High-Frequency Direct Detection of Triazole Resistance in Aspergillus fumigatus from Patients with Chronic Pulmonary Fungal Diseases in India.

Brijesh Sharma,Ashutosh Singh,Anuradha Chowdhary,Jacques F Meis,Kaushal Kumar Mahto

doi:10.3390/jof6020067

Abstract

Aspergillosis due to azole-resistant Aspergillus fumigatus is a worldwide problem with major therapeutic implications. In patients with invasive aspergillosis, a low yield of fungal cultures results in underestimation of azole resistance. To detect azole resistance in A. fumigatus, we applied the AsperGenius® Resistance multiplex real-time polymerase chain reaction (PCR) assay to detect TR34/L98H, and TR46/T289A/Y121F mutations and the AsperGenius® G54/M220 RUO PCR assay to detect G54/M220 mutations directly in bronchoalveolar lavage (BAL) samples of 160 patients with chronic respiratory diseases in Delhi, India. Only 23% of samples were culture-positive compared to 83% positivity by A. fumigatus species PCR highlighting concerns about the low yield of cultures. Notably, 25% of BAL samples (33/160 patients) had azole resistance-associated mutation by direct detection using PCR assay. Detection of resistance-associated mutations was found mainly in 59% and 43% patients with chronic pulmonary aspergillosis (CPA) and allergic bronchopulmonary aspergillosis (ABPA), respectively. Overall, a G54 mutation, conferring itraconazole resistance, was the predominant finding in 87.5% and 67% of patients with CPA and ABPA, respectively. In culture-negative, PCR-positive samples, we detected azole-resistant mutations in 34% of BAL samples. Azole resistance in chronic Aspergillus diseases remains undiagnosed, warranting standardization of respiratory culture and inclusion of rapid techniques to detect resistance markers directly in respiratory samples.

Highlights

A wide spectrum of pulmonary fungal diseases, including invasive aspergillosis (IA), chronic pulmonary aspergillosis (CPA), Aspergillus bronchitis, and allergic bronchopulmonary aspergillosis (ABPA) are primarily caused by Aspergillus fumigatus [1]
The optimal cycle threshold cut-off value for the AsperGenius® Species real-time polymerase chain reaction (PCR) was
We demonstrate an alarmingly high frequency (25%) of azole resistance among patients with chronic and allergic bronchopulmonary aspergillosis

Summary

Introduction

A wide spectrum of pulmonary fungal diseases, including invasive aspergillosis (IA), chronic pulmonary aspergillosis (CPA), Aspergillus bronchitis, and allergic bronchopulmonary aspergillosis (ABPA) are primarily caused by Aspergillus fumigatus [1]. Azoles are the first line of treatment, independently of the clinical manifestation of disease [2]. The last decade has witnessed several case series of the emergence of azole resistance in clinical settings resulting in increasing rates of treatment failure. Azole resistance is often caused by mutations in the hot spot Cyp51A gene, implicated in the ergosterol biosynthesis pathway. J. Fungi 2020, 6, 67; doi:10.3390/jof6020067 www.mdpi.com/journal/jof

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