Variation in copy number of the 28S rDNA of Aspergillus fumigatus measured by droplet digital PCR and analog quantitative real-time PCR

Alexandre Alanio; Aude Sturny-Leclère; Marion Benabou; Nicolas Guigue; Stéphane Bretagne

doi:10.1016/j.mimet.2016.06.015

Variation in copy number of the 28S rDNA of Aspergillus fumigatus measured by droplet digital PCR and analog quantitative real-time PCR

J Microbiol Methods. 2016 Aug:127:160-163. doi: 10.1016/j.mimet.2016.06.015. Epub 2016 Jun 14.

Authors

Alexandre Alanio¹, Aude Sturny-Leclère², Marion Benabou³, Nicolas Guigue⁴, Stéphane Bretagne⁵

Affiliations

¹ Parasitology-Mycology Laboratory, Lariboisière Saint-Louis Fernand Widal Hospitals, Assistance Publique-Hôpitaux de Paris (AP-HP), France; Université Paris-Diderot, Sorbonne Paris Cité, France; Institut Pasteur, Molecular Mycology Unit, CNRS URA3012, National Reference Center of Invasive Mycoses and Antifungals, France.
² Institut Pasteur, Molecular Mycology Unit, CNRS URA3012, National Reference Center of Invasive Mycoses and Antifungals, France.
³ Institut Pasteur, Human Genetics and Cognitive Functions Unit, Paris, France.
⁴ Parasitology-Mycology Laboratory, Lariboisière Saint-Louis Fernand Widal Hospitals, Assistance Publique-Hôpitaux de Paris (AP-HP), France.
⁵ Parasitology-Mycology Laboratory, Lariboisière Saint-Louis Fernand Widal Hospitals, Assistance Publique-Hôpitaux de Paris (AP-HP), France; Université Paris-Diderot, Sorbonne Paris Cité, France; Institut Pasteur, Molecular Mycology Unit, CNRS URA3012, National Reference Center of Invasive Mycoses and Antifungals, France. Electronic address: stephane.bretagne@aphp.fr.

PMID: 27316653
DOI: 10.1016/j.mimet.2016.06.015

Abstract

Droplet digital PCR (ddPCR) after DNA digestion yielded a 28S rDNA copy number of 61 to 86 copies/genome when testing 10 unrelated Aspergillus fumigatus isolates, higher than with quantitative PCR. Unfortunately, ddPCR after DNA digestion did not improve the sensitivity of our PCR assay when testing serum patients with invasive aspergillosis.

Keywords: Aspergillus fumigatus; Copy number variation; Droplet digital PCR; Real-time quantitative PCR; rDNA.

MeSH terms

Aspergillosis / diagnosis
Aspergillosis / microbiology
Aspergillus fumigatus / genetics*
DNA Copy Number Variations*
DNA, Ribosomal / genetics*
DNA, Ribosomal / isolation & purification
Humans
RNA, Ribosomal, 28S / genetics
Real-Time Polymerase Chain Reaction* / instrumentation
Real-Time Polymerase Chain Reaction* / methods
Sensitivity and Specificity

Substances

DNA, Ribosomal
RNA, Ribosomal, 28S