Genetic Manipulation of Human Fungal Pathogens

The advent of precise genome editing has transformed our understanding of human fungal pathogens by enabling targeted interrogation of gene function, virulence mechanisms and drug-resistance determinants. In recent years, tools such as CRISPR-Cas9 have been adopted across major human-associated fungal species, including Candida albicans, Candida glabrata and the emerging pathogen Candida auris. These platforms have addressed longstanding challenges of low transformation efficiencies, limited selectable markers and diploid genome architecture. They permit gene deletion, insertion and allelic replacement with unprecedented speed and reliability. Innovative plasmid-borne systems, autonomously replicating sequences and transient delivery of RNA–protein complexes have expanded the genetic toolkit for non-albicans species and accelerated the dissection of pathogenicity factors. Collectively, these approaches have laid the groundwork for large-scale functional genomics, the construction of mutant libraries and the identification of novel antifungal targets with global clinical relevance.

Research from Nature Portfolio

Seminal work has described a robust CRISPR-Cas9 system in Candida glabrata that overcomes the species’ intrinsic barriers to genetic manipulation. This strategy utilises a strain engineered to constitutively express Cas9 alongside an online programme to design highly efficient guide RNAs. Combined with molecular assays for mutant verification and in vivo infection models in Drosophila, researchers have demonstrated the involvement of specific genes in host colonisation and virulence. The system’s high editing efficiency and minimal off-target effects have set a new standard for systematic loss-of-function analysis in non-albicans yeasts and provided a blueprint for adapting CRISPR-based tools to other diploid pathogens.

Genetic Manipulation of Human Fungal Pathogens publication trend

The graph below shows the total number of articles in genetic manipulation of human fungal pathogens across all publications each year (not limited to Nature Index journals).

Technical terms

CRISPR-Cas9: A genome-editing tool comprising a guide RNA that directs the Cas9 nuclease to a specific DNA sequence to induce a double-strand break.

Shuttle vector: A plasmid engineered to replicate in two different organisms, facilitating cloning in bacteria and subsequent transformation into a target fungal species.

Autonomously replicating sequence (ARS): A DNA element that enables extrachromosomal plasmid replication in yeast without integration into the host genome.

Ribonucleoprotein complex (RNP): A pre-assembled combination of Cas9 protein and guide RNA used for transient genome editing without genomic incorporation of editing constructs.

Homologous recombination: A cellular repair pathway that uses an exogenous DNA template with sequence similarity to repair double-strand breaks, enabling precise gene edits.

References

1. Development of a Shuttle Vector That Transforms at High Frequency for the Emerging Human Fungal Pathogen: Candida auris. Journal of Fungi (2024).
2. Use of RNA-Protein Complexes for Genome Editing in Non-albicans Candida Species. mSphere (2017).
3. Genome engineering in the yeast pathogen Candida glabrata using the CRISPR-Cas9 system. Scientific Reports (2016).