Key Laboratory of Zoonoses of Jiangsu Province, Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou University, Yangzhou 225009, China
Abstract:Salmonella enteritidis is one of the zoonotic foodborne pathogens and could infect humans mainly through contaminated poultry and egg products. To explore the possibility as new genetically engineered vaccines, the double gene-deletion strain of Salmonella enteritidis C50041ΔspiCΔcrp was constructed in this study. The crp gene was deleted again by λ Red system on the base of Salmonella enteritidisΔspiC to construct Salmonella enteritidisΔspiCΔcrp. PCR and antibiotic resistance showed Salmonella enteritidisΔspiCΔcrp had been successfully constructed, and biological identification clarified that its growth and the biochemical properties had changed; LD50 was high about 1 000 folds and virulence was significantly attenuated, comparing with wild type strain C50041. It laid an important foundation for further study of Salmonella enteritidis as a live attenuated vaccine.
[1]Raphael S, Sharon MT, Wang JY, et al. Salmonella enterica serovar Enteritidis core O polysaccharide conjugated to H:g,m flagellin as a candidate vaccine or protection against invasive infection with S. Enteritidis[J]. Infect Immun, 2011, 79(10): 4240-4249. DOI: 10.1128/IAI.05484-11 [2]Mickael CS, Lam PKS, Berberov EM, et al. Salmonella enterica serovar Enteritidis tatB and tatC mutants are impaired in Caco-2 Cell invasion in vitro and show reduced systemic spread in chickens[J]. Infect Immun, 2010, 78(8): 3493-3505. DOI: 10.1128/IAI.00090-10 [3]Keller LH, Benson CE, Krotec K, et al. Salmonella Enteritidis colonization of the reproductive tract and forming and freshly laid eggs of chickens[J]. Infect Immun, 1995, 63(7): 2443-2449. [4]Mead G, Lammerding AM, Cox N, et al. Scientific and technical factors affecting the setting of Salmonella criteria for raw poultry: a global perspective[J]. J Food Prot, 2010, 73(8): 1566-1590. [5]Geng SZ, Liu NN, Jiao XA, et al. Development and identification of S06004ΔspiC mutant of Salmonella Pullorum[J]. Chin Vet Sci, 2014, 44(4): 379-386. (in Chinese) 耿士忠, 刘男男, 焦新安,等. 鸡白痢沙门菌S06004ΔspiC突变株的构建与鉴定[J]. 中国兽医科学, 2014, 44(04): 379-386. [6]Geng SZ, Jiao XA, Pan ZM, et al.An improved method to knock out the asd gene of Salmonella enterica serovar Pullorum[J]. J Biomed Biotechnol, 2009, 2009: 646380. DOI: 10.1155/2009/646380 [7]Datsenko KA, Wanner BL. One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products[J]. Proc Natl Acad Sci U S A, 2000, 97(12): 6640-6645. [8]Trach DD, Cam PD, Ke NT, et al. Investigations into the safety and immunogenicity of a killed oral cholera vaccine developed in Vietnam[J]. World Health Orgam, 2002, 80: 2-8. [9]Gunn BM, Wanda SY, Burshell D, et al. Construction of recombinant attenuated Salmonella enterica serovar Typhimurium vaccine vector strains for safety in newborn and infant mice[J]. Clin Vaccine Immunol, 2010, 17(3): 354-362. DOI: 10.1128/CVI.00412-09 [10]Fields PI, Swanson RV, Haidaris CG, et al. Mutants of Salmonella typhimurium that cannot survive within the macrophage are avirulent[J]. Proc Natl Acad Sci U S A, 1986, 83: 5189-5193.