Browsing by Author "Thornton, Christina"

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    Phenotypic and Genotypic Characterization of the Streptococci Isolated from Sputum of Adult Cystic Fibrosis Patients
    (2013-08-27) Thornton, Christina; Surette, Michael; Storey, Douglas
    The cystic fibrosis (CF) lung microbiome is a complex polymicrobial community with numerous organisms involved in disease progression. The non-pyogenic streptococci, such as the Streptococcus milleri group, have been implicated in CF disease progression and pathogenesis, but other members are less understood as there has been limited study. The objectives of this study were to characterize a collection of diverse non-pyogenic streptococci and investigate their pathogenic potential and interactions with principal CF pathogens. Genotypic analysis fully characterized 100 unique isolates from CF sputum that did not identify with known species of streptococci by partial 16S rRNA sequencing. To examine these unique isolates, I did extensive genotypic characterization with cpn60, sodA, rpoB and ddl in addition to full 16S rRNA sequencing. 48 isolates identified as novel streptococci isolates, of which 20 of the novel presented with divergent multiple 16S rRNA gene sequences. Phenotypic characterization was expanded to a larger collection of 297 non-pyogenic streptococci isolates with profiling done by several assays. I observed poor correlation between genotypic methods and there was high phenotypic heterogeneity, particularly with regards to increased β-hemolysis under anaerobic condition on human blood as compared to standard conditions. Due to chronic antibiotic use by CF patients, the assessment of the streptococci antibiotic resistome from 459 isolates was done against nine antibiotics relevant in CF treatment. High antibiotic resistance was observed against the macrolides azithromycin (56.4%) and erythromycin (51.6%). Investigation of the molecular mechanisms of macrolide resistance yielded an unexpected result in that nearly half the isolates were found to have point mutations at position 2058 or 2059 of the 23S ribosomal subunit, an uncommon molecular mechanism of resistance within the streptococci. The synergistic interactions of the oropharyngeal flora (OF) with P. aeruginosa were investigated with the construction of transposon mutant libraries in two of the synergen isolates. Mutants were obtained that displayed attenuation or up-regulation of these synergistic interactions and will be used for further study and identification. The work presented here provides the framework to elucidate the role of the diverse streptococci in CF disease progression.