Purification and characterisation of plasmodium falciparum Hypoxanthine phosphoribosyltransferase

Magister Scientiae - MScMalaria remains the most important parasitic disease worldwide. It is estimated that over 500 million infections and more that 2.7 million deaths arising from malaria occur each year. Most (90%) of the infections occur in Africa with the most affected groups being children of less than five years of age and women. this dire situation is exacerbated by the emrggence of drug resistant strains of Plasmodium falciparum. The work reported in this thesis focuses on improving the purification of PfHPRT by investigating the characteristics of anion exchange DE-52 chromatography (the first stage of purification), developing an HPLC gel filtration method for examining the quaternary structure of the protein and possible end stage purification, and initialcrystalization trials. a homology model of the open, unligaded PfHPRT is constructed using the atoomic structures of human, T.ccruz and STryphimurium HPRT as templates.South Afric

Interplay between Platelet Dysfunction and Vascular Thrombosis in Traumatic Injury

Platelets halt bleeding accompanying traumatic injury by performing primary hemostasis to repair vascular leakage at injury sites. In trauma individuals, ex vivo platelet function tests often indicate impairment despite normal count. Moreover, incubation of platelets from normal non-traumatized individuals with plasma from trauma victims demonstrates impairment suggesting association with factors in circulation. Notably, not all trauma victims die from hemorrhage. Despite laboratory evidence of dysfunction, thrombotic vascular occlusions are persistent in trauma survivors as corroborated by postmortem findings from victims who die. The time course of platelet reactions post-traumatic injury, that is, the transition from states favoring bleeding to those that facilitate thrombosis is still unclear. Of the several terminologies describing platelet behavior with regards to injury, including hyporeactivity, anergy, exhaustion, and maladaptive states, few have focused on platelet-platelet interactions. It is increasingly becoming clear that platelet interaction with injured endothelium is a probable missing link in the mechanistic explanation of vascular thrombosis post-traumatic injury. This postulate is supported by evidence of increased adhesive protein, von Willebrand factor, and released from injured endothelium. In all, this potentially explains the suboptimal response to anticoagulants or antiplatelets post-trauma. This chapter will review current knowledge on platelet functions in relation to vascular thrombosis post-trauma, the time course, mechanistic hypothesis, and response to therapeutic interventions and clinical outcomes

Controversies in Platelet Functions in Diabetes Mellitus Type 1

Individuals with diabetes mellitus (DM) are at high risk of thrombosis in which hyperactive platelets are implicated. The platelet hyperactivity has been linked to hyperglycemia. This hypothesis is supported by studies in type II diabetes mellitus showing increased sensitivity of platelets to stimulating agonists in the context of tissue resistance to high-circulating insulin. However, controversy still exists regarding the altered platelet functions in type 1 diabetes mellitus (T1DM) and the link to modifying factors such as blood glucose, hyperlipidemia, metabolic acidosis and insulin treatment. Moreover, increased insulin dosage or treatment appears to have antagonistic actions: diminished functions at low doses and enhanced activation at high doses, the switch being attributable to insulin-like growth factor. The physiological role of insulin in suppressing platelet activation is lost in T1DM, a scenario that favors increased platelet sensitivity to stimulating agonists. Furthermore, the response to antiplatelet agents and statins is sub-optimal in diabetics presenting clinical and research knowledge gap regarding the ideal antiplatelet treatment in DM in general and T1DM in particular. This chapter reviews the unique characteristics of platelet functions in T1DM highlighting the controversial areas linking unique behavior of platelets and the abnormal response to therapeutic interventions

Resistance related metabolic pathways for drug target identification in Mycobacterium tuberculosis

Criteria used to filter high priority M.tuberculosis drug targets. The genes highlighted in bold satisfied all the selection criteria. The hyphen (−) indicates exclusion from further analysis. Abbreviations used: NUI- Not under investigation, PDB- Protein Data Bank, TBSGC- TB Structural Genome Consortium. References 12-Sassetti et al., 2003; 34-Lamichhane et al., 2003. Data can be viewed in Microsoft excel. (XLS 12 kb

Evolving Paradigms in Laboratory Biomarkers of Fibrinolysis Phenotypes and Association with Post-Traumatic Vascular Thrombosis

Traumatic tissue injury triggers blood coagulation to stanch bleeding and concomitant blood clot lysis to restore vascular patency. Approximately, 40% of trauma cases potentially present with trauma-induced coagulopathy that may coexist with clot dissolution or fibrinolysis. Laboratory test results of fibrinolysis biomarkers stratify fibrinolytic phenotypes into hyperfibrinolysis, physiological, hypofibrinolysis, and fibrinolytic shutdown. However, often, there is incongruence between laboratory findings and clinical presentation of bleeding or vascular thrombosis. Increasingly, it is becoming clear that laboratory findings transiently depend on the timing of blood sampling. The spectrum of evolving fibrinolysis phenotypes, a component of nature’s adaptation to wound healing that ranges from initial promotion of blood fluidity to subsequent thrombosis, presents a clinical diagnostic dilemma with regard to the timing of antifibrinolytics or anticoagulants intervention. This chapter will review the available literature on post-traumatic fibrinolytic phenotypes, diagnostic challenges, evolution over time, clinical outcomes following therapeutic interventions, and association with vascular thrombosis

Evolution and structural analysis of Glossina morsitans (Diptera; Glossinidae) Tetraspanins

Tetraspanins are important conserved integral membrane proteins expressed in many organisms. Although there is limited knowledge about the full repertoire, evolution and structural characteristics of individual members in various organisms, data obtained so far show that tetraspanins play major roles in membrane biology, visual processing, memory, olfactory signal processing, and mechanosensory antennal inputs. Thus, these proteins are potential targets for control of insect pests. Here, we report that the genome of the tsetse fly, Glossina morsitans (Diptera: Glossinidae) encodes at least seventeen tetraspanins (GmTsps), all containing the signature features found in the tetraspanin superfamily members. Whereas six of the GmTsps have been previously reported, eleven could be classified as novel because their amino acid sequences do not map to characterized tetraspanins in the available protein data bases. We present a model of the GmTsps by using GmTsp42Ed, whose presence and expression has been recently detected by transcriptomics and proteomics analyses of G. morsitans. Phylogenetically, the identified GmTsps segregate into three major clusters. Structurally, the GmTsps are largely similar to vertebrate tetraspanins. In view of the exploitation of tetraspanins by organisms for survival, these proteins could be targeted using specific antibodies, recombinant large extracellular loop (LEL) domains, small-molecule mimetics and siRNAs as potential novel and efficacious putative targets to combat African trypanosomiasis by killing the tsetse fly vector

De novo assembly and annotation of the Amblyomma hebraeum tick midgut transcriptome response to Ehrlichia ruminantium infection

The South African bont tick Amblyomma hebraeum is a hematophagous vector for the heartwater disease pathogen Ehrlichia ruminantium in southern Africa. During feeding, the tick’s enterocytes express proteins that perform vital functions in blood digestion, including proteins that may be involved in E. ruminantium acquisition, colonization or immunity. To delineate the molecular mechanism of midgut response to E. ruminantium infection, we performed comparative analyses of midgut transcriptomes of E. ruminantium infected engorged A. hebraeum nymphs, and infected adult male and female ticks with their corresponding matched uninfected controls, before and during feeding. A total of 102,036 unigenes were annotated in public databases and their expression levels analyzed for engorged nymphs as well as unfed and partly-fed adult ticks. There were 2,025 differentially expressed genes (DEGs) in midguts, of which 1,225 unigenes were up-regulated and 800 unigenes were down-regulated in the midguts of infected ticks. Annotation of DEGs revealed an increase in metabolic and cellular processes among E. ruminantium infected ticks. Notably, among the infected ticks, there was up-regulation in the expression of genes involved in tick immunity, histone proteins and oxidative stress responses. We also observed up-regulation of glycoproteins that E. ruminantium could potentially use as docking sites for host cell entry. Insights uncovered in this study offer a platform for further investigations into the molecular interaction between E. ruminantium and A. hebraeum

Social-ecological resilience of Gazi Bay and Vanga mangrove systems, Kenya

Mangrove forests are important, yet sensitive and highly threatened coastal ecosystems that require careful management and utilization. This study assesses, compares and contrasts social and ecological resilience of two mangrove forests of Gazi Bay and Vanga and their adjacent coastal communities. Assessment was done on mangrove forests structure, key disturbances in mangroves and social systems, and changes in mangrove management regimes. Sampling was done in 10m*10m quadrants laid along belt transects perpendicular to the shorelines. Socio-economic data was collected using semi-structured interviews and household questionnaires, and further data was obtained from field observations and analysis of satellite images. A historical timeline for the two mangrove systems was created to give insight on disturbance regimes and to reveal changes in systems’ resilience over time. The results reveal that changes in mangrove management regimes have impacted resilience of mangrove systems differently. Damming of Mkurumuji River has led to loss of livelihoods and to death of Sonneratia alba trees at Gazi Bay due to excessive sedimentation. This is happening before the mangroves have had sufficient time to recover from a previous disturbance by clear cutting, presenting a major shock to the S. alba stand and fundamentally altering its state. Human population growth at Gazi Bay and Vanga is not causing a reduction in mangrove resource, as villagers are increasingly using concrete blocks as mangrove substitutes for building. Finally, pressure on Vanga mangrove forest is not due to local utilization, but due to harvesting for trade by traders living far away from the village. The study recommends that the government and other responsible stakeholders should commit to increasing people’s income-generating opportunities to reduce poverty and increase resilience of mangrove systems. Further, scientists and mangrove resource managers should recognise local knowledge and foster its complementarity with scientific knowledge.submittedVersionM-IE