Molecular epidemiology of azole-resistant Aspergillus fumigatus in respiratory infections

Aspergillus fumigatus is a ubiquitous ascomycete mould and the primary cause of aspergillosis which varies in severity and clinical presentation. Triazoles have been the most widely used antifungal agents in prophylaxis and treatment of Aspergillus-related infections. Since the late 2000s there has been a steady increase in the number of reported resistance to azole antifungals in A. fumigatus, causing a major clinical concern with subsequent treatment failure among some patients. This PhD aimed to clarify the epidemiology of azole-resistant A. fumigatus at a regional level in West London. Firstly, the prevalence of azole-resistance in a large collection of clinical A. fumigatus isolates collected over a 17 year period (1998 - 2017) was determined, which surprisingly found a low level of resistance among mixed patient population. Conversely, a two-year (2015 - 2017) passive surveillance of isolates obtained from a specialist cardio-thoracic centre located in the same region, revealed an alarmingly high prevalence of resistance mostly among patients with cystic fibrosis. Secondly, to evaluate the application of whole-genome sequencing (WGS) in interrogation of azole-resistant A. fumigatus, a global panel of 24 isolates was analysed. WGS demonstrated that SNP analysis was able to accurately identify polymorphisms in the cyp51A gene conferring resistance to triazole antifungals. The WGS further associated to a globally panmictic population in A. fumigatus. In London, WGS revealed that environmentally-driven TR34/L98H, G54W and P216L were common alterations in cyp51A. A novel TR34/L98H/T289A/G448S polymorphism was discovered here causing a pan-azole-resistant phenotype. Phylogenomic analysis of sequenced genomes including London isolates, showed two novel major clades; one clade contains azole-resistant A. fumigatus with TR34/L98H allele, while the second clade consists of most azole-susceptible isolates. Finally, this PhD characterises the phenotypic heterogeneity of A. fumigatus and showed that acidic pH tolerance exists among some clinical A. fumigatus isolates. Furthermore, phenotypic switching accompanied with enhanced growth rate found in one azole-resistant isolate indicates that azole-resistant A. fumigatus has high fitness in azole-free environments and confirms that sexual reproduction leads to transmission of TR34/L98H allele in the absence of exposure to medical or fungicidal azole agents. Overall these findings indicate that prevalence of azole-resistance is linked to specific patient populations in regional level. Presence of TR34/L98H among azole-resistant isolates suggests a spillover of environmentally acquired antifungal resistance into susceptible patient cohorts.Open Acces