L’intégrase (IN) du VIH-1 est une enzyme clé qui catalyse l’insertion stable du génome viral dans celui de la celluleinfectée. D’autre part, l’IN participe également à de nombreuses étapes du cycle viral telles que la transcriptioninverse ou la maturation virale. La compréhension des mécanismes impliqués dans la régulation de l’intégrationcellulaire au cours de l’infection est un enjeu important. L’IN fait partie du complexe de préintégration composé defacteurs cellulaires et viraux. La dynamique des interactions au sein de ce complexe régule les activités catalytiquesmais également non catalytiques de l’IN. C’est dans ce contexte de recherche de nouveaux cofacteurs de l’intégraseque nous avons identifié une interaction entre l’IN et la protéine Kinase GCN2.Mon travail de thèse s’est orienté sur trois questions autour de l’étude du rôle de cette interaction IN/GCN2.- Dans un premier temps, le rôle de la protéine kinase cellulaire GCN2 au cours du cycle viral a été étudié. Nousavons pu montrer que l’infection par le VIH-1 provoque un stress activant GCN2. Cette activation aboutit à uneinhibition de la traduction dès les premières heures de l’infection.- GCN2 est capable de phosphoryler l’IN du VIH-1 sur deux positions : les sérines en position 24 et 255. L’étude durôle de la phosphorylation de l’IN par GCN2 a permis de montrer que l’absence de phosphorylation de l’IN entraîneune stimulation de l’infection. GCN2 via la phosphorylation de l’IN a donc un effet restrictif sur l’infection par le VIH-1.- L’étude du domaine d’interaction entre l’IN et GCN2 a permis d’identifier un résidu essentiel de l’IN, l’acideglutamique en position 85 (E85). En effet, la mutation E85A de l’IN entraîne la production de virus non infectieux,suite à un défaut de maturation / HIV-1 integrase (IN) catalyzes the integration of the viral DNA into the cellular genome. Besides, IN is also involved inother steps of the viral life cycle like the reverse transcription or the viral maturation. Our group is interested in themechanisms involved in the regulation of cellular integration during the infection. IN is part of a pre-integrationcomplex composed of cellular and viral proteins. The dynamic of these interactions regulates IN activities but also noncatalytic activities such as nuclear import and tethering of the complex to the integration site. During theidentification of news partners of IN, an interaction between IN and the kinase GCN2 was characterizedMy PhD project is composed of 3 topics:- The role of the cellular kinase GCN2 was studied during the viral cycle. We showed that GCN2 is activated uponHIV-1 infection. This activation leads to a general decrease of cellular translation during the first hours of infection.-GCN2 is able to phosphorylate IN on two positions: the serines in position 24 and 255. The absence of INphosphorylation causes a stimulation of HIV-1 infection. We demonstrate that GCN2 affects the viral cycle via thephosphorylation of IN.- Binding domain analysis between IN and GCN2 led to the identification of a critical residue of IN, the glutamicacid in position 85 (E85). Mutations E85A of IN impair the viral production by inhibiting the viral maturation.

Denna studie handlar om elevers och lärares upplevelse av att arbeta med datorer i undervisningen inom ramen för ett nyligen infört en-till-en-projekt i en genomsnittlig svensk gymnasieskola år 2012. Syftet med undersökningen är att analysera och diskutera vilka för- och nackdelar lärare och elever upplevde i relation till det införda en-till-en-projektet år 2012. Studien avser vidare att diskutera vilka bakomliggande faktorer som bidrog till de problem som upplevdes, och hur detta kan relateras till den tidigare forskning på området som fanns att tillgå vid tidpunkten för studiens genomförande samt till dagens forskning. Det empiriska materialet för undersökningen består av en enkätundersökning bland fyra klasser och två lärare, samt uppföljande intervjuer med nämnda lärare samt med datoransvarig på skolan. Resultaten visar att eleverna är relativt nöjda med en-till-en-projektet. Lärarna håller i stort sett med eleverna om vilka positiva aspekter som finns av en-till-en, men är sammantaget relativt missnöjda då de har vidare ambitioner som omöjliggörs av bland annat brister i tekniken, brister i organisationen inom skolan vad gäller datorerna, samt brister i deras egen kompetens. Elevernas relativt nöjdhet kan då förklaras med att de saknar lärarnas insikt, vilket visar att elevers självutvärdering av deras egen situation inte nödvändigtvis speglar kvaliteten på det som utvärderas. Resultaten från undersökningen är i stort i linje med den forskning som fanns att tillgå 2012, men denna forskning behöver kompletteras av oberoende aktörer då den utfördes på uppdrag av initiativtagarna till två stycken en-till-en-projekt med relativt hög mediaprofil.

The overall aim of this project was to investigate HIV-1 genetic diversity in South Afri ca and to characterise the immune response in mice to a South African subtype C gp120. To investigate the relationship between subtype and mode of transmission, samples were collected from individuals infected by heterosexual and male homosexual transmission from patients attending local HIV/AIDS clinics in Cape Town (n=49) and Bloemfontein (n=4). Isolates were subtyped using heteroduplex mobility assay (HMA) based on the V3-V5 region of the env geneusing reference plasmids (2 B, 2 C and 1 D) representative of local subtypes. HMA identified four env subtypes: A, B, C and D. Subtype B viruses were found in 92.9% (26/28) of the male homosexual/bisexual group and subtype C viruses in 77.2% (17 /22) of the heterosexual group. Subtype B viruses were also identified in two heterosexual patients, one patient infected by blood transfusion and in two patients with. unknown mode of transmission. Subtype D viruses were found in one male homosexual patient and one heterosexual patient and a husband and wife couple were infected with subtype A viruses. A significant association between subtype and mode of transmission (p=<0.0001) was identified, confirming two independent epidemics. To determine the subtype distribution of HIV within urban heterosexual populations throughout South Africa, samples were collected from women attending antenatal clinics in Johannesburg (n=34), Pretoria (n=S) and Durban (n=20). Samples from Bloemfontein (n=24) were taken from individuals attending an HIV/AIDS clinic. All eighty-three samples were subtyped by HMA in the env region as before. The predominant subtype circulating within the urban heterosexual population throughout South Africa was identified as subtype C (92.8%) although subtype B was also detected (7.2%). It may thus be beneficial if a HIV vaccine for South Africa is based on a subtype C model. In addition, a rapid method for identification of HIV-1 gag subtypes was developed based on restriction fragment length polymorphism (RFLP) analysis of 400bp (p17) or 650bp (p17 and 5' p24) long PCR fragments. This strategy was appl i ed to eighty-six samples (Cape Town n=47, Johannesburg n=20, Bloemfontein n=17 and Durban n=2) previously subtyped by either sequence analysis of the gag p17 region (n=31), heteroduplex mobility assay (HMA) based on the env gene (n=76), or both (n=21). RFLP analysis identified two subtype A, twenty-five subtype B, fifty-eight subtype C and one subtype D isolates. There were no discrepancies between RFLP and sequence gag subtypes, demonstrating the reliability of this method and no discordance between gag RFLP subtypes and env HMA subtypes, indicating no recombinant viruses in the genomic regions analysed.

With the use of contemporary design tools, the modes of representation and fabrication in architecture have started to blur and overlap. Can this overlapping alter the role of and potential for material and materiality in architecture? Applying a composite logic in terms of tools, techniques and materials, the project reflects upon the tools of the architect. It presents a full scale installation, exploring material processes and contrasting a dichotomist view on form and matter. / I och med användandet av nya verktyg för design och gestaltning är det inte längre tydligt var gränsen går mellan representation och fabrikation - inom arkiteturfältet finns idag en överlappning eller sammanblandning av dessa. Kan denna överlappning förändra begreppet materialitet inom arkitekturen? Genom att tillämpa blandade logiker vad gäller verktyg, tekniker och material reflekterar projektet över arkitektens olika verktyg. Det presenterades i form av en fullskale-installation.

Hematopoiesis is coordinated by a complex regulatory network of transcription factors among them PU.1 (Spi1, Sfpi1) and GATA-1 represent key molecules. GATA-1 and PU.1 bind each other on DNA to block each others transcriptional programs to prevent development of undesired lineage during hematopoietic commitment. Murine erythroleukemia (MEL) cells, transformed erythroid precursors that are blocked from completing the late stages of erythroid differentiation, co-express GATA-1 and PU.1 and as my and others data document, are able to respond to molecular removal (down-regulation) of PU.1 or addition (up-regulation) of GATA-1 by inducing terminal erythroid differentiation. We provide novel evidence that downregulation of GATA-1 or upregulation of PU.1 induces incompletely differentiation into cell cycle arrested monocytic-like cells. Furthermore, PU.1- dependent transcriptome is negatively regulated by GATA-1 in MEL cells, including CCAAT/enhancer binding protein alpha (Cebpa) and Core-binding factor, beta subunit (Cbfb) that encode additional key hematopoietic transcription factors. Chromatin immunoprecipitation and reporter assays identified PU.1 motif sequences near Cebpa and Cbfb that are co-occupied by PU.1 and GATA-1 in the leukemic blasts. Furthermore, transcriptional regulation of these loci by...

A study was conducted (November 2012) on the communal dipping system in Mnisi, Mpumalanga Province of South Africa to detect levels of blue tick resistance to commonly used acaricides. The larvae obtained from engorged females of the one host tick Rhipicephalus (B). microplus from twelve communal dipping areas were tested against various concentrations of amitraz, chlorfenvinphos and cypermethrin using the Shaw Larval Immersion Test method. Only R. (B). microplus ticks were identified from all sample areas, indicating a displacement of the indigenous R. (B). decoloratus tick in this area. Resistance testing using the Shaw Larval Immersion Test showed that no resistance to chlorfenvinphos was detected at any of the dip tanks, which was in keeping with the absence of known use of this product in the area. An important finding was the rapid development of resistance to the pyrethroids, which had only been in use for four months prior to conducting the study. Only one area (Hlalakane) yielded a R(B).spp population that was wholly susceptible to all three compounds. Resistance to amitraz was variable, with half (six out of 12) of the dip tanks comprising susceptible R(B).spp populations and two dip tanks with emerging resistance to amitraz. Possible risk factors which caused the resistance problems are discussed and acaricide management strategies recommended. / Dissertation (MSc)--University of Pretoria, 2015. / tm2016 / Veterinary Tropical Diseases / MSc

The Mnisi community is in the north-eastern corner of the Bushbuckridge Municipal Area, Mpumalanga Province, South Africa. This community is located at the livestock/wildlife interface sharing borders with several game reserves, and livestock are likely to be exposed to diseases with a wildlife reservoir, such as Corridor disease. Known tick vectors of important diseases such as Corridor disease, redwater, heartwater and anaplasmosis are present in the area. Although the farmers frequently dip their cattle in acaricide-filled dip tanks to control the tick burden, tick-borne diseases (TBDs) are still a major problem. This study was undertaken to determine if the symptoms of cattle in poor health in the Mnisi community could be attributed to TBDs. Corridor disease has previously been identified in cattle in the Mnisi community. Recent experimental studies have shown that T. parva DNA can be detected in infected cattle that survive the disease in the field. An additional aim of the study was therefore to identify T. parva carrier cattle in the area, and to search for evidence of selection of cattle-adapted T. parva parasites in carrier cattle. The study was conducted from July 2012 to June 2013. During the study period, samples from clinically sick cattle suspected of TBDs were collected to determine the cause of their symptoms. Blood smears from the clinically sick cattle were analysed using light microscopy while some cases were subjected to histopathology and T. parva-specific quantitative real-time polymerase chain reaction (qPCR). DNA extracted from blood samples and in some cases tissue samples collected from clinically sick cattle (n=137) was tested for the presence of haemoparasite DNA using the reverse line blot (RLB) hybridization assay. To identify T. parva carrier cattle, records from Hluvukani Animal Clinic and Bushbuckridge State Veterinary office were scrutinized to identify herds that may have been exposed to T. parva infection. Blood samples (n=670) were collected from herds that had recorded Corridor disease cases in the past three years, as well as herds that may have shared grazing with buffalo from the Kruger National Park and surrounding private game reserves. The indirect fluorescent antibody test (IFAT) was used to check for T. parva antibodies. Seropositive herds were revisited, as well as herds that had confirmed Corridor disease cases during the study period, and blood samples were collected (n=432). DNA extracted from these samples was screened for the presence of T. parva DNA using the T. parva-specific qPCR. In an attempt to find evidence of selection of cattle-adapted T. parva, the p67, p104 and PIM parasite genes were amplified from qPCR positive samples, and the amplicons were cloned and sequenced. Out of the 137 clinical disease cases examined from the study area, 24 cases of TBDs were diagnosed, of which 19 were Theileria related. The RLB hybridization assay confirmed the presence of tick-borne haemoparasites in the Mnisi community: 89 of the 137 clinical disease cases (65.0%) were found positive for one or more haemoparasite (Theileria, Babesia, Anaplasma and/or Ehrlichia species) while 48 (35.0%) were negative or below the detectable limit of the test. IFAT results indicated that there is a high seroprevalence of theileriosis (63.6%) in the Mnisi community area, but this may be due to cross reactions with other Theileria parasites known to be present (e.g. T. taurotragi). Fewer cattle (13.4%) were seropositive at the highest titre tested (160), and these are most likely to be associated with T. parva. In DNA extracted from blood samples from these seropositive herds, the T. parva-specific qPCR detected T. parva in eleven samples (2.6%). Eight of the eleven cattle were re-sampled six months later, but only one was still qPCR positive. All of the p104 and PIM sequences and two of the three p67 sequences were characteristic of buffalo-type T. parva alleles previously identified, implying that the T. parva infections in the cattle were transmitted directly from buffalo to cattle, and providing no evidence of selection of cattle-type alleles in the carrier animals. The study revealed that TBDs are a problem in the Mnisi community and surrounding area. Most important of the TBDs identified was Corridor disease, a notifiable disease in South Africa, which was the cause of most deaths among the cattle that were sampled. There was no evidence for the selection of cattle-derived T. parva alleles in any of the samples from T. parva carrier cattle, but a p67 sequence obtained from a clinical case was closely related to previously-identified alleles from cattle-derived isolates. Theileria parva DNA could only be detected in carrier cattle for a limited time post-exposure, suggesting that the infection will be cleared in infected animals before larvae or nymphs are available to pick up infections the following season. However, one bovine was still qPCR positive six months post-exposure, albeit with a very high Cp value (indicating a very low parasitaemia). The selection of T. parva parasites in cattle from the diverse T. parva population in African buffalo, therefore, remains a concern in the Mnisi community area, and at other livestock/wildlife interfaces in South Africa, but the risk is probably very low. / Dissertation (MSc)--University of Pretoria, 2015. / tm2016 / Veterinary Tropical Diseases / MSc

The South African poultry industry has been beset by sporadic H6N2 avian influenza infection (sub-lineage I and II) in chickens since the early 2000s, with economic losses resulting from reduced egg production and co-infection with other pathogens. An egg-based inactivated H6N2 vaccine (AVIVAC® AI; Deltamune (Pty) Ltd.) based on a 2002 sub-lineage I isolate is available, although substantial antigenic drift has occurred in H6N2 viruses since its implementation. Globally, seasonal and pandemic plant-produced hemagglutinin (HA)- based influenza virus-like particle (VLP) vaccines are in advanced clinical trials with proven efficacy, speed of production, cost-effectiveness, scalability and safety, although not yet established for poultry. In this study, H6 avian influenza VLPs (sub-lineage I and II, respectively) were transiently produced in Nicotiana benthamiana and tested for protective efficacy in the target host. A production platform has been established for H6 VLPs in N. benthamiana by optimising protein expression and purification to maximize yield and by assessing the feasibility of large-scale production and downstream processing in a preliminary study. Subsequently, the respective plant-produced H6 VLPs were formulated into vaccines and their capacity to reduce viral replication and shedding upon challenge with a 2016 H6N2 field isolate were established in specific-pathogen-free (SPF) chickens, in comparison to the commercial H6N2 vaccine. The plant-produced sub-lineage I VLP vaccine (768 HA units/dose) was highly immunogenic (mean hemagglutination inhibition (HI) titer 10.7 log2), reduced the oropharyngeal and cloacal viral shedding by more than 100- and 6-fold, respectively, and shortened the duration of oropharyngeal shedding by at least a week in comparison to the non-vaccinated control. Due to initial low yield of sub-lineage II VLPs, the maximum antigenic mass vaccine dose (48 HA units/dose)) resulted in substantially lower HA-specific antibody titers (mean HI titer > 4 log2), but still reduced viral shedding from the oropharynx by more than 5-fold in comparison to the non-vaccinated control. In contrast, the commercial vaccine not only failed to effectively reduce shedding in comparison to the non-vaccinated control, but exacerbated oropharyngeal shedding until day 21 after viral challenge, illustrating the antigenic dissimilarity between the commercial vaccine and a recent field virus. Plant-produced VLP vaccines, which facilitates differentiation between infected and vaccination animals (DIVA), presents a new generation of poultry vaccines that is highly efficacious and cost-effective with the major advantage of producing a tailored antigenically-matched vaccine candidate within a short space of time and holds enormous potential for the poultry industry. / Thesis (PhD)--University of Pretoria, 2020. / Production Animal Studies / PhD / Unrestricted

Animal diseases have always been one of the main constraints on animal production, especially in Africa where there are a variety of tropical and subtropical diseases. Knowledge of these diseases and the development of approaches to combat them is highly relevant to the socio-economic development of Africa and its fight against poverty. Serological tests were performed to determine seroprevalence and important risk factors for occurrence of respiratory pathogens in cattle on 423 biobanked sera collected from cattle at 11 dip tanks in the Mnisi communal farming area which is on the edge of the Kruger National Park. These pathogens are known to cause significant production losses in livestock by predisposing animals to secondary infections including pneumonia. A pentavalent, indirect ELISA test was performed to estimate seroprevalence of bovine herpesvirus-1, bovine respiratory syncytial virus, bovine viral diarrhea virus, parainfluenza virus-3 and bovine adenovirus-3 infections in cattle at the wildlife-livestock interface in the Mnisi communal farming area. Previous exposure to the five pathogens was determined. Additionally, the data was analyzed using the statistical software R to determine important risk factors that predicted exposure to the pathogens in cattle, namely population factors (distance from interface and month of collection) and individual characteristics (age, sex, body condition and breed). Age and body condition of the animals were found to have an effect on seropositivity while breed, sex, spatial distribution of the animals and month of sample collection did not have an effect. Recommendations to reduce pathogen exposure and improve production are made to the livestock owners in the Mnisi community. / Dissertation (MSc)--University of Pretoria, 2016. / Veterinary Tropical Diseases / MSc / Unrestricted

Anaplasmosis, babesiosis, and heartwater are the three most important tick-borne diseases of cattle in South Africa and result in a large number of mortalities. Endemic stability contributes to disease control, but little is known about the conditions required for maintenance of endemic stability. Through the on-going Health and Demographic Surveillance System in Livestock in the study area of the Mnisi One Health Platform, Mpumalanga, a great deal of information is being collected about cattle in the area, with the eventual aim of developing mathematical models to describe and predict infections. More than 15000 cattle have been identified for tick burden assessment, serological analyses and parasite identification. However, little is known about the time-course of infection of cattle with various tick-borne haemoparasites. Therefore, this study aimed to investigate the time-course of infection in new-born calves (n=10) and the presence of haemoparasites in adult ticks over a one year period using reverse line blot (RLB) hybridization and quantitative polymerase chain reaction assays. Blood samples and adult ticks were collected monthly from new-born calves in two areas of the Mnisi communal area: five located in a peri-urban area and five at the wildlife/livestock interface. A total of 119 blood samples and 805 adult ticks were collected. The RLB results confirm the exposure of most new-born calves in the Mnisi communal area to non-pathogenic and pathogenic tick-borne haemoparasites in the genera Anaplasma, Babesia, Ehrlichia and Theileria in their first year of life. A total of 805 adult ticks were identified to species level using identification keys and molecular methods. Only two tick species, Amblyomma hebraeum and Rhipicephalus microplus, were found on the calves during the year. Non-pathogenic and pathogenic haemoparasites in the genera Anaplasma, Babesia, Ehrlichia and Theileria were detected in pooled DNA extracted from ticks that had digested their blood meal. Pathogen-specific qPCR results indicated that some of the pathogens could not be detected in the calves until six to seven months of age and A. marginale was not detected at all in three calves at the wildlife/livestock interface. These calves were either infected at levels below the detection limit of our assays, or they were not infected at all. If the latter, it is possible that exposure to related non-pathogenic haemoparasites might help to establish and maintain endemic stability. Factors such as cattle density and dipping methods within different areas in the Mnisi communal area may play a role in the number of infected tick vectors in an area, and thus in the time-course of infection in new-born calves. It is clear that detailed information for cattle in different localities in the Mnisi communal area will be required in order to build accurate mathematical models to describe and predict infections. / Dissertation (MSc)--University of Pretoria, 2019. / Veterinary Tropical Diseases / MSc / Unrestricted