Salmonella is often associated with retail poultry products. Our research evaluated the effect of temperature on the biofilm formation by Salmonella spp. and the efficacy of essential oils in controlling these biofilms on stainless steel surfaces. The sublethal concentrations of thyme, oregano and carvacrol at 0.006-0.012% suppressed biofilm formation by Salmonella spp. while concentrations at 0.05-0.1% reduced the biofilms of a three-strain mixture of Salmonella spp. by 7 logs. Carvacrol was evaluated in combination with lauric arginate for controlling the three-strain mixture of Salmonella spp. in ground turkey containing 1%, 7% or 15% fat. Higher concentrations of carvacrol (1%) or lauric arginate (2000 ppm) when applied individually did not reduce Salmonella counts in ground turkey containing 7% fat. The combined mixture of carvacrol and lauric arginate at these higher concentrations was found to be synergistic in reducing the Salmonella counts by 4 log CFU/g in ground turkey containing 7% fat.

The antimicrobial efficacy of carvacrol, trans-cinnamaldehyde, eugenol and thyme oil (0.5, 1, 2 and 5% v/v) was evaluated against Salmonella on turkey breast cutlets as 2 min dip treatments at 4°C. Carvacrol revealed maximum efficacy against Salmonella on turkey cutlets and was selected for 30s and 60s dip treatments at 4°C. Only 5% carvacrol was effective and exhibited 1.0 and 1.8 log CFU/g reduction of Salmonella with 30s and 60s dip treatments, respectively. However, carvacrol showed synergistic activity with carbon dioxide in modified atmosphere packaging (95% CO2/5% O2) against Salmonella and Campylobacter and caused 1.0-2.0 log CFU/g reductions with lower concentrations (0.25, 0.5 and 1.0%). This combination reduced the growth of lactic acid bacteria. In the third experiment, efficacy of lauric arginate (200 and 400ppm) against C. jejuni was tested on chicken breast fillets. Both these concentrations significantly reduced C. jejuni on chicken fillets (1.0-1.5 log CFU/g) at 4°C.

Aims There are few proven therapies that can protect against the inevitable ischaemia reperfusion injury (IRI) that occurs during renal transplantation. IRI increases the likelihood of delayed graft function (DGF), which negatively impacts on the long-term survival of a transplanted kidney. One enzyme of interest, heme oxygenase-1 (HO-1), degrades heme and protects against the oxidative stress that occurs secondary to IRI. Clinical renal recipients with higher HO-1 levels have improved graft function post transplant. Heme arginate (HA), a form of hemin, which has been used to treat porphyria for over 30 years, has repeatedly been shown to induce HO-1 in in vivo and in vitro macrophages. It is one of the few HO-1 inducers approved for clinical use and healthy volunteer studies confirmed that HA could also safely induce HO-1 in humans. Prior to the formal start of the MD, the University of Edinburgh successfully applied to NHS Blood and Transplant for funding to investigate whether giving HA to recipients of deceased donor renal grafts prior to transplant could upregulate HO-1 and whether this had any effect on the function and health of the grafts. This MD aims to explain the background behind the proposed study, the process of study approval, planning and trial logistics and protocol. This thesis then describes the methods of sample analysis, the results and future directions for the HOT (Heme Oxygenase-1 in renal Transplantation) study. Methods The HOT study planning and approval process took eight months and the first participant was randomised in January 2012. The study was sponsored by ACCORD, a joint company from University of Edinburgh and NHS Lothian, and recruited patients from the Edinburgh Royal Infirmary Transplant Unit. The protocol was followed to ensure that 40 recipients were randomised blind to either active (two doses 3mg kg-1 HA: pre-operatively, day 2) or placebo (NaCl: same schedule). To ensure that the primary outcome was fulfilled, recipient blood was taken daily for peripheral blood mononuclear cells (PBMC) extraction. After further blinding steps, the PBMCs were analysed for HO-1 protein and mRNA. The secondary outcome measures involved collecting urine for analysis of urinary biomarkers (KIM-1 and NGAL), taking renal graft biopsies pre-op and day 5 for renal HO-1 analysis and collecting renal function data. DGF was calculated daily. To ensure that all adverse event data was captured, the recipients were closely reviewed for 7 days and their renal function was monitored for 90 days. Results The final participant was recruited in May 2013 within the predicted timescale and to budget. This participant completed follow-up in August 2013. Of the 40 participants, three received the infusion but did not receive a transplant and therefore could not give primary outcome data. The remaining 37 did and this was analysed. Adverse events were equivalent between groups and there were no adverse reactions to HA. HA upregulated PBMC HO-1 protein at 24 hours compared to placebo: HA 11.1ng/ml (1.0- 37.0) vs. placebo 0.14ng/ml (-0.7- 0.3)(p= < 0.0001). PBMC HO-1 mRNA was also increased: HA 2.73 fold (1.8- 3.2) vs. placebo 1.41 fold (1.2- 2.2) (p=0.02). HA increased HO-1 protein immunopositivity in day 5 renal tissue compared with placebo: HA 0.21 (-24- 0.7) vs. placebo -0.03 (-76- 0.15) (p=0.02) and the percentage of HO-1 positive renal macrophages also increased: HA 50.8 cells per HPF (40.0- 59.8) vs. placebo 22.3 (0- 34.8) (p=0.012). Renal HO-1 mRNA was also increased in HA group: 2.02 (0.20- 4.03) fold increase compared to 1.68 (0.75- 10.39) fold in the placebo group but it was not significant (p= 0.451). Urinary biomarkers were reduced after HA but not significantly so. Histological injury and DGF rates were similar between the groups. Conclusion HA is safe and effective in renal transplant recipients as reported in this phase II, randomised, placebo controlled, blinded, single-centre study. The primary outcome was achieved and demonstrated for the first time that HA induces HO-1 in peripheral and renal macrophages in kidney transplant recipients. There was also evidence that HA increased HO-1 expression in renal tissue. There was no evidence that HA improved renal function or reduced injury as seen in animal models but it is recognised that the sample size was small and the study was not powered to these endpoints. Larger studies are planned to determine the impact of HO-1 upregulation on clinical outcomes and evaluate the benefit to patients at risk of IRI. The plans for HOT2 are expanded in this thesis.

Colloidal dispersions such as oil-in-water or water-in-oil emulsions have found widespread use in the food industry. Oil-in-water emulsions consist of three principal components i.e. oil dispersed in the form of droplets, water surrounding the droplets as the continuous phase, and emulsifiers comprising the interface. Because of the complicated interaction among components, it is often difficult to predict the physicochemical properties and final functionalities of emulsions. Nevertheless, the structural and functional features of emulsions allow scientists to create many unique emulsions that may serve as suitable carriers for lipophilic functional compounds. These functional compounds may include antioxidants, flavors, colors and antimicrobials, the latter which is the principal topic of this thesis. Incorporation of food antimicrobials in emulsions could create value-added emulsions that may improve the safety and quality of a variety of foods, but to date, few systematic studies on their formulation have been reported. The objective of this thesis was therefore to formulate food emulsions that are physicochemically stable and able to deliver antimicrobial compounds to microbial target sites. Two antimicrobial agents, N-α-lauroyl-L-arginine ethyl ester monohydrochloride, (LAE) and eugenol were used as model compound to be incorporated into the colloidal food dispersion. The two antimicrobials were selected because they are either amphiphilic (LAE) or predominantly lipophilic (eugenol). When emulsions were formulated with eugenol, an essential oil component, it was found that O/W emulsions were only stable when emulsions were formulated with other lipids (hexadecane, dodecane, tetradecane, and corn oil). Above a critical loading of the carrier lipid with eugenol, Ostwald’s ripening led to rapid destabilization while above this critical loading concentration, the ripening rate was greatly reduced and depended on type of carrier lipid and concentration of eugenol. Alternatively, when emulsions were formulated with LAE as emulsifier, results indicated that emulsions were not stable to aggregation and coalescence. Consequently, LAE had to be combined with a nonionic surfactant (Tween 20) to improve the emulsion stability. Higher Tween20 composition led to more stable emulsions droplets. Both systems (emulsions with either eugenol or LAE) had high antimicrobial efficacies and were able to completely inhibit microbial growth at concentrations that depended on the type of microorganisms and formulation of the emulsions. Generally, eugenol were able to more effectively inhibit the growth of E.coli O157:H7 while LAE containing emulsions were more effective against L.monocytogenes. Finally, a food emulsion was formulated that contained both antimicrobial agents; eugenol in the lipid phase and LAE in the droplet interface. Interestingly, stability of these emulsions depended both on the LAE and eugenol loading. The antimicrobial activity in this double antimicrobial emulsion was high but was principally influenced by the interfacial formulation that is the ratio of LAE to Tween 20. The combined emulsion similar to the LAE stabilized emulsion more effectively inhibited growth of L.monocytogenes.

The available antiretroviral compounds can effectively suppress the replication of HIV-1 and block the disease progression. However it is impossible to eradicate the virus from the organism as the HIV-1 integrated in the genome is not affected by the existing anti-HIV-1 drugs. Therefore, new latency reversing agents are being actively developed as part of "shock and kill" therapy to reactivate the provirus and clear the reservoir. Normosang (heme arginate; HA) is a human hemin- containing compound used to treat acute porphyria. Heme is physiologically catabolised by heme oxygenases to form iron (Fe2+ ), carbon monoxide (CO) and biliverdin that is further converted to bilirubin by biliverdin reductase. In this study, we have demonstrated that HA inhibited HIV-1 replication during the acute infection, which was accompanied by the inhibition of reverse transcription. On the other hand, HA synergised with phorbol myristyl acetate (PMA) and reactivated the HIV-1 provirus in ACH-2 cells and the HIV-1 "mini-virus" in Jurkat cell clones A2 and H12. HIV-1 ''mini-virus'' was reactivated also by HA-alone. Further, we have studied the effects of heme degradation products on latent HIV-1 reactivation when added individually. We employed addition of ascorbate to generate Fe2+ , resulting in an increased...

The available antiretroviral compounds can effectively suppress the replication of HIV-1 and block the disease progression. However it is impossible to eradicate the virus from the organism as the HIV-1 integrated in the genome is not affected by the existing anti-HIV-1 drugs. Therefore, new latency reversing agents are being actively developed as part of "shock and kill" therapy to reactivate the provirus and clear the reservoir. Normosang (heme arginate; HA) is a human hemin- containing compound used to treat acute porphyria. Heme is physiologically catabolised by heme oxygenases to form iron (Fe2+ ), carbon monoxide (CO) and biliverdin that is further converted to bilirubin by biliverdin reductase. In this study, we have demonstrated that HA inhibited HIV-1 replication during the acute infection, which was accompanied by the inhibition of reverse transcription. On the other hand, HA synergised with phorbol myristyl acetate (PMA) and reactivated the HIV-1 provirus in ACH-2 cells and the HIV-1 "mini-virus" in Jurkat cell clones A2 and H12. HIV-1 ''mini-virus'' was reactivated also by HA-alone. Further, we have studied the effects of heme degradation products on latent HIV-1 reactivation when added individually. We employed addition of ascorbate to generate Fe2+ , resulting in an increased...