This study compared the YeastOne and reference CLSI M38-A2 broth microdilution methods for antifungal susceptibility testing of species. The MICs of antifungal agents were determined for 100 isolates, including 54 (24 TR/L98H isolates), 23 , 13, and 10 isolates. The overall agreement (within 2 2-fold dilutions) between the two methods was 100%, 95%, 92%, and 90% for voriconazole, posaconazole, itraconazole, and amphotericin B, respectively. The voriconazole geometric mean (GM) MICs were nearly identical for all isolates using both methods, whereas the itraconazole and posaconazole GM MICs obtained using the YeastOne method were approximately 1 dilution lower than those obtained using the reference method. In contrast, the amphotericin B GM MIC obtained using the YeastOne method was 3.3-fold higher than that observed using the reference method. For the 24 TRL98H isolates assayed, the categorical agreement (classified according to the CLSI epidemiological cutoff values) was 100%, 87.5%, and 83.3% for itraconazole, voriconazole, and posaconazole, respectively. For four isolates, the itraconazole MICs were>8 μg/ml using the M38-A2 method due to trailing growth, whereas the corresponding itraconazole MICs obtained using the YeastOne method were all ≤0.25 μg/ml without trailing growth. These data suggest that the YeastOne method can be used as an alternative for azole susceptibility testing of species and for detecting the TR/L98H isolates but that this method fails to detect isolates exhibiting trailing growth with itraconazole. Additionally, for isolates with azole MICs that approach or that are at susceptibility breakpoints or with high amphotericin B MICs detected using the YeastOne method, further MIC confirmation using the reference CLSI method is needed.

This study delineated the characteristics of 24 (11.2%) culture-positive, influenza-associated pulmonary aspergillosis (IAPA) patients out of 215 patients with severe influenza during 2016–2019 in a medical center in southern Taiwan. Twenty (83.3%) patients did not have EORTC/MSG-defined host factors. The mean time from influenza diagnosis to Aspergillus growth was 4.4 days, and 20 (83.3%) developed IAPA within seven days after influenza diagnosis. All patients were treated in intensive care units and all but one (95.8%) received mechanical ventilation. Aspergillus tracheobronchitis was evident in 6 (31.6%) of 19 patients undergoing bronchoscopy. Positive galactomannan testing of either serum or bronchoalveolar lavage was noted in all patients. On computed tomography imaging, IAPA was characterized by peribronchial infiltrates, multiple nodules, and cavities superimposed on ground-glass opacities. Pure Aspergillus growth without bacterial co-isolation in culture was found in 17 (70.8%) patients. A. fumigatus (15, 62.5%), A. flavus (6, 25.0%), and A. terreus (4, 16.7%) were the major causative species. Three patients had mixed Aspergillus infections due to two species, and two had mixed azole-susceptible and azole-resistant A. fumigatus infection. All patients received voriconazole with an all-cause mortality of 41.6%. Of 14 survivors, the mean duration of antifungal use was 40.5 days. In conclusion, IAPA is an early and rapidly deteriorating complication following influenza that necessitates clinical vigilance and prompt diagnostic workup.

Background: Epidemiological knowledge is important to guide antifungal therapy.Objective: This multicentre study aimed to investigate the species distribution and antifungal susceptibility of Aspergillus isolates in Taiwan.Method: Four hundred and ninety-two clinical Aspergillus isolates, collected during 2016-2020, were identified by calmodulin sequencing and tested for antifungal susceptibility using CLSI M38-A3. The Cyp51A sequences of azole-resistant Aspergillus fumigatus and Aspergillus flavus isolates were analysed.Results: This collection comprised 30 species from eight Aspergillus sections-Flavi (33.5%), Nigri (26.0%), Fumigati (24.2%), Terrei (10.0%), Nidulantes (5.1%), Circumdati (0.8%), Restricti (0.2%) and Aspergillus (0.2%). Sections Fumigati, Flavi and Terrei were primarily represented by A. fumigatus (99.2%), A. flavus (95.8%) and A. terreus (100%), respectively. Section Nigri comprised nine species, mostly A. welwitschiae (60.2%), A. niger (12.5%), A. brunneoviolaceus (10.9%) and A. tubingensis (10.2%). A. fumigatus (39.6%) and A. flavus (26.4%) predominated among 53 isolates from lower respiratory samples, whereas section Nigri species (46.2%) and A. terreus (29.2%) predominated among 65 isolates from ear samples. Reduced susceptibility to amphotericin B (minimal inhibitory concentration (MIC) > 1 μg/mL) was noted in A. flavus (7.0%), A. terreus (6.1%), A. nidulans and section Circumdati (A. flocculosus, A. subramanianii and A. westerdijkiae) isolates. Acquired azole resistance was observed in seven A. fumigatus (5.9%), all of which carried TR 34 /L98H or TR 34 /L98H/S297T/F495I mutation, and three A. flavus (1.9%), one of which carried G441S mutation. Reduced susceptibility to itraconazole (MIC >1 μg/mL) was noted in 55.5% of section Nigri isolates, mainly in A. welwitschiae, A. niger and A. tubingensis, whereas A. brunneoviolaceus, A. aculeatinus and A. japonicus were hypersusceptible to azoles. Anidulafungin was active against all isolates except for one isolate. Conclusions:This study depicted the molecular epidemiology and species-specific characteristics of Aspergillus in Taiwan, which aids in appropriate antifungal therapy

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