Quantitative Risk Assessment of Oocyst Versus Bradyzoite Foodborne Transmission of Toxoplasma gondii in Brazil.

Toxoplasma gondii is a globally distributed zoonotic protozoan parasite. Infection with T. gondii can cause congenital toxoplasmosis in developing fetuses and acute outbreaks in the general population, and the disease burden is especially high in South America. Prior studies found that the environmental stage of T. gondii, oocysts, is an important source of infection in Brazil; however, no studies have quantified this risk relative to other parasite stages. We developed a Bayesian quantitative risk assessment (QRA) to estimate the relative attribution of the two primary parasite stages (bradyzoite and oocyst) that can be transmitted in foods to people in Brazil. Oocyst contamination in fruits and greens contributed significantly more to overall estimated T. gondii infections than bradyzoite-contaminated foods (beef, pork, poultry). In sensitivity analysis, treatment, i.e., cooking temperature for meat and washing efficiency for produce, most strongly affected the estimated toxoplasmosis incidence rate. Due to the lack of regional food contamination prevalence data and the high level of uncertainty in many model parameters, this analysis provides an initial estimate of the relative importance of food products. Important knowledge gaps for oocyst-borne infections were identified and can drive future studies to improve risk assessments and effective policy actions to reduce human toxoplasmosis in Brazil.

Effects of Toxoplasma gondii infection on cognition, symptoms, and response to digital cognitive training in schizophrenia.

Studies indicate that neuroscience-informed digital cognitive training can remediate cognitive impairments in schizophrenia, but the factors contributing to these deficits and response to treatment remain unclear. Toxoplasma gondii is a neuroinvasive parasite linked to cognitive decline that also presents a higher prevalence in schizophrenia. Here, we compared the cognition and symptom severity of IgG seropositive (TOXO+; n = 25) and seronegative (TOXO-; n = 35) patients who participated in a randomized controlled trial of digital cognitive training. At baseline, TOXO+ subjects presented lower global cognition than TOXO- (F = 3.78, p = 0.05). Specifically, TOXO+ subjects showed worse verbal memory and learning (F = 4.48, p = 0.03), social cognition (F = 5.71, p = 0.02), and higher antibody concentrations were associated with increased negative (r = 0.42, p = 0.04) and total (r = 0.40, p = 0.04) schizophrenia symptoms. After training, the TOXO+ group showed higher adherence to the intervention (X2 = 9.31, p = 0.03), but there were no differences in changes in cognition and symptoms between groups. These findings highlight the association between seropositivity to T. gondii and deteriorated cognition and symptoms in schizophrenia. Further research is needed to assess the specific efficacy of digital cognitive training on this population.

Microcefalia por toxoplasmose congênita em tempos de epidemia por Zika vírus no Brasil / Microcephaly due to congenital toxoplasmosis in times of Zika virus epidemic in Brazil

Descreve-se um caso de toxoplasmose congênita com microcefalia, ocorrido durante a epidemia de Zika vírus no Brasil e somente diagnosticado como toxoplasmose aos sete meses de idade. Este caso ilustra a pertinência e urgência para a implementação de políticas públicas específicas para prevenção, diagnóstico e tratamento para a toxoplasmose adquirida durante a gestação, e demonstra que quando o assunto é microcefalia por infecções congênitas no Brasil, precisamos estar atentos a outras possibilidades além da infecção por Zika vírus; em especial à toxoplasmose congênita, que é altamente prevalente em nosso país, e se for diagnosticada e tratada no devido tempo, danos neurológicos e oculares irreversíveis poderão ser evitados. Apoiamos integralmente a portaria do Ministério da Saúde que torna obrigatória a notificação de casos de toxoplasmose gestacional e congênita no Brasil. As ações que venham a ser implementadas deverão resultar em um programa nacional específico para toxoplasmose gestacional e congênita, que possa beneficiar crianças em todo o país por meio de medidas de educação preventiva em saúde, da triagem e do tratamento para gestantes e recém nascidos.

A case of congenital toxoplasmosis with microcephaly, occurred during the epidemic of Zika virus in Brazil, and only diagnosed as toxoplasmosis at seven months of age, is described. This case illustrates the pertinence and urgency for the implementation of specific public policies for prevention, diagnosis and treatment for toxoplasmosis acquired during pregnancy, and shows that when the subject is microcephaly due to congenital infections in Brazil, one must be attentive to other possibilities besides Zika virus infection; in particular to congenital toxoplasmosis, which is highly prevalent in our country, and if diagnosed and treated in due course, irreversible neurological and ocular damage may be avoided. We fully support the Ministry of Health ordinance that makes it compulsory to notify cases of gestational and congenital toxoplasmosis in Brazil. The actions that will be implemented should result in a specific national program for gestational and congenital toxoplasmosis, which will benefit children throughout the country through preventive health education, screening and treatment for pregnant women and newborns.

IgA and IgG1 reactivities assessed by flow cytometry mirror clinical aspects of infants with ocular congenital toxoplasmosis.

This study intended to apply the flow cytometric analysis of IgA and IgG reactivity and intracytoplasmic cytokine analysis to understand and decode the clinical aspects of infants with ocular congenital toxoplasmosis. The Toxoplasma gondii-infected infants (TOXO) were subdivided according to their clinical aspects based on the absence (NRL), presence of active (ARL), active/cicatricial (ACRL) or cicatricial retinochoroidal lesions (CRL) and compared to non-infected controls (NI). The reactivity of anti-T. gondii IgG subclasses resembles the clinical aspects of ocular lesions. IgG and IgG1 discriminate infants with cicatricial lesions (ACRL and CRL) from both ARL and NLR. IgG2 and IgG3 are particularly higher in ACRL and CRL as compared to NLR. No differences were observed when IgG4 reactivity was evaluated. Thus, the results indicated that the reactivity patterns of IgA, IgG and IgG subclasses are able to discriminate ARL, ACRL and CRL from NLR or NI. IgA and IgG subclasses are relevant serological biomarkers with diagnostic and prognostic applicability, respectively. Moreover, IgA and IgG1 were closely related to cytokine production by innate/adaptive immunity cells. IgA reactivity was directly associated to TNF-α-derived from neutrophils, monocytes and CD8(+) T-cells, while IgG1 was inversely correlated with IFN-γ-producing CD4(+) and CD8(+) T-cells but positively correlated with IL-10(+) B-cells. These findings provide insights on the relationship between the cytokine production by innate/adaptive immunity and the antibody pattern of infants with ocular congenital toxoplasmosis. In addition, the present study supports the use of flow cytometric serology as a potential tool for the diagnosis and monitoring of ocular lesions in T. gondii-infected infants in the clinical setting.

Waterborne toxoplasmosis investigated and analysed under hydrogeological assessment: new data and perspectives for further research.

We present a set of data on human and chicken Toxoplasma gondii seroprevalence that was investigated and analysed in light of groundwater vulnerability information in an area endemic for waterborne toxoplasmosis in Brazil. Hydrogeological assessment was undertaken to select sites for water collection from wells for T. gondii oocyst testing and for collecting blood from free-range chickens and humans for anti-T. gondii serologic testing. Serologic testing of human specimens was done using conventional commercial tests and a sporozoite-specific embryogenesis-related protein (TgERP), which is able to differentiate whether infection resulted from tissue cysts or oocysts. Water specimens were negative for the presence of viable T. gondii oocysts. However, seroprevalence in free-range chickens was significantly associated with vulnerability of groundwater to surface contamination (p < 0.0001; odds ratio: 4.73, 95% confidence interval: 2.18-10.2). Surprisingly, a high prevalence of antibodies against TgERP was detected in human specimens, suggesting the possibility of a continuous contamination of drinking water with T. gondii oocysts in this endemic setting. These findings and the new proposed approach to investigate and analyse endemic toxoplasmosis in light of groundwater vulnerability information associated with prevalence in humans estimated by oocyst antigens recognition have implications for the potential role of hydrogeological assessment in researching waterborne toxoplasmosis at a global scale.

Waterborne toxoplasmosis investigated and analysed under hydrogeological assessment: new data and perspectives for further research

We present a set of data on human and chicken Toxoplasma gondiiseroprevalence that was investigated and analysed in light of groundwater vulnerability information in an area endemic for waterborne toxoplasmosis in Brazil. Hydrogeological assessment was undertaken to select sites for water collection from wells for T. gondiioocyst testing and for collecting blood from free-range chickens and humans for anti-T. gondiiserologic testing. Serologic testing of human specimens was done using conventional commercial tests and a sporozoite-specific embryogenesis-related protein (TgERP), which is able to differentiate whether infection resulted from tissue cysts or oocysts. Water specimens were negative for the presence of viable T. gondiioocysts. However, seroprevalence in free-range chickens was significantly associated with vulnerability of groundwater to surface contamination (p < 0.0001; odds ratio: 4.73, 95% confidence interval: 2.18-10.2). Surprisingly, a high prevalence of antibodies against TgERP was detected in human specimens, suggesting the possibility of a continuous contamination of drinking water with T. gondiioocysts in this endemic setting. These findings and the new proposed approach to investigate and analyse endemic toxoplasmosis in light of groundwater vulnerability information associated with prevalence in humans estimated by oocyst antigens recognition have implications for the potential role of hydrogeological assessment in researching waterborne toxoplasmosis at a global scale.

Specific IgM, IgG and IgG1 directed against Toxoplasma gondii detected by flow cytometry and their potential as serologic tools to support clinical indirect fundoscopic presumed diagnosis of ocular disease.

In the present study we evaluated the anti-Toxoplasma gondii immunoglobulin profiles of a group of 118 individuals living in an endemic area. The aim of the study was to select biomarkers to support the ophthalmological diagnosis of retinal/retinochoroidal scars presumably caused by T. gondii infection. Overall anti-T. gondii reactivity of the IgM, IgG, IgA, IgE and IgG subclasses was investigated by flow cytometry-based anti-fixed tachyzoite antibodies (FC-AFTA) in four groups of subjects, referred to as: i) TOXO(L)--seropositive patients with retinal/retinochoroidal scars presumably caused by T. gondii infection; these patients were further subdivided according to morphological aspects of their ocular scar lesions as A, B or C; ii) TOXO(NL)--seropositive patients without ocular scar lesions; iii) NEG(L)--T. gondii seronegative patients presenting retinal lesions; and iv) NEG(NL)--T. gondii seronegative without retinal lesions (negative controls). Our data demonstrated that anti-T. gondii IgG profiles were able to discriminate the mean reactivity of TOXO(L) from all other clinical groups. Analysis of anti-T. gondii immunoglobulin profiles revealed that IgM and IgG were good biomarkers capable of discriminating between individual reactivity in patients with retinal/retinochoroidal scars presumably caused by T. gondii infection [TOXO(L)] from those caused by other clinical conditions. Furthermore, anti-T. gondii IgG1 reactivity was able to discriminate TOXO(L) from all other clinical groups. In conclusion, the pre-selected IgM, IgG and IgG1 anti-T. gondii antibody subclasses were able to segregate both TOXO(L) and the other subgroups, including the scar lesion group types (A, B, C), from other clinical conditions. These results suggest the applicability of this technique in the clinical laboratory to detect putative biomarker for diagnosis of ocular lesions in T. gondii-infected patients. Studies in other areas implementing the methods described in the present study would be of value and enable evaluation of a system for classification of presumed ocular toxoplasmosis scar lesions. This classification would make comparative studies on ocular toxoplasmosis conducted in different regions around the world possible.

Single nucleotide polymorphisms in the interferon gamma gene are associated with distinct types of retinochoroidal scar lesions presumably caused by Toxoplasma gondii infection.

The association of single nucleotide polymorphisms (SNPs) in the interferon (IFN)-γ gene ( IFNG ) with different types of retinal scar lesions presumably caused by toxoplasmosis were investigated in a cross-sectional population-based genetic study. Ten SNPs were investigated and after Bonferroni correction, only the associations between SNPs rs2069718 and rs3181035 with retinal/retinochoroidal scar lesions type A (most severe scar lesions) and C (least severe scar lesions), respectively, remained significant. The associations of two different IFNG SNPs with two different types of retinal lesions attributable to toxoplasmosis support the hypothesis that different inflammatory mechanisms underlie the development of these lesions. The in vitro analysis of IFN-γ secretion by peripheral blood mononuclear cells stimulated with Toxoplasma gondii antigens was also investigated. The association between SNP rs2069718 and type A scar lesions revealed that differential IFN-γ levels are correlated with distinct genotypes. However, no correlation was observed with IFN-γ secretion levels and the SNP rs3181035 , which was significantly associated with type C scar lesions. Our findings strongly suggest that immunogenetic studies of individuals with congenital or postnatally acquired infection are needed to better understand the role of IFN-γ and its polymorphisms in the pathogenesis of ocular toxoplasmosis.

Single nucleotide polymorphisms in the interferon gamma gene are associated with distinct types of retinochoroidal scar lesions presumably caused by Toxoplasma gondii infection

The association of single nucleotide polymorphisms (SNPs) in the interferon (IFN)-γ gene ( IFNG ) with different types of retinal scar lesions presumably caused by toxoplasmosis were investigated in a cross-sectional population-based genetic study. Ten SNPs were investigated and after Bonferroni correction, only the associations between SNPs rs2069718 and rs3181035 with retinal/retinochoroidal scar lesions type A (most severe scar lesions) and C (least severe scar lesions), respectively, remained significant. The associations of two different IFNG SNPs with two different types of retinal lesions attributable to toxoplasmosis support the hypothesis that different inflammatory mechanisms underlie the development of these lesions. The in vitro analysis of IFN-γ secretion by peripheral blood mononuclear cells stimulated with Toxoplasma gondii antigens was also investigated. The association between SNP rs2069718 and type A scar lesions revealed that differential IFN-γ levels are correlated with distinct genotypes. However, no correlation was observed with IFN-γ secretion levels and the SNP rs3181035 , which was significantly associated with type C scar lesions. Our findings strongly suggest that immunogenetic studies of individuals with congenital or postnatally acquired infection are needed to better understand the role of IFN-γ and its polymorphisms in the pathogenesis of ocular toxoplasmosis.

Association of a NOD2 gene polymorphism and T-helper 17 cells with presumed ocular toxoplasmosis.

Retinochoroiditis manifests in patients infected with Toxoplasma gondii. Here, we assessed 30 sibships and 89 parent/case trios of presumed ocular toxoplasmosis (POT) to evaluate associations with polymorphisms in the NOD2 gene. Three haplotype-tagging single-nucleotide polymorphisms (tag-SNPs) within the NOD2 gene were genotyped. The family-based association test showed that the tag-SNP rs3135499 is associated with retinochoroiditis (P = .039). We then characterized the cellular immune response of 59 cases of POT and 4 cases of active ocular toxoplasmosis (AOT). We found no differences in levels of interferon γ (IFN-γ) and interleukin 2 produced by T-helper 1 cells when comparing patients with AOT or POT to asymptomatic individuals. Unexpectedly, we found an increased interleukin 17A (IL-17A) production in patients with POT or OAT. In patients with POT or AOT, the main cellular source of IL-17A was CD4(+)CD45RO(+)T-bet(-)IFN-γ(-) T-helper 17 cells. Altogether, our results suggest that NOD2 influences the production of IL-17A by CD4(+) T lymphocytes and might contribute to the development of ocular toxoplasmosis.

Candidate gene analysis of ocular toxoplasmosis in Brazil: evidence for a role for toll-like receptor 9 (TLR9)

Toxoplasma gondii infection is an important mediator of ocular disease in Brazil more frequently than reported from elsewhere. Infection and pathology are characterized by a strong proinflammatory response which in mice is triggered by interaction of the parasite with the toll-like receptor (TLR)/MyD88 pathway. A powerful way to identify the role of TLRs in humans is to determine whether polymorphisms at these loci influence susceptibility to T. gondii-mediated pathologies. Here we report on a small family-based study (60 families; 68 affected offspring) undertaken in Brazil which was powered for large effect sizes using single nucleotide polymorphisms with minor alleles frequencies > 0.3. Of markers in TLR2, TLR5 and TLR9 that met these criteria, we found an association Family Based Association Tests [(FBAT) Z score = 4.232; p = 1.5 x 10-5; p corrected = 1.2 x 10-4] between the C allele (frequency = 0.424; odds ratio = 7; 95 percent confidence interval 1.6-30.8) of rs352140 at TLR9 and toxoplasmic retinochoroiditis in Brazil. This supports the hypothesis that direct interaction between T. gondii and TLR9 may trigger proinflammatory responses that lead to severe pathologies such as the ocular disease that is associated with this infection in Brazil.

Host immune response to Toxoplasma gondii and Ascaris lumbricoides in a highly endemic area: evidence of parasite co-immunomodulation properties influencing the outcome of both infections.

Toxoplasmosis and ascaridiasis evoke polar Th-1 and Th-2 host immune responses, respectively. A study to investigate the specific cytokine profile production by in vitro cultures of peripheral blood mononuclear cells from individuals living under precarious sanitary conditions in a highly endemic area for the parasites Toxoplasma gondii and Ascaris lumbricoides was conducted. High levels of both IFN-gamma (Th-1) and IL-13 (Th-2) were observed in groups of co-infected individuals presenting toxoplasmic ocular lesions. Significantly lower IL-10 and TGF-beta levels were produced by co-infected individuals in comparison with groups of individuals not infected with A. lumbricoides and either positive or negative for T. gondii living under good sanitary conditions (control groups). The possible influence of co-parasitism on the clinical presentation of ocular toxoplasmosis is discussed.

Host immune response to Toxoplasma gondii and Ascaris lumbricoides in a highly endemic area: evidence of parasite co-immunomodulation properties influencing the outcome of both infections

Toxoplasmosis and ascaridiasis evoke polar Th-1 and Th-2 host immune responses, respectively. A study to investigate the specific cytokine profile production by in vitro cultures of peripheral blood mononuclear cells from individuals living under precarious sanitary conditions in a highly endemic area for the parasites Toxoplasma gondii and Ascaris lumbricoides was conducted. High levels of both IFN-³ (Th-1) and IL-13 (Th-2) were observed in groups of co-infected individuals presenting toxoplasmic ocular lesions. Significantly lower IL-10 and TGF-² levels were produced by co-infected individuals in comparison with groups of individuals not infected with A. lumbricoides and either positive or negative for T. gondii living under good sanitary conditions (control groups). The possible influence of co-parasitism on the clinical presentation of ocular toxoplasmosis is discussed.

Candidate gene analysis of ocular toxoplasmosis in Brazil: evidence for a role for toll-like receptor 9 (TLR9).

Toxoplasma gondii infection is an important mediator of ocular disease in Brazil more frequently than reported from elsewhere. Infection and pathology are characterized by a strong proinflammatory response which in mice is triggered by interaction of the parasite with the toll-like receptor (TLR)/MyD88 pathway. A powerful way to identify the role of TLRs in humans is to determine whether polymorphisms at these loci influence susceptibility to T. gondii-mediated pathologies. Here we report on a small family-based study (60 families; 68 affected offspring) undertaken in Brazil which was powered for large effect sizes using single nucleotide polymorphisms with minor alleles frequencies > 0.3. Of markers in TLR2, TLR5 and TLR9 that met these criteria, we found an association Family Based Association Tests [(FBAT) Z score = 4.232; p = 1.5 x 10-5; p corrected = 1.2 x 10-4] between the C allele (frequency = 0.424; odds ratio = 7; 95% confidence interval 1.6-30.8) of rs352140 at TLR9 and toxoplasmic retinochoroiditis in Brazil. This supports the hypothesis that direct interaction between T. gondii and TLR9 may trigger proinflammatory responses that lead to severe pathologies such as the ocular disease that is associated with this infection in Brazil.

Ocular sequelae of congenital toxoplasmosis in Brazil compared with Europe.

BACKGROUND: Toxoplasmic retinochoroiditis appears to be more severe in Brazil, where it is a leading cause of blindness, than in Europe, but direct comparisons are lacking. Evidence is accumulating that more virulent genotypes of Toxoplasma gondii predominate in South America. METHODS: We compared prospective cohorts of children with congenital toxoplasmosis identified by universal neonatal screening in Brazil and neonatal or prenatal screening in Europe between 1992 and 2003, using the same protocol in both continents. RESULTS: Three hundred and eleven (311) children had congenital toxoplasmosis: 30 in Brazil and 281 in Europe, where 71 were identified by neonatal screening. Median follow up was 4.1 years in Europe and 3.7 years in Brazil. Relatively more children had retinochoroiditis during the first year in Brazil than in Europe (15/30; 50% versus 29/281; 10%) and the risk of lesions by 4 years of age was much higher: the hazard ratio for Brazil versus Europe was 5.36 (95%CI: 3.17, 9.08). Children in Brazil had larger lesions, which were more likely to be multiple and to affect the posterior pole (p<0.0001). In Brazil, visual impairment (<6/12 Snellen) was predicted for most affected eyes (87%, 27/31), but not in Europe (29%; 20/69, p<0.0001). The size of newly detected lesions decreased with age (p = 0.0007). CONCLUSIONS: T. gondii causes more severe ocular disease in congenitally infected children in Brazil compared with Europe. The marked differences in the frequency, size and multiplicity of retinochoroidal lesions may be due to infection with more virulent genotypes of the parasite that predominate in Brazil but are rarely found in Europe.

Waterborne toxoplasmosis, Brazil, from field to gene.

Water was the suspected vehicle of Toxoplasma gondii dissemination in a toxoplasmosis outbreak in Brazil. A case-control study and geographic mapping of cases were performed. T. gondii was isolated directly from the implicated water and genotyped as SAG 2 type I.

Type 1 chemokine receptor expression in Chagas' disease correlates with morbidity in cardiac patients.

Chemokines and chemokine receptors (CKRs) control the migration of leukocytes during the inflammatory process and are important immunological markers of type 1 (CCR5 and CXCR3) and type 2 (CCR3 and CCR4) responses. The coexpression of CKRs (CCR2, CCR3, CCR5, CXCR3, and CXCR4) and intracellular cytokines (interleukin-10 [IL-10], IL-4, tumor necrosis factor alpha [TNF-alpha], and gamma interferon [IFN-gamma]) on T CD4+ and CD8+ peripheral cells from individuals with indeterminate (IND) or cardiac (CARD) clinical forms of Chagas' disease after in vitro stimulation with Trypanosoma cruzi antigens, were evaluated in this study. The percentage of T CD4+ and CD8+ cells coexpressing CCR5 and IFN-gamma, CXCR3 and IFN-gamma, and CXCR3 and TNF-alpha were higher in CARD than in IND individuals; on the other hand, the percentage of T CD4+ or CD8+ cells coexpressing CCR3 and IL-10 or coexpressing CCR3 and IL-4 were lower in CARD individuals than in IND individuals. In addition, a significant positive correlation between the expression of CCR5 or CXCR3 and IFN-gamma was observed in CARD individuals contrasting with a significant positive correlation between the expression of CCR3 and IL-4 and of CCR3 and IL-10 in IND patients. These results reinforce the hypothesis that a T. cruzi-exacerbated specific type 1 immune response developed by CARD chagasic patients is associated with the development of heart pathology.