Site directed mutation of ctx operon in vcusm2, vibrio cholerae vaccine candidate strain: towards the development of a vaccine for cholera
| dc.contributor.author | Chan, Li Ann (Melissa) | |
| dc.date.accessioned | 2015-07-30T02:39:00Z | |
| dc.date.available | 2015-07-30T02:39:00Z | |
| dc.date.issued | 2008 | |
| dc.description.abstract | Cholera is an acute diarrheal disease caused by Vibrio cholerae 01 and 0139. It affects countries with scarce clean water supplies and poor sanitation systems. Reported cases and mortality rates are increasing. No vaccines are available that confers protection against the 0139 serotype. This study aims to create an attenuated 0139 vaccine strain by mutating the ctx operon in VCUSM2 through site-directed mutagenesis, rendering it non-toxigenic but retaining its immunogenicity. VCUSM2 is a hemA mutated, ALA auxotrophic 0139 strain with two copies of ctx operon in tandem. The operon was cloned into pTZ57R, ace and zot genes were deleted by inverse PCR and the cholera toxin A subunit was mutated at the 7th (arginine to lysine, R7K) and 112th (glutamate to glutamine, E112Q) amino acid positions. VCUSM14 was obtained through a series of conjugation with BW20767-}.pir E. coli harboring pWM91 carrying respective inserts, and sucrose selection. VCUSM14 does not cause fluid accumulation in rabbit ileal loop model and is a poor colonizer in the infant mouse model. CT -ELISA showed that it produces toxin molecules recognizable by polyclonal anti-CT antibodies. This strain shows potential for vaccine development once colonization abilities are improved through reinsertion of the hemA gene. | en_US |
| dc.identifier.uri | http://hdl.handle.net/123456789/928 | |
| dc.language.iso | en | en_US |
| dc.subject | Vaccine candidates | en_US |
| dc.subject | Vibrio cholerae | en_US |
| dc.title | Site directed mutation of ctx operon in vcusm2, vibrio cholerae vaccine candidate strain: towards the development of a vaccine for cholera | en_US |
| dc.type | Thesis | en_US |
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