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- ItemAn In Vitro Model of Endemic Burkitt’s Lymphoma (eBL) Pathogenesis; Cooperation of Plasmodium Falciparum and Epstein Barr Virus in DNA Damage Mediated Via Activation Induced Cytidine Deaminase.(2012-06-15) Ayivor-Djanie, ReubenPlasmodium falciparum (P. falciparum) and Epstein-Barr virus (EBV) infections are contributors in the pathogenesis of endemic Burkitt’s lymphoma (eBL), although the precise mechanism of their synergy remains elusive. Reports suggest that the role of P. falciparum is indirect, creating a permissive environment for the outgrowth of EBV. EBV on the other hand invades and immortalizes lymphocytes in vitro and upregulates activation-induced cytidine deaminase (AID), a DNA repair enzyme responsible for diversifying the antibody repertoire and a potent mutagen capable of inducing the genetic damage characteristic of eBL. It is yet to be shown how exposure to P. falciparum affects the expression of AID in lymphocytes. The aim of this work was to investigate the possible direct role of P. falciparum in eBL lymphomagenesis by exploring parasite-lymphocyte interactions and AID expression after exposure to P. falciparum and/or EBV. Malaria positive slides were examined for parasite-lymphocyte interactions and primary tonsillar mononuclear cells (MNCs) were co-cultured with RBCs infected with up to 5% parasitemia of the 3D7 strain of P. falciparum. Geimsa stained thin smears were made from these co-cultures and examined for parasite-MNC interactions over a five day period. No direct parasite-MNC interaction was observed from all slides examined. The levels of AID mRNA in MNCs was measured by qPCR after in vitro exposure to P. falciparum and/or EBV, and in the presence or absence of 2µg/ml cyclosporine. P. falciparum induced up to a 6-fold increase in AID over unstimulated controls, EBV induced a 13-fold maximum increase, and both pathogens together induced up to a 22-fold increase in AID. With cyclosporine, AID mRNA levels in the P. falciparum stimulated cultures remained unchanged. EBV alone induced a 22- fold increase in AID and both pathogens together induced a 42-fold increase in AID. DNA damage was estimated by Comet Assay and quantified with an algorithm from the Comet Assay Project Lab (CASP). DNA comets revealed that P. falciparum induced moderate DNA damage in MNCs with up to 5.6% and 10% DNA in tails of comets with and without cyclosporine respectively. Cultures stimulated with EBV recorded DNA damage of up to 16% and 13% DNA in tails of comets with and without cyclosporine respectively; and both pathogens induced DNA damage with up to 11% and 16% DNA in tails of comets with and without cyclosporine respectively. The levels of DNA damage in these cells correlated with AID levels and demonstrate that P. falciparum plays a direct role in eBL pathogenesis, by inducing AID expression to levels similar to that expressed in BL cells and cooperating with EBV to induce abnormally high levels of AID and DNA damage.
- ItemThe kinetics of mycolactone in relation to the microbiological, clinical and immunological responses to antibiotic therapy for mycobacterium ulcerans disease.(2014-11-20) Sarfo, Fred StephenBackground: Mycobacterium ulcerans is the causative agent for a chronic necrotising skin infection called Buruli ulcer. Pathology of the disease is closely linked with the elaboration of a unique lipid toxin, mycolactone, which has potent cytotoxic and immunomodulatory properties. In this study, assays were developed to detect and quantify mycolactone concentrations in tissues during curative antibiotic therapy in mice and in humans to understand its dynamics in pathogenesis and to explore its potential as a biomarker for diagnosis and monitoring of patients with Buruli ulcer disease on antibiotic therapy. The currently recommended antibiotic regimen for the management of Buruli ulcer is a combination of daily intramuscular injections of streptomycin and oral rifampicin for 8 weeks (RS8). This regimen was compared with streptomycin/rifampicin for 2 weeks followed by clarithromycin/rifampicin (RS2RC6) for 6 weeks in patients to determine the clinical and bacteriological effectiveness in a pilot study. Methods: Biopsies were obtained from infected human skin tissues and BALB/c mouse hind footpads before, during and after 8-weeks of rifampicin-containing combination antibiotic therapy. Lipids were extracted from tissues using organic solvents, mycolactone concentrations were measured using a cytotoxicity assay and mass spectrometry. Trends in mycolactone concentrations and clinical, bacteriological and immuno-histopathological responses were determined. Concentrations of cytokines in supernates of whole blood assays in humans or murine splenocytes after stimulation with mycobacterial antigens/T-cell mitogens were measured using ELISA. iv Results: Eighty-three patients with confirmed Buruli ulcer were randomized to RS8 or RS2RC6 and monitored for recurrence free-healing. Bacterial load in tissue samples before and after treatment for 6 and 12 weeks was monitored in samples obtained by 4mm punch biopsy by semi-quantitative culture. There was no difference in using RS8 or RS2RC6 with respect to healing rate or the proportion healed in each group after 4, 8, 12, 16, 20, 24 and up to 52 weeks. The success rate was 93% in each group and there was no recurrence after 12-month follow-up. There was no difference in the number of bacteria cultured at the different time points for the two regimens. Mycolactone was detectable in 92% and 77% of human samples (n=80) using cytotoxicity assays and mass spectrometry respectively. Antibiotic therapy was associated with a decline in tissue concentration of mycolactone in both human and murine-infected tissues which was paralleled by resolution of clinical lesions, reductions in bacteriological counts and restoration of local and systemic immune responses. Discussions/Conclusions: This study shows that mycolactone concentrations in tissues is closely associated with the presence of M. ulcerans and provides useful proof-of-concept data that mycolactone detection could potentially be used to monitor response to antibiotic therapy as well as for diagnosis of Buruli ulcer disease. The findings from the pilot study indicate that rifampicin combined with clarithromycin can replace rifampicin and streptomycin for the continuation phase after rifampicin-streptomycin treatment for 2 weeks without any apparent loss of efficacy. The implication is that a controlled trial of fully oral therapy using rifampicin and clarithromycin for 8 weeks (RC8) is justified.
- ItemThe kinetics of mycolactone in relation to the microbiological, clinical and immunological responses to antibiotic therapy for mycobacterium ulcerans disease.(2014) Sarfo, Fred StephenBackground: Mycobacterium ulcerans is the causative agent for a chronic necrotising skin infection called Buruli ulcer. Pathology of the disease is closely linked with the elaboration of a unique lipid toxin, mycolactone, which has potent cytotoxic and immunomodulatory properties. In this study, assays were developed to detect and quantify mycolactone concentrations in tissues during curative antibiotic therapy in mice and in humans to understand its dynamics in pathogenesis and to explore its potential as a biomarker for diagnosis and monitoring of patients with Buruli ulcer disease on antibiotic therapy. The currently recommended antibiotic regimen for the management of Buruli ulcer is a combination of daily intramuscular injections of streptomycin and oral rifampicin for 8 weeks (RS8). This regimen was compared with streptomycin/rifampicin for 2 weeks followed by clarithromycin/rifampicin (RS2RC6) for 6 weeks in patients to determine the clinical and bacteriological effectiveness in a pilot study. Methods: Biopsies were obtained from infected human skin tissues and BALB/c mouse hind footpads before, during and after 8-weeks of rifampicin-containing combination antibiotic therapy. Lipids were extracted from tissues using organic solvents, mycolactone concentrations were measured using a cytotoxicity assay and mass spectrometry. Trends in mycolactone concentrations and clinical, bacteriological and immuno-histopathological responses were determined. Concentrations of cytokines in supernates of whole blood assays in humans or murine splenocytes after stimulation with mycobacterial antigens/T-cell mitogens were measured using ELISA. iv Results: Eighty-three patients with confirmed Buruli ulcer were randomized to RS8 or RS2RC6 and monitored for recurrence free-healing. Bacterial load in tissue samples before and after treatment for 6 and 12 weeks was monitored in samples obtained by 4mm punch biopsy by semi-quantitative culture. There was no difference in using RS8 or RS2RC6 with respect to healing rate or the proportion healed in each group after 4, 8, 12, 16, 20, 24 and up to 52 weeks. The success rate was 93% in each group and there was no recurrence after 12-month follow-up. There was no difference in the number of bacteria cultured at the different time points for the two regimens. Mycolactone was detectable in 92% and 77% of human samples (n=80) using cytotoxicity assays and mass spectrometry respectively. Antibiotic therapy was associated with a decline in tissue concentration of mycolactone in both human and murine-infected tissues which was paralleled by resolution of clinical lesions, reductions in bacteriological counts and restoration of local and systemic immune responses. Discussions/Conclusions: This study shows that mycolactone concentrations in tissues is closely associated with the presence of M. ulcerans and provides useful proof-of-concept data that mycolactone detection could potentially be used to monitor response to antibiotic therapy as well as for diagnosis of Buruli ulcer disease. The findings from the pilot study indicate that rifampicin combined with clarithromycin can replace rifampicin and streptomycin for the continuation phase after rifampicin-streptomycin treatment for 2 weeks without any apparent loss of efficacy. The implication is that a controlled trial of fully oral therapy using rifampicin and clarithromycin for 8 weeks (RC8) is justified.
- ItemMechanistic Studies and Mathematical Models for Mycolactone Toxin and Autoinhibitory WASP: Model Mechanisms for Buruli ulcer Initiation and Spread.(2015-11-09) Nyarko, Peter RomeoThe Wiskott-Aldrich Syndrome Protein (WASP) has been implicated in many diseases such as Wiskott-Aldrich Syndrome (WAS) and Buruli ulcer. Mycobactrium ulcerans is the main causative organism of Buruli ulcer (BU) disease. The bacteria secretes a polyketide lipid toxin (Mycolactone). The toxin not only di uses through the cell membranes, but also binds, hijacks and disrupts the normal functions of WASP in the cytoplasm leading to over polymerization of actin lament, cytoskeletal rearrangement and eventually cell death through necrosis. In pre-ulcerative BU disease, toxins extend beyond the actual size of the lesion. A mathematical model is developed to describe the binding mechanism of the two conformations of WASP and the complexes formed using the idea of isomerization. The formulation utilizes ligand concentration-dependence (ligand-receptor), equilibrium and conservation principles. By this approach, we are able to determine the fractional response of WASP against change in concentration of its activators;
- ItemModelling Customers Response to Service offers using game Theory Case Study; MTN and Vodafone in Bolgatanga Polytechnic(2011-11-18) Issahaku, Abdul-MajeedGame theory is being found to be one of the useful tools for analysing strategic situations that involve competitors or rivals who struggle against each other for supremacy...
- ItemPerformance Assessment of Satellite and DACCIWA Optical Gauge Rainfall Products for the Analysis of Trends and Drivers of Rainfall Extremes in Ghana(KNUST, 2018-11) Atia Winifred AyinpogbillaRainfall plays an important role in the socio-economic development of any nation. Below or above normal rainfall conditions have serious consequences on key socioeconomic sectors such as agriculture. This is particularly true for countries where rain-fed agriculture is a predominant practice as such, rainfall impact studies are important. In this thesis, rainfall impact studies in Ghana were examined in three different areas. The first part of the study evaluated the performance of satellites and merged rainfall products over Ghana in order to examine their reliability as surrogate ground-based gauge measurements for areas where gauge networks are sparse. Satellite products (TAMSAT, TRMM, ARC2, CMAP, GPCP and CHIRPs) and gauge-only products (CRU and GPCC) were validated with gridded rainfall data from the Ghana Meteorological Agency (GMet) on monthly to annual time scales using a suite of statistical analysis. The results showed that, the performance of the satellites and merged rainfall products is a function of the scales and locations used in the validation. While ARC2 showed large biases on both monthly and annual time scales, all other products, especially, TRMM and TAMSAT showed relatively good skills on the monthly scale (r > 0.90) than the annual. On the countrywide basis, CHIRPs and CRU markedly revealed better skills on both time scales. In all the four agro-ecological zones of the country, the products were able to capture the respective rainfall patterns, onset, cessation and spells (wet and dry) of each zone. Thus, TRMM and TAMSAT will serve as better surrogates at monthly time scale whereas CHIRPS and CRU would be better substitutes to ground-based gauge data for both time scales over the entire country and in the four ecological zones. In the second part of the study, the performance of the new Dynamic Aerosol Chemistry-Cloud Interactions in West Africa (DACCIWA) optical rain gauge (DOG) measurements over the Ashanti region of Ghana were assessed. The study further examined the validities of subdaily rainfall data from IMERG and TRMM over the region for their exploits in the key socio-economic activities of the country such as agriculture, hydrology and water resource management. Daily rainfall data for nine co-located stations from the Ghana Meteorological agency (GMet) were used to assess the validity of the DOGs. A point-point assessment of IMERG and TRMM from 2016–2017 for the nine stations were then performed using the DOGs. The outcome of the assessment revealed that the DOGs were able to capture the diurnal and monthly rainfall patterns in the region. IMERG revealed a better skill in capturing the daily rainfall than TRMM although both products showed some similarities in the June–August (JJA) season. Both products slightly over-estimated gauge measurements (<2 mm/day) in all selected stations. The third part of the study examined the climatologies and trends of extreme rainfall events in Ghana. The study further probed the links between the rainfall extremes and SST anomalies at the oceanic basin. The trend analysis revealed that CDD, CWD, R10mm and R20mm were in the range of 5–140, 5–14, 25–60 and 2–26 days per year respectively. Moreover, PRCPTOT, R95p, R99p, SDII, RX1day and RX5day were in the range of 800-1800 mm, 150–450 mm, 30–130 mm, 6–14 mm, 30–100 mm and 60–190 mm per year respectively. The maximum of temporally averaged intensity rainfall indices have shown to cover southwestern Ghana while the minimum of these indices covered northwestern and eastern coasts of Ghana. Significant decreasing trends in wet indices were observed over the Volta lake and central Ghana while low positive trends were dominant over the northern sectors of the country. In most cases, with the exception of the CDD and CWD indices, there were significant positive (negative) correlations with the Atlantic (Pacific and Indian) basins SSTs. IOD had a dipole effect on rainfall indices with the central and southern sectors generally covered by negative correlations while northern and coastal Ghana have shown positive correlations. The R95p index over central Ghana had negative correlations with AMO while positive correlations dominated some isolated parts of the country in the south and northeast. The impacts of AMO on PRCPTOT index were insignificant over most parts of the country with a few exceptions over southeastern and northern boarder regions. These results have implications on the improvement of monthly-annual forecasts of the Ghanaian rainfall and its extremes, and also provides prior knowledge for better understanding of multidecadal modulations of global inter-annual teleconnections
- ItemPreparation and characterization of fat particles-in-water emulsions for the encapsulation of bioactive compounds using allanblackia seed fat and tween 20 and sodium alginate as emulsifying agents(2015-11-09) Badu, MercyEffective delivery of poorly water-soluble bioactive ingredients has been considered a great challenge in the pharmaceutical, food and cosmetic industries. The compartmentalized hydrophobic and hydrophilic regions of emulsion systems give them the advantage to encapsulate polar and non-polar compounds for effective delivery. However, the challenge associated with emulsions is their thermodynamic instability. This study investigated the stabilisation process of fully crystallised fat globules as a dispersed phase in an oil-in-water emulsion system prepared from a naturally occurring vegetable fat, a nonionic surfactant and a polysaccharide biopolymer. The crystallisation profiles of the fats from Allanblackia seed and shea nuts were determined. Based on the results Allanblackia seed fat was selected and used as the dispersed phase in the oil-in-water emulsion system. The emulsion was formulated in the presence of Tween 20 and sodium alginate. The mixed emulsifier system gave stability against aggregation of the fat particles hence preventing coalescing, creaming and flocculation and this was more significant at 2 % to 4 % sodium alginate content. The emulsion particle size and microstructure were dependent on the fat/water ratio and a viscoelastic system with G′ > G′′ at the lower frequencies and G′′ > G′ at higher frequencies (where G and G are….). The dispersed fat particles emulsion system was used to encapsulate sudan orange dye (a water insoluble dye) and the efficiency of the encapsulation was characterized by an increase in particle size (ranging from 246 to 250 nm) and varying size distribution. UV-Vis spectrophotometric analysis of the dye loading capacity, release rate and leaching capacity showed a successful entrapment of the dye in the fat matrix. This confirms Allanblackia seed fat as a potential solid fat for its application in the formulation of oil-in-water emulsions.
- ItemQuantum Mechanical Studies of the Mechanisms of Some Transition Metal Organometallic Reactions(2009-07-18) Tia, RichardThe mechanistic pathways of three organometallic reactions, namely the oxidation of ethylene by chromyl chloride leading to the formation of epoxide, 1,2-dichloroethane, 1,2-chlorohydrin, acetaldehyde, and vinyl alcohol precursors; the olefin metathesis reaction involving ethylene and 20 metal methylidene Cl4MCH2(M=Cr, Mo, W, Ru, Re) and metal oxo-methylidene Cl2(O)MCH2(M=Cr, Mo, W, Ru, Re) complexes; and the transition-metal-assisted formation of 1,2-dinitroso complexes of cobalt and its congeners have been studied using hybrid density functional theory at the B3LYP/LACVP* level of theory. The formation of the epoxide precursor (Cl2(O)Cr-OC2H4) was found to take place via initial [2+2] addition of ethylene across the Cr=O bond of CrO2Cl2 to form a chromaoxetane intermediate, as opposed to [3+2] addition across the two Cr=O bonds of CrO2Cl2 as suggested in earlier studies. The hitherto unexplored pathway involving initial [3+2] addition of ethylene across the Cr=O and Cr-Cl bonds of CrO2Cl2 was found to be more favorable than the [3+2] addition across the two Cr-O bonds of CrO2Cl2. The formation of the 1,2-dichloroethane precursor was found to take place via [3+2] addition of ethylene across the two Cr-Cl bonds of CrO2Cl2. The 1,2-chlorohydrin precursor was also found to originate from [3+2] addition of ethylene across the Cr-O and Cr-Cl bonds of CrO2Cl2 as opposed to [2+2] addition of ethylene across the Cr-Cl bond. Also the vinyl alcohol and acetaldehyde precursors were found to arise from a direct attack of one of the carbon atoms of ethylene on an oxygen atom of CrO2Cl2 through a triplet intermediate. In the reactions of Cl4MCH2(M=Cr, Mo, W, Ru, Re) with ethylene it was found that the formation of the metallacyclobutane through formal [2+2] cycloaddition, a key step in the olefin metathesis reaction according to the Herrison-Chauvin mechanism, is a low-barrier process in each of the complexes studied. It was also found that the active species for the formation of the metallacyclobutane is a carbene complex and not a carbenoid complex. One key factor was found to be responsible for the difference in metathesis activity in these complexes: the stability of the carbenoid complexes relative to the carbenes. In Cr and Ru, the carbenoid complexes are more stable than the carbenes and thus Cl4CrCH2 and Cl4RuCH2 are likely to exist in the lower- 21 energy carbenoid Cl3MCH2Cl form as opposed to the carbene Cl4M=CH2 form. This is likely to deplete the reaction surface of the active species of the process, making Cl4MCH2(M=Cr, Ru) not suitable for olefin metathesis. This suggests that whereas Cl4MCH2 (M = Mo, W, Re) may catalyze olefin metathesis, Cl4MCH2 (M = Cr, Ru) may not. The W and Re complexes have been found to have greater metathesis activity than the Mo complex. In the Cl2(O)MCH2 (M=Mo, W, Re) complexes the metathesis reaction has favorable energetics and is found to be more feasible than the side-reactions studied while in the Cl2(O)MCH2(M=Cr, Ru) complexes, the olefin metathesis is found to be less favorable than the side reactions. In the transition-metal-assisted formation of 1,2-dinitrosoalkanes, it was found that the activation barriers for the one-step [3+2] addition pathway for the formation 1,2-dinitrosoalkanes are generally very low while the activation barriers for the [2+2] addition of the C=C bond of the olefins across the M-N bonds of CpM(NO)2 (M=Co, Rh, Ir) to form an intermediate are generally very high. A transition state for the re-arrangement of the products of [2+2] addition to the products of [3+2] addition could not be located, indicating that the re-arrangement of the products of [2+2] addition by reductive elimination involving the second metal-nitrogen π-bond to form the observed 1,2-dintrosoalkanes as suggested in the work of Rappé and Upton may not be possible. Therefore it is concluded that the direct one-step [3+2] addition pathway proposed by Bergman and Becker for the formation of 1,2-dinitrosoalkanes is a more plausible pathway.