SC Chickens Research Articles

Comparison of Disease Susceptibility and Subclass-Specific Antibody Response in SC and FP Chickens Experimentally Inoculated with Eimeria tenella, E. acervulina, or E. maxima

Susceptibility to disease and the subclass-specific antibody response to Eimeria tenella, E. acervulina, and E. maxima were compared in two inbred strains of chickens, FP (B15B21) and SC (B2B2). FP strain was more susceptible to coccidiosis than SC chickens based on oocyst production, lesion score, and clinical signs. FP chickens infected with E. tenella had more severe cecal lesions and a significantly lower hematocrit level than SC chickens. FP chickens infected with E. acervulina excreted five times as many oocysts at 6 days postinfection as SC and showed a 71% reduction in plasma carotenoid level compared with controls (56% reduction in SC chickens). Body-weight change did not correlate with other signs of disease. Both SC and FP chickens produced high levels of serum IgM and IgG and biliary IgA. Although SC chickens had a slightly higher antibody response than FP chickens at 7 days postinoculation, both strains maintained high levels of IgM, IgG, and IgA for a prolonged period post primary inoculation. Although SC and FP chickens show different disease susceptibility to coccidiosis, they demonstrate similar antibody response.

Selective integration of avian leukosis virus in different hematopoietic tissues

Hematopoietic tissues obtained from avian leukosis virus (ALV)-infected Hyline SC chickens were analyzed for the presence of integrated viral DNA sequences. Cells were prepared from bone marrow, bursa, spleen, thymus, and peripheral blood. Following the removal of erythrocytes, cellular DNAs from each of these tissues were examined by Southern analysis. During the first few weeks of infection, DNA from the bone marrow contained as many as 0.5 copies of viral DNA per haploid genome. Cells from the bursa and peripheral blood contained between 0.05 and 0.15 copies per haploid genome. In contrast, neither splenic nor thymic DNA contained significant levels of viral DNA sequences even though infected birds developed titers of circulating virus between 10 5 and 10 6 IU/ml of plasma. DNA prepared from erythrocytes isolated from the peripheral blood of these birds contained approximately 0.4 copies of integrated viral sequences per haploid genome at 2 weeks after infection. Despite greater levels of integrated sequences in other tissues, by 9 weeks after infection viral sequences were detected only in DNA from bursal lymphocytes. Cells prepared from the spleen and thymus of infected birds were also examined for their size distribution, their internal complexity and their surface expression of immunoglobulin. None of the populations examined differed from normal, uninfected control preparations. These results suggest that ALV infection occurs primarily in the bone marrow and that the different tissues of the hematopoietic system are selectively infected. Further, these results indicate that ALV infection persists longer in bursal lymphocytes than in other hematopoietic tissues. Previous studies have demonstrated that the lymphoid tumors that develop in white leghorn chickens following ALV infection are bursal-dependent B-cell lymphomas that express immunoglobulin M. The observations presented in this communication offer, in part, an explanation for the restricted phenotype of the lymphoid tumor that develops in the SC chicken. Further, the data suggest an explanation for the bursal-dependent nature of the ALV-induced lymphoma.

Immune response during coccidiosis in SC and FP chickens. I. [formula omitted][formula omitted] assessment of T cell proliferation response to stage-specific parasite antigens

Development of cell-mediated immunity (CMI) and comparative effectiveness of different stage-specific coccidia antigens in T cell activation during avian coccidiosis were evaluated in two inbred strains of chickens using a specific in vitro T cell proliferation assay. Lymphocytes from chickens infected with different Eimeria spp. showed proliferative response to sporozoites, merozoites or Eimeria soluble antigen (Esa) excreted by cultured parasites. Detectable CMI response was observed at 21 day P.I. in chickens infected with E. tenella and E. maxima . Generally lower T cell response was observed in chickens infected with E. acervulina . Merozoites were highly immunogenic compared to sporozoites. Esa prepared from cultured parasites was as effective as whole parasites in evoking a T cell response. Although strain variation in T cell response to parasites or Esa was observed during a primary infection, substantially enhanced T cell response was observed 3 days after a secondary infection in both strains of chickens. The results of the present investigation suggest that Esa may be a major parasite antigen released to the immune system during early stages of infection and relevant to the development of protective immunity.

Cross-reactive cellular and humoral immunity to carcinogen-induced chicken fibrosarcomas. II. Effect of prior immunization with transplantable tumors on primary tumor incidence.

Primary carcinogen-induced (7,12-dimethyl-benz[a]anthracene; DMBA) tumor-bearing SC chickens (B2/B2) frequently showed antibodies in their sera which reacted with cells from their autochthonous tumors, chicken embryo fibroblasts (CEF), tumor cells from some transplantable tumor lines, and from approximately 10% of other primary tumors. Similar results were obtained by ELISA on glutaraldehyde-fixed cells and by immunofluorescence on viable cells. The serum antibody reactivity could be removed by absorption with CEF but not with non-cross-reacting primary tumor cells or a variety of normal tissues. Although sera from normal chickens never showed significant reactivity, a high percentage of sera from chickens that had been injected with DMBA but failed to develop detectable tumors showed antibody activity to a transplantable DMBA-induced tumor and to CEF. On the basis of previously established cross-reactivity patterns in protective immunity to transplantable carcinogen-induced fibrosarcomas, attempts were made to protect against chemical carcinogenesis by prior immunization with selected DMBA-induced transplantable tumors. Tumor-immune chickens showed a significant decrease in the development of tumors during the first 3 mo after injection of DMBA (p = 0.001) or methylcholanthrene (p = 0.033) when compared to controls. This resistance to tumor induction in immune chickens was correlated to the degree of tumor immunity to the immunizing tumor present 1 mo after carcinogen injection (p = 0.046). There was, however, no detectable difference in the incidence of tumors arising later than 3 mo after carcinogen injection. The reduction in tumor incidence in immune as compared to control chickens at 5 mo was therefore less striking than the reduction seen at 3 mo. Immunization with CEF and adjuvants or with adjuvants alone afforded no protection to tumor induction.

Delayed maturation of the antibody response to type 2 thymus-independent antigens in a partially inbred strain of chicken.

The ontogeny of antibody responses to trinitrophenylated (TNP) thymus-independent (TI) antigens was compared in two partially inbred strains of chicken: the SC strain (B2/B2 genotype) and the FP strain (B15/B22 genotype). In the SC chicken, maturation of both the splenic anti-TNP plaque-forming cell (PFC) response and the 19S hemagglutinating antibody response to TI type 2 (TI-2) antigens, TNP-Ficoll and TNP-dextran, were delayed to a significantly later time in ontogeny (20 wk of age) than in the FP chickens (9 wk of age). Four- to 6-wk-old SC chickens were virtually immunologically unresponsive to stimulation with TI-2 antigens. The TI-1 antigen TNP-Brucella abortus was equally immunogenic in both FP and SC chickens of different age groups tested. Kinetic studies of the primary PFC response to TNP-Ficoll in immunologically mature chickens of the SC and FP strains demonstrated a peak PFC response 4 days after antigen injection, followed by a rapid decline in numbers of splenic PFC/spleen on day 6. The results of these studies are discussed in relation to earlier observations that suggested there may be a delay or a defect in the ontogeny of the thymus in the SC chicken.

Avian leukosis virus infection: analysis of viremia and DNA integration in susceptible and resistant chicken lines.

Avian leukosis viruses induce lymphoid leukosis, a lymphoma which develops within the bursa of Fabricius several months after virus infection. Chickens from the Hyline SC and FP lines are, respectively, susceptible and resistant to avian leukosis virus-induced lymphoid leukosis. We examined plasma and cellular DNA obtained from avian leukosis virus-infected chickens for the presence of viremia and integrated viral sequences to determine whether the extent of virus infection is comparable in individuals of both lines. A less than twofold difference in the frequency of viremia was detected between chickens of the two different lines. Although the analysis of plasma samples, which were obtained at different times postinfection, demonstrated that the duration of viremia was comparable in both susceptible and resistant chickens, the onset of the viremia observed in susceptible chickens generally preceded by 1 week that observed in resistant chickens. Moreover, integrated viral sequences were detected in approximately 90% of the SC and 40% of the FP chickens. The appearance of infectious virus in the plasma was, in general, associated with the presence of integrated viral sequences in both the bursal cells and the erythrocytes obtained from the same chicken. The presence of both the viremia and the integrated viral DNA sequences was transient, suggesting a mechanism for the elimination of virus-infected cells in both susceptible and resistant chickens. Furthermore, at 5 weeks postinfection no integrated exogenous viral sequences were detected in splenic lymphocytes obtained from either chicken line, regardless of whether these chickens were viremic or had integrated viral sequences detectable in other tissues. Our results indicate that extensive avian leukosis virus replication occurs in approximately 50% of the FP and 100% of the SC chickens. Although it appears that the viral infection spreads more quickly in the SC chickens, our results afford no obvious explanation of the resistance to the development of lymphoma exhibited by FP chickens.

Differential response to avian leukosis virus infection exhibited by two chicken lines

Infection of susceptible chickens with avian leukosis virus (ALV) results in the development of bursal lymphomas. These neoplasms develop within the bursa of Fabricius following a latent period of several months. The response exhibited by two previously uncharacterized chicken lines to ALV infection has been examined. The two lines, Hyline SC and FP, responded differently to ALV infection. During a 24-week period following intravenous ALV infection, 27 of 50 SC chickens developed bursal lymphomas. No lymphomas developed in the 36 FP chickens tested. A majority of the SC chickens that developed lymphomas also exhibited widespread metastasis to the liver, spleen, and kidneys. Analysis of cellular DNA from the primary and metastasic tumors demonstrated the clonal nature of these neoplasms and revealed altered c- myc loci, as reported in other studies, suggesting the importance of this locus in the development of these tumors. Further characterization of the ALV infection of SC and FP chickens will provide an opportunity to analyze the mechanism of resistance and to contribute to the understanding of the tumorigenic process.

Immunity to carcinogen-induced transplantable fibrosarcoma in [formula omitted] chickens : V. Relationship to tumor cell-specific delayed hypersensitivity and serum antibody

Lasting immunity to the chemically induced (DMBA) fibrosarcomas (CHCT-NYU1, 2, and 4) of SC chickens ( B2 B2 ) can be obtained by injection of Corynebacterium parvum (CP)-primed chickens with tumor cells and CP in one wing and tumor cells alone in the other wing. The local delayed hypersensitivity (DH) reaction to CP in the wing inhibits local growth completely, whereas the tumor on the contralateral side shows transient growth. In the present studies, the development of tumor immunity was studied in detail by monitoring DH and antibody formation to the tumor cells and adoptive immunity with spleen cells in Winn tests. Injection of NYU1 cells alone in normal or CP-treated animals induced transient immunity in Winn tests in 50% of the animals, weak DH reactivity, and antibody detectable by immunofluorescence within the first 2 weeks. Chickens receiving both NYU1 cells and CP in one wing and NYU1 cells alone on the other side developed stronger DH reactions to the tumor cells and a higher incidence of immunity in Winn tests which was sustained throughout the period of observation. Antibody levels were similar to those of animals receiving tumor cells alone. In contrast, injection of CP and tumor cells on one side without a tumor challenge on the contralateral side did not induce detectable immunity in CP-primed chickens. Chickens immunized to NYU2 and 4 cells were also tested for DH reactivity and antibody formation. Studies on the cross-reactivity between the tumor lines showed that there was cross-reactivity at the humoral level while at the cellular level this was not apparent. However, animals immune to one tumor line rejected transplants of another tumor line. Observations on the antibody specificity(s) suggested that it was not directed against minor histocompatibility or avian sarcoma viral antigens. SC embryo fibroblasts could induce DH, and serum antibody induced by tumor cells usually reacted also with such embryo cells.

Derivation of Transplantable 7,12-Dimethylbenz[<italic>a</italic>]anthracene-Induced Chicken Fibrosarcoma Lines: Differences in Metastasizing Properties and Organ Specificity<xref ref-type="fn" rid="fn2">2</xref><xref ref-type="fn" rid="fn3">3</xref>

Several transplantable 7,12-dimethylbenz[a]anthracene-induced SC chicken fibrosarcoma (CHCT-NYU) lines were studied for their ability to grow in internal organs after iv injection (artificial metastases) into 1- to 3-week-old chickens. Some tumor lines were recently derived, whereas others were studied after many serial subcutaneous transplantations. STriking similarities as well as differences were found between tumor lines' ability to grow in various organs. Artificial metastases were seen primarily in the stomach, pancreas, lungs, heart, and muscle, and occasionally also in the kidneys and in the liver. Agammaglobulinemic recipients showed more extensive organ involvement than normal recipients of the same age. Whole-body gamma-irradation also enhanced the incidence of artificial metastases, particularly in lungs. Antibody from the serum of a primary tumor-bearing host reduced the growth of the corresponding tumor in many organs. The metastatic pattern of line CHCT-NYU4 was a relatively stable property, since there was little difference in the internal localization of these tumor cells studied before and after 15 serial subcutaneous transplants. However, intravenous transplantation of tumor cells from line CHCT-NYU4 taken from the liver, lungs, and pancreas of a single recipient established sublines with changes in organ specificity. After a few such serial transplants of liver-derived tumor, a line was derived that grew virtually in the liver alone. A subline with preference for growth in lungs was also obtained, but its ability to grow in the pancreas persisted. A pancreas-derived tumor line also grew in the liver and lungs. Subcutaneous transplants of tissue fragments of the lung-derived tumor line caused the appearance of spontaneous metastases in lungs. The incidence of spontaneous metastases with the lung-derived line was much greater than that with the liver-derived line or with the original CHCT-NYU4 line.

Transfer of Delayed Hypersensitivity in the Chicken

Delayed hypersensitisity (DH) can be transferred in FP and sc chickens with 1 × 108 spleen or peripheral blood cells from donors exhibiting moderate to strong DB reactions. Increasing the spleen cells dose from 1 to 4 × 108 per recipient had no marked enhancing effect on the results. Spleen cells from agammaglobulinemic donors were also shown to transfer DH reactions.

Chemical Carcinogen-Induced Transplantable Fibrosarcomas in Histocompatible Chickens. II. Effect of Age and Thymectomy on Tumor Incidence<xref ref-type="fn" rid="FN2">2</xref><xref ref-type="fn" rid="FN3">3</xref>

The incidence of primary fibrosarcoma 2-4 months after im injection of 7,12-dimethylbenz[a]anthracene (DMBA) was studied in normal, neonatally thymectomized and bursectomized SC (B2/B2) chickens. The tumor incidence was not significantly increased in thymectomized chickens inoculated at 1 week of age, but thymectomized animals inoculated at 4 weeks of age developed a higher incidence of tumors than did controls. Bursectomy did not affect tumor induction. Whereas thymectomy and bursectomy clearly reduced T- and B-cell immune responses, respectively, neither the carcinogen nor the presence of tumor had a detectable effect on the immune response. The effect of varying the age of chickens at the time of carcinogen injection was also studied. DMBA injected into chickens at 8-12 weeks of age produced a significantly lower incidence of tumors than did a similar dose of DMBA injected into chickens at 1-4 weeks of age. Thus DMBA-induced tumors in the chicken may present an interesting model for studies on immune surveillance.

Recruitment of null cells during graft versus host reactions in the chicken

Untreated SC (B2/B2) chicken spleen or thymus cells (2 × 10 7) caused significantly increased [ 3H]thymidine incorporation in spleens of heavily irradiated FP (B15/B21) recipient chicks on Day 4 after iv injection. Mitomycin-treated SC spleen cells or spleen cells treated with rabbit anti-T-cell serum and complement failed to raise the [ 3H]thymidine incorporation over that in uninjected, bursa cell-injected or FP spleen cell-injected controls. However, the combination of mitomycin-treated spleen or thymus cells and anti-T-treated spleen cells caused an increased [ 3H]thymidine uptake, suggesting the recruitment of non-T cells into proliferation by alloreactive mitomycin-treated T cells. Bursa cells did not proliferate during GVH reactions even though they could be shown to undergo proliferation in vivo upon mitogen (lipopolysaccharide and dextran sulfate) stimulation. In contrast, anti-T-treated spleen cells from agammaglobulinemic chickens were recruited into proliferation, suggesting that the recruited cell was not only not a T cell, but also no pre-B or B cell and most likely represented a cell of the monocyte-macrophage series.

Cytotoxicity to allogeneic cells in the chicken I. Role of macrophages in the cytotoxic effect on 51Cr-labeled red blood cells by immune spleen cells

Cytotoxic effector cells in SC and FP chicken spleens were induced by repeated intravenous immunizations with 2 × 10 7 allogeneic red blood cells (RBC). Spleen cells taken 3 days after the last of 3–8 injections caused significant 51Cr release over background from labeled RBC in 5–24 hours at 37° C. Maximal effect was obtained after 5 injections. The effector cells in intact chickens fractionated as adherent to nylon wool and to plastic surfaces and were resistant to rabbit anti-chicken thymus and C. Trypan blue in the incubation mixture greatly reduced the cytotoxic effect without raising the background significantly. The effector cell population in Aγ chickens consisted of a prominent adherent cell and a detectable non adherent T-cell component. The cytotoxic activity of nylon wool adherent cells was somewhat higher on 51Cr labeled syngeneic RBC than that of unfractionated immune spleen cells. However, adherent cells clearly had much higher activity against allogeneic RBC. It appeared therefore, that there was a macrophage-mediated specific cytotoxic effect in spleen cells from allo-immune chickens.

Immunity to transplantable carcinogen induced fibrosarcomas in B2/B2 chickens: II. Macrophage dependency of adoptive T cell mediated tumor immunity

The nature of immunity to transplantable chemically induced fibrosarcomas in SC (B2/B2) chicken was examined using Winn tests performed in the wing web. Immunity in spleen cell donors was induced by pretreatment with C. parvum or BCG followed by tumor cells + bacterial adjuvant in one and tumor cells alone in the other wing web. The T cells mediating the adoptive immunity were sensitive to anti-T + C, nylon wool nonadherent, mitomycin resistant and radiation (1000 R) sensitive. The adoptive immunity could not be expressed in heavily irradiated recipients or in hosts pretreated with trypan blue or silica. The host contribution could be reconstituted by iv injection of spleen or bone marrow cells from agamma-globulinemic (Aγ) unimmunized donors 2 days prior to Winn tests or by the local injection into wing webs of spleen cells or purified peripheral blood monocytes from Aγ donors. It was concluded that cooperation between immune donor T cells and normal monocytes of host origin mediated the inhibition of SCFS growth in Winn tests.

Immunity to transplantable carcinogen-induced fibrosarcomas in B2/B2 chickens. III. Tumor growth inhibition by local delayed hypersensitivity reactions to unrelated antigens.

Delayed hypersensitivity (DH) reaction to human Ig, corynebacterium parvum or allogeneic cells at the site of tumor cell injection, suppressed the fibrosarcoma (SCFS) growth in chickens. Spleen cells of SC chickens sensitized to human Ig or C. parvum suppressed SCFS growth when adoptively transferred with tumor cells, but only when the sensitizing antigen was present locally. This suppression did not occur in irradiated recipients, SCFS I cells injected into wattles of chickens immune to the tumor, provoked a local DH reaction. Spleen cells from donors sensitized to SCFS I adoptively transferred immunity to both SCFS I and SCFS II when cells of the two tumors were mixed together before injection, but not when SCFS II cells were injected alone. Tumor-specific and nontumor-specific DH may be essential for local suppression of fibrosarcoma growth in chickens.

Requirements for induction of delayed hypersensitivity in the chicken

Delayed hypersensitivity (DH) reactions to bovine serum albumin (BSA) or human gamma globulin (HGG) emulsions in complete Freund's adjuvant (CFA) were elicited in normal and bursectomized (BX) SC and FP chickens of varying ages. Responses in normals could be obtained with doses of BSA as low as 2 μg. Reactivity in agammaglobulinemic BX birds was generally lower than in normal animals. Attempts to increase DH with Bacillus Calmette Guérin pretreatment failed in both normal and BX birds. Cytoxan pretreatment did increase DH in normal but not in BX chickens. Similarly, replacement of whole Mycobacteria with the E. coli water soluble cell wall extracts, “Mur-Nac” and peptidoglycans, was effective in normal chickens but failed in BX birds. Modification of BSA with dodecanoyl side groups allowed the induction of DH by i.v. immunization in normal chickens unlike unsubstituted BSA. When immunization was performed with D-BSA or BSA in CFA, no significant difference was evident. Because of the generally poor health of BX birds, attempts were made to improve their responsiveness by pretreatment with normal chicken Ig or normal chicken spleen cell injections. Such pretreated BX chickens indeed gave higher DH to HGG. Thus, the inability of BX animals to respond to the same modifying influences in DH as did normal birds may have been due to an overall lack of responsiveness rather than to a role of B cell or antibody in these effects.

Chemical carcinogen-induced transplantable fibrosarcomas in histocompatible chickens. I. Incidence of tumor induction in normal and bursectomized chickens.

Transplantable fibrosarcomas were developed in two B-locus-defined chicken strains from primary tumors induced by im injection of 2 mg 7,12-dimethylbenz[a]anthracene in 0.1 ml dimethyl sulfoxide into 1- to 2-week-old chicks. Viruses were not important factors in transmission of these tumors as evidenced by 1) transplantability only within the chicken strain of origin, 2) lack of evidence for a filterable agent, 3) maintenance of donor karyotypic characteristics upon transplantation, 4) lack of DNA polymerase and avian leukosis virus group-specific protein production in vitro. Bursectomized inbred SC chickens had a higher incidence of tumor induction than did normals of the same strain. Although the exact interpretation of this finding posed some problems, as discussed, an important function of enhancing antibody in tumor growth appeared excluded.