title: Characterization and multicentric validation of a common standard for Toxoplasma gondii detection using nucleic acid amplification assays.
abstract: The molecular diagnosis of toxoplasmosis essentially relies upon laboratory-developed methods and suffers from lack of standardization, hence the large diversity of performances between laboratories. Moreover, quantifications of parasitic loads differ among centers, a fact which prevents the possible prediction of the severity of this disease as a function of parasitic loads. The objectives of this multicentric study performed in eight proficient laboratories of the Molecular Biology Pole of the French National Reference Center for Toxoplasmosis (NRC-T) were (i) to assess the suitability of a lyophilized preparation of Toxoplasma gondii as a common standard for use in this PCR-based molecular diagnosis and (ii) to make this standard available to the community. High-quality written procedures were used for the production and qualification of this standard. Three independent batches of this standard, containing concentrations ranging from 10(4) to 0.01 T. gondii genome equivalents per PCR, were first assessed: the linear dynamic range was ≥ 6 log, the intra-assay coefficients of variation (CV) from a sample containing 10 T. gondii organisms per PCR were 0.3% to 0.42%, and the interassay CV over a 2-week period was 0.76% to 1.47%. A further assessment in eight diagnostic centers showed that the standard is stable, robust, and reliable. These lyophilized standards can easily be produced at a larger scale when needed and can be made widely available at the national level. To our knowledge, this is the first quality control assessment of a common standard which is usable both for self-evaluation in laboratories and for accurate quantification of parasitic loads in T. gondii prenatal infections.
authors.affiliation: CHU (University Hospital Centre) of Montpellier/University Montpellier I, Laboratoire de Parasitologie-Mycologie, Montpellier, France Pôle Biologie Moléculaire du Centre National de Référence de la Toxoplasmose, Montpellier, France University Montpellier I, Faculty of Pharmacy, Laboratoire de Biophysique & Bioanalyse, Montpellier, France.
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Varlet-Marie
authors.affiliation: CHU (University Hospital Centre) of Montpellier/University Montpellier I, Laboratoire de Parasitologie-Mycologie, Montpellier, France Pôle Biologie Moléculaire du Centre National de Référence de la Toxoplasmose, Montpellier, France.
authors.forename: Yvon
authors.initials: Y
authors.lastname: Sterkers
authors.affiliation: Pôle Biologie Moléculaire du Centre National de Référence de la Toxoplasmose, Montpellier, France CHU A. Michallon/University Joseph Fourier Grenoble I, Laboratoire de Parasitologie-Mycologie, Département des Agents Infectieux, Grenoble, France.
authors.forename: Marie-Pierre
authors.initials: MP
authors.lastname: Brenier-Pinchart
authors.affiliation: Pôle Biologie Moléculaire du Centre National de Référence de la Toxoplasmose, Montpellier, France CHU of Toulouse, Service de Parasitologie-Mycologie, Toulouse, France.
authors.forename: Sophie
authors.initials: S
authors.lastname: Cassaing
authors.affiliation: Pôle Biologie Moléculaire du Centre National de Référence de la Toxoplasmose, Montpellier, France CHU of Dijon, Laboratoire de Parasitologie-Mycologie, Faculty of Medicine, Université de Bourgogne, UMR 1347 Aggroécologie, Dijon, France.
authors.forename: Frédéric
authors.initials: F
authors.lastname: Dalle
authors.affiliation: Pôle Biologie Moléculaire du Centre National de Référence de la Toxoplasmose, Montpellier, France CHU of Lille/Université Nord de France (UDSL), Service de Parasitologie-Mycologie, Faculty of Medicine, Institut Pasteur de Lille, Lille, France.
authors.forename: Laurence
authors.initials: L
authors.lastname: Delhaes
authors.affiliation: Pôle Biologie Moléculaire du Centre National de Référence de la Toxoplasmose, Montpellier, France Hôpitaux Universitaires de Strasbourg/University of Strasbourg, Laboratoire de Parasitologie et Mycologie Médicales, Strasbourg, France.
authors.forename: Denis
authors.initials: D
authors.lastname: Filisetti
authors.affiliation: Pôle Biologie Moléculaire du Centre National de Référence de la Toxoplasmose, Montpellier, France CHU A. Michallon/University Joseph Fourier Grenoble I, Laboratoire de Parasitologie-Mycologie, Département des Agents Infectieux, Grenoble, France.
authors.forename: Hervé
authors.initials: H
authors.lastname: Pelloux
authors.affiliation: Pôle Biologie Moléculaire du Centre National de Référence de la Toxoplasmose, Montpellier, France AP-HP, Groupe Hospitalier Pitié-Salpêtrière, Laboratoire de Parasitologie-Mycologie, Paris, France.
authors.forename: Fériel
authors.initials: F
authors.lastname: Touafek
authors.affiliation: Pôle Biologie Moléculaire du Centre National de Référence de la Toxoplasmose, Montpellier, France University Paris Descartes, AP-HP, Groupe Hospitalier Cochin, Laboratoire de Parasitologie-Mycologie, Paris, France.
authors.forename: Hélène
authors.initials: H
authors.lastname: Yera
authors.affiliation: CHU (University Hospital Centre) of Montpellier/University Montpellier I, Laboratoire de Parasitologie-Mycologie, Montpellier, France Pôle Biologie Moléculaire du Centre National de Référence de la Toxoplasmose, Montpellier, France p-bastien@chu-montpellier.fr.
authors.forename: Patrick
authors.initials: P
authors.lastname: Bastien
mesh_terms: D005602:France; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods* / Q000592:standards*; D021141:Nucleic Acid Amplification Techniques / Q000379:methods* / Q000592:standards*; D059208:Parasite Load / Q000379:methods / Q000592:standards; D012015:Reference Standards; D014122:Toxoplasma / Q000235:genetics / Q000302:isolation & purification*; D014123:Toxoplasmosis / Q000175:diagnosis*