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An enzyme-linked immunosorbent assay was constructed by using as antigens the type-specific immunodominant glycopeptidolipids of selected serotypes of Mycobacterium avium. This assay system was used to determine the prevalence of raised antibody levels to these antigens in groups of controls, human immunodeficiency (HIV)-negative and -positive homosexual men, and HIV-negative patients with active M. avium infections as a possible indicator of potential exposure and/or colonization by M. avium in these individuals. The results indicate that while antibody levels were raised in only 2.4% of control individuals, 33% of HIV-negative homosexual men and 44% of HIV-positive patients exhibited raised levels. Moreover, further examination of the HIV-positive group revealed no correlation between antiglycopeptidolipid antibody activity and helper T cell numbers. These data indicate that exposure to M. avium is prevalent among the homosexual male population, regardless of their HIV status. Moreover, the data are suggestive that the emergence of disseminated M. avium disease in HIV-positive patients may sometimes arise from earlier colonization, rather than as a newly acquired infection during terminal immunodeficiency.

Disseminated Mycobacterium avium infection is the major cause of bacteremia in patients with acquired immunodeficiency syndrome. We present here a new animal model of this disease, thymectomized C57BL/6 mice that were intravenously infused with monoclonal antibody to selectively deplete CD4+ T cells. The increased susceptibility of such animals to M. avium infection is comparable to that of C57BL/6 beige mice and thus may provide a viable alternative to the latter model. Further, using representative strains of acquired immunodeficiency syndrome-associated M. avium (serotypes 1, 4, and 8 and a rough isolate), we show that the course of such infections in thymectomized, CD4-deficient mice can be markedly restrained and in some cases the infections can be sterilized by treatment over a 120-day period with a regimen containing 40 mg of the new antimycobacterial agent rifabutin per kg (body weight).

We examined the effects of lead ingestion on in vitro and in vivo indices of immune function in mallards (Anas platyrhynchos). Twenty-four mallard drakes were randomly divided into three groups (I, II, III), then assigned to treatment or control subgroups (n = 4). On day 0, all treatment birds were dosed orally with one number 4 lead shot (mean = 0.2 g). We challenged all individuals in each group with washed sheep red blood cells (SRBC) injected intraperitoneally on days 0 (Group I), 7 (Group II) or 14 (Group III), and collected blood for analyses 7 and 8 days after SRBC challenge. We measured and compared blood lead concentrations, in vitro lymphocyte transformation responses to phytohemagglutinin A and lipopolysaccharide, and hemagglutination titers to SRBC. Mean blood lead concentrations were elevated (P less than or equal to 0.04) in treatment birds at each sampling period. Large individual variability in lymphocyte stimulation responses precluded further analysis of those data. Hemagglutination titers to SRBC were lower (P less than 0.0001) in lead-poisoned ducks than in controls, suggesting that ingested lead may have immunosuppressive effects on mallards.

The purpose of these experiments was to re-evaluate the usefulness of the thymectomized, irradiated, bone marrow reconstituted mouse (TXB mouse) as a model of the effect of immunodeficiency on resistance to mycobacterial infection. It was found that, although TXB mice had no resistance to intravenous infection with Mycobacterium tuberculosis, such mice were able to express moderate resistance to infection with the BCG vaccine, even when given in high doses. It is concluded, therefore, that rather than representing an animal devoid of immunological reactivity, the TXB mouse retains a residual capacity, albeit minimal, to express acquired immunity.

Subcutaneous BCG vaccination of mice several weeks after intravenous or subcutaneous infection with either M. avium or M. kansasii had no effect on the subsequent course of these nontuberculous mycobacterial infections. In some animals the growth of the BCG infection in the draining popliteal lymph nodes was reduced compared with the growth in controls, although these mice were as resistant as BCG-vaccinated controls to a subsequent airborne challenge infection with M. tuberculosis. Nonvaccinated previously infected mice also showed some degree of resistance to the airborne challenge infection; this resistance was more pronounced in mice infected with nontuberculous mycobacteria intravenously than in those infected subcutaneously.

Mice heavily infected with Mycobacterium bovis BCG rapidly became anergic to cutaneous injection with tuberculin. Evidence is presented suggesting that this anergy reflects an adaptive physiological change within the host in which antigen-reactive Thy-1.2+ cells become sequestered in central lymphoid tissues, with a concomitant reduction in the circulating pool. No evidence could be provided to support the suggestion that anergy was a consequence of an acquired immunosuppressive mechanism.

The results of this study demonstrate that spleen cells taken from mice at the height of the primary immune response to intravenous infection with Mycobacterium tuberculosis possess the capacity to transfer adoptive protection to M. tuberculosis-infected recipients, but only if these recipients are first rendered T cell-deficient, either by thymectomy and gamma irradiation, or by sublethal irradiation. A similar requirement was necessary to demonstrate the adoptive protection of the lungs after exposure to an acute aerosol-delivered M. tuberculosis infection. In both infectious models successful adoptive immunotherapy was shown to be mediated by T lymphocytes, which were acquired in the donor animals in response to the immunizing infection. It is proposed that the results of this study may serve as a basic model for the subsequent analysis of the nature of the T cell-mediated immune response to both systemic and aerogenic infections with M. tuberculosis.

Fifteen patients with infantile bone and joint infections were studied immunologically and clinically, 3 at the time of illness and 12 later. Abnormality of immunoglobulins, or complement, or phagocytes was found in 9 patients; 6 were within normal limits for the tests undertaken. Immunodeficiency is probably responsible for the subdued clinical signs of infection and for delayed diagnosis in some patients. It was also related to the extent of femoral head damage in infective arthritis of the hip and to the incidence of wound infection in late elective surgery.

The results of in vitro experiments designed to measure the immunocompetence of mice heavily infected with the atypical mycobacterial pathogens Mycobacterium avium and Mycobacterium simiae were compared with the results of experiments which used in vivo approaches. Blastogenic responsiveness in vitro both to mitogen and to alloantigen was severely depressed in the heavily infected mice; this responsiveness could be restored by removal of an inhibitory Thy-1.2-, nylon wool-adherent cell population. No evidence was found to support the previous contention that suppressor T cells may play a role in the inhibition of this responsiveness. These results were then compared with experiments which measured the ability of the infected animal to elicit a delayed-type hypersensitivity response to sheep erythrocytes in vivo. However, although delayed-type hypersensitivity responses in vivo were also depressed, evidence was obtained which suggested that this unresponsiveness was due to inadequate sensitization of T cells, possibly due to catabolism of antigen, rather than due to the influence of an active, immunosuppressive mechanism. Finally, despite the severely depressed ability of cells from infected mice to respond to alloantigenic stimulation in vitro, infected animals were fully able to cause the regression of a tumor implant in vivo.

The efficacy of Mycobacterium bovis BCG immunization in mice with established pulmonary infections caused by atypical mycobacteria was studied. In all four strains of Mycobacterium tested (M. kansasii, M. simiae, M. avium, and M. scrofulaceum), intravenous inoculation with 10(6) BCG had no discernible effect upon the course of atypical mycobacterial infection within the lungs; despite this, however, all BCG-vaccinated groups of mice were fully resistant to a subsequent acute aerogenic challenge with M. tuberculosis H37Rv, regardless of the presence of the pulmonary atypical mycobacterial infections. Furthermore, animals infected with M. kansasii, M. simiae, or M. avium but not vaccinated with BCG expressed considerable antituberculous resistance within the lungs, resulting in significant prolonged survival of these animals. The relevance of these findings to the expression of antituberculous resistance in human populations in areas in which atypical mycobacteria are endemic and the failure of these findings to support the hypothesis that prior contact with atypical mycobacteria might in some way jeopardize or interfere with the efficacy of subsequent BCG vaccination are discussed.

Adoptive transfer of protective immunity to an aerogenic infection with the facultative intracellular bacterium Mycobacterium tuberculosis was mediated by a population of T cells acquired in the spleen of donor mice at the height of the primary cell-mediated immune response to an immunizing infection with M. bovis bacillus Calmette-Guerin. Successful adoptive immunotherapy was ablated by prior exposure of immune donor cells to ionizing radiation or by treatment of these cells with antibody raised against the Ly-2 marker. In contrast, however, the capacity of immune donor cells to passively transfer delayed-type hypersensitivity (DTH) responses to tuberculin was unaffected by prior treatment with antibody to Ly-2, but was completely ablated by treatment by antibody to Ly-1. These results indicate, that DTH and protective anti-tuberculous immunity are dissociable phenomena, mediated by separate populations of T lymphocytes.

The relationship between natural resistance to Mycobacterium bovis BCG, expressed by the Bcg gene, and the generation of acquired resistance to this infection in various selected inbred strains of mice was investigated. Consistent with previous findings, a low dose (approximately 10(4)) of BCG Montreal grew progressively in the spleens of inbred mouse strains previously designated susceptible to BCG (Bcgs), but grew poorly in resistant strains (Bcgr). In contrast, however, little difference was observed in the growth of the organism in the liver or lungs of these mice, whereas furthermore, all animals behaved as Bcgs when infected with the World Standard preparation of BCG, BCG Pasteur. Moreover, four strains tested (Bcgr; A/J, C3H/HeJ, and Bcgs; B10.A/J, BALB/c), all showed evidence of the generation of acquired resistance to a small inoculum of BCG Montreal, as demonstrated by their substantial protection against a subsequent intravenous challenge with virulent M. tuberculosis. These findings are interpreted as being inconsistent with the Bcg gene hypothesis and call into doubt the usage of the term Bcg as a gene designation.

Mice were immunized with the cell wall of BCG suspended in an oil-in-saline emulsion, and examined against time for the emergence of T cell-mediated acquired immunity. Evidence is presented that shows that levels of acquired resistance expressed in these animals over the first month following inoculation, and which enabled them to substantially resist an intravenous challenge infection with Mycobacterium tuberculosis, were completely nonspecific in nature, in that they were equally well expressed in normal and T cell-deficient mice, and were present at a time when no protective T cell activity could be passively transferred from the inoculated host. Paradoxically, in contrast, weak but statistically significant protective immunity could be detected in the spleens of CW-immunized mice approximately 3 months after inoculation, at a time when the donor animals were devoid of resistance to rechallenge. Finally, evidence is presented that shows that the CW material, if given subcutaneously, is highly immunogenic for the generation of delayed-type hypersensitivity effector T cells; however, these cells do not themselves contribute to protective immunity.

The effect of prior vaccination of mice with Mycobacterium bovis BCG on the subsequent course of acute aerogenic infection with various environmental mycobacteria was tested. A protective effect was recorded against infection with M. avium, and M. kansasii; by contrast no effect was noted against M. simiae or M. intracellulare. The prophylactic effects of BCG were demonstrated regardless of whether vaccination was given by the intravenous or aerogenic routes. These findings support the hypothesis that BCG vaccination, if given prior to contact, can provide some degree of protection against certain non-tuberculous mycobacterial infections.

Results show that various inbred strains of mice can be segregated into two distinct groups, based on their capacity to allow a number of nontuberculous mycobacterial infections to grow in target organs following experimental intravenous infection. The first group, which allowed these infections to grow progressively, was thus designated as naturally susceptible to these infections; in contrast, those strains which were able to exert detectable bacteriostasis were designated as naturally resistant. It was then found that segregation of mouse strains based on this distinction also mirrored the capacity of these animals to generate acquired immunity to the mycobacterial infections. For example, Mycobacterium simiae grew progressively in susceptible C57BL/6 mice, subsequently triggering acquired mechanisms of immunity, whereas no evidence for acquired immunity could be found in resistant A/Tru mice infected with this organism. The possibility that acquired immunity could not be expressed in the latter strain as a result of a defect in macrophage activation was excluded. Moreover, it was found that the trait of resistance to these infections could be transferred by bone marrow cells into radiation chimeras, thus indicating that this trait was expressed by the progeny of hemopoietic precursor cells. Subsequent backcross analysis to determine the mode of inheritance of the trait of resistance to these mycobacterial infections revealed data that were consistent with the hypothesis that this resistance is controlled by more than one gene. Statistical analysis of the data by the maximum likelihood method suggested polygenic control, although in some cases the probability values suggested control by a major gene, influenced by modifier genes. These findings suggest that the previous hypothesis that the growth of mycobacterial infections in inbred strains of mice is controlled by a single gene should be reevaluated.

This study shows that memory immune T lymphocytes, harvested from young (3-month-old) mice infected intravenously with Mycobacterium tuberculosis and then exposed to protracted chemotherapy with isoniazid, are capable of adoptively protecting old (24 month) mice from a subsequent fatal challenge infection. Survival of such adoptively protected animals was prolonged, but did not extend beyond the mean survival time of uninfected old control mice. During this time the passively transferred memory T cell population retained their functional capacity to protect against subsequent tuberculosis infection. These data indicate, therefore, that reconstitution of decayed cell-mediated antimicrobial immunity in old mice in vivo with memory T cells is technically feasible, although the life span of the animal is not extended over that of control animals. They indicate, moreover, that the memory T cells remain functional in what some reports have considered a suppressive environment and show further that the macrophages with which the infused T cells interact in the aged host remain functionally able to express acquired resistance.

A number of nonliving mycobacterial preparations were tested in vivo for their capacity to generate various relevant parameters of cellular immunity. All preparations tested had some detectable activity in raising resistance to challenge with Mycobacterium tuberculosis or with Listeria monocytogenes and in conferring the ability to mount a delayed-type hypersensitivity response to tuberculin. This report presents the first evidence, however, that none of these preparations were able to generate protective T cells capable of adoptive immunization against virulent tuberculosis. These data are discussed in terms of the use of these preparations in generating M. tuberculosis-reactive T-cell lines and the application of these lines in the continuing search for an improved vaccine against tuberculosis.

This study employed an experimental mouse model of Mycobacterium tuberculosis infection to investigate the effects of aging on T cell-mediated protective cellular immunity. It was found that although mice of 3 to 18 mo of age were fully resistant to a standard immunizing dose of Mycobacterium tuberculosis, progressive mortality was observed in old (24 to 28 mo) mice. Death of these older animals was associated with an inability to contain or to eliminate the mycobacterial infection in the spleen and liver, and with an inability to prevent the progressive growth of the infection in the lungs. It was then revealed by the use of reciprocal passive cell transfer experiments that the age-related susceptibility of old mice reflected an inability to generate mediator protective T lymphocytes in response to the infection. In contrast, no evidence was obtained to indicate any defect at the effector cell (macrophage) level, as evidenced primarily by the finding that immune T cells from young mice conferred equivalent levels of immunity upon both old and young recipients. The results suggest therefore that T cell-mediated immunity undergoes an age-related decline in terms of its ability to respond to infection with Mycobacterium tuberculosis.

A variety of experimental infections with pathogenic mycobacteria are associated with the development of persistent disease, in which little or no changes in the numbers of the infectious organism can be detected. This report describes a simple experimental model designed to test the hypothesis that this persistence may reflect in part the ability of these organisms to resist the enhanced bacteriostatic and bactericidal properties acquired by host macrophages as a result of these mycobacterial infections. To examine this possibility mice were inoculated with test organisms at a time when these animals were expressing very high levels of nonspecific resistance, and hence macrophage activation, as a result of a prior intravenous infection with Mycobacterium bovis bacillus Calmette-Guerin (BCG). The results show that the test organisms fall into three groups; (a) those, such as Mycobacterium tuberculosis, which were sensitive to the presence of activated macrophages, (b) those, such as Mycobacterium avium and Mycobacterium kansasii, which were insensitive, and (c) one organism, Mycobacterium intracellulare, in which progressive growth of the infection was significantly improved. These results are consistent with the hypothesis that some mycobacteria, particularly those associated with persistent disease, possess an intrinsic resistance to host bactericidal and bacteriostatic mechanisms in vivo.

Ten-day-old chickens infected with an avian osteopetrosis virus [MAV-2(O)] were more susceptible to challenge with Listeria monocytogenes than virus-free chickens, as demonstrated by reduced bacterial clearance from their spleens. Reduced clearance of L. monocytogenes was observed throughout a 26-day period after MAV-2(O) infection.

The results herein show that mice infected with Mycobacterium tuberculosis and then exposed to a protracted course of isoniazid chemotherapy possess a heightened state of acquired resistance to subsequent challenge with the homologous organism. Our results provide the first evidence, moreover, that this resistance is mediated by a long-lived, cyclophosphamide- and irradiation-resistant L3T4+ Lyt-2- lymphocyte capable of giving rise to an accelerated re-emergence of resistance in the animal upon rechallenge. Evidence is also provided to show that triggering of this memory-immune T cell population in the re-challenged host was associated with the rapid emergence of non-specific resistance to secondary bacterial infection; however, the accelerated emergence of this population was only observed if the challenge inoculum consisted of the living organism. The relevance of this latter finding to strategies for vaccine development is discussed.

Factors that govern the induction of suppressor T cells after stimulation with concanavalin A (Con A) were investigated in a two-stage culture system. Normal mouse spleen cells were incubated with Con A in the presence of a variety of drugs and then assayed for suppressive activity by means of a secondary anti-sheep erythrocyte response in vitro. The inclusion of inhibitors of mitosis (vinblastine sulphate or mytomycin C) or protein synthesis (cycloheximide or pactamycin) into normal spleen cell cultures containing Con A failed to inhibit the subsequent development of suppressor cells. Furthermore, spleen cells from mice previously irradiated with 900 rad or injected with cyclophosphamide expressed a level of suppressor activity after Con A stimulation which was equivalent to that of normal spleen cells. However, the inclusion of drugs that inhibit microtubule or microfilament function (colchicine or cytocholasin B) did prevent suppressor cell induction. Kinetic studies also revealed that significant suppressor activity was detectable in normal spleen cells after only 3 h exposure to Con A. These results indicate that the induction of suppressor T cells in this system is a maturation event involving changes in the cell membrane and is entirely independent of protein synthesis and cellular proliferation.

This study documents the formation of leukocyte foci in the livers of mice infused with either normal or immune T cells and then challenged intravenously with Listeria monocytogenes. The results show that the transfer of antilisterial resistance occurred before mononuclear cell granuloma formation and was associated instead with the appearance of foci of infiltrating lymphocytes and neutrophils. Numbers of these foci remained low in mice which received immune cells but increased progressively until death in mice which received normal cells. These findings do not support the previous hypothesis that a major component of acquired resistance against Listeria infection involves the rapid generation of mononuclear cell granuloma formation under the control of immune T cells.

The present study was designed to attempt to construct an animal model of the recrudescence of pulmonary tuberculosis by following the course of infection in mice exposed at 3 months of age to a low dose aerosol of Mycobacterium tuberculosis Erdman. The results show that such mice died of overwhelming lung disease at a median age of 24 months; in contrast, uninfected mice lived 4 to 6 months longer. Examination of infected mice over the period of 20 to 24 months of age provided evidence to suggest that recrudescence over this period of time was a general phenomenon within the infected mice, and was characterized by increases in lung bacterial numbers from approximately 10(4) to 10(8) organisms per animal. Experiments designed to investigate the underlying basis of this recrudescence revealed that the capacity of the old mice to mount acquired immunity to the infection exhibited a substantial age-related decline in these animals when compared with young mice exposed to similar aerogenic doses. The data suggest, therefore, that "spontaneous" recrudescence of pulmonary tuberculosis in old mice reflects their inability to recall anamnestic acquired immunity to the reemerging infection.

The purpose of the present study was to examine further the recent hypothesis that subcutaneous infection with Mycobacterium kansasii resulted in the generation of a cell-mediated hypersensitivity reaction ('Koch' reaction) which could, it was argued, subsequently interfere with the generation of acquired immunity following vaccination of the animal with BCG. The results of the present study were unable to confirm this hypothesis in that they show, firstly, that subcutaneous M. kansasii infection was associated with the development of substantial Arthus-like reactivity which masked the detection of any subsequent delayed response, and that furthermore, attempts to adoptively transfer this form of delayed reaction by means of passive transfer of cells were unsuccessful. Furthermore, the results show that, despite the presence of the M. kansasii infection, BCG-vaccinated animals were fully resistant to subsequent aerosol-delivered challenge with virulent M. tuberculosis.

Memory immunity in mice to BCG vaccination is mediated by Thy-1.2+ L3T4+ Lyt-2- cyclophosphamide-resistant T-lymphocytes. The time required for the emergence of acquired memory immunity was inversely proportional to the BCG inoculum size, although the level of memory expressed, once established, was equivalent regardless of the original dose of BCG administered. In mice given a high intravenous dose (10(8)) of BCG, an apparently biphasic memory response was observed, initially peaking on day 15-20, then declining for 10 days or so before increasing again to maximal levels between day 30 and 60 of the infection. This trough in resistance was not due to an active immunosuppressive mechanism; instead, it is suggested that it represents an initial loss or consumption of memory T-cells generated early during the infection which are stimulated to give rise to a state of active immunity as a result of the persisting high mycobacterial load.

The generation of acquired immunity to BCG in mice was compared to that in animals receiving an admixture of BCG and killed Mycobacterium leprae. No significant qualitative differences were observed between the two groups in terms of their generation of delayed sensitivity, of protective T cells, and of immunological memory. In addition, the admixture was as effective as BCG in conferring protective immunity against certain nontuberculous mycobacteria, and in one case, that of M. avium 706, significantly augmented protection.

The results shown here demonstrate that in mice heavily infected with Mycobacterium bovis BCG Pasteur, mitogen-induced levels of interleukin-2 (IL-2) correlate temporally with the state of immunity that is being expressed in the animal during the course of the infection. Active immunity, which is conferred by populations of both CD4+ (L3T4) and CD8+ (Lyt-2) T lymphocytes, and memory immunity, which is mediated by a population of CD4+ T lymphocytes, were identified and distinguished in terms of their sensitivity to cyclophosphamide therapy, their ability to passively transfer specific resistance to infection with virulent Mycobacterium tuberculosis, and their capacity to produce and/or absorb IL-2. In this regard, concanavalin A (Con A)-stimulated L3T4+ and Lyt-2+-enriched splenocytes exhibited an apparent depression in measurable levels of IL-2 when harvested during the first 40 days of the infection, which could be explained by the subsequent observation that these T cells were capable of rapidly absorbing a known quantity of recombinant IL-2 in vitro. Detectable levels of IL-2 in these mitogen-stimulated supernatants began to rise after Day 25, which was temporally associated with a gradual shift from active immunity, to immunity mediated by cyclophosphamide-resistant memory T cells, which did not absorb IL-2 in vitro. These data indicate that fluctuations in apparent IL-2 production reflect changes in the type of immunity being expressed, rather than than some form of defect in IL-2 production.

Adoptive immunization of T cell-deficient recipient mice with M. tuberculosis-specific memory immune T lymphocytes conferred upon these animals the ability to express significantly enhanced resistance both to the homologous infection, and to three strains of nontuberculous mycobacteria. These results support the hypothesis, therefore, that antigenic determinants possessed by the four mycobacterial strains that are relevant to the generation of protective cellular immunity are identical or closely crossreactive.

The activity of the rifampin derivative rifabutin (ansamycin; LM427) was tested in vivo against experimental infections of mice with members of the Mycobacterium avium complex (MAC) and a highly virulent strain of Mycobacterium tuberculosis. Rifabutin was moderately effective against an environmental serotype 8 strain of MAC and of significant effectiveness against low dose aerogenic infections with two strains of higher virulence. The drug was also highly effective in causing the rapid resolution of an intravenous M. tuberculosis infection. These data thus contest a previous negative report on the activity of rifabutin in mouse models of M. avium complex.

The growth of Mycobacterium avium 702 in the spleens and livers of four inbred strains of mice varied such that the mice could be separated into naturally susceptible (BALB/c and C57BL/6) and naturally resistant (A/Tru and DBA/2) strains. This phenomenon was independent of the size of the infecting inoculum of bacteria in that both low (10(4))- and high (10(7))-dose inocula of M. avium grew progressively in susceptible strains and were eliminated from the target organs of resistant strains. Resistance and susceptibility were also demonstrated in in vitro preparations of macrophages from these strains of mice. Over a 7-day period, replication of M. avium in susceptible mouse macrophages was far greater than that in resistant macrophages. Evidence was obtained to suggest that toxic oxygen metabolites were not responsible for this difference. Though no difference was found in the rate of clearance of M. avium from the blood of susceptible or resistant mice, resident macrophages from susceptible mice ingested more M. avium in vitro than did resident macrophages from resistant animals. Growth of M. avium in spleens of susceptible mice induced a large influx of phagocytes, whereas this was not observed in resistant mice. In contrast to this it was found that, after injection of a variety of inflammatory agents, influx of leukocytes into the peritoneal cavity could not be used to distinguish susceptible and resistant strains of mice.

The results of this study show that an isogeneic barrier to the successful adoptive immunotherapy of Mycobacterium tuberculosis infected mice is progressively lost as these animals age. This was shown in this study by the demonstration that increasing levels of transferred acquired specific resistance could be conferred on normal old recipients of greater than 22 months of age, whereas in young recipients such transfers were unsuccessful without the prior exposure of recipients to sublethal levels of whole-body ionizing gamma irradiation. These data indicate that strategies of adoptive immunotherapy in the elderly may not have to overcome the age-related isogeneic physiological barrier to this procedure.

The activation of suppressor T cells by Concanavalin A (Con A) was characterized by changes in expression of T cell surface markers. Treatment of normal mouse spleen cells with monoclonal antibody to Ly-1 prior to activation with Con A consistently reduced the resulting suppressor cell activity as assayed by mixed culture with primed spleen cells responding to sheep erythrocytes. Addition of indomethacin to anti-Ly-1 depleted spleen cells together with Con A had no effect on the resulting suppressor cell activity, whereas its addition to normal spleen cells together with Con A significantly abrogated suppressor T cell induction. In contrast, treatment of normal spleen cells with anti-Ly-1 following 24h culture with Con A had no effect on resulting suppressor cell activity. Treatment of spleen cells either prior to or following culture with Con A with monoclonal antibody to Ly-2 completely abrogated suppressor T cell activity. Suppressor T cells activated by Con A were depleted significantly by fractionation on columns containing insolubilized histamine. In contrast, fractionation of normal spleen cells on histamine columns followed by culture with Con A had no effect on subsequent suppressor T cell induction. Furthermore, direct addition of histamine (5 x 10(-4) M - 5 x 10(-8) M) to Con A preculture had no effect on resultant suppressor T cell generation, suggesting that possible immunodulatory actions of histamine may be expressed at the effector rather than during induction stages of Con A-induced suppressor T cell activation.

Mice infected i.v. with the virulent Erdman strain of Mycobacterium tuberculosis exhibited three distinct phases of infection within the spleen. These consisted of a primary phase, characterized by the progressive growth of the organism; a secondary phase, in which the viable organism was progressively eliminated; and a tertiary phase, characterized by a chronic or slowly recrudescing disease state. Passive transfer experiments, in which T cell-enriched spleen cells from immune donors were infused into T cell-deficient recipients and were measured for their capacity to adoptively protect these mice from challenge with M. tuberculosis, provided evidence that at least three separate populations of protective T cells were acquired in response to the infection within the time frame of the experiments. These populations of T cells could be distinguished in that they differed in their expression of the L3T4 and Lyt-2 cell surface molecules, in terms of their kinetic profiles of emergence and loss, and (c) in terms of their susceptibility to cyclophosphamide. The results may suggest that different populations of protective T cells can be generated at different times during the infection as various classes of antigens (for example, metabolic or structural antigens) become available for presentation by host macrophages. It is hypothesized, furthermore, that the kinetics of emergence and loss of these various populations may reflect switching in the mode of immunity being expressed, particularly during the chronic phase of the infection, from that of a state of active immunity to one of immunologic memory.

Mice heavily infected with Mycobacterium bovis bacillus Calmette-Guérin (BCG) rapidly generated an acquired cellular immune response to this infection, as characterized primarily by the emergence of a splenic T-cell population capable of passively transferring substantial levels of adoptive protection against a challenge infection with M. tuberculosis. The emergence of this protective T-cell population was temporally associated with considerable levels of DNA synthesis in vivo in both the spleen and liver, and with the development of an acquired capacity within the animal to express very high levels of nonspecific resistance to secondary intracellular bacterial infection. Concomitant with the emergence of this acquired response, splenic T cells from infected animals became severely unresponsive to blastogenic in vitro stimulation with the mitogen phytohemagglutinin, and possessed the capacity to suppress the responsiveness of normal T cells in cocultures. Both the unresponsiveness of T cells from infected mice and their immunosuppressive activity in vitro could be essentially ablated by supplementation of the tissue culture medium with a supernatant containing very high titers of the T-cell growth factor interleukin 2 (IL-2). Furthermore, T cells harvested from these animals at the peak of in vitro unresponsiveness exhibited a substantial capacity to absorb or consume IL-2 from IL-2-containing supernatants. It is hypothesized, on the basis of these findings, that mice heavily infected with BCG acquire an IL-2-dependent T-cell population within the spleen in response to this infection, and that the observed in vitro blastogenic unresponsiveness of spleen cells which contain this population may be an artefactual effect arising from the reduction or consumption of available IL-2 within the sustaining culture medium. The relevance of these findings is discussed with particular regard to clinical situations, such as lepromatous leprosy, in which restorative strategies involving the in vivo use of IL-2 are presently being postulated.

The results of this study show that acquired protective immunity to Listeria monocytogenes, measured as the capacity of splenic T cells to passively transfer acquired resistance to normal syngeneic recipients, was mediated by a population of cyclophosphamide-sensitive CD4- CD8+ T cells. The results further show, in contrast, that memory immunity to listeriosis, which was defined as resistance transferred by T cells harvested from infected animals that had received ampicillin chemotherapy, was shown to be mediated by cyclophosphamide-resistant CD4+ CD8- T cells.

The course of infection with Mycobacterium bovis BCG Pasteur was followed against time in groups of mice vaccinated by either the aerogenic or subcutaneous route. The generation of acquired protective immunity and immunological memory was determined in each group by adoptive immunisation procedures. In addition, subcutaneously vaccinated mice were tested for their ability to resist an aerogenic challenge with a lethal dose of M. tuberculosis. No overall qualitative differences in the magnitude or longevity of antituberculosis immunity in mice vaccinated by the two procedures were observed. It is concluded that aerogenic vaccination offers no immunological advantage over vaccination by the subcutaneous route.