The cis and trans signals important in BPV replication were identified using a transient replication assay, the mobility shift assay, and a comparison between the BPV and LPV genomes. Replication of deleted BPV genomic clones, which contain the natural left (3’ OH end of the viral minus strand) and right (5’ PO, end of the viral minus strand) BPV termini, defined the minimum size of the BPV origin of replication (ori) to be the terminal 171 nucleotides of each terminus. Clones containing duplicate termini or altered left ends were also shown to replicate. The BPV ori was determined to have two domains identified by a computer analysis of homologus regions between these termini. Three proteins were identified that bind to the left terminal 171 nucleotides in the hairpin conformation. Inhibition of the formation of the DNA-protein complexes with competitor DNA localized two potential binding sites that correspond to the domains mentioned above. Two of the DNA-protein complexes were formed by BPV-coded proteins as determined by inhibition of the complex by anti-BPV antibodies. The third complex resulted from binding of a host cell S-phase protein that is a likely candidate for the S-phase factor required for autonomous parvovirus replication. The BPV ori thus appears to function by binding both cellular and viral proteins for the initiation of DNA synthesis from the hairpinned termini. The comparison of the BPV and LPV genome sequence suggest that the genomic organization of LPV may be more like BPV than that of the rodent parvovirus minute virus of mice; and therefore, LPV may contain similar cis signals. / Ph. D.

Mannheimia haemolytica is an opportunistic pathogen in cattle and the main bacterial agent in bovine respiratory disease. Despite its economic importance, few studies have characterized the genetic diversity of M. haemolytica, particularly from feedlots. Three genotyping techniques (BOX-PCR, (GTG)5-PCR and PFGE) were compared to discriminate M. haemolytica and strains from the family Pasteurellaceae. PFGE was the most discriminating and repeatable, although BOX-PCR was most accurate in clustering isolates together according to species. Mannheimia haemolytica was isolated from nasal swab samples collected from cattle upon entry and exit from two feedlots in southern Alberta. These were characterized by PFGE and antimicrobial susceptibility using a disk-diffusion assay. Select gene determinants were screened for using PCR. PFGE analysis revealed the isolates to be highly diverse. Ten percent of the isolates exhibited resistance. At present, the development and spread of antimicrobial resistance in M. haemolytica observed within the feedlots examined appears to be low. / xi, 116 leaves : ill. ; 29 cm