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African horse sickness (AHS) is a lethal disease of horses. The aetiological agent is African horsesickness virus (AHSV) (genus Orbivirus; family: Reoviridae). Immunity to all nine serotypes is needed for full protection. The AHSV virion is composed of seven structural proteins and 10 dsRNA genes. The outer capsid protein, VP2, determines the serotype and elicits protective neutralising antibodies (NAbs). The existing polyvalent attenuated vaccine has some drawbacks. The most important ones are the exclusion of serotypes 5 and 9. Also, vaccinated and naturally infected horses cannot be differentiated. This impedes the international movement of horses. Recombinant su.bunit vaccines should solve both problems. Previously, it was established that baculovirus-expressed AHSV VP2 induces NAbs and partial protection. The main aim of this investigation was to improve the level of protection and develop a prototype VP2-based AHSV -5 vaccine to supplement the current attenuated vaccine and pave the way for developing a VP2-based subunit vaccine that incorporates all nine AHSV serotypes. Preliminary challenge experiments in horses indicated that AHSV -5 rVP2 provides incomplete protection against a lethal AHSV-5 challenge. Probable causes and solutions for this problem had to be sought. Analysis of the baculovirus-expressed AHSV-5 VP2 showed that most (ca. 90 %) is aggregated and that only the soluble part (ca. 10 %) elicits NAbs in guinea pigs and full protection in horses. Preliminary small-scale production studies indicated that solubility of the AHSV-5 rVP2 could be improved considerably by optimising in vitro infection conditions. A significant finding was that the safety and efficacy of soluble AHSV -5 rVP2 is determined by the adjuvant used. Saponin-based adjuvants rendered the best results, albeit with dose-related side effects in horses. Saponin Q was tolerated best. These results for the first time implicated AHSV VP2 in stimulating a protective cell-mediated immunity. Based on these results, it is recommended that a candidate AHSV-5 VP2-based subunit vaccine, consisting of 50 µg rVP2 and 3.0 mg Saponin Q for the primary immunisation followed by a 50 µµg rVP2 and 0.6 mg Saponin Q booster, be formulated for the purpose of field trials. The knowledge generated during this study, combined with the recent cloning of the VP2 genes of all nine AHSV serotypes, provides a route for the development of a complete recombinant vaccine that will offer protection against all nine AHSV serotypes and could well free the restraint on the import and export of horses to and from South Africa. / Thesis (PhD (Microbiology))--University of Pretoria, 2005. / Biochemistry / unrestricted

Please read the abstract in the section 00front of this document / Dissertation (MSc (Genetics))--University of Pretoria, 2006. / Genetics / unrestricted

The neuroparalytic syndrome called botulism is caused by the neurotoxins produced by bacteria in the genus Clostridium. There are seven toxigenic types of C. botulinum (A to G) based on antigenically distinct toxins produced by different strains of the organism. Animal botulism is caused by C. botulinum type C and D neurotoxins and has a severe economic impact on cattle farming in South Africa and neighbouring countries. Current treatment regimes include the use of acetylcholine for symptomatic treatment, but this is unfortunately very seldom successful. All indications are that there is no cure for this disease and that effective control can only be achieved through development of efficacious vaccines. The botulinum vaccine currently in use in South Africa contains an adjuvanted toxoid form of the type C and o neurotoxins. However, this bivalent vaccine relies on problematic anaerobic cultivation of the Clostridium bacterium followed by isolation, purification and inactivation of the toxin by treatment with formalin. Apart from the fastidious growth requirements of this organism, it has been reported that the production of toxin by these cells declines rapidly and eventually ceases, following laboratory passaging of the bacterial cultures. In addition, improper inactivation of the toxins may also lead to the demise of animals following vaccination. Thus, there exists a great need for a safe, effective and inexpensive vaccine against botulism. To investigate the potential of types C and D botulinum neurotoxins as efficacious recombinant vaccine candidates against botulism, full-length copies of the genes were obtained by polymerase chain reaction (PCR) amplification from bacteriophage DNA isolated from Clostridium botulinum type C (Stockholm) and D (South Africa) cultures. The full-length genes were cloned and subsequently sequenced to verify their integrity. By making use of PCR-based site-directed mutagenesis procedures, three amino acid mutations were introduced in the zinc-binding motif of the respective neurotoxins. Mutation of this domain has previously been reported to successfully detoxify type C neurotoxin. The wild-type and mutant genes were subsequently expressed in insect cells using the BAC-to-BAC™ baculovirus system. Although, unique protein bands corresponding to the size of the neurotoxins could not be seen in Coomassie brilliant blue-stained gels, Western blot analysis indicated immunoreactive material for wild-type and mutant type C corresponding to the size of the type C toxin light chain. However, there was no conclusive evidence to support the successful expression of the full-length wild-type and mutant type D genes. / Dissertation (MSc (Microbiology))--University of Pretoria, 2006. / Microbiology and Plant Pathology / unrestricted

Thesis (MSc (Psychology))--University of Stellenbosch, 2006. / HIV vaccines are currently being developed and tested worldwide. This thesis reports on a qualitative study that was conducted to determine the concerns and problems regarding participation in future HIV vaccine trials. The sample for the study was selected from a peri-urban township, Masiphumelele, in Cape Town, Western Cape province, South Africa. The HIV-prevalence rate in Masiphumelele is 25%. A total of 10 participants between the ages of 19 and 30 were recruited for the present study. All participants’ first language was Xhosa and seven of them had English as a second language. Owing to a language barrier, an interpreter assisted the interviewer in conducting the interviews in the preferred language of the participants. Participants were recruited by convenience sampling and were asked to participate in two semi-structured interviews, under confidential conditions. The first interview addressed knowledge regarding HIV/AIDS, HIV vaccines and HIV clinical trials. The second interview identified the concerns and problems participants had regarding participation in future HIV vaccine trials. The interviews were recorded, transcribed and entered into Atlas ti., a computer program that assists in the analysis of textual data. The analysis of the data focused on the content of participants’ concerns about barriers to participation and their perspectives on facilitators to participation. The data collected on concerns and problems which, may influence participants’ willingness to participate in future HIV vaccine trials, was divided into two overarching themes, namely, barriers to participation and facilitators to participation. The barriers to participation included physical symptoms, stigma and discrimination, trypanophobia, distrust, psychological distress, sexual disinhibition and family responsibilities. The facilitators to participation included altruism, own protection from HIV infection, hopefulness, medical incentives, determining of HIV status, acquisition of knowledge, and equal treatment of participants in the experimental group and the placebo control group resulting from a double-blinded randomised trial. The question of participants’, recruited in the present study, willingness to participate in a future HIV vaccine trial are discussed in terms of Bronfenbrenner’s (1979) theoretical work on ecological systems, the social learning theory and the Health Belief Model (HBM). These theoretical frameworks deal with individuals, their behaviour and their environment, and how these influence one another. The significance and future direction of this line of research helps to overcome the barriers to participation and enhance the facilitators to participation. Thus, the intended result of such efforts is to maximise individuals’ participation in future HIV vaccine trials.

Thesis (PhD)--Stellenbosch University, 2008. / ENGLISH ABSTRACT: Background South Africa is the country with the largest number of HIV infections in the world. As behaviour change initiatives have been suboptimal in curbing the spread of the pandemic, an HIV vaccine is likely to be an important development as a biological agent may circumvent some of the challenges of initiating widespread behaviour change. The development of an HIV vaccine will require several thousands of HIV negative participants who are at high risk of HIV infection to participate in HIV vaccine clinical trials. Before recruitment for such trials may begin, various scientific, ethical, and sociobehavioural issues need to be considered. One of the key sociobehavioural issues concerns the willingness of individuals at high risk of HIV infection to participate in HIV vaccine trials. However, a psychometric measure of willingness to participate (WTP) has not been constructed, and there is a paucity of theory to guide studies of WTP. Objectives The first objective of this study was to construct a psychometric measure of WTP in an HIV vaccine trial, and to derive the exploratory factor structure of the measure. The second objective was to examine the extent to which the Theory of Planned Behavior (TPB) could predict variance in WTP, and to determine whether the TPB was strengthened by the inclusion of mistrust of researchers, knowledge of HIV vaccines and HIV vaccine trials, altruism, and perceived risk of HIV infection as additional predictor variables. Methodology This study was a research survey with a cross-sectional design. A convenience sample of 399 participants was recruited from an urban-informal settlement near Cape Town. As 79 of the questionnaires were poorly completed, the final sample size was 320. To develop a measure of WTP in an HIV vaccine trial, an item pool was developed whereby items directly reflected inhibitors and facilitators of WTP. After an iterative process of refinement, the final scale consisted of 35 items and was named the Willingness to Participate Scale (WTPS). A principal component Kaiser normalised exploratory factor analysis (EFA) was conducted on the items that constituted the WTPS. This procedure was performed to identify latent factors which were informed by the items of the scale. To test the predictive capacity of the TPB and the additional predictor variables, a two-step linear hierarchical multiple regression analysis was performed. At step 1, the TPB variables were entered simultaneously. At step 2, the TPB variables along with the additional predictor variables were entered simultaneously. Results The WTPS demonstrated excellent internal consistency (α = 0.90) and initial construct validity, as evidenced by the presence of seven latent factors. The factors accounted for 53.15% of the variance in WTP and were: (i) Social approval and trust; (ii) Stigmatisation; (iii) Personal costs; (iv) Personal gains; (v) Personal risks; (vi) Convenience; and (vii) Safety. The TPB significantly accounted for 6.4% (R² = 0.06) of the variance in WTP [F(3, 316) = 7.16, p < 0.001], yielding a small effect size (ƒ2 = 0.06). The TPB, together with mistrust, knowledge of HIV vaccines and HIV vaccine trials, altruism, and perceived risk of HIV infection as additional predictor variables significantly accounted for 10.2% (R² = 0.10) of the variance in WTP [F(7, 312) = 5.06, p < 0.001], yielding a small to medium effect size (ƒ2 = 0.11). Subjective norms, mistrust of researchers, altruism, and perceived risk of HIV infection were significant independent predictors of WTP. Conclusion Against the backdrop of the study limitations, the results of this study provide initial support for the reliability and construct validity of the WTPS among the most eligible trial participants in the Western Cape of South Africa. The findings also suggest that the TPB may not be an appropriate theoretical framework for predicting WTP in an HIV vaccine trial in this context. Nonetheless, normative pressure by others, mistrust of researchers, altruism, and perceived risk of HIV infection may influence WTP in this population. Implications for future research are discussed. / AFRIKAANSE OPSOMMING: Agtergrond Suid afrika is die land met die hoogste getal HIV infeksies in die wêreld. Vir die ontwikkeling van 'n HIV entstof, sal daar vereis word dat duisende HIV negatiewe deelnemers, wat 'n hoë risko kans staan om HIV infeksie op te doen, moet deelneem aan 'n kliniese HIV vaksine proeftog. Verskeie, wetenskaplike, etiese en sosiale gedrags punte moet oorweeg word, voor die werwing van sulke proefnemings. Een van die hoof aspekte van sosiale gedrags punte is die bereidwillgheid van 'n individu om blootgestel te word aan die HIV infeksie tydens die proeftog. 'n Psigometriese skatting van bereidwiligheid om deel te neem (BODTN) is egter nog nie gekonstruktureer nie en daar is 'n skaarste/geringheid in studies om as gids te dien vir die BODTN. Doel Die eerste doel van hierdie studie was om 'n psigometriese skatting van BODTN in ’n HIV vaksiene proefneming te konstruktureer, en om die ondersoekings oorsaak/faktor struktuur daarvan te meet en af te lei. Die tweede doel was om ondersoek in te stel of die omvang van die Teorie van Beplande Gedrag (TBG) verskille kan voorspel in die BODTN, en om vas te stel of die TBG versterk word deur die insluiting van wantroue in navorsers, kennis van HIV vaksienes en HIV vaksiene proefnemings, altruïsme, en die begrypbare risko van HIV infeksie as adisionele voorspellers. Metode Hierdie studie is 'n navorsings ondersoek met 'n deursneeproef ontwerp. ’n Grieflike aantal van 399 deelnemers was gewerf van 'n informele nedersetting naby Kaapstad. Die finale getal was 320 omdat 79 nie die vraelys korrek on volledig ingevul het nie. Na 'n interaktiewe proses van suiwering/verfyning, het die finale skaal uit 35 items bestaan en word die skaal benoem na die Willingness to Participate Scale (WTPS). Die prinsipale komponent Kaiser normaliseer EFA wat gedoen was op die items wat die WTPS konstitueer. Hierdie prosedure was gedoen om die latente faktore te identifiseer wat beskikbaar gestel was deur die items van die skaal. Om die voorspelbare kapasiteit van die TBG en die adisionele voorspelbare verskille te toets, het ons 'n twee stap hiërargiese veelvoudige agteruitgaan analise gebruik. By stap 1 was die TBG veranderlikes gelyktydig ingedruk. By stap 2 is die TBG veranderlikes tesame met die adisionele voorspellers in gedruk. Resultate Die WTPS het uitstekende interne konsistensie en 'n aanvanklike geldigheid gedemonstreer, soos bewys deur die teenwoordigheid van die 7 latente faktore. Die faktore verantwoord 53.15% van die verskil in WTP en was: (i) Sosiale aanvaarding en vertroue; (ii) Stigma; (iii) Persoonlike koste; (iv) Persoonlike wins/profyt; (v) Persoonlike risiko's; (vi) Gerieflikheid; en (vii) Veiligheid. Die TBG verantwoord 6.4% (R²=0.06) van die verskil in BODTN [F(3.316) = 7.16, p<0.001] met 'n toegewende klein groote uitwerking/uitslag. Die TBG tesame met wantroue, kennis van HIV vaksienes en HIV vaksiene proefnemings, altruïsme, en begrypbare risko van HIV infeksie as adisionele voorspellers, verwantwoord 10.2% (R²=0.10) van die verskil in BODTN [F(7.312 = 5.06, p<0.001], met 'n toegewende klein tot medium groote uitwerking/uitslag (f²=0.11). Subjektiewe norme, wantroue in navorsers, altruïsme, en 'n beprypbare risko van HIV infeksie was betekenisvolle, onafhanklike voorspellers van die BODTN. Gevolgtrekking Teen die agtergrond van die studie beperkinge, het die resultate van hierdie studie ondersteuning voorsien aan die vertroubaarheid en konstruktiewe geldigheid van die WTPS onder die mees geskikste proef deelnemers in die Wes Kaap van Suid Afrika. Die bevinding stel ook voor dat die TBG nie altyd 'n geskikte teoretiese raamwerk is vir die voorspelling van die BODTN in 'n HIV vaksiene proefneming in hierdie konteks is nie. Des nie teen staande, normale druk van ander, wanrtroue in navorsers, altruïsme en die begrypbare infeksie van HIV kan die populasie deur die BODTN beinvloed word. Implikasies vir toekomstige navorsing is bespreek.

Novel vaccine strategies for the presentation of immunologically important epitopes to the immune system are continuously being developed. Two such systems include the particulate protein and live viral vector delivery systems. In his study the long-term objective is to explore the African horsesickness virus (AHSV) serotype 9 viral protein 7 (VP7) and the Lumpy skin disease (LSDV) viral vector as two different vaccine strategies, particularly in view of the development of an HIV-1 vaccine. Consequently two very specific objectives were outlined in this study. The first was to express the HIV¬1 subtype C strain Du 151 gp41 epitopes ALDSWK and RVLAIERYLKD on the surface of the AHSV-9 VP7 particulate protein crystalline structures. A longer-term aim is to synthesise large quantities of these chimeric VP7 crystals in order to assess the immune response against the inserted epitopes. Secondly, the efficiency of the LSDV bi-directional promoter pA7LA8R in expressing chimeric VP7 proteins was to be evaluated by utilising the late element of this promoter to determine expression levels. Nucleotide sequences encoding the ALDSWK and RVLAIERYLKD epitopes were amplified from the HIV-1 subtype C strain Du 151 gp160 gene utilising PCR. These sequences were cloned individually as well as in combination into a multiple cloning site (549-566bp) present in the AHSV-9 VP7 gene. Recombinant pFASTBAC vectors PFASTBAC-VP7-MT 177-RVLAIERYLKD, PFASTBAC-VP7-MT 177-ALDSWK AND PFASTBAC-VP7-MT-177-RVLAIERYLKD-ALDSWK were identified, sequenced and used in the generation of recombinant baculoviruses utilising the BAC-to-BAC™ Baculovirus expression system. Expression of all three chimeric proteins, VP7-ALDSWK, VP7-RVLAIERYLKD and VP7- RVLAIERYLKD-ALDSWK was detected in infected Sf9 insect cells utilising SDS-PAGE. Further investigations will involve high-level expression of these proteins, which in turn will allow their characterisation as well as solubility, scanning electron and immunogenicity studies. In order to evaluate the efficiency of the LSDV bi-directional promoter, the AHSV-9 VP7 gene was cloned under the control of the late element (pA7L) of this promoter. The recombinant pHSsgpt-VP7 transfer vector was subsequently transfected into lamb testis cells infected with wild type LSDV in order to generate recombinant LSDV-VP7. Several rounds of recombinant virus selection in the presence of mycophenolic acid resulted in the loss of the LSDV-VP7 recombinant. Due to this unforeseen result, the expression of the VP7 protein from the late element of the pA7LA8R bi¬directional promoter could not be quantified and the efficiency of this promoter was not determined. The loss of LSDV recombinants, which contain a gene under the control of the late promoter element pA7L, has occurred previously and is suspected to be because of the instability of these recombinants. Due to the difficulties inherent in working with the LSDV viral vector delivery system, it has subsequently been decided to explore an alternate poxviral vector system. The focus in this study is now being shifted onto the promising Modified Vaccinia Ankara (MVA) viral vector system. / Dissertation (MSc (Genetics))--University of Pretoria, 2006. / Genetics / unrestricted

Rabies is a fatal but preventable neurotropic disease of potentially all mammals. The disease is caused by lyssaviruses. Rabies is recognized as the 10th most common lethal infectious disease in the world, rendering it one of the most feared zoonotic diseases known to man. Nevertheless, rabies can be prevented by application of pre- or post exposure treatments. Rabies vaccines have been available since the time of Pasteur, more that one hundred years ago. Since, vaccine research focused on the development of safer and more effective vaccines. Topics of current interest in the field of rabies vaccinology were addressed in this study. A primary concern regarding the disease is human mortalities, in the range of 60 000, reported every year. Most of these are linked to exposure to rabid dogs. In addition, a great number of post exposure treatments are administered each year at great costs. Despite availability of efficacious biologics, several factors influence the optimal use and accessibility of these agents in the countries of interest, with cost and availability being the major contributing factors. A proven approach is mass oral vaccination of target animals, such as dogs, which indirectly infers protection to susceptible hosts, including man. Currently available vaccines present several disadvantages of use though, including issues of safety or doubtful stability. Safer but effective alternative vaccines that could be used in oral baits would be valuable. Here the use of two candidate host restricted poxvirus vaccine vectors were explored, particularly also in regard to oral innocuity. The construction, convenient isolation and use of a recombinant Lumpy skin disease virus (Neethling strain) expressing rabies virus glycoprotein in a mouse model were investigated. In addition, a recombinant Modified Vaccinia virus Ankara expressing rabies virus glycoprotein was prepared and tested as a vaccine in mice, dogs and raccoons. In both cases it was clear that the severe attenuation of these viruses did affect the efficacy of the recombinant vaccines in the non-permissive hosts. With the recombinant MVA a clear dosage effect could be shown, and equivalent humoral responses could only be attained at much higher titers of vaccine virus as with replication competent counterparts. Secondly, the cross-protection of rabies vaccines across the spectrum of lyssaviruses was addressed. Lyssaviruses can be divided into two groups based on sequence analysis and pathogenesis. Viruses belonging to the so-called phylogroup II, are the Mokola, Lagos and West Caucasian Bat viruses. Classic rabies biologics fail to fully protect against the viruses attributed to a lack of cross-neutralization. Here, cross-protection and cross-reactive immune responses induced by recombinant vaccinia viruses expressing rabies, Mokola or West Caucasian Bat virus glycoproteins, in single or dual combinations, were investigated. As expected, there was a lack of cross-protection of rabies and Mokola glycoprotein vaccines. There was also a clear lack of cross-protection of West Caucasian Bat virus glycoprotein vaccine and rabies and Mokola viruses. The dual antigen expressing vaccines did not appear to offer any additional protective effect in the tested model. The Mokola virus glycoprotein vaccines induced neutralizing antibody responses that significantly cross-neutralized Lagos Bat virus. / Thesis (PhD (Microbiology))--University of Pretoria, 2006. / Microbiology and Plant Pathology / unrestricted

Since diffusion models of social network are widely used in studying epidemiology, in this thesis, we apply diffusion models to study the contact immunity generated by attenuated vaccines.Oral polio vaccine (OPV) is a typical attenuated vaccine for polio that can produce contact immunity and therefore help protect more individuals than vaccinees. / To better capture the utilization of OPV’s contact immunity, we model the community as a social network, and formulate the task of maximizing the contact immunity effect as an optimization problem on graphs, which is to find a sequence of vertices to be “vaccinated” to maximize the total number of vertices “infected” by the attenuated virus. Furthermore, as immune defiicient patients may suffer from the live attenuated virus in the vaccine, we develop models in consideration of this restriction, and study related problems. / We present polynomial-time algorithms for these problems on trees, and show the intractability of problems on general graphs. / 社交網絡的擴散模型被廣泛運用于對流行病學的研究，在本文中，我們使用擴散模型對減毒活疫苗產生的接觸性免疫進行研究。口服脊髓灰質炎疫苗（OPV）是一種典型的減毒活疫苗，它可以在人群中產生接觸性免疫，使得更多未接種疫苗的人獲得免疫力。 / 爲了更好的刻畫OPV 產生的接觸性免疫，我們將社區模型化為社交網絡，從而將接觸性免疫效應最大化的任務轉化爲圖優化問題，即通過發現頂點的一個「接種」序列來最大化被減活病毒「感染」的頂點數量。此外，因爲減毒疫苗中的活病毒會使患有免疫缺陷的病人患病，我們考慮在此因素限制下的模型，并研究相關的問題。 / 我們給出這些問題在樹上的多項式時間算法，并證明其在一般圖上的複雜性。 / Ma, Chenglong. / Thesis M.Phil. Chinese University of Hong Kong 2015. / Includes bibliographical references (leaves 40-47). / Abstracts also in Chinese. / Title from PDF title page (viewed on 12, September, 2016). / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only.

In the history of public health vaccines have proven to be among the most effective disease prevention tools. It is clear that in the fight against HIV that new and powerful preventive technology such as a vaccine is badly needed. Ethically, however the processes of developing a vaccine against HIV have been distinctly different from that of any previous pharmaceutical products. HIV vaccine trials can be ethically complex for a number of reasons. In 2004 the HIV I AIDS Vaccine Ethics Group undertook a research initiative that aimed to collect data from various South African stake holders of HIV vaccine trials to ascertain what they perceived as the ethical challenges related to HIV vaccine trials. A quantitative content analysis on the data from 31 semistructured interviews revealed that the ethical issue listed spontaneously by most of the respondents was that of informed consent. Further probing and discussion on informed consent identified a number of sub issues which the respondents thought would pose important challenges to HIV vaccine trials in the South African context. This study undertook to do a more in-depth qualitative analysis of the data to ascertain whether the challenges and concerns the stakeholders have are consistent with or different to those already identified in the literature and ethical guidelines on informed consent in medical research. What variables may be impacting on the position stakeholders take was also of interest. Results indicated that many concerns relating to the substantive and procedural elements of informed consent were consistent with those debated in the literature. These issues related to first person consent, the voluntariness of participants' consent, practicing cultural sensitivity, dealing with language issues, promoting and assessing understanding of material disclosed, issues around the vulnerability of .. participants, children and adolescents' capacity to consent and the role of the media. More specific to the South African context, stakeholders were concerned about the legal framework under which the trials take place, the general lack of education and training about HIV vaccine trials, a lack of communication and coordination between stakeholder groups, and the historical influences of apartheid on black South African participants' capacity to consent. The main variables that appeared to impact on the position stakeholders took related to the role the stakeholders play within the trials, the philosophical position underpinning their ethical viewpoints, stakeholders' understanding of vulnerability and capacity to consent, and how they view the universality or relativity of ethical issues. / Thesis (M.A.)-University of KwaZulu-Natal, Pietermaritzburg, 2008.