Human coronaviruses, namely HCoV-229E and HCoV-OC43, have been identified as causal agent of upper respiratory tract infections for decades. The significance of human coronaviruses studies re-emerged after the SARS pandemic. Two novel human coronaviruses, HCoV-NL63 and HCoV-HKU1, were identified after the SARS pandemic. Up till now there are a total of five human coronaviruses being identified. After the discovery of HCoV-HKU1 in 2005, molecular epidemiology and genome studies identified the first natural recombination in human coronaviruses, which has led to the generation of different genotypes. A similar phenomenon was also observed in HCoV-OC43 in a subsequent study, resulting in the emergence of a novel genotype associated with pneumonia. Although HCoV-229E has been discovered for over forty years, information regarding its evolution and epidemiology is little and scattered. In this study, 32 HCoV-229E strains were collected from nasopharyngeal aspirates over a period of 8 years (from April 2004 to January 2012). Three genes, including RdRp (RNA-dependent-RNA polyermase), S (spike) and N (nucleocapsid), were sequenced and analyzed. Phylogenetic studies showed the existence of genetic drift among six chronological groups, including group 1 from 1979 to 1982, group 2 from 1982 to 1984, group 3 from 1990 to 1992, group 4 from 2001 to 2005, group 5 from 2005 to 2008 and group 6 from 2011 to 2012. One particular strain in 2006, HCoV-229E-HK06-24 displayed an incongruent position between the S and N gene, suggesting a possible recombination between group 4 and group 5. Additionally, five strains from 2011 to 2012 showed incongruent positions in RdRp comparing to other strains, which are suspected to be novel group 6. This study revealed the first evidence for a possible natural recombination event in HCoV-229E. The predominate group 6, which is genetically different from previous strains, may have been arisen by genetic drift. Further surveillance is required to monitor the genetic changes in HCoV-229E. / published_or_final_version / Microbiology / Master / Master of Medical Sciences

Thesis (M. Phil.)--University of Hong Kong, 2007. / Also available in print.

Thesis (M. Phil.)--University of Hong Kong, 2008. / Also available in print.

Thesis (M.Sc. (Microbiology))--Mahidol University, 1971.

Thesis (Ph. D.)--University of Hong Kong, 2010. / Includes bibliographical references (leaves 151-172). Also available in print.

Thesis (M. Med. Sc.)--University of Hong Kong, 2006.

abstract: Coronaviruses are medically important viruses that cause respiratory and enteric infections in humans and animals. The recent emergence through interspecies transmission of severe acute respiratory syndrome coronavirus (SARS-CoV) strongly supports the need for development of vaccines and antiviral reagents. Understanding the molecular details of virus assembly is an attractive target for development of such therapeutics. Coronavirus membrane (M) proteins constitute the bulk of the viral envelope and play key roles in assembly, through M-M, M-spike (S) and M-nucleocapsid (N) interactions. M proteins have three transmembrane domains, flanked by a short amino-terminal domain and a long carboxy-terminal tail located outside and inside the virions, respectively. Two domains are apparent in the long tail - a conserved region (CD) at the amino end and a hydrophilic, charged carboxy-terminus (HD). We hypothesized that both domains play functionally important roles during assembly. A series of changes were introduced in the domains and the functional impacts were studied in the context of the virus and during virus-like particle (VLP) assembly. Positive charges in the CD gave rise to viruses with neutral residue replacements that exhibited a wild-type phenotype. Expression of the mutant proteins showed that neutral, but not positive, charges formed VLPs and coexpression with N increased output. Alanine substitutions resulted in viruses with crippled phenotypes and proteins that failed to assemble VLPs or to be rescued into the envelope. These viruses had partially compensating changes in M. Changes in the HD identified a cluster of three key positive charges. Viruses could not be recovered with negatively charged amino acid substitutions at two of the positions. While viruses were recovered with a negative charge substitution at one of the positions, these exhibited a severely crippled phenotype. Crippled mutants displayed a reduction in infectivity. Results overall provide new insight into the importance of the M tail in virus assembly. The CD is involved in fundamental M-M interactions required for envelope formation. These interactions appear to be stabilized through interactions with the N protein. Positive charges in the HD also play an important role in assembly of infectious particles. / Dissertation/Thesis / Ph.D. Microbiology 2010

published_or_final_version / Microbiology / Doctoral / Doctor of Philosophy

published_or_final_version / abstract / Medicine / Master / Doctor of Medicine

published_or_final_version / abstract / Microbiology / Doctoral / Doctor of Philosophy