Biomedical subjects
Publications and source records attributed to L D Bacon.
A chicken MHC class I (B-F) cDNA from SPAFAS line 11 embryonic liver tissue was isolated and characterized by nucleotide sequencing. Comparing this sequence with previously described B-F cDNAs highlights clustered nucleotide substitutions in exon 3, encoding amino acids located on the alpha-helical region of the alpha 2 domain.
The objectives were to investigate whether or not B haplotypes influence vaccinal immunity against Marek's disease (MD) in commercial chickens and to evaluate whether retrospective analysis would detect the influence. This method involved evaluating the B haplotypes of turkey herpesvirus (HVT)-vaccinated sick vs normal chickens from a flock afflicted with MD symptoms. An analysis of the retrospective data disclosed that MD symptoms were present in a higher proportion of B2B19 than B2B21 chickens. A prospective study was then conducted with blood-typed chickens of the strain vaccinated with HVT or HVT + 301B bivalent MD vaccines prior to inoculation of the very virulent Md5 virus. The bivalent vaccine provided better protection than HVT alone, but with either vaccine fewer B2B21 chickens developed MD lesions. We conclude that the B haplotype influence on vaccinal immunity against MD previously demonstrated in B-congenic strains of chickens is also significant in commercial chickens and that the influence can be detected through analysis of B haplotypes in sick vs normal chickens of an affected flock.
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A turkey subline at the Ohio Agricultural Research and Development Center was developed by DNA typing of the MHC using a chicken MHC Class II probe, and it segregated for specific MHC genotypes. Histocompatibility was examined between turkeys of known MHC genotype using skin graft procedures, mixed lymphocyte reactions (MLR), and graft-versus-host reactions (GVHR). Skin grafts were exchanged among 3-wk-old turkeys and it was found that when birds shared DNA patterns (genotypes), the skin grafts were usually accepted. In contrast, skin grafts were always rejected when birds did not share the identical DNA pattern. Similarly, MLR only occurred when the lymphocytes were derived from birds that did not share the same DNA pattern. Lastly, GVHR were examined in embryos injected with either sire or dam blood. The GVHR in embryos was dependent on the parental MHC genotype. Four MHC haplotypes were identified in the turkey subline. The turkey MHC has been designated MhcMega-B, and each of the haplotypes, Mega-B(1) through Mega-B(4).
Chickens of the B2, B5, B15, B19, and B21 B-congenic haplotypes differ in disease resistance. Complementary DNA from B-congenic chicken strains have been analyzed for allelic diversity of expressed Class II MHC genes. The predicted amino acid sequences of eight genes from five haplotypes were subjected to Wu-Kabat variability analysis. The B-L gene polymorphic regions and conserved regions are highly similar to the human leukocyte antigen Class II genes. Therefore, the present analysis reveals candidate polymorphisms important in determining the spectrum of antigenic peptides presented to T helper cells, and allelic differences possibly important in resistance to avian disease.
Coccidia antigen-specific T lymphocyte proliferation induced by Eimeria acervulina was measured in chickens congenic at the major histocompatibility complex and in two unrelated lines. Sporozoites and merozoites induced high proliferation of lymphocytes following primary infection, with similar changes seen for splenic or peripheral blood lymphocytes. Highest antigen-specific proliferation after both primary and secondary infection was seen in Line 15I5, which contributed the background genome to the congenic lines. In general, merozoites elicited higher proliferation responses than sporozoites. Analyses of differences between uninfected and infected chickens showed that, at some points in the infection cycle, proliferation of lymphocytes was greater and at other times less for infected than uninfected chickens.
Genetic selection for early humoral immune responsiveness against two simultaneous antigens, namely, heat-killed Escherichia coli and Newcastle disease virus vaccine (NDV), was performed in a heterogenic population of broiler chickens. The humoral immune response was measured prior to 24 days of age, depending on the vaccine. Chicks were divided into two populations according to their antibody response: a population of high responders to both antigens and a population of low responders. After four generations of selection, a significant difference was observed in the immune response of the two populations, and therefore, an attempt was made to identify possible differences in the segregation of major histocompatibility complex genes. Using alloantisera prepared in B-congenic White Leghorn, the high responders exhibited a high percentage of chickens having erythrocytes agglutinated with B5 antisera, and the low responders exhibited a high percentage of chicks with erythrocytes agglutinated with B15 and B19 antisera. The B13 antisera agglutinated cells of similar proportions of chickens in the two populations. A random commercial broiler flock was screened with the antisera, and correlation was high between B haplotype and antibody level to E. coli following vaccination. The present work indicated the possibility of a direct association between the genetic characteristics represented in the B complex and humoral antibody response in a broiler population.
The present review briefly outlines the effector cells and molecules influencing cell-mediated immunity. Emphasis is placed on the genes determining these molecules in mammals and in the chicken, including genes for cytokines and cytokine receptors. The review concludes with a tabular presentation containing examples of specific genetic influences on cell-mediated immunity in chickens and mammals.
The influence of the chicken major histocompatibility (B) complex (MHC) on monocyte and macrophage recruitment and activation was examined using fully developed 15I5-B congenic White Leghorn lines (ten backcross generations). The phagocytic activity of Sephadex-elicited peritoneal macrophages for sheep red blood cells (SRBCs) was highest in lines 15.7-B2 and 15.P-B13 and lowest in 15.15I-B5 and 15.N-B21. The same pattern of phagocytic activity was obtained when LPS (E. coli) was used as the in vivo elicitor-activator of peritoneal macrophages. Lines with B2 and B13 haplotypes had elevated percentages of phagocytic macrophages and a higher internalization activity per cell than did B5 and B21 congenic chickens. Differential peritoneal macrophage function between congenic lines was further supported by quantitation of superoxide anion release. B2 and B13 haplotypes were associated with high activity in contrast with B5, which was low, and 15I5 (B15) and B21 which were intermediate for superoxide anion release by macrophages. In vitro activation of blood monocytes with LPS resulted in similar line differences for SRBC phagocytic activity as were observed with in vivo Sephadex and LPS activation. In contrast, chemotaxis of blood mononuclear leukocytes to f-met-leu-phe produced a reciprocal response pattern among the haplotypes. Cells from lines with haplotypes B5 and B21 were superior to those of B2, B13, and B15 congenic lines in their directed migration towards this chemoattractant. All functional differences occurred despite similarities among lines in the cellular profiles of both elicited peritoneal exudate cells and isolated blood mononuclear cells.
Eight 15I5 B-congenic White Leghorn chicken lines, containing haplotypes B2, B5, B12, B13, B15, B19, and B21, were subjected to molecular genotyping with chicken B-F (Class I) and B-L (Class II) major histocompatibility complex (MHC) probes. Genomic DNA was digested with restriction enzymes, hybridized with a Class I or Class II probe, and analyzed for restriction fragment length polymorphisms. Digestion with HindIII or EcoRI yielded no B-L polymorphisms. Digestion with PvuII or BglII and hybridization with a B-L or B-F probe produced polymorphisms that were shared between several haplotypes, although the haplotypes with similar patterns were clustered differently between Class I and Class II probes. The genetic variation seen for B-L and B-F probe hybridization of PvuII digests was much less than that previously demonstrated for B-G probing of PvuII digests of the same lines. Description of MHC Class I and II restriction patterns of the well-characterized 15I5 B-congenic lines will aid in identification of genes important in disease resistance.
Intramuscular immunization with the recombinant p250 surface antigen of Eimeria acervulina merozoite (rEAMZp250) or oral inoculation with live recombinant Escherichia coli expressing the rEAMZp250 protein resulted in antigen-specific T-cell and humoral responses and conferred a significant reduction in mucosal parasitism compared to immunization with the negative control antigen preparation. Among the major histocompatibility complex (MHC) (B)-congenic chickens receiving intramuscular immunization, strain .6-2 (B2-B2) showed significant (P less than 0.05) protection to live E. acervulina challenge compared to the other strains examined. In contrast, strains .C-12(B12B12) and .P-13 (B13B13) showed significant protection among the groups given live recombinant E. coli. In general, strains showing enhanced T-cell responses to the rEAMZp250 protein were better protected compared to those showing minimal or no T-cell responses. Thus the results suggest that the B haplotypes of the host and the mode of antigen presentation influence the outcome of protection following an immunization of chickens with recombinant coccidial antigen.
The relative importance of MHC genes and background genes in the genetic control of disease susceptibility and the development of protective immunity to E. tenella infection was investigated in eight different strains of 15I5-B congenic and four inbred chicken strains. RPRL 15I5-B congenic chickens that share a common genetic background but express different B haplotypes demonstrated wide variations in disease susceptibility and the development of acquired resistance to E. tenella infection. Infection of chickens sharing a common B haplotype but expressing different genetic backgrounds showed quite contrasting levels of susceptibility to secondary E. tenella infection. In all chicken strains examined, infected chickens developed high levels of serum and biliary anti-coccidial antibodies regardless of their B haplotypes. Furthermore, no correlation between antibody levels and the phenotypically expressed levels of disease resistance was demonstrated. These findings lend support to the view that interaction of MHC genes and non-MHC genes influences the outcome of host response to E. tenella infection.
Our objective was to determine whether the cell-associated herpesvirus vaccines used in chickens to control Marek's disease tumors can augment development of lymphoid leukosis (LL) induced by exogenous avian leukosis virus (ALV). Various single or mixed Marek's disease vaccines were inoculated at day 1, and ALV was injected at 1 to 10 days, with chickens of several experimental or commercial strains. Development of LL was monitored at 16 to 48 weeks in various experiments. In several strains of chickens we repeatedly found that the widely used serotype 3 turkey herpesvirus vaccine did not augment LL in comparison with unvaccinated controls. However, LL development and incidence were prominently augmented in several chicken strains vaccinated with serotype 2 vaccines, used alone or as mixtures with other serotypes. In one chicken strain, augmentation was demonstrated after natural exposure to ALV or serotype 2 Marek's disease virus viremic shedder chickens. Augmentation of LL by virulent or attenuated Marek's disease viruses of serotype 1 was intermediate in effect. Serotype 2 Marek's disease virus augmentation of LL was prominent in three laboratory lines and one commercial strain of White Leghorns, but it was not observed in an LL-resistant laboratory line or four commercial strains susceptible to ALV infection. Chickens developed similar levels of viremia and neutralizing antibodies to ALV regardless of the presence of augmentation of LL, suggesting that the mechanism of enhanced LL did not result from differences in susceptibility or immune response to ALV. We postulate that the serotype 2 herpesviruses may augment LL through one of several possible influences on bursal cells that are subsequently transformed by exogenous ALV.
Six Ea-B-congenic lines of chickens were used to study the role of the B histocompatibility complex in susceptibility and immunity to Eimeria tenella and E. acervulina. The B haplotypes were 2, 2, 5, 12, 13, or 19. Parental line 15I5 with the B15 haplotype (15I5-15) and a commercial White Leghorn (CWL) line were also used. The B-congenic lines were generally less susceptible to E. tenella than the CWL line based on weight gain, cecal lesion score, plasma pigment, and packed cell volume. Within the congenic lines, the 15I5-15 line was more resistant to E. tenella than the 15.6-2 and 15.7-2 lines. Using a level of infection of E. acervulina resulting in moderate disease, the 15.7-2 line was more susceptible than all other lines based on weight gain, and the 15.6-2 and 15.7-2 lines had greater intestinal lesion scores than the 15.15I-5 or 15I5-15 lines. The CWL line was most susceptible based on lesion score. Thus 15I5-15 chickens were more resistant to primary infection by several criteria than the 15.7-2 or 15.6-2 chickens infected with either coccidial species. A single immunization with 100 oocysts of E. tenella produced less immunity in the congenic lines than in the CWL, whereas four immunizations resulted in immunity in all lines except 15.15I-5. Immunization with E. acervulina produced good immunity to a challenge infection in all lines. These results suggest that the B complex has a detectable influence on resistance and susceptibility to avian coccidiosis, but it may play only a minor role in the development of immunity to a challenge infection.
The roles of major histocompatibility complex (MHC) and non-MHC-linked genes in the genetic control of disease susceptibility and the development of protective immunity to Eimeria acervulina infection were investigated in six 15I5-B congenic and four different strains of chickens characterized for the MHC. When oocyst production was assessed, wide variations were noted following initial and challenge infections among the strains of chickens tested. In general, 15.N-21, 15.P-13, B21, B19, SC, and FP chickens were protected following challenge infection whereas 15I5, 15.P-19, 15.7-2, and 15.6-2 chickens were not. Strains of chickens sharing a same B haplotype on different genetic backgrounds did not show comparable levels of protection. These results lead to the view that non-MHC-linked genes have a profound influence on the outcome of the host response to E. acervulina infection. Chickens infected twice at 1-month intervals by an oral inoculation with E. acervulina developed both coccidial-specific antibody and T-cell responses. E. acervulina infected chickens showed T-cell-mediated immune responses to the intact sporozoites as well as to recombinant proteins, p130 of sporozoites and p150 of merozoites. Both p130 and p150 antigens have been identified and characterized previously. Sera obtained from all infected chickens recognized the p150 merozoite protein, but not the p130 sporozoite protein in immunoblots. In general, the cellular response, but not the antibody response to the p150 recombinant surface merozoite antigen correlated with the degree of protection following the challenge infection. These results suggest that the strains of chickens having improved protection against challenge infection demonstrate higher T-cell responses to the recombinant surface merozoite protein, p150.
The chemotactic activity of chicken blood mononuclear leukocytes was examined in partially-developed 15I5-B-congenic chicken lines using Enterobacter cloacae culture supernatant and formyl-methionyl-leucyl-phenylanine (f-met-leu-phe) (10(-5)M). Cells from seven different coded B-congenic lines were used to study each chemoattractant in vitro. Mononuclear cells from lines .15I-B5,.C-B12 and the background line 15I5 (B15) exhibited a significantly greater directed migration to bacterial supernatant than did cells from four lines carrying the B2, B2, B13, and B19 haplotypes, respectively. Similarly response to f-met-leu-phe was greatest in lines .15I-B5,.C-B12 and .N-B21, with the same four lines exhibiting a significantly lower response. Since f-met-leu-phe was originally isolated from bacteria, the results indicate that these lines possess differential chemotactic responses to certain bacterially-derived chemoattractants. Major histocompatibility complex (MHC) differences between the lines may serve as a genetic basis for the differential responses. Extrapolation of these results to other chemotactic-receptor systems would require further examination.
A dominant sex-linked gene, K, regulates slow feathering (SF), whereas a recessive allele, k+, determines rapid feathering (RF) in chickens. This trait provides a convenient and inexpensive approach to gender identification of White Leghorn (WL) chicks at hatch, i.e., in a sex-linked mating using k+/k+ males mated with K/- females, the K/k+ male chicks are SF, and the k+/- females are RF. However, in many WL strains, female progeny of SF dams produce fewer eggs and have higher mortality than progeny of RF dams. This loss in productivity has been attributed to higher infection and shedding rates for leukosis viruses (ALV) in SF than in RF dam lines. Because infectious endogenous viruses (EV) can induce immunological tolerance to ALV, we examined the expression and distribution of ev genes in SF and RF siblings from heterozygous K/k+ sires and k+/- dams. Infectious ALV and EV were detected by cocultivation of frozen heparinized blood cells on selected chick embryo fibroblasts and culture supernatants were tested for viral antigen by enzyme-linked immunosorbent assay tests. Specific ev genes were identified as restriction fragment length polymorphisms after hybridization with a recombinant plasmid containing the complete genome of a Rous-associated virus. It was concluded that ev21 and K genes are tightly linked because, in different WL crosses, all SF chicks inherited ev21 but RF siblings uniformly lacked ev21. Alternatively, the K gene in WL may be a mutation resulting from the insertion of ev21 in the k+ gene. The SF chicks which harbor ev21 expressed infectious EV21; evidence that EV21 may influence susceptibility to ALV is presented.
A monoclonal antibody (MAb), designated P2M11, that detects a monomorphic determinant of chicken class II antigens was produced from the fusion of P3X63 myeloma cells with spleen cells from BALB/c mice immunized with chicken splenic lymphocytes. Flow cytometric analyses of lymphocytes from the SC and FP strains of chickens showed 30 to 50% staining of bursa cells, 15 to 20% staining cells, and less than 5% staining of thymus cells. Addition of MAb P2M11 to splenic of T cell cultures stimulated with allogeneic cells or concanavalin A resulted in a significant inhibition of the T cell proliferation responses. Immunoprecipitation of 35S-methionine-labeled spleen cell extracts using MAb P2M11 identified molecules with apparent molecular weights of approximately 28,000, 30,000, and 32,000 by sodium dodecyl-polyacryl-amide gel electrophoresis. Taken together, these data indicate that the antigens detected by MAb P2M11 are similar in cell distribution and structure to chicken Ia antigens encoded by B-L genes. Using this MAb, a strain difference was demonstrated in the tissue distribution of Ia antigen positive lymphocytes in the spleens but not the thymuses of 15I5-B congenic and inbred strains of chickens. This difference may be due to the genes associated with B-complex genes.