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. I. Use of bacterial adjuvants in induction of immunity demonstrable in Winn tests.

Tumor line specific immunity could be induced in chickens with the use of BCG or CP. Direct contact between bacteria and tumor cells was required for immunity induction, but succeeded only in animals which also had been previously injected with the same bacterial vaccine. While other methods such as iv injected live mitomycin treated tumor was capable of inducing immunity, only BCG or CP mediated immunity was capable of being adoptively transferred in the Winn assay. Bursectomized agamma-globulinemic chickens also produced strong transferable immunity. Crossreactivity between transplant lines was detectable upon rechallenge of immune donors but was not evident upon adoptive immunity transfer. Immunity was detectable in the spleen and in peripheral blood lymphocytes but could not be transferred by thymus cells.     

Cross-reactive cellular and humoral immunity to carcinogen-induced chicken fibrosarcomas. I. Protective cross-immunity between transplantable tumor lines

Dimethylbenz[a]anthracene (DMBA)-induced transplantable fibrosarcomas in B2 homozygous chickens (SC line) grow progressively in normal chickens, but are rejected by chickens immunized previously with irradiated tumor cells and Corynebacterium parvum. Tumor-immune chickens resist challenge by the immunizing tumor lines as well as by some, but not all, fibrosarcoma lines. The pattern of cross-reactivity between four DMBA-induced transplantable tumor lines was examined in detail. Ability to reject a tumor challenge correlated very well (p less than 0.001) with the presence of delayed-type hypersensitivity (DTH) to that tumor. Immunization with one of two of the DMBA-induced lines tested also caused rejection of transplantable tumors developed from methylcholanthrene-induced and benzo(a)pyrene-induced primary fibrosarcomas. Although immunization with tumor caused DTH to chicken embryo fibroblasts (CEF), immunization with CEF failed to cause protective immunity or DTH to tumors. Presence of protective immunity, where tested, also correlated with the ability of spleen cells from immune donors to inhibit tumor growth in Winn tests. Humoral immunity exhibited even greater cross-reactivity than did cellular immunity. Distinct patterns of cross-reactivity were nevertheless observed with respect to the serum antibodies as detected in ELISA. Two of these patterns were also observed in several sera from primary tumor-bearing chickens, both including reactivity with CEF. Such reactivity was absent from normal chicken sera.     

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.     

Assessment of syngeneic cell-mediated immunity to mouse mammary tumors by three different assays

Summary The purpose of this study was to compare the MCT, CRT, and Winn assays for assessment of CMI to syngeneic tumors. Spontaneous and MMTV-induced mammary tumors in Balb/c and Balb/cfC₃H mice were used. LNC were obtained at various times relative to S.C. injection of sensitizing tumor cells and surgical removal of the outgrowth. In direct comparison to Winn assays, CRT often gave false negatives (i.e., LNC could inhibit tumor growth in the Winn assay without being cytolytic), but when LNC were cytolytic in the CRT, they inhibited in Winn assays. Thus, a positive CRT was associated with a positive Winn assay. In contrast, results from MCT tests were generally not associated with those obtained by CRT or Winn assays. This divergence in assays was independent of differences in status of the LNC donor. With all three assays, stimulation of tumor growth or increased survival was occasionally observed. For the CRT this was manifested in the release of smaller amounts of ⁵¹Cr in the presence of immune LNC than in the presence of normal LNC. For MCT and Winn assays it was seen in better tumor growth after treatment with LNC than after treatment with medium alone. Detection of this phenomenon was independent among the three assays.Data from the CRT and Winn assays were analyzed to see whether they correlated with in vivo behavior of the sensitizing tumors. Detection of CMI by these tests was associated with tumors growing out after relatively long latency periods, but was independent of tumor growth rate. This is in contrast to our previously reported analysis of MCT data, in which detection of CMI was associated with slower growth rate but was independent of the latency period (Hager et al., 1978).Abbreviations CMI cell-mediated immunity - CRT chromium-release test - MCT microcytotoxicity test - MMTV murine mammary tumor virus - LDA lymphocyte-dependent antibody - LNC lymph node cells - SBF serum-blocking factors - SC subcutaneous - TCM tissue culture medium     

Inhibition of pulmonary metastases of B16 melanoma with irradiated tumor cells and BCG

Summary When the tumor-bearing leg of C57BL/6J mice was amputated 16 days after SC inoculation of 10⁶B16 melanoma cells, all the amputated mice died of pulmonary metastases. Transfer of lungs from the amputated to normal syngeneic mice revealed tumor cells in the lungs just after amputation. Repeated weekly injections of BCG and irradiated tumor cells, beginning 24 h after amputation of the tumor-bearing limb, prolonged the survival only of mice presensitized to BCG. Injections of BCG or irradiated melanoma cells alone, of neuraminidase- and mitomycin C-treated tumor cells or of Levamisole had no effect, but injections of ConA-coated tumor cells slightly prolonged the survival of the amputated mice. Both BCG and B16 cells induced humoral and cell-mediated immunity but there was no cross-reactivity between BCG and B16 cells. Abbreviations used: ConA, concanavalin A; SC, subcutaneous; IP, intraperitoneal; IV, intravenous; ID, intradermal; IT, intratumoral; PBS, phosphate-buffered saline (0.01 M sodium phosphate, pH 7.1); VCN, Vibrio cholerae neuraminidase; HBSS, Hank's balanced salt solution; RPMIM, Roswell Park Memorial Institute medium     

Passive transfer of tumor immunity with spleen cells from guinea pigs cured of hepatocarcinoma by nonspecific immunotherapy

Summary The purpose of this study was to evaluate cell-mediated tumor immunity in strain-2 guinea pigs cured of line-10 hepatocarcinoma by oil-in-water emulsions containing phenol-water extracts from either BCG or the Re mutant of Salmonella typhimurium (Re ET) admixed with mycobacteria glycolipid (P3). Treatment with these emulsions produced complete regression of established tumor nodules and prevented the growth of lymph node metastases in 25 of the 28 animals inoculated intradermally (ID) with 10⁶ line-10 cells and given intralesional immunotherapy 6 days later. No tumor regression was observed in animals given phenol-water extracts alone. Spleen cells, taken from guinea pigs cured of line-10 by BCG extract + P3 or Re ET + P3, were tested for their influence on tumor growth by means of an in vivo adoptive neutralization test (Winn test). Cell transfer was accomplished by the subcutanous injection of various concentrations of spleen cells admixed with 10⁵ viable line-10 cells. The results showed that as few as 10⁷ immune spleen cells completely inhibited the growth of 10⁵ tumor cells in 46–54% of the animals. The best tumor growth inhibition (77–85%) was observed in animals given 5 × 10⁷ immune cells admixed with 10⁵ tumor cells. The onset of transferrable tumor immunity was earlier in animals treated with the BCG extract + P3 than in those given the Re ET + P3. However, the duration of detectable tumor immunity was longer in the latter group. In contrast, no inhibition of tumor growth was observed in animals given spleen cells from normal or tumor-bearing guinea pigs. Moreover, spleen cells obtained from guinea pigs immunized with BCG extract + P3 or Re ET + P3 emulsions only and admixed with line-10 cells failed to transfer tumor immunity to normal animals. Thus, results from this study clearly demonstrated that cell-mediated tumor immunity was elicited in animals cured of line-10 tumor with combinations of P3 and phenol-water extracts of either BCG or Re mutant of S. typhimurium and that sensitized spleen cells effectively transferred systemic tumor immunity to normal recipients.     

Suppressor T cells with histamine type II receptors in chickens bearing chemically induced fibrosarcomas

Immunity to a local (wing web) challenge with transplantable, chemically induced tumors, such as CHCT-NYU-4, can be transferred to histocompatible SC chickens with intravenously injected splenic T cells from tumor-immune chickens. Simultaneous iv injection of splenic T cells from chickens bearing progressively growing CHCT-NYU-4 prevents the expression of this adoptive systemic tumor immunity. The splenic suppressor T cells are B-L(Ia) positive and adhere to dishes coated with cimetidine-protein conjugate, suggesting that they bear type II histamine receptors (H2R). Intravenous injection of gamma-irradiated CHCT-NYU-4 cells 7 days prior to a local challenge with the same tumor promotes growth of a normally suboptimal tumor cell dose such that the incidence of progressive tumor growth is significantly increased. Concomitant treatment with the H2R antagonist, ranitidine, inhibits tumor growth of an optimal tumor dose challenge and promotes induction of tumor immunity. However, this drug cannot reverse the effect of iv injected tumor cells. Another drug with reported antisuppressor cell activity, cyclophosphamide (50 mg/kg), injected 1 day prior to challenge causes a transient inhibition of tumor growth. Injection of this drug on Day 7 does not have a significant effect by itself but, in combination with the Day-1 pretreatment, a significant inhibition of tumor growth by size and tumor incidence measurements is obtained. These results indicate that manipulation of immunity to transplantable fibrosarcomas in the chicken is possible with drugs acting on suppressor cells.     

The presence of immunoregulatory cells in chicken thymus: function in B and T cell responses.

The presence of immunoregulatory cells in chicken thymus was studied by using several different systems. Chickens injected with large numbers of syngeneic thymocytes were tested for their ability to produce antibody to heterologous red cells. Similar chickens were studied for their ability to reject allogeneic skin grafts. In separate studies, mixtures of thymocytes with spleen cells or with peripheral blood leukocytes were assayed for their ability to respond to PHA or to produce a graft-vs-host reaction in embryonic chicks. These studies indicated that immunoregulatory cells exist in chicken thymus, which displays both helper and suppressor activity. The suppressor cells were more prevalent or more easily detectable in young birds and in chickens with intact bursas. The helper function of thymocytes was seen to better advantage with cells derived from older animals and from bursectomized donors.     

T cell tolerance in the chicken. I. Parameters affecting tolerance induction to human gamma-globulin in agammaglobulinemic and normal chickens.

T cell-mediated delayed hypersensitivity (DH) to human gamma-globulin (HGG) can be induced in chickens by subcutaneous injection of the antigen in complete Freund's adjuvant (CFA). In the present work, it has been demonstrated that specific tolerance of the cells mediating this DH can readily be induced in both normal and bursectomized (BX) FP strain chickens by simple i.v. injection of soluble antigen, regardless of the presence of antibody production to the tolerogen. A significant degree of tolerance at the DH and helper T cell levels could be generated in BX birds by injection of as little as 0.5 mg of HGG; such a dose only induced tolerance in normal birds when it had been previously deagregated by ultracentrifugation. Regular, nondeaggregated antigen could produce tolerance in normal animals, but only at doses of greater than 5 mg. The tolerizing injection induced a primary antibody response in normal birds in all cases, but a secondary response could not be obtained in animals rendered tolerant at the T cell level. Establishment of tolerance appeared to be very rapid, and animals remained refractory to induction of DH for at least 3 weeks after the tolerizing injection. The mode in which the antigen was presented to the animals appeared to be crucial in determining whether tolerance or sensitivity would be established.     

Influence of different routes of anti-tumor immunization: alternative induction of tumor immunity and tumor enhancement.

Chickens and quails were immunized in parallel either i.v. or intramuscularly (i.m.) with lectin column-purified antigens from chick embryo cells that were transformed in vitro by avain sarcoma virus (ASV). After five to six injections, immunity of the animals was tested by challenge with ASV into the wing webs. Whereas tumor growth was inhibited after i.v. immunization with respect to incidence rate and time of tumor appearance, tumor growth was enhanced after i.m. injection. Animals that were injected with normal cell antigens served as controls. Spleen cells from only those animals that were immunized i.v. exerted a cytotoxic effect in vitro against ASV-transformed cells, whereas spleen cells from i.m. injected animals, in contrast, suppressed such cytotoxicity. The search for serum blocking or arming factors suggested that sera from i.m. injected animals block cellular cytotoxicity whereas sera from i.v. immunized animals render normal spleen cells cytotoxic (arming effect). The use of viruses from different subgroups and of antigens from gp85-lacking ASV-transformed cells indicates that immune effects were obtained against tumor cell surface antigens that differ from the antigen that is involved in virus neutralization (s-gp85).     

Augmentation of immune response to syngeneic fibrosarcoma T241 with in vivo protection

Summary Lewis T241 fibrosarcoma, a syngeneic tumor in C57 BL/6J mice, was found to be poorly immunogenic. When tumor-bearing animals (TBA) were challenged with tumor cells either concomitantly or after excision of a growing tumor no protection was observed. In vivo (Winn) neutralization assays also showed a lack of tumor immunogenicity. However, in vitro studies showed that a significant proliferative response could be elicited from the spleen cells of TBA when these cells were cultured with either mitomycin-C-treated tumor cells or KCl tumor extract. Similarly, macrophage migration inhibition factor (MIF) was produced by TBA spleen cells upon incubation with KCl tumor extract, but no cell-mediated cytotoxicity to T241 target cells was observed with various lymphoid cell populations at any stage of tumor growth. Immunization of syngeneic animals with Vibrio cholerae neuraminidase(VCN)-treated, irradiated tumor cells alone or admixed with Freund's complete adjuvant (FCA) resulted in decreased tumor growth and fewer pulmonary metastases following challenge with 10⁶ tumor cells. No complete tumor rejection was observed. In contrast, 13 of 16 animals immunized with irradiated tumor cells admixed with FCA rejected 10⁵ tumor cells. Animals that grew tumors had significantly reduced tumor growths and pulmonary metastases. Lymph node and peritoneal exudate cells (PEC) of immunized animals showed significant cytotoxicity to T241 cells.     

DNA vaccination against the idiotype of a murine B cell lymphoma: mechanism of tumor protection

Several studies have shown that immunization with DNA, which encodes the idiotypic determinants of a B cell lymphoma, generates tumor-specific immunity. Although induction of antiidiotypic Abs has correlated with tumor protection, the effector mechanisms that contribute to tumor protection have not been clearly identified. This study evaluated the tumor protective effects of humoral and cellular immune mechanisms recruited by idiotype-directed DNA vaccines in the 38C13 murine B cell lymphoma model. Antiidiotypic Abs induced by DNA vaccination supported in vitro complement-mediated cytotoxicity of tumor cells, and simultaneous transfer of tumor cells and hyperimmune sera protected naive animals against tumor growth. However, in vitro stimulation of immune splenocytes with tumor cells failed to induce idiotype-specific cytotoxicity, and following vaccination, depletion of CD4 or CD8 T cell subsets did not compromise protection. Furthermore, protection of naive recipients against tumor challenge could not be demonstrated either by a Winn assay approach or by adoptive transfer of spleen and lymph node cells. Thus, in this experimental model, current evidence suggests that the tumor-protective effects of DNA vaccination can be largely attributed to idiotype-specific humoral immunity.     

Increased growth of RSV-induced tumors in chickens partially tolerant to MHC alloantigens

Chickens with B2B2 MHC genotypes were made partically tolerant to B5 MHC cell-surface antigens and the fate of their Rous-sarcoma-virus (RSV)-induced tumors was determined. B2B2 chickens partially tolerant to viable or lysed white blood cells (WBC) or viable red blood cells (RBC) from B5B5 chickens had a significantly higher incidence of tumor progression than untreated, PBS-treated, or B2B2 chickens inoculated with WBC from other B2B2 chickens. The criteria for tolerance were absence of antibody titer to the cell type inoculated and acceptance of allografts from B5B5 donors by B2B2 chickens. Graft-vs-host reactions occurred only in B2B2 chickens inoculated with viable WBC from B5B5 chickens. It appears that B2B2 chickens partially tolerant to B5 antigens failed to mount a successful immune response to RSV-induced tumors partly because of a B5 MHC antigen(s) cross-reacted with a tumor associated antigen(s) thereby severely limiting B2B2 host recognition of the tumor as foreign. Since WBC and RBC cell-surface antigens appear to contribute similarly to the effect, the B-F- region of the MHC may be involved.     

Regulation of the immune response to tumor antigens. IX. In vitro Lyt-1+2- cell proliferative responses to cellbound or subcellular tumor antigen

We investigated the cellular basis of immune reactivity to the S1509a fibrosarcoma in tumor-immune A/J mice. In a Winn assay, immune Lyt-1+2- T cells are capable of retarding S1509a tumor growth in naive A/J mice. In vitro proliferation to S1509a is also mediated by tumor-immune Lyt-1+2- T cells. This response is specific to the immunizing tumor and appears 5 to 7 days after reexposure to the tumor in vivo. Proliferation also requires the presence of a population of adherent cells. In fact, adherent peritoneal exudate cells pulsed with tumor membrane fragments derived from S1509a cells can stimulate proliferation. Proliferation is blocked by the addition of anti-I-Ak monoclonal antibody to the culture medium without complement or by treatment of the responder population with anti-I-Ak and complement. In vitro responsiveness is also inhibited by the presence of tumor-specific suppressor T cells in vivo. These observations suggest in vitro proliferation may provide a potential means of defining tumor antigens and cell-surface structures involved in tumor immunity.     

Eimeria acervulina: evaluation of the cellular and antibody responses to the recombinant coccidial antigens in B-congenic chickens

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.     

Differential ability of tumor-unique and cross-reactive antigen(s) on two murine hepatoma cell lines to induce Lyt-1+2- T cells responsible for in vivo protective immunity

The role of the tumor-unique determinant(s) on two syngeneic murine hepatoma cells in inducing in vivo protective immunity was investigated in comparison with that of the tumor-cross-reactive determinant(s). Induction of vaccinia-reactive helper T cells in C3H/He mice by intraperitoneal (i.p.) inoculation of viable vaccinia virus and then immunization with vaccinia-infected syngeneic MH134 or MH129 tumor cells resulted in the production of potent anti-MH134 or -MH129 antibody as well as the generation of in vivo protective immunity. Neither antibody reacted with other syngeneic plasmacytoma or fibrosarcoma cells, but both cross-reacted appreciably with the other hepatoma cells as well reacted strongly as with the tumor cells used for immunization. The absorptions of anti-MH134 and -MH129 antisera with the respective hepatoma cells abolished their reactivities with both the corresponding hepatoma cells and the other hepatoma cells. In contrast, the absorption of these antisera with the other tumor cells resulted in loss of their cross-reactivities with the other hepatoma cells, but not loss of their specific reactivity to the respective hepatoma cells. Although in these hematoma systems, the above-mentioned immunization protocol resulted in in vivo induction of protective immunity and generation of antibodies, in vivo immunity as observed by Winn assays was mediated by Lyt-1+2- T cells and was specific for each type of hepatoma cells. These results indicate that these two types of hepatoma cells bear two kinds of antigenic determinants, one kind unique to each hepatoma and the other kind cross-reactive with the other hepatoma cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Studies of delayed hypersensitivity to L. Monocytogenes in mice: nature of cells involved in passive transfers

C3Hf/Umc mice were immunized by an intravenous injection of a sublethal dose of live Listeria monocytogenes. The animals developed delayed-type hypersensitivity (DH) concomitant with infectious immunity to this organism. Delayed hypersensitivity could be transferred to normal lethally irradiated mice with spleen cells from immune animals. The immune cells cells responsible for transfer of adoptive immunity were susceptible to in vitro cytolytic action of anti-theta iso-antibody and complement, since such treatment rendered these cells incapable of further passive transfer of specific immunity to Listeria. The acquired DH to Listeria persisted in mice after 900 R lethal irradiation, provided normal syngeneic bone marrow cells were also administered, thus indicating the persistance of a cell population in the immune irradiated mice, resistant to effects of radiation. The radio resistant nature of this immune cell population was further demonstrated by passive transfer with spleen cells, derived from preimmunized lethally irradiated mice to normal syngeneic mice or to lethally irradiated nonimmunized hosts reconstituted with normal bone marrow which then responded to antigenic challenge with DH.Treatment of the immune radio resistant spleen cells in vitro with anti-theta and complement eliminated passive transfers of DH by these cells; however, this effect was less obvious than similar treatment of the immune, nonirradiated, spleen cells.     

The effect of bursectomy and thymectomy on the course of avian influenza virus infection

The effect of hormonal bursectomy and neonatal surgical thymectomy on the course of an avian influenza virus infection in chickens was studied. Analysis of the immunologic status of the chickens prior to infection included assay of natural agglutinins to rabbit red blood cells and induced agglutinins to sheep red blood cells, serum immunoelectrophoresis patterns, and in vitro effects of phytohemagglutinin on lymphocyte transformation. At 6 wk of age the chickens received the influenza virus intratracheally. Daily temperatures and mortality were recorded and HAI antibody titers were measured 7 and 14 days later. Completely thymectomized chickens were characterized by a failure of lymphocyte transformation to take place in two successive studies and absence of thymic remnants at autopsy. Bursectomy was associated with a significantly higher occurrence of temperature elevation (P < 0.05) and mortality (P < 0.001). Thymectomy had no significant effect on the course of the virus infection. Preliminary findings with passive administration of serum from immune animals also suggested a role for antibody in host resistance. These studies suggest cell-mediated immunity is less important than humoral immunity in recovery from avian influenza virus infection.