Bovis BCG Infection Research Articles

Preexisting inflammation due to Mycobacterium bovis BCG infection differentially modulates T-cell priming against a replicating or nonreplicating immunogen.

Induction of T-cell memory by vaccination ensures long-term protection against pathogens. We determined whether on-going inflammatory responses during vaccination influenced T-cell priming. A preexposure of mice to Mycobacterium bovis BCG impaired their subsequent ability to prime T cells against Listeria monocytogenes. This was characterized by a decrease in L. monocytogenes-specific gamma interferon (IFN-gamma)-secreting CD4(+) and CD8(+) T cells. The intensity of T-cell priming towards L. monocytogenes depended on the extent of L. monocytogenes expansion, and a cessation of this expansion caused by M. bovis BCG-induced inflammation resulted in impairment in T-cell priming. A challenge of M. bovis BCG-infected mice with a higher L. monocytogenes dose increased L. monocytogenes survival and restored T-cell priming towards L. monocytogenes. Impairment in T-cell priming towards L. monocytogenes due to M. bovis BCG-induced inflammation resulted in a compromised protective efficacy in the long term after mice were rechallenged with L. monocytogenes. Preexisting inflammation selectively impaired T-cell priming for replicating immunogens as CD8(+) T-cell response to ovalbumin administered as an inert antigen (ovalbumin-archaeosomes) was enhanced by M. bovis BCG preimmunization, whereas priming towards ovalbumin administered as a live immunogen (L. monocytogenes-ovalbumin) was impaired. Thus, depending on the nature of the immunogen, the presence of prior inflammatory responses may either impede or boost vaccine efficacy.

Hepatoprotective role of Ganoderma lucidum polysaccharide against BCG-induced immune liver injury in mice.

To examine the effect of ganoderma lucidum polysaccharide (GLP) on the immune liver injury induced by BCG infection, and investigate the relationship between degrees of hepatic damage and NO production in mice. Immune hepatic injury was markedly induced by BCG-pretreatment (125 mg.kg(-1), 2-week, iv) or by BCG-pretreatment plus lipopolysaccharide (LPS, 125 microg.kg(-1), 12-hour,iv) in mice in vivo. Hepatocellular damage induced by BCG-pretreated plus inflammatory cytokines mixture (CM), which was included TNF-alpha, IL-1beta, IFN-gamma and LPS in culture medium in vitro. Administration of GLP was performed by oral or incubating with culture medium at immune stimuli simultaneity. Liver damage was determined by activity of alanine aminotransferase (ALT) in serum and in hepatocytes cultured supernatant, by liver weight changes and histopathological examination. NO production in the cultured supernatant was determined by the Griess reaction. Moreover, inducible nitric oxide synthase (iNOS) protein expression was also examinated by immunohistochemical method. Immune hepatic injury was markedly induced by BCG or BCG plus inflammatory cytokines in BALB/c mice in vivo and in vitro. Under BCG-stimulated condition, augment of the liver weight and increase of the serum/supernatant ALT level were observed, as well as granuloma forming and inflammatory cells soakage were observed by microscopic analysis within liver tissues. Moreover, NO production was also increased by BCG or/and CM stimuli in the culture supernatant, and a lot of iNOS positive staining was observed in BCG-prestimulated hepatic sections. Application of GLP significantly mitigated hepatic tumefaction, decreased ALT enzyme release and NO production in serum/supernatant, improved the pathological changes of chronic and acute inflammation induced by BCG-stimuli in mice. Moreover, the immunohistochemical result showed that GLP inhibited iNOS protein expression in BCG-immune hepatic damage model. The present study indicates that NO participates in immune liver injury induced by Mycobacterium bovis BCG infection. The mechanisms of protective roles by GLP for BCG-induced immune liver injury may be due to influence NO production in mice.

Effects of Mycobacterium bovis BCG infection on regulation of L-arginine uptake and synthesis of reactive nitrogen intermediates in J774.1 murine macrophages.

The generation of nitric oxide (NO) by activated macrophages is believed to control mycobacterial infection in the murine system. In this study we examined the effect of Mycobacterium bovis BCG infection on the L-arginine-dependent NO pathway in J774.1 murine macrophages. We have confirmed previous results by demonstrating that stimulation of J774.1 with lipopolysaccharide (LPS) and gamma interferon (IFN-gamma) results in an increase in the uptake of 3H-labeled L-arginine and a concomitant increase in the production of NO. We have also shown that BCG can mimic LPS treatment, leading to enhanced L-[3H]arginine uptake by IFN-gamma-stimulated macrophages. Lipoarabinomannan, a component of the BCG cell wall that is structurally similar to LPS, is not responsible for the uptake stimulation in IFN-gamma stimulated macrophages. Although we demonstrated that there was a 2.5-fold increase in NO production by macrophages 4 h after LPS-IFN-gamma stimulation, BCG infection (with or without IFN-gamma stimulation) did not lead to the production of NO by the macrophages by 4 h postinfection. At 24 h postinfection, the infected macrophages that were stimulated with IFN-gamma produced amounts of NO similar to those of macrophages stimulated with LPS-IFN-gamma. This suggests that there are multiple regulatory pathways involved in the production of NO. Finally, our data suggest that increased expression of the arginine permease, MCAT2B, after 4 h of LPS-IFN-gamma treatment or BCG infection-IFN-gamma treatment is not sufficient to account for the increases in L-[3H]arginine uptake detected. This suggests that the activity of the L-arginine transporter(s) is also altered in response to macrophage activation.

CD40 Ligand Expression in Mycobacterium bovis BCG Infection and Its Regulation by Cytokines : A Direct Role of Interleukin 12

Background Activation and clonal expansion of T cells require not only the recognition of processed antigen on the surface of the antigen-presenting cell (APC) by T-cell receptor (TCR), but also involve co-stimulatory signals that are provided by the simultaneous engagement of cell surface molecules expressed by both the APC and the T cell. Interaction between CD40 and its ligand (CD40L) is known to mediate host immune response and T-cell-mediated effector functions in mycobacterial infections in mice. In this work, we investigated the capacity of Mycobacterium bovis ( M. bovis) BCG to induce the expression of CD40L on human T cells. Methods Human cells were obtained from healthy adults by centrifugation using Ficoll/Hypaque. Cells (1 × 10 6) were incubated in RPMI medium with BCG. After incubation at 37°C in 5% CO 2 atmosphere for 40 h, cells were collected and double-stained with anti-CD40L-PE and anti-CD4-FITC or anti-CD8-FITC mAb. The quantification of positively stained population was based on samples stained with isotype control antibodies analyzed on a FACScan. Results M. bovis BCG stimulation induced a significant amount of CD40L expression on CD4+ T cells, while CD40L was not significantly detected on human CD8+ T cells. The highest CD40L expression on BCG-activated T cells in synergism with interleukin-12 endogenous occurred after a 40-h cell culture with a dose of 10 μg/mL of BCG (84.86 ± 11.77; mean ± standard deviation [SD]). This CD40L expression on BCG-activated human T cells was significantly inhibited by anti-IL-12 mAb (5.08 ± 1.7; mean ± SD). In contrast, anti-IFN-γ and anti-IL-2 mAb did not have an important role in this expression. Conclusions These results indicate that the capacity of BCG to induce CD40L expression on human cells represents a novel mechanism underlying the regulation of cellular responses against tuberculosis. Furthermore, the results showed a direct role of IL-12 to enhance the expression of CD40L on BCG-activated human cells.

Latency-associated peptide of transforming growth factor beta enhances mycobacteriocidal immunity in the lung during Mycobacterium bovis BCG infection in C57BL/6 mice.

Latency-associated peptide of transforming growth factor beta (TGF-beta) (LAP) was used to determine whether in vivo modulation of TGF-beta bioactivity enhanced pulmonary immunity to Mycobacterium bovis BCG infection in C57BL/6 mice. LAP decreased BCG growth in the lung and enhanced antigen-specific T-cell proliferation and gamma interferon mRNA expression. Thus, susceptibility of the lung to primary BCG infection may be partially mediated by the immunosuppressive effects of TGF-beta.

Fatal Mycobacterium bovis BCG Infection in TNF-LT-α-Deficient Mice

Neutralization of TNF or disruption of TNF-R1 leads to fatal Mycobacterium bovis BCG infection. Here we used TNF-LT-α-deficient mice to test whether a complete disruption of TNF and LT-α reduces further host resistance to BCG infection. The bacterial burden especially in the lungs of TNF-LT-α-deficient mice was significantly increased and the mice succumbed to infection between 8 and 10 weeks. In the absence of TNF-LT-α the granulomatous response was severely impaired and delayed. The cells in the granulomas of TNF-LT-α-deficient mice expressed low levels of MHC class II and ICAM-1. They contained a few T cells and F4/80-positive macrophages expressing little iNOS and acid phosphatase activity. By contrast, the lethal action of endotoxin was dramatically reduced in BCG-infected TNF-LT-α-deficient mice. In summary, in the absence of TNF-LT-α the recruitment and activation of mononuclear cells in response to BCG infection were significantly delayed and reduced resulting in immature granulomas allowing uncontrolled fatal infection.

A role for lymphotoxin β receptor in host defense against Mycobacterium bovis BCG infection

A role for lymphotoxin β receptor in host defense against Mycobacterium bovis BCG infection

Effect of the injection of an extract of Ascaris suum on macrophage activation during the early phase of Mycobacterium bovis BCG infection in C57Bl/6 mice

Injection of an Ascaris suum extract (Asc) affects both the humoral and cellular immune responses to unrelated antigens when it is co-administered with these antigens. In the present study we evaluated the effect of Asc on macrophage activation in the early phase of Mycobacterium bovis BCG (Pasteur strain TMCC 1173) infection in C57Bl/6 mice. C57Bl/6 mice were injected intraperitoneally (ip) with 0.1 mg BCG (BCG group) or BCG plus 1 mg Asc (BCG + Asc group). The peritoneal exudates were obtained at 2, 7 and 14 days after infection. The numbers of IFN-gamma-secreting cells were assessed by the ELISPOT assay. Nitric oxide (NO) production was measured by the Griess method and by the evaluation of NADPH diaphorase activity in the peritoneal exudates. The administration of Asc extract increased NADPH diaphorase activity (2 days: control = 0, BCG = 7%, BCG + Asc = 13%, and Asc = 4%; 7 days: control = 4, BCG = 13%, BCG + Asc = 21%, and Asc = 4.5%) and TNF-alpha levels (mean +/- SD; 2 days: control = 0, BCG = 169 +/- 13, BCG + Asc = 202 +/- 37, and Asc = 0; 7 days: control = 0, BCG = 545 +/- 15.5, BCG + Asc = 2206 +/- 160.6, and Asc = 126 +/- 26; 14 days: control = 10 +/- 1.45, BCG = 9 +/- 1.15, BCG + Asc = 126 +/- 18, and Asc = 880 +/- 47.67 pg/ml) in the early phase of BCG infection. Low levels of NO production were detected at 2 and 7 days after BCG infection, increasing at 14 days (mean +/- SD; 2 days: control = 0, BCG = 3.7 +/- 1.59, BCG + Asc = 0.82 +/- 0.005, Asc = 0.48 +/- 0.33; 7 days: control = 0, BCG = 2.78 +/- 1.54, BCG + Asc = 3.07 +/- 1.05, Asc = 0; 14 days: control = 0, BCG = 9.05 +/- 0.53, BCG + Asc = 9.61 +/- 0.81, Asc = 10.5 +/- 0.2 (2 x 10(6)) cells/ml). Furthermore, we also observed that Asc co-injection induced a decrease of BCG-colony-forming units (CFU) in the spleens of BCG-infected mice during the first week of infection (mean +/- SD; 2 days: BCG = 1.13 +/- 0.07 and BCG + Asc = 0.798 +/- 0.305; 7 days: BCG = 1.375 +/- 0. 194 and BCG + Asc = 0.548 +/- 0.0226; 14 days: BCG = 0.473 +/- 0.184 and BCG + Asc = 0.675 +/- 0.065 (x 10(2)) CFU). The present data suggest that Asc induces the enhancement of the immune response in the early phase of BCG infection.

CTLA-4 blockade enhances the immune response induced by mycobacterial infection but does not lead to increased protection.

The murine immune response to a pulmonary mycobacterial infection is slow to develop, allowing bacterial numbers to increase in the lung for several weeks after infection. We sought to enhance the protective immune response induced during Mycobacterium bovis BCG infection by administering an antibody that blocks the interaction of CTLA-4 with its ligands, CD80 and CD86. We found that injection of anti-CTLA-4 monoclonal antibody (MAb) greatly enhanced and accelerated the immune response, as measured by increased cellularity of the draining mediastinal lymph nodes, and enhanced antigen-inducible proliferation and gamma interferon production by mediastinal lymphocytes in vitro. However, despite the apparently enhanced immune response in the mediastinal lymph node following treatment with anti-CTLA-4 MAb, there was no improvement in clearance of mycobacteria in the lungs, liver, or spleen. Examination of the primary site of infection, the lung, revealed that CTLA-4 blockade had no effect on the number or function of lymphocytes infiltrating the infected lung tissue. Taken together, these data suggest that in vivo CTLA-4 blockade enhances mycobacterial-infection-induced lymphocyte expansion and effector cell cytokine production in the draining lymph node but does not alter the number or function of lymphocytes at the primary site of infection and therefore does not lead to enhanced clearance of the infection.

Cytolytic T-cell responses to human dendritic cells and macrophages infected with Mycobacterium bovis BCG and recombinant BCG secreting listeriolysin

Cytolytic T-cell responses from 63 normal blood donors were monitored in a Mycobacterium bovis BCG infection system in vitro. We wanted to know whether cultured dendritic cells were capable of potentiating the cytolytic T-cell responses to M. bovis BCG. Infected cultured dendritic cells were up to ten times more effective antigen-presenting cells than macrophages in proliferative assays, while cytolytic T-cell induction did not differ significantly between dendritic cells and macrophages. Separated CD4 + and CD8 + T-cell subsets contributed equally to lysis of infected targets. Experiments comparing wild-type M. bovis BCG strain with two new recombinant M. bovis BCG strains secreting listeriolysin revealed statistically significant higher maximal lysis values for recombinant M. bovis BCG. We conclude from our in vitro infection system with mycobacteria that dendritic cells are superior to macrophages in proliferative assays but equal to macrophages in their ability to induce cytolytic T-cell responses. Moreover, our data suggest that recombinant M. bovis BCG vaccine strains secreting listeriolysin improve cytolytic T-cell responses.

Effect of fetal calf serum on cytokine release by bone marrow-derived macrophages during infection with intracellular bacteria

Bone marrow-derived macrophages (BMM) comprise a population of quiescent cells which can be activated by defined signals. Here, we directly compare the release of chemokines and monokines by BMM raised either in serum-supplemented or in serum-free medium in response to Listeria monocytogenes EGD or Mycobacterium bovis BCG infection. We focused on this issue because there have been several controversial reports on the production of cytokines by BMM due to different in vitro culture conditions. Culture in serum-supplemented medium primed BMM for release of monocyte chemoattractant protein (MCP)-1, interleukin (IL)-6, IL-12, but had no effect on macrophage inflammatory protein (MIP)-1oc and tumor necrosis factor (TNF)-α production in response to L. monocytogenes infection. After challenge infection with M. bovis, BMM raised and stimulated in serum-supplemented medium secreted higher levels of MCP-1, MIP-1oc, IL-6, TNF-α but not of IL-12 as compared to BMM cultured and infected in a serum-free medium. The effects of serum could be partially mimicked by interferon-γ. Because the serum components responsible for BMM priming are undefined, BMM cultured under serum-free conditions provide an appropriate cell population for defining macrophage activating signals.

The hygiene hypothesis in the development of atopy and asthma--still a matter of controversy?

The remarkable increase in asthma and atopy prevalThere are now also experimental data to support the hygiene hypothesis. Erb and coworkers demonences during the last few decades in Western societstrated in a murine model of allergen-induced airway ies cannot be explained by changes in genetic factors eosinophilia that M. bovis-BCG infection suppressed or by improvements in diagnostic prodedures only. the local Th2 immune response (IL–5 production by Environmental factors, particularly those associated T cells and eosinophil accumulation into the airwith a Westernized lifestyle, are considered to be ways), and that this effect was dependent upon IFN-c involved in this increase. In the late 1980s, Strachan signaling; IFN-c produced during the immune found that repeated infections in early life may response against mycobacteria appeared to suppress prevent the development of hay fever.1 He further the deleterious Th2 response in the lung.9 suggested that the most significant cause of the increase in atopic diseases in Western countries is the decreased exposure to cross-infections among younger siblings as the result of decreased family Infections that may have an size.2 Concomitantly, Romagnani3 suggested that the immunomodulatory effect rising prevalence of atopy could be related to the strong reduction in childhood infections, particularly It is uncertain which infections may have an immunotuberculous infections that stimulate the production modulatory role in early life, but micro-organisms of cytokines antagonistic to Th2 cell differentation.3 that elicit a vigorous cell-mediated immunity (a This ‘hygiene hypothesis’ has now gained wide Th1-type response), particularly intracellular bacteria acceptance as the most plausible explanation of the and viruses, would be the most logical candidates. increase in atopic diseases. Several recent studies It is probably fundamental that the infection be have provided further evidence that certain invasive invasive, protracted or repeated. Data obtained from infections, such as Mycobacterium tuberculosis,4 human studies thus far have suggested a role for the hepatitis A5 and measles6 infection in childhood may intracellular bacterium, M. tuberculosis, and for the prevent the development of atopy and atopic diseases two single-stranded RNA viruses, measles and hepatin later life. By contrast, no similar effect has been itis A, which generally elicit a lifelong immunity.10 found for vaccination. Neither measles vaccination7 Short-lived immunity is often associated with localnor BCG vaccination in children with atopic heredized mucosal infections (e.g. common cold infecity8 has shown any significant effect on the subtions), whereas long-term protective immunity is a sequent development of atopy. It appears that only feature of many systemic infections (e.g. measles, certain natural infections in childhood that are of polio and mumps).11 Local mucosal infections, even sufficient intensity might be able to decrease susceptithough occurring recurrently, seem not to play a significant role here. However, there is some evidbility to atopy.

Frequencies of IFNγ- and IL-4-producing cells during Mycobacterium bovis BCG infection in two genetically susceptible mouse strains: role of [formula omitted] T cells and NK1.1 cells

Frequencies of IFN γ- and IL-4-producing spleen cells in response to Mycobacterium bovis BCG infection were determined in C57BL 6 and BALB c mice. Both mouse strains express equal innate susceptibility to M. bovis BCG (Bcg s), but differ in their NK1.1 and T-cell activities. M. bovis BCG infection induced higher frequencies ( f ≈ 1 500 ) of antigen-induced IFN γ-secreting spleen cells in C57BL 6 mice as compared to BALB c mice ( f ≈ 1 8000 ). Concanavalin A stimulated almost equal numbers of IFN γ-secreting cells in both mouse strains ( f ≈ 1 50 ). Treatment with anti-NK1.1 mAb of M. bovis BCG-infected C57BL 6 mice did not alter frequencies of IFN γ-secreting cells. Equally low numbers of antigen-induced IL-4-producing cells ( f ≈ 1 3000 ) were determined in both C57BL 6 and BALB c mice during M. bovis BCG infection and treatment of C57BL 6 mice with anti-NK1.1 mAb had no measurable effect on IL-4 producers. Finally, frequencies of IFNγ-producing cells were markedly reduced (10-fold) in M. bovis BCG-infected TCR-β − − gene deletion mutants as compared to their heterozygous controls. Our findings verify that M. bovis BCG infection primarily induces IFNγ-secreting α β T cells of TH1 type and show that the frequencies of these IFNγ producers differ in the two Bcg s mouse strains C57BL 6 and BALB c .

Immune response to Mycobacterium bovis bacille Calmette Guérin infection in major histocompatibility complex class I‐ and II‐deficient knock‐out mice: contribution of CD4 and CD8 T cells to acquired resistance

Knock-out mice with defined major histocompatibility complex (MHC) deficiencies were infected intravenously with Mycobacterium bovis bacille Calmette Guérin (M. bovis BCG) to assess the relative impact of MHC class I- and II-dependent immune responses. Heterozygous control mice were capable of controlling growth of M. bovis BCG, although infection progressed chronically, as assessed by determination of colony-forming units. Furthermore, infected controls developed granulomatous lesions at the site of mycobacterial growth and delayed-type hypersensitivity (DTH) reactions after challenge with purified protein derivative of tuberculin. In vitro, spleen cells from heterozygous control mice produced high concentrations of interferon-gamma (IFN-gamma) after restimulation with mycobacterial antigens. In contrast, the MHC class II-deficient A beta-/- mice, which are virtually devoid of functional CD4 T cells, succumbed to M. bovis BCG infection. Furthermore, A beta-/- mice lacked DTH reactions to tuberculin and only few minute picnotic lesions were formed in livers of infected mice. Finally, spleen cells from infected A beta-/- mice failed to produce measurable IFN-gamma concentrations after restimulation in vitro with various mycobacterial antigen preparations. The capacity of beta 2-microglobulin (beta 2m)-deficient mice, which are devoid of CD8 alpha/beta T cells, to inhibit growth of M. bovis BCG was only slightly affected at low inocula, although significantly higher colony-forming units were detected in spleens. These knock-out mice developed strong DTH responses to tuberculin and their spleen cells produced high levels of IFN-gamma once reactivated by mycobacterial antigens. Furthermore, in livers of infected beta 2m-deficient mice, extravascular infiltrates developed which were more diffuse than those in infected control littermates. Remarkably, the beta 2m-deficient mice were substantially more susceptible to higher inocula of M. bovis BCG than their control littermates. Our data formally prove the essential role of MHC class II-dependent immune mechanisms in all relevant aspects of immunity to M. bovis BCG. In addition, our findings emphasize an important contribution of MHC class I-dependent immunity to effective anti-mycobacterial protection. We assume that CD4 T cells are highly effective in containing M. bovis BCG within distinct granulomatous lesions, but fail to eradicate their intracellular pathogens. It appears most likely that CD8 T cells are also required to achieve this goal.

Nitric oxide production and mycobacterial growth inhibition by murine alveolar macrophages: the sequence of rIFN-γ stimulation and Mycobacterium bovis BCG infection determines macrophage activation

Prestimulation with recombinant-interferon-γ (rIFN-γ) followed by Mycobacterium bovis BCG infection induced high nitric oxide production and potent mycobacterial growth inhibition in murine alveolar macrophages. Reversal of the sequence of treatments caused opposite effects, suggesting that a sequential 2-step process comprising first rIFN-γ stimulation and second mycobacterial infection is operative in the activation of alveolar macrophages.

Assessment of mycobacterial infection and multiplication in macrophages by polymerase chain reaction

The amplification of mycobacterium-specific DNA sequences from samples obtained from infected patients by the polymerase chain reaction (PCR) has been useful in the clinical diagnosis of mycobacterial diseases. Using 20 bp oligonucleotide primers that recognizw a 123 bp repeated sequence present in M. bovis and M. tuberculosis DNA, we describe in detail the conditions of the PCR reaction that allow an assessment of the mycobacterial content of infected macrophages. The results of the highly reproducible, time-efficient PCR technique show good correlation with the widely used colony forming unit (CFU) and [ 3H]uracil incorporation methods for the detection of Mycobacterium. Our method allows an assessment of the level of M. bovis BCG infection from a variety of sources including peritoneal macrophages and macrophage lines, within a few hours, making it the assay of choice for rapid determination of the level of mycobacterial growth in infected cells, in experimental models of mycobacterial infection.

Memory T cell-mediated resistance to Mycobacterium tuberculosis infection in innately susceptible and resistant mice

The memory T cell immune response to Mycobacterium tuberculosis infection was examined in strains of mice which vary in their natural susceptibility to Mycobacterium bovis BCG infection. Naturally susceptible (NS) C57BL/6 and naturally resistant (NR) B6D2 F1 hybrid mice were infected with a sublethal dose of M. tuberculosis and then given antibiotic therapy beginning 2 weeks postinfection. T cells from both strains of mice transferred significant levels of resistance to syngeneic mice challenged aerogenically with M. tuberculosis. This memory response was not substantially reduced by depletion of either L3T4+ or Lyt2+ T cells from the donor mice but was ablated by depletion of both T cell subsets. Cyclophosphamide pretreatment of C57BL/6 memory T cell donors also ablated the resistance transferred to recipient mice. In contrast, B6D2 memory T cells were not affected by cyclophosphamide treatment, suggesting that differences may exist in the metabolic state of the memory T cells in the two donor strains, despite the fact that they both develop similar levels of acquired resistance to a subsequent tuberculous challenge.

Immunomodulation of mouse macrophage killing of Mycobacterium avium in vitro

When C57BL/6 mice were infected intravenously with Mycobacterium avium, bacterial growth continued within the spleen until more than 10(8) CFU/g of tissue were attained. This contrasted with Mycobacterium bovis BCG infections where growth declined after 2 weeks. In vivo M. avium-infected splenic macrophages were harvested from chronically infected mice and cultured in vitro for 4 days at 37 degrees C. The number of viable mycobacteria within the resulting macrophage monolayers decreased when cultured in the presence of autologous sensitized T cells and an exogenous source of interleukin-2 (recombinant interleukin-2; 50 U/ml) compared with untreated controls (P less than 0.05). Incubation of the infected macrophages with autologous T cells and soluble M. avium antigens also significantly reduced the number of viable organisms. These results indicate that the mycobactericidal activity of M. avium-infected macrophages can be enhanced in a way that may have important therapeutic implications for patients infected with this opportunistic pathogen.

Mechanisms of recrudescence of Mycobacterium bovis BCG infection in mice.

The capacity of various immunosuppressive agents to cause a recrudescence of the replication of Mycobacterium bovis BCG in the spleens of chronically infected mice was investigated. The actions of three corticosteroid preparations, cyclosporin A, and anti-T-cell subset monoclonal antibodies were compared. Treatment of mice with hydrocortisone acetate, which depressed the number of splenic lymphocytes and suppressed T-cell responses, most effectively exacerbated the stationary BCG counts, at 4 to 6 months after infection. The magnitude of reactivation was more pronounced in innately resistant CBA/Ca mice than in the susceptible C57BL/6 strain of mice. Splenic bacterial counts were also amplified by anti-L3T4 antibody when the antibody was injected at the chronic phase, whereas cyclosporin A had an effect only during the initial 6 weeks after BCG infection. Cultures of spleen cells from chronically infected mice showed a significant increase in the numbers of viable BCG recovered after 7 days of incubation in the presence of dexamethasone but not with cyclosporin A. The observed differences between the tested immunosuppressive agents indicate that the stationary bacterial counts during chronic BCG infection are maintained by discrete T-cell actions on the infected macrophages.

Immunosuppressive effect of cyclosporin A on Mycobacterium bovis BCG infections in mice.

The effect of increasing doses of cyclosporin A (CsA) given to mice infected intravenously with Mycobacterium bovis BCG was investigated. Development of both tuberculin hypersensitivity and acquired antituberculous resistance was suppressed in a dose-responsive manner. Daily dosages at 100 mg/kg of body weight prevented any reduction in the BCG counts within the lungs, liver, or spleen. This effect was associated with lowered nonspecific resistance to a Listeria monocytogenes challenge and a decline in specific protective immunity adoptively transferred to naive recipients. CsA treatment had no effect on antilisterial activity by activated macrophages or on the antituberculous immunity expressed by specific memory T cells. CsA treatment inhibited the ability of BCG-vaccinated mice to produce gamma interferon (IFN-gamma) after a secondary stimulation with live BCG or with lipopolysaccharide. Spleen cells from BCG-infected mice which were exposed to daily treatment with CsA showed reduced IFN-gamma production in response to purified protein derivative or concanavalin A stimulation, suggesting that the immunosuppressive effect of CsA on BCG-infected mice was expressed by inhibiting the development of effector T cells responsible for the production of IFN-gamma.