Generation of safety enhanced Edwardsiella tarda ghost vaccine.
A dual vector expressing the ghost-inducing PhiX174 lysis E gene and the bacterial DNA degrading staphylococcal nuclease A (SNA) gene was constructed to solve the problem of remnant antibiotic resistance genes and genomic DNA with intact pathogenic islands in the final product of Edwardsiella tarda ghosts (ETG). The SNA (devoid of secretion signal sequence and the nuclease B amino terminus sequence), fused with the 26 amino acid N-terminal sequence of the lambda phage Cro gene, showed successful degradation of bacterial nucleic acids. Furthermore, the nuclease activity of SNA in E. tarda was enhanced by codon optimization of the SNA gene using site-directed mutagenesis. ETG were generated via coexpression of the SNA gene and lysis gene E under the control of each lambdaP(R) promoter. The ghost bacteria generation system we describe is advantageous as it allows the use of a single plasmid, improves safety and vaccine purity by limiting residual genetic content from the ghost bacteria, and reduces production costs through cheap means of induction that use only temperature shifts.
Lee
Dong Jin
DJ
Department of Aquatic Life Medicine, Pukyong National University, Pusan 608-737, Korea.
Kwon
Se Ryun
SR
Zenke
Kosuke
K
Lee
Eun Hye
EH
Nam
Yoon Kwon
YK
Kim
Sung Koo
SK
Kim
Ki Hong
KH
D000818:Animals; D001428:Bacterial Vaccines / Q000276:immunology*; D004269:DNA, Bacterial / Q000378:metabolism; D020609:Edwardsiella tarda / Q000276:immunology*; D004756:Enterobacteriaceae Infections / Q000517:prevention & control / Q000662:veterinary*; D004926:Escherichia coli / Q000235:genetics; D005393:Fish Diseases / Q000517:prevention & control*; D015964:Gene Expression Regulation, Bacterial; D008836:Micrococcal Nuclease / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D013997:Time Factors; D015164:Vaccines, Inactivated / Q000276:immunology; D014764:Viral Proteins / Q000235:genetics
D001428:Bacterial Vaccines; D004269:DNA, Bacterial; C029555:E protein, bacteriophage X174; D015164:Vaccines, Inactivated; D014764:Viral Proteins; D008836:Micrococcal Nuclease