The etiological agent of rabies encephalitis belongs to the genus Lyssavirus in the Rhabdoviridae family. Lyssaviruses are negative sense, single stranded RNA viruses and cause an estimated 55 000 human deaths per year with 44% of these deaths occurring in Africa (WHO, 2005). With intense research effort and increased sequence information it is becoming evident that the Lyssavirus genus is much more diverse than initially thought and therefore diagnostic methods need to be modified accordingly. The African continent sustains a diverse variety of lyssaviruses, however, most countries in Africa do not have active surveillance or necessary diagnostic tools and therefore rabies-related lyssaviruses are underreported. Previous studies have indicated that real-time PCR has improved sensitivity and rapidity over conventional molecular diagnostic methods with the added advantage of allowing accurate estimations of viral load in a wide variety of samples. Several realtime PCR assays have been developed; however, none were specifically aimed at detection of lyssaviruses present on the African continent. This study was therefore aimed at evaluating certain molecular diagnostic methods for the detection of African lyssaviruses. Furthermore, the application of real-time PCR for various fields in lyssavirus research i.e. diagnostics, surveillance and pathogenicity studies were evaluated. This study revealed two different hemi-nested PCR assays capable of detecting representatives of African lyssaviruses. A real-time PCR was developed that was successful for the detection of African lyssaviruses. In addition, a quantitative assay and internal control was successfully employed for confirming ante-mortem human rabies diagnosis as well as post-mortem animal rabies diagnosis in formalin fixed brain material. As such the real-time PCR assay developed in this study could therefore be routinely used for ante-mortem diagnosis and as a confirmatory test for post-mortem diagnosis. The ability of this assay to detect and quantify all currently known African lyssaviruses not only offers improved surveillance capacity, but offers unique potential as a sensitive tool to track virus movement in pathogenicity studies. These aspects are important in our search for a better understanding of the complex epidemiological and viral characteristics of African lyssaviruses. Copyright / Dissertation (MSc)--University of Pretoria, 2010. / Microbiology and Plant Pathology / unrestricted

La faune sauvage a été depuis longtemps incriminée dans la survenue de zoonoses et joue le rôle de réservoir d'agents pathogènes (virus Nipah, Hendra, Ebola, Hantaan etc.) pour l'homme. Les îles tropicales et subtropicales du Sud-Ouest de l'Océan Indien (SOOI) constituent l'une des 34 régions reconnues comme « hotspot » de biodiversité au niveau mondial. Elles sont caractérisées par un très fort endémisme de la faune sauvage surtout sur l'Ile de Madagascar. Le caractère multi-insulaire de la région du SOOI, la diversité de ses biotopes et ses disparités biogéographiques et humaines offrent un champ d'investigation unique pour explorer « in natura » la dynamique évolutive des agents infectieux et les relations hôtes-virus. Nos travaux de recherche ont porté sur deux modèles de virus à ARN de polarité négative, les paramyxovirus et les lyssavirus. Le premier modèle viral nous a permis d'aborder les questions relatives à la dynamique de transmission virale au sein de communauté d'hôtes, plus particulièrement, les chauves-souris et les petits mammifères terrestres de Madagascar et d'identifier les facteurs agissant sur cette dynamique de transmission et de diversification virale, en particulier les facteurs bio-écologiques associés à leurs hôtes. Le second modèle viral, les lyssavirus, nous a permis de décrire sur l'ensemble des îles du SOOI échantillonés, la circulation virale dans ce système multi-insulaire diversifié, au sein des chauves-souris dont la plupart des espèces sont endémiques à cette région. Dans l'ensemble, nos investigations ont permis de mettre en évidence des échanges viraux (« host-switch ») importants entre chauves-souris, petits mammifères terrestres endémiques de Madagascar et les rongeurs introduits, le rôle de ces mammifères en tant que réservoir viral majeur et souligner le rôle disséminateur de Rattus rattus. Par ailleurs, nous avons pu identifier ce phénomène de « host-switch » comme étant le mécanisme macro-évolutif prépondérant et l'importance des facteurs biotiques et abiotiques à l'origine de la dynamique de transmission et de la diversification virale observée chez les paramyxovirus de chauves-souris de Madagascar. / Since many decades, the wild fauna has been incriminated as an important reservoir of many zoonotic pathogens (Nipah, Hendra, Ebola, Hantaan viruses etc.) at risk for humans. Tropical and subtropical islands of the South West Indian Ocean (SWIO) are part of the 34 areas of the world recognized as "hotspot" of biodiversity. They are characterized by a strong wildlife endemism especially on Madagascar. The multi-island structure of the SWIO region, the diversity of its biotopes and its biogeographical human disparities, offer a unique opportunity to investigate "in natura" the evolutionary dynamics of infectious agents and the host-virus relationships. Our research has focused on two models of negative RNA viruses, paramyxoviruses and lyssaviruses. The first virus model allowed us to address issues related to the dynamics of viral transmission within a host community, in particular, bats and small terrestrial mammals of Madagascar and to identify the driving factors, especially bio-ecological factors associated with their hosts, affecting the dynamic of transmission and of viral diversification. The second model allowed us to describe on the islands of the SWIO, the intense circulation of bats lyssaviruses in this multi-island system which bats are endemic to this region. Overall, our investigations highlighted (i) intense viral exchanges ("host-switch") between bats, endemic terrestrial small mammals and introduced rodents from Madagascar, (ii) the role of these mammals as major viral reservoir and (iii) the key role played by Rattus rattus as viral spreader. Furthermore, we identified both the phenomenon of "host-switch" as the major macro-evolutionary mechanism among bat paramyxoviruses from Madagascar and the importance of biotic and abiotic factors in shaping the transmission dynamics and viral diversification.