Investigating Coccidioides specific pH response and targeted gene deletion using CRISPR-Cas9

Miller, Karis Jolee (2021) Investigating Coccidioides specific pH response and targeted gene deletion using CRISPR-Cas9. Masters thesis, Northern Arizona University.

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Abstract

Valley fever is caused by the fungal pathogen, Coccidioides, which has two known species, C. immitis and C. posadasii. This xerotolerant fungus is associated with alkaline soils in arid to semi-arid regions of the Americas that have elevated average temperatures and low annual rainfall. Due to the soil being highly alkaline, Coccidioides likely utilizes intracellular signaling mechanisms to survive these conditions. In this study, we describe the first investigation into the pH sensing mechanism in Coccidioides, the pH-responsive Pal/RIM pathway. We tested wild type Coccidioides growth in varying pH environments using both liquid and solid media, as well as generating a PacC knock-out strain using the CRISPR-Cas9 method. RNA expression data was also collected from the varying pH media to compare the Pal pathway transcription factor, PacC, activity. Our study establishes that wild type Coccidioides grows on media ranging from pH 7-9. In vitro-assembled CRISPR/Cas9 was used to create a PacC knockout to investigate functional effects upon loss of PacC function. We were able to successfully delete the PacC transcription factor utilizing our CRISPR-Cas9 method. This study provides novel insight into how this organism utilizes a highly conserved fungal pH sensing pathway and will lead to future insight on the pathogenesis and lifecycle of Coccidioides.

Item Type:	Thesis (Masters)
Publisher’s Statement:	© Copyright is held by the author. Digital access to this material is made possible by the Cline Library, Northern Arizona University. Further transmission, reproduction or presentation of protected items is prohibited except with permission of the author.
Keywords:	Valley feaver, Fungal infections, Concidicides, pH, Pathogenesis
Subjects:	Q Science > QP Physiology
MeSH Subjects:	C Diseases > C08 Respiratory Tract Diseases
NAU Depositing Author Academic Status:	Student
Department/Unit:	Graduate College > Theses and Dissertations College of the Environment, Forestry, and Natural Sciences > Biological Sciences
Date Deposited:	05 Jul 2022 21:46
Last Modified:	05 Jul 2022 21:46
URI:	https://openknowledge.nau.edu/id/eprint/5800

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