Comparative Analysis of Fungal Proteomes for Identifying Resistance Genes: A Preliminary Step Towards Experimental Validation

Antimicrobial resistance (AMR) has been declared a global threat to public and environmental health by the World Health Organization (WHO). AMR investigation in fungal organisms has been limited to mainly clinically relevant species, leaving atypical and environmental isolates neglected. Due to this, this study aimed to employ computational methods for the identification of potential proteins associated with antifungal resistance genes. This served as the initial phase to determine a specific list of genes to look for functioning as a molecular catalog of specific sequences to experimentally validate in environmental fungal isolates currently available at the Laboratorio de Bioinformática Aplicada at the UNA. Our approach involved the analysis of ten proteomes, where we aimed to establish orthologous relationships with a well-validated database of antifungal genes; additionally, we characterized this orthologous by identifying the most frequent functional features. To achieve this, we used Orthofinder version 2.5.2 and observed that 96 284 genes (87.6% of the total) were assigned to 12 005 orthogroups. Four orthogroups (OG0000000, OG0000004, OG0000378 and OG0000630) were the only ones that contained sequences from all the analyzed species. Uncharacterized and hypothetical proteins (UP and HP respectively) were the most prevailing attributes, followed by ergosterol fungal cell wall and MFS and ABC transporters, showing sequences related to different mechanisms of antifungal resistance in the analyzed species. Nonetheless, we determined that many AMR proteins are present in non-pathogenic fungal genome sequences, which is of particular interest as these are usually not studied. AMR in fungi is complex as it is related to molecular and metabolic mechanisms rather than only specific molecules, this is integrated into eukaryotic intrinsic genomic complexity and fungal lack of knowledge This study represents the initial phase in a series of ongoing investigations at the LABAP aimed at advancing our understanding of AMR in environmental fungi. Furthermore, our findings lay the groundwork for the development of a future surveillance pipeline for environmental fungi resistance genes to validate in Costa Rica.