title: Development of a real-time PCR assay for quantification of Acanthamoeba trophozoites and cysts.
abstract: Free-living amoebae have been found to be a reservoir for various pathogenic bacteria in aquatic environments. For example, the Acanthamoeba genus renders possible the intracellular multiplication of Legionella pneumophila, which is responsible for legionellosis. It consequently matters to quantify Acanthamoeba cells and thereby enhance our assessment of the risk of contamination. The classical microbiological method of quantification relies on amoebae growth and most probable number calculation. We have developed a real-time PCR assay based on a TaqMan probe that hybridizes onto 18S rDNA. This probe is specific to the Acanthamoeba genus. The assay was successful with both the trophozoite and the cyst forms of Acanthamoeba. Highly sensitive, it proved to permit detection of fewer than 10 cells, even those that are not easily cultivable, such as the cyst forms.
authors.affiliation: Equipe de Microbiologie, Laboratoire de Chimie de l'Eau et de l'Environnement, UMR CNRS 6008, Université de Poitiers, 40 avenue du recteur Pineau, 86022 Poitiers, France.
authors.forename: Delphine
authors.initials: D
authors.lastname: Rivière
authors.forename: Florence Ménard
authors.initials: FM
authors.lastname: Szczebara
authors.forename: Jean-Marc
authors.initials: JM
authors.lastname: Berjeaud
authors.forename: Jacques
authors.initials: J
authors.lastname: Frère
authors.forename: Yann
authors.initials: Y
authors.lastname: Héchard
mesh_terms: D000048:Acanthamoeba / Q000235:genetics* / Q000254:growth & development* / Q000302:isolation & purification; D048248:Acanthamoeba castellanii / Q000235:genetics / Q000254:growth & development / Q000302:isolation & purification; D000818:Animals; D017931:DNA Primers / Q000235:genetics; D016054:DNA, Protozoan / Q000032:analysis* / Q000302:isolation & purification; D017125:Genes, Protozoan; D016133:Polymerase Chain Reaction / Q000379:methods*; D012337:RNA, Ribosomal, 18S / Q000235:genetics; D012680:Sensitivity and Specificity; D016415:Sequence Alignment; D013045:Species Specificity
chemical_list: D017931:DNA Primers; D016054:DNA, Protozoan; D012337:RNA, Ribosomal, 18S