The CD28/B7 interaction is not required for resistance to Toxoplasma gondii in the brain but contributes to the development of immunopathology.

Infection of C57BL/6 mice with Toxoplasma gondii leads to chronic encephalitis characterized by infiltration into the brain of T cells that produce IFN-gamma and mediate resistance to the parasite. Our studies revealed that expression of B7.1 and B7.2 was up-regulated in brains of mice with toxoplasmic encephalitis (TE). Because CD28/B7-mediated costimulation is important for T cell activation, we assessed the contribution of this interaction to the production of IFN-gamma by T cells from brains and spleens of mice with TE. Stimulation of splenocytes with Toxoplasma Ag or anti-CD3 mAb resulted in production of IFN-gamma, which was inhibited by 90% in the presence of CTLA4-Ig, an antagonist of B7 stimulation. However, production of IFN-gamma by T cells from the brains of these mice was only slightly reduced (20%) by the addition of CTLA4-Ig. To address the role of the CD28/B7 interaction during TE, we compared the development of disease in C57BL/6 wild-type (wt) and CD28-/- mice. Although the parasite burden was similar in wt and CD28-/- mice, CD28-/- mice developed less severe encephalitis and survived longer than wt mice. Ex vivo recall responses revealed that mononuclear cells isolated from the brains of chronically infected CD28-/- mice produced less IFN-gamma than wt cells, and this correlated with reduced numbers of intracerebral CD4+ T cells in CD28-/- mice compared with wt mice. Taken together, our data show that resistance to T. gondii in the brain is independent of CD28 and suggest a role for CD28 in development of immune-mediated pathology during TE.     

Treatment with anti-L3T4 (CD4) monoclonal antibody reduces the inflammatory response in toxoplasmic encephalitis

Toxoplasma gondii is an important cause of disease of the central nervous system in patients with AIDS. Among the variety of immunologic disorders encountered by AIDS patients is a depletion of CD4+ subpopulation of lymphocytes. In order to determine the role of this population of T lymphocytes in the generation of toxoplasmic encephalitis, mice chronically infected with T. gondii were treated with mAb GK1.5 directed against the cell surface glycoprotein L3T4 (CD4) of T lymphocytes. Histopathologic sections of brains of control and treated animals were examined at regular intervals during and after completion of treatment. The results demonstrated significantly less inflammation in brains of mice during treatment with GK1.5 mAb. In addition, recrudescence of the inflammatory process occurred after discontinuation of treatment. Similar results were observed in experiments in which different strains of mice and T. gondii were used.     

Decreased resistance of B cell-deficient mice to infection with Toxoplasma gondii despite unimpaired expression of IFN-gamma, TNF-alpha, and inducible nitric oxide synthase

The role of B cells in resistance against Toxoplasma gondii was studied using B cell-deficient (muMT) mice. Following peroral infection with 10 cysts of the ME49 strain, all muMT mice survived the acute stage of the infection but died between 3 and 4 wk after infection. In contrast, all control mice were alive at 8 wk after infection. At the stage during which muMT animals succumbed to the infection, parasite replication and pathology were most evident in their brains; small numbers of tachyzoites were also detectable in their lungs. Significantly greater numbers of T. gondii cysts and areas of inflammation associated with tachyzoites were observed in brains of muMT than in control mice. Large areas of necrosis associated with numerous tachyzoites were observed only in brains of muMT mice. Anti-T. gondii IgG Abs were detected only in sera of control mice, whereas similar levels of IFN-gamma were detected in sera of both strains of mice. Amounts of mRNA for IFN-gamma, IL-10, and inducible NO synthase in the brain did not differ between infected muMT and control mice. Expression of mRNA for TNF-alpha was increased in brains of muMT mice. Administration of polyclonal rabbit anti-T. gondii IgG Ab prevented early mortality and pathology associated with tachyzoites in the brain in the infected muMT mice. These results indicate that B cells play an important role, most likely through their production of specific Abs, in resistance to persistent active (tachyzoite) infection with T. gondii in mice, especially in the brain and lung.     

Induction of tumor necrosis factor-alpha and inducible nitric oxide synthase fails to prevent toxoplasmic encephalitis in the absence of interferon-gamma in genetically resistant BALB/c mice

Following infection with Toxoplasma gondii, certain strains of mice, such as BALB/c, are genetically resistant to development of toxoplasmic encephalitis (TE) and establish a latent chronic infection as do humans. Thus, these animals appear to be a suitable model to analyze the mechanism of resistance to TE. Since the mechanism for their genetic resistance is unknown, we examined the role of interferon-gamma (IFN-gamma) tumor necrosis factor-alpha (TNF-alpha) and inducible nitric oxide synthase (iNOS) in the resistance using BALB/c-background IFN-gamma-deficient (IFN-gamma(-/-)) mice. IFN-gamma(-/-) and control mice were infected with the ME49 strain of T. gondii and treated with sulfadiazine to establish chronic infection. After discontinuing sulfadiazine, the IFN-gamma(-/-) mice all died, whereas the control mice all survived. Histological studies revealed remarkable inflammatory changes associated with large numbers of tachyzoites in brains of the IFN-gamma(-/-) mice but not in the control mice after discontinuation of sulfadiazine. Large amounts of mRNA for tachyzoite-specific SAG1 were detected in brains of only the IFN-gamma(-/-) mice. IFN-gamma mRNA was detected in brains of only the control mice, whereas mRNA for TNF-alpha and iNOS were detected in brains of both strains of mice. The amounts of the mRNA for TNF-alpha and iNOS did not differ between these mice. Treatment of IFN-gamma(-/-) mice with recombinant IFN-gamma prevented development of TE. These results demonstrate that IFN-gamma is crucial for genetic resistance of BALB/c mice against TE and that TNF-alpha and iNOS are insufficient to prevent TE in the absence of IFN-gamma.     

Identification of a 200- to 300-fold repetitive 529 bp DNA fragment in Toxoplasma gondii, and its use for diagnostic and quantitative PCR

We have identified a novel 529bp fragment that is repeated 200- to 300-fold in the genome of Toxoplasma gondii. This 529bp fragment was utilised for the development of a very sensitive and specific PCR for diagnostic purposes, and a quantitative competitive-PCR for the evaluation of cyst numbers in the brains of chronically infected mice. The 529bp fragment was found in all 60 strains of T. gondii tested, and it discriminates DNA of T. gondii from that of other parasites. Toxoplasma gondii DNA was detected in amniotic fluid of patients, as well as in various tissues from infected mice. Polymerase chain reaction with the 529bp fragment was more sensitive than with the 35-copy B1 gene. For the quantitative competitive-PCR, a 410-bp competitor molecule was co-amplified with similar efficiency as the 529bp fragment. Quantitative competitive-PCR produced a linear relationship between the relative amounts of PCR product and the number of tachyzoites in the range of 10(2)-10(4) tachyzoites and 100-3000 tissue cysts. A highly significant correlation between visual counting of brain cysts and quantitative competitive-PCR was obtained in mice chronically infected with Toxoplasma. Thus, quantitative competitive-PCR with the 529bp fragment can be used as an alternative for the tedious visual counting of brain cysts in experimental animals. With the quantitative competitive-PCR, furthermore, we could confirm the copy number of the 529bp fragment in tachyzoites and estimate the number of bradyzoites per cyst.     

Simultaneous depletion of CD4+ and CD8+ T lymphocytes is required to reactivate chronic infection with Toxoplasma gondii.

C57BL/6 mice chronically infected with an avirulent strain (ME-49) of Toxoplasma gondii were used to study the mechanisms by which T lymphocytes and IFN-gamma prevent reactivation of latent infection. Infected animals were treated with mAb, either anti-CD8, anti-CD4, anti-CD4 plus anti-CD8, anti-IFN-gamma, or anti-CD4 plus anti-IFN-gamma and the mice followed for survival, histopathology, cyst numbers, and spleen cell cytokine responses. In agreement with previously published findings, treatment with anti-IFN-gamma antibodies fully reactivated the asymptomatic infection, inducing massive necrotic areas in the brain with the appearance of free tachyzoites and death of all animals within 2 wk. Mice treated with the combination of anti-CD4 plus anti-CD8 antibodies showed augmented pathology and mortality nearly identical to the anti-IFN-gamma- treated animals. In contrast, treatment with anti-CD4 or anti-CD8 mAb alone failed to result in significantly enhanced brain pathology or mortality. In additional experiments, full reactivation of infection was observed in mice treated with anti-CD4 plus anti-IFN-gamma indicating that CD4+ lymphocytes are not required for the pathology resulting from IFN-gamma neutralization. Cytokine measurements on parasite Ag-stimulated spleen cells from mAb-treated mice indicated that both CD4+ and CD8+ cells produce IFN-gamma whereas only CD4+ cells contribute to parasite Ag-induced IL-2 synthesis. Together, these results suggest that CD4+ and CD8+ lymphocytes act additively or synergistically to prevent reactivation of chronic T. gondii infection probably through the production of IFN-gamma.     

Microglia and macrophages as innate producers of interferon-gamma in the brain following infection with Toxoplasma gondii

We previously reported the requirement of interferon-gamma (IFN-gamma) expression by cells other than T and natural killer (NK) cells in the brain, in addition to T cells, for prevention of toxoplasmic encephalitis following infection with Toxoplasma gondii. In the present study, we analysed the identity of the IFN-gamma-producing non-T, non-NK cells in the brain using infected athymic nude and SCID mice that lack T cells but express IFN-gamma in their brains. Intracellular staining for IFN-gamma followed by flow cytometry revealed that approximately 45-60% of the cells expressing IFN-gamma in their brains were positive for CD11b or F4/80 on their surfaces. Smaller portions of the cells were positive for pan-NK marker. Further smaller portions were positive for CD11c, and these cells were less than 5% of the IFN-gamma-expressing cells in brains of infected SCID mice. In addition to IFN-gamma proteins, large amounts of mRNA for IFN-gamma were detected in CD11b+ cells purified from brains of infected mice, but it was not the case in the cells obtained from uninfected animals. In infected SCID mice depleted of NK cells by treatment with anti-asialo-GM1 antibody, cells expressing IFN-gamma in their brains were all positive for CD11b, and the IFN-gamma-producing cells were detected in both CD45low and CD45high populations. These results suggest that CD11b+ CD45low microglia and CD11b+ CD45high blood-derived macrophages are the major non-T, non-NK cells which express IFN-gamma in the brain of mice infected with T. gondii.     

Toxoplasma gondii: determination of the onset of chronic infection in mice and the in vitro reactivation of brain cysts

Toxoplasma gondii is an intra-cellular parasite that infects humans through vertical and horizontal transmission. The cysts remain dormant in the brain of infected humans and can reactivate in immunocompromised hosts resulting in acute toxoplasmic encephalitis which may be fatal. We determined the onset and progression of brain cysts generation in a mouse model following acute toxoplasmosis as well as the ability of brain cysts to reactivate in vitro. Male Balb/c mice, (uninfected control group, n = 10) were infected orally (study group, n = 50) with 1000 tachyzoites of T. gondii (ME49 strain) and euthanized at 1, 2, 4, 8 and 16 weeks post infection. Brain tissue was harvested, homogenized, stained and the number of brain cysts counted. Aliquots of brain homogenate with cysts were cultured in vitro with confluent Vero cells and the number of cysts and tachyzoites counted after 1 week. Brain cysts but not tachyzoites were detected at week 2 post infection and reached a plateau by week 4. In vitro Vero cells culture showed similar pattern for cysts and tachyzoites and reactivation of cyst in vitro was not influenced by the age of the brain cysts.     

The immunocytochemical demonstration of Toxoplasma antigen in the brains of congenitally infected mice

The peroxidase anti-peroxidase immunocytochemical staining method was used to identify Toxoplasma antigen in paraffin embedded sections of the brains of 22 mice congenitally infected with the parasite. Intact Toxoplasma tissue cysts were readily demonstrated in the brain in all cases. In 4 of the 22 infected mice there was evidence of rupture of the cyst wall and/or presence of extra-cystic Toxoplasma antigen. Further support for the extra-cystic location of Toxoplasma antigen was obtained by electron microscopy of reprocessed tissue which revealed endozoites in the area immediately surrounding a ruptured cyst. The possible implications of these findings in relation to the pathogenesis of congenital toxoplasmic meningo-encephalitis are discussed.     

The induction and kinetics of antigen-specific CD8 T cells are defined by the stage specificity and compartmentalization of the antigen in murine toxoplasmosis

Toxoplasma gondii forms different life stages, fast-replicating tachyzoites and slow-growing bradyzoites, in mammalian hosts. CD8 T cells are of crucial importance in toxoplasmosis, but it is unknown which parasite stage is recognized by CD8 T cells. To analyze stage-specific CD8 T cell responses, we generated various recombinant Toxoplasma gondii expressing the heterologous Ag beta-galactosidase (beta-gal) and studied whether 1) secreted or cytoplasmic Ags and 2) tachyzoites or bradyzoites, which persist intracerebrally, induce CD8 T cells. We monitored the frequencies and kinetics of beta-gal-specific CD8 T cells in infected mice by MHC class I tetramer staining. Upon oral infection of B6C (H-2(bxd)) mice, only beta-gal-secreting tachyzoites induced beta-gal-specific CD8 T cells. However, upon secondary infection of mice that had received a primary infection with tachyzoites secreting beta-gal, beta-gal-secreting tachyzoites and bradyzoites transiently increased the frequency of intracerebral beta-gal-specific CD8 T cells. Frequencies of splenic and cerebral beta-gal-specific CD8 T cells peaked at day 23 after infection, thereafter persisting at high levels in the brain but declining in the spleen. Splenic and cerebral beta-gal-specific CD8 T cells produced IFN-gamma and were cytolytic upon specific restimulation. Thus, compartmentalization and stage specificity of an Ag determine the induction of CD8 T cells in toxoplasmosis.     

Endogenous IFN-gamma is required for resistance to acute Trypanosoma cruzi infection in mice

In order to study the role of endogenous IFN-gamma in Trypanosoma cruzi infection in mice, a potent murine IFN-gamma-specific mAb was injected i.p. on days -1, 7, and 14, relative to infection. Irrespective of the parasite inocula (100 or 25,000), groups of antibody-treated mice had significantly greater cumulative mortality rates than did appropriate controls. In antibody-treated mice, mean survival times were also significantly shorter, and maximum mean parasitemia levels were significantly higher, than in controls. Moreover, the number of amastigote nests in tissues was higher than in control mice and attained a maximum at the same time as parasitemia. As evident from kinetic studies of neutralizing activity, injected mAb were rapidly consumed in infected, but not in noninfected, mice, which is suggestive of massive IFN-gamma production during the early parasitemic phase of the disease. Nevertheless, IFN-gamma remained undetectable in the sera of infected but untreated mice. Unexpectedly, however, a peak of IFN-like antiviral activity, characterizable as a mixture of IFN-gamma and IFN-beta, appeared in mAb-treated mice that survived to infection at a time when neutralizing activity of injected mAb had drastically decreased in the circulation. We hypothesize that this high level of artificially induced endogenous IFN-gamma, not neutralized by the amounts of injected mAb, was due to the more intense parasite multiplication occurring in mAb-treated mice, which in turn may have induced an increased amount of various cytokines. TNF-alpha was not found in the serum of our mice. The humoral immune response entered its exponential phase at a time point later than that when protection by endogenous IFN-gamma was evident. Treatment with IFN-gamma-specific antibody, as applied in our study, failed to affect the level of different Ig isotypes or of T. cruzi-specific antibodies. Our study clearly indicates that IFN-gamma is produced early in acute T. cruzi infection and exerts a protective effect that is probably independent from the humoral immune response.     

Correlation between increased susceptibility to primary Toxoplasma gondii infection and depressed production of gamma interferon in pregnant mice.

To explore a possible mechanism of pregnancy-associated suppression of T cell-mediated immunity to Toxoplasma gondii, acquired resistance and gamma interferone (IFN-gamma) production in pregnant mice were compared with those in virgin mice after infection with the S-273 strain of this protozoan parasite. The 50% lethal dose of this strain was less than 200 tachyzoites for pregnant mice and 2,800 organisms for virgin controls. Toxoplasma-induced production of both IFN-alpha and IFN-gamma in the bloodstream of pregnant mice was significantly depressed as compared with that in virgin controls. The administration of recombinant murine IFN-gamma (rMuIFN-gamma) resulted in a significant decrease of mortality and parasitic growth in the organs of pregnant mice infected with a lethal dose of S-273 strain tachyzoites. Thus, the impairment of T cell-mediated immune responses was evident in pregnant mice from the impaired IFN-gamma-generating capacity and poor survival rate after primary infection with Toxoplasma. When mice with chronic Toxoplasma infection were injected with specific antigen, the resultant production of IFN-gamma was also significantly suppressed during pregnancy. However, there was no direct correlation between the serum levels of IFN-gamma and susceptibility to reinfection, since the mortality rate of chronically infected pregnant mice after the challenge with the high virulent RH strain was not significantly higher than that of virgin controls.     

Congenital toxoplasmosis in a Indo-Pacific bottlenose dolphin (Tursiops aduncus)

Toxoplasma gondii was identified in tissues of a stillborn late-term fetus from an Indo-Pacific bottlenose dolphin (Tursiops aduncus). Myocardial necrosis and nonsuppurative inflammation in the heart and nonsuppurative necrotizing encephalitis were associated with tachyzoites and tissue cysts. The diagnosis was confirmed by immunohistochemical staining with anti-T. gondii-specific polyclonal rabbit serum.     

Evidence for Finely-Regulated Asynchronous Growth of Toxoplasma gondii Cysts Based on Data-Driven Model Selection

Toxoplasma gondii establishes a chronic infection by forming cysts preferentially in the brain. This chronic infection is one of the most common parasitic infections in humans and can be reactivated to develop life-threatening toxoplasmic encephalitis in immunocompromised patients. Host-pathogen interactions during the chronic infection include growth of the cysts and their removal by both natural rupture and elimination by the immune system. Analyzing these interactions is important for understanding the pathogenesis of this common infection. We developed a differential equation framework of cyst growth and employed Akaike Information Criteria (AIC) to determine the growth and removal functions that best describe the distribution of cyst sizes measured from the brains of chronically infected mice. The AIC strongly support models in which T. gondii cysts grow at a constant rate such that the per capita growth rate of the parasite is inversely proportional to the number of parasites within a cyst, suggesting finely-regulated asynchronous replication of the parasites. Our analyses were also able to reject the models where cyst removal rate increases linearly or quadratically in association with increase in cyst size. The modeling and analysis framework may provide a useful tool for understanding the pathogenesis of infections with other cyst producing parasites.     

TCR V beta 8+ T cells prevent development of toxoplasmic encephalitis in BALB/c mice genetically resistant to the disease

BALB/c are genetically resistant to development of toxoplasmic encephalitis (TE) when infected with Toxoplasma gondii, whereas CBA/Ca mice are susceptible. We compared TCR Vbeta chain usage in lymphocytes infiltrated into brains between these animals following infection. TCR Vbeta8(+) cells were the most frequent T cell population in brains of infected, resistant BALB/c mice, whereas TCR Vbeta6(+) T cells were more prevalent than Vbeta8(+) T cells in brains of infected, susceptible CBA/Ca mice. Adoptive transfer of Vbeta8(+) immune T cells, obtained from infected BALB/c mice, prevented development of TE and mortality in infected athymic nude mice that lack T cells. In contrast, adoptive transfer of Vbeta6(+) immune T cells did not prevent development of TE or mortality in the nude mice. The protective activity of Vbeta8(+) immune T cells was greater than that of the total Vbeta8(-) population. In addition, Vbeta8(+) immune T cells produced markedly greater amounts of IFN-gamma than did the Vbeta8(-) population after stimulation with tachyzoite lysate Ags in vitro. Thus, Vbeta8(+) T cells appear to play a crucial role in the genetic resistance of BALB/c mice against development of TE.     

Administration of anti-IL-4 monoclonal antibody 11B11 increases the resistance of mice to Listeria monocytogenes infection.

The role of endogenous IL-4 in resistance to Listeria monocytogenes infection was investigated by in vivo administration of an anti-IL-4 mAb (11B11). Mice treated with 0.01 to 0.4 mg of anti-IL-4 mAb before L. monocytogenes challenge demonstrated a significantly reduced peak bacterial burden in their livers and spleens and accelerated bacterial clearance from these organs. In addition, histopathologic damage to the liver was reduced. Maximal protection was achieved by i.p. injection of 0.1 mg of anti-IL-4 mAb 2 or 24 h before L. monocytogenes challenge; treatment with anti-IL-4 mAb after injection of L. monocytogenes had no effect on antilisterial resistance. Anti-IL-4 mAb-treated and control Listeria-infected mice exhibited similar patterns of IFN-gamma, IL-2, and IL-4 mRNA, as determined by polymerase chain reaction amplification of RNA extracted from spleen cells. In both anti-IL-4 mAb-treated and control mice, IFN-gamma, IL-2, and IL-4 mRNA were produced within 4 h after challenge. Cytokine mRNA levels were similar for anti-IL-4 mAb-treated and control mice, except for the greater amount of IFN-gamma mRNA in the anti-IL-4 mAb-treated mice at 4 h after L. monocytogenes challenge. IFN-gamma and IL-2 mRNA levels were sustained for at least 5 days, whereas IL-4 mRNA was undetectable by 3 days after challenge. There were no significant differences in the amounts of IL-4 and IFN-gamma detected in culture supernatants of spleen cells from anti-IL-4 mAb-treated and control Listeria-infected mice. These results suggest that endogenous IL-4, a cytokine thought to be produced principally by Th2 cells, has a deleterious effect on host defense against the facultative intracellular pathogen L. monocytogenes. Administration of an anti-IL-4 mAb increases antilisterial resistance without causing a detectable shift to a Th1 type of cytokine response.     

Detection of the initial site of Toxoplasma gondii reactivation in brain tissue

Detection of the initial site of Toxoplasma gondii reactivation in brain tissue is difficult because the number of latent cysts is small and reactivation is a transient event. To detect the early stage of reactivation in mouse brain tissue, we constructed a cyst-forming strain of T. gondii in the tachyzoite stage, specifically expressing red fluorescence. The PLK strain of T. gondii was stably transfected with a red fluorescent protein gene, DsRed Express, under the control of a tachyzoite-specific SAG-1 promoter and the resulting parasite was designated as PLK/RED. Tachyzoites of PLK/RED growing in Vero cells showed red fluorescence. When C57BL/6J mice were i.p. infected with tachyzoites of PLK/RED, red fluorescent tachyzoites were detected in their brains at the fourth day p.i. However, red fluorescent tachyzoites were not detected in BALB/c mice latently infected with PLK/RED, although non-fluorescent cysts were detected in their brains. After treatment of latently infected mice with dexamethasone for 1 month, the mice showed neurological symptoms. In mice with symptoms, red fluorescent tachyzoites were again detected in their brains and in other organs. To detect the initial site of reactivation, BALB/c mice latently infected with the strain were treated with dexamethasone for 3 weeks, and brains were excised before any symptoms appeared. Excised brains were examined for red fluorescence-positive sites. By a histological study of red fluorescent-positive sites, we detected a cyst containing red fluorescent zoites, which still had a PAS stain-positive cyst wall. A few red fluorescent zoites breaking away from the cyst were also observed. The stage-specific expression of fluorescent protein facilitates detection of a rare transient event and makes it possible to detect the initial site of reactivation.     

Toxoplasma gondii: in vivo expression of BAG-5 and cyst formation is independent of TNF p55 receptor and inducible nitric oxide synthase functions

Wild type, TNFRp55(-/-), iNOS(-/-) and IFN-gamma(-/-) mice were infected with Toxoplasma gondii strain ME-49, and the central nervous system (CNS), lungs, liver, spleen, heart and kidneys were examined for the presence of parasites expressing tachyzoite-specific (SAG-1) and bradyzoite-specific (BAG-5) antigens. During the acute phase of infection, the peripheral organs, but not the CNS, of the IFN-gamma(-/-) mice are heavily parasitized by tachyzoites and there are no signs of parasites expressing BAG-5. In contrast, the tissues from TNFRp55(-/-) and inducible nitric oxide synthase (iNOS)(-/-) mice, mainly the CNS, presented high numbers of parasites expressing SAG-1 and/or BAG-5. Tachyzoite transformation into bradyzoite and cyst development was shown to be normal in the tissues from TNFRp55(-/-) and iNOS(-/-) mice, as indicated by the high numbers of BAG-5/PAS positive cysts. Consistently, reactivation of infection in IFN-gamma(-/-) mice was rapid and characterized by a dramatic increase in SAG-1, contrasting with slow course in the TNFRp55(-/-) or iNOS(-/-) mice associated with a relatively small increase in SAG-1- and/or BAG-5-positive parasites. In conclusion, our results suggest that the control of multiplication of tachyzoites is largely dependent on endogenous IFN-gamma with partial involvement of TNFRp55 and iNOS. In contrast, induction of BAG-5 expression and cyst formation during toxoplasmosis seems to be dependent on IFN-gamma, but independent of TNFRp55 and iNOS functions.     

The effect of anti-IFN-gamma antibody on the protective effect of Lyt-2+ immune T cells against toxoplasmosis in mice

The effect of an IFN-gamma mAb on the protective activity of immune T cells against Toxoplasma infection was examined in a murine model of toxoplasmosis. Mice that received anti-IFN-gamma antibody and immune spleen cells all died of toxoplasmosis after challenge with Toxoplasma tachyzoites. In contrast, mice that received normal IgG and immune spleen cells all survived the infection. The protective activity of Lyt-2+ immune T cells, previously shown to be the principal mediators of resistance against Toxoplasma in mice was completely ablated by the anti-IFN-gamma mAb. These results suggest that IFN-gamma is the major mediator of the resistance against Toxoplasma infection in mice which is conferred by immune T cells.     

Toxoplasma gondii: inconsistent dissemination patterns following oral infection in mice

Since Toxoplasma gondii is transmitted in the wild through the ingestion of infective cysts, oral infection is a preferred model for studying the natural mode of parasite dissemination and pathogenesis. Using luciferase-expressing strains of T. gondii and in vivo imaging, we observed different patterns of disease progression in mice depending of the method of oral infection. Oral gavage of infective cysts (e.g., bradyzoites) resulted in an inconsistent pattern of parasite dissemination; in the majority (20/29) of infected mice, luciferase-derived signal (indicating high numbers of Toxoplasma tachyzoites) was first observed in the right chest area. At later time points this signal spread to other parts of the mouse, including the abdominal area. In the remaining mice (9/29), parasites were first observed replicating in the abdominal area, as might be expected. In contrast, when mice were infected naturally (either via ingestion of whole brains from previously infected mice or brain cyst homogenate-soaked bread), parasites were first observed replicating in the abdominal area in all mice examined (10/10). Based on the inconsistency of infections initiated with oral gavage, it is recommended that natural feeding be used to infect mice when a consistent oral infection is desired.