Date of Award

Winter 2023

Project Type

Dissertation

Program or Major

Microbiology

Degree Name

Doctor of Philosophy

First Advisor

Louis S Tisa

Second Advisor

Julie G Bryce

Third Advisor

Kelley Thomas

Abstract

Introduction: Blastococcus saxobsidens is an endolithic bacterium associated with biodegradation of stone monuments across the Mediterranean basin. Recently, strains of the bacterium have been isolated from high altitude deserts in Chile, volcanic glass in Iceland, and from masonry of several cultural heritage sites across Europe and the Americas. Blastococcus saxobsidens global dispersion has been linked to patterns of wind distributed dust particles. The physiology that this bacterium uses to colonize a wide range of rock types in such distinct locations has not been previously explored however. This project has established that B. saxobsidens colonizes and respires a variety of chemically distinct rock types. An RNAseq approach was used to capture their transcriptome during rock colonization. The resulting profiles, which represent snapshots of global gene expression, were compared to the profile of B. saxobsidens cells grown in liquid Czapek broth without rocks. The differential gene expression from these profiles distinguished lithotrophy and other rock-associated genes which were highly expressed in cells grown on rocks. Aims: This project focused on four primary aims: 1) Show that B. saxobsidens cells will colonize and respire rock types beyond the sandstones they were initially isolated from. 2) Identify the rock associated genes and operons that are expressed while B. saxobsidens cells colonize and respire rock. 3) Assess differential gene expression of B. saxobsidens transcriptomes from growth on three different rock types. 4) Establish a B. saxobsidens RNAseq informed metabolic-network resource and compare rock-associated genes across a Geodermatophilaceae STAG phylogeny. Results: The tetrazolium bioassay indicated that B. saxobsidens respired 7 of the 14 different rock types tested to varying degrees. Cell growth led to decreased sample pH which indicates acid excretion is part of the rock degradation cascade which confirmed previous observations. No single or synergized elemental composition was identified as a driving force for rock respiration. RNAseq differential gene expression analysis determined that B. saxobsidens transcriptome reflected specific interactions with the rock substrate it was exposed to. The adaptations were relatively slow and minimal differences were seen at 14 days of incubation between different rocks. By the 28th day of incubation on rock, 3,532 genes, representing 70% of the coding genome, were differentially expressed across the 3 different rock types. Each biofilm incubated for 28-days on rocks differentially uniquely expressed: 476 (red sandstone), 532 (white sandstone), and 361 (biotite rich gneiss) genes when compared to cells grown in the absence of rocks. The transcriptomes were similarly split between up- and down- regulated genes which represented transitions in metabolism rather than metabolic decline due to starvation. A BLAST2GO metabolic-pathway network was created as a resource to further understand the diversity of metabolic processes taking place during B. saxobsidens rock respiration and colonization. With it, lithotrophy, chemoautotrophy, chemotaxis, motility, and adhesion phenotypes were identified from the RNAseq expression data. This confirmed that the cells incubated on rocks for 14 and 28 days heavily invested in chemoautotrophy & lithotrophy when compared to cells grown in Czapek complete medium. The expressed rock associated operons and genes from B. saxobsidens were reviewed across a STAG phylogenetic tree. A pattern of similarity emerged between Blastococcus and Modestobacter. The similarities between the two accessory genomes are likely attributable to the endolithic nature of the two genera.

Download

Share

COinS