The success of our first-line antimalarial treatments is threatened by increased drug resistance in Plasmodium parasites. This makes the development of novel drugs critical to combat malaria. Historically, natural products have been an excellent source of novel antimalarial compounds and thus are an ideal place to search for potential drugs. Filamentous members of the bacterial phylum, Actinobacteria, are well-known antibiotic producers, but their antimalarial potential has not been well investigated. This makes these actinobacteria a potentially valuable source of novel antimalarial compounds. To evaluate the antimalarial potential of the filamentous actinobacteria, uncharacterized environmental actinobacterial strains from the Meyers laboratory culture collection, as well as the type strains of new actinobacterial species identified and characterized in the Meyers laboratory, were screened for antiplasmodial activity against drug-sensitive Plasmodium falciparum, NF54. Liquid cultures were extracted using the mid-polar solvent, ethyl acetate, with the aim of discovering drug-like molecules that can be administered orally. Thirty-one strains of actinobacteria belonging to eight genera (Actinomadura, Amycolatopsis, Gordonia, Kribbella, Micromonospora, Nocardia, Nonomuraea, and Streptomyces) were screened revealing fourteen active strains. Eight strains were identified for further study as the displayed antiplasmodial efficacy matching predefined criteria. Of these eight candidates, Streptomyces strain PR3 was selected, as it showed excellent antiplasmodial efficacy, no cytotoxicity against Chinese Hamster Ovary (CHO) or liver HepG2 cell lines, no haemotoxicity, and was easy to culture. Bioassay-guided fractionation of the crude extracts of strain PR3, supported by high-resolution mass spectrometry (HRMS) and nuclear magnetic resonance (NMR) analysis, was conducted to isolate and identify the compounds responsible for the antiplasmodial activity. During purification by solid phase extraction (SPE), a novel class of compounds was isolated. The structure of these compounds was elucidated by HRMS and NMR analysis and determined to be a series of crown ethers with a methylated backbone. These methylated crown ethers (MCE) were not produced by strain PR3, but by the cyclization of polypropylene glycol (PPG) oligomers from Amberlite® XAD-16N 20–60 mesh resin under aqueous conditions. The MCEs displayed weak antiplasmodial activity against P. falciparum NF54, without cytotoxicity against the Chinese Hamster Ovary, HepG2 cell lines, nor human erythrocytes. To the author's knowledge, the MCEs are novel compounds, and this is the first time the cyclization of PPG oligomers into crown ethers has been reported. As the MCEs were not responsible for strain PR3's potent antiplasmodial activity, further study was conducted. Using the Global Natural Product Social molecular networking (GNPS) workflow, genome mining, and NMR analysis, it was revealed that the cyclodepsipeptides, valinomycin, montanastatin, and nine other novel analogues were responsible for the high antiplasmodial activity detected. A review of the literature revealed that the structure of four of these analogues had been predicted, based on MS/MS and the biosynthesis of valinomycin. Using the same described biosynthetic logic and MS/MS analysis, two new cyclodepsipeptides, compounds 1054 and 1068, were elucidated. Unfortunately, chromatographic systems developed were unable to purify the cyclodepsipeptides, and individual evaluation of their antiplasmodial efficacy and host selectivity was not possible. The fraction containing the cyclodepsipeptides exhibited strong antiplasmodial activity against the drug-sensitive, NF54 and multidrug-resistant K1, strains of P. falciparum. No cytotoxicity was displayed against the CHO cell line and no haemotoxicity was seen against human erythrocytes. Moderate toxicity was exhibited against the liver HepG2 cell line; however, the selectivity index of the cyclodepsipeptides suggested that they are selectively targeting the Plasmodium parasites. Overall, these results are positive, and further study of the individual cyclodepsipeptides is warranted. During the investigation, discrepancies were noticed between different fractions in terms of antiplasmodial activity. These fractions contained both the MCEs and, cyclodepsipeptides along with a range of impurities, yet they displayed potent antiplasmodial activity. Further study suggested that combination of the MCEs and cyclodepsipeptides elicits a synergistic response and improves antiplasmodial efficacy. This was determined independently using two models, the fixed-ratio isobologram method and the CompuSyn programme based on the massaction law principle. The workflow developed during this investigation demonstrates how new technologies can be used to dereplicate and elucidate bioactive natural products. This workflow can be utilized to continue this research and identify new natural products that can combat malaria

This research examined the ways in which families constructed presenting problems in the talk of their first session at a psychology clinic. It also looked at the ways in which they constructed individual subjectivities during the interviews. Six families self-selected for therapy and were then invited to participate in the study. The analysis was based on the clinical interviews undertaken by therapists.

An increase in the provision of psychotherapy by mental health professionals was the basis of this study. It was presumed that a comparison of therapists' expectancies would provide a reflection of their approaches to psychotherapy. The current study investigated the effects of profession, education/training and experience on therapists' expectancies regarding the client's needs, the therapists' directiveness, the therapists' level of interpretation and the client's outcome. / There were 64 randomly selected subjects participating in the experiment. The subjects were identified in terms of being either a counselor or clinical psychologist, having a doctorate or a master's degree, and having (LESSTHEQ) or > 7 years of experience. Each subject viewed video interviews of two clients with different problems. The subjects completed the Therapist Expectancy Inventory (TEI) after viewing each interview. / A 2 x 2 x 2 factorial design with repeated measures was employed for the study, with a separate ANOVA performed for each of the 4 dependent variables. An alpha level of .05 was designated in all of the analyses. Additional descriptive statistics were obtained to enhance the primary analyses. / The results of the investigation suggest there are similarities as well as differences among the various groups of therapists regarding their expectancies. Results of the analyses indicated that the greatest amount of variation (with a p < .05) among the expectancies of the therapists occurred first regarding a diagnostic factor (expectancy of client's needs), followed by a prognostic factor (expectancy of client's outcome), which was followed in turn by a process factor (expectancy of therapist's directiveness). A second process factor (expectancy of therapist's level of interpretation) did not yield any significant variation. / Some of the results could be explained readily while other findings were less explicable and could only be afforded speculation. An inconclusive body of literature regarding the factors which were studied would not permit a definitive explanation of some of the findings. It would seem that further research is necessary to support or refute the current data. / The overall contribution of the study may have been limited by the confounding effects of intervening variables, and a possible inherent bias in the make-up of the instrument that was used to measure therapists' expectancies. / Further research should include other mental health professions--e.g., psychiatry and social work. It is also recommended that another means of analyzing the clinical judgments of therapists be used along with the TEI in the future. / Source: Dissertation Abstracts International, Volume: 47-01, Section: B, page: 0377. / Thesis (Ph.D.)--The Florida State University, 1985.

Sustaining a burn injury is one of the most traumatic accidents a child or adolescent can experience. In South Africa paediatric burn injury is a significant social problem and incident rates tend to be elevated in informal settlements because of the reliance on fossil fuels (paraffin, wood, and coal) for heating and cooking. The role of mental health professionals in helping young burn survivors and their families through their traumatic ordeal is of paramount importance. There is currently a shortage of specialised psychosocial assistance for burn patients and their families in South Africa. Given the enormous challenge of paediatric burn injury in South Africa it is alarming that so little research has been undertaken in the field. The present knowledge regarding this issue is largely reliant on international research, which cannot always be generalised to the specific challenges posed by the South African context. The purpose of this study was to provide local insight into the psychosocial adjustment of burn survivors that can inform intervention programmes in this field. In order to explore the psychosocial adjustment of paediatric burn survivors during the school reintegration process it was decided to interview the burn survivors, their parents and their school teachers. The sample comprised 7 paediatric burn survivors and their respective caregivers and teachers. The study was conducted using a qualitative paradigm. Data was collected by conducting semi-structured interviews with the participants and their caregivers and teachers. Data was analysed by conducting a thematic analysis of the transcribed narratives of the participants, their caregivers and teachers. The study yielded important information about the social contexts of a small group of burn survivors and their families and the challenges that they face during the school re-integration process and psychosocial adjustment in generaL The majority of the families in this study were experiencing extreme socio-economic difficulties. This was a strong contributing factor in their limitations to fulfil their roles in supporting the burn survivor. The mental health difficulties of parents as well as participants before and after the burn injury showed that the burn injury often meant an additional stress factor to already existing problems. It was found that the burn injury also poses an additional challenge to most of the interviewed teachers, who have to cope with multiple challenges in their classes and therefore have very limited resources (time, empathy, attention) to attend to the specific challenges of the re-integration of burn survivors. The different layers of problems that were experienced by participants, their families and teachers highlighted the reality of the South African context.

Background: Tuberculosis (TB) is the most common opportunistic infection in children with human immunodeficiency virus (HIV) infection in developing countries, and co-treatment for HIV infection and TB is frequently indicated. Efavirenz and lopinavir/ritonavir (ratio 1:1) as part of antiretroviral therapy are used in combination with rifampicin-based antitubercular treatment in South African TB/HIV co-infected children. Adult studies show that concomitant rifampicin significantly reduces efavirenz and lopinavir plasma concentrations. However, the pharmacokinetics (PK) of efavirenz and lopinavir/ritonavir are poorly characterized in children, especially African children and no study has evaluated the effect of rifampicin-based antitubercular treatment on efavirenz and lopinavir/ritonavir plasma concentrations in children. Although therapeutic drug monitoring (TDM) is recommended in selected patients (including young children and patients receiving concomitant antitubercular treatment), TDM is seldom available in resource-constrained countries. There is an urgent need to develop a field friendly method which requires small volumes of blood, and inexpensive processing and storage conditions. Furthermore, because HIV replicates in the cells, efavirenz and lopinavir need to penetrate into these infected cells to inhibit viral replication. Therefore, directly measurement of intracellular concentrations of these drugs in HIV-infected children could provide better understanding of drug exposure at the action site. It is also important to evaluate the effects of frequently co-administered drugs on intracellular accumulation of efavirenz and lopinavir. Objectives: 1) To evaluate efavirenz and lopinavir/ritonavir plasma concentrations and determine the effects of rifampicin on efavirenz and lopinavir/ritonavir PK in HIV-infected African children with and without rifampicin-based antitubercular treatment. 2) To develop and validate the dried blood spot (DBS) method as an alternative to conventional plasma methods of drug concentration measurement in TDM. 3) To evaluate in vivo intracellular concentrations of efavirenz and lopinavir/ritonavir in HIV-infected children with and without concomitant antitubercular treatment. 4) To determine the in vitro modulation effects on the intracellular accumulation of efavirenz IV and lopinavir in human peripheral blood mononuclear cells (PBMCs) by drug efflux protein inhibitors, as well as frequently co-administered rifampicin and ritonavir (at low dose; as pharmacoenhancer). Methods: 1) Plasma efavirenz and lopinavir/ritonavir concentrations were measured by validated liquid chromatography/tandem mass spectrometry (LC/MS/MS) method in TB/HIV co-infected children during and after rifampicin-based antitubercular treatment as well as in a group of controls (HIV-infected children without TB). Children in the efavirenz study (n= 30) were receiving standard doses of efavirenz as part of antiretroviral treatment. Trough concentrations (Cmin) of efavirenz were estimated by extrapolation of the log-linear concentration-time line to 24 hours after the previous dose. Children in the lopinavir/ritonavir study were receiving additional ritonavir (lopinavir: ritonavir ratio 1:1) during antitubercular treatment (n= 15), and standard doses of lopinavir/ritonavir (LPV/r; ratio 4:1) after antitubercular treatment, and in controls (n= 15). The PK of lopinavir and ritonavir were characterized from concentration-time curves using WinNonlin version 4.1 by non-compartmental analysis. 2) Aliquots of 50 &Icirc;¼ L of whole blood from the efavirenz and lopinavir/ritonavir studies were dried onto filter paper. The drug concentrations were analyzed using validated LC/MS/MS method. The effects of high temperature and direct sunlight on the stabilities of these antiretroviral drugs in DBS samples were tested. 3) Intracellular concentrations of efavirenz, lopinavir and ritonavir were measured in trough concentrations of 11 TB/HIV co-infected children using a validated LC/MS/MS method. Six children were receiving double dose of LPV/r (4:1) with concomitant rifampicin; 5 children were receiving standard doses of efavirenz with rifampicin-based antitubercular treatment, 3 of them had intracellular concentrations measured again after completing rifampicin-based antitubercular treatment. 4) in vitro intracellular accumulation of efavirenz and lopinavir were measured in human PBMCs in the absence and presence of P-glycoprotein inhibitors (verapamil at 50 &Icirc;¼ M, V furosemide at 50 &Icirc;¼ M and cyclosporine A at 20 &Icirc;¼ M) and frequently co-administered drugs at levels representing the average concentrations found in patients (ritonavir at 5 mg/L and rifampicin at 4 mg/L). The concentrations of efavirenz and lopinavir in PBMCs were determined by LC/MS/MS. Results and Conclusions: 1) The co-administration of rifampicin did not significantly reduce efavirenz estimated Cmin concentrations. A high proportion of children with and without concomitant antitubercular treatment had sub-therapeutic efavirenz concentrations despite being correctly dosed according to the manufacturer's instructions, raising concerns about the adequacy of current efavirenz dosing recommendations in children. The lopinavir key PK parameter, Cmin, was not significantly different in same group of children during and after rifampicin-based antitubercular treatment or compared to HIV-infected children without tuberculosis. The recommended minimum therapeutic concentration was achieved in 87% of children during antitubercular treatment and in 92% without concomitant antitubercular treatment. Therefore, in the context of limited options, LPV/r with additional ritonavir (ratio 1:1) is an acceptable approach to treat young children receiving concomitant rifampicin-based antitubercular treatment, although safety remains a concern and hepatic alanine transaminase levels should be monitored regularly. 2) Plasma and DBS concentrations of efavirenz, lopinavir and ritonavir were strongly correlated. The median (interquartile range, IQR) DBS/plasma concentration ratios for efavirenz, lopinavir and ritonavir were 0.93 (IQR 0.83, 1.08), 0.73 (IQR 0.61, 0.90) and 1.05 (IQR 0.74, 1.21), respectively. PK parameters of efavirenz and ritonavir were closely similar between DBS and plasma; whereas lopinavir pre-dose and Cmin (at 12 hours after lopinavir intake) concentrations were 16% lower in DBS samples. The 3 antiretroviral drugs in DBS samples were stable at 37 deg C for 7 days and with exposure to direct sunlight for 2 hours. DBS can be used as an alternative field-friendly method for efavirenz, lopinavir and ritonavir concentration monitoring. However, pre-dose and Cmin concentrations of lopinavir in DBS samples need to be increased by 16% when used to predict plasma concentrations. VI 3) In vivo median intracellular/plasma concentration ratios for efavirenz, lopinavir and ritonavir amongst 11 TB/HIV co-infected children during antitubercular treatment were 0.91 (IQR 0.54, 1.19), 0.22 (IQR 0.09, 0.31) and 4.17 (IQR 1.30, 7.33), respectively. Two children had efavirenz intracellular/plasma concentration ratios during vs. after antitubercular treatment: 1.00 vs. 0.61 and 0.27 vs. 0.79. 4) Furosemide significantly increased efavirenz and lopinavir accumulation in healthy human PBMC samples by 1.2- 1.5 fold. Whereas, neither verapamil nor cyclosporin A had significant effects on efavirenz or lopinavir intracellular accumulation. Despite being an inducer of P-glycoprotein, rifampicin increased the accumulation of both efavirenz and lopinavir to different extents in all 3 PBMC samples. The low-dose ritonavir (at the concentration found in HIV-infected patients) had no effect on intracellular accumulation of efavirenz and lopinavir at therapeutic concentrations.

Includes bibliographical references (leaves 150-175). / Includes abstract. / Protective immunity against mycobacterial infection requires an effective cytolytic response, in addition to an intact Type l (Th1) cytokine pathway. Natural killer (NK) cells and cytolytic T-cells (CTL) are essential components of protective immunity against tuberculosis (TB) and mediate granule-dependent killing of infected cells. Granulysin, an antimicrobial protein, and perforin, a pore-forming molecule, have been found to co-localise in the granules of these two cell types. Granulysin has been shown to be directly cytotoxic to extracellular Mycobacterium tuberculosis (M.tb) and, together with perforin, is cytolytic against intracellular mycobacteria. This project evaluated the role of these two cytolytic mediators in TB.

This is a study of the role of oxygen free radicals in isoniazid induced toxicity and its implications on the safety of isoniazid containing regimens in elderly patients. During isoniazid metabolism, toxic metabolites are produced which, together with isoniazid, may lead to toxicity by mechanism (s) yet unknown. Understanding the mechanism of toxicity is important for effective management and prevention of isoniazid induced toxicity. Elderly patients take priority for this consideration because isoniazid is more toxic in these people. Consequently, "the therapeutic importance of isoniazid metabolism in elderly patients" was found a suitable title for this work. lsoniazid metabolites; acetylisoniazid, monoacetyl-hydrazine and diacetylhydrazine were synthesized and characterized by thin layer chromatography, high performance liquid chromatography (HPLC), ultra-violet absorption spectra and mass spectrometry. Thereafter, these compounds together with hydrazine were used in the subsequent experiments.

No description available.

Although several dozen different compounds are able to transiently alter chloroquine resistance via chemosensitisation, the phenomenon has never evolved beyond laboratory practice as a result of in vivo difficulties. Chemosensitising compounds either need to be administered at doses which are toxic to the host in order to reverse resistance, or the drug is so highly bound to serum proteins that there is an insufficient circulating quantity available to restore sensitivity. Nine chemosensitisers were evaluated in vitro against several resistant isolates of the malaria parasite in order to develop a cocktail treatment of three compounds which could reverse resistance additively or synergistically when used at low doses with chloroquine. This would bypass any toxicity issues which might arise from the use of a high dose of a single agent. Six of the chemosensitisers were selected for combination into six different cocktails which were tested in vitro. Each cocktail contained one antidepressant, one antihistamine and one antipsychotic. Low doses of each drug were able to alter resistance to a small extent singly and in combination; this was shown by determining the effect of drugs and cocktails on both chloroquine transport using radiolabelled chloroquine, and chloroquine efficacy using the lactate dehydrogenase assay for parasite viability. The reversal activity was shown to be additive in the cocktail treatments and not synergistic, and was highly dose-dependent. There was no direct correlation between the change in chloroquine transport and the extent of resistance reversal. The chemosensitisers' effect on chloroquine transport was evaluated in a mouse model of malaria and shown to be similar to that seen against cultured human parasites; following this, the cocktails were tested for efficacy in mice infected with chloroquine-resistant malaria. Five of the six cocktails were able to significantly alter parasite survival in the mice in conjunction with a low dose of chloroquine. Drug levels in the mice were quantified via mass spectrometry and liquid chromatography in order to correlate the efficacy data. One of the compounds in the failed treatment was shown to circulate at low levels in the animals and this is possibly why that treatment, although effective in vitro, did not yield a result in vivo.

BACKGROUND: Antiretroviral therapy (ART), notably efavirenz and lopinavir, have been associated with metabolic abnormalities known to increase cardiovascular risk. Efavirenz and lopinavir pharmacokinetics demonstrate considerable interindividual variability, which in part, may be explained by host genetic factors. Mitochondrial DNA (mtDNA) variation influences ART related metabolic complications. However, the associations between genetic polymorphisms and pharmacokinetics of antiretroviral drugs, and their associations with metabolic complications, are incompletely understood. We explored associations of mitochondrial DNA (mtDNA) haplogroups and ART related metabolic complications, characterized relationships between genetic polymorphisms and plasma efavirenz concentrations, and investigated associations between plasma efavirenz/lopinavir concentrations and lipid and glucose concentrations in HIVinfected Black South Africans. METHODS: We collected clinical and laboratory data from HIV infected patients on ART from Cape Town. We sequenced the mitochondrial genome and determined African mtDNA haplogroups. We genotyped 241 polymorphisms in genes potentially relevant to efavirenz metabolism and transport. We measured steady state efavirenz and lopinavir concentrations and used regression analyses to determine associations with metabolic parameters.