Molecular Profiling of Fungi using specific primers

Accurate information about fungal identification is very important in various fields, such as medical, agronomic, environmental, and industrial. Traditional morphological and biochemical-based assays can be time-consuming, nonspecific, and fail to detect cryptic or uncultured species. Conversely, molecular profiling based particularly on the use of specific primers in polymerase chain reaction (PCR) methods offers a highly efficient alternative, as it is sensitive, highly specific, and relatively fast. This paper presents general aspects of fungal molecular profiling, the main genetic regions commonly employed: Internal Transcribed Spacer (ITS), Large Subunit (LSU), and Beta-tubulin(β-tubulin), and how to use specific primers for varied species and genera. It also discusses their incorporation into sophisticated technologies such as quantitative PCR (qPCR), multiplex PCR (mPCR), or Next-generation sequencing (NGS) methodologies, including metabarcoding. The discussion incorporates clinical diagnostics, crop disease management, environmental monitoring, and food safety applications, but also addresses some challenges like those linked to primer bias, cross-reactivity, or the lack of reference data. In addition, recent technologies such as Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-based methods for nucleic acid detection and AI-assisted primer design provide enormous potential for this field. The study emphasizes the continued importance of multidisciplinary approaches in fungal diagnosis.