings, re-isolation of MG and serology. Results indicated that under the conditions of this experiment, treatment with 7.5 mg Difloxacin per kg body weight for 5 days was already effective against pathogenic MG infection. The dose of 10 mg/kg Difloxacin was equally effective as a dose of 10 mg/kg Enrofloxacin in treating respiratory symptoms.     

Experimental autoimmune myasthenia gravis induction in B cell-deficient mice

Experimental autoimmune myasthenia gravis (EAMG) is an animal model for human myasthenia gravis (MG). Autoantibody-induced functional loss of nicotinic acetylcholine receptor (AChR) at the postsynaptic membrane is an important pathogenic feature of both MG and EAMG. To evaluate the extent at which the humoral immune response against AChR operates in the pathogenesis of EAMG, we immunized B cell knockout (muMT) and wild-type C57BL/6 mice with AChR and complete Freund's adjuvant. The ability of AChR-primed lymph node cells to proliferate and secrete IFN-gamma in response to AChR and its dominant peptide alpha146-162 were intact in muMT mice as in wild-type mice. Similar amounts of mRNA for IFN-gamma, IL-4 and IL-10 in AChR-reactive lymph node cells were detected in muMT and wild-type mice. However, muMT mice had no detectable anti-AChR antibodies and remained completely free from clinical EAMG. We conclude that B cells are critically required for the genesis of clinical EAMG, but not for AChR-specific T cell priming.     

Altered alveolar macrophage function in calorie-restricted rats

The capacity of four chicken lines (Y11, L2, B13, PA12) to control Salmonella enteritidis (SE) phage type 4 (PT4) systemic colonization was investigated. Thirteen-week-old chickens were intravenously inoculated with 10(6) SE colony-forming units, and the levels of SE colonization were determined at various time intervals after inoculation in liver, spleen, genital organs, and ceca. The course of SE infection showed a rapid contamination of liver, spleen, and genital organs, whereas the ceca were infected later. A significant (P < 0.001) effect of the chicken line on levels of SE was detected on day 3 postinoculation (PI) in liver and ceca, on day 10 PI in ceca, and on day 15 PI in spleen. Because an early control of systemic Salmonella infection by the Ity/Nramp1 gene has been demonstrated in mice, we aimed to study the early resistance of chickens to SE. As a consequence, we then focused our study on the between- and within-line variabilities of SE levels on day 3 PI. According to the SE levels in liver on day 3 PI, the chicken lines could be classified as susceptible (Y11 and L2) or resistant (PA12 and B13). This early variability was explored in resistant B13 and susceptible L2 lines. Differences between these two lines were confirmed in liver but not in ceca. A large within-line variability was observed in all organs of these two lines. The genetic origin of this variability will have to be determined as a prerequisite to an eventual selection.     

Protection against lethal Japanese encephalitis virus infection of mice by immunization with the highly attenuated MVA strain of vaccinia virus expressing JEV prM and E genes

Genes encoding the glycosylated precursor of the membrane (prM) and envelope (E) proteins of a Korean strain of Japanese encephalitis virus (JEV) were inserted into the genome of the host-range restricted, highly attenuated, and safety-tested MVA strain of vaccinia virus. MVA recombinants containing the JEV genes, under strong synthetic or modified H5 vaccinia virus promoters, were isolated. Synthesis of JEV prM and E proteins was detected by immunofluorescence microscopy, flow cytometry, and polyacrylamide gel electrophoresis. Mice inoculated and boosted by various routes with either of the MVA recombinants produced JEV neutralizing antibodies, that had titres comparable with those induced by an inactivated JEV vaccine, as well as haemagglutination-inhibiting antibodies. Mice immunized with 2 x 10(6) infectious units of MVA/JEV recombinants by intramuscular or intraperitoneal routes were completely protected against a 10(5) LD50 JEV challenge at 9 weeks of age.     

Induction of antigen-specific antibodies in vaginal secretions by using a nontoxic mutant of heat-labile enterotoxin as a mucosal adjuvant

Immunization of the female reproductive tract is important for protection against sexually transmitted diseases and other pathogens of the reproductive tract. However, intravaginal immunization with soluble antigens generally does not induce high levels of secretory immunoglobulin A (IgA). We recently developed safe mucosal adjuvants by genetically detoxifying Escherichia coli heat-labile enterotoxin, a molecule with a strong mucosal adjuvant activity, and here we describe the use of the nontoxic mutant LTK63 to induce a response in the mouse vagina against ovalbumin (Ova). We compared intravaginal and intranasal routes of immunization for induction of systemic and vaginal responses against LTK63 and Ova. We found that LTK63 is a potent mucosal immunogen when given by either the intravaginal or intranasal route. It induces a strong systemic antibody response and IgG and long-lasting IgA in the vagina. The appearance of vaginal IgA is delayed in the intranasally immunized mice, but the levels of vaginal anti-LTK63 IgA after repeated immunizations are higher in the intranasally immunized mice than in the intravaginally immunized mice. LTK63 also acts as a mucosal adjuvant, inducing a serum response against Ova, when given by both the intravaginal and intranasal routes. However, vaginal IgA against Ova is stimulated more efficiently when LTK63 and antigen are given intranasally. In conclusion, our results demonstrate that LTK63 can be used as a mucosal adjuvant to induce antigen-specific antibodies in vaginal secretions and show that the intranasal route of immunization is the most effective for this purpose.     

Pentadecapeptide BPC 157 positively affects both non-steroidal anti-inflammatory agent-induced gastrointestinal lesions and adjuvant arthritis in rats

Besides a superior protection of the pentadecapeptide BPC 157 (an essential fragment of an organoprotective gastric juice peptide BPC) against different gastrointestinal and liver lesions, an acute anti-inflammatory and analgetic activity was also noted. Consequently, its effect on chronic inflammation lesions, such as adjuvant arthritis, and non-steroidal anti-inflammatory agents (NSAIAs)-induced gastrointestinal lesions was simultaneously studied in rats. In gastrointestinal lesions (indomethacin (30 mg/kg s.c.), aspirin (400 mg/kg i.g.) and diclofenac (125 mg/kg i.p.) studies, BPC 157 (10 micrograms or 10 ng/kg i.p.) was regularly given simultaneously and/or 1 h prior to drug application (indomethacin). In the adjuvant arthritis (tail-application of 0.2 mL of Freund's adjuvant) studies (14 days, 30 days, 1 year) BPC 157 (10 micrograms or 10 ng/kg i.p.), it was given as a single application (at 1 h either before or following the application of Freund's adjuvant) or in a once daily regimen (0-14th day, 14-30th day, 14th day-1 year). Given with the investigated NSAIAs, BPC 157 consistently reduced the otherwise prominent lesions in the stomach of the control rats, as well as the lesions in the small intestine in the indomethacin groups. In the adjuvant arthritis studies, the lesion's development seems to be considerably reduced after single pentadecapeptide medication, and even more attenuated in rats daily treated with BPC 157. As a therapy of already established adjuvant arthritis, its salutary effect consistently appeared already after 2 weeks of medication and it could be clearly seen also after 1 year of application. Taking together all these results, the data likely point to a special anti-inflammatory and mucosal integrity protective effect.     

Experimental studies on the growth of cultured epithelium and endothelium of rabbit cornea exposed to low temperature

Three strains of Mycoplasma synoviae (MS) (Olson-WVU 1853, reported in 1956; and Massachusetts 9895 and 5044, respectively isolated in 1957 and 1972) were used for experimental inoculation of chickens to evaluate the various mycoplasma serologic tests. The MS plate agglutination and hemagglutination-inhibition (HI) tests were highly sensitive for detection of infection; reactions persisted for test periods extending to 63 weeks. Correlation between MS HI-tube and micro methods was good. HI titers were highest in sera from birds inoculated with strain 9895. All three strains produced cross-reactions with M. Gallisepticum (MG) plate antigens, which were detectable for 2 to 12 weeks following MS inoculation; strain 5044 produced the lowest percentage of cross-reactions. MG HI titers of 16 were observed occasionally in sera from all three group inoculated with strain 1853. It appears from these limited serologic studies in chickens that there may be antigenic differences among the three MS strains.     

Human immune responses to influenza virus vaccines administered by systemic or mucosal routes

Healthy adult volunteers were immunized by parenteral or oral routes with trivalent inactivated influenza vaccine (A/Chile/1/83 (H1N1), A/Mississippi/1/85 (H3N2), and B/Ann Arbor/1/86), or intranasally with live attenuated, cold-adapted influenza type A/Texas/1/85 (H1N1) reassortant virus. In all volunteers, cells spontaneously secreting IgA, IgG or IgM antibodies specific to influenza virus were detected in peripheral blood on days 6-13 after immunization, and specific IgA, IgG and IgM antibodies to influenza vaccine were measured in sera and external secretions (saliva and nasal lavage). Following systemic immunization, a raise in specific antibodies of all isotypes was observed in sera beginning on day 13. Although small variations in IgA and IgM antibodies in saliva and nasal lavages were detected, antigen-specific IgG significantly increased between days 13 and 27. Intranasal administration of attenuated virus induced IgA and IgG antibodies in serum as well as in secretions. Serum antibodies were not substantially influenced by oral immunization, only a small increase in all isotypes was observed in volunteers' sera 21 days after ingestion of vaccine. However, in secretions, antigen-specific IgA and IgG responses were detected one week after immunization and reached a peak response on day 20. These studies show that different routes of immunization can be effective for the induction of specific antibodies, and support the concept of the common mucosal immune system in humans by demonstrating that the oral or intranasal administration of antigen-induced specific antibodies of IgA isotype in external secretions, preceded by the transient appearance in peripheral blood of specific antibody-producing cells.     

Dimethoxyphenylethylamine and tetrahydropapaverine are toxic to the nigrostriatal system

Experimental allergic neuritis (EAN) is a T cell mediated animal model of Guillain-Barré syndrome, characterized by inflammation and demyelination of the peripheral nervous system (PNS). To study the involvement of immunoregulatory cytokines, we induced EAN in Lewis rats by immunizing with bovine PNS myelin (BPM) and Freund's complete adjuvant. mRNA expression of the cytokines IL-1beta, IL-6, IL-10, IL-12, TNF-alpha and TNF-beta, and the cytolytic effector molecule cytolysin was examined in lymph node mononuclear cells (MNC) over the course of EAN by in situ hybridization after culture without antigen and in the presence of BPM, the myelin P2 protein, the control antigen acetylcholine receptor, or the mitogen PHA. Three patterns of cytokine mRNA expressing MNC in relation to clinical EAN could be distinguished: (i) IL-1beta mRNA expressing cells peaked already on day 3 post immunization (p.i.), and BPM- and P2-reactive TNF-alpha, and BPM-reactive IL-6 mRNA expressing cells were also detected already on day 7 p.i., i.e., before onset of clinical EAN; (ii) BPM- and P2-reactive TNF-alpha peaked together with P2-reactive TNF-beta, IL-6 and IL-12 mRNA expressing cells at height of clinical EAN, consistent with a disease-promoting role for these four cytokines; (iii) high levels of BPM- and P2-reactive IL-10 and cytolysin mRNA expressing cells were observed only during recovery (day 28 p.i.), consistent with a disease down-regulating role of IL-10 and cytolysin. The results suggest a major proinflammatory role for IL-1beta, TNF-alpha, TNF-beta, IL-6 and IL-12 and a disease down-regulating function of IL-10 as well as cytolysin in EAN.     

Twenty-four-hour rhythms in immune responses in rat submaxillary lymph nodes and spleen: effect of cyclosporine

Twenty-four-hour variations in cellularity, lipopolysaccharide (LPS)- and concanavalin A (Con A)-induced cell proliferation, and natural killer (NK) activity were examined in submaxillary lymph nodes and spleen of rats injected with Freund's complete adjuvant or its vehicle and kept under light from 08:00 to 20:00 h daily. A significant daily variation in cellularity was detected, exhibiting maxima at 09:00 h in submaxillary lymph nodes (nonimmunized and immunized rats) and at 13:00 h in spleen (immunized rats only). Submaxillary lymph node LPS- and Con A-mitogenic effect displayed maximal activity during daytime (peak at 13:00-17:00 h). In spleen, the maxima for 24-h rhythm in LPS-induced cell proliferation and NK activity occurred at midnight and at early morning (09:00 h), respectively. Con A-induced spleen cell proliferation peaked at midday in nonimmunized rats only. Injection of the immunosuppressive drug cyclosporine decreased Freund's adjuvant-induced augmentation of LPS and Con A mitogenic effect in both tissues and diminished spleen cell number. Cyclosporine blunted circadian rhythms in submaxillary lymph node Con A response and cell number, while it shifted the maximum in LPS effect to peak at 01:00 h. Cyclosporine also suppressed the circadian changes in LPS- and Con A-induced spleen cell proliferation, but not those found in NK activity. The results indicate the existence of 24-h rhythms in immune responses of rat submaxillary lymph nodes and spleen with maxima at different times of the day and that were significantly affected by cyclosporine injection.     

Induction of systemic and mucosal immune responses to human immunodeficiency virus type 1 by a DNA vaccine formulated with QS-21 saponin adjuvant via intramuscular and intranasal routes

Induction of mucosal and cell-mediated immunity is critical for development of an effective vaccine against human immunodeficiency virus (HIV). We compared intramuscular and intranasal immunizations with a DNA vaccine encoding env of HIV-1 and evaluated the QS-21 saponin adjuvant for augmentation of the systemic and mucosal immune responses to HIV-1 in a murine model. Vaccination via the two routes elicited comparable systemic immune responses, and QS-21 consistently enhanced antigen-specific serum immunoglobulin G2a (IgG2a) production, delayed-type hypersensitivity reaction, and cytolytic activity of splenocytes. Intestinal secretory IgA production and cytolytic activity of the mesenteric lymph node cells are preferentially elicited by intranasal immunization, and QS-21 augmented these activities as well. This adjuvant augmented production of interleukin-2 (IL-2) and gamma interferon (IFN-gamma) associated with decrease in IL-4 synthesis by antigen-restimulated splenocytes. The serum immunoglobulin subtype profile showed a dominant IgG2a response and less strong IgG1 and IgE production in a QS-21 dose-dependent manner. As expected, enhancements of humoral and cell-mediated immune responses by QS-21 were abrogated by treatment with anti-IL-2 and anti-IFN-gamma monoclonal antibodies. These results suggest that the intranasal route of DNA immunization is more efficient than the intramuscular route in inducing mucosal immunity mediated by sIgA and mesenteric lymphocytes. Furthermore, QS-21 is able to act as a mucosal adjuvant in DNA vaccination and demonstrates its immunomodulatory property via stimulation of the Th1 subset. This study emphasizes the importance of the route of immunization and the use of an adjuvant for effective DNA vaccination against HIV-1.     

Epitope polarity and adjuvants influence the fine specificity of the humoral response against Semliki Forest virus specific peptide vaccines

The humoral response to synthetic peptide vaccines against Semliki Forest virus (SFV) in H-2d BALB/c mice was investigated with the enzyme linked immunosorbent assay and the pepscan technique. The peptide vaccines consisted of amino acid sequences 240-255 (B) and 137-151 (T) of the E2 membrane protein of SFV colinearly synthesized in either orientation T-B or B-T. Sequence B contains an epitope inducing humoral immunity to lethal SFV infection and sequence T contains a H-2d restricted T-helper cell epitope. With sera from mice immunized subcutaneously with peptide T-B, and Quil A as adjuvant, two immunodominant B-cell epitopes were identified, FVPRAD, at position 240-246 and PHYGKEI, at position 145-151. However, with peptide B-T and Quil A as adjuvant for immunization the epitope PHYGKEI was clearly immunodominant, but antibodies elicited against this epitope were not reactive with SFV-infected L cells in contrast to the antibodies elicited by epitope FVPRAD. An additional epitope EPARKGKVH, at position 247-255, was identified with sera from mice immunized subcutaneously with either peptide T-B or B-T and Montanide ISA 740 as an adjuvant. Monoclonal antibodies selected for reactivity with SFV-infected L cells did bind also to epitope FVPRAD. Interestingly, this epitope could induce antibodies cross-reactive with a synthetic peptide derived from macrophage migration inhibitory factor that shares amino acid residues VPRA at position 9-12 with the protective B-cell epitope FVPRAD. The present study shows clearly that the fine specificity of the humoral response against peptide vaccines is differentially influenced by both adjuvant and epitope polarity which may affect vaccine efficacy. Further, the study reminds us that potentially autoimmune antibodies could be induced by vaccines.