Laboratory diagnosis of Toxoplasma gondii infection with direct and indirect diagnostic techniques.

Background: Toxoplasma gondii infection, common parasitic zoonoses, is an important cause of spontaneous abortion, mental retardation, encephalitis, ocular disease and death worldwide. Today the major diagnostic techniques for the toxoplasmosis are serological assays, but its have many limitations. Aim : The goal in this study is to improve the diagnostic accuracy of T. gondii infection, using direct (Real Time PCR) and indirect (IgM, IgA, IgG and IgG avidity) diagnostic techniques. Materials and Methods: In the period between 2007 and 2008, 96 non consecutive different clinical samples (38 blood, 40 amniotic fluids, 8 cerebrospinal fluids, 10 vitreous humors) and 96 sera have been studied simultaneously through molecular biology and serological techniques. Results: Direct and indirect diagnostic techniques used in this study for laboratory diagnosis of T. gondii infection were always concordant. Conclusions : The high correlation between direct and indirect diagnostic techniques exhibit that serologic techniques are accurate diagnostic assays as screening test in laboratory diagnosis of toxoplasmosis.

Avaliação comparativa entre o desempenho dos reagentes paratoxoplasmose AxSYM, VIDAS e VIDIA em um país latinoamericano com alta soroprevalência / Comparative evaluation of AxSYM, VIDAS and VIDIA toxoplasmosis reagent performance in a high seroprevalence Latin American country

Aims: The purpose of this study was to compare the performance of three automated immunoassays for the detection of IgM and IgG Toxoplasma gondii antibodies using sera of pregnant women living in Colombia, a Latin American country with a high seroprevalence. Methods: A total of 905 sera were tested for IgM antibodies and 914 for IgG antibodies with AxSYM, VIDAS and VIDIA immunoassays. Discrepancies were resolved by using the dye test for IgG antibodies, and the ISAGA test for IgM. Results: The overall agreement between AxSYM, VIDAS and VIDIA assays was excellent for detection of IgG and IgM antibodies, and discrepancies were relatively rare (3.6% and 5.5% of sera for IgG and IgM antibodies, respectively). The performance of the three immunoassays was similar for the detection of IgG antibodies with high sensitivity (100.00% for VIDIA, 99.59% for VIDAS, 99.38% for AxSYM) and specificity (99.04% for VIDIA, 98.82% for AxSYM, 98.57% for VIDAS). The specificity for IgM antibodies was excellent for the three immunassays (99.88% for VIDIA, 99.76% for AxSYM and VIDAS). The sensitivity of the detection of IgM antibodies was higher with VIDIA (95.12%) than with VIDAS (76.74%) and AxSYM (61.90%) assays. The correlation between IgG titers was limited between AxSYM and VIDAS assays and between AxSYM and VIDIA assays, but was excellent between VIDIA and VIDAS assays. Conclusions: Our study performed with Latin American sera confirmed the excellent specificity of AxSYM, VIDAS and VIDIA assays for the detection of IgG and IgM antibodies already reported in other countries. The sensitivity of the detection of IgG antibodies was slightly higher with VIDIA than with VIDAS and AxSYM assays. The sensitivity of the detection of IgM antibodies was higher with VIDIA than with VIDAS and AxSYM assays.

Triagem para toxoplasmose na gestação: um ano de experiência em um laboratório de referência italiano / Screening for toxoplasmosis during pregnancy: one-year experience in an Italian reference laboratory

Pavia, IRCCS Foundation, San Matteo Polyclinic Pavia, a reference laboratory for diagnosis of toxoplasmosis, in the investigation of women with suspected acute toxoplasmosis. Methods: All sera were tested with LIAISON® Toxo IgM and IgG II, Toxo IgG Avidity II kits (DiaSorin, Saluggia, Italy), VIDAS Toxo IgG II and Toxo IgG Avidity (bioMérieux, Marcy l?Etoile, France), IgM ISAGA (bioMérieux, Marcy l?Etoile, France) and ETI-TOXOK-A reverse PLUS (DiaSorin, Saluggia, Italy). When required (IgG negative/IgM positive women), IgG/IgM Western Blot II (LDBio, Lyon, France) was also performed. Prenatal diagnosis on amniotic fluid was done by nested PCR. All newborns were followed up to one year of age in order to exclude or confirm the diagnosis of congenital toxoplasmosis. All pregnant women with acute or undetermined stages of infection were treated. Results: In the course of 2007, 236 women with suspected acute (IgM-positive) Toxoplasma infection were followed up. In the reference laboratory, 91 women had test results indicating acute toxoplasmosis, and 10 had undetermined status of infection. These 101 patients represented 42.8% of the 236 women referred. Acute toxoplasmosis could be excluded in the remaining 135 patients, of whom 53 were non-immune. Three infected newborns were observed, all from mothers tested for the first time during the third trimester of pregnancy. Conclusions: The role of a reference laboratory in suspected toxoplasmosis acquired during pregnancy is crucial to date the infection and discriminate between seroconversion and false positive anti-Toxoplasma IgM antibodies. This avoids unnecessary anxiety in immune women, provides correct counseling about primary prevention and periodic testing for seronegative ones, and allows early treatment and follow-up of pregnant women with acute infection and their newborns.

Dificuldades observadas em um centro de referência no diagnóstico e manejo de gestantes com toxoplasmose / Difficulties observed in a reference center in the diagnosis and management of pregnant women with toxoplasmosis

Aims: To evaluate the difficulties met in the care of pregnant women with toxoplasmosis diagnosis in antenatal care services. Methods: Longitudinal prospective study with 262 pregnant women referred to the Toxoplasmosis Clinic at Instituto de Pesquisa Clínica Evandro Chagas-Fundação Oswaldo Cruz, between January 2005 and July 2009. Results: Most women (91.2%) were in the second and third trimesters of pregnancy, and 81.3% were referred by public health services. The average delay was 113.4 days in the collection of the first sample for serological tests in antenatal care, 52.1 days for referral and 160.6 days in starting treatment. Younger women (P=0.03) and those from the public health system were referred later (P<0.05). Treatment was initiated at the origin for only 16% of the pregnant women, and 5% of these did not receive the recommended dose of spiramycin. At the Reference Center there was a low rate of confirmation of the serological tests performed in the health services of origin. It was found that 12.6% of pregnant women with an initial diagnosis of acute toxoplasmosis were susceptible to infection by Toxoplasma gondii. These tests were considered false positives. Conclusions: This study highlights the difficulties met in the management of pregnant women with toxoplasmosis in the antenatal care, including the quality of diagnostic tests and the need for greater emphasis on continuing education of health professionals.

Objetivos: avaliar as dificuldades encontradas no atendimento de gestantes com diagnóstico de toxoplasmose por parte de serviços de atendimento pré-natal. Métodos: estudo longitudinal, prospectivo, com 262 gestantes encaminhadas ao Ambulatório de Toxoplasmose do Instituto de Pesquisa Clínica Evandro Chagas-Fundação Oswaldo Cruz, entre janeiro de 2005 e julho de 2009. Resultados: a maioria das gestantes foram encaminhadas no segundo ou terceiro trimestre de gestação (91,2%) e por serviços públicos de saúde (81,3%). O tempo médio de demora na coleta de sangue para os testes sorológicos no pré-natal foi de 113,4 dias. Houve demora média de 52,1 dias para o encaminhamento e 160,6 dias para o início do tratamento. Mulheres mais jovens (P=0,03) e aquelas provenientes do sistema público de saúde (P<0,000) foram encaminhadas mais tardiamente. O tratamento foi iniciado na origem em apenas 16% das gestantes, e 5% destas não receberam a dose preconizada de espiramicina. No Centro de Referência houve baixa confirmação dos testes sorológicos realizados nos serviços de saúde de origem. Constatou-se que 12,6% das gestantes com diagnóstico inicial de toxoplasmose aguda eram suscetíveis à infecção por Toxoplasma gondii, sendo os testes considerados falso-positivos. Conclusões: este estudo destaca dificuldades observadas no manejo de gestantes com toxoplasmose por parte do atendimento pré-natal da rede básica de saúde, incluindo a atenção quanto à qualidade dos testes diagnósticos e a necessidade de maior ênfase na educação continuada dos profissionais de saúde.

Toxoplasmose e gestação: resultados perinatais e associação do teste de avidez de IgG com infecção congênita em gestantes com IgM anti-Toxoplasma gondii reagente / Toxoplasmosis and pregnancy: perinatal results and association of the IgG avidity test with congenital infection in pregnant women with positive anti-Toxoplasma gondii IgM

Objetivos: verificar os desfechos perinatais em gestantes com toxoplasmose aguda e se houve associação entre os resultados dos testes de avidez para anticorpos IgG anti-Toxoplasma gondii e a presença ou ausência de infecção fetal/neonatal. Métodos: um estudo transversal incluiu gestantes com diagnóstico sorológico de toxoplasmose apresentando IgM específica reagente, atendidas no Ambulatório de Gestação de Alto Risco da Faculdade de Medicina da Universidade Federal de Mato Grosso do Sul, no período de novembro de 2002 a novembro de 2007. Resultados do teste de avidez de IgG demonstrando índices superiores a 30% foram considerados alta avidez, enquanto valores inferiores a 30% foram considerados baixa avidez. Definiram-se como sendo de infecção fetal e/ou neonatal os casos com resultado positivo para a reação em cadeia da polimerase no líquido amniótico ou com IgM específica para toxoplasmose reagente no sangue do recém-nascido. Resultados: considerando-se todas as gestantes referidas para o ambulatório de gestação de alto risco no período estudado, a frequência de gestantes com IgM anti-Toxoplasma gondii reagente foi de 10,8% (176/1.634). A taxa de infecção congênita nessas pacientes foi de 4% (7/176). O teste de avidez de IgG foi realizado em 162 gestantes (92%), encontrando-se avidez alta em 144 (88,9%). Houve associação (p=0,003) entre avidez alta e ausência de toxoplasmose fetal/neonatal na amostra estudada, com razão de prevalência de 13,4 (intervalo de confiança [IC] 95% 2,2-86,6). O Valor preditivo positivo do teste de avidez (probabilidade de infecção congênita com avidez baixa) foi de 22% (IC 95% 6%-47%), enquanto o valor preditivo negativo (probabilidade de ausência da infecção congênita com avidez alta) foi de 98% (IC 95% 94%-99%).

Aims: To verify the perinatal outcomes in pregnant women with acute toxoplasmosis, and to determine if there was association between the results of Toxoplasma gondii-specific IgG avidity test and the presence or absence of fetal/neonatal infection. Methods: A cross-sectional study included pregnant women with serological diagnosis of acute toxoplasmosis (presenting a positive Toxoplasma gondii-specific IgM test) attended at the outpatient unit for high-risk pregnancy of the Faculty of Medicine, Federal University of Mato Grosso do Sul, Brazil, in the period from November 2002 to November 2007. Test results demonstrating IgG avidity index above 30% were considered high avidity, while values below 30% were considered low avidity. Fetal and/or neonatal infection was defined by positive result for the polymerase chain reaction in amniotic fluid, or by a positive Toxoplasma gondii-specific IgM test in the newborn?s serum. Results: Considering all pregnant women referred to the outpatient unit for high-risk pregnancy in the period of study, frequency of pregnant women with positive Toxoplasma gondii-specific IgM was 10.8% (176/1.634). The rate of congenital infection in these patients was 4% (7/176). The IgG avidity test was performed in 162 patients (92% of the 176 pregnant women with positive IgM), and the avidity was high in 144 (88.9%). There was an association (p=0.003) between high avidity and no fetal/neonatal toxoplasmosis in our sample, with a prevalence ratio of 13.4 (confidence interval [CI] 95% 2.2-86.6). The positive predictive value of the avidity test (probability of congenital infection with a low avidity) was 22% (95% 6%-47%), while the negative predictive value (probability of absence of congenital infection with a high avidity) was 98% (95% CI 94% -99%).