Date of Award

Spring 5-6-2019

Document Type

Thesis

Degree Name

Bachelor of Arts

Department

Biochemistry & Molecular Biology

First Advisor

Joan Slonczewski

Second Advisor

Karen Hicks

Third Advisor

Chris Bickford

Abstract

Haloarchaea are polyextremophiles capable of surviving a wide range of conditions inhibitory to most other microbial life. These conditions include, but are not limited to, salt stress, relatively high levels of gamma and UV irradiation, temperature stress, pH stress, and desiccation. The hardiness of the halophiles makes them important model organisms for the astrobiological community, and the feasibility of microbial life on Mars has been brought into question. The purpose of this experiment is to assess haloarchaeal adaptation to iron and acid stress in order to model how these microbes may adapt to some stressors that would have been found in the oxidizing, iron-rich, acidic brine pools on the surface of Mars. We conducted a laboratory evolution project in which we evolved Halobacterium sp. NRC-1 for 500 generations in a set of four conditions, with four evolving populations per condition. The four conditions correspond to a neutral pH control, mild acid stress (pH 6.3), iron and acid stress (600 μM ferrous sulfate, pH 6.3), and an iron stress condition (600 μM ferrous sulfate). 16 clones from generation 500 of evolution were isolated, analyzed for phenotypic changes as evidence for mutation, and their DNA was sequenced using Illumina Next-Generation sequencing. We found generalized loss of gas vesicles and motility, coupled with a striking acid-adapted phenotype in acid-evolved strain J3-1. Genotypic analysis of all 16 clones revealed 378 mutations, with patterns of high variability due to ISH element activity, large deletions, and loss of function mutations in proton pumps or transporters in multiple strains.

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