Evaluation of a New Histoplasma spp. Quantitative RT-PCR Assay

Laboratory diagnosis of histoplasmosis is based on various methods, including microscopy, culture, antigen, and DNA detection of Histoplasma capsulatum var. capsulatum or Histoplasma capsulatum var. duboisii. To improve sensitivity of existing real-time quantitative PCR (qPCR) assays, we developed a...

Full description

Saved in:

Bibliographic Details
Published in:	The Journal of molecular diagnostics : JMD Vol. 23; no. 6; pp. 698 - 709
Main Authors:	Alanio, Alexandre, Gits-Muselli, Maud, Lanternier, Fanny, Sturny-Leclère, Aude, Benazra, Marion, Hamane, Samia, Rodrigues, Anderson Messias, Garcia-Hermoso, Dea, Lortholary, Olivier, Dromer, Françoise, Bretagne, Stéphane, Woimant, Marine Gosset, blanchard, Geneviève, Silhadi, Souad, Vignier, Nicolas, Pitsch, Aurelia, Jidar, Kaoutar, Traversier, Nicolas, Poisson, Didier, Lecointre, Claire, Foulet, Françoise, Botterel, Françoise, Ammar, Nawel Ait, Amsellem, Gabriel, Frederic, Poirier, Philipe, Cornu, Marjorie, Loridant, Severine, Morio, Florent, Boutoille, David, Jeddi, Fakhri, Hasseine, Lilia, Ouissa, Rachida, Toubas, Dominique, Bailly, Eric, Désoubeaux, Guillaume, Ronez, Emily, Foulon, Guillaume, Lefrançois, Sebastien, Bonnal, Christine, Paugam, André, Dougados, Maxime, Desroches, Marine, Barazzutti, Hélène, Paleiron, Nicolas, rabodonirina, Meja, Catherinot, Emilie, Cardot-Martin, Emilie, Hiesse, Chrisian, Salvator, Hélène, Aguilar, Claire, Gigandon, Anne, de Montpreville, Thomas, Bougnoux, Marie-Elisabeth, Sitterlé, Emilie, Fekkar, Arnaud, Imbert, Sébastien, Bleibtreu, Alexandre, Senghor, Yaye, Denis, Blandine, Molina, Jean-Michel, Liegeon, Geoffroy, Munnier, Anne-Lise, Malphettes, Marion, Denis, Julie, Berlioz-Arthaud, Alain, Lange, Franciska, Chiaruzzi, Myriam, Epelboin, Loic
Format:	Journal Article
Language:	English
Published:	United States Elsevier Inc 01-06-2021
Subjects:	DNA, Fungal - analysis Genes, Fungal Histoplasma - genetics Humans Limit of Detection Prospective Studies Real-Time Polymerase Chain Reaction - methods Reproducibility of Results Reverse Transcriptase Polymerase Chain Reaction - methods
Online Access:	Get full text
Tags:	Add Tag No Tags, Be the first to tag this record!

Description
Summary:	Laboratory diagnosis of histoplasmosis is based on various methods, including microscopy, culture, antigen, and DNA detection of Histoplasma capsulatum var. capsulatum or Histoplasma capsulatum var. duboisii. To improve sensitivity of existing real-time quantitative PCR (qPCR) assays, we developed a new RT-qPCR assay that allows amplification of whole nucleic acids of Histoplasma spp. validated on suspected cases. The limit of detection was 20 copies, and the specificity against 114 fungal isolates/species was restricted to Histoplasma spp. Whole nucleic acids of 1319 prospectively collected consecutive samples from 907 patients suspected of having histoplasmosis were tested routinely between May 2015 and May 2019 in parallel with standard diagnostic procedures performed in parallel. Forty-four had proven histoplasmosis attributable to H. capsulatum var. capsulatum (n = 40) or H. capsulatum var. duboisii (n = 4) infections. The results of RT-qPCR were positive in 43 of 44 patients (97.7% sensitivity) in at least one specimen. Nine of 863 cases (99% specificity) were RT-qPCR positive and therefore classified as possible cases. RT-qPCR was positive in 13 of 30 (43.3%) blood samples tested in proven cases. A positive RT-qPCR result in blood was significantly associated with H. capsulatum var. capsulatum progressively disseminated histoplasmosis with a positive RT-qPCR result in 92.3% of the immunocompromised patients with disseminated disease. This new Histoplasma RT-qPCR assay enabling amplification of H. capsulatum var. capsulatum and H. capsulatum var. duboisii is highly sensitive and allows the diagnosis of histoplasmosis advantageously from blood and bronchoalveolar lavage fluid.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23
ISSN:	1525-1578 1943-7811
DOI:	10.1016/j.jmoldx.2021.02.007