A thesis submitted to the Faculty of Health Sciences, University of the Witwatersrand, Johannesburg, in fulfilment of the requirements for the degree of Doctor of Philosophy Johannesburg 2017. / Introduction: Human rhinovirus (HRV) is the most prevalent virus detected in children with respiratory symptoms; however, its aetiological role during disease episodes remains unclear as detection of HRV is also ubiquitous among asymptomatic children. We evaluated the clinical epidemiology of HRV-associated disease among children hospitalised with severe and very severe pneumonia together with community control children living in Africa and Southeast Asia. In addition, we explored the associations between the molecular subtyping and nasopharyngeal viral loads of the HRV species and their ability to cause viraemia as potential markers for HRV disease. Methods: Using a case-control study conducted in seven countries, we compared the clinical characteristics of children (1-59 months of age) hospitalised with HRV-associated pneumonia between August 2011 - January 2014 and age-frequency matched controls. Nasopharyngeal swabs from the cases and controls were tested for HRV, together with 27 other respiratory pathogens, with quantitative real-time PCR assays. The 5’ NCR region of the HRV positive samples were sequenced to determine the species/strains of HRV and phylogenetic analysis was performed. Additionally, the blood samples from a limited number of cases (n=210) and controls (n=212) were tested for the presence of HRV viraemia and the 5’ NCR sequence of positive blood samples were further characterised. Results: Overall, HRV detection was 1.45-fold (aOR 95% CI: 1.29-1.62) higher among children hospitalised with pneumonia (24%) compared to controls (21%, P<0.005); including being 2.08-fold (28% vs 18%, aOR 95% CI: 1.75-2.47) more associated with case status among children 12-59 months of age. The HRV-associated cases were younger (13.1 months) than controls with HRV infection (15.4 months, P=0.001) and more likely to be malnourished (30% vs. 12%, P<0.001) and HIV-1 exposed (10% vs. 8%, P=0.046). HRV nasopharyngeal viral load was significantly higher among cases compared to controls (3.7 vs. 3.5 log10 copies/mL, P<0.001). Also, HRV viraemia was 7.02-fold (aOR 95% CI 1.70-28.94) more prevalent among cases (7%) compared to controls (2%, P=0.007). Moreover, HRV nasopharyngeal viral loads ≥4 log10 copies/mL differentiated between viraemia positive and negative cases. There was, however, no difference in the molecular subtyping of the HRV species prevalence among cases (HRV-A:48%; HRV-B:7%; HRV-C:45%) and controls (HRV-A:45%; HRV-B:10%; HRV-C:45%, P=0.496); as well as no evidence of seasonal or temporal clustering of the HRV species over time. Among cases, HRV detection was less likely to be associated with presence of radiographically confirmed pneumonia (40% vs 46%, P=0.001) or hospital stay >3 days (52% vs 61%, P=0.001). It was, however, positively associated with older age (13.1 months vs. 11.3 months, P<0.001) and presence of wheeze (46% vs. 31%, P<0.001) compared to the HRV uninfected cases. HRV was the sole virus detected in the 53% of cases and generally there were no differences in severity or clinical presentation among cases with HRV mono-infections compared to those with HRV-mixed infections. The HRV mono-infections, however, were associated with a 2.83-fold (aOR 95% CI: 1.44-5.53) higher case fatality ratio than cases with HRV and other viral mixed infections (10% vs. 5%, P=0.002). The HRV-associated case fatalities were more likely to have markers of bacterial co-infections compared to the HRV-associated cases that survived. Among the HRV species, HRV-C compared to HRV-A cases were older (12.1 vs. 9.4 months, P=0.033), more likely to present with wheeze (35% vs. 25%, P=0.031) and 2.59-fold (aOR 95% CI: 1.23-5.95) more likely to be associated with viraemia (12% vs. 2%, P=0.025). Conversely, the HRV-A infected cases were more likely to have radiographically confirmed pneumonia (46%) compared to HRV-C infected cases (36%, P=0.040) and HRV-A mono-infected cases were more likely to have hospital stay of >3 days (72%) than HRV-C mono-infected cases (54%, P=0.039). Conclusion: HRV detection, especially among children 1-5 years of age, was associated with severe lower respiratory tract infection; however, HRV detection was ubiquitous with a high degree of genetic diversity among both cases and controls. Thus the true etiologic role of HRV during childhood disease, especially among infants, remains uncertain. Nonetheless, HRV nasopharyngeal viral loads ≥4log10 copies/mL in conjunction with HRV viraemia are potential markers for HRV-associated severe respiratory disease. Among cases, HRV-A was associated with radiographically confirmed pneumonia and generally more severe disease than HRV-C which was more associated with viraemia and wheezing disease. / MT2017

Ce travail de pharmacochimie est consacré à la synthèse de nouvelles molécules benzéniques en vue d’étudier leurs propriétés pharmacologiques in vitro sur le Rhinovirus humain 14 mais également d’en déduire des relations de structure activité. En effet, le fil conducteur du projet de thèse est un travail de pharmacomodulation en collaboration avec le Rega Institute for Medical Research de Louvain. La méthode principale de synthèse de ces structures est basée sur la méthodologie TDAE ou Tétrakis(DiméthylAmino)Ethylène, appliquée sur des substrats dérivés du chlorure de nitrobenzyle. La synthèse et l’évaluation biologique de plus de 100 molécules a permis de décrire 5 hits dérivés du 4,5-diméthoxybenzène présentant des activités biologiques intéressantes in vitro (Concentration Effective Médiane de 1,5 à 4,3 μM ; index de sélectivité de 6 à 92). Les différentes stratégies adoptées lors de ce travail de pharmacochimie ont permis d’étendre l’étude des réactions par transfert monoélectronique sur de nouveaux substrats. Ainsi, le 1-(3-chloroprop-1-ynyl)-4-nitrobenzène a fait l’objet d’une étude en méthodologie LD-SRN1 avec des nitroalcanes, nitrocycloalcanes ainsi qu’avec des anions sulfinates ; ces travaux ont permis de décrire de nouveaux substrats insaturés originaux avec de bons rendements. De plus, la réactivité de ce substrat original a été évaluée en méthodologie TDAE avec des aldéhydes aromatiques. Ces travaux ont été valorisés par la synthèse de diarylbutynols originaux et ouvrent de nombreuses perspectives de recherche sur ce même noyau. / This pharmacochemistry work aims at synthesizing of new benzenic derivatives molecules with the scope to study both the chemical and antirhinoviruses 14 pharmacological properties in vitro. In fact, this work was focused on the pharmacomodulation of benzonitrile derivatives in collaboration with Rega Institute for Medical Research group. One hundred structural analogues were synthesized and a structure-activity-relationship was established. Biological assays showed five molecules with interesting anti-hRV 14 activities (EC50 from 1.5 to 4.3 and selectivity index from 6 to 92).These products derived from the TDAE-initiated reaction of various nitrobenzyl chloride analogues. Different strategies directed toward this pharmacochemistry project permitted to study single electron transfer (SET) reaction on original substrates. In this way, we explored the concept of LD-SRN1 on a propargylic chloride derivative such as 1-(3-chloroprop-1-ynyl)-4-nitrobenzene with nitronate and sulfinate anions. This latter compound also constitutes a potential substrate for the preparation of a propargylic anion using the TDAE strategy. So, we examined the use of TDAE methodology in alkyne series with various aromatic aldehydes in the presence of TDAE. These last works opened the scope of single electron transfer (SET) reactions.