Chronic Toxoplasmosis Research Articles

Protection against LethalNeospora caninumInfection in Mice Induced by Heterologous Vaccination with amic1 mic3KnockoutToxoplasma gondiiStrain

Neospora caninum and Toxoplasma gondii are closely related, obligate intracellular parasites infecting a wide range of vertebrate hosts and causing abortion and neonatal morbidity and mortality. Several lines of evidence suggest that cross immunity between these two pathogens could be exploited in the design of strategies for heterologous vaccination. We assessed the ability of an attenuated strain of T. gondii ("mic1-3KO strain") conferring strong protection against chronic and congenital toxoplasmosis to protect mice against lethal N. caninum infection. Mice immunized with mic1-3KO tachyzoites by the oral and intraperitoneal routes developed a strong cellular Th1 response and displayed significant protection against lethal heterologous N. caninum infection, with survival rates of 70% and 80%, respectively, whereas only 30% of the nonimmunized mice survived. We report here the acquisition of heterologous protective immunity against N. caninum following immunization with a live attenuated mic1-3KO strain of T. gondii.

12/15-Lipoxygenase-Dependent Myeloid Production of Interleukin-12 Is Essential for Resistance to Chronic Toxoplasmosis

Interleukin-12 (IL-12) is critical for resistance to Toxoplasma gondii during both the acute and chronic stages of infection. However, the cellular and molecular pathways that regulate IL-12 production during chronic toxoplasmosis are incompletely defined. We recently discovered that 12/15-lipoxygenase (12/15-LOX), which oxidizes unsaturated lipids in macrophages, is a novel and selective regulator of IL-12 production. We now demonstrate the essential role of this enzyme in the chronic phase of toxoplasmosis. Although 12/15-LOX-deficient mice were resistant to acute T. gondii infection, 80% of 12/15-LOX-deficient mice died during chronic toxoplasmosis, compared to no deaths in wild-type controls. The morbidity of chronically infected 12/15-LOX mice was associated with an increase in brain inflammation and parasite burden. These data suggest that the evolution of the immune response to T. gondii is accompanied by an increasing requirement for 12/15-LOX-mediated signaling. Consistent with this conclusion, 12/15-LOX activity was enhanced during chronic, but not acute, toxoplasmosis. Furthermore, the enhanced susceptibility of 12/15-LOX-deficient mice to chronic toxoplasmosis was associated with reduced production of IL-12 and gamma interferon (IFN-gamma) that was not evident during acute infection. Importantly, ex vivo IFN-gamma production by 12/15-LOX-deficient splenocytes could be rescued by the addition of recombinant IL-12. These data establish that 12/15-LOX is a critical mediator of the chronic type 1 inflammatory response and that immune mediators can be subject to distinct cellular and/or molecular mechanisms of regulation at different stages of inflammation.

Toxoplasma gondii: P30 peptides recognition pattern in human toxoplasmosis

In this study, human sera reactivity against nine peptides derived from the Toxoplasma gondii P30 protein was assessed by ELISA in patients with different clinical forms of toxoplasmosis. Same as has been reported in mice, sera from congenital, ocular and chronic asymptomatic toxoplasmosis patients recognized more strongly peptides from the protein’s carboxy-terminus, being peptide 2017 (amino acids 301–320) the one most strongly recognized by sera from patients with ocular toxoplasmosis. Serum samples collected from 13 patients without ocular infection, 13 with inactive chorioretinal scars, 6 with active ocular infection and 10 seronegative individuals were then screened for anti-2017 IgG. Peptide 2017 was recognized by all patients’ samples but not by sera from T. gondii-seronegative individuals. No statistically significant differences were found between the absorbance levels of groups with and without lesions or with active or inactive ocular lesions, as determined by ANOVA.

Haematological changes of cats with Toxoplasma gondii-specific antibodies

The purpose of this study is to access the usefulness of haematological parameters in diagnosing of chronic toxoplasmosis in cats. Antibodies of Toxoplasma gondii were detected in 50 cats using an enzyme-linked immunosorbent assay. Immunoglobulin M (IgM) and immunoglobulin G (IgG) were measured with the cutoff point for T. gondii considered to be 1:64. Fifteen (30%) of the cats were found to be seropositive. No statistically significant difference was found amongst different age groups, but the male cats showed significantly higher antibody titres than female cats. A significant positive correlation was found between IgM and IgG of the cats. Comparison of haematological data between two groups of the cats (IgM <1/64, n = 35; IgM ≥1/64, n = 15) showed that packed cell volume (PCV), red blood cell (RBC) and monocyte values in cats with higher IgM titres were significantly higher than cats with lower IgM. It is suggested that in cases with higher values of PCV, RBC and monocyte in routine complete blood count profile of apparently normal cats, toxoplasmosis should be considered. Further studies with more cases are needed to ensure the diagnostic values of these measured parameters.

Value of 2 IgG avidity commercial tests used alone or in association to date toxoplasmosis contamination

Toxoplasmosis acquired during pregnancy exposes the fetus to congenital toxoplasmosis. Avidity tests are commonly used to date time of infection to evaluate the fetal risk and to offer prenatal diagnosis. This study evaluated and compared 2 commercial avidity tests: Platelia™ Toxo IgG Avidity (Bio-Rad, Marnes la Coquette, France) and Liaison® Toxo IgG Avidity II (Diasorin, Saluggia, Italy) kits. In complement, a study of specific IgG and IgM in the 2 systems was carried out. Sensitivity and specificity of the avidity tests were 94.4% and 87.8% for the Liaison® and 91.3% and 98.5% for the Platelia™ methods, respectively. Percentages of complete discrepancies, partial discrepancies, and agreement between both tests were 1.1%, 23.6%, and 75.3%, respectively. Moreover, the combination of both avidity tests may be useful in some cases. Indeed, that strategy permitted to confirm without delay a chronic toxoplasmosis in 23 cases and avoid treatment in these pregnant women.

Vaccination contre la toxoplasmose chez les animaux de rente

Toxoplasmosis is a worldwide zoonotic disease caused by the protozoa Toxoplasma gondii. It is transmitted to man by the ingestion of contaminated and undercooked meat. In sheep and goats, toxoplasmosis causes numerous abortions. A thorough analysis of the seroprevalence of toxoplasmosis in different animal species will help consumer information and thus limit the risks of transmission. The vaccination of farm animals may help reduce the transmission to man, as well as prevent abortion in ewes. A naturally attenuated live T. gondii vaccine is available for the prevention of abortions in ewes, but its virulence is not fully controlled, and there is a risk of reversion to a pathogenic strain. Molecular biology techniques have lead to the development of attenuated strains through the deletion of targeted genes, which are unlikely to revert to their initial virulence. A strain called Mic1-3KO was shown to be effective against congenital and chronic toxoplasmosis in mice. It is also effective against congenital toxoplasmosis in ewes. This vaccine approach remains promising.

Protective Th1 immune responses against chronic toxoplasmosis induced by a protein–protein vaccine combination but not by its DNA–protein counterpart

Vaccine-induced protection against toxoplasmosis is correlated with cellular immune responses to Toxoplasma gondii, both in animals and man. The goal of the current study was to evaluate whether the combination of a recombinant protein and a plasmid DNA vaccine could offer an advantage over the protein mixture, and protect outbred mice against infection with T. gondii. To this purpose, the chimeric protein rEC2, encoding antigenic fragments of surface-associated proteins MIC2, MIC3 and SAG1, was combined with pGRA7 plasmid DNA or rGRA7 protein. High levels of antibodies were elicited by both vaccine formulations. The protein–DNA vaccine elicited a polarized Th1/Th2 immune response, characterized by IFN-γ and IL-10, and afforded low protection (24%) against brain cyst formation. In contrast, the protein–protein vaccine elicited a Th1-focused immune response, characterized by IFN-γ and IL-2 production, conferring a strong protection (79%) against brain cyst formation in chronic toxoplasmosis. We show here that GERBU adjuvanted protein vaccines confer better protection against toxoplasmosis than the protein–DNA heterologous vaccine.

Use of GRA6-Derived Synthetic Polymorphic Peptides in an Immunoenzymatic Assay To Serotype Toxoplasma gondii in Human Serum Samples Collected from Three Continents

Serotyping is a simple typing method that consists of an immunoenzymatic assay (enzyme-linked immunosorbent assay [ELISA]) using synthetic polymorphic peptides derived from Toxoplasma gondii antigens. We developed a new ELISA based on GRA6 C-terminal polymorphic peptides. Serum samples from 41 human infections due to 23 archetypal (type I, II, or III) and 18 nonarchetypal strains were selected in order to validate this approach. For 20 out of the 23 archetypal infections, there was a clear correlation between microsatellite genotype and GRA6 serotyping. All infections due to nonarchetypal strains were misclassified as archetypal strain infections. The GRA6 C-terminal peptides from these strains were analyzed to explain this misclassification. A second group of 455 patients with acute and chronic toxoplasmosis due to unknown genotypes from different European, African, and Latin American countries were included in this study, and the strain type predicted by this method. The results suggest that serotyping is a promising method for typing strains, although limitations exist for African and South American strains as a consequence of higher peptide polymorphism. Other peptides from different markers must be studied in order to discriminate archetypal from nonarchetypal strains.

Use of the serum reactivity against Toxoplasma gondii excreted–secreted antigens in cerebral toxoplasmosis diagnosis in human immunodeficiency virus-infected patients

Despite the development of serological and molecular methods in recent years, the diagnosis of cerebral toxoplasmosis in human immunodeficiency virus (HIV)-infected patients still presents difficulties. In the present study, we investigated whether cerebral toxoplasmosis induced changes in the reactivity of serum toward Toxoplasma gondii excreted-secreted antigens (ESA) in order to develop an assay for evaluating HIV-infected patients with cerebral toxoplasmosis. The antigen selection was based on those produced by tachyzoites, since it is the form of the organism responsible for disseminating the infection, as well as stimulation of the humoral and cellular immune responses. By using an ELISA containing pooled ESA recovered from infected culture supernatants with tachyzoites-RH strain (ESA-ELISA), we found that ESA had a high specificity for sera from patients with cerebral toxoplasmosis. The reactions were compared with an ELISA using crude tachyzoites antigen, widely used in traditional serology. The assays were performed on 293 serum samples separated as follows: 100 sera from patients with cerebral toxoplasmosis and AIDS (symptomatic), 99 sera from individuals with chronic toxoplasmosis (asymptomatic) and 94 sera from healthy individuals without toxoplasmosis (control). The crude tachyzoite antigen in ELISA was able to distinguish both groups of sera with toxoplasmosis, as similar reactivity were observed in sera from patients with cerebral toxoplasmosis and those from chronic individuals. In contrast, ESA-ELISA distinguished sera from symptomatic and asymptomatic individuals (three times more reactive in the former group, 12.6 versus 4.2). The assays were reproducible based on immunoblotting and statistical analysis. These data suggest the utility of ESA-ELISA in the diagnosis of cerebral toxoplasmosis in HIV-infected patients, since it provided clear evidence that anti-ESA antibodies are present principally in patients with active infection. The absence of a significant amount of antibodies distinguished the patients without clinical symptoms of infection.

Anti-P30 IgA antibodies as prenatal markers of congenital toxoplasma infection

This study extends a previous study and confirms that the detection of anti-P30 IgA antibodies is very helpful in the diagnosis of acute acquired or congenital toxoplasmosis. Moreover, we demonstrate that an anti-P30 IgA response can be mounted in the fetuses infected by Toxoplasma gondii during their intra-uterine life as early as week 23 of gestation. A double-sandwich ELISA described in our previous work was used to detect anti-P30 IgA antibodies in 1378 human serum samples collected from 551 patients, including 162 fetuses whose mothers had been infected by T. gondii during pregnancy, 46 congenitally infected and 90 uninfected newborns and 253 women suspected of having been infected during pregnancy, including the mothers of fetuses and newborns previously described. Anti-P30 IgA antibodies were detected in all cases of acute toxoplasmosis but in no case of chronic toxoplasmosis: in the majority of cases, the IgA antibody titre fell below cut-off in 3-9 months. Among the 46 congenitally infected newborns, anti-P30 IgA antibodies were detected in sera of 41 infected newborns (38 at birth, two in the first months of life, one in the seventh month of life), while anti-P30 IgM antibodies were detected in only 30 cases at birth and in one case during the first month of life. Among 162 fetuses, anti-P30 IgA response was observed in five infected fetuses, but was not detected in either 152 uninfected fetuses or in five fetuses considered as infected. The absence or presence of anti-P30 IgA antibodies in the fetus is discussed in relation to the date of maternal infection and collection of the fetal blood. It clearly appears from our study that the combined testing of both IgM and IgA in the fetus and the newborn is essential for a more efficient diagnosis of infection.

Reverse Enzyme-Linked Immunosorbent Assay Using Monoclonal Antibodies against SAG1-Related Sequence, SAG2A, and p97 Antigens fromToxoplasma gondiiTo Detect Specific Immunoglobulin G (IgG), IgM, and IgA Antibodies in Human Sera

The present study aimed to evaluate the performance of three monoclonal antibodies (MAbs) in reverse enzyme-linked immunosorbent assays (ELISAs) for detecting immunoglobulin G (IgG), IgM, and IgA antibodies against Toxoplasma gondii in 175 serum samples from patients at different stages of T. gondii infection, as defined by both serological and clinical criteria, as follows: recent (n = 45), transient (n = 40), and chronic (n = 55) infection as well as seronegative subjects (n = 35). The results were compared with those obtained by indirect ELISA using soluble Toxoplasma total antigen (STAg). Our data demonstrated that MAb A3A4 recognizes a conformational epitope in SAG1-related-sequence (SRS) antigens, while A4D12 and 1B8 recognize linear epitopes defined as SAG2A surface antigen and p97 cytoplasmatic antigen, respectively. Reverse ELISA for IgG with A3A4 or A4D12 MAbs was highly correlated with indirect ELISA for anti-STAg IgG, whereas only A4D12 reverse ELISA showed high correlation with indirect ELISA for IgM and IgA isotypes. To our knowledge, this is the first report analyzing the performance of a reverse ELISA for simultaneous detection of IgG, IgM, and IgA isotypes active toward native SAG2A, SRS, and p97 molecules from STAg, using a panel of human sera from patients with recent and chronic toxoplasmosis. Thus, reverse ELISA based on the capture of native SAG2A and SRS antigens of STAg by MAbs could be an additional approach for strengthening the helpfulness of serological tests assessing the stage of infection, particularly in combination with highly sensitive and specific assays that are frequently used nowadays for diagnosis of toxoplasmosis during pregnancy or congenital infection in newborns.

Toxoplasma gondii: Serological characterization and immunogenicity of recombinant surface antigen 2 (SAG2) expressed in the yeast Pichia pastoris

The full length surface antigen 2 (SAG2) gene of the protozoan parasite Toxoplasma gondii was cloned and intracellularly expressed in the Pichia pastoris expression system. The molecular weight of the expressed recombinant SAG2 (36 kDa) was much larger than the native SAG2 (22 kDa). This discrepancy in size was due to hyperglycosylation, as deglycosylation assay reduced the size of the recombinant SAG2 to 22 kDa. Despite being hyperglycosylated, the recombinant SAG2 reacted strongly with pooled anti- Toxoplasma human serum, pooled anti- Toxoplasma mouse serum and a SAG2-specific monoclonal antibody. The glycosylated recombinant SAG2 was further evaluated in Western blot and in-house enzyme-linked immunosorbent assay (ELISA) using 80 human serum samples, including confirmed early acute (IgM positive, IgG negative; n = 20), acute (IgM positive, IgG positive; n = 20) and chronic (IgM negative, IgG positive; n = 20) toxoplasmosis patients, and toxoplasmosis negative control patients ( n = 20). Results of the Western blot showed that the recombinant SAG2 reacted with all 60 samples of the toxoplasmosis cases but not with the Toxoplasma-negative samples. The sensitivity of in-house ELISA was 80%, 95% and 100% for early acute, acute and chronic patients’ serum samples, respectively. Vaccination study showed that serum from mice immunised with the glycosylated recombinant SAG2 reacted specifically with the native SAG2 of T. gondii. The mice were significantly protected against lethal challenge with live T. gondii RH strain tachyzoites ( P < 0.01) and their survival time was increased compared to controls. Therefore, the present study shows that the P. pastoris-derived recombinant SAG2 was specific and suitable for use as antigen for detecting anti- Toxoplasma IgG and IgM antibodies. The vaccination study showed that recombinant SAG2 protein was immunoprotective in mice against lethal challenge.

Malondialdehyde, Glutathione, and Nitric Oxide Levels in Toxoplasma gondii Seropositive Patients

The aim of this study was to investigate the difference in the serum malondialdehyde (MDA), glutathione (GSH), and nitric oxide (NO) levels between normal and T. gondii-infected patients. To this end, MDA, GSH, and NO levels in the sera of 37 seropositive patients and 40 participants in the control group were evaluated. In Toxoplasma ELISA, IgG results of the patient group were 1,013.0 +/- 543.8 in optical density (mean +/- SD). A statistically significant difference was found between patients and the control group in terms of MDA, GSH, and NO levels. A decrease in GSH activity was detected, while MDA and NO levels increased significantly. Consequently, it is suggested that the use of antioxidant vitamins in addition to a parasite treatment shall prove useful. The high infection vs control ratio of MDA and NO levels probably suggests the occurrence as a mechanism of tissue damage in cases of chronic toxoplasmosis. Moreover, it is recommended that the patient levels of MDA, GSH, and NO should be evaluated in toxoplasmosis.

Decreased infarct size after focal cerebral ischemia in mice chronically infected with Toxoplasma gondii

To determine whether Toxoplasma gondii infection could modify biological phenomena associated with brain ischemia, we investigated the effect of permanent middle cerebral artery occlusion (MCAO) on neuronal survival, inflammation and redox state in chronically infected mice. Infected animals showed a 40% to 50% decrease of infarct size compared with non-infected littermates 1, 4 and 14 days after MCAO. The resistance of infected mice may be associated with increased basal levels of anti-inflammatory cytokines and/or a marked reduction of the MCAO-related brain induction of two pro-inflammatory cytokines, tumor necrosis factor-alpha and interferon-gamma (IFNγ). In addition, potential anti-inflammatory/neuroprotective factors such as nerve growth factor, suppressor of cytokine signaling-3, superoxide dismutase activity, uncoupling protein-2 and glutathione (GSH) were upregulated in the brain of infected mice. Consistent with a role of GSH in central cytokine regulation, GSH depletion by diethyl maleate inhibited Toxoplasma gondii lesion resistance by increasing the proinflammatory cytokine IFNγ brain levels. Overall, these findings indicate that chronic toxoplasmosis decisively influences both the inflammatory molecular events and outcome of cerebral ischemia.

Comparison of certain blood components and immunoglobulin in women who are infected and not infected with the disease of constipation

A comparative study was carried out about toxoplasmosis in Neinava governorate and it was revealed the effect of toxoplasmosis on some blood components represented by haemoglobin level and the number of leucocytes and blood platelets. Heamoglobin level decreased in women with acute and chronic toxoplasmosis (1 0.2 g/1 00 mg± 1.4 and 10.5 g/1 00 ml+ 1.9) respectively, while the level remain normal in women with old infection(l3.1g /100 ±0.8) compared with uninfected woman. A slight decrease in leucocytes count in chronic infection( 9.6 X 10 L± 3.11 and 9.2 X 10. cell/L 2.7) was observed respectively, while in woman with old infection no noticeable change occurred in leucocytes count(8.9 X10 cell/L ± 5.4) compared with uninfected women. As concern plateletscount neither increase nor decrease was observed( 260X 10 ceel/±5.4) in case of acute toxoplasmosis and (258X 10 pl/L ±4.1) in case with chronic toxoplasmosis while in old infection(264 X 10 pl/3 .26) compared withuninfected. Estimation of immunoglobulin level was carried out for IgG, IgM, IgA in serum of infected females(acute, chronic and old ) compared with uninfected. It was revealed from the results that IgA level (390.2 mg/1 00 ml±43.3) and less than that in old infections(300.2 mg/100 ml± 41.2) while IgM increased in case of acute infection(30 1.13 mg/1 00 ml± 36.1) and less in old infections(227 mg/ 100 ml±18.5) while IgG has increased to reach 1925.3 mg/ 100 ml±l9.2) in case of chronic infections and less than that in acute infections(l731. 7 mg/1 00 ml± 16. 7) compared with uninfected.It is revealed also that the number of positive cases of hyper autoimmunoglobulins was ( 4) cases out of (30) cases ( 13.3%)in case of aborted females infected with acute toxoplasmosis whom did not responds to treatment while no positive case was detected in chronic toxoplasmosis . From the results of statistical analysis using chi-square no significant difference or correlation between infection and hyperautoimmunoglobulins was detected while after using Z-test for comparing percentages a significant relationship between the type of infection with toxoplasmosis (both acute and chronic) with hyperautoimmunoglobulins in aborted women whom they did not respond to treatment properly, was found.

Meta-analysis of three case controlled studies and an ecological study into the link between cryptogenic epilepsy and chronic toxoplasmosis infection

A meta-analysis was performed on three case controlled studies which examined the relationship between latent toxoplasmosis gondii infection in the immunocompetent host and cryptogenic epilepsy. Further comparison was also made by examining the seroprevalence of toxoplasmosis rates for 17 various countries, cities or regions against the prevalence rates for epilepsy in those regions. The results for the meta-analysis showed a log-odds ratio of 4.8 which approximates to a similar relative risk, (CI 2.6 to 7.8), with CI for all three studies being above 1. Seroprevalence rates for toxoplasmosis and prevalence rates of epilepsy showed a strong association (p<0.001). The prevalence of toxoplasmosis is an important factor in the prevalence of epilepsy with a probable link in the cryptogenic epilepsies. An area with a reduced burden of toxoplasmosis will also have a reduced burden of epilepsy. Neuropathophysiology findings from various studies show a common physical relationship of microglial nodule formation in Toxoplasma gondii infection and epilepsy. This analysis raises the possibility that one of the many causes of epilepsy may be an infectious agent, or that cryptogenic epilepsy may be a consequence of latent toxoplasmosis infection. This raises the possibility that public health measures to reduce toxoplasmosis infection may also result in a reduction in epilepsy.

BALB/c mice resistant to Toxoplasma gondii infection proved to be highly susceptible when previously infected with Myocoptes musculinus fur mites

The immune response induced by Toxoplasma gondii is characterized by Th1 immune mechanisms. We previously demonstrated that C57BL/6 mice infested with Myocoptes musculinus and infected with T. gondii by intraperitoneal route undergo accelerated mortality according to Th2 immune mechanisms induced by the acarian. To evaluate whether infection with M. musculinus influences T. gondii-induced Th1 response in a resistant mouse lineage, BALB/c, which develops latent chronic toxoplasmosis in a way similar to that observed in immunocompetent humans, this study was done. The animals were infected with T. gondii ME-49 strain 1 month after M. musculinus infestation, being the survival and the immune response monitored. The double-infected displayed higher mortality rate if compared with the mono-infected mice. In addition, infection with M. musculinus changed the T. gondii-specific immune response, converting BALB/c host to a susceptible phenotype. Spleen cells had increased the levels of IL-4 in double-infected mice. This alteration was associated with severe pneumonia, encephalitis and wasting condition. In addition, a higher tissue parasitism was observed in double-infected animals. It can be concluded that infection with these two contrasting parasites, M. musculinus and T. gondii, may convert an immunocompetent host into a susceptible one, and such a host will develop severe toxoplasmosis.

IgG avidity assay firms up the diagnosis of acute toxoplasmosis on the first serum sample in immunocompetent pregnant women

Rapid diagnosis of acute toxoplasmosis during pregnancy permits timely treatment and prevents or attenuates congenital toxoplasmosis. Specific IgM antibodies to Toxoplasma as marker of acute infection are often poorly informative, meaning that a complementary technique is needed to reach a diagnosis on the first sample. Here we evaluated 2 commercial kits designed to assist with the diagnosis of acute toxoplasmosis: Platelia® Toxo IgG Avidity Complementary Reagents and Platelia® Toxo IgA, both from BIO-RAD (Marnes La Coquette, France). We tested 2 groups of subjects: 36 patients with acute toxoplasmosis and 55 patients with chronic toxoplasmosis. The IgG avidity test had a sensitivity of 100% (36/36), a specificity of 92.7% (51/55), a positive predictive value of 90%, and a negative predictive value of 100%. Among the immunocompetent women population, the avidity test had perfect sensitivity and specificity, and positive and negative predictive values of 100%. The IgA test had a sensitivity of 88.8% (32/36) and a specificity of 85.4% (47/55), and positive and negative predictive values of 80% and 92.1%, respectively. When the 2 tests were combined, there was only 1 case in which the diagnosis of chronic toxoplasmosis could not be confirmed. The IgG avidity test can therefore be used to rapidly distinguish between chronic and acute infection on the first sample from a pregnant woman, provided there is no underlying immunodepression and no ongoing antitoxoplasmic treatment. In these 2 situations, the results must be interpreted with care, and other serologic markers, including IgA, should be tested. Determination of a pregnant woman's status on a first serum sample allows therapeutic and preventive management to be started without delay.

Toxoplasma gondii: DNA vaccination with genes encoding antigens MIC2, M2AP, AMA1 and BAG1 and evaluation of their immunogenic potential

A combination of antigenic regions of microneme proteins have been previously reported as being protective against chronic toxoplasmosis. In this work, we evaluated immune responses induced by immunizing BALB/c and C57BL/6 mice intradermally with plasmid DNA encoding the protein sequences of Toxoplasma gondii AMA1, MIC2, M2AP and BAG1. Mice immunized with the AMA1 gene developed high levels of serum IgG2a and c antibodies as well as cellular immune responses associated with IFN-γ synthesis suggesting a modulated Th1 type of response. Immunization with the AMA1 gene resulted in a partial but significant protection against the acute phase of toxoplasmosis compared to MIC2, M2AP and BAG1 genes. Therefore, the AMA1 gene appears to generate a strong specific immune response and also provides effective protection against toxoplasmosis more than the MIC2, M2AP and BAG1 genes.

Organ- and Disease-Stage-Specific Regulation ofToxoplasma gondii-Specific CD8-T-Cell Responses by CD4 T Cells

Toxoplasma gondii induces a persistent central nervous system infection, which may be lethally reactivated in AIDS patients with low CD4 T-cell numbers. To analyze the role of CD4 T cells for the regulation of parasite-specific CD8 T cells, mice were infected with transgenic T. gondii expressing the CD8 T-cell antigen beta-galactosidase (beta-Gal). Depletion of CD4 T cells prior to infection did not affect frequencies of beta-Gal(876-884)-specific (consisting of residues 876 to 884 of beta-Gal) CD8 T cells but resulted in a pronounced reduction of intracerebral beta-Gal-specific gamma interferon (IFN-gamma)-producing and cytolytic CD8 T cells. After cessation of anti-CD4 treatment a normal T. gondii-specific CD4 T-cell response developed, but IFN-gamma production of intracerebral beta-Gal-specific CD8 T cells remained impaired. The important supportive role of CD4 T cells for the optimal functional activity of intracerebral CD8 T cells was also observed in mice that had been depleted of CD4 T cells during chronic toxoplasmosis. Reinfection of chronically infected mice that had been depleted of CD4 T cells during either the acute or chronic stage of infection resulted in an enhanced proliferation of beta-Gal-specific IFN-gamma-producing splenic CD8 T cells. However, reinfection of chronically infected mice that had been depleted of CD4 T cells in the acute stage of infection did not reverse the impaired IFN-gamma production of intracerebral CD8 T cells. Collectively, these findings illustrate that CD4 T cells are not required for the induction and maintenance of parasite-specific CD8 T cells but, depending on the stage of infection, the infected organ and parasite challenge infection regulate the functional activity of intracerebral CD8 T cells.