Adherence to antiretroviral therapy in children in Zimbabwe: a randomized control trial to validate a new self-reported adherence monitoring tool
- Authors: Mugore, Linnetie
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/54734 , vital:26607
- Description: Background: Among children taking antiretroviral therapy (ART), self-reports have been widely reported to over-estimate adherence levels. Pill count adherence levels are often lower than self-reported levels, with unannounced home pill count adherence being lower than facility based pill count adherence. There is often poor agreement between pill count adherence levels and those measured using other objective adherence measuring methods such as Medication Event Monitoring Systems (MEMS®), which is widely viewed as the gold standard for adherence measurement. Objectives: The aim of this study was to design and evaluate a new self-reported paediatric adherence monitoring tool, assess the feasibility of using pill count methods in monitoring adherence and identify challenges to reporting adherence among children on ART in rural and urban Zimbabwe. Methods A dual centre, superiority, parallel design RCT was conducted to evaluate the newly-developed visually- and verbally-cued „past 10 days‟ tool for the assessment of adherence in children on ART at two sites in Zimbabwe; Harare Central Children‟s Hospital in an urban setting, and Murambinda Mission Hospital, a rural site. Child-caregiver pairs presenting to one of these facilities for the child‟s review of ART and refill of the medication were recruited, signed informed consent obtained, and were randomised for self-reported adherence monitoring into either the experimental group („new 10-day tool‟) or the control group („PACTG-style‟ self-report tool). Data (demographic, socioeconomic, and reported adherence) were collected in individual interviews with child-caregiver pairs. Additional adherence monitoring methods used for both groups included the Morisky-8-Item Medication Adherence Scale (MMAS-8) and a facility based pill count. FGDs were held with groups of caregivers and groups of children ≥13 years of age to understand reasons for non-adherence as well as issues around reporting non-adherence. Superiority testing was conducted by comparing adherent proportions and their confidence intervals (95% CI). Further concurrent validity test was done using the Mann-Whitney U test to evaluate the relationship between the new tool and the MMAS-8 scores. Agreement between the child and caregiver reports of adherence was used as a test of reliability of the new tool using the kappa statistic. Socio-demographic, clinical and care-related factors associated with adherence were identified using reported adherence in both child and caregiver groups in a logistic regression model. Two pill count methods were assessed for feasibility using the proportions of children with complete data for calculating adherence levels, and their CI and a comparison of the two methods, a routinely-used method and one that incorporated the reported residual quantity (RRQ) of medication at last refill. Results : Analysis included 245 child-caregiver pairs, 123 in the experimental group and 122 in the control group. The median age for children was 9 years. In the experimental group, adherence by caregiver and child reports ranged from 94.3% - 98.4% and 78.4% - 96.1%, and those in the control group ranged from 89.2% - 97.5% and 71.2% - 98.1%, respectively. There was no significant difference between adherence levels in the two groups. Adherence levels measured by both the experimental and control tools were found to be associated with MMAS-8 adherence levels (p <0.05). Agreement between child- and caregiver-reported adherence was moderate though significant (kappa; 0.407, p <0.05). Only about half of the children had adequate data to compute pill counts. Proportions adherent at 95% cut-off were 39% by the „routine pill count‟ and 58% by the „Pill count RRQ‟. Being an orphan was associated with child reported-adherence whereas use of non-human reminders, having a maternal relative as a primary caregiver and knowledge of dose frequency, were all associated with caregiver-reported adherence. Major causes of non-adherence mentioned during the FGDs included interference of medication administration times with scheduling of routine socio-economic activities and lack of support from some non-biological caregivers. Reporting of non-adherence appeared to be hampered by perceptions of negative reactions by healthcare workers to these reports and by caregivers being unaware that the child missed some doses. Conclusions: The „new 10-day tool‟ was not shown to be superior to the „PACTG-style tool‟ in detecting non-adherence, however this new tool was found to be a valid and reliable adherence monitoring tool that included a moderately long recall period of 10 days, can be applied without the need for the respondent to remember names of individual medicines in the
- Full Text:
- Date Issued: 2014
- Authors: Mugore, Linnetie
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/54734 , vital:26607
- Description: Background: Among children taking antiretroviral therapy (ART), self-reports have been widely reported to over-estimate adherence levels. Pill count adherence levels are often lower than self-reported levels, with unannounced home pill count adherence being lower than facility based pill count adherence. There is often poor agreement between pill count adherence levels and those measured using other objective adherence measuring methods such as Medication Event Monitoring Systems (MEMS®), which is widely viewed as the gold standard for adherence measurement. Objectives: The aim of this study was to design and evaluate a new self-reported paediatric adherence monitoring tool, assess the feasibility of using pill count methods in monitoring adherence and identify challenges to reporting adherence among children on ART in rural and urban Zimbabwe. Methods A dual centre, superiority, parallel design RCT was conducted to evaluate the newly-developed visually- and verbally-cued „past 10 days‟ tool for the assessment of adherence in children on ART at two sites in Zimbabwe; Harare Central Children‟s Hospital in an urban setting, and Murambinda Mission Hospital, a rural site. Child-caregiver pairs presenting to one of these facilities for the child‟s review of ART and refill of the medication were recruited, signed informed consent obtained, and were randomised for self-reported adherence monitoring into either the experimental group („new 10-day tool‟) or the control group („PACTG-style‟ self-report tool). Data (demographic, socioeconomic, and reported adherence) were collected in individual interviews with child-caregiver pairs. Additional adherence monitoring methods used for both groups included the Morisky-8-Item Medication Adherence Scale (MMAS-8) and a facility based pill count. FGDs were held with groups of caregivers and groups of children ≥13 years of age to understand reasons for non-adherence as well as issues around reporting non-adherence. Superiority testing was conducted by comparing adherent proportions and their confidence intervals (95% CI). Further concurrent validity test was done using the Mann-Whitney U test to evaluate the relationship between the new tool and the MMAS-8 scores. Agreement between the child and caregiver reports of adherence was used as a test of reliability of the new tool using the kappa statistic. Socio-demographic, clinical and care-related factors associated with adherence were identified using reported adherence in both child and caregiver groups in a logistic regression model. Two pill count methods were assessed for feasibility using the proportions of children with complete data for calculating adherence levels, and their CI and a comparison of the two methods, a routinely-used method and one that incorporated the reported residual quantity (RRQ) of medication at last refill. Results : Analysis included 245 child-caregiver pairs, 123 in the experimental group and 122 in the control group. The median age for children was 9 years. In the experimental group, adherence by caregiver and child reports ranged from 94.3% - 98.4% and 78.4% - 96.1%, and those in the control group ranged from 89.2% - 97.5% and 71.2% - 98.1%, respectively. There was no significant difference between adherence levels in the two groups. Adherence levels measured by both the experimental and control tools were found to be associated with MMAS-8 adherence levels (p <0.05). Agreement between child- and caregiver-reported adherence was moderate though significant (kappa; 0.407, p <0.05). Only about half of the children had adequate data to compute pill counts. Proportions adherent at 95% cut-off were 39% by the „routine pill count‟ and 58% by the „Pill count RRQ‟. Being an orphan was associated with child reported-adherence whereas use of non-human reminders, having a maternal relative as a primary caregiver and knowledge of dose frequency, were all associated with caregiver-reported adherence. Major causes of non-adherence mentioned during the FGDs included interference of medication administration times with scheduling of routine socio-economic activities and lack of support from some non-biological caregivers. Reporting of non-adherence appeared to be hampered by perceptions of negative reactions by healthcare workers to these reports and by caregivers being unaware that the child missed some doses. Conclusions: The „new 10-day tool‟ was not shown to be superior to the „PACTG-style tool‟ in detecting non-adherence, however this new tool was found to be a valid and reliable adherence monitoring tool that included a moderately long recall period of 10 days, can be applied without the need for the respondent to remember names of individual medicines in the
- Full Text:
- Date Issued: 2014
An investigation into the development of knowledge and strategies for the teaching of visual literacy in under-resourced Eastern Cape schools
- Authors: Mbelani, Madeyandile
- Date: 2014
- Subjects: Uncatalogued
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/64401 , vital:28540
- Description: This thesis reports on a multiple case study PhD project that aimed to investigate meaningful and critical development of knowledge and strategies to teach visual literacy, a component of English First Additional Language (FAL) in six under-resourced schools of the Eastern Cape of South Africa. The study begins by locating visual literacy within a broad framework of literacy as a social practice, and discusses its importance. Further, it discusses complexities of making sense of and teaching visual literacy, especially for the majority of in-service teachers who experienced visual literacy neither as learners nor as teacher trainees. The gap between the curriculum and teachers’ classroom practices is what triggered this study to adopt a transformative paradigm. The main research question is, “How can teacher professional development in English Language Teaching advance in-service teachers’ knowledge of and strategies for meaningful and critical teaching and learning of visual literacy?” To respond to this question, I drew on cultural historical activity theory (CHAT) and critical realism (CR) to design four phases of this study that incorporated the seven stages of an expansive learning cycle. These phases focussed on exploring and expanding teachers’ sense making and teaching of visual literacy. I collected data through interviews, document analysis, videoed lessons and change laboratory (CL) workshops. I designed a data analysis tool that brought together CHAT, CR, multimodal social semiotics, critical discourse analysis and pedagogical discourse to make sense of the data. Through a process of reflexivity, the study illuminated layers of factors that constrained meaningful and critical teaching of visual literacy in the empirical, the actual and the real domains of reality. These factors include teachers’ unconscious reproduction of discourses of domination, their intolerance of diverse cultural discourses, resistance to curriculum change, and the fact that they are comfortable with the status quo. I brought these factors to CL workshops for expansive learning. The study contributes in-depth insight into English FAL in-service teacher development in the area of visual literacy. By locating the study within meaning making and teaching of visual literacy, it was possible to interrogate access, diversity, domination and design in teachers’ classroom practices. As a result of this study participants were made aware of the extent to which these factors enabled or hindered meaningful and critical teaching. Participants repositioned themselves as subjects of the activity system, thereby mobilising their agency to take control of the structures and cultures that condition their teaching.
- Full Text:
- Date Issued: 2014
- Authors: Mbelani, Madeyandile
- Date: 2014
- Subjects: Uncatalogued
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/64401 , vital:28540
- Description: This thesis reports on a multiple case study PhD project that aimed to investigate meaningful and critical development of knowledge and strategies to teach visual literacy, a component of English First Additional Language (FAL) in six under-resourced schools of the Eastern Cape of South Africa. The study begins by locating visual literacy within a broad framework of literacy as a social practice, and discusses its importance. Further, it discusses complexities of making sense of and teaching visual literacy, especially for the majority of in-service teachers who experienced visual literacy neither as learners nor as teacher trainees. The gap between the curriculum and teachers’ classroom practices is what triggered this study to adopt a transformative paradigm. The main research question is, “How can teacher professional development in English Language Teaching advance in-service teachers’ knowledge of and strategies for meaningful and critical teaching and learning of visual literacy?” To respond to this question, I drew on cultural historical activity theory (CHAT) and critical realism (CR) to design four phases of this study that incorporated the seven stages of an expansive learning cycle. These phases focussed on exploring and expanding teachers’ sense making and teaching of visual literacy. I collected data through interviews, document analysis, videoed lessons and change laboratory (CL) workshops. I designed a data analysis tool that brought together CHAT, CR, multimodal social semiotics, critical discourse analysis and pedagogical discourse to make sense of the data. Through a process of reflexivity, the study illuminated layers of factors that constrained meaningful and critical teaching of visual literacy in the empirical, the actual and the real domains of reality. These factors include teachers’ unconscious reproduction of discourses of domination, their intolerance of diverse cultural discourses, resistance to curriculum change, and the fact that they are comfortable with the status quo. I brought these factors to CL workshops for expansive learning. The study contributes in-depth insight into English FAL in-service teacher development in the area of visual literacy. By locating the study within meaning making and teaching of visual literacy, it was possible to interrogate access, diversity, domination and design in teachers’ classroom practices. As a result of this study participants were made aware of the extent to which these factors enabled or hindered meaningful and critical teaching. Participants repositioned themselves as subjects of the activity system, thereby mobilising their agency to take control of the structures and cultures that condition their teaching.
- Full Text:
- Date Issued: 2014
Development of styrene based imprinted sorbents for selective clean-up of metalloporphyrins in organic media
- Authors: Awokoya, Kehinde Nurudeen
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/54521 , vital:26580
- Description: Most crude oils contain traces of vanadium and nickel complex with porphyrins (VTPP and NTPP) within their asphaltene fraction. Although these metals are only present in trace quantities, they have a significant and detrimental impact on the refining process by degrading the quality of intermediate and end products. Therefore, their selective removal is highly desirable. This thesis presents the development of nickel porphyrin, nickel vanadyl porphyrin imprinted nanofibers and vanadyl porphyrin imprinted polymer (MIP) particles for application as selective sorbents. Computational model based on the combination of molecular dynamics simulations and quantum mechanics was successfully applied to the styrene functional monomer selection. The particle was prepared by bulk polymerization and the nanofibers by a novel approach combining molecular imprinting and electrospinning technology. The morphologies, thermal stabilities and porosities of the imprinted sorbents were studied using SEM, TGA, and BET nitrogen gas adsorption. Chloroform was found to swell the polymer particles to a higher degree than methanol and acetonitrile. The adsorption characteristics of the imprinted sorbents best fitted with Freundlich isotherm model. The imprinted sorbents recorded high extraction efficiencies (EEs) of > 99 % in selectively extracting the metalloporphyrins. The impact of the template on the affinity of recognition for NTPP was evaluated. The results showed that the NTPP adsorption capacity increased as the molar ratio of NTPP to styrene was increased from 1:1 to 3:1. The optimal ratio of template to functional monomer which yielded the best specific affinity and highest recovery (99.9 %) was 3:1. The effects of trifluoroacetic acid (TFA), dichloromethane (DCM), dimethyl sulphoxide (DMSO), pentane (PEN) on electro-spinnability of the polymer solutions and the morphological appearance of the nanofibers were investigated. The imprinted nanofibers exhibited the same selectivity specialism for both NTPP and VTPP. A remarkable stability in relation to reusability was observed when imprinted nanofibers were used, as they could be reused nine times without incurring any significant loss in removal efficiency. The results were validated by analysing a certified reference material. The imprinted sorbents were therefore found to be selective sorbents that are well suited for handling trace metals in organic media.
- Full Text:
- Date Issued: 2014
- Authors: Awokoya, Kehinde Nurudeen
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/54521 , vital:26580
- Description: Most crude oils contain traces of vanadium and nickel complex with porphyrins (VTPP and NTPP) within their asphaltene fraction. Although these metals are only present in trace quantities, they have a significant and detrimental impact on the refining process by degrading the quality of intermediate and end products. Therefore, their selective removal is highly desirable. This thesis presents the development of nickel porphyrin, nickel vanadyl porphyrin imprinted nanofibers and vanadyl porphyrin imprinted polymer (MIP) particles for application as selective sorbents. Computational model based on the combination of molecular dynamics simulations and quantum mechanics was successfully applied to the styrene functional monomer selection. The particle was prepared by bulk polymerization and the nanofibers by a novel approach combining molecular imprinting and electrospinning technology. The morphologies, thermal stabilities and porosities of the imprinted sorbents were studied using SEM, TGA, and BET nitrogen gas adsorption. Chloroform was found to swell the polymer particles to a higher degree than methanol and acetonitrile. The adsorption characteristics of the imprinted sorbents best fitted with Freundlich isotherm model. The imprinted sorbents recorded high extraction efficiencies (EEs) of > 99 % in selectively extracting the metalloporphyrins. The impact of the template on the affinity of recognition for NTPP was evaluated. The results showed that the NTPP adsorption capacity increased as the molar ratio of NTPP to styrene was increased from 1:1 to 3:1. The optimal ratio of template to functional monomer which yielded the best specific affinity and highest recovery (99.9 %) was 3:1. The effects of trifluoroacetic acid (TFA), dichloromethane (DCM), dimethyl sulphoxide (DMSO), pentane (PEN) on electro-spinnability of the polymer solutions and the morphological appearance of the nanofibers were investigated. The imprinted nanofibers exhibited the same selectivity specialism for both NTPP and VTPP. A remarkable stability in relation to reusability was observed when imprinted nanofibers were used, as they could be reused nine times without incurring any significant loss in removal efficiency. The results were validated by analysing a certified reference material. The imprinted sorbents were therefore found to be selective sorbents that are well suited for handling trace metals in organic media.
- Full Text:
- Date Issued: 2014
Ecological consequences of non-native fish invasion in Eastern Cape headwater streams
- Authors: Ellender, Bruce Robert
- Date: 2014
- Subjects: Fishery management -- South Africa -- Eastern Cape Fishes -- Conservation -- South Africa Introduced fishes -- South Africa -- Eastern Cape Introduced organisms
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/69065 , vital:29380
- Description: The introduction, spread and concomitant impacts of non-native species are a global problem. Fish are among the most widely introduced vertebrate groups, with their impacts affecting multiple levels of organisation- from individuals, populations and communities, to entire ecosystems. In South Africa, the largest perceived threat to range-restricted endemic headwater stream fishes is said to be invasion by non-native fishes, however, as is the case elsewhere, invasive impacts are often a case of risk perception rather than actual risk analysis. Two range-restricted headwater species, the Eastern Cape redfin Pseudobarbus afer and the Border barb Barbus trevelyani are redlisted by the International Union for the Conservation of Nature (IUCN) as ‘Endangered’, primarily due to invasion by non-native fishes. To investigate invasions in South Africa, and provide a quantitative estimate of the impact of non-native fishes on the two imperilled endemics, P. afer and B. trevelyani, the overall aims of this thesis were to: (A) Provide a literature review on non-native fish invasions in South Africa; (B) Using two case studies on the headwaters of the perennial Keiskamma and episodic Swartkops River systems, investigate the naturalisation-invasion continuum to provide a holistic view of the invasion process in these variable environments. The specific thesis objectives were: (1) Reviewing current knowledge of invasive impacts of non-native fishes in South Africa; (2) Investigating invasibility of headwater stream environments by non-native fishes; (3) Determining the establishment success of non-native fishes, (4) Assessing the spatial and temporal impacts of invasion; (5) Understanding mechanisms responsible for non-native fish impacts; (6) Investigating the threat of non-native fish invasion on the genetic diversity of two the two headwater fishes, P. afer and B. trevelyani. Results from the literature review of fish invasions (Chapter 1) showed that South Africa has a long history of non-native fish introductions, spanning two and a half centuries. Currently, 55 species have been introduced or translocated. Many of these introduced species have become fully invasive (36%). Their impacts also span multiple levels of biological organisation. There was a general paucity of studies on fish invasions (38 studies), however, of those conducted, reviewed studies placed emphases on invasive impacts (25 studies) and the transport, introduction, establishment and spread stages of the invasion process were largely ignored. The two study systems, the Swartkops and Keiskamma Rivers, were heavily invaded and numbers of introduced species surpassed that of natives (Chapter 2, 3 and 5). Headwater streams had varying invasibility and a number of non-native species were successfully established (Chapter 2, 3, 5 and 6). The remainder of the invasions were casual incursions into headwater streams from source populations in mainstream and impoundment environments which were invasion hotspots. Irrespective of establishment, four predatory invaders (largemouth bass Micropterus salmoides, smallmouth bass M. dolomieu, brown trout Salmo trutta and rainbow trout Oncorhynchus mykiss) impacted heavily on native fish communities (Chapter 3, 4 and 5). Two broad types of invasion were documented, top down invasion by non-native O. mykiss and S. trutta and upstream invasion by M. salmoides and M. dolomieu (Chapter 3 and 5). Their impacts included changes in community structure, extirpation from invaded stream reaches resulting in contracted distribution, and isolation and fragmentation of native fish populations. The impacts of non-native predatory fishes were particularly acute for P. afer and B. trevelyani. Where non-native predatory fish occurred, P. afer and B. trevelyani had been extirpated (Chapter 3 and 5). As a result both native species exhibited contracted distributions (>20% habitat loss due to invasion). Upstream invasion by centrarchids isolated and fragmented P. afer populations into headwater refugia, while top down invasion by salmonids excluded B. trevelyani from invaded, more pristine stream reaches, by forcing the species into degraded unsuitable lower stream reaches. Predation also disrupted population processes such as adult dispersal for P. afer, and centrarchid-invaded zones acted as demographic sinks, where adults dispersing through invaded reaches were rapidly depleted. While the Mandela lineage of P. afer exhibited little within or between drainage genetic structuring, B. trevelyani was >4% divergent between drainages, and up to 2% divergent between streams within the Keiskamma River system (Chapter 7). The distribution of genetic diversity for B. trevelyani also indicated that the loss of diversity was imminent without immediate conservation interventions. This thesis has provided conclusive evidence that native fishes are vulnerable to invasion and that non-native predatory fishes have significant impacts on native fishes in Eastern Cape headwater streams. If management and conservation measures are implemented, the unwanted introduction and spread of non-native fishes may be restricted, allowing native fishes opportunities for recovery.
- Full Text:
- Date Issued: 2014
- Authors: Ellender, Bruce Robert
- Date: 2014
- Subjects: Fishery management -- South Africa -- Eastern Cape Fishes -- Conservation -- South Africa Introduced fishes -- South Africa -- Eastern Cape Introduced organisms
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/69065 , vital:29380
- Description: The introduction, spread and concomitant impacts of non-native species are a global problem. Fish are among the most widely introduced vertebrate groups, with their impacts affecting multiple levels of organisation- from individuals, populations and communities, to entire ecosystems. In South Africa, the largest perceived threat to range-restricted endemic headwater stream fishes is said to be invasion by non-native fishes, however, as is the case elsewhere, invasive impacts are often a case of risk perception rather than actual risk analysis. Two range-restricted headwater species, the Eastern Cape redfin Pseudobarbus afer and the Border barb Barbus trevelyani are redlisted by the International Union for the Conservation of Nature (IUCN) as ‘Endangered’, primarily due to invasion by non-native fishes. To investigate invasions in South Africa, and provide a quantitative estimate of the impact of non-native fishes on the two imperilled endemics, P. afer and B. trevelyani, the overall aims of this thesis were to: (A) Provide a literature review on non-native fish invasions in South Africa; (B) Using two case studies on the headwaters of the perennial Keiskamma and episodic Swartkops River systems, investigate the naturalisation-invasion continuum to provide a holistic view of the invasion process in these variable environments. The specific thesis objectives were: (1) Reviewing current knowledge of invasive impacts of non-native fishes in South Africa; (2) Investigating invasibility of headwater stream environments by non-native fishes; (3) Determining the establishment success of non-native fishes, (4) Assessing the spatial and temporal impacts of invasion; (5) Understanding mechanisms responsible for non-native fish impacts; (6) Investigating the threat of non-native fish invasion on the genetic diversity of two the two headwater fishes, P. afer and B. trevelyani. Results from the literature review of fish invasions (Chapter 1) showed that South Africa has a long history of non-native fish introductions, spanning two and a half centuries. Currently, 55 species have been introduced or translocated. Many of these introduced species have become fully invasive (36%). Their impacts also span multiple levels of biological organisation. There was a general paucity of studies on fish invasions (38 studies), however, of those conducted, reviewed studies placed emphases on invasive impacts (25 studies) and the transport, introduction, establishment and spread stages of the invasion process were largely ignored. The two study systems, the Swartkops and Keiskamma Rivers, were heavily invaded and numbers of introduced species surpassed that of natives (Chapter 2, 3 and 5). Headwater streams had varying invasibility and a number of non-native species were successfully established (Chapter 2, 3, 5 and 6). The remainder of the invasions were casual incursions into headwater streams from source populations in mainstream and impoundment environments which were invasion hotspots. Irrespective of establishment, four predatory invaders (largemouth bass Micropterus salmoides, smallmouth bass M. dolomieu, brown trout Salmo trutta and rainbow trout Oncorhynchus mykiss) impacted heavily on native fish communities (Chapter 3, 4 and 5). Two broad types of invasion were documented, top down invasion by non-native O. mykiss and S. trutta and upstream invasion by M. salmoides and M. dolomieu (Chapter 3 and 5). Their impacts included changes in community structure, extirpation from invaded stream reaches resulting in contracted distribution, and isolation and fragmentation of native fish populations. The impacts of non-native predatory fishes were particularly acute for P. afer and B. trevelyani. Where non-native predatory fish occurred, P. afer and B. trevelyani had been extirpated (Chapter 3 and 5). As a result both native species exhibited contracted distributions (>20% habitat loss due to invasion). Upstream invasion by centrarchids isolated and fragmented P. afer populations into headwater refugia, while top down invasion by salmonids excluded B. trevelyani from invaded, more pristine stream reaches, by forcing the species into degraded unsuitable lower stream reaches. Predation also disrupted population processes such as adult dispersal for P. afer, and centrarchid-invaded zones acted as demographic sinks, where adults dispersing through invaded reaches were rapidly depleted. While the Mandela lineage of P. afer exhibited little within or between drainage genetic structuring, B. trevelyani was >4% divergent between drainages, and up to 2% divergent between streams within the Keiskamma River system (Chapter 7). The distribution of genetic diversity for B. trevelyani also indicated that the loss of diversity was imminent without immediate conservation interventions. This thesis has provided conclusive evidence that native fishes are vulnerable to invasion and that non-native predatory fishes have significant impacts on native fishes in Eastern Cape headwater streams. If management and conservation measures are implemented, the unwanted introduction and spread of non-native fishes may be restricted, allowing native fishes opportunities for recovery.
- Full Text:
- Date Issued: 2014
Electrospun fiber based colorimetric probes for aspartate aminotransferase and I7ß-estradiol
- Authors: Pule, Bellah Oreeditse
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/54869 , vital:26623
- Description: Fabrication, characterization and application of electrospun polymer composite based colorimetric probes are presented in this thesis. The first part of the thesis involved the development of a protocol for in situ reduction of gold trication (Au³+) into metallic gold atoms with sodium borohydride. The prepared PS-Au NPs showed an SPR band at 542 nm. Furthermore the absorbance of the colloidal Au NPs in polystyrene exhibited a good linear correlation (r2 = 0.9934) to E2 concentration in the range 5 to 50 ppb. The lowest naked eye detection limit was found to be 0.5 ppb and could further be easily monitored by UV-vis spectrophotometer. Upon interaction with E2 Au NPs aggregated to give nanoparticle clusters, confirmed through TEM analysis. Different concentrations of Au NPs were found to have a significant effect on the conductivity of the PS-Au NPs solution. At low concentrations of Au NPs (0.002, 0.015 and 0.025% w/v) PS-Au NPs solution could be electrospun without clogging. The FE-SEM images showed a non-beaded morphology of PS-Au NPs composite fibers. Upon interaction of the colorimetric probe strips with various E2 concentrations it was observed that with increasing E2 concentrations (50 ng/ml to 1000 µg/ml) the colour of the probe changed gradually from white to shades of pink and eventually to shades of blue at higher E2 concentrations. The visible cut-off concentration was 100 ng/ml. The second component of the thesis focussed on the development of diazonium dye-nylon 6 colorimetric probe for aspartate aminotransferase. At optimal pH 7.4 the enzyme was stable, highly active and catalyzed a reaction that was susceptible to detailed kinetic analysis by continuous optical methods. The KM values for L-aspartate, a- ketoglutarate and oxaloacetate were 2.60, 0.59 and 0.066 mM, respectively. On the basis of these KM values the solid-state colorimetric probe was developed. A colour change occurred when an electrospun dye-N 6 probes were exposed to visibly detectable concentrations of oxaloacetate, an AST-catalyzed reaction product. While monitoring AST activity at 530 run, a linear relation was obtained between oxaloacetate concentrations ranging from 0.4 - 7.4 µg/ml. Naked eye detection limit of 2.4 µg/ml oxalaoacetate equivalence of 10 times the normal AST activity was attained. The colorimetric probe was in addition, tested against co-substrates aspartate, ketoglutarate and a variety of other compounds such as alanine, pryruvate, as well as glutamic, malaic and succinic acids known to interfere with AST activity. Each compound elicited a distinct and unambiguous colour change upon interaction with the colorimetric probe. Further X-ray powder diffraction (XRD), duNouy ring tensiometer, Brunauer- Emmett- Teller (BET) and energy dispersive X-ray spectroscopy (EDS/EDX) characterization confirmed composition and stability of the colorimetric probes. Colorimetric probes developed in this thesis are relatively cost effective, simple and "rugged" for measurement of analytes with visual detection without sample pretreatment in matrices, such as plasma and dairy effluents. The probes warrant further investigation as they have shown potential and offer a promising solid-state platform for both clinical diagnostics and environmental monitoring.
- Full Text:
- Date Issued: 2014
- Authors: Pule, Bellah Oreeditse
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/54869 , vital:26623
- Description: Fabrication, characterization and application of electrospun polymer composite based colorimetric probes are presented in this thesis. The first part of the thesis involved the development of a protocol for in situ reduction of gold trication (Au³+) into metallic gold atoms with sodium borohydride. The prepared PS-Au NPs showed an SPR band at 542 nm. Furthermore the absorbance of the colloidal Au NPs in polystyrene exhibited a good linear correlation (r2 = 0.9934) to E2 concentration in the range 5 to 50 ppb. The lowest naked eye detection limit was found to be 0.5 ppb and could further be easily monitored by UV-vis spectrophotometer. Upon interaction with E2 Au NPs aggregated to give nanoparticle clusters, confirmed through TEM analysis. Different concentrations of Au NPs were found to have a significant effect on the conductivity of the PS-Au NPs solution. At low concentrations of Au NPs (0.002, 0.015 and 0.025% w/v) PS-Au NPs solution could be electrospun without clogging. The FE-SEM images showed a non-beaded morphology of PS-Au NPs composite fibers. Upon interaction of the colorimetric probe strips with various E2 concentrations it was observed that with increasing E2 concentrations (50 ng/ml to 1000 µg/ml) the colour of the probe changed gradually from white to shades of pink and eventually to shades of blue at higher E2 concentrations. The visible cut-off concentration was 100 ng/ml. The second component of the thesis focussed on the development of diazonium dye-nylon 6 colorimetric probe for aspartate aminotransferase. At optimal pH 7.4 the enzyme was stable, highly active and catalyzed a reaction that was susceptible to detailed kinetic analysis by continuous optical methods. The KM values for L-aspartate, a- ketoglutarate and oxaloacetate were 2.60, 0.59 and 0.066 mM, respectively. On the basis of these KM values the solid-state colorimetric probe was developed. A colour change occurred when an electrospun dye-N 6 probes were exposed to visibly detectable concentrations of oxaloacetate, an AST-catalyzed reaction product. While monitoring AST activity at 530 run, a linear relation was obtained between oxaloacetate concentrations ranging from 0.4 - 7.4 µg/ml. Naked eye detection limit of 2.4 µg/ml oxalaoacetate equivalence of 10 times the normal AST activity was attained. The colorimetric probe was in addition, tested against co-substrates aspartate, ketoglutarate and a variety of other compounds such as alanine, pryruvate, as well as glutamic, malaic and succinic acids known to interfere with AST activity. Each compound elicited a distinct and unambiguous colour change upon interaction with the colorimetric probe. Further X-ray powder diffraction (XRD), duNouy ring tensiometer, Brunauer- Emmett- Teller (BET) and energy dispersive X-ray spectroscopy (EDS/EDX) characterization confirmed composition and stability of the colorimetric probes. Colorimetric probes developed in this thesis are relatively cost effective, simple and "rugged" for measurement of analytes with visual detection without sample pretreatment in matrices, such as plasma and dairy effluents. The probes warrant further investigation as they have shown potential and offer a promising solid-state platform for both clinical diagnostics and environmental monitoring.
- Full Text:
- Date Issued: 2014
The characterization of DNAJC3: elucidating the function of the TPR domains
- Mutsvunguma, Lorraine Zvichapera
- Authors: Mutsvunguma, Lorraine Zvichapera
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/55874 , vital:26751
- Description: DNAJC3 is a novel member of the DNAJ family with two domains linked to co-chaperone functions, namely the tetratricopeptide repeat (TPR) and J domain. Out of the two domains, the TPR domains are the least characterized. Therefore, the aim of this study was to characterize and elucidate additional functions of DNAJC3 TPR domains through in silico, in vitro and ex vivo approaches. Through multiple sequence and structural alignment as well as electrostatic potential analysis, DNAJC3 TPR domain were found to be most similar to TPR-containing proteins with Hsp90 or Hsp70 independent functions. In vitro pull down assays illustrated that DNAJC3 TPR domains did not interact with either cytosolic Hsp90 and Hsp70 or Grp78 and Grp94 directly, however a potential indirect interaction with Grp94 and Hsp90 was observed in mammalian lysates, via pull down assays; suggesting the formation of a complex between the proteins mediated by a specific substrate. DNAJC3 TPR domains were found to bind indiscriminately to both native and heat denatured substrates in a dose dependent manner. DNAJC3 TPR domains bound to β-galactosidase with greater affinity than malate dehydrogenase (MDH), suggesting that DNAJC3 TPR domains might exhibit substrate specificity that has not been reported before. Preliminary ex vivo analysis of DNAJC3 in mammalian cells showed that induced stress conditions did not alter the cytosolic or endoplasmic reticulum (ER) localization, or levels of DNAJC3 protein, suggesting that the protein is not stress inducible. However, protein levels of DNAJC3 were dramatically reduced by Hsp90 inhibitor novobiocin at 500 μM. Transient knockdown DNAJC3 did not change the protein levels of either Grp78 or Grp94, but decreased the protein levels of Hsp70/Hsp90 organizing protein HOP. On the other hand, protein levels of DNAJC3 were increased in HOP depleted cells. In conclusion, this study was the first to experimentally demonstrate that DNAJC3 TPR domains do not interact directly with Hsp90, Hsp70, Grp78 or Grp94, and therefore DNAJC3 is unlikely to participate in traditional co-chaperone interactions with those proteins via its TPR domain. However, the J domain is known to interact with Grp78. The discovery that DNAJC3 TPR domains resemble that of TPR-containing proteins with functions independent of Hsp90 or Hsp70 suggests that DNAJC3 might link the Hsp70/Grp78 chaperone machinery to non co-chaperone related functions, which requires further analysis.
- Full Text:
- Date Issued: 2014
- Authors: Mutsvunguma, Lorraine Zvichapera
- Date: 2014
- Language: English
- Type: text , Thesis , Doctoral , PhD
- Identifier: http://hdl.handle.net/10962/55874 , vital:26751
- Description: DNAJC3 is a novel member of the DNAJ family with two domains linked to co-chaperone functions, namely the tetratricopeptide repeat (TPR) and J domain. Out of the two domains, the TPR domains are the least characterized. Therefore, the aim of this study was to characterize and elucidate additional functions of DNAJC3 TPR domains through in silico, in vitro and ex vivo approaches. Through multiple sequence and structural alignment as well as electrostatic potential analysis, DNAJC3 TPR domain were found to be most similar to TPR-containing proteins with Hsp90 or Hsp70 independent functions. In vitro pull down assays illustrated that DNAJC3 TPR domains did not interact with either cytosolic Hsp90 and Hsp70 or Grp78 and Grp94 directly, however a potential indirect interaction with Grp94 and Hsp90 was observed in mammalian lysates, via pull down assays; suggesting the formation of a complex between the proteins mediated by a specific substrate. DNAJC3 TPR domains were found to bind indiscriminately to both native and heat denatured substrates in a dose dependent manner. DNAJC3 TPR domains bound to β-galactosidase with greater affinity than malate dehydrogenase (MDH), suggesting that DNAJC3 TPR domains might exhibit substrate specificity that has not been reported before. Preliminary ex vivo analysis of DNAJC3 in mammalian cells showed that induced stress conditions did not alter the cytosolic or endoplasmic reticulum (ER) localization, or levels of DNAJC3 protein, suggesting that the protein is not stress inducible. However, protein levels of DNAJC3 were dramatically reduced by Hsp90 inhibitor novobiocin at 500 μM. Transient knockdown DNAJC3 did not change the protein levels of either Grp78 or Grp94, but decreased the protein levels of Hsp70/Hsp90 organizing protein HOP. On the other hand, protein levels of DNAJC3 were increased in HOP depleted cells. In conclusion, this study was the first to experimentally demonstrate that DNAJC3 TPR domains do not interact directly with Hsp90, Hsp70, Grp78 or Grp94, and therefore DNAJC3 is unlikely to participate in traditional co-chaperone interactions with those proteins via its TPR domain. However, the J domain is known to interact with Grp78. The discovery that DNAJC3 TPR domains resemble that of TPR-containing proteins with functions independent of Hsp90 or Hsp70 suggests that DNAJC3 might link the Hsp70/Grp78 chaperone machinery to non co-chaperone related functions, which requires further analysis.
- Full Text:
- Date Issued: 2014
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