Thesis (M. Phil.)--University of Hong Kong, 2008. / Includes bibliographical references (leaf 83-101) Also available in print.

Thesis (M. Phil.)--University of Hong Kong, 2008. / Includes bibliographical references (leaf 83-101) Also available online.

Thesis (M.Med.Sc.)--University of Hong Kong, 2002. / Includes bibliographical references (leaves 42-47). Also available in print.

Thesis (M. Phil.)--University of Hong Kong, 2005. / Title proper from title frame. Also available in printed format.

Thesis (Ph.D.)--McMaster University, 2004. / Advisor: John P. Capone. Includes bibliographical references (p. 179-[208]). Available also online.

Influenza A virus (IAV) can evolve from low virulence in animal hosts to become highly virulent in humans. Pandemic Influenza A viruses such as the 1918 Spanish Influenza caused over 50 million deaths worldwide. However the genetic determinants of IAV host adaptation and virulence are largely uncharacterized. The IAV NS1 protein is a multifunctional interferon antagonist and a known virulence factor. We hypothesized that NS1 mutations selected upon IAV evolution to a novel host contribute to host adaptation by mechanisms involving increased gene expression and IFN antagonism. To this end, I phenotypically characterized the NS1 mutations selected upon adaptation of A/Hong Kong/1/1968 (H3N2) (HK-wt) to increased virulence in the mouse. Sequencing the NS genome segment of mouse-adapted variants revealed eleven mutations in the NS1 gene and four in the overlapping NEP gene. Using the HK-wt virus and reverse genetics to express recombinant HK NS1 mutant viruses, I demonstrated that all NS1 mutations were adaptive and enhanced virus replication (up to 100 fold) in mouse cells and/or lungs. All but one NS1 mutant was associated with increased virulence measured by survival and weight loss in the mouse. Ten of twelve NS1 mutants significantly enhanced IFN-β antagonism to reduce the level of IFN-β production relative to HK-wt in infected mouse lungs at 1 day post infection, where nine mutants induced viral yields in the lung that were ≥ HK-wt (up to 16 fold increase). Eight of 12 NS1 mutants had decreased binding affinity to the cleavage and polyadenylation specificity factor (CPSF30). The majority of mutant NS1 genes demonstrated increased viral polymerase activity and viral protein production in mouse cells. Viral protein production and viral growth were also assessed in human and canine cell lines; however these adaptive phenotypes were more robust in infected mouse cells. Adaptive NS1 mutations also increased cytoplasmic cellular localization of the NS1 protein in infected cells in a host cell-specific manner. Evaluation of phenotypic trends associated with the NS1 mutants demonstrated an inverse correlation between CPSF30 binding affinity and viral polymerase activity enhancement. This study demonstrates that NS1 is a multifunctional virulence factor subject to adaptive evolution.

The sequence of 3553 nucleotides corresponding to the 3'-terminal region of potato virus S (PVS) has been determined from cloned cDNA. The sequence obtained contains six open reading frames with the potential to encode proteins of Mr 10,734, Mr 32,515, Mr 7,222, Mr 11,802, Mr 25,092 and at least Mr 41,052. The amino acid sequence of the 33K ORF has been confirmed to be that of the viral coat protein gene. The nucleotide sequence of this ORF was obtained from expression plasmids which were isolated by binding with a specific monoclonal antibody to PVS, and the expression of coat protein fusion products was verified by Western blots of bacterial cell lysates. The deduced amino acid sequence of a 70 amino acid portion from the central region of the PVS coat protein was 59% identical to the analogous region of potato virus X. In addition, the 7K, 12K and 25K ORF's displayed significant sequence homology with similar sized ORF's from a number of potexviruses. The partial 41K ORF was homologous with the C-terminal portion of the viral replicase proteins of potato virus X and white clover mosaic virus. While the biological functions of the 12K and 25K non-structural proteins coded for by PVS and members of the potexvirus group remain unknown, the 12K protein displays a hydropathicity profile consistent with a membrane associated protein and the 25K protein contains a conserved sequence motif found in a number of nucleoside triphosphate binding proteins. Members of the carlavirus group are distinguished from the potexviruses by the presence of a small [11K (PVS, potato virus M) - 16K (lily symptomless virus)] 3' terminal ORF which appears to contain a sequence motif similar to the 'zinc-finger' domain found in many nucleic acid binding proteins. The coat protein gene from potato virus S (PVS) was introduced into Nicotiana debneyii tobacco as well as a commercial potato cultivar, 'Russet Burbank', by leaf disc transformation using Agrobacterium tumefaciens. Transgenic plants expressing the viral coat protein were highly resistant to subsequent infection following mechanical inoculation with the Andean or ME strains of PVS as indicated by a lack of accumulation of virus in the upper leaves. The coat protein mediated protection afforded by these transgenic plants was sufficient to prevent the accumulation of virus in the tissues of non-transformed 'Russet Burbank' shoots which had been grafted onto transgenic plants inoculated with PVS, and in reciprocal grafts, transgenic shoots accumulated less than 2% (6 weeks after grafting) of the concentration of PVS found in non-transformed shoots similarly grafted onto plants systemically infected with PVS. These transgenic plants also displayed a measure of resistance to inoculation with a related carlavirus from potato, potato virus M. In agreement with previous reports for plants expressing PVX coat protein, plants expressing PVS coat protein were also protected from inoculation with PVS RNA. These results provide further evidence that coat protein mediated protection for these two groups of viruses, which share similar genome organizations, may involve inhibition of some early event in infection, other than, or in addition to, virus uncoating. Specific monoclonal antibodies were prepared against a C-terminal derived 18 kDa portion of the 25K protein of PVS expressed as an in-frame chimeric fusion protein with the glutathione S-transferase gene. The in vivo expression of this non-structural protein in virus infected tissue, as well as tissue from transgenic tobacco (var Xanthi-nc) engineered to contain the entire 25K gene, was verified by Western immunoblot labelling. / Medicine, Faculty of / Biochemistry and Molecular Biology, Department of / Graduate

No description available.

Thesis (Ph. D.)--University of Missouri--Columbia, 1996. / Typescript. Vita. Includes bibliographical references (leaves : 131-150). Also available on the Internet.

子宮頸癌是全球女性中第三常見的癌症。人類乳頭瘤狀病毒（HPV）已被證實為引致子宮頸癌的主要因素。目前已發現了150多種HPV。HPV-52在世界上較為少見，但在亞洲，特別是東亞地區，卻相當流行。 / 本回顧性研究收集了303個HPV-52陽性的子宮頸樣本，其中185個來自香港，118個來自韓國首爾。我們通過對HPV基因組中E6、E7、L1和LCR區域進行擴增和測序，以檢測HPV-52變異株的序列多樣性和致癌風險。 / L1-LCR-E6-E7串聯片段佔據了HPV-52基因組全長的41%。由191條該種序列構建的系統發育樹顯示，HPV-52變異株進化成四個世系。原型系A進化系在香港和首爾都很少見，只占全部樣本的3.7%。B進化系（89.5%）則是最普遍的HPV-52病毒系。E6的最大序列差異為1.6%，L1（2.3%），E7（3.4%）和LCR（4.8%）依次增大。因此，E6作為最保守的基因組區域可作為HPV-52通用引物PCR的靶點，而E7更適宜作為特定變異株的PCR靶點。此外，在短片段序列中發現了可識別HPV-52進化系和進化枝的單核苷酸突變。它們可用於擴增斷裂的DNA片段或大規模實驗中。再者，進化壓力分析顯示E6、E7和L1三個編碼區域都經歷了強烈的淨化選擇作用。 / HPV-52進化系和常見變異株在香港和首爾的分佈情況沒有顯著差異。但E6中的nt 356G>A、nt 378A>C和nt 467C>A (N122K) 核苷酸突變只出現在香港樣本，而L1的nt 6239G>A以及LCR的nt 7395G>A和nt 7911A>C核苷酸突變只在首爾樣本中發現。HPV-52 E6的N122K突變對子宮頸癌有較高的致癌風險（P-value = 0.002）。E6中的nt 378A>C (P-value = 0.014) 同義突變, 以及LCR中的nt 7665G>A (P-value < 0.001)和nt 94G>A (P-value = 0.007)突變，亦與高致癌風險相關。LCR中的nt 7911A>C (P-value = 0.007)和nt 19T>C (P-value = 0.008) 突變則對子宮頸癌的發展有較低風險。HPV-52 E7或L1中的突變與子宮頸癌的發展無明顯關係。上述結果需要通過進一步研究證實。針對HPV-52序列變異的病毒學和作用機理的深入研究是必要的。 / Cervical cancer is the third most common cancer in women worldwide. It has been proven that human papillomavirus (HPV) is the primary causative agent of cervical cancer. To date, more than 150 HPV types have been characterized. HPV-52 is rare around the world but frequently detected in Asia, especially East Asia. / This retrospective study analyzed 303 cervical samples that 185 were collected from Hong Kong, and 118 were collected from Seoul, Korea. All samples were positive for HPV-52. HPV gene regions of E6, E7, L1 and LCR were amplified and sequenced to determine sequence diversity and risk association of HPV-52 variants between the two cities. / The 191 concatenated L1-LCR-E6-E7 sequences that comprised 41% of the whole HPV-52 genome displayed four distinct clusters. The prototype-like lineage A was rare in both cities, only found in 3.7% of all samples. Lineage B (89.5%) was found to be the most prevalent lineage. The maximum sequence divergence of E6 was 1.6%, followed by L1 (2.3%), E7 (3.4%) and LCR (4.8%). E6 being the most conserved region could be a target for HPV-52 consensus PCR, and E7 could be a target for variant-specific PCR. Besides, several single-nucleotide substitutions diagnostic for HPV-52 lineage and clade classification were identified within a few short fragments. They might be useful when handling fragmented DNA and being a more feasible approach in large-scale studies. Moreover, analysis of evolutionary pressure indicated that all the three encoding regions, E6, E7 and L1, underwent strong purifying selection. / No significant difference in the distribution pattern of HPV-52 lineages and common variants between Hong Kong and Seoul was observed. But nucleotide substitutions nt 356G>A, nt 378A>C and nt 467C>A (N122K) were only found in Hong Kong samples; whereas nt 6239G>A, nt 7395G>A and nt 7911A>C were exclusively found in samples from Seoul. A significantly higher risk for cervical cancer was found for the HPV-52 E6 variant N122K (P-value = 0.002). A synonymous substitution of E6, nt 378A>C (P-value = 0.014), as well as two nucleotide substitutions of LCR, nt 7665G>A (P-value < 0.001) and nt 94G>A (P-value = 0.007), were also associated with a significant increase in risk for cervical cancer. Two substitutions found to confer a lower risk for cervical cancer were nt 7911A>C (P-value = 0.007) and nt 19T>C (P-value = 0.008), both of which located at LCR. No significant associations between HPV-52 E7 or L1 variants and cervical cancer development were observed. Further studies are needed to confirm these findings, and in-depth investigations into the virological and functional implications of HPV-52 sequence variations are warranted. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Zhang, Chuqing. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 124-137). / Abstracts also in Chinese. / Abstract --- p.i / Acknowledgements --- p.v / Table of contents --- p.vii / List of Figures --- p.ix / List of Tables --- p.x / Abbreviations --- p.xii / Chapter Chapter One --- Introduction --- p.1 / Chapter 1.1 --- History of Human Papillomavirus --- p.2 / Chapter 1.2 --- Biology of Human Papillomavirus --- p.4 / Chapter 1.2.1 --- Genome structure --- p.4 / Chapter 1.2.2 --- Protein function --- p.6 / Chapter 1.2.3 --- Latent and lytic life cycle --- p.9 / Chapter 1.2.4 --- Classification --- p.10 / Chapter 1.3 --- Epidemiology of Human Papillomavirus --- p.14 / Chapter 1.3.1 --- Global burden --- p.14 / Chapter 1.3.2 --- Transmission --- p.18 / Chapter 1.3.3 --- Clinical course --- p.19 / Chapter 1.3.4 --- Prevention --- p.23 / Chapter 1.4 --- Human Papillomavirus Type 52 --- p.25 / Chapter 1.5 --- Objectives --- p.26 / Chapter Chapter Two --- Materials and Methods --- p.27 / Chapter 2.1 --- Study Design --- p.28 / Chapter 2.2 --- Study population --- p.29 / Chapter 2.3 --- DNA extraction --- p.31 / Chapter 2.4 --- Polymerase chain reaction --- p.32 / Chapter 2.4.1 --- Long-fragment PCR approach --- p.33 / Chapter 2.4.2 --- Short-fragment PCR approach --- p.40 / Chapter 2.4.3 --- Purification of PCR products --- p.46 / Chapter 2.5 --- Nucleotide sequencing --- p.47 / Chapter 2.6 --- Data analysis --- p.48 / Chapter 2.6.1 --- Phylogenetic analysis --- p.48 / Chapter 2.6.2 --- Statistical analysis --- p.49 / Chapter Chapter Three --- Results --- p.50 / Chapter 3.1 --- Phylogeny of HPV-52 --- p.53 / Chapter 3.1.1 --- Concatenated sequence of L1-LCR-E6-E7 --- p.53 / Chapter 3.1.2 --- E6 gene --- p.56 / Chapter 3.1.3 --- E7 gene --- p.59 / Chapter 3.1.4 --- L1 gene --- p.62 / Chapter 3.1.5 --- Long control region --- p.67 / Chapter 3.2 --- Nucleotide sequence variation of HPV-52 --- p.70 / Chapter 3.2.1 --- E6 gene --- p.70 / Chapter 3.2.2 --- E7 gene --- p.73 / Chapter 3.2.3 --- L1 gene --- p.75 / Chapter 3.2.4 --- Long control region --- p.81 / Chapter 3.3 --- Geographical distribution of HPV-52 variants --- p.86 / Chapter 3.4 --- Risk association of HPV-52 variants --- p.96 / Chapter Chapter Four --- Discussion --- p.105 / Chapter 4.1 --- Strengths and weaknesses --- p.107 / Chapter 4.2 --- Phylogeny of HPV-52 variants --- p.109 / Chapter 4.2.1 --- Variant lineage classification system of HPV-52 --- p.109 / Chapter 4.2.2 --- Sequence variability of HPV-52 --- p.110 / Chapter 4.2.3 --- Evolutionary pressure on HPV-52 --- p.111 / Chapter 4.3 --- Nucleotide sequence variations of HPV-52 --- p.113 / Chapter 4.3.1 --- E6 gene --- p.113 / Chapter 4.3.2 --- E7 gene --- p.114 / Chapter 4.3.3 --- L1 gene --- p.116 / Chapter 4.3.4 --- Long control region --- p.117 / Chapter 4.4 --- Conclusions --- p.121 / References --- p.124 / Appendices --- p.138