Subdominant/cryptic CD8 T cell epitopes contribute to resistance against experimental infection with a human protozoan parasite

During adaptive immune response, pathogen-specific CD8(+) T cells recognize preferentially a small number of epitopes, a phenomenon known as immunodominance. Its biological implications during natural or vaccine-induced immune responses are still unclear. Earlier, we have shown that during experimental infection, the human intracellular pathogen Trypanosoma cruzi restricts the repertoire of CD8(+) T cells generating strong immunodominance. We hypothesized that this phenomenon could be a mechanism used by the parasite to reduce the breath and magnitude of the immune response, favoring parasitism, and thus that artificially broadening the T cell repertoire could favor the host. Here, we confirmed our previous observation by showing that CD8(+) T cells of H-2(a) infected mice recognized a single epitope of an immunodominant antigen of the trans-sialidase super-family. In sharp contrast, CD8(+) T cells from mice immunized with recombinant genetic vaccines (plasmid DNA and adenovirus) expressing this same T. cruzi antigen recognized, in addition to the immunodominant epitope, two other subdominant epitopes. This unexpected observation allowed us to test the protective role of the immune response to subdominant epitopes. This was accomplished by genetic vaccination of mice with mutated genes that did not express a functional immunodominant epitope. We found that these mice developed immune responses directed solely to the subdominant/cryptic CD8 T cell epitopes and a significant degree of protective immunity against infection mediated by CD8(+) T cells. We concluded that artificially broadening the T cell repertoire contributes to host resistance against infection, a finding that has implications for the host-parasite relationship and vaccine development.     

Infection with Trypanosoma cruzi restricts the repertoire of parasite-specific CD8+ T cells leading to immunodominance

Interference or competition between CD8(+) T cells restricted by distinct MHC-I molecules can be a powerful means to establish an immunodominant response. However, its importance during infections is still questionable. In this study, we describe that following infection of mice with the human pathogen Trypanosoma cruzi, an immunodominant CD8(+) T cell immune response is developed directed to an H-2K(b)-restricted epitope expressed by members of the trans-sialidase family of surface proteins. To determine whether this immunodominance was exerted over other non-H-2K(b)-restricted epitopes, we measured during infection of heterozygote mice, immune responses to three distinct epitopes, all expressed by members of the trans-sialidase family, recognized by H-2K(b)-, H-2K(k)-, or H-2K(d)-restricted CD8(+) T cells. Infected heterozygote or homozygote mice displayed comparably strong immune responses to the H-2K(b)-restricted immunodominant epitope. In contrast, H-2K(k)- or H-2K(d)-restricted immune responses were significantly impaired in heterozygote infected mice when compared with homozygote ones. This interference was not dependent on the dose of parasite or the timing of infection. Also, it was not seen in heterozygote mice immunized with recombinant adenoviruses expressing T. cruzi Ags. Finally, we observed that the immunodominance was circumvented by concomitant infection with two T. cruzi strains containing distinct immunodominant epitopes, suggesting that the operating mechanism most likely involves competition of T cells for limiting APCs. This type of interference never described during infection with a human parasite may represent a sophisticated strategy to restrict priming of CD8(+) T cells of distinct specificities, avoiding complete pathogen elimination by host effector cells, and thus favoring host parasitism.     

A DNA-priming protein-boosting regimen significantly improves type 1 immune response but not protective immunity to Trypanosoma cruzi infection in a highly susceptible mouse strain

BALB/c or C57Bl/6 mice immunized with plasmids containing Trypanosoma cruzi genes developed specific immune responses and protective immunity against lethal parasitic infection. In contrast, in the highly susceptible mouse strain A/Sn, DNA vaccination reduced the peak parasitemia but promoted limited mouse survival after challenge. In the present study, we tested whether the immunogenicity and protective efficacy of vaccination could be improved by combining DNA and recombinant protein immunization regimens. A/Sn mice immunized with plasmid p154/13 which harbours the gene encoding Trypanosoma cruzi trans-sialidase developed a predominant type 1 immune response. In contrast, immunization with the recombinant Trypanosoma cruzi trans-sialidase protein adsorbed to alum generated a typical type 2 immune response. Simultaneous administration of both p154/13 and recombinant Trypanosoma cruzi trans-sialidase protein also led to a predominant type 2 immune response. Sequential immunization consisting of two priming doses of p154/13 followed by booster injections with recombinant Trypanosoma cruzi trans-sialidase protein significantly improved specific type 1 immune response, as revealed by a drastic reduction of the serum IgG1/IgG2a ratio and by an increase in the in vitro interferon-gamma secretion by CD4 T cells. Our observations confirm and extend previous data showing that a DNA-priming protein-boosting regimen might be a general strategy to enhance type 1 immune response to DNA vaccines. Upon challenge with Trypanosoma cruzi, no improvement in protective immunity was observed in mice immunized with the DNA-priming protein-boosting regimen when compared to animals that received DNA only. Therefore, our results suggest that in this experimental model there is no correlation between the magnitude of type 1 immune response and protective immunity against Trypanosoma cruzi infection.     

Trypanosoma cruzi: cellular and antibody response against the parasite in mice immunized with a 19-amino acid synthetic peptide

Several monoclonal antibodies were prepared against the flagellar fraction of Trypanosoma cruzi epimastigotes (Tulahuén strain, stock Tul 2). One of them, FCH-F8-4, has previously shown biologic activity against the parasite (complement-mediated lysis and neutralization of the trypomastigote infectivity). Immunopurified antigens using this monoclonal antibody elicited a protective immune response in mice. Two recombinant cDNA clones were detected with this anti-flagellar fraction monoclonal antibody on a lambda gt11 expression library prepared from T. cruzi epimastigote mRNA. The insert of one of these cDNA clones, lambda(FCH-F8-4)1 (150 bp) coded for a 19-amino acid peptide (PAFLGCSSRFSGSFSGVEP). This insert hybridized with a 5.0-kb mRNA from epimastigotes. The beta-galactosidase fusion protein was produced in lysogenic bacteria. The monoclonal antibody recognized the epitope present in the fusion protein after western blotting of the crude lysate. A synthetic peptide (SP4) containing the complete sequence of lambda(FCH-F8-4)1 was constructed on solid phase. This peptide was able to inhibit the ELISA reactivity (in a range from 13 to 52%) of flagellar fraction immunized mouse sera and when administered (coupled to KLH or alone) to BALB/c mice with Bordetella pertussis as adjuvant, it induced a humoral and cellular immune response which was detected by ELISA, immunofluorescence, blotting, and DTH reactions against T. cruzi antigens. The immune response obtained indicates that this synthetic peptide resembles the parasite antigen conformation and could be useful for diagnosis purposes or be able to elicit immunoprotection against T. cruzi infection.     

Trypanosoma cruzi: immune response in mice immunized with parasite antigens

The humoral and cellular immune responses were studied in mice immunized with flagellar fraction (F), F plus Bordetella pertussis as adjuvant (F-Bp), and microsomal (Mc) subcellular fractions from the epimastigote forms of Trypanosoma cruzi. The immune response was studied before and after the challenge with 50 bloodstream forms of T. cruzi, Tulahuén strain. The immunization with F-Bp, but not with Mc or F and Bp separately, protected mice, in terms of parasitemia and mortality, from the challenge with the parasite. Before the challenge, levels of specific antibodies in mice immunized with F-Bp were higher than in mice immunized with F or Mc. Antibody levels 17 days after the infection were similar in the three groups of mice while nonimmunized mice reached lower levels. Early during the infection nonimmunized infected mice lacked delayed-type hypersensitivity (DTH) responses to parasite antigens and to concanavalin A (Con A). Mice immunized with F-Bp, however, presented positive DTH responses to parasite antigens and Con A both, before and after the challenge with T. cruzi. DTH reaction was transferred with spleen cells. Mice immunized with Mc behaved similarly to infected nonimmunized animals in their reactivity to parasite antigens. These results indicated striking differences between protected and nonprotected mice in humoral and cellular immune responses during experimental T. cruzi infection.     

Protective immunity against the protozoan Leishmania chagasi is induced by subclinical cutaneous infection with virulent but not avirulent organisms

Protective immunity against Leishmania major is provided by s.c. immunization with a low dose of L. major promastigotes or with dihydrofolate-thymidylate synthase gene locus (DHFR-TS) gene knockout L. major organisms. Whether these vaccine strategies will protect against infection with other Leishmania species that elicit distinct immune responses and clinical syndromes is not known. Therefore, we investigated protective immunity to Leishmania chagasi, a cause of visceral leishmaniasis. In contrast to L. major, a high dose s.c. inoculum of L. chagasi promastigotes was required to elicit protective immunity. Splenocytes from mice immunized with a high dose produced significantly greater amounts of IFN-gamma and lower TGF-beta than mice immunized with a low dose of promastigotes. The development of protective immunity did not require the presence of NK cells. Protection was not afforded by s.c. immunization with either attenuated L. chagasi or with L. major promastigotes, and s.c. L. chagasi did not protect against infection with L. major. Subcutaneous immunization with DHFR-TS gene knockouts derived from L. chagasi, L. donovani, or L. major did not protect against L. chagasi infection. We conclude that s.c. inoculation of high doses of live L. chagasi causes a subclinical infection that elicits protective immune responses in susceptible mice. However, L. chagasi that have been attenuated either by long-term passage or during the raising of recombinant gene knockout organisms do not elicit protective immunity, either because they fail to establish a subclinical infection or because they no longer express critical antigenic epitopes.     

In vivo administration of recombinant IFN-gamma induces macrophage activation, and prevents acute disease, immune suppression, and death in experimental Trypanosoma cruzi infections

Recombinant murine IFN-gamma (rMu-IFN-gamma) was demonstrated to be a potent in vivo activator of mouse peritoneal macrophages to kill Trypanosoma cruzi in vitro and to be capable of conferring protection against death from acute T. cruzi infection. Following i.p. injections of rMu-IFN-gamma, resident peritoneal macrophages were cultured and infected with T. cruzi in vitro. Numbers of intracellular parasites were determined at different times thereafter. Ten or 100 micrograms (1 microgram = 6.5 X 10(5) U) of Mu-IFN-gamma, injected both 24 and 4 h before macrophage harvest, induced up to 99% inhibition of T. cruzi. One microgram of rMu-IFN-gamma was not effective under these conditions. In vitro inhibition of T. cruzi by peritoneal macrophages occurred by 24 h after infection and continued until at least 120 h after infection. There were no significant differences in initial parasite uptake by macrophages from IFN-gamma-treated or control mice, indicating that the rMu-IFN-gamma induced parasite killing. One i.p. dose of 10 micrograms was as effective as two doses if the single injection was given 24 h before macrophage harvest. In subsequent experiments, mice were given multiple injections of 10 micrograms rMu-IFN-gamma beginning 24 h before or 2 h after infection with virulent T. cruzi. Mice treated with rMu-IFN-gamma had significantly lower parasitemias and decreased morbidity compared with control mice. Proliferative responses to Con A and antibody responses to SRBC were not significantly lowered in IFN-gamma-treated mice, in contrast to untreated infected controls. All of the IFN-gamma-treated mice survived acute T. cruzi infection, whereas 100% of saline-treated infected mice died. It was demonstrated in this study that rMu-IFN-gamma activated mouse macrophages in vivo to kill T. cruzi and that rMu-IFN-gamma significantly reduced morbidity and immune suppression, and eliminated mortality resulting from acute infection with this parasite.     

Influence of adjuvants in inducing immune responses to different epitopes included in a multiepitope,multivalent, multistage Plasmodium falciparum candidate vaccine (FALVAC-1) in outbred mice

FALVAC-1, a vaccine against Plasmodium falciparum was developed by joining 21 epitopes from P. falciparum vaccine antigens and an universal T helper epitope from tetanus toxoid. Since adjuvants influence different aspects of immune responses, in this study we investigated the effect of four adjuvants aluminum hydroxide (alum), nonionic copolymer adjuvant P1005 (water-in-oil emulsion), CpG oligodeoxynucleotides (ODN), and QS-21 in eliciting immune responses in outbred mice. QS-21 and copolymer adjuvants were the best formulations in inducing higher and long-lasting antibody titers to the whole vaccine compared to alum and CpG. QS-21 was the only adjuvant to elicit predominantly IgG2a response and antibodies reactive with all epitopes incorporated in the vaccine construct. Vaccine elicited antibodies recognized sporozoites and asexual blood-stage parasites. FALVAC-1 immunized mice induced lymphoproliferative and IFN-gamma response to the vaccine. QS-21 and CpG adjuvants were able to elicit T proliferative responses to 20 of the 22 epitopes in the vaccine. In conclusion, this study demonstrated that with suitable adjuvant such as QS-21, it is possible to elicit immune responses to most of the epitopes included in the FALVAC-1 vaccine.     

BCG expressing LCR1 of Leishmania chagasi induces protective immunity in susceptible mice

Cellular immune responses are required for protective immunity against Leishmania chagasi. Immunization strategies using live intracellular bacteria (e.g., bacille-Calmette Guerin strain of Mycobacterium bovis) expressing recombinant antigens can induce cellular immune responses to these antigens. Previous studies demonstrated that the L. chagasi antigen LCR1 stimulates IFN-gamma production from T cells of infected BALB/c mice, and immunization with recombinant LCR1 partially protects against L. chagasi infection. To determine whether live bacteria could enhance the immunization potential of LCR1, we engineered BCG expressing LCR1 (BCG-LCR1). Subcutaneous immunization with BCG-LCR1, but not with BCG containing plasmid only (BCG-pMV261), elicited better protective immunity against L. chagasi infection than LCR1 protein alone. BCG-LCR1 administered intraperitoneally did not protect. Splenocytes from mice immunized s.c. with either BCG-LCR1 or BCG-pMV261 and then infected with L. chagasi promastigotes had increased antigen-induced IFN-gamma and reduced IL-10 production compared to splenocytes of control mice. We propose that BCG-LCR1 promotes a Th1-type protective immune response, and it may be a useful component of a Leishmania vaccine.     

The surface protein superfamily of Trypanosoma cruzi stimulates a polarized Th1 response that becomes anergic

Trypanosoma cruzi is an obligate intracellular parasite that chronically infects mammals. Extracellular mammalian stage trypomastigotes simultaneously express and release multiple members of the parasite's surface protein superfamily; these extracellular proteins should stimulate MHC class II-restricted CD4 T cells. The surface protein superfamily, however, encodes variant epitopes that may inhibit this CD4 response. In this report the surface protein-specific CD4 response was investigated. CD4 cells isolated from acutely and chronically infected mice did not proliferate when stimulated with surface proteins. Adoptive transfer of surface protein-specific CD4 clones or immunization with a peptide encoding a surface protein T cell epitope protected mice during T. cruzi infection. These data strongly suggested that surface proteins were expressed and presented to CD4 cells during infection. Limiting dilution analysis identified an expanded population of surface protein-specific CD4 cells during the acute and chronic infection. These surface protein-specific CD4 cells did not produce IL-2 or IL-4, but did produce IFN-gamma. Enzyme-linked immunospot analyses confirmed that many of the surface protein-specific CD4 cells produce IFN-gamma. Together these results suggest that during T. cruzi infection a potentially protective CD4 response becomes anergic. It is possible that this anergy is induced by variant T cell epitopes encoded by the surface protein superfamily.     

The Trypanosoma cruzi Tc52-released protein induces human dendritic cell maturation,signals via Toll-like receptor 2, and confers protection against lethal infection

The intracellular protozoan parasite Trypanosoma cruzi is the etiological agent of Chagas disease. We have recently identified a T. cruzi-released protein related to thiol-disulfide oxidoreductase family, called Tc52, which is crucial for parasite survival and virulence. In vitro, Tc52 in combination with IFN-gamma activates human macrophages. In vivo, active immunization with Tc52 relieves the immunosuppression associated to acute infection and elicits a specific immune response. As dendritic cells (DC) have a central role in the initiation of immune responses, we investigated whether Tc52 may modulate DC activity. We show that Tc52 induces human DC maturation. Tc52-treated immature DC acquire CD83 and CD86 expression, produce inflammatory chemokines (IL-8, monocyte chemoattractant protein-1, and macrophage-inflammatory protein-1 alpha), and present potent costimulatory properties. Tc52 binds to DC by a mechanism with the characteristics of a saturable receptor system and signals via Toll-like receptor 2. While Tc52-mediated signaling involves its reduced glutathione-binding site, another portion of the molecule is involved in Tc52 binding to DC. Finally, we report that immunization with Tc52 protects mice in vivo against lethal infection with T. cruzi. Together these data evidence complex molecular interactions between the T. cruzi-derived molecule, Tc52, and DC, and suggest that Tc52 and related class of proteins might represent a new type of pathogen-associated molecular patterns. Moreover, the immune protection data suggest that Tc52 is among candidate molecules that may be used to design an optimal multicomponent vaccine to control T. cruzi infection.     

Trypanosoma cruzi: molecular identification and characterization of new members of the Tc13 family. Description of the interaction between the Tc13 antigen from Tulahuén strain and the second extracellular loop of the beta(1)-adrenergic receptor

Trypanosoma cruzi Tc13 antigens belong to the trans-sialidase superfamily. Their sequences have been described only partially and, up to now, their physiological activity has not been elucidated. Here we present two new members of this family from the Tulahuén strain (Tc13 Tul) and the CL Brener clone (Tc13 CL), being the latter the first Tc13 sequence fully described. Alignment of all Tc13 sequences allowed us to define two sub-families that differ in the number of repeats and the presence or absence of the GPI addition site. Chromoblots demonstrate that Tc13 antigens are mainly located in chromosome III and its homologous. Pull down assays suggest that recombinant MBP-Tc13 Tul interacts with the second extracellular loop of the beta(1)-adrenergic receptor. This is the first evidence that a Tc13 antigen acts as a ligand interacting with a neurotransmitter receptor. These observations might add some light to the development of chagasic pathology.     

Induction of B- and T-cell responses to cruzipain in the murine model of Trypanosoma cruzi infection

Trypanosoma cruzi, the causative agent of Chagas' disease, is an important cause of heart disease in Latin America. The parasite is transmitted mucosally, with both intra- and extracellular life stages in the human host. Cruzipain, the major cysteinyl proteinase of T. cruzi, has been shown to be antigenic in both humans and mice during infection with the parasite. We extend these observations, showing here that multiple murine immune subsets of potential importance for vaccine-induced protection can be induced by cruzipain. Cruzipain-specific serum IgG responses were induced during chronic infection with T. cruzi. In addition, T. cruzi mucosal infection stimulated the development of cruzipain-specific secretory IgA detectable in fecal extracts from infected mice. Cruzipain-specific type 1 cytokine responses characterized by the production of IFN-gamma but not IL-4 were also detectable during murine infection. Furthermore, immunization of mice with a DNA vaccine encoding cruzipain was shown to stimulate cytotoxic T lymphocyte (CTL) responses capable of recognizing and lysing T. cruzi-infected cells. The induction of serum antibody, mucosal IgA, Th1 cytokine and CTL responses by cruzipain in mice supports the use of this parasite protein for further efforts in T. cruzi vaccine development.     

Cutting edge: small molecule CD40 ligand mimetics promote control of parasitemia and enhance T cells producing IFN-gamma during experimental Trypanosoma cruzi infection

Host resistance to Trypanosoma cruzi infection depends on a type 1 response characterized by a strong production of IL-12 and IFN-gamma. Amplifying this response through CD40 triggering results in control of parasitemia. Two newly synthesized molecules (<3 kDa) mimicking trimeric CD40L (mini CD40Ls(-1) and (-2)) bind to CD40, activate murine dendritic cells, and elicit IL-12 production. Wild-type but not CD40 knockout mice exhibited a sharp decrease of parasitemia and mortality when inoculated with T. cruzi mixed with miniCD40Ls. Moreover, the immunosuppression induced by T. cruzi infection was impaired in mice treated with miniCD40Ls, as shown by proliferation of splenic lymphocytes, percentage of CD8(+) T cells, and IFN-gamma production. Mice surviving T. cruzi infection in the presence of miniCD40L(-1) were immunized against a challenge infection. Our results indicate that CD40L mimetics are effective in vivo and promote the control of T. cruzi infection by overcoming the immunosuppression usually induced by the parasites.     

High-avidity, high-IFNγ-producing CD8 T-cell responses following immune selection during HIV-1 infection

HIV-1 mutations, which reduce or abolish CTL responses against virus-infected cells, are frequently selected in acute and chronic HIV infection. Among population HIV-1 sequences, immune selection is evident as human leukocyte antigen (HLA) allele-associated substitutions of amino acids within or near CD8 T-cell epitopes. In these cases, the non-adapted epitope is susceptible to immune recognition until an escape mutation renders the epitope less immunogenic. However, several population-based studies have independently identified HLA-associated viral changes, which lead to the formation of a new T-cell epitope, suggesting that the immune responses that these variants or 'neo-epitopes' elicit provide an evolutionary advantage to the virus rather than the host. Here, we examined the functional characteristics of eight CD8 T-cell responses that result from viral adaptation in 125 HLA-genotyped individuals with chronic HIV-1 infection. Neo-epitopes included well-characterized immunodominant epitopes restricted by common HLA alleles, and in most cases the T-cell responses against the neo-epitope showed significantly greater functional avidity and higher IFNγ production than T cells for non-adapted epitopes, but were not more cytotoxic. Neo-epitope formation and emergence of cognate T-cell response coincident with a rise in viral load was then observed in vivo in an acutely infected individual. These findings show that HIV-1 adaptation not only abrogates the immune recognition of early targeted epitopes, but may also increase immune recognition to other epitopes, which elicit immunodominant but non-protective T-cell responses. These data have implications for immunodominance associated with polyvalent vaccines based on the diversity of chronic HIV-1 sequences.     

CTLA-4 blockage increases resistance to infection with the intracellular protozoan Trypanosoma cruzi

Recent studies have revealed an important role for CTLA-4 as a negative regulator of T cell activation. In the present study, we evaluated the importance of CTLA-4 to the immune response against the intracellular protozoan, Trypanosoma cruzi, the causative agent of Chagas' disease. We observed that the expression of CTLA-4 in spleen cells from naive mice cultured in the presence of live trypomastigote forms of T. cruzi increases over time of exposure. Furthermore, spleen cells harvested from recently infected mice showed a significant increase in the expression of CTLA-4 when compared with spleen cells from noninfected mice. Blockage of CTLA-4 in vitro and/or in vivo did not restore the lymphoproliferative response decreased during the acute phase of infection, but it resulted in a significant increase of NO production in vivo and in vitro. Moreover, the production of IFN-gamma in response to parasite Ags was significantly increased in spleen cells from anti-CTLA-4-treated infected mice when compared with the production found in cells from IgG-treated infected mice. CTLA-4 blockade in vivo also resulted in increased resistance to infection with the Y and Colombian strains of T. cruzi. Taken together these results indicate that CTLA-4 engagement is implicated in the modulation of the immune response against T. cruzi by acting in the mechanisms that control IFN-gamma and NO production during the acute phase of the infection.     

Suppression of cell-mediated immunity in experimental Chagas' disease.

The effect of acute infection with the Tulahuén strain of Trypanosoma cruzi on the cellular immune response in Swiss mice was studied. Mice were immunized with either Freund's complete adjuvant or oxazolone, a skin sensitizing agent, and subsequently skin-tested with either BCG protoplasm or oxazolone to detect delayed hypersensitivity. Depression of the response to these antigens was observed in infected mice during the stage of marked parasitemia. Mice which were responsive to oxazolone before infection lost their ability to respond as the infection progressed. When immunized with live attenuated T. cruzi before infection with virulent organisms, mice developed a greater than normal sensitivity to oxazolone and survived infection. These experiments do not conclude whether immunosuppression due to infection with T. cruzi is directed toward induction or expression of the cell-mediated immune response to the antigens employed.     

DNA immunization with the ribosomal P2beta gene of Trypanosoma cruzi fails to induce pathogenic antibodies

Patients with chronic Chagas' heart disease (cChHD) develop a strong IgG response against the C-terminal region of the Trypanosoma cruzi ribosomal P2beta protein (TcP2beta). These antibodies have been shown to exert an in vitro chronotropic effect on cardiocytes through stimulation of the beta1-adrenergic receptor (beta1-AR). Moreover, the presence of antibodies recognizing the TcP2beta C-terminus was associated with cardiac alterations in mice immunized with the corresponding recombinant protein. Here, we demonstrate that DNA immunization could be used to modulate the specificity of the anti-TcP2beta humoral response in order to avoid the production of pathogenic antibodies. After DNA injection, we detected IgG antibodies that were directed only to internal epitopes of the TcP2beta molecule and that did not exert anti-beta1-AR functional activity, measured as an increase in intracellular cAMP levels of transfected COS-7 cells. Accordingly, DNA-immunized mice did not present electrocardiographic alterations. These data demonstrate that anti-TcP2beta antibodies elicited by DNA immunization are completely different in their specificity and functional activity from those produced during T. cruzi infection.     

Trypanosoma cruzi: influence of predominant bacteria from indigenous digestive microbiota on experimental infection in mice

To verify the influence of some predominant components from indigenous microbiota on systemic immunological responses during experimental Chagas disease, germ-free NIH Swiss mice were mono-associated with Escherichia coli, Enterococcus faecalis, Bacteroides vulgatus or Peptostreptococcus sp. and then infected with the Y strain of Trypanosoma cruzi. All the mono-associations predominantly induced a Th1 type of specific immune response to the infection by T. cruzi. A direct correlation was observed between a higher survival rate and increased IFN-gamma and TNF-alpha production (P<0.05) in E. faecalis-, B. vulgatus-, and Peptostreptococcus-associated mice. Moreover, higher levels of anti-T. cruzi IgG1 and anti-T. cruzi IgG2a were also found in mono-associated animals after infection. On the other hand, with the exception of E. faecalis-associated mice, mono-association induced a lower IL-10 production after infection (P<0.05) when compared with germ-free animals. Interestingly, spleen cell cultures from non-infected germ-free and mono-associated mice spontaneously produced higher levels (P<0.05) of IL-10 than cultures from infected mono-associated mice, except again for E. faecalis-associated animals. In conclusion, the presence of the components of the indigenous microbiota skews the immune response towards production of inflammatory cytokines during experimental infection with T. cruzi in gnotobiotic mice. However, the degree of increase in production of cytokines depends on each bacterial component.