Payvand Parvizi

MODULATION OF ANTIBODY-MEDIATED IMMUNE RESPONSE BY PROBIOTICS IN CHICKENS

ABSTRACT Probiotic bacteria, including Lactobacillus acidophilus and Bifidobacterium bifidum, have been shown to enhance antibody responses in mammals. The objective of this study was to examine the effects of a probiotic product containing the above bacteria in addition to Streptococcus faecalis on the induction of the chicken antibody response to various antigens, both systemically and in the gut. The birds received probiotics via oral gavage and subsequently were immunized with sheep red blood cells (SRBC) and bovine serum albumin (BSA) to evaluate antibody responses in serum or with tetanus toxoid (TT) to measure the mucosal antibody response in gut contents. Control groups received phosphate-buffered saline. Overall, BSA and SRBC induced a detectable antibody response as early as week 1 postimmunization (p.i.), which lasted until week 3 p.i. Probiotic-treated birds had significantly (P ≤ 0.001) more serum antibody (predominantly immunoglobulin M [IgM]) to SRBC than the birds that were not treated with probiotics. However, treatment with probiotics did not enhance the serum IgM and IgG antibody responses to BSA. Immunization with TT resulted in the presence of specific IgA and IgG antibody responses in the gut. Again, treatment with probiotics did not change the level or duration of the antibody response in the gut. In conclusion, probiotics enhance the systemic antibody response to some antigens in chickens, but it remains to be seen whether probiotics have an effect on the generation of the mucosal antibody response.

Effects of lactobacilli on cytokine expression by chicken spleen and cecal tonsil cells.

ABSTRACT Lactobacillus acidophilus, Lactobacillus reuteri, and Lactobacillus salivarius are all normal residents of the chicken gastrointestinal tract. Given the interest in using probiotic bacteria in chicken production and the important role of the microbiota in the development and regulation of the host immune system, the objective of the current study was to examine the differential effects of these bacteria on cytokine gene expression profiles of lymphoid tissue cells. Mononuclear cells isolated from cecal tonsils and spleens of chickens were cocultured with one of the three live bacteria, and gene expression was analyzed via real-time quantitative PCR. All three lactobacilli induced significantly more interleukin 1β (IL-1β) expression in spleen cells than in cecal tonsil cells, indicating a more inflammatory response in the spleen than in cecal tonsils. In cecal tonsil cells, substantial differences were found among strains in the capacity to induce IL-12p40, IL-10, IL-18, transforming growth factor β4 (TGF-β4), and gamma interferon (IFN-γ). In conclusion, we demonstrated that L. acidophilus is more effective at inducing T-helper-1 cytokines while L. salivarius induces a more anti-inflammatory response.

Oral Treatment of Chickens with Lactobacilli Influences Elicitation of Immune Responses

ABSTRACT Commensal microbes in the intestine are in constant interaction with host cells and play a role in shaping the immune system. Lactobacillus acidophilus, Lactobacillus reuteri, and Lactobacillus salivarius are members of the chicken intestinal microbiota and have been shown to induce different cytokine profiles in mononuclear cells in vitro. The objective of the present study was to examine the effects of these bacteria individually or in combination on the induction of antibody- and cell-mediated immune responses in vivo. The birds received lactobacilli weekly via oral gavage starting on day of hatch and subsequently, at 14 and 21 days, were immunized with sheep red blood cells (SRBC), keyhole limpet hemocyanin (KLH), Newcastle disease virus vaccine, and infectious bursal disease virus vaccine. Antibody responses in serum were measured weekly for 4 weeks beginning on the day of primary immunization. The cell-mediated immune response was evaluated at 21 days postimmunization by measurement of gamma interferon (IFN-γ) production in splenocytes stimulated with inactivated vaccine antigens. L. salivarius-treated birds had significantly more serum antibody to SRBC and KLH than birds that were not treated with probiotics. L. salivarius-treated birds also had decreased cell-mediated immune responses to recall antigen stimulation. L. reuteri treatment did not significantly affect the systemic immune response, while L. acidophilus treatment increased the antibody response to KLH. These results indicate that systemic antibody- and cell-mediated immune responses can be modulated by oral treatment with lactobacilli but that these bacteria may vary in their ability to modulate the immune response.

Transcriptional analysis of host responses to Marek’s disease virus infection in genetically resistant and susceptible chickens

Mareks disease virus (MDV) is a cell-associated oncogenic herpesvirus that targets B cells and T cells, inducing lymphoid tumours in chickens. Genetic resistance to Mareks disease (MD) is regulated in a polygenic fashion. In this study, we sought to compare the gene expression profiles following infection of birds that are genetically resistant or susceptible to MD (with the B21 and B19 haplotypes respectively at the MHC locus), including comparisons to uninfected controls. On days 4, 7, 14 and 21 post-infection, gene expression profiles in spleen tissue were obtained using a chicken immune-specific microarray. A number of genes showed significant (P <or= 0.05) differential expression across time and treatments. These included the chemokine AH221, B-cell marker Bu-1, IgG, IgA, IgM, MHC class II beta chain, granzyme A (GZMA) and signal transducers and activators of transcription 2 (STAT2) genes. In several comparisons, genes such as GZMA and STAT2 were induced in infected birds regardless of their genetic background. However, only immunoglobulin genes were differentially expressed by >or=2-fold in resistant compared with susceptible infected chickens. IgM and IgG were significantly induced on day 7 post-infection in susceptible chickens compared to resistant birds, whereas both of these genes were repressed in susceptible birds on day 14 post-infection. Overall, gene expression profiles in the chicken spleen observed after MDV infection were dependent on time and host genetic background. These gene expression profiles provide a platform for defining novel candidate genes for resistance or susceptibility to MD.

Host responses in the bursa of Fabricius of chickens infected with virulent Marek's disease virus

The bursa of Fabricius serves as an important tissue in the process of Mareks disease virus (MDV) pathogenesis, since B cells of the bursa harbor the cytolytic phase of MDV replication cycle. In the present study, host responses associated with MDV infection in the bursa of Fabricius of chickens were investigated. The expression of MDV phosphoprotein (pp)38 antigen, MDV glycoprotein (gB) and MDV viral interleukin (vIL)-8 transcripts was at the highest at 4 days post-infection (d.p.i.) and then showed a declining trend. On the contrary, the expression of meq (MDV EcoRI Q) gene as well as the viral genome load increased gradually until day 14 post-infection. The changes in viral parameters were associated with significantly higher infiltration of macrophages and T cell subsets, particularly CD4+ T cells into the bursa of Fabricius. Of the genes examined, the expression of interferon (IFN)-alpha, IFN-gamma genes and inducible nitric oxide synthase (iNOS) was significantly up-regulated in response to MDV infection in the bursa of Fabricius. The results suggest a role for these cells and cytokines in MDV-induced responses in the bursa of Fabricius.

Prophylactic treatment with Toll-like receptor ligands enhances host immunity to avian influenza virus in chickens.

Avian influenza viruses (AIV) pose a threat towards the health of both poultry and humans. To interrupt the transmission of the virus, novel prophylactic strategies must be considered which may reduce the shedding of AIV. One potential is the prophylactic use of Toll-like receptor (TLR) ligands. Many cells of the immune system express TLRs, and cellular responses to TLR stimulation include activation and the production of cytokines. TLR ligands have been employed as prophylactic treatments to enhance host resistance to pathogens both in mammals and chickens. Therefore, the present study was conducted to determine whether TLR ligands may be used prophylactically in chickens to enhance host immunity to AIV. Chickens received intramuscular injections of either low or high doses of the TLR ligands poly I:C, lipopolysaccharide (LPS) and CpG ODN. Twenty-four hours post-treatment, chickens were infected with the low pathogenic avian influenza virus H4N6, and both oropharyngeal and cloacal virus shedding were assessed on days 4 and 7 post-infection. To identify potential correlates of immunity, spleen and lungs were collected on days 2, 4 and 7 post-infection for RNA extraction. The results suggested that all of the TLR ligand treatments induced a significant reduction in virus shedding, with the TLR3 ligand poly I:C conferring the greatest AIV immunity compared to control birds, followed by CpG ODN and LPS. Furthermore, transcriptional analysis of gene expression in the spleen and lungs suggest IFN-α and IL-8 as correlates of immunity conferred by poly I:C, and IFN-γ for CpG ODN and LPS. In conclusion, TLR ligands, have the ability to enhance host immunity against AIV, and future studies should consider exploring the combinatory effects of poly I:C and CpG ODN prophylaxis in conjunction with AIV vaccination.

Enhancement of immunogenicity of a virosome-based avian influenza vaccine in chickens by incorporating CpG-ODN

Influenza virosomes are virus-like particles, representing a platform for vaccine development. In this study, we examined the immunogenicity of avian influenza virosomes with or without inclusion of recombinant chicken interferon-gamma (rChIFN-γ) or CpG-ODN in chickens. Immunization with virosomes adjuvanted with CpG-ODN elicited the highest haemagglutination inhibition antibody titres, as well as IgG and IgA serum antibody responses. Moreover, Virosomes+CpG-ODN formulation induced an antigen-specific spleen cell proliferation and IFN-γ expression. In conclusion, our results demonstrated that virus-specific antibody- and cell-mediated responses may be induced in chickens immunized with virosomes and these responses can be enhanced by incorporating CpG-ODN in the virosome vaccine formulation.

Cytokine gene expression in splenic CD4+ and CD8+ T cell subsets of genetically resistant and susceptible chickens infected with Marek's disease virus

T cells from the spleens of B(19)/B(19) and B(21)/B(21) chickens infected with MDV JM-16 strain were fractionated by flow cytometry at 4, 10, 21 days post infection (d.p.i.). The expression of cytokine and viral genes (meq and glycoprotein B (gB)) was measured by real-time RT-PCR. It was determined that CD4(+) and CD8(+) T cells had both become infected with Mareks disease virus (MDV) in both chicken lines. There was significantly higher expression of meq in CD4(+) T cells compared to CD8(+) T cells at 10 and 21 d.p.i. Furthermore, at 10 and 21 d.p.i., there was a tendency for higher expression of meq in both T cell subsets of B(19) chickens compared to those of B(21) chickens. There were temporal changes in the expression of cytokines, interferon (IFN)-gamma, interleukin (IL)-18, IL-6, and IL-10, in various T cell subsets. Among these changes, there was an increase in IL-10 expression in both T cell subsets at different time points, especially in the susceptible line at 10 and 21 d.p.i. Our results indicate that cytokines could be differentially induced by MDV in CD4(+) and CD8(+) T cell subsets and that IL-10 may play a role in the modulation of immune response to MDV. However, an association between cytokine gene expression in T cell subsets and resistance or susceptibility to MD was not established.

Expression of cytokine genes following pre- and post-hatch immunization of chickens with herpesvirus of turkeys.

Induction of immune response as characterised by expression of cytokine genes in the spleen following immunization of pre- and post-hatch chickens with herpesvirus of turkeys (HVT) vaccine was studied. The pattern of expression of IFN-gamma and IL-10 genes in pre-hatch immunized chickens was different from that observed in post-hatch HVT immunized chickens. This expression pattern of cytokine genes was associated with significantly higher HVT transcripts in pre-hatch immunized chickens than in post-hatch immunized chickens. In conclusion, HVT immunization in chickens, irrespective of the age of immunization, stimulates host response characterised by the expression of cytokine genes, such as IFN-gamma and IL-10 in the spleen. However, the age of immunization appears to influence the temporal pattern of IFN-gamma and IL-10 expression as well as replication of HVT.

Cytokine gene expression in splenic CD4+ and CD8+ T-cell subsets of chickens infected with Marek's disease virus.

Specific-pathogen free chickens were infected with the RB1B strain of Mareks disease virus (MDV) and T cells from the spleens of infected as well as age-matched controls were fractionated by flow cytometry at 4, 10, and 21 days post-infection (d.p.i.). Real-time quantitative reverse transcription PCR was used to assess the amount of cytokine transcripts as well as viral genes meq and glycoprotein B (gB). There was an increase in the number of CD4(+) T cells, as well as a significant increase in the expression of the viral meq gene in CD4(+) T cells, which coincided with the presence of tumors in various organs of infected birds. It was also observed that there was a significant upregulation in the amount of the gene expression of interferon (IFN)-gamma, interleukin (IL)-18, and IL-6 at 4 and 21 d.p.i. in CD4(+) and CD8(+) T-cell subsets. The expression of IL-10 was upregulated as well. The outcome of the cytokine milieu inclined towards the induction of a type I immune response at 4 and 21 d.p.i. Our study indicates that MDV-associated cytokine profiles vary in CD4(+) and CD8(+) T-cell subsets, and that cytokines including IFN-gamma, IL-18, IL-6, and IL-10 may play a role in the elicitation of an immune response to MDV.