Study on Genetic Inactivation Bacterial Ghosts of Pasteurella multocida based PhiX174 Gene E Lysis Cassette Mediated

[Objective] To study on genetic inactivation bacterial ghosts of Pasteurella multocida based PhiX174 gene E lysis cassette mediated. [ Method ] Recombinant pPBA1100-e was constructed by which the gene E of bacteriophage Phix174 and temperature sensitivity expressing control system hybridized with plasmid pPBA1100 by genetic engineering method. Recombinant was transformed to Pasteurella multocida and lysis gene E expressed by temperature induction. Recombinant was detected by restriction endonuclease. Cell morphology of bacterial ghost of Pasteurella mul- tocida was observed by scanning electron microscopy and inactivation ratio was estimated by CFU analysis. I Result~ The results indicated that the recombination plasmid presented three bands by restriction endonuclease and agarose electrophoresis and that molecular weight of every band ac- corded with theoretical value. The result of SEM observing showed that recombination plasmid expressed successfully in P. multocida and produced bacterial ghost. The result of CFU detecting demonstrated that inactivation ratio of P. multocida reached 99 per cent. ~Conclusion~ This study pro- vided technical bases for the preparation of antigen vaccine of natural bacterial outer membrane protein.

Cancer cell membrane-coated bacterial ghosts for highly efficient paclitaxel delivery against metastatic lung cancer

Chemotherapy is one of the major approaches for the treatment of metastatic lung cancer,although it is limited by the low tumor delivery efficacy of anticancer drugs.Bacterial therapy is emerging for cancer treatment due to its high immune stimulation effect;however,excessively generated immunogenicity will cause serious inflammatory response syndrome.Here,we prepared cancer cell membrane-coated liposomal paclitaxel-loaded bacterial ghosts(LP@BG@CCM)by layer-by-layer encapsulation for the treatment of metastatic lung cancer.The preparation processes were simple,only involving film formation,electroporation,and pore extrusion.LP@BG@CCM owned much higher 4T1 cancer cell toxicity than LP@BG due to its faster fusion with cancer cells.In the 4T1 breast cancer metastatic lung cancer mouse models,the remarkably higher lung targeting of intravenously injected LP@BG@CCM was observed with the almost normalized lung appearance,the reduced lung weight,the clear lung tissue structure,and the enhanced cancer cell apoptosis compared to its precursors.Moreover,several major immune factors were improved after administration of LP@BG@CCM,including the CD4^(+)/CD8a^(+)T cells in the spleen and the TNF-α,IFN-γ,and IL-4 in the lung.LP@BG@CCM exhibits the optimal synergistic chemo-immunotherapy,which is a promising medication for the treatment of metastatic lung cancer.

Construction of Salmonella Pullorum ghost by co-expression of lysis gene E and the antimicrobial peptide SMAP29 and evaluation of its immune efficacy in specific-pathogen-free chicks

In this study, a safety enhanced Salmonella Pullorum （S. Pullorum） ghost was constructed using an antimicrobial peptide gene, and evaluated for its potential as a Pullorum disease （PD） vaccine candidate. The antimicrobial peptide SMAP29 was co-expressed with lysis gene E to generate S. Pullorum ghosts. No viable bacteria were detectable either in the fermentation culture after induction of gene E- and SMAP29-mediated lysis for 24 h or in the lyophilized ghost products. Specific-pathogen- free （SPF） chicks were intraperitoneally immunized with ghosts at day 7 of age and no mortality, clinical symptoms or signs of PD such as anorexia, depression and diarrhea were observed. On challenge with a virulent S. Pullorum strain at 4 wk post-immunization, a comparatively higher level of protection was observed in the S. Pullorum ghost immunized chickens with a minimum of pathological lesions and bacterial loads compared to the birds in inactivated vaccine groups. In addition, immunization with the S. Pullorum ghosts induced a potent systemic IgG response and was associated with significantly increased levels of cytokine IFN-y and IL-4 and relative percentages of CD4＋ and CD8＋ T lymphocytes. Our results indicate that SMAP29 can be employed as a new secondary lethal protein to enhance the safety of bacterial ghosts, and to prepare a non-living bacterial vaccine candidate that can prevent PD in chickens.