Isolation and characterization of cDNA clones for chicken major histocompatibility complex class II molecules.

The chicken major histocompatibility complex (MHC), the B complex, is being intensively analysed at the DNA level. To further probe the molecular structure of chicken MHC class II genes, cDNA clones coding for chicken MHC class II (B-L) beta chain molecules were isolated from an inbred G-B2 Leghorn chicken spleen and liver. Twenty-nine cDNA clones were isolated from the spleen and eight cDNA clones were isolated from the liver. Based on restriction maps, most clones could be clustered into one family of genes. Four cDNA clones were sequenced (S7, S10 and S19 from the spleen and L1, which was identical to S19, from the liver). Complete amino acid sequences of B-L beta chain molecules were predicted from the nucleotide sequences of the cDNA clones. Although both the nature and the location of the conserved residues were similar in chicken and mammalian sequences, some species-specific differences were found, suggesting that the structures of the B-L molecules of this haplotype are similar, but not identical, to their mammalian counterparts.

Selecting for efficiency of egg production using food-consumption records in layer-type chickens.

1. The purpose of the experiment was to evaluate the use of food consumption records in selection to improve layer-type chickens. 2. Twelve subpopulations of Leghorns, including 5878 tested birds, were used in a selection experiment which lasted 5 generations. Four were selected using an index (I1) that combined information on body weight (BW), egg-mass output (EM), and individual food-consumption records (FC). Four were selected using an index (I2) that combined information just on BW and EM. The remaining 4 served as controls, with selection differentials set to zero in each generation. 3. Differences between the selected groups, control-corrected each generation, measured the "food-record effect". 4. Including food-consumption records in the selection criterion resulted in increased body weight (7%), reduction in food consumption (6 to 9%), increase in food efficiency (egg mass/food consumption) (2 to 5%), and increase in income over food costs (10%), relative to selection not involving food records.

Isolation and characterization of three class II MHC genomic clones from the chicken.

A genomic library was constructed from sperm DNA from an individual of the inbred chicken line G-B2, MHC haplotype B6. The library was screened with a chicken class II probe (beta 2 exon specific) and three MHC class II beta chain genomic clones were isolated. The restriction maps of the three clones showed that each of the three clones was unique. The position of the beta chain sequence was located in each of the three genomic clones by Southern blot hybridization. Subclones containing the beta chain gene were produced from each of the genomic clones and the orientation of the leader peptide, beta 1, beta 2, transmembrane, and cytoplasmic exons was determined by Southern blot hybridization and nucleotide sequencing. The complete nucleotide sequence of two of the three subclones was determined. Comparison of the nucleotide and predicted amino acid sequences of the two subclones with other class II beta chain sequences showed that the B6 chicken beta chain genes are evolutionarily related to the class II beta chain genes from chickens of other MHC haplotypes, and to class II beta chain genes from other species. Analysis of Southern blots of B6 chicken DNA, as well as the isolation of the three beta chain genes, suggests that chickens of the B6 haplotype possess at least three MHC class II beta chain genes.

Analysis of B-G and immune response genes in the Iowa State University S1 chicken line by hybridization of sperm deoxyribonucleic acid with a major histocompatibility complex class II probe.

Sperm DNA was isolated from chickens of the Iowa State University S1 line. Birds were from sublines selected for B-G antigen, humoral immune response to glutamic acid-alanine-tyrosine (IrGAT), and response to Rous sarcoma virus-induced (RSV) tumors. The DNA was digested with restriction enzymes and subjected to Southern blot analysis with a DNA probe specific for a class II gene of the chicken major histocompatibility complex (MHC). Restriction fragment length polymorphisms (RFLP) associated with B-G antigen type were found after digestion of DNA with three enzymes, PvuII, BglII, or Sau3A, out of a total of 15 tested. No RFLP were shown to be associated with IrGAT or RSV type. This study shows that RFLP analysis of DNA may be a useful addition to or alternative to serological evaluation of MHC haplotype in the chicken.

Inbreeding effects on fertility and hatchability associated with the formation of sublines.

The deleterious effect of inbreeding on fertility and hatchability was studied in a population of Leghorns developed primarily for immunogenetics studies. The 1983 population averaged 42% inbreeding and traced back to only two sires and five dams of the 1965 base population. Up to 1975 (Period 1), the population was maintained as a single-unit closed flock, during which time inbreeding increased an average of 1.44% per year. After 1975 (Period 2), the population was split into five sublines. Inbreeding then increased at the average rate of 2.7% per year. The regression of fertility on inbreeding was -.31 +/- .04 in Period 1 and -.17 +/- .06 in Period 2. Corresponding values for hatchability of fertile eggs were +.08 +/- .05 in Period 1 and -.24 +/- .07 in Period 2. The regression of hatchability of total eggs set was -.15 +/- .04 in Period 1 and -.35 +/- .10 in Period 2. The evidence implies that both fertility and hatchability have a heritable basis. As expected, the deleterious effects of inbreeding on reproduction was more severe in Period 2, with sublining.

Research note: differences in major histocompatibility complex gene frequencies associated with feed efficiency and laying performance.

Associations between egg production and feed efficiency and the genes of the major histocompatibility complex (MHC) of the chicken were investigated. Breeders were selected on the basis of indexes incorporating information on body weight and egg mass, with or without feed consumption information. Over 1,000 progeny were serotyped for the erythrocyte antigen B (Ea-B) after the 6th generation of selection. The B2 and B13 haplotypes accounted for over 75% of the Ea-B gene pool in all lines. Comparing each index-selected line with the control B2 significantly increased in frequency at the expense of B13. This study further implicates MHC-linked genes as important in the physiology of growth and reproduction in the chicken.

Molecular analysis of the chicken major histocompatibility complex gene and gene products.

New technologies are enabling researchers to conduct analysis of the molecular structure of the genes and gene products of the major histocompatibility complex (MHC). Two of these new technologies are the production of monoclonal antibodies by hybridomas and analysis of deoxyribonucleic acid (DNA) by restriction fragment length polymorphisms (RFLP) on Southern blots. Monoclonal antibodies, because of their exquisite specificity, can be used to isolate purified MHC antigens for further study. The RFLP analysis can be used to examine differences in the MHC at the DNA level. This paper describes the two aforementioned technologies and their application to molecular analysis of the chicken MHC.

Subregions and functions of the chicken major histocompatibility complex.

The chicken major histocompatibility complex (MHC) exerts genetic influence over a variety of important biological functions including immune response, disease resistance, growth and development, aging, and reproduction. The chicken MHC possesses at least three subregions encoding distinct gene products. The B-G subregion encodes antigens unique to erythrocyte surfaces. The B-L and B-F subregions encode cell surface glycoproteins homologous to mammalian Class II and Class I antigens, respectively. Class I and Class II molecules are crucial for recognition of self vs. nonself and for cell communication, and therefore are fundamental for all immune responses. Studies of chromosomal recombinants have been particularly useful in eliciting the structure and function of subregions of the chicken MHC.

Association of Marek's disease with Ea-B and immune response genes in subline and F2 populations of the Iowa State S1 Leghorn line.

Chickens from the Iowa State S1 White Leghorn line, selected for characteristics of Ea-B serotype, humoral immune response to glutamic acid-alanine-tyrosine (Ir-GAT), and response to Rous sarcoma virus (RSV)-induced tumors, were evaluated for genetic resistance to Marek's disease (MD). In the first two trials, sublines that were triple homozygous for the three traits were challenged with MD virus. Birds of the B1B1 blood type were significantly (P less than .001) more resistant to MD than chickens of the B19B19 blood type. High responders to GAT were significantly (P less than .001) more resistant to MD virus than low responders. The RSV classification had no detectable association with MD resistance. Chickens challenged with MD virus in the third trial were an F2 population produced from inter se matings of S1 chickens heterozygous for the three traits under selection. Data from this trial confirmed increased MD resistance of chickens possessing the B1B1 blood type when associated with genes encoding high immune response to GAT.

A chicken sex-limited protein that crossreacts with the fourth component of complement.

Plasma samples from more than 300 inbred chickens were screened by using an immunofixation technique with antibody against the fourth component of complement (C4) from humans. Precipitation patterns of plasma from adult male and sexually immature birds, either male or female, were identical. Plasma from egg-laying hens demonstrated a distinctly different precipitation pattern compared with plasma of other birds, with one additional band appearing 14 to 9 days before production of the first egg. The banding pattern could not be induced in males by progesterone injection and remained unchanged in molted female birds.

Performance of layer-type chickens as related to body conformation and composition. 1. A static analysis of shank length and body weight at 20 weeks of age.

The growth of an animal part (y) relative to the whole (x), is expressed in terms of an exponential equation, y = alpha x beta, where alpha and beta are growth constants. For the present study x is body weight and y is shank length determined for 10 lines of chickens at 20 weeks of age. The derived values of alpha and beta, assumed to be limiting relative growth constants for bone, were estimated by least squares of the log-transformed (linear) exponential equation. From the means of alpha and beta, calculated for each of seven Leghorn and three Fayoumi lines, individual values of alpha i and beta i were calculated for more than 24,000 pedigreed birds included in 9 years of records. The individual values (i) were calculated as: (formula; see text) If the initial growth constant, alpha, is the primary hereditary growth parameter governing conformation, then the parameter, beta, should not be as highly heritable as alpha. To test this hypothesis, the heritability (h2) of both alpha and beta were calculated for each of the 10 lines. For alpha sire component h2 values ranged from .12 +/- .05 to .45 +/- .10 and for beta from .05 to .41. The pooled h2 estimate of alpha over lines (.31 +/- .02) was not significantly different from that for beta (.31 +/- .02). Compared with the original data on body weight and shank length, the transformation significantly reduced the heritability estimated. We conclude that the parameters, alpha and beta, are useful only in describing phenotypic and not genetic differences in relative growth.

Genetic linkage of subgroup C Rous sarcoma virus-induced tumour expression in chickens to the IR-GAT locus of the B complex.

Two B complex genotypes, B1B1 and B19B19, of outbred line S1, were tested for low and high immune response to GAT, from which four recombinants were recovered: B1B1 GAT-hi and -lo, and B19B19 GAT-hi and -lo. Also included in the study were birds of B2B2 genotype with an intermediate level of immune response to GAT. A total of 225 birds of these groups were challenged with the Bryan strain of Rous Sarcoma virus subgroup C, RSV (RAV-7), by inoculation into the wing web at five weeks of age. The B1B1 genotype had the lowest percentage of regressors (17.6%), B19B19 had the highest (42.2%), and the B2B2 genotype was intermediate (23.7%). Combining the results of GAT response over the B1B1 and B19B19 genotypes, 14.0% of GAT-lo and 37.8% of GAT-hi regressed their tumours, respectively. The highly significant (P less than or equal to 0.01) difference between the combined GAT-hi and -lo groups would suggest that the Rs locus controlling tumour regression induced by the subgroup C virus is closely linked to the region controlling immune response to GAT, but the data also provides evidence that the B-F region of the B complex also plays an important role in RSV-induced tumour regression.

Influence of B complex versus non B genetic background on cellular resistance and on Rous sarcoma virus-induced tumour regression in chickens.

Six of eight inbred lines, segregating for the same B complex alleles, permitted a comparison of genetic backgrounds on resistance to Bryan high titre Rous sarcoma virus (RSV) of subgroups A, B and C. Tests were based both on wing web challenge (WWC), using 682 five-week-old chicks, and on the chorioallantoic membranes (CAMs) of 606 twelve-day-old embryos (CAM test). Absence of tumour development on WWC and of pock formation on CAMs indicated cellular resistance to RSV (primary mechanism of genetic resistance). The differential response of tumours (regressive or progressive tumour growth in WWC) measured the secondary mechanism of genetic resistance. The results verified previously reported studies that the tv loci for cellular resistance map outside the B histocompatibility complex, and that variation in RSV-induced tumour regression was mostly but not entirely linked to the B complex. Although cellular resistance and tumour regression are not wholly independent events, the data generally support the dual mechanism hypothesis of resistance. The ratios of variance between lines with the same B serotype, sigma2L, to that between serotypes with common inbred line background, sigma2B, demonstrated that sigma5L were 3.2 and 17.3-fold larger than sigma2B, respectively, to the CAM and WWC tests of cellular resistance (tv genes). In contrast, sigma2B was 5-fold larger than sigma2L for tumour regression (rs genes). The results further indicate that segregation at both the tv and rs loci is greater than expected within these inbred lines in view of their high coefficients of inbreeding. Theoretical expectations show that for eight inbred lines, each with an inbreeding coefficient of 80% assuming initial gene frequencies of .5, the probability of complete gene fixation at the same tv locus is .43 and the probability that only one line is segregating is .38. Finally, consideration is given to the heritability of cellular resistance and tumour regression when viewed as quantitative traits in a selection programme. As expected, the heritable fraction of variance is substantial even when the frequency of an unfavourable dominant allele, as tv, is low.

Significance of body weight as a performance parameter.

The laying performance of different body weight classes, defined on both a genetic and phenotypic scale, were compared. The distribution of pedigreed Leghorn breeders of both sexes were truncated on the basis of 30 week body weight into two classes: light (L) or heavy (H). The progeny then were classified according to parental mating types as either L X L, L X H, or H X H. Differences among these progeny groups defined the genetic scale. The phenotypic scale was derived separately by phenotypic truncation of the progeny distribution into light (L), medium (M), and heavy (H) body weight classes at 30 weeks. Differences in rate of lay and in egg mass output between classes were in poor agreement with respect to the two scales. Light birds were poorest egg producers on the phenotypic scale but were the best egg producers on the genetic scale. Superior egg production by the heavy-bird class during the late spring and early summer months of the year could not be demonstrated. Deviations in body weight sharply reflected poorer performance on the phenotypic scale but not the genetic scale. This emphasizes the dominant role of management, feeding, and disease control over the genetic influence in maintaining optimum size for satisfactory productivity.

Genetic control of serum immunoglobulin G levels in the chicken.

Serum IgG (7S) levels differed significantly for chickens from 10 different inbred lines. Within lines differences between B blood groups were statistically significant. The genetic control of serum IgG was further examined using birds from B complex haplotypes marked at the B locus and the Ir-GAT locus. Birds from each of five subgroup haplotypes (B1B1 Ir-GAT-Lo and -Hi, B19B19 Ir-GAT-Lo and -Hi, and B2B2 Ir-GAT intermediate) were tested for levels of serum IgG at 3, 6, 9, and 21 weeks of age. The rate and level of IgG reached in the serum was more than two-fold greater in the GAT-Hi birds than in the GAT-Lo. The Ir region of the B complex exerts some control over the ontogenesis of IgG, though most of the genetic variation seems not to be B complex associated.

Biochemical polymorphic systems in inbred lines of chickens: a survey.

Biochemical polymorphic gene frequency profiles from blood samples of two outbred and seven inbred lines of chickens were studied for hemoglobin, albumin, transferrin, alkaline phosphatase, esterase II, and leucine aminopeptidase, and from egg samples of these lines for ovoalbumen, ovoglobulin (G2 and G3), and conalbumen. Complete gene fixation was found for hemoglobin, albumin, transferrin, ovalbumin, ovoglobulin, and conalbumin. The same alleles were fixed in each system in each line. For four systems, a particular allele within a system predominated in seven populations; gene frequencies ranged from 0.60 to 0.98. For esterases I and II, the genes Es-IB and Es-IIS ranged in frequency between 0.82 and 0.97, and between 0.64 and 0.93, respectively. For ovoglobulin, G2B ranged between 0.75 and 1.00 with four lines fixed for this allele. The rather remarkable similarity of gene frequency profiles among lines, several of which are only remotely related, suggests that certain characteristic polymorphic frequencies for these biochemical polymorphisms possess higher adaptive values in an evolutionary sense.

Immune response and disease resistance in chickens. II. Marek's disease and immune response to GAT.

Genetic control of resistance of susceptibility to Marek's disease in chickens was found to be associated with immune response to the amino acid polymer GAT. Thus, both B1B1 and B19B19 GAT low responders had significantly greater incidence of MD than the B1B1 and B19B19 GAT high responder counterparts. The results point to gene(s) controlling MD resistance mapping within the immune response region of the B complex, the major histocompatibility system of the chicken. Patterns of general mortality demonstrate that the low viability, observed over several years among B1B1 birds, likewise is associated with the immune response region of the B complex.

Immune response and disease resistance in chickens. I. Selection for high and low titer to Salmonella pullorum antigen.

A selection experiment for high and low anti-Salmonella pullorum antibody titer was carried out over four generations within the B1B1 blood group genotype in chickens. The study was aimed primarily at identifying different response patterns controlled by immune response genes linked to the B system, the major histocompatibility complex of the chicken. Maximal divergence was obtained in the third generation of selection when agglutination titers of 1/320 and 1/80 in B1B1 high and low responders, respectively, were reached. Immune response of S. pullorum was deduced to be controlled by polygenes. The B1B1 population, selected for high immune response to S. pullorum antigen, consistently had greater total mortality as well as greater susceptibility to challenge with Marek's disease virus compared with B1B1 population selected for low response. This, however, is believed to be a consequence of random drift of genes for disease resistance in the relatively small populations and not a direct consequence of selection for high or low S. pullorum titer.