Date of Award

8-2021

Document Type

Thesis

Degree Name

Master of Science in Biology

Department

Biology

First Reader/Committee Chair

Dr. Paul M. Orwin

Abstract

Approximately 10% of all bacterial genomes sequenced thus far contain a secondary replicon. This considerable genetic reservoir contains many potentially mobilizable elements, allowing for the formation of many unique secondary replicons. This property of bacterial populations vastly increases the genomic diversity available to species that effectively take up and maintain these replicons. Members of the genus Variovorax have extensive heterogeneity in genome architecture, including sequenced isolates containing plasmids, megaplasmids, and chromids. Using available Illumina data on the NCBI database, we have completed these assemblies using 3rd generation sequencing methods on 17 members of this genus. We have sequenced, assembled, and evaluated these now complete Variovorax genomes to examine the diversity of elements. From these assemblies 9 chromids, 7 megaplasmids, 2 plasmids, and an integrated megaplasmid were identified using genomic frequency characteristics. We observed in the genomic characteristics data that there is a pattern of evolution where a plasmid is picked up by the organism and over evolutionary time, there is expansion of the replicon in size through acquisition of mobile elements and interreplicon transfer. As this process goes on, the secondary replicons’ genomic frequency characteristics regress toward the chromosome. Given the evolutionary pattern seen, we were interested to find out if the high levels of heterogeneity present in Variovorax are due to factors controlling secondary replicon maintenance present within the primary chromosome. Using two strains of Variovorax paradoxus strains, VAI-C (multipartite genome) and EPS (single chromosome) and two plasmids pRU1105 and pBBR5pemIKpBAD (which contains a Toxin-Antitoxin system), were used to evaluate the maintenance of plasmids in the absence of selection. Surprisingly, the Toxin-Antitoxin system had a deleterious effect and showed no additional phenotype in either organism.

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