A peptide originated from Toxoplasma gondii microneme 8 displaying serological evidence to differentiate recent from chronic human infection.

Toxoplasmosis is able to cause death and/or sequelae in foetuses from pregnant women and immunocompromised individuals. The early diagnosis, able to differentiate acute from chronic phases, is essential to define the treatment against this disease and minimize the risk of complications. Here we describe a peptide derived from microneme 8 (pMIC8) protein of Toxoplasma gondii, able to distinguish the phase of infection. By using human and mice serum samples with different infection times, we assessed the ability of pMIC8 to interact with antibodies present in early of infection, and compared the results obtained with soluble antigen of T. gondii (STAg). The results showed that pMIC8 was recognized more precisely with antibodies present in serum samples from individuals with time of infection below 3 months, followed by those between 4 and 6 months of infection. Based on these results, it is possible to conclude that the association of immunoassays using STAg and pMIC8 as antigen preparations can be used to distinguish acute from chronic infections.

Proposed panel of diagnostic tools for accurate temporal classification of symptomatic T. gondii infection.

Serological tests available for the diagnosis of acute Toxoplasma gondii infection have limitations in establishing the temporal diagnosis of acute toxoplasmosis. The present analytical-descriptive investigation comprises of a prospective longitudinal cohort study to search for accurate biomarkers to distinguish acute, early and late convalescent T. gondii infection. Classic methods (immunofluorescence-IFA along with Enzyme-linked immunosorbent-ELISA and fluorescent-ELFA assays) for IgM, IgA, IgG and IgG avidity were employed in parallel with flow cytometry-based anti-fixed T. gondii tachyzoites serology (FC-AFTA-IgM, IgG, IgG avidity and IgG subclasses). The results reemphasized the limitations of IgM & IgG IFA, IgG ELFA, IgG & IgG subclasses FC as well as IgA ELISA biomarkers for the temporal diagnosis of acute toxoplasmosis. Receiver Operating-characteristics features (ROC-curves) were employed to adjust conventional cut-offs aiming at establishing a novel protocol to discriminate more accurately the different phases of toxoplasmosis. Conversely, IgM presented high diagnostic co-positivity for acute toxoplasmosis (97% for ELISA, 96% for ELFA and 95% for FC-AFTA) along with moderate co-negativity for detection of late convalescent toxoplasmosis (82%, 76% and 79%, respectively). IgG avidity (ELFA and FC-AFTA) outstand with the highest performance indices with 91% and 96% co-negativity for assessing acute toxoplasmosis and 91% and 98% co-positivity for late convalescent toxoplasmosis, respectively. Multivariate analysis generated a three-step algorithm comprising IgM ELFA screening followed by ELFA and FC-AFTA IgG avidity with high accuracy in discriminating acute from late convalescent infection. Together, these findings demonstrate the applicability of the proposed panel of diagnostic tools for accurate temporal classification of T. gondii infection.

Establishing tools for early diagnosis of congenital toxoplasmosis: Flow cytometric IgG avidity assay as a confirmatory test for neonatal screening.

The aim of this study was to evaluate the performance of conventional serology (Q-Preven™ and ELFAVIDAS™) and flow cytometry-based serologic tools for early serologic diagnosis of congenital toxoplasmosis. The study groups included prospectively confirmed cases of congenital toxoplasmosis (TOXO=88) and age-matching non-infected controls (NI=15).The results demonstrated that all samples tested positive/indeterminate for anti-T. gondii IgM screening at birth using air-dried whole blood samples. Serum samples collected at 30-45days after birth tested positive for ELFAVIDAS™ IgG in both groups. While all NI tested negative for ELFAVIDAS™ IgM and IgA, only 78% and 36% of TOXO tested positive for IgM and IgA, respectively. Flow cytometry-based anti-T. gondii IgM, IgA and IgG reactivity displayed moderate performance with low sensitivity (47.6%, 72.6% and 75.0%, respectively). Regardless the remarkable specificity of IgG1, IgG2 and IgG3 subclasses for early diagnosis, weak or moderate specificity was observed (Se=73.9%, 60.2% and 83.0%, respectively). The analysis of IgG avidity indices (AI) demonstrated the highest performance among the flow cytometry-based methods (Se=96.6%; Sp=93.3%), underscoring the low avidity index (AI<60%) within TOXO (97.0%) in contrast with the high avidity index (AI>60%) in NI (93%). Analysis of anti-T. gondii IgG and IgG3 reactivity for mother:infant paired samples may represent a relevant complementary tests for early diagnosis. In conclusion, a feasible high-standard algorithm (Accuracy=97.1%) was proposed consisting of Q-Preven™ IgM screening at birth, followed by ELFAVIDAS™ IgM and flow cytometric IgG avidity analysis at 30-45days after birth as a high performance tool for early serological diagnosis of congenital toxoplasmosis.

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.

Perfil clínico e epidemiológico e desfecho reprodutivo em gestantes infectadas pelo hivatendidas na maternidade de um hospital universitário em Vitória, Brasil / Clinical and epidemiological profile and reproductiveoutcome in hiv-infected pregnant women assisted at auniversity hospital maternity in Vitória, Brazil

A infecção pelo vírus da imunodeficiência humana (HIV), assim como a Síndrome da Imunodeficiência Adquirida (Aids), uma epidemia mundial,pode acarretar graves consequências em termos de morbidade e mortalidade materna e fetal. Objetivos: Descrever o perfil clínico e epidemiológico, e o desfecho reprodutivo em gestantes infectadas pelo HIV. Métodos: Estudo de corte transversal, com 109 gestantes infectadas pelo HIV que tiveram terminação na maternidade de um hospital universitário em Vitória, Espírito Santo, entre novembro de 2001 e maio de 2012. Os dados foram extraídos de prontuários médicos e registros públicos. Resultados: Os achados mais marcantes entre os casos foram idade materna média de 28 anos, pardas e negras (76,1%), até 8anos do Ensino Fundamental (63,3%), ocupação do lar (59,4%) e casada/união estável (70,6%). Eram nulíparas 24,1%, e 15,7% com 3 ou mais partos, 33%tiveram o diagnóstico de infecção pelo HIV durante a gestação atual, sendo 53,7% das gestantes com critérios para Aids. O parto cesáreo ocorreu em 82,6%dos casos, parto pretermo em 17,4%, baixo peso ao nascer em 23,9% e morte perinatal em 4,6% dos recém-nascidos. Conclusão: Observou-se nesta casuística a ocorrência de um perfil de gestantes de baixo nível socioeconômico. O parto pretermo e a morte perinatal foram mais comuns que na população em geral,sinalizando para a necessidade de ações preventivas durante o acompanhamento da gestante infectada pelo HIV para redução desses eventos.

The infection by the human immunodeficiency virus (HIV), as well as the acquired immune deficiency syndrome (Aids), a worldwide epidemic,may lead to serious consequences in terms of maternal and fetal morbidity and mortality. Objective: To describe the clinical and epidemiological profiles and the reproductive outcome in HIV-infected pregnant women. Methods: Cross-sectional study, with 109 pregnant women infected by HIV who had their termination in auniversity hospital maternity in Vitória, Espírito Santo, from November 2001 to May 2012. The data were extracted from medical and public records. Results: Themost prominent findings among the cases were average maternal age of 28 years, non-white (76.1%), up to 8 years of elementary school (63.3%), housewives (59.4%)and marital status married/cohabitation (70.6%). The nulliparous were 24.1%, and 15.7% had 3 or more childbirths, 33% had a diagnosis of HIV infection duringpregnancy, and 53.7% of pregnant women met the criteria for Aids. The cesarean occurred in 82.6% of cases, preterm birth in 17.4%, and low birth weight in 23.9%and perinatal death in 4.6% of the newborns. Conclusion: It has been observed, in this casuistry, a pregnant women profile of low socioeconomic level. Preterm birthand perinatal death were more common than in the general population, indicating the need for preventive actions for monitoring the HIV infected pregnant women inorder to reduce these events

Flow cytometry-based algorithm to analyze the anti-fixed Toxoplasma gondii tachyzoites IgM and IgG reactivity and diagnose human acute toxoplasmosis.

In the present study we evaluated the performance of a flow cytometry-based algorithm as a new serological approach to detect antibodies to T. gondii and specific IgG avidity to diagnose acute toxoplasmosis. The results showed that using FC-AFTA-IgM assay, all serum samples from patients with acute toxoplasmosis demonstrated seropositivity, whereas 90% of patients with chronic infection and 100% of non-infected individuals presented negative results. Thus, only 10% of patients with chronic toxoplasmosis showed residual IgM, in contrast with other methodologies used to diagnosis acute toxoplasmosis. On the order hand, FC-AFTA-IgG assay as well as FC-AFTA-IgG subclasses is unlikely to discriminate acute from chronic toxoplasmosis. We have also evaluated the performance of FC-AFTA-IgG avidity as a tool to exclude chronic toxoplasmosis in patients with positive FC-AFTA-IgM. Our data showed an excellent performance of FC-AFTA-IgG avidity employing the cut-off of 60% for Avidity Index (AI) with sensitivity and specificity of 100%. All serum samples from patients presenting acute toxoplasmosis showed low avidity index (AI≤60%), whereas all chronic patients showed high avidity index (AI>60%). The outstanding performance indexes of this novel flow cytometry-based algorithm support its use as a non-conventional alternative serological approach to diagnose human acute toxoplasmosis.

Mortalidade perinatal hospitalar: classificaçäo de óbitos do Hospital Universitário Cassiano Antonio Moraes: Vitória, Espírito Santo (1992-1993) / Mortality perinatal hospitalar: classification of the deaths of the University Hospital Cassiano Antonio Moraes: Vitória, Espírito Santo (1992-1993)

A Mortalidade Perinatal (MPN) säo considerados pela OMS uma prioridade em saúde infantil sendo seu conhecimento fundamental para sua prevençäo. A multiplicidade de causas que envolvem o óbito perinatal dificulta a confecçäo de uma classificaçäo que contemple apenas o evento nosológico desencadeante do óbito. Dentre as várias classificaçöes propostas, a de Wigglesworth tem como pressuposto básico a estratificaçäo dos óbitos por faixa de peso, que isoladamente e a variável que mais se correlaciona com a MPN, e sua classificaçäo em 5 grupos mutualmente exclusivos, dispostos abaixo. Além do mais, ela baseia-se em achados clínicos, prescindindo de necropsia, cujos dados também podem ser utilizados, se disponíveis. Grupo 1- Óbitos ocorridos antes do trabalho de parto (TP); Grupo 2- Malformaçöes Congénitas (MC); Grupo 3- Condiçöes relacionadas ao parto prematuro/imaturidade; Grupo 4- Condiçöes asfixicas desenvolvidas durante o TP e tocotraumatismo e, Grupo 5- Condiçöes especificas como isoimunizaçäo Rh e infecçöes congénitas. Identificamos e dimensionamos os problemas relacionados a MPN no Hospital Universitário Cassiano Antonio Moraes (HUCAM) da Universidade Federal do Espirito Santo. Paralelamente ele avaliou a exequibilidade e o grau de concordância da classificaçäo de Wigglesworth utilizando dados clínicos isoladamente e associados aos achados anatomopatológicos. Foram estudados todos os 152 óbitos perinatais ocorridos no HUCAM de janeiro de 1992 a dezembro de 1993, período em que nasceram vivos 4171 bebês pesando mais de 1000g. Dos 152 óbitos perinatais 89 eram natimortos e 63 neomortos, correspondendo a uma taxa de natimortalidade de 20,9/1000, neomortalidade de 15,1/1000 e MPN de 34,2/1000 nascimento únicos. A classificaçäo de Wigglesworth possibilitou identificar e dimensionar os principais problemas perinatais relacionados aos óbitos estudados, apontando áreas críticas no cuidado de saúde prestado no HUCAM. Mostrou também ser um excelente mecanismo de monitoraçäo da MPN, oferecendo subsídios para o planejamento de açöes de saúde na área perinatal.