Identification of Genetic, Phenotypic, and Transcriptomic Differences in Virulence-attenuated Strains Relative to Fully Virulent Strains of Listeria monocytogenes.
Cook, Peter W.
Listeria monocytogenes is a human and animal pathogen capable of causing disease outcomes from gastroenteritis to death. In addition, it is resistant to several common food environmental stressors, and capable of growth at low temperatures. The studies presented herein attempt to differentiate strains of L. monocytogenes using genomic, phenotypic, and transcriptomic information. Chapter 1 first describes aspects of L. monocytogenes that are integral in the understanding of virulence and pathogenicity, differentiation of strains, and provides an understanding the mechanisms of evolution that may occur in the food processing environment. Chapther 2 describes the first study that used a set of virulence-attenuated strains isolated from food-associated environments that were identified with known naturally occurring premature stop codons in internalin A (inlA), an important virulence gene. Several different mutation types have been identified both across and within the lineages of L. monocytogenes. These lineages are genetically distinct suggesting the premature stop codon in inlA has arisen through either a common mechanism or has resulted from convergent mutations and could provide independent clonal complexes, or clonally related organism, with fitness increases through the loss of the primary interaction with the host. The set of strains selected were subjected to whole genome sequencing to identify any additional loss-of-function mutations, or other changes to the genome that could provide additional support for the loss of the primary host pathogenicity route. Chapter 3 describes the second study that selected two virulence-attenuated strains and two fully virulent strains from the nearest neighbor clonal complex to compare the transcriptional activity of these strains to one another. The strains were selected to represent the virulence-attenuated subpopulation or the fully virulent subpopulation based on the results of the first study including their increased biomass formation or swarming motility, having the two most common inlA PMSC types, and being isolated from food processing facilities. The transcriptomic experiment was performed by comparing each of the strains transcriptome on stainless steel to itself under planktonic conditions. The temperature, moisture, and sheer force over the coupons were included to be sure the comparisons were between planktonic and fully adherent L. monocytogenes. We were able to identify several gene categories across the clonal complexes that contained genes potentially non-functional due to premature stop codons in the whole genome sequencing data, and the transcriptomic data suggests broad differences in the number and content of differentially expressed genes between the fully virulent and virulence-attenuated strains.