Discovery of a HapE mutation that causes azole resistance in Aspergillus fumigatus through whole genome sequencing and sexual crossing.

Azole compounds are the primary therapy for patients with diseases caused by Aspergillus fumigatus. However, prolonged treatment may cause resistance to develop, which is associated with treatment failure. The azole target cyp51A is a hotspot for mutations that confer phenotypic resistance, but in a...

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Autores principales:	Simone M T Camps, Bas E Dutilh, Maiken C Arendrup, Antonius J M M Rijs, Eveline Snelders, Martijn A Huynen, Paul E Verweij, Willem J G Melchers
Formato:	article
Lenguaje:	EN
Publicado:	Public Library of Science (PLoS) 2012
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/a0f56d7baf5d45fd85b31e063bd0b19f
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spelling	oai:doaj.org-article:a0f56d7baf5d45fd85b31e063bd0b19f2021-11-18T08:06:56ZDiscovery of a HapE mutation that causes azole resistance in Aspergillus fumigatus through whole genome sequencing and sexual crossing.1932-620310.1371/journal.pone.0050034https://doaj.org/article/a0f56d7baf5d45fd85b31e063bd0b19f2012-01-01T00:00:00Zhttps://www.ncbi.nlm.nih.gov/pmc/articles/pmid/23226235/?tool=EBIhttps://doaj.org/toc/1932-6203Azole compounds are the primary therapy for patients with diseases caused by Aspergillus fumigatus. However, prolonged treatment may cause resistance to develop, which is associated with treatment failure. The azole target cyp51A is a hotspot for mutations that confer phenotypic resistance, but in an increasing number of resistant isolates the underlying mechanism remains unknown. Here, we report the discovery of a novel resistance mechanism, caused by a mutation in the CCAAT-binding transcription factor complex subunit HapE. From one patient, four A. fumigatus isolates were serially collected. The last two isolates developed an azole resistant phenotype during prolonged azole therapy. Because the resistant isolates contained a wild type cyp51A gene and the isolates were isogenic, the complete genomes of the last susceptible isolate and the first resistant isolate (taken 17 weeks apart) were sequenced using Illumina technology to identify the resistance conferring mutation. By comparing the genome sequences to each other as well as to two A. fumigatus reference genomes, several potential non-synonymous mutations in protein-coding regions were identified, six of which could be confirmed by PCR and Sanger sequencing. Subsequent sexual crossing experiments showed that resistant progeny always contained a P88L substitution in HapE, while the presence of the other five mutations did not correlate with resistance in the progeny. Cloning the mutated hapE gene into the azole susceptible akuB(KU80) strain showed that the HapE P88L mutation by itself could confer the resistant phenotype. This is the first time that whole genome sequencing and sexual crossing strategies have been used to find the genetic basis of a trait of interest in A. fumigatus. The discovery may help understand alternate pathways for azole resistance in A. fumigatus with implications for the molecular diagnosis of resistance and drug discovery.Simone M T CampsBas E DutilhMaiken C ArendrupAntonius J M M RijsEveline SneldersMartijn A HuynenPaul E VerweijWillem J G MelchersPublic Library of Science (PLoS)articleMedicineRScienceQENPLoS ONE, Vol 7, Iss 11, p e50034 (2012)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Simone M T Camps Bas E Dutilh Maiken C Arendrup Antonius J M M Rijs Eveline Snelders Martijn A Huynen Paul E Verweij Willem J G Melchers Discovery of a HapE mutation that causes azole resistance in Aspergillus fumigatus through whole genome sequencing and sexual crossing.
description	Azole compounds are the primary therapy for patients with diseases caused by Aspergillus fumigatus. However, prolonged treatment may cause resistance to develop, which is associated with treatment failure. The azole target cyp51A is a hotspot for mutations that confer phenotypic resistance, but in an increasing number of resistant isolates the underlying mechanism remains unknown. Here, we report the discovery of a novel resistance mechanism, caused by a mutation in the CCAAT-binding transcription factor complex subunit HapE. From one patient, four A. fumigatus isolates were serially collected. The last two isolates developed an azole resistant phenotype during prolonged azole therapy. Because the resistant isolates contained a wild type cyp51A gene and the isolates were isogenic, the complete genomes of the last susceptible isolate and the first resistant isolate (taken 17 weeks apart) were sequenced using Illumina technology to identify the resistance conferring mutation. By comparing the genome sequences to each other as well as to two A. fumigatus reference genomes, several potential non-synonymous mutations in protein-coding regions were identified, six of which could be confirmed by PCR and Sanger sequencing. Subsequent sexual crossing experiments showed that resistant progeny always contained a P88L substitution in HapE, while the presence of the other five mutations did not correlate with resistance in the progeny. Cloning the mutated hapE gene into the azole susceptible akuB(KU80) strain showed that the HapE P88L mutation by itself could confer the resistant phenotype. This is the first time that whole genome sequencing and sexual crossing strategies have been used to find the genetic basis of a trait of interest in A. fumigatus. The discovery may help understand alternate pathways for azole resistance in A. fumigatus with implications for the molecular diagnosis of resistance and drug discovery.
format	article
author	Simone M T Camps Bas E Dutilh Maiken C Arendrup Antonius J M M Rijs Eveline Snelders Martijn A Huynen Paul E Verweij Willem J G Melchers
author_facet	Simone M T Camps Bas E Dutilh Maiken C Arendrup Antonius J M M Rijs Eveline Snelders Martijn A Huynen Paul E Verweij Willem J G Melchers
author_sort	Simone M T Camps
title	Discovery of a HapE mutation that causes azole resistance in Aspergillus fumigatus through whole genome sequencing and sexual crossing.
title_short	Discovery of a HapE mutation that causes azole resistance in Aspergillus fumigatus through whole genome sequencing and sexual crossing.
title_full	Discovery of a HapE mutation that causes azole resistance in Aspergillus fumigatus through whole genome sequencing and sexual crossing.
title_fullStr	Discovery of a HapE mutation that causes azole resistance in Aspergillus fumigatus through whole genome sequencing and sexual crossing.
title_full_unstemmed	Discovery of a HapE mutation that causes azole resistance in Aspergillus fumigatus through whole genome sequencing and sexual crossing.
title_sort	discovery of a hape mutation that causes azole resistance in aspergillus fumigatus through whole genome sequencing and sexual crossing.
publisher	Public Library of Science (PLoS)
publishDate	2012
url	https://doaj.org/article/a0f56d7baf5d45fd85b31e063bd0b19f
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Discovery of a HapE mutation that causes azole resistance in Aspergillus fumigatus through whole genome sequencing and sexual crossing.

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