Aspergillus spp.: suscetibilidade aos antifúngicos isolados e em associação por diferentes metodologias e detecção de mecanismos de resistência a azólicos

Abstract

Aspergillosis is a group of diseases among the main causes of morbidity and mortality in immunosuppressed patients. Aspergillus fumigatus is the most common species that causes invasive and non-invasive aspergillosis in humans, followed by Aspergillus flavus, Aspergillus niger and Aspergillus terreus. Azole antifungals are the main treatment of these infections, with voriconazole being the drug of choice in invasive aspergillosis. Echinocandins and amphotericin B are also active against Aspergillus and are currently used in specific situations or in association with azoles. Azole resistance has been increasingly documented among medical centers around the world, mainly associated with A. fumigatus, and has become a major therapeutic problem. The development of resistance has been associated with mutations in the cyp51A gene of A. fumigatus. Two pathways of resistance selection have been studied, the pathway in the patient, and more recently, an environmental route. In order to overcome resistance, alternatives such as the use of combination drugs have been studied in the treatment of patients with aspergillosis refractory to standard treatment. This study aimed to determine the susceptibility profile of A. fumigatus and A. flavus, isolated from patients with aspergillosis or from the environment against azole antifungals, echinocandins and amphotericin B, alone and in combination, by using different methodologies. In addition, resistance mechanisms of clinical or environmental strains of A. fumigatus that showed resistance to one or more azole antifungals were investigated. The main results showed that in either environmental or clinical strains some azole resistant profiles were detected, while the echinocandins were very active against both species of Aspergillus. Amphotericin B showed more pronounced activity against A. fumigatus in comparison to A. flavus. In addition, Etest and agar plates with azole methodologies have been found to be capable of determining the minimum inhibitory concentration (MIC) and detecting resistance of A. fumigatus to isavuconazole, respectively, in accordance with the standard broth microdilution method. The interactions between azole and echinocandin antifungal agents against A. fumigatus varied depending on the methodology used, making it very important to standardize these methods to obtain concordant results. However, the synergistic interactions found in vitro and in vivo showed a favorable effect to the use of these associations in the treatment of aspergillosis. In the azole-echinocandin combinations against A. flavus, a higher percentage of synergistic interactions were observed when the MIC was read while by reading the minimum effective concentration (MEC) indifference was the main outcome. Furthermore, mutations linked to the cyp51A gene were found in one clinical and one environmental itraconazole-resistant A. fumigatus strains. In conclusion, combinations between azoles and echinocandins are promising for use in the treatment of patients with aspergillosis refractory to standard treatment; different Aspergillus susceptibility profiles to antifungals tested alone and in association can be found depending on the methodology used, making it necessary to standardize the different tests in order to have easier and faster methodologies that facilitate their use in routine laboratories. Finally, the first report of the presence of mechanisms of resistance to azoles linked to the cyp51A gene in Brazilian Aspergillus isolates generates concern with the management of aspergillosis in clinical centers of Brazil in order to avoid that the resistance rates increase by the indiscriminate use of azoles in the clinic and in agriculture.