Optimizing in vitro spherulation cues in the fungal pathogen Coccidioides
ABSTRACT Coccidioides spp. are part of a group of thermally dimorphic fungal pathogens, which grow as filamentous cells (hyphae) in the soil and transform to a different morphology upon inhalation into the host. The Coccidioides host form, the spherule, is unique and highly undercharacterized due to...
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American Society for Microbiology
2025-01-01
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| Series: | mSphere |
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| Online Access: | https://journals.asm.org/doi/10.1128/msphere.00679-24 |
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| author | Christina M. Homer Elena Ochoa Mark Voorhies Anita Sil |
| author_facet | Christina M. Homer Elena Ochoa Mark Voorhies Anita Sil |
| author_sort | Christina M. Homer |
| collection | DOAJ |
| description | ABSTRACT Coccidioides spp. are part of a group of thermally dimorphic fungal pathogens, which grow as filamentous cells (hyphae) in the soil and transform to a different morphology upon inhalation into the host. The Coccidioides host form, the spherule, is unique and highly undercharacterized due to both technical and biocontainment challenges. Each spherule arises from an environmental spore (arthroconidium), matures, and develops hundreds of internal endospores, which are released from the spherule upon rupture. Each endospore can then go on to form another spherule in a cycle called spherulation. One of the foremost technical challenges has been reliably growing spherules in culture without the formation of contaminating hyphae and consistently inducing endospore release from spherules. Here, we present optimization of in vitro spherule growth and endospore release, by closely controlling starting cell density in the culture, using freshly harvested arthroconidia, and decreasing the concentration of multiple salts in spherulation media. We developed a minimal medium to test spherule growth on various carbon and nitrogen sources. We defined a critical role for the dispersant Tamol in both early spherule formation and prevention of the accumulation of a visible film around spherules. Finally, we examined how the conditions under which arthroconidia are generated influence their transcriptome and subsequent development into spherules, demonstrating that this is an important variable to control when designing spherulation experiments. Together, our data reveal multiple strategies to optimize in vitro spherulation growth, enabling characterization of this virulence-relevant morphology.IMPORTANCECoccidioides spp. are thermally dimorphic fungal pathogens found in the Southwest United States, Mexico, Central America, and South America. Coccidioides can infect both immunocompetent and immunocompromised people and can cause a devastating disseminated infection, including meningitis, with 30% mortality despite all currently available treatments. In this work, we tackle one of the current largest technical barriers to studying the fungus Coccidioides: reliably growing its host form in vitro. Our work is impactful because we have created a set of foundational tools for the burgeoning field of Coccidioides pathogenesis research. We have carefully optimized conditions that allow the development of Coccidioides in vitro into its pathogenic form. This work will open up many lines of investigation into the molecules that underlie Coccidioides pathogenesis. |
| format | Article |
| id | doaj-art-4e09a3ae90cc4fa0971f0b7f2cb5f0df |
| institution | Kabale University |
| issn | 2379-5042 |
| language | English |
| publishDate | 2025-01-01 |
| publisher | American Society for Microbiology |
| record_format | Article |
| series | mSphere |
| spelling | doaj-art-4e09a3ae90cc4fa0971f0b7f2cb5f0df2025-01-28T14:00:56ZengAmerican Society for MicrobiologymSphere2379-50422025-01-0110110.1128/msphere.00679-24Optimizing in vitro spherulation cues in the fungal pathogen CoccidioidesChristina M. Homer0Elena Ochoa1Mark Voorhies2Anita Sil3Division of Infectious Diseases, University of California San Francisco, San Francisco, California, USADepartment of Microbiology and Immunology, University of California San Francisco, San Francisco, California, USADepartment of Microbiology and Immunology, University of California San Francisco, San Francisco, California, USADepartment of Microbiology and Immunology, University of California San Francisco, San Francisco, California, USAABSTRACT Coccidioides spp. are part of a group of thermally dimorphic fungal pathogens, which grow as filamentous cells (hyphae) in the soil and transform to a different morphology upon inhalation into the host. The Coccidioides host form, the spherule, is unique and highly undercharacterized due to both technical and biocontainment challenges. Each spherule arises from an environmental spore (arthroconidium), matures, and develops hundreds of internal endospores, which are released from the spherule upon rupture. Each endospore can then go on to form another spherule in a cycle called spherulation. One of the foremost technical challenges has been reliably growing spherules in culture without the formation of contaminating hyphae and consistently inducing endospore release from spherules. Here, we present optimization of in vitro spherule growth and endospore release, by closely controlling starting cell density in the culture, using freshly harvested arthroconidia, and decreasing the concentration of multiple salts in spherulation media. We developed a minimal medium to test spherule growth on various carbon and nitrogen sources. We defined a critical role for the dispersant Tamol in both early spherule formation and prevention of the accumulation of a visible film around spherules. Finally, we examined how the conditions under which arthroconidia are generated influence their transcriptome and subsequent development into spherules, demonstrating that this is an important variable to control when designing spherulation experiments. Together, our data reveal multiple strategies to optimize in vitro spherulation growth, enabling characterization of this virulence-relevant morphology.IMPORTANCECoccidioides spp. are thermally dimorphic fungal pathogens found in the Southwest United States, Mexico, Central America, and South America. Coccidioides can infect both immunocompetent and immunocompromised people and can cause a devastating disseminated infection, including meningitis, with 30% mortality despite all currently available treatments. In this work, we tackle one of the current largest technical barriers to studying the fungus Coccidioides: reliably growing its host form in vitro. Our work is impactful because we have created a set of foundational tools for the burgeoning field of Coccidioides pathogenesis research. We have carefully optimized conditions that allow the development of Coccidioides in vitro into its pathogenic form. This work will open up many lines of investigation into the molecules that underlie Coccidioides pathogenesis.https://journals.asm.org/doi/10.1128/msphere.00679-24Coccidioidesfungal pathogenesisarthroconidiamorphologydevelopment |
| spellingShingle | Christina M. Homer Elena Ochoa Mark Voorhies Anita Sil Optimizing in vitro spherulation cues in the fungal pathogen Coccidioides mSphere Coccidioides fungal pathogenesis arthroconidia morphology development |
| title | Optimizing in vitro spherulation cues in the fungal pathogen Coccidioides |
| title_full | Optimizing in vitro spherulation cues in the fungal pathogen Coccidioides |
| title_fullStr | Optimizing in vitro spherulation cues in the fungal pathogen Coccidioides |
| title_full_unstemmed | Optimizing in vitro spherulation cues in the fungal pathogen Coccidioides |
| title_short | Optimizing in vitro spherulation cues in the fungal pathogen Coccidioides |
| title_sort | optimizing in vitro spherulation cues in the fungal pathogen coccidioides |
| topic | Coccidioides fungal pathogenesis arthroconidia morphology development |
| url | https://journals.asm.org/doi/10.1128/msphere.00679-24 |
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