dAzole resistance in Aspergillus fumigatus is an increasing problem. The TR34 L98H and TR46 Y121F T289A mutations that can occur in patients without previous azole exposure have been reported in Europe, Asia, the Middle East, Africa, and Australia. Here, we report the detection of both the TR34 L98H and TR46 Y121F T289A mutations in confirmed A. fumigatus isolates collected in institutions in the United States. These mutations, other mutations known to cause azole resistance, and azole MICs are reported here. Invasive aspergillosis remains a major problem in immunocompromised individuals, including solid organ transplant recipients, those undergoing hematopoietic stem cell transplant, and patients receiving highly immunosuppressive chemotherapies (1-3). In patients with structural damage to the lungs, such as those who have had tuberculosis or sarcoidosis, chronic pulmonary aspergillosis is also a significant problem, the prevalence of which has been estimated to be approximately 3 million patients worldwide (4-7). The azoles are a mainstay in the treatment of invasive aspergillosis, as the members of this class that have activity against Aspergillus species, itraconazole, posaconazole, voriconazole, and isavuconazole, may be given orally, and the treatment of this invasive fungal infection is often prolonged. However, prolonged therapy may predispose patients to adverse effects and drug interactions associated with these agents and increase the potential for the development of drug-resistant organisms (8). Over the last several years, concern has been growing regarding azole resistance in Aspergillus fumigatus (9, 10). Azole-resistant A. fumigatus isolates recovered from patients failing therapy have been reported in several countries around the world. Mutations in the CYP51A gene, which encodes the Cyp51 enzyme responsible for the last step in ergosterol biosynthesis, have been the resistance mechanism documented in clinical isolates, and these mutations have been found in isolates recovered from patients who have had long exposures to azoles (11, 12). However, resistant isolates harboring tandem repeats of various sizes in the promoter region of the CYP51A gene, along with nonsynonymous point mutations leading to amino acid changes in the Cyp51 enzyme, have been recovered from azole-naive patients with invasive aspergillosis (8, 12, 13). These resistance mechanisms, which include TR34 L98H and TR46 Y121F T289A, have been linked to the environmental use of azoles in agriculture and the preservation of various materials (14, 15), and they have been found in various countries, including many in Europe, India, China, Iran, Tanzania, and Australia (13, 16-21). However, these mutations have not yet been reported in isolates collected in the United States (22). Our objective was to evaluate the Cyp51-associated mechanisms of azole resistance in a collection of A. fumigatus isolates from institutions across the United States.The antifungal susceptibility database in the Fungus Testing Laboratory at the University of ...