The bursa of Fabricius (BF), which is unique to birds, serves as the central humoral immune organ and plays a significant role in B lymphocyte differentiation. In this study, a new bursal peptide (BP-IV) was isolated from BF, which promoted colony-forming unit pre-B formation and regulated B cell differentiation. BP-IV also exerted immunomodulatory effects on antigen-specific immune responses via both humoral and cellular immunity in chicken and mice that had been immunized with inactivated avian influenza virus (AIV; H9N2 subtype), including enhancing AIV-specific antibody and cytokine production. The results of this study provided novel insights into the use of a potential candidate reagent for B cell development and future immuno-pharmacological use.
Animals ; Birds ; Bursa of Fabricius* ; Cell Differentiation ; Chickens* ; Immunity, Cellular ; Influenza in Birds ; Lymphocytes ; Mice ; Stem Cells

Infectious bursal disease virus (IBDV) causes a highly contagious and immunosuppressive disease of chicken. Agar gel immunodiffusion using IBDV antigen extracted from bursa of Fabricius of infected chicken has been used officially for diagnosis of IBDV in Korea. In this study, in order to replace the IBDV whole virus antigen with non-infectious antigen, recombinant VP2 protein (rVP2) of IBDV was produced using recombinant baculovirus expression system. Purified baculovirus-expressed rVP2 was used as an antigen in an agar gel immunodiffusion (AGID). rVP2 antigen precipitated specifically IBDV antibodies. AGID using rVP2 antigen detected anti-IBDV antibodies from 6 dpi to 28 dpi (termination of the experiment) when specific pathogen free chickens were experimentally infected with IBDV 52/70 strain. This was consistent with result by AGID using IBDV antigen, virus neutralization test (VNT) and a commercial ELISA kit (except for one serum). The sensitivity of rVP2 was the same with that of IBDV antigen when field sera (n=324) were tested by AGID. However, AGID using rVP2 antigen detected maternal antibodies from broiler chickens (n=20) on a broiler farm up to 15 days old, although the detection rate of the AGID was relatively low compared to a commercial ELISA kit. Our results indicate that IBDV whole virus antigen from IBDV infected chickens would be replaced with recombinant VP2 protein as an antigen for AGID.
Agar ; Animals ; Antibodies ; Baculoviridae ; Bursa of Fabricius ; Chickens ; Enzyme-Linked Immunosorbent Assay ; Immunodiffusion ; Infectious bursal disease virus ; Korea ; Neutralization Tests ; Specific Pathogen-Free Organisms ; Sprains and Strains ; Staphylococcal Protein A ; Viruses

This study describes the histochemical characteristics and ultrastructure of mast cells from tongue, proventriculus, ileum and fabricius bursa, in pheasant (Phasianus colchicus) by light and electron microscopy. We compared the stainability of 4 different methods, toluidine blue, alcian blue, congo red and alkaline Giemsa, to stain mast cell granules from fixed pheasant organs in three different fixatives, 10% neutral buffered formalin, Carnoy's solution or half-strength Karnovsky's solution. Mast cells in all experimental organs were not stained with 4 different staining methods after fixation in 10% neutral buffered formalin but well stained in fixed organs with half-strength Karnovsky's solution. The mast cells had many metachromatic granules stained with toluidine blue or alkaline Giemsa and orthochromatic granules stained with alcian blue or congo red in tissues fixed in half-strength Karnovsky's solution. In electron microscopy, pheasant mast cells were oval, triangular, spindle-like or irregular and had a few finger-like cytoplasmic processes. There were the membrane-bounded secretory granules and the well-developed organelles in mast cells. Internal large granules were oval or irregular, and had variable shape; some higher or lower electron density with homogeneous appearance; some had a particular appearance, and a few showed reticular or spongy-like structure. This indicates that 10% neutral buffered formalin or Carnoy's fixation may be inadequate for detection of mast cells in pheasant, whereas the half-strength Karnovsky's fixation provides metachromatic or orthochromatic staining of mast cell granules.
Alcian Blue ; Animals ; Bursa of Fabricius ; Congo Red ; Cytoplasm ; Fixatives ; Formaldehyde ; Ileum ; Mast Cells* ; Microscopy, Electron ; Organelles ; Proventriculus ; Secretory Vesicles ; Tolonium Chloride ; Tongue

The bursa of Fabricius (BF) is a central humoral immune organ unique to birds. Four bursal peptides (BP-I, BP-II, BP-III, and BP-IV) have been isolated and identified from the BF. In this study, the immunoadjuvant activities of BPs I to IV were examined in mice immunized with H9N2 avian influenza virus (AIV) vaccine. The results suggested that BP-I effectively enhanced cell-mediated immune responses, increased the secretion of Th1 (interferon gamma)- and Th2 (interleukin-4)-type cytokines, and induced an improved cytotoxic T-lymphocyte (CTL) response to the H9N2 virus. BP-II mainly elevated specific antibody production, especially neutralizing antibodies, and increased Th1- and Th2-type cytokine secretion. BP-III had no significant effect on antibody production or cell-mediated immune responses compared to those in the control group. A strong immune response at both the humoral and cellular levels was induced by BP-IV. Furthermore, a virus challenge experiment followed by H&E staining revealed that BP-I and BP-II promoted removal of the virus and conferred protection in mouse lungs. BP-IV significantly reduced viral titers and histopathological changes and contributed to protection against H9N2 AIV challenge in mouse lungs. This study further elucidated the immunoadjuvant activities of BPs I to IV, providing a novel insight into immunoadjuvants for use in vaccine design.
Adjuvants, Immunologic ; Animals ; Antibodies, Neutralizing ; Antibody Formation ; Birds ; Bursa of Fabricius ; Cytokines ; Immunity, Cellular ; Immunity, Humoral ; Influenza A Virus, H9N2 Subtype ; Influenza in Birds ; Lung ; Mice ; Peptides ; T-Lymphocytes, Cytotoxic

The bursa of Fabricius (BF) is the acknowledged central humoral immune organ in birds. Bursal septpeptide II (BSP-II) is an immunomodulatory bioactive peptide isolated from BF. To understand the effects of BSP-II on immune induction, gene expression profiles of hybridoma cells treated with BSP-II were evaluated. Pathway analysis showed that regulated genes were involved in cytokine-cytokine receptor interactions, T cell receptor signaling pathway, and pathway in cancer. It was observed that BSP-II reduced tumor cells proliferation and stimulated p53 expression. These results indicate potential mechanisms underlying the effects of the humoral immune system on immune induction, including antitumor activities. Our study has provided a novel insight into immunotherapeutic strategies for treating human tumors.
Animals ; Antineoplastic Agents/*pharmacology ; Avian Proteins/*pharmacology ; Bursa of Fabricius/immunology ; Cell Proliferation/drug effects ; Chickens/*immunology ; Hybridomas/drug effects ; Immunologic Factors/*pharmacology ; Oligonucleotide Array Sequence Analysis/veterinary ; Signal Transduction/*drug effects ; *Transcriptome

The present study was undertaken to determine the viability and infectivity of oocysts of Cryptosporidium baileyi that had been stored from 1 to 40 months at 4 degrees C preserved in 2.5% potassium dichromate solution. Oocysts of C. baileyi were purified from the feces of experimentally infected chickens using discontinuous sucrose gradients. Subsequently, the purified oocysts were suspended in 2.5% potassium dichromate solution at a concentration of 1 x 10 (7) organism/ml, and their viabilities were assessed by nucleic acid staining, histologic examination, and infectivity to 2-day-old chickens. All chickens inoculated with oocysts that had been stored for 1-18 months developed patent infections, while chickens infected with older oocysts remained uninfected. Between 5.8% and 82.2% of the oocysts, stored at 4 degrees C in 2.5% potassium dichromate solution, were found to be viable, as determined by nucleic acid staining. Parasite colonization in the bursa of Fabricius was detected in the microvillus border of bursal epithelium. The finding that C. baileyi oocysts remain infective to chickens for at least 18 months offers important time-saving advantages to investigators who frequently require large numbers of oocysts.
Animals ; Bursa of Fabricius/parasitology ; Chickens/*parasitology ; Coloring Agents ; Cryptosporidiosis/parasitology/pathology/*veterinary ; Cryptosporidium/drug effects/*growth & development/pathogenicity ; Feces/parasitology ; Oocysts/drug effects/*growth & development/pathogenicity ; *Organic Chemicals ; *Potassium Dichromate/pharmacology ; Poultry Diseases/parasitology/pathology ; Preservation, Biological/*methods ; Staining and Labeling

The aim of this work was to investigate developmental changes in cell proliferation and apoptosis in normal duck bursa of Fabricius using flow cytometry and immunohistochemistry. Studies were carried out on Tianfu ducks on days 24 and 27 of embryogenesis (E24 and E27) along with days 20, 70, and 200 of postnatal development (P20, P70, and P200). Results showed that the percentage of G0/G1 bursa cells significantly increased between E24 and P200 while the percentage of cells in the S phase or G2 + M phase as well as the proliferating index obviously decreased during the same period. Proliferation cell nuclear antigen was detected in lymphocyte and interfollicular epithelium. The proliferative lymphocyte density tended to decrease from E24 to P200. Apoptotic bodies in macrophages, free apoptotic bodies, or nuclei with condensed chromatin in lymphocytes in follicles were identified by transferase-mediated dUTP nick-end labeling. Both flow cytometry and microscopic analysis reveal that the proportion of apoptotic cells and apoptotic lymphocyte density increased from E24 to P20, fell on P70, then rose again on P200. Our foundings demonstrate that cell proliferation decreases and apoptosis increases with age. These changes may account for duck bursa development and involution.
Animals ; *Apoptosis ; Bursa of Fabricius/*cytology/embryology/growth & development/*physiology ; Cell Proliferation ; Ducks/embryology/*physiology ; Embryo, Nonmammalian/cytology/embryology ; Embryonic Development ; Epithelium/physiology ; Female ; Flow Cytometry/veterinary ; Immunohistochemistry/veterinary ; Lymphocytes/physiology ; Male

To screen the interactive proteins with IBDV Gt VP2 protein from cDNA library of B Lymphoid cells of the bursa of Fabricius. The expression cDNA library plasmids was transformed to the yeast competent cells, which have the bait plasmid-Gt VP2. After testing for growth in synthetic complete medium lacking histidine and uracil and for production of beta-galactosidase (X-gal), we obtained 16 positive clones. We searched the gene sequences of positive clones in the NCBI website. The blast results showed that five positive clones were the gallus sequences. They were Gallus gallus breed mitochondrial DNA, O_G1cNAc transferase, Tumor protein p53 binding protein, Stathmin and Chondroitin sulfate Ga1NAcT-2, respectively. This study is helpful for the further identifying the receptors of IBDV in B Lymphoid cells of the bursa of Fabricius.
Animals ; B-Lymphocytes ; metabolism ; virology ; Bursa of Fabricius ; metabolism ; Chickens ; DNA, Mitochondrial ; metabolism ; Gene Library ; Infectious bursal disease virus ; Protein Binding ; Protein Interaction Mapping ; Receptors, Virus ; metabolism ; Tumor Suppressor Protein p53 ; metabolism ; Two-Hybrid System Techniques ; Viral Structural Proteins ; genetics ; metabolism

OBJECTIVETo study on the effect of been pollen on development of immune organ of animal.

METHODA total of 144 one day-old broilers were randomly divided into 2 groups, in which each group included 72 chickens. The control group was fed on the basal diet for 42 days, and that of experiment group supplemented 1.5% bee pollen. Six chickens in each group were selected and slaughtered at 7, 14, 21, 28, 35, 42 days respectively, and the thymuses, cloacal bursa and spleens were obtained, weighted, fixed in Bouin liquid and made into paraffin section.

RESULTCompared with control group, the weight and the relative weight of thymuses, cloacal bursa and spleens of experiment group increased significantly (P < 0.05) or extremely significantly (P < 0.01). In experiment group, the cortex of thymic lobule, bursa nodule and Periarterial Lymphatic Sheaths thicken obviously; the volume of bursa nodule, splenic nodule and ellipsoid augmented, and the germinal center of splenic nodule were obvious; the thymic corpuscle increased; the plica of cloacal bursa developed well and the degenerating of it retarded.

CONCLUSIONThe diet supplemented bee pollen could boost the early development of thymus and cloacal bursa, retard the degenerating of cloacal bursa and promote the immune response of spleen.

Animal Nutritional Physiological Phenomena ; Animals ; Bees ; Bursa of Fabricius ; anatomy & histology ; growth & development ; Chickens ; growth & development ; immunology ; Female ; Male ; Organ Size ; Pollen ; Random Allocation ; Spleen ; anatomy & histology ; growth & development ; Thymus Gland ; anatomy & histology ; growth & development

Studies were performed to determine the effects of Bcell suppression on the pathogenesis of Subgroup J avian leukosis virus (ALV-J) in broiler chickens. Neonatal chickens were treated with cyclophosphamide (CY) or PBS, and then infected with ALV-J (ADOL-7501) at 2 weeks of age. CY treatment induced B cell specific immunosuppression throughout the experiment confirmed by decreased bursal weight, intact lymphocyte mitogenetic activity stimulated by Con A and increased relative subpopulation of CD3-positive cells as measured by flow cytometry. Chickens in this experiment had Mareks disease virus exposure prior to three weeks of age as determined by the presence of lymphocytic infiltration and antibody. Virus neutralizing antibody against ALV-J was first observed at 6 weeks post-infection in some of the infected chickens in the PBS group. As expected, none of the chickens from the CY group and uninfected chickens developed virus-neutralizing antibody. The viremic status was measured by real time RT-PCR using SYBR green I dye. The percentage of viremic chickens was significantly higher, and more chickens had high titered viremia, in the CY treated group. No neoplastic foci consistent with ALVJ infection were observed in any of the experimental chickens. The frequency and intensity of viral antigen expression determined by immunohistochemistry was significantly higher in tissues from CY treated birds than those of PBS treated chickens at 3 weeks post-infection. This study showed that B cell specific immunosuppression with CY treatment in chickens resulted in increase in viremia and viral antigen load in tissues.
Animals ; Avian Leukosis/*immunology/virology ; Avian leukosis virus/genetics/*immunology ; Body Weight/physiology ; Bursa of Fabricius/immunology ; *Chickens ; Concanavalin A/immunology ; Cyclophosphamide/*pharmacology ; Flow Cytometry/veterinary ; Immunocompromised Host ; Immunohistochemistry/veterinary ; Immunophenotyping/veterinary ; Immunosuppressive Agents/*pharmacology ; Lymphocyte Activation/drug effects/immunology ; Organic Chemicals/chemistry ; Poultry Diseases/immunology/*virology ; RNA, Viral/chemistry/genetics ; Random Allocation ; Reverse Transcriptase Polymerase Chain Reaction/veterinary ; Spleen/immunology/virology ; Statistics, Nonparametric ; Viremia/veterinary