猪霍乱沙门菌C500△asd缺失株的构建及其作为疫苗活载体的初步应用
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摘要
从上世纪70年代遗传工程技术兴起以来,出现了许多新的疫苗研究策略。减毒沙门菌作为疫苗载体具有安全、高效的特性,已受到医学与兽医学界的广泛重视。减毒沙门菌的主要优点包括它可以经黏膜途径免疫(口服或鼻内),操作方便,对接种宿主刺激小。此外,沙门菌为兼性胞内寄生细菌,能有效呈递抗原,激发机体抗沙门菌和外源蛋白的特异性体液与细胞免疫反应,并能同时诱导黏膜免疫与全身免疫。为避免在基因工程活菌苗的研制中使用抗生素作为选择压,将减毒沙门菌用于宿主载体平衡致死系统中,使在没有抗生素存在的情况下,重组质粒能稳定存在于宿主体内。这样的疫苗设计为防控重大的动物疫病带来了新的希望。
猪霍乱沙门菌减毒株C500 (Salmonella choleraesuis C500 strain)是通过化学致弱用于预防仔猪副伤寒的有效活菌苗,其免疫原性强,安全性好,使用40余年来使我国仔猪副伤寒得到了较好的控制。因此,利用减毒的猪霍乱沙门菌C500构建宿主载体平衡致死系统用于运送重要疾病病原基因疫苗很有实际应用价值。
新城疫(Newcastle disease, ND)是由副粘病毒科禽腮腺炎病毒属中的新城疫病毒(Newcastle disease viruses, NDV)引起多种禽类发生感染和高度死亡的一种急性、接触性传染病,常给养禽业造成重大经济损失。在许多国家对该病的防控主要靠疫苗的免疫接种。NDV的融合蛋白F介导病毒与宿主细胞膜的融合,是病毒入侵细胞的重要分子。本研究构建了以猪霍乱沙门菌C500△asd缺失株为运送载体的NDV F基因工程菌,以小鼠和鸡为动物模型,评价了其免疫特性,初步探索了猪霍乱沙门菌C500△asd缺失株做为活疫苗载体的可行性。
1猪霍乱沙门菌C500△asd缺失株宿主载体平衡致死系统的构建与鉴定
根据GenBank中已发表的猪霍乱沙门菌asd基因的序列,设计两对引物分别扩增asd基因的上下游片段,然后构建含缺失1,107 bp的asd基因与蔗糖敏感基因(sacB)的重组自杀性质粒,与猪霍乱沙门菌C500接合转移,两步法筛选无抗性的△asd缺失株,用PCR证实基因组asd基因的缺失突变。该缺失株生长必需外源DAP(二氨基庚二酸)。进一步鉴定猪霍乱沙门菌C500△asd缺失株的表型、生长特性、毒力等,结果表明猪霍乱沙门菌C500△asd缺失株与其亲本株猪霍乱沙门菌C500的表型、生长特性基本一致,而其毒力有了进一步的下降。将携带asd基因的红色荧光蛋白质粒转化猪霍乱沙门菌C500△asd缺失株后,在体外传至100代后仍可高效表达红色荧光蛋白,说明该缺失株可以用来作为宿主载体平衡致死系统来高效表达外源基因,为深入研究以猪霍乱沙门菌C500△asd缺失株作为疫苗载体奠定了基础。
2新城疫病毒F蛋白在猪霍乱沙门菌C500△add缺失株中的表达及部分免疫特性分析
复苏本实验室保存的重组菌X4550(pYA3334-F),进行PCR和SDS-PAGE鉴定,阳性菌提取重组质粒(pYA3334-F)电转化猪霍乱沙门菌C500△asd缺失株,普通LB平板筛选阳性克隆,命名为猪霍乱沙门菌C500△asd(pYA3334-F).SDS-PAGE电泳检测到F蛋白的表达。动物免疫实验结果表明,重组菌猪霍乱沙门菌C500△asd(pYA3334-F能在小鼠和鸡体内诱导产生针对F蛋白的特异性抗体,显示本研究构建的以猪霍乱沙门菌C500△asd缺失株运送的外源抗原具有良好的免疫原性,能够激发有效的免疫应答,初步证明该系统做为活疫苗载体的可行性。
Since 1970s, many new vaccine strategies were developed with the advent of genetic engineering, so the safer and more efficient live vaccine using attenuated Salmonella spp. as the vector received wide attention in medical and veterinary medicine. The major advantage of the attenuated Salmonella spp. is that it can be administered by mucosal route (oral or intranasal), easily genetically manipulated and cause little side effect on animals. In addition, as an intracellular invasive bacterium, Salmonella spp. can effectively present antigen, induce specific humoral and cellular immune response against Salmonella spp. and heterologous antigen, and trigger both mucosal and systemic immune at the same time. To avoid using antibiotic as the screening marker in the construction of genetic engineering live vaccine, nutrition vicious attenuated Salmonella spp. based on balanced-lethal host-vector system was designed to deliver the expressed protein of foreign gene. Such vaccine design brings new hope to animal under the threat of contagious disease.
Strain C500 of Salmonella choleraesuis is an avirulent vaccine strain attenuated by chemical method, it has good immunogenicity and safety and has been used to prevent piglet paratyphoid in China for over 40 years. Strain Salmonella choleraesuis C500 used as a delivery system for heterologous antigens based on the host-vector balanced lethal system would have great prospect in the near future.
Newcastle disease (ND) is a highly contagious disease that causes substantial economic losses for poultry. Newcastle disease virus (NDV) belongs to the newly defined genus Avulavirus in the family of Paramyxoviridae. Its vaccine is frequently used for the prevention and control of ND in some countries when the disease spreads widely. The F protein, which mediates fusion of viral envelope and cellular membrane, is an important factor for viruse invasion and also a good protective immunogen. In this work, Salmonella choleraesuis C500△asd mutant was constructed, and used as a delivery system for F gene based on the balanced-lethal host-vector system. The immune responses of i.n. and i.m. vaccination with the recombinant Salmonella choleraesuis C500 vaccine strain expressing F antigen of NDV both in mouse and chicken were invaluated, the results showed that both of them produced specific antibody against F protein. It suggests that this balanced-lethal host-vector system of Salmonella choleraesuis could express foreign gene(s), delivery the target protein and intiate specific immunity, and could be used as vaccine vector against infectious disease.
1 Construction and characterization of Salmonella choleraesuis C500△asd host-vector balanced lethal system
Two sets of primers were designed and synthesized based on the sequences of asd gene of Salmonella choleraesuis in GenBank, then the upstream and downstream fragments of asd gene were amplified and cloned from Salmonella choleraesuis C500 genome DNA. The recombinant suicide plasmid was constructed in which asd gene was replaced by Cm gene. Strains with the deletion of asd gene were selected by two-step protocol and named Salmonella choleraesuis C500△asd mutant, which could not live without additive DAP. The mutant was confirmed by PCR method. Its phenotypes, growth properties and virulence characteristics were further verified, the results showed that the chracteristics of△asd mutant were consistent with those of its wild type, and its virulence had a drop compared with its parent strain. The dsRED plasmid was transformed into Salmonella choleraesuis C500△asd mutant by electroporation to generate Salmonella choleraesuis C500△asd (pYA3334-dsRed), the recombinant could express dsRED stably and efficiently after 100 generations in vitro. The results showed that the Salmonella choleraesuis C500△asd mutant was successfully constructed and it could be used as a vector based on the host-vector balanced-lethal system to express foreign gene(s) stably and efficiently, this study would pave a way for developing Salmonella choleraesuis C500 vaccine strain as a live vectored vaccine.
2 Expression of F protein of Newcastle disease virus in Salmonella choleraesuis C500△asd mutant and analysis of its immunogenicity
The recombinant Salmonella typhimurium X4550(pYA3334-F) was indentified by PCR and SDS-PAGE, the positive plasmid pYA3334-F was extracted and electroporated into Salmonella choleraesuis C500△asd mutant, the recombinant was screened and designated as Salmonella choleraesuis C500△asd (pYA3334-F), SDS-PAGE was conducted to detect the expression of F gene in recombinant Salmonella choleraesuis C500△asd (pYA3334-F). It was shown that Salmonella choleraesuis C500Aasd (pYA3334-F) induced specific antibody against F protein of NDV both in mouse and chicken.
In summary, the F protein expressed in attenuated Salmonella choleraesuis C500△asd had good immunogenity, and could initiate humoral and celluar immune responses. Salmonella choleraesuis C500△asd might be used as a live vaccine vector.
猪霍乱沙门菌减毒株C500 (Salmonella choleraesuis C500 strain)是通过化学致弱用于预防仔猪副伤寒的有效活菌苗,其免疫原性强,安全性好,使用40余年来使我国仔猪副伤寒得到了较好的控制。因此,利用减毒的猪霍乱沙门菌C500构建宿主载体平衡致死系统用于运送重要疾病病原基因疫苗很有实际应用价值。
新城疫(Newcastle disease, ND)是由副粘病毒科禽腮腺炎病毒属中的新城疫病毒(Newcastle disease viruses, NDV)引起多种禽类发生感染和高度死亡的一种急性、接触性传染病,常给养禽业造成重大经济损失。在许多国家对该病的防控主要靠疫苗的免疫接种。NDV的融合蛋白F介导病毒与宿主细胞膜的融合,是病毒入侵细胞的重要分子。本研究构建了以猪霍乱沙门菌C500△asd缺失株为运送载体的NDV F基因工程菌,以小鼠和鸡为动物模型,评价了其免疫特性,初步探索了猪霍乱沙门菌C500△asd缺失株做为活疫苗载体的可行性。
1猪霍乱沙门菌C500△asd缺失株宿主载体平衡致死系统的构建与鉴定
根据GenBank中已发表的猪霍乱沙门菌asd基因的序列,设计两对引物分别扩增asd基因的上下游片段,然后构建含缺失1,107 bp的asd基因与蔗糖敏感基因(sacB)的重组自杀性质粒,与猪霍乱沙门菌C500接合转移,两步法筛选无抗性的△asd缺失株,用PCR证实基因组asd基因的缺失突变。该缺失株生长必需外源DAP(二氨基庚二酸)。进一步鉴定猪霍乱沙门菌C500△asd缺失株的表型、生长特性、毒力等,结果表明猪霍乱沙门菌C500△asd缺失株与其亲本株猪霍乱沙门菌C500的表型、生长特性基本一致,而其毒力有了进一步的下降。将携带asd基因的红色荧光蛋白质粒转化猪霍乱沙门菌C500△asd缺失株后,在体外传至100代后仍可高效表达红色荧光蛋白,说明该缺失株可以用来作为宿主载体平衡致死系统来高效表达外源基因,为深入研究以猪霍乱沙门菌C500△asd缺失株作为疫苗载体奠定了基础。
2新城疫病毒F蛋白在猪霍乱沙门菌C500△add缺失株中的表达及部分免疫特性分析
复苏本实验室保存的重组菌X4550(pYA3334-F),进行PCR和SDS-PAGE鉴定,阳性菌提取重组质粒(pYA3334-F)电转化猪霍乱沙门菌C500△asd缺失株,普通LB平板筛选阳性克隆,命名为猪霍乱沙门菌C500△asd(pYA3334-F).SDS-PAGE电泳检测到F蛋白的表达。动物免疫实验结果表明,重组菌猪霍乱沙门菌C500△asd(pYA3334-F能在小鼠和鸡体内诱导产生针对F蛋白的特异性抗体,显示本研究构建的以猪霍乱沙门菌C500△asd缺失株运送的外源抗原具有良好的免疫原性,能够激发有效的免疫应答,初步证明该系统做为活疫苗载体的可行性。
Since 1970s, many new vaccine strategies were developed with the advent of genetic engineering, so the safer and more efficient live vaccine using attenuated Salmonella spp. as the vector received wide attention in medical and veterinary medicine. The major advantage of the attenuated Salmonella spp. is that it can be administered by mucosal route (oral or intranasal), easily genetically manipulated and cause little side effect on animals. In addition, as an intracellular invasive bacterium, Salmonella spp. can effectively present antigen, induce specific humoral and cellular immune response against Salmonella spp. and heterologous antigen, and trigger both mucosal and systemic immune at the same time. To avoid using antibiotic as the screening marker in the construction of genetic engineering live vaccine, nutrition vicious attenuated Salmonella spp. based on balanced-lethal host-vector system was designed to deliver the expressed protein of foreign gene. Such vaccine design brings new hope to animal under the threat of contagious disease.
Strain C500 of Salmonella choleraesuis is an avirulent vaccine strain attenuated by chemical method, it has good immunogenicity and safety and has been used to prevent piglet paratyphoid in China for over 40 years. Strain Salmonella choleraesuis C500 used as a delivery system for heterologous antigens based on the host-vector balanced lethal system would have great prospect in the near future.
Newcastle disease (ND) is a highly contagious disease that causes substantial economic losses for poultry. Newcastle disease virus (NDV) belongs to the newly defined genus Avulavirus in the family of Paramyxoviridae. Its vaccine is frequently used for the prevention and control of ND in some countries when the disease spreads widely. The F protein, which mediates fusion of viral envelope and cellular membrane, is an important factor for viruse invasion and also a good protective immunogen. In this work, Salmonella choleraesuis C500△asd mutant was constructed, and used as a delivery system for F gene based on the balanced-lethal host-vector system. The immune responses of i.n. and i.m. vaccination with the recombinant Salmonella choleraesuis C500 vaccine strain expressing F antigen of NDV both in mouse and chicken were invaluated, the results showed that both of them produced specific antibody against F protein. It suggests that this balanced-lethal host-vector system of Salmonella choleraesuis could express foreign gene(s), delivery the target protein and intiate specific immunity, and could be used as vaccine vector against infectious disease.
1 Construction and characterization of Salmonella choleraesuis C500△asd host-vector balanced lethal system
Two sets of primers were designed and synthesized based on the sequences of asd gene of Salmonella choleraesuis in GenBank, then the upstream and downstream fragments of asd gene were amplified and cloned from Salmonella choleraesuis C500 genome DNA. The recombinant suicide plasmid was constructed in which asd gene was replaced by Cm gene. Strains with the deletion of asd gene were selected by two-step protocol and named Salmonella choleraesuis C500△asd mutant, which could not live without additive DAP. The mutant was confirmed by PCR method. Its phenotypes, growth properties and virulence characteristics were further verified, the results showed that the chracteristics of△asd mutant were consistent with those of its wild type, and its virulence had a drop compared with its parent strain. The dsRED plasmid was transformed into Salmonella choleraesuis C500△asd mutant by electroporation to generate Salmonella choleraesuis C500△asd (pYA3334-dsRed), the recombinant could express dsRED stably and efficiently after 100 generations in vitro. The results showed that the Salmonella choleraesuis C500△asd mutant was successfully constructed and it could be used as a vector based on the host-vector balanced-lethal system to express foreign gene(s) stably and efficiently, this study would pave a way for developing Salmonella choleraesuis C500 vaccine strain as a live vectored vaccine.
2 Expression of F protein of Newcastle disease virus in Salmonella choleraesuis C500△asd mutant and analysis of its immunogenicity
The recombinant Salmonella typhimurium X4550(pYA3334-F) was indentified by PCR and SDS-PAGE, the positive plasmid pYA3334-F was extracted and electroporated into Salmonella choleraesuis C500△asd mutant, the recombinant was screened and designated as Salmonella choleraesuis C500△asd (pYA3334-F), SDS-PAGE was conducted to detect the expression of F gene in recombinant Salmonella choleraesuis C500△asd (pYA3334-F). It was shown that Salmonella choleraesuis C500Aasd (pYA3334-F) induced specific antibody against F protein of NDV both in mouse and chicken.
In summary, the F protein expressed in attenuated Salmonella choleraesuis C500△asd had good immunogenity, and could initiate humoral and celluar immune responses. Salmonella choleraesuis C500△asd might be used as a live vaccine vector.
引文
[1]Sharpe S, Fooks A, Lee J, et al. Single oral immunization with replication deficient recombinant adenovirus elicits long lived transgene specific cellular and humoral immune responses [J].Virology, 2002,293:210-216.
[2]Merchlinsky M and Moss B. Introduction of foreign DNA into the vaccine virus genome by in vitro ligation:recombination independent selectable cloning vectors [J]. Virology,1992,190(1): 522-526.
[3]Mackett M, Smit hL, Moss B, et al. Vaccinia virus:a selectable eukaryotic cloning and expression vector [J]. Proc Natl Acad Sci USA,1982,79:7415-7419.
[4]Brochier B, Aubert MF, Pastoret PP, et al. Field use of a vaccinia-rabies recombinant vaccine for the control of sylvatic rabies in Europe and North America [J]. Rev Sci Tech,1996,5(3): 947-997.
[5]王文玲,黄保英,邓瑶,等.甲型流感病毒M2基因在痘苗病毒天坛株中的表达[J].病毒学报,2007,24(5)：377-383.
[6]Christina JK, Janice C, Matthew R, et al. Use of recombinant poxviruses to stimulate antimelanoma T Cell reactivity [J].Annu Surg Oncol,1998,5 (1):64-76.
[7]Fooks AR. Development of oral vaccines for human us [J]. Curr Opin Mol Ther,2000,2:80-86.
[8]Matsunaga Y, Wakatsuki Y, Tabata Y, et al. Oral immunization with sizepurified microsphere beads as a vehicle selectively induces systemic tolerance and sensitization [J]. Vaccine,2000,19: 579-588.
[9]Schoen C, Stritzker J, Goebel W, et al. Bacteria as DNA vaccine carriers for genetic immunization [J]. Int J Med Microbiology,2009,294:319-335.
[10]Dietrich G, Spreng S, Gentschev I, et al. Bacterial systems for the delivery of eukaryotic antigen expression vectors [J]. Antisense Nucleic Acid Drug Dev,2000,10(5):391-399.
[11]Jones BD, Ghori N, Falkow S, et al. Salmonella typhimurium initiates murine infection by penetrating and destroying the specialized epithelial M cells of the Peyer's patches [J]. Exp Med, 1994,180:15-23.
[12]Pehheiter KL, Mathur N, Giles D, et al. Non-invasive Salmonella typhimurium mutants are avirulent because of an inability to enter and destroy M cells of ileal Peyer's patches [J]. Mol Microbiol,1997,24:697-709.
[13]Vasquez TA, Jones CJ, Baumler AJ, et al. Extraintestinal dissemination of Salmonella by CD 18 expressing phagocytes [J]. Nature,1999,401:804-808.
[14]Rescigno M, Urbano M, Valzasina B, et al. Dendritic cells express tight junction proteins and penetrate gut epithelia monolayers to sample bacteria [J]. Nat Immunol,2001,2:361-367.
[15]Saunders BM and Cooper AM. Restraining mycobacteria:Role of granulomas in mycobacterial infections [J]. Immunol Cell Biol,2000,78:334-341.
[16]Sirard JC, Niedergang F, Kraehenbuhl P, et al. Live attenuated Salmonella:a paradigm of mucosal vaccines [J]. Immunol Rev,1999,171:5-26.
[17]Hopkins SA, Niedergang F, Corthesy-Theulaz IE, et al. A recombinant Salmonella typhimurium vaccine strain is taken up and survives within murine Peyer's patch dendritic cells [J]. Cell Microbiol,2000,2(2):59-68.
[18]Fastad IN, Halstense TS, Fausa O, et al. Heterogeneity of M-cell-associated B and T cells in human Peyer's patch dendritic cells [J]. Immunology,1994,83(3):457-464.
[19]Clark MA, Jepson MA, Simmons NL, et al. Selective binding and transcytosis of Ulex europaeus lectin by mouse Peyer's patch M-cells in vivo [J]. Cell Tissue Res,1995,282(3): 455-461.
[20]Vazquez TA, Jones CJ, Baumler AJ, et al. Extraintestinal dissemination of Salmonella by CD 18-expressing phagocytes [J]. Nature,1999,401(67):804-808.
[21]Rescigno M, Urbano M, Valzasina B, et al. Dendritic cells express tight junction proteins and penetrate gut epithelial monolayersto sample bacteria [J]. Nat Immunol,2001,2(4):361-367.
[22]Paglia P, Medina E, ArioliI, et al. Gene transfer in dendritic cells, induced by oral DNA vaccination with Salmonella typhimurium, results in protective immunity against a murine fibrosarcoma [J]. Blood,1998,92(9):3172-3176.
[23]Flo J, Tisminetzky S, Baralle F. Oral transgene vaccination mediated by attenuated Salmonella is an effective method to prevent herpes simplex virus induced disease in mice [J].Vaccine,2001, 19(13):1772-1782.
[24]Buchmeier, NA and Heffron F. Induction of Salmonella stress proteins upon infection of macrophages [J]. Infect Immun,1989,57(1):1-7.
[25]Yrlid U, Svensson M, Hakansson A, et al. In vivo activation of dendritic cells and T cells during Salmonella enterica serovar Typhimurium infection [J]. Infect Immun,2001,69(9): 5726-5735.
[26]Figdor CG, van Kooyk Y, Adema GJ, et al. C-type lectin receptors on dendritic cells and Langerhans cells [J]. Nat Rev Immunol,2002,2:77-84.
[27]Jarrossay D, Napolitani G, Colonna M, et al. Specialization and complementarity in microbial molecule recognition by human myeloid and plasmacytoid dendritic cells [J].Immunol,2001,31: 3388-3393.
[28]Kadowaki N, Ho S, Antonenko S, et al. Subsets of human dendritic cell precursors express different toll-like receptors and respond to different microbial antigens [J]. Exp Med,2001,194: 863-869.
[29]Giannasca PJ and Neutral MR. Interactions of microorganisms with intestinal M cells: mucosal invasion and induction of secretory immunity [J]. Infect Agents Dis,1993,2(4):242-248.
[30]RescignoM, Urbano M, Valzasina B, et al. Dendritic cells express tight junction proteins and penetrate gut epithelial monolayersto sample bacteria [J]. Nat Immunol,2001,2(4):361-367.
[31]Yrlid U and Wick MJ. Salmonella-induced apoptosis of infected macrophages results in presentation of a bacteria-encoded antign after uptake by bystander dendritic cells [J]. J Exp Med, 2000,191(4):613-624.
[32]Coombes BK and Manhony JB. Dendritic cells discoveries provide new insight into the cellular immunobiology of DNA vaccines [J]. Immunol Lett,2001,78(2):103-111.
[33]Heath W and Carbone FR. Cross-presentation in viral immunity and selftolerance [J]. Nat Rev Immunol,2001,1:126-134.
[34]Vecino WH, Morin PM, Agha R, et al. Mucosal DNA vaccination with highly attenuated Shigella is superior to attenuated Salmonella and comparable to intramuscular DNA vaccination for T cells against HIV [J]. Immunol Lett,2002,82(3):197-204.
[35]Curtiss R. Ⅲ, Zhang X, Wanda SY, et al. Induction of host immune responses using Salmonella vectored vaccines [J]. ASM Press,2007:297-313.
[36]徐引弟,郭爱珍,陈焕春,等.减毒沙门菌载体的研究进展[J].畜牧兽医科技信息,2007,12：4-6.
[37]黄小波,曹三杰,文心田,等.减毒沙门菌载体及其在疫苗研究中的应用进展[J].中国人兽共患病学报,2008,24(5)：478-480.
[38]Toebe CS,Clements JD, Cardenas L, et al. Evaluation of immunogenicity of an oral Salmonella vaccine expressing recombinant Plasmodium berghei merozoite surface protein[J]. Am J Trop Med Hyg,1997,56(2):192-199.
[39]Somner EA, Ogun SA, Sinha KA, etal. Expression of disulphide-bridge-dependent conformational epitopes and immunogenicity of the carboxy-terminal 19kDa domain of Plasmodium yoelii merozoite surface protein-1 in live attenuated Salmonella vaccine strains [J]. Microbiol,1999,145:221-229.
[40]Chen ZH, Zhao PW, Wu SM, et al. Comparison of immune responses induced by recombinant attenuated Salmonella typhi carrying eukaryotic expression plasmid or prokaryotic expression plasmid of HCV core protein [J]. Sheng Wu Gong Cheng Xue Bao,2007,23(5):862-866.
[41]Figueroa BN, Uzzau S, Maloriol D, et al. Variable assortment of prophages provides at ransferable repertoire of pathogenic determinants in Salmonella [J]. Mol Microbiol,2001,39(2): 260-271.
[42]Sznol M, Lin SL, Bermudes D, et al. Use of preferentially replicating bacterial for the treatment of cancer [J]. Clin Med,2000,105(8):1027-1030.
[43]Covhlovius B, Stassar MG, Schreurs MW, et al. Oral DNA vaccination:antigen uptake and presentation by dendritic cell selicit s protective immunity [J]. Immunology Letters,2008,80: 89-96.
[44]Domi N, Bernal G, Tierez A, et al. Salmonella enterica serovar Choleraesuis derivatives harbouring deletions in rpoS and phoP regulatory genes are attenuated in pigs, and survive and multiply in porcine intestinal macrophages and fibroblasts respectively [J]. Vet Microbiol,2008, 130 (4):298-311.
[45]Shiau AL, Chen C, Liao Y, et al. Prothymosinenhances protective immune responses induced by oral DNA vaccination [J]. Vaccine,2001,19 (28/29):3947-3956.
[46]Shiau AL, Chen CC, Yo YT, et al. Enhancement of humoral and cellular immune responses by an oral Salmonella choleraesuis vaccine expressing porcine prothymosin-α [J]. Vaccine,2005,23 (48/49):5563-5571.
[47]白杨,王继德,张兆山,等.表达幽门杆菌黏附素保守区的减毒鼠伤寒沙门菌株的构建[J].生物工程学报,2003,19(4)：434-438.
[48]Gunn JS, Ryan SS, VanVelkinburgh JC, et al. Genetic and functional analysis of a peptide resistance, and oral virulence of Salmonella enterica serovar typhimurium [J]. Infect Immun,2000, 68 (11):6139-6146.
[49]Schriefer A, Maltez JR, Silva N, et al. Expression of a pilin subunit BfpA of the bundleforming pilus of enteropathogenic Escherichia coli in an aroA live Salmonella vaccine strain [J]. Vaccine, 1999,17(7-8):770-778.
[50]Qu D, Wang S, Cai W. Protective effect of a DNA vaccine delivered in attenuated Salmonella typhimurium against Toxoplasma gondii infection in mice [J]. Vaccine,2008,26:4541-4548.
[51]Wang J, Michel V, Leclerc C, et al. Immunogenicity of viral Bcell epitopes inserted into two surface loops of the Escherichia coli K12 LamB protein and expressed in an attenuated aroA strain of Salmonella typhimurium [J]. Vaccine,1999,17(1):1-12.
[52]李文桂,陈雅棠.细菌重组减毒鼠伤寒沙门菌疫苗的研究进展[J].动物医学进展,2004,25(2)：49-53.
[53]谢明权,覃宗华,蔡建平,等.EnMIC2重组减毒沙门菌的构建及免疫保护效果研究[J].畜牧兽医学报,2005,36(7)：705-710.
[54]王芳,焦新安,潘志明,等.融合表达淋巴细胞脉络丛脑膜炎病毒和卵清白蛋白CD8+T细胞表位的减毒鼠伤寒沙门菌的构建与鉴定[J].中国预防兽医学报,2004,26(1)：76-89.
[55]Zoller M and Christ O. Prophylactic tumor vaccination:comparison of effector mechanisms initiated by protein versus DNA vaccination [J]. Immunol,2001,166(5):3440-3450.
[56]Niethammer AG, Xiang R, Ruehlmann JM, et al. Targeted interleukin therapy enhances protective immunity induced by an autologous oral DNA vaccine against murine melanoma [J]. Cancer Res,2001,61(16):6178-6184.
[57]Xiang R, Silletti S, Lode HN, et al. Protective immunity against human carcinoembryonic antigen(CEA) induced by an oral DNA vaccine in CEA-transgenicmice [J]. Clin Cancer Res,2001, 7(3 Suppl):854-864.
[58]Niethammer AG, Primus FJ, Xiang R, et al. An oral DNAvaccine against human carcinoembryonic antigen (CEA) prevents growth and dissemination of Lewis lung carcinoma in CEA transgenic mice [J]. Vaccine,2002,20(3-4):421-429.
[59]Xiang R, Primus FJ, Ruehlmann JM, et al. A dual-function DNA vaccine encoding carcinoembryonic antigen and CD40 ligand trimer induces T cell-mediated protective immunity against colon cancer in carcinoembryonic antigen-transgenic mice [J]. Immunol,2001,167(8): 4560-4565.
[60]Schriefer A, Maltez JR, Silva N, et al. Expression of a pilin subunit BfpA of the Bundle forming pilus of enteropathogenic Escherichia coli in an aroA live Salmonella vaccine strain [J]. Vaccine,1999,17(7-8):770-778.
[61]Qu D, Wang S, Cai W, et al. Protective effect of a DNA vaccine delivered in attenuated Salmonella typhimurium against Toxoplasma gondii infection in mice [J]. Vaccine,2008,26: 4541-4548.
[62]Wang J, Michel V, Leclerc C, et al. Immunogenicity of viral B-cell epitopes inserted into two Surfaces loops of the Escherichia coli K12 LamB protein and expressed in an attenuated aroA Strain of Salmonella typhimurium [J]. Vaccine,1999,17(1):1-12.
[63]Hormaeche C E, Khan C M, Villarreal B, et al. Salmonella vaccines:Mechanisms of immunity and their use as carriers of recombinant antigens [J]. Chichester:JohnWilley&Sons,1995, 17(1):155-178.
[64]Galan JE, Nakayama K, Curtiss R.III. Cloning and characterization of the asd gene of Salmonella typhimurium:use in stable maintenance of recombinant plasmids in Salmonella vaccine [J]. Gene,1990,94:29-35.
[65]Holmgren J, Czerkinsky C, Lycke N, et al. Mucosal immunity:implications for vaccine development [J]. Immuno biol,1992,184:157-179.
[66]Gentschev I, Dietrich G, Spreng S, et al. Use of the alpha-hemolysin secretion system of Escherichia coli for antigen delivery in the Salmonella typhi Ty21a vaccine strain [J]. Int J Med Microbiol,2004,294(6):63-371.
[67]陈弟诗.猪霍乱沙门菌携带的含APP apxⅡA基因的双启动子表达载体pEPR-apxⅡA的构建[D].四川农业大学报,2007,184：157-179.
[68]Stabel TJ, Mayfield JE, Tabatabai LB, et al. Oral immunization of mice with attenuated Salmonella typhimurium containing a recombinant plasmid which codes for production of a 31-kilodalton protein of Brucella abortus [J]. Infect Immun,1990,58(7):2048-2055.
[69]Roland K, Karaca K, Sizemore D, et al. Expression of Escherichia coli antigens in Salmonella typhimurium as a vaccine to prevent airsacculitis in chickens [J]. Avian Dis,2004,48(3):595-605
[70]Zhao N, Xue Y, Wu B, et al. Subcutaneous vaccination with attenuated Salmonella enterica serovar Choleraesuis C500 expressing recombinant filamentous hemagglutinin and pertactin antigens protects mice against fatal infections with both Salmonella enterica serovar Choleraesuis and Bordetella bronchiseptica [J]. Infect Immun,2008,76(5):2157-2163.
[71]刘明秋,牛晓峰,严维耀,等.以减毒猪霍乱沙门菌为载体的抗O型口蹄疫病毒重组活菌苗在家兔体内免疫应答的初步研究[J].复旦学报：自然科学版,2005,44(4)：484-489.
[72]张小荣,潘志明,焦新安,等.运送鹅源新城疫病毒DNA疫苗的减毒沙门菌及其免疫原性[J].农业生物技术学报,2004,12(4)：412-415.
[73]Shata MT, Stevcev AL, Agwale S, et al. Recent advances with recombinant bacterial vaccine vectors [J]. Mol Med Today,2000,6(2):66-71.
[74]Baud D, Ponci F, Bobst M, et al. Improved efficiency of a Salmonella-based vaccine against human papillomavirus type 16 virus-like particles achieved by using a codon-optimized version of L1[J]. J Virol,2004,78(23):12901-12909.
[75]Woo PC, Wong LP, Zheng BJ, et al. Unique immunogenicity of hepatitis B virus DNA vaccine presented by live-attenuated Salmonella typhimurium [J].Vaccine,2001,19(22): 2945-2954.
[76]Karpenko LI, Nekrasova NA, Ilyichev AA, et al. Comparative analysis using a mouse model of the immunogenicity of artificial VLP and attenuated Salmonella strain carrying a DNA-vaccine encoding HIV-1 polyepitope CTL-immunogen [J]. Vaccine,2004,22 (14):1692-1699.
[77]乔红伟,孙金福,韩文瑜,等.猪霍乱沙门菌载体介导猪瘟病毒DNA免疫研究[J]．生物工程学报,2005,21(6)：865-870.
[78]杜爱芳,王素华,胡松华,等.鸡柔嫩艾美耳球虫5401基因真核表达载体的构建及在Vero细胞中的表达[C]//中国畜牧兽医学会家畜寄生虫学分会第五次代表大会暨第八次学术研讨会论文集,2004.
[79]杜爱芳,王素华,胡松华,等.减毒沙门菌为载体传递鸡球虫DNA疫苗的安全性和稳定性试验[C]//中国畜牧兽医学会家畜寄生虫学分会第五次代表大会暨第八次学术研讨会论文集.2004.
[80]Pogonka T, Klotz C, Kovacs F, et al. A single dose of recombinant Salmonella typhimurium induces specific humoral immune responses against heterologous Eimeria tenella antigens in chicken [J]. Int J Parasitol,2003,33(1):81-88.
[81]Stager S, Gottstein B, Muller N, et al. Systemic and local antibody response in mice induced by a recombinant peptide fragment from Giardia lamblia variant surface protein(VSP) H7 produced by a Salmonella typhimurium vaccine strain [J]. Int J Parasitol,1997,27(8):965-971.
[82]Sadoff JC, Ballou WR, Baron LS, et al. Oral Saimonella typhimurium vaccine expressing circumsporozoite protein protects against malaria [J]. Science,1988,240(4850):336-338.
[83]Jain KK. Use of bacteria as anticancer agents [J]. Expert OpinBiol Therap,2001,1(2): 291-300.
[84]Yuhua L, Kunyuan G, Hui C, et al. Oral cytokine gene therapy against murine tumor using attenuated Salmonella typhimurium [J]. Int J Cancer,2001,94(3):438-443.
[85]Urashima M, Suzuki H, Yuza Y, et al. An oral CD40 ligand gene therapy against lymphoma using attenuated Salmonella typhimurium [J]. Blood,2000,95(4):1258-1263.
[86]Cochlovius B and Stassar J. Oral DNA vaccination:antigen uptake and presentation by dendritic cells elicits protective immunity [J]. Immunol Lett,2002,80(2):89-96.
[87]Foltis BA, Sahar DA, Kim AS, et al. Cyclooxygenase-2 inhibition augments the hepatic antitumor effect of oral Salmonella typhimurium in a model of mouse metastatic colon cancer [J]. Dis Colon Rectum,2002,45(8):1023-1028.
[88]Tacket CO, Sztein MB, Wasseman SS, et al. Phase 2 clinical trial of attenuated Salmonella enterica serovar typhi oral live vector vaccine CVD 908-htrA in U.S volunteers [J]. Infect Immun, 2000,68(3):1196-1201.
[89]Krieg AM, Yi AK, Schorr J, et al. The role of CpG dinucleotides in DNA vaccines [J].Trends in Microbiology,1998,6(1):23-27.
[90]Klinman DM, Verthelyi D, Take shita F, et al. Immune recognition of foreign DNA:A cure for bioterrorism Immunity [J]. Immun,1999,11(2):123-129.
[1]房晓文等.仔猪副伤寒弱毒疫苗研究.研究报告汇辑,中国兽医药品监察所,1978,4：1-11.
[2]焦新安,刘秀梵,张如宽,等.抗沙门菌A-F血清群主要O抗原单克隆抗体及初步应用[J].中国人兽共患病杂志,1989：411-414.
[3]Juliet P M, Lynne B, Peter F R, et al.Analysis of microbial community functional diversity using sole-carbon-source utilization profiles-a critique [J]. FEMS Microbio Ecolo,2002,42(3):1-14.
[4]Rodriguez-Lazaro D, Hemandez M, Eeve T, et al. A rapid and direct real time PCR based method for identification of Salmonella spp. [J]. J Microbiol Methods,2003,54(3):381-390.
[5]郑玲,林旭,占丽钦,等.大肠杆菌asd基因PCR引物设计及其模板PCR扩增[J].福建医科大学报,1998,32(2)：145-147.
[6]王恒梁,冯尔玲,林云,等.痢疾福氏asd基因的克隆及其序列分析[J].微生物学免疫学进展,2000,128(1)：15-18.
[7]李眅,李淑琴,刘纯杰,等.鼠伤寒沙门菌口asd基因的克隆及序列分析[J].生物技术通讯,2003,14(3)：175-177.
[8]Sambrook J and Russell DW.分子克隆手册[M].黄培堂等译.北京：科学出版社.2002.
[9]Philippe N, Alcaraz JP, Coursange E, et al. Improvenment of pCVD442, a suicide plasmid for gene allele exchange in bacteria [J]. Plasmid,2004,51:246-255.
[10]Nakayama K, Kelly SM, Curtiss R.Ⅲ, et al. Construction of an ASD+ expression-cloning vector stable maintenance and high level expression of cloned genesin a Salmonella vaccine strain [J].Biol Technol,1988,6:693-697.
[11]Spreng S, Dietrich G, WeidingerG, et al. Rational design of Salmonella based vaccination strategies [J]. Methods,2006,38(5):133-143.
[12]Everest P, Frankel G, Li J, et al. Expression of LacZ from the htrA, nirB and groE promoters in a Salmonella vaccine strain:influence of growth in mammalian cells [J]. FEMS Microbiol Lett,1995, 126(10):97-102.
[13]Galan JE, Nakayama K, Curtiss R.III, et al. Cloning and characterization of the asd gene of Salmonella typhimurium:use in stable maintenance of recombinant plasmids in Salmonella vaccine strains [J]. Gene,1990,94(5):29-35.
[14]Huang CB, Feng WD, Xue MQ, et al. Oral administration of the live vaccine against Swine Paratyphoid [J]. Sci Agric Sin,1981,6(1):89-94.
[15]Schleifer KH and Kandler O. Peptidoglycan types of bacterial cell walls and their taxonomic implication [J]. Bacteriol Rev,1972,36 (4):404-407.
[16]李铁民,江崎孝行.伤寒沙门菌肠毒素基因突变株的构建[J].中国人兽共患病杂志,1999,15(5)：12-14.
[1]蔡宝祥主编.家畜传染病学[M].中国农业出版社,2001：304-309.
[2]吴明华,李维绍.鸡新城疫免疫研究[J].畜牧兽医科技信息,2006：317-318.
[3]梁华,于立辉.新城疫病毒F蛋白的研究进展[J].畜牧兽医科技信息,2005：577-578.
[4]殷震,刘景华.动物病毒学[M].北京：科学出版社,1997：739-741.
[5]Gravel KA, Morrison TG Interacting domains of the HN and F protein of Newcastle Disease Virus [J]. J Virol,2003,7720:110400-110409.
[6]Aruna PA, Subbia HE, Sateesh KT, et al. Loss of N-linked glycosylation from the hemagglutin neuraminidase protein alters virulence of Newcastle Disease Virus [J]. J Virol,2004,7810: 4965-4975.
[7]Collinms WE, Strong LA, Alexander DJ, et al. Evaluation of the molecular basis of pathogenicity of the variant Newcastle Disease Virus estermed pigeon PMV-virus [J]. Arch Virol, 1994,13(34):403-411.
[8]Sakaguchi M, Nakamura H, Sonoda K, et al. Protection of chickens from Newcastle disease by vaccination with a linear plasmid DNA expressing the F protein of Newcastle Disease Virus [J]. Vaccine,1996,14(8):747-752.
[9]许发芝,仲大莲,余为一,等.新城疫病毒(NDV)F基因的克隆及其真核表达重组质粒的构建[J].安徽农业大学,2003,30(3)：280-284.
[10]吴艳涛,刘秀梵.新城疫基因工程疫苗研究[J].中国兽药杂志,2001,35(4)：56-59.
[11]邓明俊,张彦明,梁荣,等.鸡新城疫病毒F基因高效真核表达质粒的构建[J]．西北农林科技大学学报,2003,31(2)：9-12.
[12]刘兆球,姜世金,常维山,等.新城疫病毒F基因的真核表达及其免疫原性检测[J].畜牧与兽医,2005,37(10)：5-7.
[13]程相朝,李慧琴,吴庭才,等.新城疫病毒的一般生物学特性及其致病的分子基础[J].河南农业科学,2005,3(1)：161-65.
[14]Sambrook J and Russell DW.分子克隆手册[M].黄培堂等译.北京科学出版社.2002.
[15]Sizemore DR, Branstrom AA, Sadoff JC, et al. Attenuated Shigella as a DNA delivery vehicle for DNA mediated immunization [J]. Science,1995,219(3):270-299.
[16]Sizemore DR, Branstrom AA, Sadoff JC, et al. Attenuated bacteria as a DNA delivery vehicle for DNA mediated immunization [J].Vaccine.1997,36(2):804-807.
[17]Darji A, Guzman CA, Gerstel B, et al. Oral somatic transgene vaccination using attenuated S.typhimurium [J]. Cell,1997,36(2):91-765.
[18]Pasetti MF, Anderson RJ, Noriega FR, et al. Attenuated Salmonella typhi vaccine strain CVD915 as a live vector utilizing prokaryotic or eukaryotic expression systems to deliver foreign antigens and elicit immune responses [J]. Clin Immunol,1999,921:76-89.
[19]Shiau AL, Chen YL, Liao CY, et al. Prothymosin alpha enhances protective immune responses induced by oral DNA vaccination against pseudorabies delivered by Salmonella choleraesuis [J]. Vaccine,2001,192(29):3947-3956.
[20]Yang CY, Shieh HK, Lin YL, et al. Newcastle disease virus isolated from recent outbreaks in Taiwan phylogenetically related to viruses(Genotype V) from recent outbreaks in Western Europe [J]. Avian Disease,1999,43:125-130.
[21]Sakaguchi M, Nakamura H, Sonoda K, et al. Protection of chickens from Newcastle disease by vaccination with a linear plasmid DNA expressing the F protein of Newcastle Disease Virus [J]. Vaccine,1996,14:747-752.
[22]Peeters BP, de Leeuw OS, KochG, et al. Rescue of Newcastle Disease Virus from cloned cDNA: Evidence that cleavability of the fusion protein is a major determinant for virulence [J]. Journal of Virology,1999,73:5001-5009.
[2]Merchlinsky M and Moss B. Introduction of foreign DNA into the vaccine virus genome by in vitro ligation:recombination independent selectable cloning vectors [J]. Virology,1992,190(1): 522-526.
[3]Mackett M, Smit hL, Moss B, et al. Vaccinia virus:a selectable eukaryotic cloning and expression vector [J]. Proc Natl Acad Sci USA,1982,79:7415-7419.
[4]Brochier B, Aubert MF, Pastoret PP, et al. Field use of a vaccinia-rabies recombinant vaccine for the control of sylvatic rabies in Europe and North America [J]. Rev Sci Tech,1996,5(3): 947-997.
[5]王文玲,黄保英,邓瑶,等.甲型流感病毒M2基因在痘苗病毒天坛株中的表达[J].病毒学报,2007,24(5)：377-383.
[6]Christina JK, Janice C, Matthew R, et al. Use of recombinant poxviruses to stimulate antimelanoma T Cell reactivity [J].Annu Surg Oncol,1998,5 (1):64-76.
[7]Fooks AR. Development of oral vaccines for human us [J]. Curr Opin Mol Ther,2000,2:80-86.
[8]Matsunaga Y, Wakatsuki Y, Tabata Y, et al. Oral immunization with sizepurified microsphere beads as a vehicle selectively induces systemic tolerance and sensitization [J]. Vaccine,2000,19: 579-588.
[9]Schoen C, Stritzker J, Goebel W, et al. Bacteria as DNA vaccine carriers for genetic immunization [J]. Int J Med Microbiology,2009,294:319-335.
[10]Dietrich G, Spreng S, Gentschev I, et al. Bacterial systems for the delivery of eukaryotic antigen expression vectors [J]. Antisense Nucleic Acid Drug Dev,2000,10(5):391-399.
[11]Jones BD, Ghori N, Falkow S, et al. Salmonella typhimurium initiates murine infection by penetrating and destroying the specialized epithelial M cells of the Peyer's patches [J]. Exp Med, 1994,180:15-23.
[12]Pehheiter KL, Mathur N, Giles D, et al. Non-invasive Salmonella typhimurium mutants are avirulent because of an inability to enter and destroy M cells of ileal Peyer's patches [J]. Mol Microbiol,1997,24:697-709.
[13]Vasquez TA, Jones CJ, Baumler AJ, et al. Extraintestinal dissemination of Salmonella by CD 18 expressing phagocytes [J]. Nature,1999,401:804-808.
[14]Rescigno M, Urbano M, Valzasina B, et al. Dendritic cells express tight junction proteins and penetrate gut epithelia monolayers to sample bacteria [J]. Nat Immunol,2001,2:361-367.
[15]Saunders BM and Cooper AM. Restraining mycobacteria:Role of granulomas in mycobacterial infections [J]. Immunol Cell Biol,2000,78:334-341.
[16]Sirard JC, Niedergang F, Kraehenbuhl P, et al. Live attenuated Salmonella:a paradigm of mucosal vaccines [J]. Immunol Rev,1999,171:5-26.
[17]Hopkins SA, Niedergang F, Corthesy-Theulaz IE, et al. A recombinant Salmonella typhimurium vaccine strain is taken up and survives within murine Peyer's patch dendritic cells [J]. Cell Microbiol,2000,2(2):59-68.
[18]Fastad IN, Halstense TS, Fausa O, et al. Heterogeneity of M-cell-associated B and T cells in human Peyer's patch dendritic cells [J]. Immunology,1994,83(3):457-464.
[19]Clark MA, Jepson MA, Simmons NL, et al. Selective binding and transcytosis of Ulex europaeus lectin by mouse Peyer's patch M-cells in vivo [J]. Cell Tissue Res,1995,282(3): 455-461.
[20]Vazquez TA, Jones CJ, Baumler AJ, et al. Extraintestinal dissemination of Salmonella by CD 18-expressing phagocytes [J]. Nature,1999,401(67):804-808.
[21]Rescigno M, Urbano M, Valzasina B, et al. Dendritic cells express tight junction proteins and penetrate gut epithelial monolayersto sample bacteria [J]. Nat Immunol,2001,2(4):361-367.
[22]Paglia P, Medina E, ArioliI, et al. Gene transfer in dendritic cells, induced by oral DNA vaccination with Salmonella typhimurium, results in protective immunity against a murine fibrosarcoma [J]. Blood,1998,92(9):3172-3176.
[23]Flo J, Tisminetzky S, Baralle F. Oral transgene vaccination mediated by attenuated Salmonella is an effective method to prevent herpes simplex virus induced disease in mice [J].Vaccine,2001, 19(13):1772-1782.
[24]Buchmeier, NA and Heffron F. Induction of Salmonella stress proteins upon infection of macrophages [J]. Infect Immun,1989,57(1):1-7.
[25]Yrlid U, Svensson M, Hakansson A, et al. In vivo activation of dendritic cells and T cells during Salmonella enterica serovar Typhimurium infection [J]. Infect Immun,2001,69(9): 5726-5735.
[26]Figdor CG, van Kooyk Y, Adema GJ, et al. C-type lectin receptors on dendritic cells and Langerhans cells [J]. Nat Rev Immunol,2002,2:77-84.
[27]Jarrossay D, Napolitani G, Colonna M, et al. Specialization and complementarity in microbial molecule recognition by human myeloid and plasmacytoid dendritic cells [J].Immunol,2001,31: 3388-3393.
[28]Kadowaki N, Ho S, Antonenko S, et al. Subsets of human dendritic cell precursors express different toll-like receptors and respond to different microbial antigens [J]. Exp Med,2001,194: 863-869.
[29]Giannasca PJ and Neutral MR. Interactions of microorganisms with intestinal M cells: mucosal invasion and induction of secretory immunity [J]. Infect Agents Dis,1993,2(4):242-248.
[30]RescignoM, Urbano M, Valzasina B, et al. Dendritic cells express tight junction proteins and penetrate gut epithelial monolayersto sample bacteria [J]. Nat Immunol,2001,2(4):361-367.
[31]Yrlid U and Wick MJ. Salmonella-induced apoptosis of infected macrophages results in presentation of a bacteria-encoded antign after uptake by bystander dendritic cells [J]. J Exp Med, 2000,191(4):613-624.
[32]Coombes BK and Manhony JB. Dendritic cells discoveries provide new insight into the cellular immunobiology of DNA vaccines [J]. Immunol Lett,2001,78(2):103-111.
[33]Heath W and Carbone FR. Cross-presentation in viral immunity and selftolerance [J]. Nat Rev Immunol,2001,1:126-134.
[34]Vecino WH, Morin PM, Agha R, et al. Mucosal DNA vaccination with highly attenuated Shigella is superior to attenuated Salmonella and comparable to intramuscular DNA vaccination for T cells against HIV [J]. Immunol Lett,2002,82(3):197-204.
[35]Curtiss R. Ⅲ, Zhang X, Wanda SY, et al. Induction of host immune responses using Salmonella vectored vaccines [J]. ASM Press,2007:297-313.
[36]徐引弟,郭爱珍,陈焕春,等.减毒沙门菌载体的研究进展[J].畜牧兽医科技信息,2007,12：4-6.
[37]黄小波,曹三杰,文心田,等.减毒沙门菌载体及其在疫苗研究中的应用进展[J].中国人兽共患病学报,2008,24(5)：478-480.
[38]Toebe CS,Clements JD, Cardenas L, et al. Evaluation of immunogenicity of an oral Salmonella vaccine expressing recombinant Plasmodium berghei merozoite surface protein[J]. Am J Trop Med Hyg,1997,56(2):192-199.
[39]Somner EA, Ogun SA, Sinha KA, etal. Expression of disulphide-bridge-dependent conformational epitopes and immunogenicity of the carboxy-terminal 19kDa domain of Plasmodium yoelii merozoite surface protein-1 in live attenuated Salmonella vaccine strains [J]. Microbiol,1999,145:221-229.
[40]Chen ZH, Zhao PW, Wu SM, et al. Comparison of immune responses induced by recombinant attenuated Salmonella typhi carrying eukaryotic expression plasmid or prokaryotic expression plasmid of HCV core protein [J]. Sheng Wu Gong Cheng Xue Bao,2007,23(5):862-866.
[41]Figueroa BN, Uzzau S, Maloriol D, et al. Variable assortment of prophages provides at ransferable repertoire of pathogenic determinants in Salmonella [J]. Mol Microbiol,2001,39(2): 260-271.
[42]Sznol M, Lin SL, Bermudes D, et al. Use of preferentially replicating bacterial for the treatment of cancer [J]. Clin Med,2000,105(8):1027-1030.
[43]Covhlovius B, Stassar MG, Schreurs MW, et al. Oral DNA vaccination:antigen uptake and presentation by dendritic cell selicit s protective immunity [J]. Immunology Letters,2008,80: 89-96.
[44]Domi N, Bernal G, Tierez A, et al. Salmonella enterica serovar Choleraesuis derivatives harbouring deletions in rpoS and phoP regulatory genes are attenuated in pigs, and survive and multiply in porcine intestinal macrophages and fibroblasts respectively [J]. Vet Microbiol,2008, 130 (4):298-311.
[45]Shiau AL, Chen C, Liao Y, et al. Prothymosinenhances protective immune responses induced by oral DNA vaccination [J]. Vaccine,2001,19 (28/29):3947-3956.
[46]Shiau AL, Chen CC, Yo YT, et al. Enhancement of humoral and cellular immune responses by an oral Salmonella choleraesuis vaccine expressing porcine prothymosin-α [J]. Vaccine,2005,23 (48/49):5563-5571.
[47]白杨,王继德,张兆山,等.表达幽门杆菌黏附素保守区的减毒鼠伤寒沙门菌株的构建[J].生物工程学报,2003,19(4)：434-438.
[48]Gunn JS, Ryan SS, VanVelkinburgh JC, et al. Genetic and functional analysis of a peptide resistance, and oral virulence of Salmonella enterica serovar typhimurium [J]. Infect Immun,2000, 68 (11):6139-6146.
[49]Schriefer A, Maltez JR, Silva N, et al. Expression of a pilin subunit BfpA of the bundleforming pilus of enteropathogenic Escherichia coli in an aroA live Salmonella vaccine strain [J]. Vaccine, 1999,17(7-8):770-778.
[50]Qu D, Wang S, Cai W. Protective effect of a DNA vaccine delivered in attenuated Salmonella typhimurium against Toxoplasma gondii infection in mice [J]. Vaccine,2008,26:4541-4548.
[51]Wang J, Michel V, Leclerc C, et al. Immunogenicity of viral Bcell epitopes inserted into two surface loops of the Escherichia coli K12 LamB protein and expressed in an attenuated aroA strain of Salmonella typhimurium [J]. Vaccine,1999,17(1):1-12.
[52]李文桂,陈雅棠.细菌重组减毒鼠伤寒沙门菌疫苗的研究进展[J].动物医学进展,2004,25(2)：49-53.
[53]谢明权,覃宗华,蔡建平,等.EnMIC2重组减毒沙门菌的构建及免疫保护效果研究[J].畜牧兽医学报,2005,36(7)：705-710.
[54]王芳,焦新安,潘志明,等.融合表达淋巴细胞脉络丛脑膜炎病毒和卵清白蛋白CD8+T细胞表位的减毒鼠伤寒沙门菌的构建与鉴定[J].中国预防兽医学报,2004,26(1)：76-89.
[55]Zoller M and Christ O. Prophylactic tumor vaccination:comparison of effector mechanisms initiated by protein versus DNA vaccination [J]. Immunol,2001,166(5):3440-3450.
[56]Niethammer AG, Xiang R, Ruehlmann JM, et al. Targeted interleukin therapy enhances protective immunity induced by an autologous oral DNA vaccine against murine melanoma [J]. Cancer Res,2001,61(16):6178-6184.
[57]Xiang R, Silletti S, Lode HN, et al. Protective immunity against human carcinoembryonic antigen(CEA) induced by an oral DNA vaccine in CEA-transgenicmice [J]. Clin Cancer Res,2001, 7(3 Suppl):854-864.
[58]Niethammer AG, Primus FJ, Xiang R, et al. An oral DNAvaccine against human carcinoembryonic antigen (CEA) prevents growth and dissemination of Lewis lung carcinoma in CEA transgenic mice [J]. Vaccine,2002,20(3-4):421-429.
[59]Xiang R, Primus FJ, Ruehlmann JM, et al. A dual-function DNA vaccine encoding carcinoembryonic antigen and CD40 ligand trimer induces T cell-mediated protective immunity against colon cancer in carcinoembryonic antigen-transgenic mice [J]. Immunol,2001,167(8): 4560-4565.
[60]Schriefer A, Maltez JR, Silva N, et al. Expression of a pilin subunit BfpA of the Bundle forming pilus of enteropathogenic Escherichia coli in an aroA live Salmonella vaccine strain [J]. Vaccine,1999,17(7-8):770-778.
[61]Qu D, Wang S, Cai W, et al. Protective effect of a DNA vaccine delivered in attenuated Salmonella typhimurium against Toxoplasma gondii infection in mice [J]. Vaccine,2008,26: 4541-4548.
[62]Wang J, Michel V, Leclerc C, et al. Immunogenicity of viral B-cell epitopes inserted into two Surfaces loops of the Escherichia coli K12 LamB protein and expressed in an attenuated aroA Strain of Salmonella typhimurium [J]. Vaccine,1999,17(1):1-12.
[63]Hormaeche C E, Khan C M, Villarreal B, et al. Salmonella vaccines:Mechanisms of immunity and their use as carriers of recombinant antigens [J]. Chichester:JohnWilley&Sons,1995, 17(1):155-178.
[64]Galan JE, Nakayama K, Curtiss R.III. Cloning and characterization of the asd gene of Salmonella typhimurium:use in stable maintenance of recombinant plasmids in Salmonella vaccine [J]. Gene,1990,94:29-35.
[65]Holmgren J, Czerkinsky C, Lycke N, et al. Mucosal immunity:implications for vaccine development [J]. Immuno biol,1992,184:157-179.
[66]Gentschev I, Dietrich G, Spreng S, et al. Use of the alpha-hemolysin secretion system of Escherichia coli for antigen delivery in the Salmonella typhi Ty21a vaccine strain [J]. Int J Med Microbiol,2004,294(6):63-371.
[67]陈弟诗.猪霍乱沙门菌携带的含APP apxⅡA基因的双启动子表达载体pEPR-apxⅡA的构建[D].四川农业大学报,2007,184：157-179.
[68]Stabel TJ, Mayfield JE, Tabatabai LB, et al. Oral immunization of mice with attenuated Salmonella typhimurium containing a recombinant plasmid which codes for production of a 31-kilodalton protein of Brucella abortus [J]. Infect Immun,1990,58(7):2048-2055.
[69]Roland K, Karaca K, Sizemore D, et al. Expression of Escherichia coli antigens in Salmonella typhimurium as a vaccine to prevent airsacculitis in chickens [J]. Avian Dis,2004,48(3):595-605
[70]Zhao N, Xue Y, Wu B, et al. Subcutaneous vaccination with attenuated Salmonella enterica serovar Choleraesuis C500 expressing recombinant filamentous hemagglutinin and pertactin antigens protects mice against fatal infections with both Salmonella enterica serovar Choleraesuis and Bordetella bronchiseptica [J]. Infect Immun,2008,76(5):2157-2163.
[71]刘明秋,牛晓峰,严维耀,等.以减毒猪霍乱沙门菌为载体的抗O型口蹄疫病毒重组活菌苗在家兔体内免疫应答的初步研究[J].复旦学报：自然科学版,2005,44(4)：484-489.
[72]张小荣,潘志明,焦新安,等.运送鹅源新城疫病毒DNA疫苗的减毒沙门菌及其免疫原性[J].农业生物技术学报,2004,12(4)：412-415.
[73]Shata MT, Stevcev AL, Agwale S, et al. Recent advances with recombinant bacterial vaccine vectors [J]. Mol Med Today,2000,6(2):66-71.
[74]Baud D, Ponci F, Bobst M, et al. Improved efficiency of a Salmonella-based vaccine against human papillomavirus type 16 virus-like particles achieved by using a codon-optimized version of L1[J]. J Virol,2004,78(23):12901-12909.
[75]Woo PC, Wong LP, Zheng BJ, et al. Unique immunogenicity of hepatitis B virus DNA vaccine presented by live-attenuated Salmonella typhimurium [J].Vaccine,2001,19(22): 2945-2954.
[76]Karpenko LI, Nekrasova NA, Ilyichev AA, et al. Comparative analysis using a mouse model of the immunogenicity of artificial VLP and attenuated Salmonella strain carrying a DNA-vaccine encoding HIV-1 polyepitope CTL-immunogen [J]. Vaccine,2004,22 (14):1692-1699.
[77]乔红伟,孙金福,韩文瑜,等.猪霍乱沙门菌载体介导猪瘟病毒DNA免疫研究[J]．生物工程学报,2005,21(6)：865-870.
[78]杜爱芳,王素华,胡松华,等.鸡柔嫩艾美耳球虫5401基因真核表达载体的构建及在Vero细胞中的表达[C]//中国畜牧兽医学会家畜寄生虫学分会第五次代表大会暨第八次学术研讨会论文集,2004.
[79]杜爱芳,王素华,胡松华,等.减毒沙门菌为载体传递鸡球虫DNA疫苗的安全性和稳定性试验[C]//中国畜牧兽医学会家畜寄生虫学分会第五次代表大会暨第八次学术研讨会论文集.2004.
[80]Pogonka T, Klotz C, Kovacs F, et al. A single dose of recombinant Salmonella typhimurium induces specific humoral immune responses against heterologous Eimeria tenella antigens in chicken [J]. Int J Parasitol,2003,33(1):81-88.
[81]Stager S, Gottstein B, Muller N, et al. Systemic and local antibody response in mice induced by a recombinant peptide fragment from Giardia lamblia variant surface protein(VSP) H7 produced by a Salmonella typhimurium vaccine strain [J]. Int J Parasitol,1997,27(8):965-971.
[82]Sadoff JC, Ballou WR, Baron LS, et al. Oral Saimonella typhimurium vaccine expressing circumsporozoite protein protects against malaria [J]. Science,1988,240(4850):336-338.
[83]Jain KK. Use of bacteria as anticancer agents [J]. Expert OpinBiol Therap,2001,1(2): 291-300.
[84]Yuhua L, Kunyuan G, Hui C, et al. Oral cytokine gene therapy against murine tumor using attenuated Salmonella typhimurium [J]. Int J Cancer,2001,94(3):438-443.
[85]Urashima M, Suzuki H, Yuza Y, et al. An oral CD40 ligand gene therapy against lymphoma using attenuated Salmonella typhimurium [J]. Blood,2000,95(4):1258-1263.
[86]Cochlovius B and Stassar J. Oral DNA vaccination:antigen uptake and presentation by dendritic cells elicits protective immunity [J]. Immunol Lett,2002,80(2):89-96.
[87]Foltis BA, Sahar DA, Kim AS, et al. Cyclooxygenase-2 inhibition augments the hepatic antitumor effect of oral Salmonella typhimurium in a model of mouse metastatic colon cancer [J]. Dis Colon Rectum,2002,45(8):1023-1028.
[88]Tacket CO, Sztein MB, Wasseman SS, et al. Phase 2 clinical trial of attenuated Salmonella enterica serovar typhi oral live vector vaccine CVD 908-htrA in U.S volunteers [J]. Infect Immun, 2000,68(3):1196-1201.
[89]Krieg AM, Yi AK, Schorr J, et al. The role of CpG dinucleotides in DNA vaccines [J].Trends in Microbiology,1998,6(1):23-27.
[90]Klinman DM, Verthelyi D, Take shita F, et al. Immune recognition of foreign DNA:A cure for bioterrorism Immunity [J]. Immun,1999,11(2):123-129.
[1]房晓文等.仔猪副伤寒弱毒疫苗研究.研究报告汇辑,中国兽医药品监察所,1978,4：1-11.
[2]焦新安,刘秀梵,张如宽,等.抗沙门菌A-F血清群主要O抗原单克隆抗体及初步应用[J].中国人兽共患病杂志,1989：411-414.
[3]Juliet P M, Lynne B, Peter F R, et al.Analysis of microbial community functional diversity using sole-carbon-source utilization profiles-a critique [J]. FEMS Microbio Ecolo,2002,42(3):1-14.
[4]Rodriguez-Lazaro D, Hemandez M, Eeve T, et al. A rapid and direct real time PCR based method for identification of Salmonella spp. [J]. J Microbiol Methods,2003,54(3):381-390.
[5]郑玲,林旭,占丽钦,等.大肠杆菌asd基因PCR引物设计及其模板PCR扩增[J].福建医科大学报,1998,32(2)：145-147.
[6]王恒梁,冯尔玲,林云,等.痢疾福氏asd基因的克隆及其序列分析[J].微生物学免疫学进展,2000,128(1)：15-18.
[7]李眅,李淑琴,刘纯杰,等.鼠伤寒沙门菌口asd基因的克隆及序列分析[J].生物技术通讯,2003,14(3)：175-177.
[8]Sambrook J and Russell DW.分子克隆手册[M].黄培堂等译.北京：科学出版社.2002.
[9]Philippe N, Alcaraz JP, Coursange E, et al. Improvenment of pCVD442, a suicide plasmid for gene allele exchange in bacteria [J]. Plasmid,2004,51:246-255.
[10]Nakayama K, Kelly SM, Curtiss R.Ⅲ, et al. Construction of an ASD+ expression-cloning vector stable maintenance and high level expression of cloned genesin a Salmonella vaccine strain [J].Biol Technol,1988,6:693-697.
[11]Spreng S, Dietrich G, WeidingerG, et al. Rational design of Salmonella based vaccination strategies [J]. Methods,2006,38(5):133-143.
[12]Everest P, Frankel G, Li J, et al. Expression of LacZ from the htrA, nirB and groE promoters in a Salmonella vaccine strain:influence of growth in mammalian cells [J]. FEMS Microbiol Lett,1995, 126(10):97-102.
[13]Galan JE, Nakayama K, Curtiss R.III, et al. Cloning and characterization of the asd gene of Salmonella typhimurium:use in stable maintenance of recombinant plasmids in Salmonella vaccine strains [J]. Gene,1990,94(5):29-35.
[14]Huang CB, Feng WD, Xue MQ, et al. Oral administration of the live vaccine against Swine Paratyphoid [J]. Sci Agric Sin,1981,6(1):89-94.
[15]Schleifer KH and Kandler O. Peptidoglycan types of bacterial cell walls and their taxonomic implication [J]. Bacteriol Rev,1972,36 (4):404-407.
[16]李铁民,江崎孝行.伤寒沙门菌肠毒素基因突变株的构建[J].中国人兽共患病杂志,1999,15(5)：12-14.
[1]蔡宝祥主编.家畜传染病学[M].中国农业出版社,2001：304-309.
[2]吴明华,李维绍.鸡新城疫免疫研究[J].畜牧兽医科技信息,2006：317-318.
[3]梁华,于立辉.新城疫病毒F蛋白的研究进展[J].畜牧兽医科技信息,2005：577-578.
[4]殷震,刘景华.动物病毒学[M].北京：科学出版社,1997：739-741.
[5]Gravel KA, Morrison TG Interacting domains of the HN and F protein of Newcastle Disease Virus [J]. J Virol,2003,7720:110400-110409.
[6]Aruna PA, Subbia HE, Sateesh KT, et al. Loss of N-linked glycosylation from the hemagglutin neuraminidase protein alters virulence of Newcastle Disease Virus [J]. J Virol,2004,7810: 4965-4975.
[7]Collinms WE, Strong LA, Alexander DJ, et al. Evaluation of the molecular basis of pathogenicity of the variant Newcastle Disease Virus estermed pigeon PMV-virus [J]. Arch Virol, 1994,13(34):403-411.
[8]Sakaguchi M, Nakamura H, Sonoda K, et al. Protection of chickens from Newcastle disease by vaccination with a linear plasmid DNA expressing the F protein of Newcastle Disease Virus [J]. Vaccine,1996,14(8):747-752.
[9]许发芝,仲大莲,余为一,等.新城疫病毒(NDV)F基因的克隆及其真核表达重组质粒的构建[J].安徽农业大学,2003,30(3)：280-284.
[10]吴艳涛,刘秀梵.新城疫基因工程疫苗研究[J].中国兽药杂志,2001,35(4)：56-59.
[11]邓明俊,张彦明,梁荣,等.鸡新城疫病毒F基因高效真核表达质粒的构建[J]．西北农林科技大学学报,2003,31(2)：9-12.
[12]刘兆球,姜世金,常维山,等.新城疫病毒F基因的真核表达及其免疫原性检测[J].畜牧与兽医,2005,37(10)：5-7.
[13]程相朝,李慧琴,吴庭才,等.新城疫病毒的一般生物学特性及其致病的分子基础[J].河南农业科学,2005,3(1)：161-65.
[14]Sambrook J and Russell DW.分子克隆手册[M].黄培堂等译.北京科学出版社.2002.
[15]Sizemore DR, Branstrom AA, Sadoff JC, et al. Attenuated Shigella as a DNA delivery vehicle for DNA mediated immunization [J]. Science,1995,219(3):270-299.
[16]Sizemore DR, Branstrom AA, Sadoff JC, et al. Attenuated bacteria as a DNA delivery vehicle for DNA mediated immunization [J].Vaccine.1997,36(2):804-807.
[17]Darji A, Guzman CA, Gerstel B, et al. Oral somatic transgene vaccination using attenuated S.typhimurium [J]. Cell,1997,36(2):91-765.
[18]Pasetti MF, Anderson RJ, Noriega FR, et al. Attenuated Salmonella typhi vaccine strain CVD915 as a live vector utilizing prokaryotic or eukaryotic expression systems to deliver foreign antigens and elicit immune responses [J]. Clin Immunol,1999,921:76-89.
[19]Shiau AL, Chen YL, Liao CY, et al. Prothymosin alpha enhances protective immune responses induced by oral DNA vaccination against pseudorabies delivered by Salmonella choleraesuis [J]. Vaccine,2001,192(29):3947-3956.
[20]Yang CY, Shieh HK, Lin YL, et al. Newcastle disease virus isolated from recent outbreaks in Taiwan phylogenetically related to viruses(Genotype V) from recent outbreaks in Western Europe [J]. Avian Disease,1999,43:125-130.
[21]Sakaguchi M, Nakamura H, Sonoda K, et al. Protection of chickens from Newcastle disease by vaccination with a linear plasmid DNA expressing the F protein of Newcastle Disease Virus [J]. Vaccine,1996,14:747-752.
[22]Peeters BP, de Leeuw OS, KochG, et al. Rescue of Newcastle Disease Virus from cloned cDNA: Evidence that cleavability of the fusion protein is a major determinant for virulence [J]. Journal of Virology,1999,73:5001-5009.