Chicken Antibodies Research Articles

Immunoprophylactic evaluation of recombinant gametocyte 22 antigen of Eimeria tenella in broiler chickens

Gametocyte proteins are being explored as potential vaccine candidates against Eimeria sp. in chicken since they are the components of the resilient oocyst wall. The aim of this study was to investigate the immunoprophylactic efficacy of recombinant Eimeria tenella gametocyte antigen 22 (EtGam22) in chickens against homologous oocyst challenge. Broiler chicks were subcutaneously immunized individually with 100μg of recombinant EtGam22 adjuvanted with Montanide ISA 71 VG at 7days of age and boosted 2weeks later. The immunized chickens were challenged individually with 1 × 104 sporulated oocysts of E. tenella 1week post-booster immunization. The anti-EtGam22 IgY and serum cytokine response was measured post-immunization. The results showed that the anti-EtGam22 IgY antibody, serum IFN-γ, IL-2, TGF-β, and IL-4 levels in chickens vaccinated with recombinant protein were significantly increased post-immunization as compared to unimmunized challenged controls (P < 0.05). The peripheral blood lymphocyte proliferation activity was also found significantly higher in EtGam22-immunized group on day 28, i.e., pre-challenge (P < 0.05). Upon homologous oocyst challenge, chickens immunized with rEtGam22 exhibited a significant drop in the total oocyst output per bird (246.78 ± 36.9 × 106, 45.23% reduction) and a significantly higher weight gain (497.7 ± 19.2g) as compared to unimmunized challenged controls. Taken together, these data indicate that EtGam22 is a potent immunogen for use as a subunit vaccine against cecal coccidiosis in chickens as it induces a diverse and robust immune response involving multiple cytokines and strong antibody titers.

Protein A and Protein G Purification of Antibodies.

Protein A and Protein G are immunoglobulin-binding proteins expressed in Staphylococcus aureus and Streptococcus sp., respectively, that have been adapted for use in purifying large amounts of IgG. They are available covalently attached to affinity resins such as 4% cross-linked agarose, making them suitable for low-pressure antibody isolation. Protein A is not recommended for the isolation of mouse mAbs because it lacks affinity for mouse IgG1, or for the isolation of antibodies from sheep, goat, chicken, hamster, or rat. IgGs from most species bind to Protein G at near physiological pH and ionic strength with a higher affinity than IgG binding to Protein A. Therefore, the pH required to dissociate bound IgG is lower, resulting in the loss of activity for some antibodies. If this is observed, Protein A may be an alternative if the IgG from the species being isolated can be purified using Protein A. Neither Protein A nor Protein G can be used for the isolation of chicken antibodies.

Generation and characterization of avian-derived anti-human CD19 single chain fragment antibodies

The human cluster of differentiation 19 (CD19) is highly expressed in most leukemia, rendering is a promising therapeutic target. In this study, we generated anti-CD19 single-chain variable fragments (scFv) from immunized chickens by phage display technology. After constructing a scFv antibody library with 2.5 × 108 compositional diversity for panning, one representative scFv clone S2 which can specifically recognize to the CD19 protein was isolated and characterized. The binding reactivity of the scFv S2 to the endogenous CD19 protein of the ARH-77 leukemia cancer cell was verified through flow cytometry and the binding affinity of scFv S2 is 6.9 × 10−8 M determined by the surface plasmon resonance system. Compared with the chicken germline, hyper mutation in the complementarity-determining regions (CDRs) suggested that scFv S2 could be generated through an antigen-driven humoral response. By molecular modeling, the possible CDR configurations of scFv S2 were constructed rationally. Furthermore, the characteristics of chicken antibodies of a protein database were investigated. The findings in this study contribute to antibody development and engineering because they reveal the geometric structures and properties of the CDRs in chicken antibodies.

In planta expression of nanobody-based designer chicken antibodies targeting Campylobacter.

Campylobacteriosis is a widespread infectious disease, leading to a major health and economic burden. Chickens are considered as the most common infection source for humans. Campylobacter mainly multiplies in the mucus layer of their caeca. No effective control measures are currently available, but passive immunisation of chickens with pathogen-specific maternal IgY antibodies, present in egg yolk of immunised chickens, reduces Campylobacter colonisation. To explore this strategy further, anti-Campylobacter nanobodies, directed against the flagella and major outer membrane proteins, were fused to the constant domains of chicken IgA and IgY, combining the benefits of nanobodies and the effector functions of the Fc-domains. The designer chimeric antibodies were effectively produced in leaves of Nicotiana benthamiana and seeds of Arabidopsis thaliana. Stable expression of the chimeric antibodies in seeds resulted in production levels between 1% and 8% of the total soluble protein. These in planta produced antibodies do not only bind to their purified antigens but also to Campylobacter bacterial cells. In addition, the anti-flagellin chimeric antibodies are reducing the motility of Campylobacter bacteria. These antibody-containing Arabidopsis seeds can be tested for oral passive immunisation of chickens and, if effective, the chimeric antibodies can be produced in crop seeds.

Chimeric Newcastle disease virus-vectored vaccine protects chickens against H9N2 avian influenza virus in the presence of pre-existing NDV immunity.

A chimeric Newcastle disease virus (NDV) vector (NDV/AI4-TFHN) was constructed with the replacement of the ectodomains of the fusion and hemagglutinin-neuraminidase proteins by those from avian paramyxovirus type 2. The chimeric virus induced high antibody response in chickens pre-immunized with NDV. A recombinant vaccine candidate, NDV/AI4-TFHN-H9, expressing the hemagglutinin of H9N2 avian influenza virus, was generated, on the basis of the chimeric NDV vector mentioned above. The NDV/AI4-TFHN-H9 vaccine elicited H9-specific hemagglutination inhibition antibodies in chickens pre-immunized with NDV vaccine, and reduced the numbers of chickens shedding virus after H9N2 challenge. NDV/AI4-TFHN-H9 could serve as an alternative vaccine for the prevention of H9N2 infection in commercial poultry flocks.

The antigenicity and cholesteroid nature of mycolic acids determined by recombinant chicken antibodies

Mycolic acids (MA) are major, species-specific lipid components of Mycobacteria and related genera. In Mycobacterium tuberculosis, it is made up of alpha-, methoxy- and keto-MA, each with specific biological functions and conformational characteristics. Antibodies in tuberculosis (TB) patient sera respond differently towards the three MA classes and were reported to cross-react with cholesterol. To understand the antigenicity and cholesterol cross-reactivity of MA, we generated three different chicken -derived phage-displayed single-chain variable fragments (scFv) that reacted similarly towards the natural mixture of MA, but the first recognized all three classes of chemically synthetic MAs, the second only the two oxygenated types of MAs and the third only methoxy MA. The cholesterol cross-reactivity was investigated after grafting each of the three scFv types onto two configurations of constant chain domains–CH1-4 and CH2-4. Weak but significant cross-reactivity with cholesterol was found only with CH2-4 versions, notably those two that were also able to recognize the trans-keto MA. The cholesteroid nature of mycobacterial mycolic acids therefore seems to be determined by the trans-keto MA subclass. The significantly weaker binding to cholesterol in comparison to MA confirms the potential TB diagnostic application of these antibodies.

Abstract 3822: Characterization of the first chicken-derived anti-PD-1 clinical stage antibody with a unique epitope and promising anticancer activity

Abstract Inhibition of immunologic checkpoints like Programmed Cell Death 1 (PD-1) has shown clinical efficacy in a broad range of cancers by improving or restoring T-cell activity. Anti-PD-1 antibodies show great promise in treating cancer malignances when administered alone or in combination with other immune activating approaches. However, high protein sequence identity between human and mammalian species used for antibody generation often disfavor generation of antibodies against functionally conserved epitopes, or prevents isolating antibodies cross reacting with ortholog species used for evaluating potential toxicity. Chickens are phylogenetically distant from mammals and are better at generating antibodies against epitopes that are conserved in mammals. Because chickens generate antibodies from a very restricted set of V-gene germline genes that are diversified by “gene conversion”, we envisioned that high throughput humanization of antibody frameworks was achievable by “mass CDR grafting” after recovering antibodies by immunization and B-cell cloning. Wild type chickens were immunized with PD-1 antigen, and a repertoire of 120 antibodies was generated with Symplex™ technology, by combining single B-cell FACS sorting and high throughput RT-PCR cloning of cognate VH and VL chains. The isolated PD-1 repertoire was cloned with an inert Fc backbone and humanized by a combination of in silico CDR grafting and gene synthesis. Humanized antibodies were expressed and screened for retained binding affinity and functionality in T-cell based assays. We successfully generated a humanized PD-1 antibody repertoire and found that most antibodies retained affinity and functionality similar to that of parental chicken antibodies. Furthermore, the antibody repertoire displayed broad binding epitope coverage on PD-1, often with strong pM affinity, and showed biophysical properties acceptable for drug development. Our lead antibody Sym021 blocked PD-L1 and PD-L2 ligand binding and downstream PD-1 signaling, resulting in elevated T-cell cytokine production in vitro. Moreover, Sym021 bound human PD-1 with much stronger affinity of 30 pM compared to clinical PD-1 mAbs nivolumab and pembrolizumab, while also cross reacting to cynomolgous and mouse PD-1. This enabled direct testing of Sym021 in syngenic mouse in vivo models and evaluation of preclinical toxicology in cynomolgus monkeys. Detailed epitope mapping showed that the epitope recognized by Sym021 was unique compared to clinical antibodies pembrolizumab and nivolumab. These results supported entry of PD-1 targeting Sym021 into clinical trials. Citation Format: Torben Gjetting, Monika Gad, Camilla Fröhlich, Maria C. Melander, Gunther Galler, Johan Lantto, Thomas Bouquin, Ivan D. Horak, Michael Kragh, Mikkel W. Pedersen, Klaus Koefoed. Characterization of the first chicken-derived anti-PD-1 clinical stage antibody with a unique epitope and promising anticancer activity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 3822.

IgY antialérgenos específicos del grupo 1 de ácaros del polvo doméstico inducidos por oligopéptidos sintéticos no glicosilados

Introduction: The use of specific antibodies capable of detecting allergens of the group 1 of house dust mites represents a potential strategy to reduce exposure and clinical symptomatology associated with asthma and allergic rhinitis. Objective: To produce and purify chicken antibodies specific for the dust mites Dermatophagoides sp. and B. tropicalis using the IgY technology. Materials and methods: We designed and synthesized oligopeptides showing immunogenic epitopes of Der p1, Der f1, and Blo t1. These were used to produce IgY antibodies in Hy Line Brown chickens. IgY were extracted from egg yolk using thiophilic chromatography. The immunogenicity and specificity were assayed by indirect ELISA and Dot Blot. Results: We obtained high reactivity of IgY antibodies against epitopes of allergens present in whole body mites extracts of D. farinae, D. pteronyssinus, and B. tropicalis. The highest IgY levels were registered between days 32 and 40 after immunization. The antibodies showed high immunoreactivity and specificity towards D. farinae proteins with detection limits above 0.03 μg of mite proteins under the experimental conditions used. Purified IgY did not show significant reactivity when binding to Periplaneta americana extract. Conclusion: The IgY technology allowed the production of specific antibodies against house dust mites group 1 allergens using non-glycosylated synthetic peptides. To our knowledge, this is the first time that this immunochemicals are used in the detection of mites of medical relevance.

Seroprevalence study of infectious laryngotracheitis virus antibody of commercial layer in Gazipur Districts of Bangladesh

This research work was conducted to detect the prevalence of Infectious Laryngotracheitis (ILT) Virus-specific antibody in chickens from Gazipur district in Bangladesh at Department of Microbiology, Hajee Mohammad Danesh Science &amp; Technology University, Dinajpur and Poultry Care Lab, Garzipur from January to June 2012. A total number of 232 sera sample of commercial layer chicken were collected from 17 different commercial layer farms at different ages. The layers prognosed for sampling had not been previously vaccinated against ILTV. The indirect enzyme linked immunosorbent assay (iELISA) was performed to estimate the Infectious Laryngotracheitis (ILT) Virus-specific antibody. Out of 232 samples, 189 (81.47%) samples were found to positive for Infectious Laryngotracheitis (ILT) Virus-specific antibody. In 17 different commercial farms prevalence based on age were 75%, 87.5%, 87.5%, 90%, 81.25%, 80%, 100%, 70%, 81.25%, 81.25%, 90%, 93.08%, 87.5%, 75%, 75%, 68.75% and 75% in the age limit 08, 09, 11, 12, 13, 14, 15, 16, 17, 17, 19, 21, 25, 31, 35, 44 and 51 weeks respectively and farms showed high level of ILT virus specific antibodies (IgG). This result showed that in 15 weeks of age prevalence was highest position i.e; 100%. The result of this study indicate that there were a high prevalence of Infectious Laryngotracheitis (ILT) Virus circulating at Gazipur district in Bangladesh.Asian J. Med. Biol. Res. March 2018, 4(1): 1-6

Prevalence of interfering antibodies in dogs and cats evaluated using a species-independent assay.

Interfering antibodies in human serum and plasma are known to react with mammalian antibodies in immunoassays and cause false-positive test results. Although this phenomenon was recently shown in companion animals, knowledge regarding immunoassay interference in veterinary medicine is very limited. The aims of this study were to set up a species-independent immunoassay procedure to detect interference in serum samples, to screen for interference in a cross-section of canine and feline patient samples from an animal hospital, and to determine if the detected interference could be neutralized using an immunoassay based on nonmammalian reagents. A 2-site sandwich-type interference assay was set up using commercially available mouse reagents. A total of 369 serum samples from 320 dogs and 263 samples from 218 cats were analyzed using the interference assay. Multiple samples were submitted from 36 dogs and 39 cats. Nineteen samples identified as interference-positive were analyzed in an assay using chicken antibodies. Interference was detected in samples from 28 dogs (9%) and 10 cats (5%) screened with the interference assay. Except for 1 cat, consistent results were obtained for all 75 dogs and cats that submitted more than 1 sample. The interference was eliminated when analyzed in the chicken-based assay (P<.001). Substances with reactivity toward mouse IgG can be detected in serum samples from dog and cat patients using a 2-site interference assay. The detected substances are most likely interfering antibodies, possibly originating from immunization with other mammalian species.

Serological survey of Infectious Bursal Disease Antibodies amongst birds in Keffi, Nassarawa State, Nigeria

Infectious Bursal Disease (IBD) is an acute, immunosuppressive disease of significant threat to the poultry industry in Nigeria and vaccination has been reported as the best method of control. This study was conducted between November and December 2015 to establish the occurrence of infectious bursal disease antibodies in exotic chickens from some selected backyard poultry farms and local chickens slaughtered in keffi central market, Nassarawa State. Sera collected randomly from two hundred birds were subjected to Agar Gel Precipitation Test (AGPT) for qualitative analysis at the virology laboratory (FCVMLT), National Veterinary Research Institute, Vom. The assay showed 35% of exotic birds had IBD virus antibodies while local birds indicated absence of detectable antibodies with significant P&lt;0.05 breed association (Χ2= 42.5, df=2). However, there was no association (P&gt;0.05) with age in both breeds and IBDV antibodies (Χ2=0.802, 3.13, df=1). The presence of antibodies in exotic breeds suggests evidence of post vaccination protective antibodies due to sero-conversion rather than anti-IBDV antibodies associated with disease. Absence of infectious bursal disease virus antibodies in local birds suggests high vulnerability to potential IBD outbreaks posing a risk of financial losses to stakeholders. In conclusion, this sero-survey of Infectious bursal disease antibodies in exotic birds suggest evidence of vaccination and the need to vaccinate local birds. Awareness campaign program amongst rural and backyard poultry keepers in the study community on the need to protect their commercial exotic and free scavenging birds against infectious bursal disease is thus recommended.

Frequency of anti-Toxocara antibodies in broiler chickens in southern Brazil.

The aim of this study was to evaluate the presence of anti-Toxocara antibodies in naturally infected broiler chickens (n = 189) from the state of Paraná, southern Brazil. The chickens were reared in a semi-intensive system by small family farmers (n = 7). An enzyme-linked immunosorbent assay (ELISA) was performed to detect the presence of anti- Toxocara spp. IgY after serum adsorption with Ascaridia galli antigens. An overall seroprevalence of 67.7% (128/189; 95% CI = 61.1-74.4) was observed. The frequency of positive animals by farm ranged from 29.6% to 100%. The optical density and reactivity index values observed in ELISA test indicated the possible chronicity of infection of the evaluated chickens. Associations between the presence of antibodies and the area where the chickens were reared (p = 0.382) or the population density of dogs on the farm (p = 0.785) were not observed. This study shows a high prevalence of Toxocara spp. antibodies in broiler chickens reared in semi-intensive systems and provides evidence that chickens are a good indicator of environmental contamination by larva migrans agents. Further studies are necessary to assess the risk factors associated with poultry infection and the likelihood of toxocariasis transmission to humans via the ingestion of free-range chicken meat.

Phosphoethanolamine-N-methyltransferase is a potential biomarker for the diagnosis of P. knowlesi and P. falciparum malaria.

BackgroundPlasmodium knowlesi is recognised as the main cause of human malaria in Southeast Asia. The disease is often misdiagnosed as P. falciparum or P. malariae infections by microscopy, and the disease is difficult to eliminate due to its presence in both humans and monkeys. P. knowlesi infections can rapidly cause severe disease and require prompt diagnosis and treatment. No protein biomarker exists for the rapid diagnostic test (RDT) detection of P. knowlesi infections. Plasmodium knowlesi infections can be diagnosed by PCR.Methods and principal findingsPhosphoethanolamine-N-methyltransferase (PMT) is involved in malaria lipid biosynthesis and is not found in the human host. The P. falciparum, P. vivax and P. knowlesi PMT proteins were recombinantly expressed in BL21(DE3) Escherichia coli host cells, affinity purified and used to raise antibodies in chickens. Antibodies against each recombinant PMT protein all detected all three recombinant proteins and the native 29 kDa P. falciparum PMT protein on western blots and in ELISA. Antibodies against a PMT epitope (PLENNQYTDEGVKC) common to all three PMT orthologues detected all three proteins. Antibodies against unique peptides from each orthologue of PMT, PfCEVEHKYLHENKE, PvVYSIKEYNSLKDC, PkLYPTDEYNSLKDC detected only the parent protein in western blots and P. falciparum infected red blood cell lysates or blood lysates spiked with the respective proteins. Similar concentrations of PfPMT and the control, PfLDH, were detected in the same parasite lysate. The recombinant PfPMT protein was detected by a human anti-malaria antibody pool.ConclusionPMT, like the pan-specific LDH biomarker used in RDT tests, is both soluble, present at comparable concentrations in the parasite and constitutes a promising antimalarial drug target. PMT is absent from the human proteome. PMT has the potential as a biomarker for human malaria and in particular as the first P. knowlesi specific protein with diagnostic potential for the identification of a P. knowlesi infection.

Genomic Region Containing Toll-Like Receptor Genes Has a Major Impact on Total IgM Antibodies Including KLH-Binding IgM Natural Antibodies in Chickens.

Natural antibodies (NAb) are antigen binding antibodies present in individuals without a previous exposure to this antigen. Keyhole limpet hemocyanin (KLH)-binding NAb levels were previously associated with survival in chickens. This suggests that selective breeding for KLH-binding NAb may increase survival by means of improved general disease resistance. Genome-wide association studies (GWAS) were performed to identify genes underlying genetic variation in NAb levels. The studied population consisted of 1,628 adolescent layer chickens with observations for titers of KLH-binding NAb of the isotypes IgM, IgA, IgG, the total KLH-binding (IgT) NAb titers, total antibody concentrations of the isotypes IgM, IgA, IgG, and the total antibodies concentration in plasma. GWAS were performed using 57,636 single-nucleotide polymorphisms (SNP). One chromosomal region on chromosome 4 was associated with KLH-binding IgT NAb, and total IgM concentration, and especially with KLH-binding IgM NAb. The region of interest was fine mapped by imputing the region of the study population to whole genome sequence, and subsequently performing an association study using the imputed sequence variants. 16 candidate genes were identified, of which FAM114A1, Toll-like receptor 1 family member B (TLR1B), TLR1A, Krüppel-like factor 3 (KLF3) showed the strongest associations. SNP located in coding regions of the candidate genes were checked for predicted changes in protein functioning. One SNP (at 69,965,939 base pairs) received the maximum impact score from two independent prediction tools, which makes this SNP the most likely causal variant. This SNP is located in TLR1A, which suggests a fundamental role of TLR1A on regulation of IgM levels (i.e., KLH-binding IgM NAb, and total IgM concentration), or B cells biology, or both. This study contributes to increased understanding of (genetic) regulation of KLH-binding NAb levels, and total antibody concentrations.

The Versatility of Framework Regions of Chicken VH and VL to Mutations.

To identify the interchangeability of VH and VL framework region (FR) residues, we artificially introduced random mutations at all residue positions in a chicken monoclonal antibody, which has only one functional VH and Vλ gene. When we classified the amino acids into 5 groups by their physicochemical properties, all FR residues could be replaced by another group except L23 (C), H36 (W), H86 (D), H104 (G), and H106 (G). Eighty-two (50.9%), 48 (29.8%), 17 (10.6%), and 9 FR residues (5.6%) could be replaced by 4, 3, 2, and 1 group(s), individually, without significant loss of reactivity. We also confirmed a similar level of versatility with 2 different chicken antibodies. This high level of versatility on FR residues has not been predicted because it has not been observed in the 150 chicken antibodies that we previously generated or in the 1,269 naïve chicken VH sequences publically available. In conclusion, chicken antibody FR residues are highly interchangeable and this property can be applied for improving the physicochemical property of antibody including thermal stability, solubility and viscosity.

Inactivated Vaccine Trial of &amp;lt;i&amp;gt;Mycoplasma gallisepticum&amp;lt;/i&amp;gt; in Ethiopia

The study and entire laboratory works were conducted from December 2014 to April 2015 in National Veterinary Institute, Bishoftu, Ethiopia. Formaldehyde inactivated Montanide ISA70 based Mycoplasma gallisepticum (MG) trial vaccine strain was confirmed the identity with known primer using PCR from locally isolates of National Veterinary Institute of Ethiopia. This study was aimed to develop formaldehyde inactivated Montanide ISA70 based MG vaccine in Ethiopia. It can help to device strategies in controlling the disease mainly through developing more effective vaccine which will replace the currently being imported vaccines by some farms. After culturing procedure, oil based inactivated MG trial vaccine was produced in suitable clean and secure accommodation. In this study, among different isolates, local isolate of Samuel farm in NVI was prepared and evaluated in chickens. The amount of immune antigen per 0.5 ml of the dose was 107 Colony forming units (CFU) of the bacteria. The trail vaccine was prepared and evaluated at the age of 16 weeks of chickens; the chickens were randomly divided into three groups (A, B and C), each having twenty birds (10 male and 10 female). Each of group B was vaccinated group of imported-live vaccine with 30 μl intraocularly for comparing with inactivated trial vaccine, each bird of group C was inoculated with 0.5 ml indigenous or trial vaccine subcutaneously at mid neck region and group A was used as a control then challenge tests were performed. After challenge test, among non-vaccinated chickens (control or group A) 2 chickens were died (10%), thicken and cloudy appearance of the air sac showed 18 (90%), 2 chickens were not showed thickened and cloudy air sack (10%). Although among vaccinated group (inactivated vaccine or group C), all chickens did not show clinical signs or post mortem changes (100%). From attenuated imported live vaccine (group B), no clinical signs or post mortem changes were observed (100%). It was concluded that oil based MG vaccine induces protective level of anti MG antibodies in chickens.

Suppression of Highly Pathogenic Avian Influenza A/H5N1 Infection Using Migratory Antibody Passed from Mother to Chick

Avian influenza is the most contagious disease not only in poultry, but also in humans. Avian influenza in humans occurs mainly in Southeast Asia, but no human-to-human pandemic has occurred. Meanwhile, outbreaks of avian influenza in poultry occur on a global scale and cause a large economic loss. Migration antibodies passed from mother birds via eggs are said to be an important component of the immune system that protects birds from infection. Thus, the immunity status of mother birds can determine the ability of offspring to defend against infection. In this study, we investigated the presence of anti-avian influenza virus antibody in chickens hatched on a poultry farm in Indonesia and examined the involvement of migratory antibodies in protecting against virus infection by infectious experiments of highly pathogenic avian influenza in chickens. Blood was collected from randomly selected chicks, and antibodies against avian influenza virus were evaluated in all birds. Since these young birds had no history of vaccination, the antibodies were deemed to have been transferred from the mother birds. The enzyme-linked immunosorbent assay antibody titer in each bird varied. Infection of these birds with highly pathogenic avian influenza virus A/H5N1 intra-nasally resulted in a high mortality rate in chicks with low antibody titers but a low mortality rate in chicks with high antibody titers. These findings indicate that migratory antibody prevented highly pathogenic avian influenza A/H5N1 infection in chicks, suggesting that such a preventive effect could also be expected with outdoor natural infection.

Anti-Müllerian hormone receptor type 2 is expressed in gonadotrophs of postpubertal heifers to control gonadotrophin secretion.

Preantral and small antral follicles may secret anti-Müllerian hormone (AMH) to control gonadotrophin secretion from ruminant gonadotrophs. The present study investigated whether the main receptor for AMH, AMH receptor type 2 (AMHR2), is expressed in gonadotrophs of postpubertal heifers to control gonadotrophin secretion. Expression of AMHR2 mRNA was detected in anterior pituitaries (APs) of postpubertal heifers using reverse transcription-polymerase chain reaction. An anti-AMHR2 chicken antibody was developed against the extracellular region near the N-terminus of bovine AMHR2. Western blotting using this antibody detected the expression of AMHR2 protein in APs. Immunofluorescence microscopy using the same antibody visualised colocalisation of AMHR2 with gonadotrophin-releasing hormone (GnRH) receptor on the plasma membrane of gonadotrophs. AP cells were cultured for 3.5 days and then treated with increasing concentrations (0, 1, 10, 100, or 1000pgmL-1) of AMH. AMH (10-1000pgmL-1) stimulated (P<0.05) basal FSH secretion. In addition, AMH (100-1000pgmL-1) weakly stimulated (P<0.05) basal LH secretion. AMH (100-1000pgmL-1) inhibited GnRH-induced FSH secretion, but not GnRH-induced LH secretion, in AP cells. In conclusion, AMHR2 is expressed in gonadotrophs of postpubertal heifers to control gonadotrophin secretion.

Detection of White Root Disease (Rigidoporus Microporus) in Various Soil Types in the Rubber Plantations Based on The Serological Reaction

The Conventional detection of White Root Disease (Rigidoporus microporus, WRD) still uses the visual method based on an abnormal color of leaf or mycelium growth on the tap root neck. The method was less effective and less efficient. The serological technique uses yolk chicken antibodies induced by immunization with mycelium extract. The purpose of this research was to examine the consistency of selected antibodies in detecting root fungi at various soil types in the rubber plantations. This research used a Completely Randomized Design non-factorial with twelve (12) treatments and two (2) replications. The results showed that the antibodies could detect WRD in various soils types. The serological detection was higher precisely than visual observation. The development of WRD mycelium varies depending on the soil types and it was different in the each estate area. In addition, this research is expected to get a serology kit to detect early symptoms of WRD in the rubber plants.

Construction of Chicken Antibody Libraries.

Recombinant antibody libraries based on chicken immunoglobulin genes are potentially valuable sources of phage displayed scFvs for use in veterinary diagnostics and research. The libraries described in this chapter are based on chicken variable heavy and light chain immunoglobulin genes joined by a short flexible linker cloned in the phagemid vector pHEN1. The resulting phagemids produce either scFvs displayed on the surface of the fusion phage subsequent to co-infection with helper phage, or soluble scFvs following IPTG induction. This chapter provides detailed and proven methods for the construction of such libraries.