Early cell-mediated immune responses to Marek's disease virus in two chicken lines with defined major histocompatibility complex antigens

N2a and P2a chickens, resistant and susceptible to Marek's disease (MD), respectively, were used to examine relationships between major histocompatibility complex (MHC)-restricted cytotoxic T lymphocytes (CTL) and natural killer (NK)-like cell activity with resistance to infection with Marek's disease virus (MDV). Ten-day-old chickens were infected with MDV and euthanatized at selected times to evaluate for NK cell and MHC-restricted cytotoxicity. The N2a MDV-infected chickens had an early cell-mediated immune response characterized by a sustained NK-like cytotoxicity that coincided with a measurable MHC-cytotoxicity that was lower than controls. Although MHC-restricted and NK cell cytotoxicity was demonstrated in P2a MDV-infected chickens at 8 dpi, both abruptly decreased and remained low for the remainder of the 20-day experiment. The critical time point that may determine the resistance to MD appears to be within the first 2 weeks post-infection. Improvement of the chicken NK cell activity may be a good candidate for both selection and immunomodulation MD control programs.     

Association of the major histocompatibility complex with avian leukosis virus infection in chickens.

1. Association of the B blood group, the major histocompatibility complex (MHC) in chickens, with avian leukosis virus (ALV) infection shown by shedding of group-specific (gs) antigen was studied in an Australorp line selected for short oviposition interval to improve egg production. Three haplotypes (B8a, B9a and B21) were segregating in this line at frequencies of 66.7, 15.6 and 17.8%, respectively, averaged over three generations. 2. The relative risk (odds ratio) of a hen becoming a gs-antigen shedder was calculated for progenies of the dams shedding gs-antigen and those of non-shedding dams separately and pooled over three generations. In the progenies of shedding dams, the relative risk was not significantly different from 1.0 for the three haplotypes. In contrast, in the progenies of non-shedding dams, the relative risk was 0.67, 0.48 and 2.53 for B8a, B9a and B21, respectively, with the last two ratios being significantly different from 1.0. 3. The average effect of haplotype substitution on probability of shedding was estimated from a linear logistic model. The estimates (relative to zero for B8a) for B9a and B21, respectively, were -0.26 and 0.03 among the progenies of shedding dams, and -0.16 and 0.87 among the progenies of non-shedding dams. The last estimate only was highly significant. 4. These results suggest that the three haplotypes were similar in susceptibility to congenital infection through hatching eggs, but differed in susceptibility to post-hatching infection from other infected birds.     

Structure, function and disease susceptibility of the bovine major histocompatibility complex

The major histocompatibility complex (MHC) of cattle is known as the bovine leukocyte antigen (BoLA) and is located on chromosome 23. BoLA has been linked to variation in resistance to disease including bovine leukemia virus‐induced lymphoma and mastitis. Moreover, BoLA appears to influence other traits such as milk yield, growth and reproduction, which are not often measured in humans, and variations in individual immune response to antigen. The BoLA appears to be organized in a similar way to the MHC region in humans, but there are notable differences. A major rearrangement within the class II region has led to the division of the BoLA into two distinct subregions of chromosome 23 separated by about a third of the chromosome’s length. The class IIa subregion contains functionally expressed DR and DQ genes, while the class IIb subregion contains the genes of undefined status such as DYA, DYB, DMA, DMB, DOB, DOA, TAP1, TAP2, LAP2 and LMP7. In addition, one pair of human class II genes (DP) does not appear to have an equivalent in cattle, and there is one pair of DY genes that seem to be found only cattle, sheep and goats. In humans, three classical, polymorphic class I genes (HLA‐A, ‐B and ‐C ) are each present on all haplotypes. However, in cattle, none of the four (or more) classical class‐I genes identified are consistently expressed, and haplotypes differ from one to another in both the gene number and composition. These variations in both class I and II are likely to play an important role in cattle immune responses. This review summarizes current knowledge of the structural and functional features and disease association of BoLA genes.     

The effect of serologically defined major histocompatibility complex haplotypes on Marek's disease resistance in commercially bred White Leghorn chickens

In commercial pure white leghorn lines, A, B, and C, the effects on resistance against a virulent strain of Marek's disease virus were assessed for B19 and B21 haplotypes of the chicken major histocompatibility complex. B haplotypes were identified by direct hemagglutination using alloantisera raised against erythrocyte antigens. In homozygous B21 female chicks from lines A and B, mortality upon challenge with virus was 16% and 9%, respectively; in B19 chicks, mortality was 42% and 60%, respectively. Intermediate mortality was observed in heterozygous B19/B21 birds. When line A and B hens were crossed with B15/B15 or B5/B19 cocks from line C, differences between B19 and B21 were significant only in the progeny from B5/B19 sires. Therefore, it was concluded that selection for major histocompatibility complex-associated disease resistance markers may be useful only when B haplotypes complement each other in commercial line crosses and when interactions with genetic background do not severely obscure the differential haplotype effects, as are observed within pure lines.     

Testing the immunogenicity of the dermal antigen in Marek's disease virus]

An experiment was performed to study the immunogenicity of the dermal antigen of Marek's disease virus, extracted from the skin of 30-day-old chickens, infected with Marek's disease virus on the first day of life. Three kinds of samples were tested: (1) dermal antigen centrifuged at 10 000 g per 0.5 h, (2) dermal antigen centrifugated at 10 000 g per 0.5 h and 100 000 g per 1 h, (3) dermal antigen treated like sample (2) and partly purified by DEAE-cellulose chromatography. Samples (1) and (2) were inoculated to two-day-old chickens and the vaccination was repeated, using complete Freund's adjuvant, 21 days later. Sample (3) was inoculated to two-day-old chickens with DEAE-dextran. All the three groups were challenged together with the controls (non-vaccinated chickens) on the seventh day after the first vaccination. A reduction of mortality was observed in the chickens vaccinated with and re-vaccinated with sample (1) (23.07%) and in the chickens vaccinated with sample (3) (30.76%). The chickens of the latter group were the last to start dying from Marek's disease--only after the 10th week of life. In the chickens which had been vaccinated and revaccinated with sample (2) the mortality was not reduced. The study is continued, with particular emphasis on the relationship of DEAE-dextran to protection against Marek's disease.     

Major histocompatibility complex class I is downregulated in Marek's disease virus infected chicken embryo fibroblasts and corrected by chicken interferon

The major histocompatibility complex (MHC) is a part of the immune system which presents epitopes of intracellular antigens on the cell surface. MHC molecules have receptor-ligand binding affinities with T lymphocytes, permitting the latter to detect foreign intracellular infectious agents. Some pathogens, such as herpesviruses, have developed strategies of evading the host response by MHC. This pressure on the immune system brought, in turn, improvements in the antigen-presenting pathway, for example through the effect of interferon (IFN), which can upregulate MHC expression. The main objective of this work was on the one hand, to determine the abilities of three strains of Marek's disease virus (MDV), a chicken herpesvirus, in interfering with the expression of MHC class I molecules in chicken embryo fibroblasts. On the other hand, we analyzed the ability of IFN to reinstate this important immune capability to the infected cells. Our results show that only an oncogenic serotype 1 strain of MDV (RB1B) was able to markedly decrease MHC class I expression, and that addition of IFN reversed this MDV effect.     

Effect of acyclovir on the replication of turkey herpesvirus and Marek's disease virus

The toxicity of acyclovir for chick embryo fibroblasts and its effect on the replication of turkey herpesvirus (strain FC 126) and Marek's disease virus (strain HPRS 16) multiplied on fibroblast culture was studied. The influence of using acyclovir on the development of the tumour process in birds infected with a virulent Marek's disease virus was also determined. Acyclovir used in doses below 12.5 micrograms ml-1 proved to be nontoxic for chick embryo fibroblast culture. It inhibited in vitro replication of turkey herpesvirus and Marek's disease virus. It was also shown to diminish the development of tumours in birds infected with Marek's disease virus.     

Effect of diluting Marek's disease vaccines on the outcomes of Marek's disease virus infection when challenged with highly virulent Marek's disease viruses

Dilution of Marek's disease (MD) vaccines is a common practice in the field to reduce the cost associated with vaccination. In this study we have evaluated the effect of diluting MD vaccines on the protection against MD, vaccine and challenge MD virus (MDV) kinetics, and body weight when challenged with strains Md5 (very virulent MDV) and 648A (very virulent plus MDV) by contact at day of age. The following four vaccination protocols were evaluated in meat-type chickens: turkey herpesvirus (HVT) at manufacturer-recommended full dose; HVT diluted 1:10; HVT + SB-1 at the manufacturer-recommended full dose; and HVT + SB-1 diluted 1:10 for HVT and 1:5 for SB-1. Vaccine was administered at hatch subcutaneously. One-day-old chickens were placed in floor pens and housed together with ten 15-day-old chickens that had been previously inoculated with 500 PFU of either Md5 or 648A MDV strains. Chickens were individually identified with wing bands, and for each chicken samples of feather pulp and blood were collected at 1, 3, and 8 wk posthatch. Body weights were recorded at 8 wk for every chicken. Viral DNA load of wild-type MDV, SB-1, and HVT were evaluated by real time-PCR. Our results showed that dilution of MD vaccines can lead to reduced MD protection, reduced relative body weights, reduced vaccine DNA during the first 3 wk, and increased MDV DNA load. The detrimental effect of vaccine dilution was more evident in females than in males and was more evident when the challenge virus was 648A. However, lower relative body weights and higher MDV DNA load could be detected in chickens challenged with strain Md5, even in the absence of obvious differences in protection.     

Immunogenetics and the major histocompatibility complex.

The poultry immune system is a complex system involving many different cell types and soluble factors that must act in concert to give rise to an effective response to pathogenic challenge. The complexity of the immune system allows the opportunity for genetic regulation at many different levels. Cellular communication in the immune response, the production of soluble factors, and the rate of development of immune competency are all subject to genetic influences. The genes of the major histocompatibility complex (MHC) encode proteins which have a crucial role in the functioning of the immune system. The MHC antigens of chickens are cell surface glycoproteins of three different classes: Class I (B-F), Class II (B-L) and Class IV (B-G). The MHC antigens serve as essential elements in the regulation of cell-cell interactions. The MHC has been shown to influence immune response and resistance to autoimmune, viral, bacterial and parasitic disease in chickens. The MHC has been the primary set of genes identified with genetic control of immune response and disease resistance, but there are many lesser-characterized genes outside of the MHC that also regulate immunoresponsiveness. Polygenic control has been identified in selection experiments that have produced lines of chickens differing in antibody levels or kinetics of antibody production. These lines also differ in immunoresponsiveness and resistance to a variety of diseases. Understanding the genetic bases for differences in immunoresponsiveness allows the opportunity selectively to breed birds which are more resistant to disease. Indirect markers that can be used for this selection can include the MHC genes and immune response traits that have been associated with specific or general resistance to disease.     

不同马立克氏病抗性鸡攻毒后IL-18基因的差异表达

本试验致力于评价鸡白细胞介素-18(IL-18)基因的表达模式与马立克氏病抗性遗传性状的相关性。MDV-1分别感染7日龄普通品系和抗性品系的霞烟鸡,攻毒后第4、7、10、14、21、28、35天采集外周血并分离外周血淋巴细胞,利用实时定量PCR技术对外周血淋巴细胞中IL-18转录水平进行相对定量分析。结果表明,普通霞烟鸡外周血淋巴细胞中具有相对高水平的IL-18mRNA,而马立克氏病抗性霞烟鸡外周血淋巴细胞中则具有相对低水平的IL-18mRNA。结果显示,鸡IL-18基因的表达模式与马立克氏病(MD)抗性遗传性状可能相关。     

Oral infection with chicken anemia virus in 4-wk broiler breeders: lack of effect of major histocompatibility B complex genotype

The pathologic consequences of chicken anemia virus (CAV) oral inoculation in 4-wk-old broiler breeders of different major histocompatibility B complex (MHC) genotypes were evaluated. MHC B complex was determined by hemagglutination and sequence-based typing. Clinical signs, serology, gross lesions, histopathologic analysis, and CAV genome quantification were used to evaluate disease progression. Clinical disease was not apparent in the inoculated broilers throughout the experimental period. At 14 days postinoculation, antibodies against CAV were detected in 26.4% (29/110) of the inoculated birds. The distribution of percent positive was 34.6% (9/26) and 32.3% (10/31) of the chickens with B A9/A9 and B A9/A4 MHC genotypes, respectively, and seroconversion in six other genotypes was 19% (10/53). These differences among MHC genotypes for specific seroconversion rate were not statistically significant. CAV genomes were detected in the thymus of 87.7% (93/110) of the inoculated birds with no statistically significant differences between MHC genotypes. Mild thymic lymphocytolysis, lymphedema, and medullary hemorrhage were observed in the inoculated chickens. Histomorphometric analysis showed that cortical lymphocyte-to-parenchyma ratios did not differ between inoculated and uninoculated groups or among MHC genotypes. Similar findings have been reported previously in white-leghorn chickens of similar age, suggesting that broilers show a similar resistance to the effects of CAV infection at this age. The absence of significant clinical and pathological changes in the orally inoculated broilers at this age contrasts with CAV-associated thymus damage seen frequently in condemned commercial broilers at harvest.     

Marek's disease in turkeys. II. Characterization of the viral glycoprotein B gene and antigen of a turkey strain of Marek's disease virus

Marek's disease virus (MDV) causes immunosuppression and tumors in chickens. As sporadic cases of Marek's disease (MD) were recorded in turkeys, the antigenic and genomic characteristics of the MDV glycoprotein B (gB) gene and antigen of turkeys were compared to the chicken MDV gB. The whole chicken and turkey gB genes were sequenced and found identical. By immunoblotting of infected-cell culture lysates using chicken convalescent and gB monoclonal antibodies, the antigenic epitopes of the chicken and turkey viruses were found to differ. The turkey MDV had a unique epitope, compared to the chicken MDV and compared with our previous findings. While the chicken MDV had two epitope types, heat-labile but dithiothreitol (DTT)-stable and heat-stable but DTT-labile, the turkey MDV gB epitope is both heat and DTT-labile.     

Influences of major histocompatibility complex class I haplotypes on avian influenza virus disease traits in Thai indigenous chickens

Natural infections with influenza viruses have been reported in a variety of animal species including humans, pigs, horses, sea mammals, mustelids and birds. Occasionally, devastating pandemics occur in domestic chickens (broiler and layers) and in humans. From November 2003 to March 2004 in many countries in Asia, there were outbreaks of H5N1 avian influenza virus, causing death of infected patients, and devastating the poultry industry. Some groups of Thai indigenous chickens survived and were therefore classified as resistant. These traits were related to immunogenetics, in particular, the major histocompatibility complex (MHC) class I and class II molecules. The chicken MHC class I were investigated as candidate genes for avian influenza virus disease resistance. Seven hundred and thirty Thai indigenous chickens from smallholder farms in the rural area of avian influenza virus disease outbreaks in the central part of Thailand were used in the present study. They were separated into two groups, 340 surviving chickens and 390 dead chickens (resistant and susceptible). Genomic DNA were precipitated from blood samples and feathers. The DNA were used to amplify the MHC class I gene. Data were analyzed using χ² analysis to test significant differences of influences of MHC class I haplotypes on avian influenza virus disease traits. The results represented nine MHC class I haplotypes: A1, B12, B13, B15, B19, B21, B2, B6, and BA12, and included 10 of their heterozygotes. The homozygous B21 from these collected samples had a 100% survival rate and they were the major survival group. In addition, the heterozygous B21 also had a high survival rate because of co‐dominant expression of these genes. In contrast, the homozygous B13 had a 100% mortality rate and they were the major mortality group. These results confirmed that MHC class I haplotypes influence avian influenza virus disease‐resistant traits in Thai indigenous chicken. The MHC genes can be used as genetic markers to improve disease‐resistant traits in chicken.     

Quantitative measures of disease in broiler breeder chicks of different major histocompatibility complex genotypes after challenge with infectious bursal disease virus

Criteria for evaluating genetic differences in resistance and susceptibility to infectious bursal disease (IBD) within a commercial broiler breeder line of chickens were compared. Line A broiler breeder chickens were challenged with graded doses of Animal and Plant Health Inspection Service (APHIS) strain IBD virus (IBDV) and evaluated at 2 time points, 3 days postinoculation (PI) and 10 days PI. Measures obtained at both time points included bursa to body weight, bursa histology, bursa lymphocyte count, and percentage of T cells in the bursa. Furthermore, viral load in the bursa was determined 3 days PI and anti-IBDV antibody titers, 10 days PI. A dose of 50 50% embryo infective dose caused IBD in about half the line A birds at the 10-day time point, and this dose was chosen for further studies. The data were analyzed for correlation among the various measures. Comparison of the 3-day- and 10-day-PI bursa lymphocyte counts indicated that birds challenged with low doses of virus suffered lymphocyte depletion at the 3-day time point, but many or all (depending on the dose) recovered by the 10-day time point. With a viral dose that caused bursal atrophy in about half the birds by 10 days PI, families segregating for 2 major histocompatibility complex (MHC) haplotypes were compared in terms of resistance to IBD. Results indicated that there was no difference among the 3 MHC genotypes in incidence of IBD by any of the disease measures.     

Homologies between the major histocompatibility complex of man and cattle: Consequences for disease resistance and susceptibility

Summary

The major histocompatibility complex (MHC) of mammals number of mostly duplicated contains a large genes. In the HLA system (the MHC of man), which is by far the best‐studied major histocompatibility system so far, roughly 20 genes have been defined and mapped. They code for three classes of proteins: HLA‐A, ‐B and ‐C (Class I), HLA‐DP, ‐DQ and ‐DR (Class II) and serum complement components C2, C4 and Bf (Class III). Furthermore, the region contains genes for 21‐hydroxylase (21‐OH) and tumor necrosis factor (TNF).

The MHC thus forms a chromosomal segment containing seva‐al clusters of genes of only partially defined biological significance, but ondoubtedly playing a role in disease suscepti‐ bility. In view of the recently obtained structural information on BoLA, the MHC of cattle, it is hypothesized that susceptibility to diseases in cattle is associated with BoLA in thesame way as human diseases.

Finally, new technical and conceptual developments in the field of MHC research and their application to the BoLA system are discussed.     

Homologies between the major histocompatibility complex of man and cattle: consequences for disease resistance and susceptibility

The major histocompatibility complex (MHC) of mammals contains a large number of mostly duplicated genes. In the HLA system (the MHC of man), which is by far the best-studied major histocompatibility system so far, roughly 20 genes have been defined and mapped. They code for three classes of proteins: HLA-A, -B and -C (Class I), HLA-DP, -DQ and -DR (Class II) and serum complement components C2, C4 and Bf (Class III). Furthermore, the region contains genes for 21-hydroxylase (21-OH) and tumor necrosis factor (TNF). The MHC thus forms a chromosomal segment containing several clusters of genes of only partially defined biological significance, but ondoubtedly playing a role in disease susceptibility. In view of the recently obtained structural information on BoLA, the MHC of cattle, it is hypothesized that susceptibility to diseases in cattle is associated with BoLA in the same way as human diseases. Finally, new technical and conceptual developments in the field of MHC research and their application to the BoLA system are discussed.     

Effects of reticuloendotheliosis virus and Marek's disease virus infection and co-infection on IFN-gamma production in SPF chickens

Two experiments were used to examine the potential role of IFN-gamma in chickens infected with reticuloendotheliosis virus (REV) and Marek's disease virus (MDV). First, chickens were infected with REV and/or MDV at 5 days of age and examined from 3 to 50 days post-infection (dpi). In REV+MDV co-infection chickens, IFN-gamma ELISA demonstrated a 3-fold increase at 7 dpi compared to the controls, while REV alone caused a 5-fold increase, the IFN-gamma levels peaked, and then gradually decreased. IFN-gamma levels significantly decreased in MDV infection at 3 dpi and 15 dpi. Second, experiments were designed to determine the effects of different viruses and ConA on IFN-gamma production. For REV- or MDV-infected chickens, the IFN-gamma levels decreased slightly after adding ConA. This is the first report of IFN-gamma production in SPF chickens infected with REV and MDV measured by directly quantitative method.