Antimicrobial-resistant bacteria are a major source of ailment in modern-day nosocomial settings, with numerous risks including leading to possible further drug resistance or spreading to those who cannot fight off the infection due to immune suppression or dysfunction. Previous work in our laboratory has determined that Klebsiella pneumoniae possesses inhibitory effects on the growth of a variety of bacteria that contain antimicrobial-resistant properties in the Enterobacteriaceae family, a major source of nosocomial antimicrobial-resistance. This novel property of K. pneumoniae inhibits the growth of Citrobacter freundii, Enterobacter aerogenes, and Enterobacter cloacae through an anti-microbial protein. The antimicrobial protein secreted from K. pneumoniae has been shown to reduce the density and growth of C. freundii, E. aerogenes, and E. cloacae in both biofilm and planktonic forms. The work performed in this thesis has shown that the antimicrobial protein is plasmid mediated by introducing a transposon (Tn5) to the plasmid to provide resistant selection and a possible way to create a mutant knockout to find the exact location of the gene in the plasmid. Upon transposon mutagenesis of the plasmid, it was electroporated into Rec- E. coli. The E. coli were then able to produce the antimicrobial protein allowing the formation of zones of inhibition when screened on C. freundii, E. aerogenes, and E. cloacae lawns. Upon confirmation that the plasmid mediates the anti- microbial protein, the plasmid was sent for sequencing to further characterize the gene responsible for coding the anti-microbial protein. This novel antimicrobial protein has high sequence similarity to bacteriocins and, thus far, is a novel and uncharacterized protein of plasmid origin found in only in this particular strain of K. pneumoniae. Further research involving this new bacteriocin could aid in the development of treatments for the highly drug resistant Enterobacteriaceae family members.

Firstly, the aims of the study were to investigate the influences of various factors, sex differences, Cytomegalovirus/Epstein-Barr virus (CMV/EBV) serostatus and vitamin D concentrations on respiratory illness incidence and immune function during the winter months in a student cohort of endurance athletes. In Chapter 3, the findings of the study concur with recent reports of illness incidence at major competitive games which indicate that female athletes may be more susceptible than their male counterparts to upper respiratory tract illness (URTI) symptoms and that lower oral-respiratory mucosal immunity may, in part, account for this. It was also found that previous coinfection with CMV and EBV might promote protective immune surveillance to lower the risk of URTI. In addition, it can be concluded that athletes with low plasma vitamin D concentrations may have a higher risk of URTI and suffer more severe symptoms when URTI is present. This may be due to impaired mucosal and systemic immunity as secretory immunoglobulin A (SIgA) secretion, cathelicidin levels and antigen-stimulated pro-inflammatory cytokine production appear to be increased by vitamin D-dependent mechanisms. A series of follow-up studies were also conducted to examine the effect of vitamin D on mucosal and systemic immunity in athletes. In Chapter 4, it was reported that the influence of vitamin D on circulating cytokines might be different in athletes compared with non-athletes and that both pro-inflammatory and anti-inflammatory cytokine production by multi-antigen stimulated whole blood culture were not influenced by 1,25-dihydroxy vitamin D (1, 25(OH)2D) iconcentrations within the normal healthy range. In Chapter 5, it was found that 5000 IU of vitamin D3 supplementation daily appears to have a beneficial effect in up-regulating the expression of SIgA and cathelicidin in athletes during a winter training period. Nevertheless, the findings reported in Chapter 6 showed that there were no significant effects of vitamin D status and a 4-week period of daily high does vitamin D3 supplementation on salivary antimicrobial protein (AMP) responses to prolonged exercise. In conclusion, a series of studies in this thesis have demonstrated the influence of various factors (sex differences, CMV/EBV serostatus and vitamin D concentrations) on susceptibility to URTI among athletes. Moreover, it was suggested that vitamin D3 supplementation could have a positive effect on immune function and lead to decreased incidence of respiratory infections.

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