Aspergillus/isolation & purification; Air Pollution, Indoor; Public health; Real-time PCR; SYBR® green; Performance assessment
Abstract :
[en] Currently, contamination of indoor environment by fungi and molds is considered as a public health problem. The monitoring of indoor airborne fungal contamination is a common tool to help understanding the link between fungi in houses and respiratory problems. Classical analytical monitoring methods, based on cultivation and microscopic identification, depend on the growth of the fungi. Consequently, they are biased by difficulties to grow some species on certain culture media and under certain conditions or by noncultivable or dead fungi that can consequently not be identified. However, they could have an impact on human health as they might be allergenic. Since molecular methods do not require a culture step, they seem an excellent alternative for the monitoring of indoor fungal contaminations. As a case study, we developed a SYBR(R) green real-time PCR-based assay for the specific detection and identification of Aspergillus versicolor, which is frequently observed in indoor environment and known to be allergenic. The developed primers amplify a short region of the internal transcribed spacer 1 from the 18S ribosomal DNA complex. Subsequently, the performance of this quantitative polymerase chain reaction (qPCR) method was assessed using specific criteria, including an evaluation of the selectivity, PCR efficiency, dynamic range, and repeatability. The limit of detection was determined to be 1 or 2 copies of genomic DNA of A. versicolor. In order to demonstrate that this SYBR(R) green qPCR assay is a valuable alternative for monitoring indoor fungal contamination with A. versicolor, environmental samples collected in contaminated houses were analyzed and the results were compared to the ones obtained with the traditional methods.
Disciplines :
Biochemistry, biophysics & molecular biology
Author, co-author :
Libert, X.; Platform Biotechnology and Molecular Biology, Scientific Institute of Public Health (WIV-ISP), J. Wytsmanstraat 14, 1050 Brussels, Belgium
Chasseur, C.; Health and Environment, Scientific Institute of Public Health (WIV-ISP), J. Wytsmanstraat 14, 1050 Brussels, Belgium
Bladt, S.; Cellule Régionale d ’ Intervention en Pollution Intérieure (CRIPI), Brussels Environment (IBGE), Avenue du Port 86C/3000, 1000 Bruxelles, Belgium
Packeu, A.; Mycology and Aerobiology, Scientific Institute of Public Health (WIV-ISP), J. Wytsmanstraat 14, 1050 Brussels, Belgium
Bureau, Fabrice ; Université de Liège > Département des sciences fonctionnelles (DSF) > GIGA-R : Biochimie et biologie moléculaire
Roosens, N. H.; Platform Biotechnology and Molecular Biology, Scientific Institute of Public Health (WIV-ISP), J. Wytsmanstraat 14, 1050 Brussels, Belgium
De Keersmaecker, S. C. J.; Platform Biotechnology and Molecular Biology, Scientific Institute of Public Health (WIV-ISP), J. Wytsmanstraat 14, 1050 Brussels, Belgium
Language :
English
Title :
Development and performance assessment of a qualitative SYBR(R) green real-time PCR assay for the detection of Aspergillus versicolor in indoor air.
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