Fungi represent an important group of eukaryotic microorganisms and play a pivotal role in processes such as organic matter cycling (including bioremediation and spoilage of fruits and vegetable) and plant growth promotion. During the various steps of their growth, some of them can produce different products of industrial interest, such as primary metabolites, secondary metabolites, enzymes, organic acids, alcohols, antibiotics, and biopolymers. They can have different morphologies, such as filamentous and unicellular (yeast) or both (dimorphic fungi), according to the environmental conditions. Fungal morphology is crucial for industrial processes and for their use in biotechnological processes (models for recombinant proteins). Fungi are characterized by a complex taxonomy mainly due to the presence of a teleomorphic and anamorphic phase during their life cycle. Some groups of fungi have essential ecosystem functions or can colonize a wide range of ecosystems, including extreme environments. The lifestyle of most of them requires molecular identification in environmental studies as well as their isolation and characterization for their further application in biotechnology. During the past decades, molecular tools have rapidly evolved, which has greatly improved our knowledge about fungal diversity in diverse habitats and ecosystems. This chapter provides an overview of the diversity of fungi as well as their roles in ecosystems and their application in biorefineries.