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Generation of Fluorescent Protein Fusions in Candida Species
Published on: March 4, 2017
Genetic Modification of Closely Related Candida Species
Eugenio Mancera1,2, Corey Frazer3, Allison M Porman3
1Departamento de Ingeniería Genética, Centro de Investigación y de Estudios Avanzados del Instituto Politécnico Nacional, Unidad Irapuato, Irapuato, Mexico.
Abstract:
Species from the genus Candida are among the most important human fungal pathogens. Several of them are frequent commensals of the human microbiota but are also able to cause a variety of opportunistic infections, especially when the human host becomes immunocompromised. By far, most of the research to understand the molecular underpinnings of the pathogenesis of these species has focused on Candida albicans, the most virulent member of the genus. However, epidemiological data indicates that related Candida species are also clinically important. Here, we describe the generation of a set of strains and plasmids to genetically modify C. dubliniensis and C. tropicalis, the two pathogenic species most closely related to C. albicans. C. dubliniensis is an ideal model to understand C. albicans pathogenesis since it is the closest species to C. albicans but considerably less virulent. On the other hand, C. tropicalis is ranked among the four most common causes of infections by Candida species. Given that C. dubliniensis and C. tropicalis are obligate diploids with no known conventional sexual cycle, we generated strains that are auxotrophic for at least two amino acids which allows the tandem deletion of both alleles of a gene by complementing the two auxotrophies. The strains were generated in two different genetic backgrounds for each species - one for which the genomic sequence is available and a second clinically important one. In addition, we have adapted plasmids developed to delete genes and epitope/fluorophore tag proteins in C. albicans so that they can be employed in C. tropicalis. The tools generated here allow for efficient genetic modification of C. dubliniensis and C. tropicalis, and thus facilitate the study of the molecular basis of pathogenesis in these medically relevant fungi.
Insights
New genetic tools enable the study of Candida dubliniensis and Candida tropicalis pathogenesis. These resources facilitate research into these important human fungal pathogens, complementing existing Candida albicans research.
Area of Science:
- Medical Mycology
- Molecular Biology
- Fungal Pathogenesis
Background:
- Species within the genus *Candida* are significant human fungal pathogens, often causing opportunistic infections in immunocompromised individuals.
- While *Candida albicans* is extensively studied, related species like *C. dubliniensis* and *C. tropicalis* are also clinically relevant.
- *C. dubliniensis* serves as a less virulent model for *C. albicans* pathogenesis, while *C. tropicalis* is a leading cause of candidiasis.
Purpose of the Study:
- To develop novel genetic tools for the genetic modification of *C. dubliniensis* and *C. tropicalis*.
- To facilitate comparative studies of pathogenesis between *C. albicans*, *C. dubliniensis*, and *C. tropicalis*.
- To enable detailed molecular investigations into the mechanisms underlying candidiasis caused by these species.
Main Methods:
- Generation of auxotrophic strains for *C. dubliniensis* and *C. tropicalis* to enable tandem gene allele deletion.
- Creation of strains in two distinct genetic backgrounds for each species, including clinically relevant isolates.
- Adaptation of existing gene deletion and protein tagging plasmids from *C. albicans* for use in *C. tropicalis*.
Main Results:
- A comprehensive set of strains and plasmids for efficient genetic manipulation of *C. dubliniensis* and *C. tropicalis* has been successfully generated.
- These tools support the study of obligate diploid fungi lacking a conventional sexual cycle.
- The developed resources are applicable across different genetic backgrounds, enhancing their utility.
Conclusions:
- The newly developed genetic tools significantly advance the capacity for molecular research in *C. dubliniensis* and *C. tropicalis*.
- These resources will accelerate the understanding of pathogenic mechanisms in these medically important *Candida* species.
- This work provides a foundation for comparative genomics and functional studies, crucial for combating candidiasis.
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