Contribution of molecular diagnosis to congenital toxoplasmosis

doi:10.1016/j.diagmicrobio.2013.02.008

Diagnostic Microbiology and Infectious Disease

Volume 76, Issue 2, June 2013, Pages 244-247

https://doi.org/10.1016/j.diagmicrobio.2013.02.008 Get rights and content

Abstract

We evaluated the performance of three real-time polymerase chain reaction (PCR) assays on 73 samples from mothers and children with congenital toxoplasmosis. PCR assays had significantly higher sensitivity in prenatal period than in birth period when targeting the 529-bp repeat element (81.3% versus 36.0%) or the B1 gene (64.6% versus 20.0%).

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Acknowledgments

This study was funded by the Molecular Biology pole of the French National Reference Centre for Toxoplasmosis (Centre National de Référence de la Toxoplasmose). The authors thank Ray Pierce and Jay Krugman for their attentive critical reading of the manuscript, and declare having no conflict of interest.

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Identification of a 200- to 300-fold repetitive 529 bp DNA fragment in Toxoplasma gondii, and its use for diagnostic and quantitative PCR
Int J Parasitol
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Comparison of loop-mediated isothermal amplification (LAMP) and real-time PCR method targeting a 529-bp repeat element for diagnosis of toxoplasmosis
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Diagnosis of congenital toxoplasmosis by polymerase chain reaction on neonatal peripheral blood
Diagn Microbiol Infect Dis
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Comparison between two amplification sets for molecular diagnosis of toxoplasmosis by real-time PCR
J Clin Microbiol
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Diagnosis of congenital toxoplasmosis: comparison of targets for detection of Toxoplasma gondii by PCR
J Clin Microbiol
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Classification system and case definitions of Toxoplasma gondii infection in immunocompetent pregnant women and their congenitally infected offspring. European Research Network on Congenital Toxoplasmosis
Eur J Clin Microbiol Infect Dis
(1996)

There are more references available in the full text version of this article.

Cited by (34)

Real-time PCR in the diagnosis of congenital toxoplasmosis
2023, Brazilian Journal of Infectious Diseases
The diagnosis of congenital toxoplasmosis presents limitations and therefore new options are necessary. The analysis of amniotic fluid by real-time PCR has already proved effective for confirmation of fetal infection. However, its performance in other biological samples is not clear yet. The aim of this study is to better understand the role of real-time PCR in the blood of the mother and newborn as well as in the amniotic fluid and placenta in the diagnosis of congenital toxoplasmosis. This is a descriptive cohort study of pregnant women with toxoplasmosis followed up in Rio de Janeiro, Brazil. Real-time PCR was performed in samples of maternal blood, amniotic fluid, placenta, and blood of newborns. In addition, histopathological examination of placentas was performed, and data collected from babies were collected. 116 pregnant women were followed up and 298 samples were analyzed. One (0.9%) pregnant woman presented positive PCR in the blood, 3 (3.5%) in the amniotic fluid, 1 (2.3%) in the placenta and no newborn had positive PCR in the blood. Histopathological study was suggestive of toxoplasmosis infection in 24 (49%) placentas. Six (5.2%) newborns were diagnosed with congenital toxoplasmosis, and only cases with positive PCR in the amniotic fluid had correlation of the PCR result with the diagnosis of congenital infection. Both maternal and blood samples of newborns and placenta did not prove to be promising in the diagnosis of congenital toxoplasmosis. Further studies are needed to evaluate the real role of molecular diagnosis in other biological materials rather than the amniotic fluid.
Molecular diagnosis of Toxoplasma gondii
2023, Molecular Medical Microbiology, Third Edition
A PCR assay with high sensitivity and specificity for the detection of swine toxoplasmosis based on the GRA14 gene
2021, Veterinary Parasitology
Toxoplasma gondii, an intracellular apicomplexan protozoan parasite, can infect all warm-blooded animals. Infected swine are considered one of the most important sources of T. gondii infection in humans. Rapidly and effectively diagnosing T. gondii infection in swine is essential. PCR-based diagnostic tests have been fully developed, and very sensitive and specific PCR is crucial for the diagnosis of swine toxoplasmosis. In this study, we used the T. gondii dense granule protein 14 (GRA14) gene as a target to design specific primers and established a high-specificity and high-sensitivity PCR detection method for swine toxoplasmosis. Notably, this PCR method could detect T. gondii tachyzoite DNA in the acute infection phase. The GRA14 gene PCR assay detected a minimum of 2.35 tachyzoites of T. gondii and can be used for T. gondii detection in blood, tissue, semen, urine and waste feed specimens. A total of 5462 blood specimens collected from pigs in 5 provinces and autonomous regions in southern China during 2016–2017 were assessed by the newly established GRA14 gene PCR method. The overall T. gondii infection rate was 18.9 % (1033/5462). According to the statistical analysis of different regions in China, the positive rates of swine toxoplasmosis from 2016 to 2017 were highest in the Shaanxi, Fujian and Guangdong areas, at 31.7 % (44/139), 21.9 % (86/391) and 18.8 % (874/4645), respectively. Specimens collected in 2017 had a higher positive rate (19.1 %) than those collected in 2016 (16.1 %). In addition, specimens collected in autumn (39.4 %), spring (22.8 %) and winter (18.2 %) had higher positive rates than those collected in summer (3.8 %). These results indicate that the new PCR method based on the T. gondii GRA14 gene has utility for the diagnosis of swine toxoplasmosis and can facilitate the diagnosis of toxoplasmosis in clinical laboratories.
Toxoplasmosis and pregnancy
2020, Journal de Pediatrie et de Puericulture
La toxoplasmose est une parasitose encore fréquente en France, dont il convient de ne pas méconnaître les manifestations cliniques, les conséquences à court ou à long terme, et les moyens diagnostiques. La prévention de la toxoplasmose congénitale repose sur un dépistage sérologique des femmes enceintes et des recommandations adaptées pour éviter une infection. Dans le contexte d’une primo-infection ou d’une suspicion d’infection précoce chez la femme enceinte, la datation est indispensable pour adapter la prise en charge en fonction du risque. Le traitement, éventuellement modifié après les résultats du diagnostic prénatal, permet de limiter la fréquence et la sévérité des infections congénitales. Dans tous les cas, même si la PCR toxoplasmose est négative dans le liquide amniotique, le dépistage postnatal est indispensable et repose essentiellement sur l’imagerie et le suivi sérologique du le nouveau-né pendant au moins 8 mois. Les techniques sérologiques sont variées et hétérogènes, et l’avis d’un laboratoire spécialisé est nécessaire, afin d’adapter au mieux la prise en charge de la femme enceinte et de son bébé.
Toxoplasmosis is a parasitic infection still frequent in France, the clinical manifestations of which, the short- or long-term consequences, and the diagnostic tools should not be overlooked. The prevention of toxoplasmosis is based on serological screening of pregnant women and hygienic recommendations. When a primary infection is suspected or documented in a pregnant woman, the estimated time of infection is essential and allows to adapt patient management. The maternal treatment, possibly modified after the result of prenatal diagnosis, reduces the frequency of vertical transmission and the severity of congenital infection. Postnatal screening of neonates is necessary, even if Toxoplasma PCR is negative in the amniotic fluid, and is based on imaging and serological follow-up of the newborn during at least 8 months. The serological techniques are numerous and heterogeneous, and require careful interpretation when specific IgM are detected, taking advise from a reference laboratory to adapt the management of the pregnant woman and her baby.
Congenital toxoplasmosis: Review of the CNR Toxoplasmosis, Pasteur Institute of Algeria
2019, Journal de Pediatrie et de Puericulture
L’infection toxoplasmique au cours de la grossesse peut être transmise au fœtus et entraîner une toxoplasmose congénitale.
Nous discutons sur le plan clinique et biologique 49 cas de toxoplasmose congénitale diagnostiqués sur une période de 16 ans (2002–2017).
L’infection pergravidique a eu lieu le premier trimestre dans 29 cas, au deuxième trimestre dans 4 cas et au troisième trimestre dans 5 cas. Pour 11 gestantes la date présumée de contamination n’a pas été précisée. La spiramycine a été prescrite pour 31 gestantes. L’échographie fœtale n’a pas été contributive dans notre série et le diagnostic anténatal non fait. Quarante-sept grossesses ont abouti et donné naissance à 47 nouveau-nés dont 40 étaient asymptomatiques et 7 ont présenté des formes patentes. Les sept nouveau-nés ont présenté une atteinte neurologique avec éruption cutanée dans deux cas et associée à une atteinte oculaire dans deux cas. Deux avortements spontanés à 4,5 et 5 mois de gestation. Le sérodiagnostic était positif pour les 47 nouveau-nés avec une meilleure sensibilité du western blot (100 %) par rapport à la recherche d’IgM (27,65 %). Trente et un nouveau-nés (65,95 %) ont été traités par la pyriméthamine-sulfadoxine. L’évolution était marquée par deux décès précoces, une choriorétinite dans 4 cas et une atteinte neurologique dans 3 cas.
L’information et la sensibilisation des femmes enceintes doivent être améliorées afin d’assurer une meilleure prise en charge de la toxoplasmose congénitale en Algérie.
During pregnancy, toxoplasmic infection can be transmitted to the fetus and lead to congenital toxoplasmosis.
We discuss clinically and biologically 49 cases of congenital toxoplasmosis diagnosed over a period of 16 years (2002–2017).
Per-pregnancy infection occurred in the first trimester in 29 cases, in the second trimester in four cases and in the third trimester in five cases. For 11 pregnant, the presumed date of contamination was not specified. Spiramycin was prescribed for 31 pregnant women. Fetal ultrasound was not contributive in our series and prenatal diagnosis not done. Forty-seven pregnancies led to the birth of 47 newborns, of which 40 were asymptomatic and 7 presented patent forms. The seven neonates had neurological involvement with rash in two cases and ocular involvement in two cases. Two spontaneous abortions at 4.5 and 5 months of gestation. The serodiagnosis was positive for the 47 newborns with a better sensitivity of the western blot (100%) compared to the search for IgM (27, 65%). Thirty-one newborns (65.95%) were treated with pyrimethamine sulfadoxine. The course was marked by two early deaths, chorioretinitis in four cases and neurological involvement in three cases.
The information and awareness of pregnant women must be improved to ensure better management of congenital toxoplasmosis in Algeria.
Evaluation of five automated and one manual method for Toxoplasma and human DNA extraction from artificially spiked amniotic fluid
2018, Clinical Microbiology and Infection
Citation Excerpt :
A total of 1296 PCRs were realized, including 972 Toxoplasma PCRs, of which 576 were reactions with DNA extracts from AF samples at the 1 T/mL concentration, 216 PCRs for the detection of inhibitors and 108 were human β-globin PCRs. Two different proficient Toxoplasma real-time PCR assays, targeting the 529-bp genomic repeat element [10], were performed: (a) TaqMan probe detection on an ABI Prism 7000 instrument (Applied Biosystems-Fisher Scientific, Illkirch, France) in one laboratory [1,2,8] and (b) FRET probes on a LightCycler 2.0 system (Roche) in the second laboratory [8]. Five microlitres of DNA extracts was added to each amplification reaction.
Molecular detection of Toxoplasma gondii plays a crucial role in the prenatal and neonatal diagnosis of congenital toxoplasmosis (CT). Sensitivity of this diagnosis is partly related to the efficiency of parasite DNA extraction and amplification. DNA extraction methods with automated platforms have been developed. Therefore, it is essential to evaluate them in combination with adequate PCR amplification assays.
In this multisite study, we investigated the suitability of two recent automated procedures for the isolation of Toxoplasma DNA from amniotic fluid (AF) (Magtration system 12GC, PSS and Freedom EVO VacS, Tecan), compared with three other automated procedures (MagNAPure Compact, Roche, BioRobot EZ1, Qiagen and modified NucliSens easyMAG, bioMérieux) and with the manual DNA extraction QIAamp DNA Mini kit (Qiagen). Two Toxoplasma PCR assays targeting the ‘529-bp’ repeat DNA element were used, based upon dual hybridization (FRET) or hydrolysis (TaqMan) probes. A total of 1296 PCRs were performed including 972 Toxoplasma PCRs.
We showed variable efficacy (4.2%–100% positive results) among the DNA extraction procedures in isolating up to five T. gondii cells/mL in AF samples. Moreover, for a given DNA extraction method, variable results were obtained among the two Toxoplasma PCR assays for detecting up to five T. gondii cells/mL: when using TaqMan PCR, all the automated systems yielded more than 60% positive results. Nevertheless, when testing the DNA extracts in triplicate, four out of six extraction methods allowed a satisfactory detection of low amounts of T. gondii DNA (≥33% of positive results) independently of the PCR assay used.
Despite the influence of the subsequent PCR method used, this study should help microbiologists in the choice of DNA extraction methods for the detection of T. gondii in amniotic fluid. The extraction method should be checked as adequate for the PCR assay used.

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Diagnostic Microbiology and Infectious Disease

Abstract

Section snippets

Acknowledgments

References (13)

Identification of a 200- to 300-fold repetitive 529 bp DNA fragment in Toxoplasma gondii, and its use for diagnostic and quantitative PCR

Int J Parasitol

Comparison of loop-mediated isothermal amplification (LAMP) and real-time PCR method targeting a 529-bp repeat element for diagnosis of toxoplasmosis

Vet Parasitol

Diagnosis of congenital toxoplasmosis by polymerase chain reaction on neonatal peripheral blood

Diagn Microbiol Infect Dis

Comparison between two amplification sets for molecular diagnosis of toxoplasmosis by real-time PCR

J Clin Microbiol

Diagnosis of congenital toxoplasmosis: comparison of targets for detection of Toxoplasma gondii by PCR

J Clin Microbiol

Classification system and case definitions of Toxoplasma gondii infection in immunocompetent pregnant women and their congenitally infected offspring. European Research Network on Congenital Toxoplasmosis

Eur J Clin Microbiol Infect Dis

Cited by (34)

Real-time PCR in the diagnosis of congenital toxoplasmosis

Molecular diagnosis of Toxoplasma gondii

A PCR assay with high sensitivity and specificity for the detection of swine toxoplasmosis based on the GRA14 gene

Toxoplasmosis and pregnancy

Congenital toxoplasmosis: Review of the CNR Toxoplasmosis, Pasteur Institute of Algeria

Evaluation of five automated and one manual method for Toxoplasma and human DNA extraction from artificially spiked amniotic fluid

NoteContribution of molecular diagnosis to congenital toxoplasmosis

Abstract

Section snippets

Acknowledgments

Int J Parasitol

Vet Parasitol

Diagn Microbiol Infect Dis

Comparison between two amplification sets for molecular diagnosis of toxoplasmosis by real-time PCR

J Clin Microbiol

Diagnosis of congenital toxoplasmosis: comparison of targets for detection of Toxoplasma gondii by PCR

J Clin Microbiol

Classification system and case definitions of Toxoplasma gondii infection in immunocompetent pregnant women and their congenitally infected offspring. European Research Network on Congenital Toxoplasmosis

Eur J Clin Microbiol Infect Dis

Note
Contribution of molecular diagnosis to congenital toxoplasmosis