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1.
Oral administration of F4 fimbriae of Escherichia coli induces intestinal mucosal immune responses in F4 receptor-positive (F4R(+)) pigs, but not in F4R(-) pigs. We examined whether F4 fimbriae in F4R(-) animals behave like a food antigen and can induce oral tolerance. Therefore, F4R(+) and F4R(-) pigs were fed 2mg of F4 and challenged i.m. to evaluate the effect of oral F4 on the systemic immune system. As control antigen, two different oral doses (2 and 600 mg) of OVA were used. Thirty days after the i.m. OVA challenge, the OVA-specific serum IgG titre in 600 mg-fed pigs was lower than that in non-fed animals, indicating that tolerance was induced. Conversely, in the 2mg-fed pigs a rapid increase of OVA-specific IgG with higher titres than those in non-fed pigs was seen following challenge, indicating a priming of the systemic immune system. A similar priming was seen in both F4-fed F4R(-) and F4R(+) pigs. Upon challenge, non-fed pigs displayed a primary immune response with a slow increase of F4-specific serum IgG, whereas F4-fed F4R(-) and F4R(+) pigs showed secondary responses with a rapid increase of serum IgG. This was expected in F4R(+) pigs, as in these animals oral F4 induces F4-specific antibody-secreting cells in the spleen, suggesting a priming of the systemic immune system. However, also the F4-fed F4R(-) pigs displayed a secondary response, despite the failure to detect a response upon oral F4 administration. These findings suggest that the F4 antigen, at a dose of 2 mg, behaves like a common food antigen in F4R(-) pigs, namely it induces a systemic priming.  相似文献   

2.
Oral immunization of both humans and animals with non-replicating soluble antigens often results in the induction of oral tolerance. However, receptor-dependent uptake of orally administered soluble antigens can lead to the induction of an antigen-specific immune response. Indeed, oral immunization of pigs with recombinant FaeG (rFaeG), the adhesin of the F4(K88) fimbriae of enterotoxigenic Escherichia coli (ETEC), induces an F4-specific humoral and cellular immune response. This response is accompanied with a reduction in the excretion of F4(+)E. coli following challenge. To improve the immune response against F4, rFaeG was orally co-administered with the mucosal adjuvant cholera toxin (CT). Oral immunization of pigs with rFaeG and CT significantly improved the induction of an F4-specific humoral and cellular immune response and also significantly reduced the faecal F4(+)E. coli excretion following F4(+) ETEC challenge as compared to rFaeG-immunized pigs. Therefore, the present study demonstrates that CT can act in pigs as a mucosal adjuvant for antigens that bind to the intestinal epithelium by a CT-receptor-independent mechanism.  相似文献   

3.
Very few antigens have been described that induce an intestinal immunity when given orally. Our laboratory demonstrated that oral administration of isolated F4 (K88) fimbriae of Escherichia coli to F4-receptor positive (F4R(+)) pigs induces protective mucosal immunity against challenge infection. However, presence of F4-receptors (F4R) on villous enterocytes is a prerequisite for inducing the immune response, as no F4-specific antibody-secreting cells (ASC) can be induced in F4R(-) pigs. In this study, the in vivo binding of isolated F4 fimbriae (F4) to the gut epithelium was examined in F4R(+) and F4R(-) pigs. It was further investigated whether binding of F4 to the F4R results in endocytosis in and translocation across the gut epithelium using microscopy. F4 did not adhere to the intestinal epithelium of F4R(-) pigs, whereas it strongly adhered to the villous epithelium and the follicle-associated epithelium (FAE) of the jejunum and ileum of F4R(+) pigs. Following binding to F4R, F4 was endocytosed by villous enterocytes, follicle-associated enterocytes and M cells. Transcytosis of F4 across the epithelium resulted in the appearance of F4 in the lamina propria and dome region of the jejunal and ileal PP. This is the first study showing transcytosis of fimbriae across the gut epithelium. This receptor-dependent transcytosis can explain the success of F4 fimbriae as oral immunogen for inducing protective immunity in F4R(+) pigs strengthening the importance of receptor-dependent endocytosis and translocation in oral vaccine strategies. Further identification of the receptor responsible for this transport is in progress.  相似文献   

4.
F4 enterotoxigenic Escherichia coli (ETEC) cause diarrhoea and mortality in piglets leading to severe economic losses. Oral immunization of piglets with F4 fimbriae induces a protective intestinal immune response evidenced by an F4-specific serum and intestinal IgA response. However, successful oral immunization of pigs with F4 fimbriae in the presence of maternal immunity has not been demonstrated yet. In the present study we aimed to evaluate the effect of maternal immunity on the induction of a systemic immune response upon oral immunization of piglets. Whereas F4-specific IgG and IgA could be induced by oral immunization of pigs without maternal antibodies and by intramuscular immunization of pigs with maternal antibodies, no such response was seen in the orally immunized animals with maternal antibodies. Since maternal antibodies can mask an antibody response, we also looked by ELIspot assays for circulating F4-specific antibody secreting cells (ASCs). Enumerating the F4-specific ASCs within the circulating peripheral blood mononuclear cells, and the number of F4-specific IgA ASCs within the circulating IgA+ B-cells revealed an F4-specific immune response in the orally immunized animals with maternal antibodies. Interestingly, results suggest a more robust IgA booster response by oral immunization of pigs with than without maternal antibodies. These results demonstrate that oral immunization of piglets with F4-specific maternal antibodies is feasible and that these maternal antibodies seem to enhance the secondary systemic immune response. Furthermore, our ELIspot assay on enriched IgA+ B-cells could be used as a screening procedure to optimize mucosal immunization protocols in pigs with maternal immunity.  相似文献   

5.
Enteric viruses are a major cause of diarrhea in animals and humans. Among them, rotaviruses are one of the most important causes of diarrhea in young animals and human infants. A lack of understanding of mechanisms to induce intestinal immunity and the correlates of protective immunity in neonates has impaired development of safe and effective vaccines against enteric viruses. Studies of candidate vaccines using an adult mouse model of subclinical enteric viral infections often do not predict vaccine efficacy against disease evaluated in neonatal large animals. A series of studies have been conducted using a neonatal gnotobiotic pig model of rotavirus infection and diarrhea to identify correlates of protective immunity and to evaluate traditional and novel vaccine approaches for the induction of mucosal immune responses and protection to enteric viruses. Gnotobiotic pigs recovered from infection with virulent Wa human rotavirus (HRV) (mimic natural infection) had high numbers of intestinal IgA rotavirus-specific primary antibody-secreting cells (ASCs) and memory B-cells (to recall antigen) measured by ELISPOT assay, which correlated with complete protection against rotavirus challenge. Most short-term IgA memory B-cells were resident in the ileum, the major site of rotavirus replication. Spleen, not the bone marrow, was the major resident site for longer-term IgG memory B-cells. Candidate rotavirus vaccines evaluated in pigs for their ability to induce intestinal or systemic ASC and protection against rotavirus infection and diarrhea included attenuated live virus, inactivated virus, and baculovirus-expressed double-layered rotavirus-like particles (2/6-VLPs). In combination with those candidate vaccines, various adjuvants, delivery systems, and immunization routes were tested, including incomplete Freund's adjuvant for i.m. immunization, and a mutant Escherichia coli heat labile enterotoxin R192G (mLT) for i.n. immunization. It was shown that orally administered replicating vaccines were most effective for priming for intestinal IgA ASC and memory B-cell responses, but i.n. administered non-replicating 2/6-VLPs plus mLT were effective as booster vaccines. We conclude that protective immunity depends on the magnitude, location, viral protein-specificity, and isotype of the antibody responses induced by vaccination. Therefore highly effective enteric viral vaccines should: (i) induce sufficient levels of intestinal IgA antibodies; (ii) include viral antigens that induce neutralizing antibodies; and (iii) require the use of effective mucosal adjuvants or antigen delivery systems for non-replicating oral or i.n. vaccines.  相似文献   

6.
Four- to 8-week-old SPF pigs were immunized, using antigens of Ascaris suum incorporated into liposomes, via intestinal cannula or orally. Avridine was also incorporated in the liposomes in one experiment and interleukin-2 (IL-2) injected into pigs in another experiment. A priming dose of embryonate eggs (80-470 eggs/pig) were given in four of six experiments. Compared to control animals, the greatest protection of pigs to migrating ascarid larvae from a challenge dose of 10,000 embryonated eggs occurred where pigs received (1) a priming dose of eggs plus second-stage ascarid larval wall incorporated into liposomes, with or without avridine or IL-2, or (2) a priming dose of eggs plus ascarid intestinal aminopeptidase incorporated into liposomes with IL-2. The degree of protection was not statistically significant due, in part, to the variability in the responses of animals in the same treatment groups and the small number of animals per group. In general, only low titers of specific serum antibodies were detected and specific antibodies were not detected in the intestinal washing.  相似文献   

7.
Oral immunization is the most effective way of inducing immune responses in the intestinal tract. Biodegradable microspheres have been used extensively for the delivery of antigens to the Peyer's patches (PPs) within the gut-associated lymphoid tissue (GALT). We evaluated various formulations of alginate microspheres for their capacity to induce mucosal immune responses in vivo. Multiple intestinal "loops" each containing a single PP, were surgically prepared in lambs. We have previously showed that PP in individual intestinal loops function as independent sites for the induction of immune responses. This animal model provides a system for directly comparing different antigen formulations within the same animal. Individual intestinal loops were injected with a model antigen, porcine serum albumin (PSA) encapsulated in three different formulations of alginate micropsheres. Three weeks after immunization, PSA-specific immune responses were assayed with antibody secreting cell (ASC) ELISPOT, lymphocyte proliferative responses (LPRs), IFN-gamma production and antibody secreted into intestinal loops. PSA encapsulated in alginate micropsheres or in saline induced humoral immune responses as indicated by the presence of numerous ASC. However, PSA-specific T-cell responses (LPR and IFN-gamma production) were not induced.  相似文献   

8.
Mucosal immunization is advantageous over other routes of antigen delivery because it can induce both mucosal and systemic immune responses. In this study, we have developed fimbriae protein of enterotoxigenic Escherichia coli (ETEC) F41 was stably expressed on the surface Lactobacillus casei 525. The method of expressing vaccine antigens in L. casei induces both systemic and mucosal immunity after oral or intranasal administration. We demonstrate that an oral or intranasal vaccine based on live recombinant L. casei 525 protects infant mice from ETEC F41 infection. This platform technology can be applied to design oral or intranasal vaccine delivery vehicles against several microbial pathogens.  相似文献   

9.
We previously showed that an intradermal (i.d.) FaeG DNA prime (2x)-oral F4 protein boost immunization induces a systemic response and weakly primes a mucosal IgG response in pigs, especially when plasmid vectors encoding the A and B subunit of the E. coli thermo-labile enterotoxin (LT) are added to the DNA vaccine. In the present study, we evaluated whether addition of 1alpha,25-dihydroxyvitamin D(3) (vitD(3)) to the DNA vaccine could further enhance this mucosal priming and/or modulate the antibody response towards IgA. To further clarify priming of systemic and mucosal responses by the i.d. DNA vaccination, we firstly compared the localization of the F4-specific antibody response in pigs that were orally boosted with F4 to that in pigs that received a third i.d. DNA immunization and secondly evaluated cytokine mRNA expression profiles after i.d. DNA vaccination. The i.d. DNA prime (2x)-oral F4 boost immunization as well as the 3 i.d. DNA vaccinations induced mainly a systemic response, with a higher response observed following the heterologous protocol. Co-administration of vitD(3), and especially of the LT vectors, enhanced this response. Furthermore, only the heterologous immunization resulted in a weak mucosal priming, which appeared to require the presence of the LT vectors or vitD(3) as adjuvants. In addition, the LT vectors strongly enhanced the FaeG-specific lymphocyte proliferation and this was accompanied by the absence of a clear IL-10 response. However, despite two DNA immunizations in the presence of these adjuvants and an oral F4 boost, we failed to demonstrate the secretory IgA response needed to be protective against enterotoxigenic E. coli.  相似文献   

10.
Recent findings demonstrate that priming by levamisole of weaned pigs experimentally vaccinated against postweaning colibacillosis (PWC) contributes to immune protection from challenge-induced clinical disease through stimulation of the mesenteric lymph node cells that participate in cell-mediated immunity. With the objective of better understanding the mechanisms by which levamisole induces protective mucosal cell-mediated immune response to vaccination against PWC, it was tested whether the drug synergizes experimental F4ac+ Escherichia coli oral vaccine in stimulating T cells also in the jejunal lamina propria (JLP) and ileal Peyer's patch (IPP) upon virulent challenge. Commercial crossbred pigs weaned at 4 weeks were allocated into two equal groups. The experimental group was i.m. primed with levamisole at an immunostimulatory dose of 2.5 mg/kg once daily, for 3 consecutive days, and controls received saline. Both groups were vaccinated orally with the vaccinal E. coli strain on day 0 and challenged with the virulent E. coli strain 7 days later. All pigs were killed on postchallenge day 6. The results determined by immunophenotyping of isolated cells indicate that priming by levamisole of the vaccinated weaned pigs selectively recruited and activated T cells in the IPP, a lymphoid organ-generating B lymphocytes. The pig IPP is normally populated with up to 5% of CD3+ T cells and CD6 is an activation antigen expressed exclusively by T cells in swine. Therefore, a significantly higher number of CD3+ (P < 0.01) and CD6+ (P < 0.001) cells observed within the IPP follicles of the primed-vaccinated vs. unprimed-vaccinated challenge-infected pigs suggest enhanced T cell-mediated immunity in this B-cell compartment induced by the potentiating action of the drug and vaccine. The ability of levamisole to influence interaction between activated T cells and B cells in the IPP of primed-vaccinated weaned pigs, and the possibility that this interaction plays a role in regulating B-cell maturation within the IPP follicles, are discussed.  相似文献   

11.
五种旋毛虫抗原对猪的免疫保护作用研究   总被引：4，自引：0，他引：4  
本实验研究了旋毛虫肌幼虫可溶性粗抗原、排泄分泌抗原（ＥＳ）、表面抗原（ＳＡ）及成虫ＥＳ、ＳＡ５种抗原对猪的免疫保护作用。结果５种抗原对猪均具有一定程度的免疫原性，可诱导猪体产生对攻击感染的抵抗力（减虫率），其中肌幼虫粗抗原为５５．２０％；肌幼虫ＥＳ为４２．５６％，肌幼虫ＳＡ为７２．２１％；成虫ＥＳ为３２．９２％；成虫ＳＡ为４２．１７％。免疫５种抗原后用肌幼虫“Ｂ”抗原、新生幼虫可溶性抗原及成虫可溶性抗原进行ＥＬＩＳＡ检测，均可测出血清抗体应答反应，其中以相应抗原测出的抗体应答较强烈。免疫５种抗原后猪外周血液中Ｂ淋巴细胞减少，Ｔｈ及Ｔｓ增加，Ｔｈ／Ｔｓ比值降低，呈暂时的细胞免疫抑制现象。  相似文献   

12.
The study asks whether, in fish, antigens encountered early in life can prime the immune system to yield memory responses on subsequent challenge with the same antigen and, if so, whether positive immunity or immunological tolerance is induced. The direct immersion method of vaccination was used to prime 4 week old carp, Cyprinus carpio, and was compared with priming by injection. Three different forms of antigen were used: the thymus dependent antigen, human gamma globulin (HGG) in soluble and in particulate (latex bound) form; also the putative thymus independent bacterin, formalin-killed Aeromonas salmonicida. The thymus dependent antigens were also used on 9 month old animals. In 4 week old carp, A. salmonicida vaccine delivered either by direct immersion or intraperitoneally (i.p.) yielded enhanced serum antibody levels and heightened proliferative responses in the lymphoid tissue of the spleen and kidney. Latex-bound HGG applied by direct immersion was found to partially suppress secondary antibody production while still eliciting enhanced proliferation. The decrease in antibody production following direct immersion priming of young fish with latex-bound HGG was not nearly as marked as the tolerance induced following priming with latex-bound HGG by the i.p. route and, unlike the tolerance induced by the injection route, may possibly still occur in older fish. When HGG was applied to young carp in soluble form by direct immersion it was ineffective and failed to influence memory induction. This is in contrast to the antibody tolerance, accompanied by an enhanced proliferative response following challenge, which resulted from administration of the soluble antigen by injection in the young fish. The status of the immune system in these antibody-tolerant fish is still far from clear. This highlights the need for further investigation of the role of cell-mediated reactions and local immunity in the immune responses of fish.  相似文献   

13.
The mucosal immune system is exposed to a range of antigens associated with pathogens, to which it must mount active immune responses. However, it is also exposed to a large number of harmless antigens associated with food and with commensal microbial flora, to which expression of active, inflammatory immune responses to these antigens is undesirable. The mucosal immune system must contain machinery capable of evaluating the antigens to which it is exposed and mounting appropriate effector or regulatory responses. Since the immune system is likely to have evolved initially in mucosal tissues, the requirement to prevent damaging allergic responses must be at least as old as the adaptive immune system, and studies of the mechanisms should include a range of non-mammalian species. Despite the importance for rational design of vaccines and for control of allergic reactions, the mechanisms involved are still largely unclear. It is not clear that the classical experimental protocol of "oral tolerance" is, in fact, measuring a biologically important phenomenon, nor is it clear whether tolerance is regulated in the evolutionarily recent organised lymphoid tissue (the lymph nodes) or the more ancient, diffuse architecture in the intestine. The capacity of the immune system to discriminate between "dangerous" and "harmless" antigens appears to develop with age and exposure to microbial flora. Thus, the ability of an individual or a group of animals to correctly regulate mucosal immune responses will depend on age, genetics and on their microbial environment and history. Attempts to manipulate the mucosal immune system towards active immune responses by oral vaccines, or towards oral tolerance, are likely to be confounded by environmentally-induced variability between individuals and between groups of animals.  相似文献   

14.
Porcine postweaning Escherichia coli enteritis is a cause of significant morbidity and mortality in pigs worldwide, and effective prevention remains an unsolved problem. This study examined the correlation between susceptibility of pigs to experimental infection with an E. coli F18 strain and the porcine intestinal F18 receptor genotypes. Thirty-one pigs classified as either belonging to the susceptible or the resistant genotype were inoculated with cultures of an E. coli O138:F18 isolated from a pig with postweaning diarrhoea. Susceptibility to colonisation and diarrhoea was assessed by clinical observations, faecal shedding of the challenge strain, histopathology and microscopic adhesion tests. Ten of 14 (71.4%) genetically susceptible pigs and one of 17 (5.9%) resistant pigs developed diarrhoea attributable to the challenge strain. There was no difference in susceptibility between homozygotic and heterozygotic susceptible pigs. Faecal shedding of the challenge strain correlated with the genetic receptor profile. Twenty pigs examined immunohistochemically revealed focal to extensive small intestinal mucosal colonisation by E. coli O138:F18 in nine of 10 susceptible and three of 10 resistant pigs. Results of in vitro adhesion assays performed with F18 cells on enterocyte preparations from 24 pigs, showed complete concordance with the F18 genotypes. In conclusion, this study showed a high correlation between the porcine intestinal F18 receptor genotypes and susceptibility to disease. However, pigs of the resistant F18 receptor genotype were not entirely protected against intestinal colonisation by E. coli F18.  相似文献   

15.
The study examined the effects of stressors on the responses of 3 and a half-week old piglets that had been given an oral dose of enterotoxigenic Escherichia coli (ETEC) and a novel harmless antigen (ovalbumin). Removal from the sow (WEAN), a short-term cold stressor (12;C for 48 hours) (TEMP) and mixing with non-littermates (MIX) were assessed in terms of the effects on faecal shedding of ETEC, immune responses, weight gain and an ACTH stimulation test. WEAN and TEMP reduced weight gain and all stressors increased faecal shedding of ETEC. All stressors increased the IgG responses to F4(K88)ac antigens and WEAN and TEMP increased the IgA responses to the same antigens, probably as a result of increased intestinal proliferation of ETEC. None of the stressors, however, had significant effects on antibody responses to ovalbumin or on lymphocyte proliferation assays. The results indicate that stressors influence the faecal shedding of ETEC in young piglets by a mechanism that may not involve modulation of immune responses.  相似文献   

16.
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17.
The allograft inflammatory factor (AIF-1/daintain) is a hormone-like peptide produced by activated monocytic cells in a variety of traumatic, inflammatory and degenerative lesions. Gut-derived AIF-1 has been shown to modulate insulin production and to attenuate autoimmune diabetes. As the localization of this gastrointestinal peptide in the porcine duodenum is not known and the pig is a convenient model for the study of nutritional modulation of the mucosal immune compartment, we have localized expression of AIF-1 by immunohistology in the duodenum of either malnourished (energy and protein supply 50% of demands, n = 5) or optimally fed pigs (n = 5). AIF-1 macrophages were predominantly located at the villus tip. The number of positively stained cells per high-power field was significantly (P < or = 0.001) higher in the malnourished pigs (74.6 +/- 2.44; least square means +/- SEM) compared to optimally fed pigs (32.56 +/- 1.99). It is likely that the effect in malnourished pigs can be explained by a more pronounced antigen contact of macrophages due to loss of epithelial integrity. Thus, AIF-1 is a novel marker for the study of the nutritional regulation of the mucosal immune system of the pig. AIF-1 expression in the duodenum was further validated by polymerase chain reaction and sequencing. Surprisingly, we detected a slight deviation from the original sequence (probably representing an allelic variation) and an AIF-1 splice variant, previously not known to occur in pigs.  相似文献   

18.
Due to almost identical lipopolysaccharide (LPS) O-antigens, infections with Yersinia enterocolitica serotype O:9 (YeO:9) cause false positive serological reactions (FPSR) in tests for Brucella and thus cause problems in National Brucella surveillance programs. As LPS are strong inducers of antibody responses it was hypothesized that cell-mediated immune responses to non-LPS antigens of the two bacteria can be used to separate immune responses to these two biologically very different infections. Following subclinical experimental infections with Brucella suis biovar 2, high interferon-gamma (IFN-gamma) assay responses with a commercial Brucella melitensis antigen preparation (Brucellergene OCB) preceded the development of antibodies. High IFN-gamma responses in the seven B. suis inoculated pigs with serological evidence of infection were consistent throughout a 20-week post-inoculation observation period. In contrast, IFN-gamma responses in two B. suis inoculated pigs without bacteriological or serological evidence of infection were below a cut-point of 25pg/ml at all samplings. IFN-gamma responses in repeated samplings from 5 uninfected control pigs and 18 pigs experimentally infected with YeO:9 were all negative, except for solitary false positives in 3.7% of the samples from both the experimentally YeO:9 infected pigs and control pigs. Skin tests using the same commercial Brucella antigen confirmed the ability of cell-mediated immune responses to differentiate between the two infections. In addition, a field evaluation of the diagnostic use of cell-mediated immune responses by IFN-gamma assay and skin test to resolve serological suspicions of Brucella was conducted in an YeO:9 infected pig herd. Following a screening of 200 pigs 39 pigs were identified with false positive serological Brucellosis reactions. While 36 of the 39 FPSR pigs were also FPSR in a second test, none of the pigs were test positive in whole blood IFN-gamma assay or Brucellergene OCB skin test. In conclusion, use of IFN-gamma assay and skin test as measurements of cell-mediated immune responses to non-LPS Brucella antigens were specific and sensitive in discriminating subclinical experimental infections with B. suis from both natural and experimental infections with YeO:9.  相似文献   

19.
The intestinal and mammary immune system in pigs   总被引：1，自引：0，他引：1  
Evidence exists from studies in other species for a link between the intestinal and mammary immune systems. This was examined in pigs by various methods including analysis of lymphocyte subsets in intestinal and mammary tissues and lymphocyte migration studies. It was concluded that in the pig both a common mucosal immune response and a genuine local immune response exist in the mammary gland.  相似文献   

20.
Enterotoxigenic and verotoxigenic F18+ Escherichia coli colonising the pig small intestine, adhere to receptors on intestinal villous enterocytes by F18 fimbriae. The aim of the present study was to define the F18R nature. The knowledge on the nature of this receptor could be important for the development of receptor-based treatments against F18+ E. coli-induced disease. The adhesion of F18+ E. coli to pig intestinal villous enterocytes was analysed in an in vitro assay. The adhesion of F18+ E. coli but not of F4ac+ E. coli was strongly inhibited by monoclonal antibodies (mAb) with blood group H-2 specificity. Conversely, blood group H-1 specific mAb could not inhibit the adhesion of F18+ E. coli nor F4ac+ E. coli. Moreover, the blood group H-2 trisaccharide strongly inhibited the adhesion of F18+ E. coli, but only partially the adhesion of F4ac+ E. coli. These data demonstrate that the F18 receptor contains the blood group antigen H-2 (-fuc-(1-2)-β-Gal-(1-4)-GlcNAc) as major carbohydrate.  相似文献   

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