Molecular epidemiology of and vaccine development against foot-and-mouth disease virus in Hong Kong
Abstract of thesis entitled
MOLECULAR EPIDEMIOLOGY OF
AND VACCINE DEVELOPMENT AGAINST
FOOT-AND-MOUTH DISEASE VIRUS IN HONG KONG
Raymond Kin-hi Hui
for the degree of Doctor of Philosophy
at The University of Hong Kong
in December 2004
Foot-and-mouth disease (FMD) is an endemic viral infectious disease in the Hong Kong porcine industry. This disease is caused by the Foot-and-mouth disease virus (FMDV) and disease outbreak causes a great impact on the local agricultural economy. Vaccination is a prompt and an effective measure to prevent the disease. To develop a specific vaccine against local strains of virus, epidemiology of the disease was first studied by nucleotide sequence analysis of the G-H loop in the VP1 region of the viral genome. Among 1056 samples collected from 42 local pig farms during the past four years, 109 were positive for the disease. Sequence data reveal that viruses isolated in Hong Kong belong to the Cathay topotype showing that the virus is introduced from neighboring Asian countries. The non-synonymous and the total substitution rates of FMDV circulating in Hong Kong were 1.26 ?10-2 and 4.83 ?10-3 substitutions/nucleotide/year respectively. Linear regression analysis estimates that the virus was introduced to Hong Kong between 1962 and 1968. Sequence data also indicate that geographical factors affect the spread of the virus, but the major causative event is most likely caused by human activities.
In order to quantify FMDV precisely during vaccine development, a Sybr green real-time RT-PCR assay was developed. Sensitivity of the assay was 2.9 log units higher than that of conventional plaque assay and the dynamic detection range of the assay was at least eight orders of magnitude, and it was also capable of detecting a single copy of viral genome molecule. The assay also reflects the level of neutralizing antibody in serum samples by monitoring the virus proliferation in virus neutralization assay. Melting curve analysis of amplicons reveals that a single nucleotide change in the amplicon contributed to alternation in melting temperature by 0.5 ?. This characteristic can be applied as a tool in quality control during inactivated vaccine production.
The effectiveness of various vaccine candidates was investigated. In this study, vaccine candidates designed according to the local virus strain were used to immunize BALB/c mice. FMDV-specific antibodies and their neutralizing activity were evaluated with ELISA and real-time RT-PCR mediated neutralizing assay respectively. Among these candidates, Escherichia coli expressing the VP1 protein elicited the antibody titer and neutralizing activity most similar as compared to those immunized with a commercial inactivated vaccine. The same viral gene fragment immunized in form of pcDNA3.1-SP DNA plasmid and recombinant adenovirus also induced humoral responses; nevertheless, the specific antibody titer was lower than those vaccinated with bacterial expressed protein and conferred only partial neutralization in cell culture.
In summary, strains of FMDV circulating in Hong Kong pig farms are localized, and therefore a specific vaccine is needed to protect the animals. Bacterial expressed VP1 protein designed according to local virus strains is a potential cost-effective vaccine candidate.
School:The University of Hong Kong
School Location:China - Hong Kong SAR
Source Type:Master's Thesis
Keywords:foot and mouth disease china hong kong virus diseases molecular epidemiology vaccines
Date of Publication:01/01/2005