Efficient production of safety-enhanced Escherichia coli ghosts by tandem expression of PhiX 174 mutant gene E and staphylococcal nuclease A gene

The application of bacterial ghosts as vaccines is limited because of their low lysis efficiency and production and the presence of pathogenic islands and/or antibiotic resistance genes within ghost preparations. To overcome these problems, a new lysis plasmid with fusion gene of mutant gene E and staphylococcal nuclease A gene (mE-L-SNA) were constructed and characterized. The new plasmid pBV-mELS could efficiently induce the genetic inactivation of Escherichia coli cultures, accompanied by the intracellular degradation of the genetic material of host cells, devoid of the presence of pathogenic islands and antibiotic resistance genes within ghost preparations. Furthermore, the lysis efficiency of the plasmid pBV-mELS was not affected by bacterial concentration and could reach 99.99995% for E. coli at late-log phase. However, when the 74-bp non-encoding region of the gene mE-L-SNA were deleted or the first T nucleotide of the gene mE-L-SNA were substituted, these resulting genes lost the function of bacteriolysis, which suggested the 74-bp region of the gene mE-L-SNA, especially the first T nucleotide, played a crucial role in enhancement of bacteriolysis. The lysis system with the gene mE-L-SNA had predominance for large-scale production of safety-enhanced bacterial ghosts. The strategy may provide a promising avenue for efficient production of safe bacterial ghost vaccines.