Molecular detection of Aspergillus and corresponding azole resistance markers: systematic comparison of two commercial PCR test kits, AsperGenius and MycoGENIE

Daniel Goldenberger*1, Nina Khanna2, Elisabeth Wehrle3,2, Helena Seth-Smith4,1, Doris Hohler1, Vladimira Hinic1, Adrian Egli4,1

Author address: 

1 Clinical Microbiology, University Hospital Basel, Basel, Switzerland, 2 Department of Infectious Diseases & Hospital Epidemiology, University Hospital Basel, 3 Department of Infectious Diseases & Hospital Epidemiology, Hospital Delémont, 4 Applied Microbiology Research, Department of Biomedicine, University of Basel

Abstract: 

Background:

Azole resistance in Aspergillus fumigatus has emerged as a global health problem. Conventional methods such as culture have a reduced sensitivity and phenotypic susceptibility testing is laborious and time-consuming. Novel molecular diagnostic tools have the potential to provide more reliable results within hours.

Materials/methods:

We compared two commercially available diagnostic test kits for detection of A. fumigatus / species DNA as well as identification of distinct molecular azole resistance markers: the AsperGenius$(r)\$(AG) and MycoGENIE® (MG) tests. The assays were evaluated with six azole-susceptible and seven azole-resistant A. fumigatus reference isolates which were also analyzed using whole genome sequencing. Twelve Aspergillus non-fumigatus strains were used to test specificity. In addition, 46 clinical samples (31 BALs, 13 biopsies, 2 miscellaneous) from 41 patients tested positive by an in-house ITS region-based Aspergillus PCR were investigated.

Results:

Analysis of all Aspergillus culture isolates showed expected findings in both tests. The analytical sensitivity for A. fumigatus was from 1.5 to 15 fg in both tests, corresponding to 0.05 to 0.5 organisms per reaction. However, detection of azole resistance resulted in a reduced sensitivity of factor 10 to 100. The detection of Aspergillus organisms in clinical samples is shown in the table. In 18/37 A. fumigatus positive samples (48.6%), azole testing was successful by the AG test from which 14 showed wildtype (sensitive) markers and 4 TR34/L98H mutations. These four samples originated from a patient with simultaneous isolation of TR34/L98H-harbouring A. fumigatus. MG could not differentiate between a sensitive or a negative result and in contrast to AG 4/4 TR34/L98H-positive clinical samples could not be identified.

Conclusions:

In accordance with previous studies, the MG test shows a slightly higher sensitivity for detection of A. fumigatus DNA in comparison to the AG assay. However, azole susceptibility results from clinical samples reveals a reduced reliability in the MG assay.

Table: Aspergillus spp.-positive clinical samples (n=46)
MG assayAG assayClinical samples
A.fumigatusA.fumigatusAspergillus spp.No.
+++37
+--4
--+3
---2
Total46
2019

abstract No: 

P2192

Full conference title: 

European Congress of Clinical Microbiology & Infectious Diseases 2019
    • ECCMID 29th (2019)