DETECTION OF SUSCEPTIBILITY AND RESISTANCE TO POLYENE AND AZOLE CLASS ANTIFUNGALS BY THE MINIMAL PROFILE CHANGE CONCENTRATION (MPCC) METHOD IN Aspergillus spp. FROM PATIENTS WITH CHRONIC AND INVASIVE PULMONARY ASPERGILLOSIS.

• Grant number: 24/17286-9
• Support Opportunities: Scholarships abroad - Research Internship - Post-doctor
• Start date: March 03, 2025
• End date: January 02, 2026
• Field of knowledge: Health Sciences - Medicine - Pathological Anatomy and Clinical Pathology
• Principal Investigator: Gil Benard
• Grantee: Tiago Alexandre Cocio
• Supervisor: Nelson Manuel Viana da Silva Lima
• Host Institution: Faculdade de Medicina (FM). Universidade de São Paulo (USP). São Paulo , SP, Brazil
• Institution abroad: Universidade do Minho (UMinho), Portugal
• Associated to the scholarship: 22/14747-0 - MOLECULAR EPIDEMIOLOGY AND AZOLE RESISTANCE MECHANISM IN ASPERGILLUS SPP.: A STUDY OF ACUTE INVASIVE AND POST-TB CHRONIC PULMONARY ASPERGILOSIS ISOLATES IN TWO REGIONS OF SÃO PAULO STATE., BP.PD

Abstract

Species of the genus Aspergillus spp. are considered fungal microorganisms with the highest frequency of isolation in clinical and environmental samples. These can cause infections in humans, leading to cutaneous and disseminated clinical manifestations, primarily affecting the lungs. The treatment of pulmonary aspergillosis has been carried out with antifungals from the azole class (itraconazole, voriconazole, and posaconazole) and the polyene class (amphotericin B). The therapeutic and prophylactic use of these antifungals has alarmingly increased in recent years, as has the number of species and variants of the genus Aspergillus that have acquired a high Minimum Inhibitory Concentration (MIC) against the main antifungals used in the treatment of the disease. To determine the MIC of antifungals in Aspergillus spp., "Antifungal Susceptibility Testing (AFST)" is used, following protocols that utilize the broth microdilution method with modifications according to different fungi (yeasts vs. filamentous), standardized by institutions such as the European Committee on Antibiotic Susceptibility Testing (EUCAST). To perform these tests, fungal cultures are needed, with defined periods of exposure to the antifungal, and analysis of the results within 72 hours. The delay in obtaining results is considered by many to be a negative factor. Therefore, there is a need to standardize more efficient and effective AFST tests using technological resources, such as the use of mass spectrometry, to obtain MIC values against relevant antifungals precisely and quickly. One of the recently used technologies is the MALDI-TOF MS technique, which determines the minimal profile change concentration (MPCC) by detecting proteomic changes in a given microorganism (bacteria and fungi) after exposure to antimicrobials at different concentrations. The study will include 80 clinical isolates of Aspergillus spp. that are part of the proponent's postdoctoral project. These isolates were collected at the Hospital das Clínicas of the University of São Paulo's Faculty of Medicine in São Paulo and Ribeirão Preto. The in vitro susceptibility test by broth microdilution will be based on the EUCAST E.DEF 9.4 protocol for filamentous fungi. Aspergillus spp. are being subjected to MIC determination against the antifungals amphotericin B, itraconazole, voriconazole, and posaconazole using the microdilution method on plates, so that the MIC values can serve as validation data to evaluate the efficacy of MPCC. In addition to the MPCC, isolates of Aspergillus spp. with high MIC to azoles will undergo sequencing of the cyp51A gene and its promoter region. Considering the scarcity of information in the literature on the use of MPCC in Aspergillus spp. using the EUCAST E.DEF 9.4 broth microdilution protocol for filamentous fungi, this proposal will test an innovative technique for evaluating the susceptibility of Aspergillus spp. to antifungals of importance for the treatment of patients with pulmonary aspergillosis.