Azole resistance in Aspergillus fumigatus. The first 2-year's Data from the Danish National Surveillance Study, 2018-2020

Abstract

Background: Azole resistance complicates treatment of patients with invasive aspergillosis with an increased mortality. Azole resistance in Aspergillus fumigatus is a growing problem and associated with human and environmental azole use. Denmark has a considerable and highly efficient agricultural sector. Following reports on environmental azole resistance in A. fumigatus from Danish patients, the ministry of health requested a prospective national surveillance of azole-resistant A. fumigatus and particularly that of environmental origin.

Objectives: To present the data from the first 2 years of the surveillance programme.

Methods: Unique isolates regarded as clinically relevant and any A. fumigatus isolated on a preferred weekday (background samples) were included. EUCAST susceptibility testing was performed and azole-resistant isolates underwent cyp51A gene sequencing.

Results: The azole resistance prevalence was 6.1% (66/1083) at patient level. The TR₃₄ /L98H prevalence was 3.6% (39/1083) and included the variants TR₃₄ /L98H, TR₃₄ ³ /L98H and TR₃₄ /L98H/S297T/F495I. Resistance caused by other Cyp51A variants accounted for 1.3% (14/1083) and included G54R, P216S, F219L, G54W, M220I, M220K, M220R, G432S, G448S and Y121F alterations. Non-Cyp51A-mediated resistance accounted for 1.2% (13/1083). Proportionally, TR₃₄ /L98H, other Cyp51A variants and non-Cyp51A-mediated resistance accounted for 59.1% (39/66), 21.2% (14/66) and 19.7% (13/66), respectively, of all resistance. Azole resistance was detected in all five regions in Denmark, and TR₃₄ /L98H specifically, in four of five regions during the surveillance period.

Conclusion: The azole resistance prevalence does not lead to a change in the initial treatment of aspergillosis at this point, but causes concern and leads to therapeutic challenges in the affected patients.

Keywords: Aspergillus fumigatus; TR34/L98H; antifungal susceptibility; azole resistance; environmental route; itraconazole; medical route; voriconazole.

FIGURE 1

MIC values for the included Aspergillus fumigatus isolates. Susceptible isolates (S) are shown green when susceptible at azole resistance screening. Susceptible isolates with an MIC are shown in blue, resistant isolates in red and isolates in the ATU for which the classification depends on the susceptibility of either itraconazole or voriconazole, respectively, are indicated in black. MIC values above 4 mg/L are shown as >4 mg/L. Isolates with no MICs for posaconazole (n = 1) and isavuconazole (n = 365) are not included in the diagrams

FIGURE 2

Cyp51A amino acid profiles found in the 66 patients with at least one resistant isolate. Each patient is shown only once. Some patients harbour resistant isolates with a Cyp51A resistance mechanism and resistant isolates with a non‐Cyp51A related mechanism (WT) (for example the two patients with P216S and wild‐type isolates)

FIGURE 3

Proportion of resistant Aspergillus fumigatus isolates and associated underlying resistance mechanism across the five Danish Regions. Each Region represented is the Region of the health care facility from which the isolate was referred. As some health care services are centralised this will not in all cases represent the patients' place of residence or the place where the resistant fungus was acquired. Total numbers of isolates were for Capital n = 910, Zealand n = 91, Southern Denmark n = 326, Central Jutland n = 419 and Northern Jutland n = 74. The resistance mechanism remained uncharacterised in five isolates from five patients whom were known to harbour other cyp51A mutant isolates (blue bar). These included four resistant isolates that did not undergo cyp51A sequencing of which three isolates derived from a patient who had other isolates with M220K, and one isolate from a patient who had other isolates with P216S, and one isolate with F46Y/M172/E427K from a patient who also had isolates with TR₃₄/L98H