| Raccoon rabies is one of the most important zoonotic diseases in North America. The objective of this thesis was to characterize the virus and its variants and track the virus in raccoons following infection. The entire genome of a raccoon rabies virus (RRV) isolated in Canada was sequenced and compared to other lyssaviruses. This is the first report that describes the RRV polymerase (L) gene sequence, which encompasses half of the genome and encodes a 2128 amino acid product. The L gene includes an additional methionine immediately prior to the normal start codon due to a nucleotide substitution and three highly conserved coding regions, blocks II, III and IV. To gain insight into the incursion of RRV into three Canadian provinces, Ontario, New Brunswick and Quebec, a collection of 192 viral isolates, from throughout its North American range was characterized. Targeting the most variable region of the RRV genome, 119 variants were identified and phylogenetic analysis supported the history of RRV spread. In Canada, RRV outbreaks emerged from neighbouring U.S. States, with no evidence of viral movement between the three Canadian provinces. To study the extent of RRV infection in a Point Infection Control (PIC) area in Ontario, we developed a TaqMan real-time polymerase chain reaction (PCR) assay to detect RRV in brain tissues. Ten out of the 721 animals were positive for the RRV, while negative for the virus with the direct fluorescent antibody test. In order to trace how the RRV spreads in the infected host, raccoons were injected with the virus in the hind limb muscle and the animals were either euthanized on specific days following infection or served as controls. A TaqMan real-time RT-PCR assay was developed and used to analyze RNA. The RRV progressed from the muscle tissue into the central nervous system with an estimated velocity of 52.5-71.3 mm/day, resulting in a short incubation period in the raccoon host. Viral RNA could be detected 16 and 12 days, respectively, prior to the onset of the first clinical rabies symptoms, confirming the usefulness of real-time RT-PCR in early rabies diagnosis.;Keywords: Lyssavirus, rabies virus; raccoon rabies strain; complete genome sequence; L gene; G-L non-coding region; phylogenetic analysis; rabies management; TaqMan real-time RT-PCR; direct fluorescent antibody test; incubation period; viral quantification... |
