鳗弧菌展示系统和平衡—致死宿主载体系统的开发
|
摘要
鳗弧菌是一类可以在世界范围内引起淡水和海水鱼类疾病大爆发的重要病原菌。本实验室前期工作从鳗弧菌野生株MVM425出发,通过重组DNA技术得到一株减毒株,初步的免疫试验表明,MVAV6203有较大的潜力开发成为成功的减毒活菌疫苗。
为了在鳗弧菌减毒活疫苗的基础上开发潜在的多价重组疫苗,将保护性抗原蛋白递呈到载体表面,需要引入高效的表面展示系统。本实验利用来源于鳗弧菌的外膜蛋白作为锚定元件,构建了一系列新型表面展示系统。该类系统的锚定元件由两部分组成：1)鳗弧菌外膜脂蛋白Wza信号肽及其N端的7个氨基酸或者大肠杆菌主要外膜脂蛋白Lpp信号肽以及N端的九个氨基酸；2)鳗弧菌外膜蛋白Omporfl, OmpU或Omp26La的跨膜区。将绿色荧光蛋白作为报告基因插入到这些系统中,构建的六个表面展示系统中有四个可以成功地将GFP展示到大肠杆菌细胞的表面；与Wza-Omporfl融合的来源于大黄鱼虹彩病毒的主要衣壳蛋白基因,也可以成功地展示到鳗弧菌减毒株MVAV6203的表面；通过将来源于迟钝爱德华氏菌的EseB抗原蛋白与六个展示系统融合,并转入MVAV6203中,可以将EseB展示到鳗弧菌表面,从而构建了一系列多价疫苗候选株。之后选用候选株AV/pW-26La-B,利用斑马鱼对其进行免疫保护评价,结果显示该菌株可以保护斑马鱼免受鳗弧菌感染,对迟钝爱德华氏菌感染的前期也能起到一定的保护作用。这一结果证明,可以通过在减毒鳗弧菌中利用表面展示将外源保护性抗原展示到细胞表面的方法开发多价疫苗。
同时我们还将MCP融合到大肠杆菌α-溶血素分泌系统中,在大肠杆菌和鳗弧菌MVAV6203中考察其分泌情况。在大肠杆菌宿主中,可以检测到约400μg／L的MCP被分泌出来,然而鳗弧菌中却没有分泌。
为了优化鳗弧菌疫苗株,以开发不含抗性标记的重组减毒鳗弧菌疫苗,我们尝试通过缺失relA、天冬氨酸半醛脱氢酶基因asd并插入TTaraC PBAD c2的方法在鳗弧菌中构建平衡-致死宿主载体系统。我们发现鳗弧菌中含有两个具有功能的asd基因asdA和asdB,且asdA和asdB都可以互补来源于其他菌种的asd突变株,但是asdA和asdB双突变对鳗弧菌却不致死。对AsdA和AsdB的生物学特性进行研究发现,AsdA与AsdB的3D结构有所不同,AsdB的结构更趋向于同一些革兰氏阳性菌的ASD结构相似,两个ASD的序列分析也显示了相同的结果。同时对鳗弧菌三磷酸甘油醛脱氢酶GAPDH的分析发现,该蛋白是被分泌到胞外的。
Vibrio anguillarum is an important bacterial fish pathogen, responsible for both marine and freshwater fish epizootics throughout the world. In our previous work, several attenuated V. anguillarum strains derived from wild-type V. anguillarum strain MVM425 were constructed with recombinant DNA technology and proved to be excellent live vaccine candidates against Vibrio pathogens.
For developing potential multivalent recombinant vaccines based on attenuated V. anguillarum live vaccine, efficient surface display systems are needed to display protective antigens onto the surface of live carrier. In this work, a series of novel cell surface display systems were examined by employing V. anguillarum outer membrane protein as anchoring motifs. These display systems consist of two parts. The first is the signal sequence and first eleven N-terminal amino acids of V. anguillarum outer membrane lipoprotein Wza, or the signal sequence and first nine N-terminal amino acids of the mature major Escherichia coli lipoprotein Lpp, and the second part is the transmembrane domains of V. anguillarum outer membrane proteins Omporfl, OmpU or Omp26La. Green fluorescent protein (GFP) was inserted to the systems and the results of GFP surface localization confirmed that four of the six surface display systems could successfully display GFP on Escherichia coli surface. The major capsid protein (MCP) of large yellow croaker iridovirus (LYCIV) was fused with Wza-Omporfl system and was displayed on the surface of an attenuated V. anguillarum MVAV6203, and a putative antigen protein EseB from pathogenic Edwardsiella tarda was successfully expressed on the surface of MVAV6203 to get multivalent vaccine candidates. The immune protection evaluation in zebra fish (Danio rerio) demonstrated that the V. anguillarum EseB-display strain AV/pW-26La-B could trigger a full protection against V. anguillarum infection and an early protection against E. tarda infection in the immunized fish. These results suggest that surface display of heterologous protective antigens in attenuated V. anguillarum could be used as a tool to develop potential V. anguillarum vector vaccine.
At the same time, the secretion of MCP based on the E. coliα-haemolysin transport system was investigated in both E. coli and attenuated V. anguillarum MVAV6203. With the aid of E. coli a-haemolysin transport system, about 400μg/L MCP was secreted into culture supernatant in E. coli, while intracellularly expressed MCP in MVAV6203 was not secreted.
In order to optimize the V. anguillarum vaccine strain and develop an antibiotic-sensitive recombinant attenuated Vibrio vaccine, we try to develop a balanced-lethal host-vector system by delete the V. anguillarum relA gene, aspartate-semialdehyde dehydrogenase gene asd and insert TTaraC PBAD c2. We found that V. anguillarum contained two asd genes, asdA and asdB. Each of two genes functioned to complement different asd mutants from other bacterial species, and double deletions of asdA and asdB in V. anguillarum were not lethal. In the 3D structure analysis of AsdA and AsdB it was shown that AsdB isoform is quite different from the AsdA isoform and in fact more closely resembles the Gram-positive spASD structure. The result was confirmed in their alignment analysis. We also found that in V. anguillarum the glyceraldehyde-3-phosphate dehydrogenase can be secreted to the supernatant.
为了在鳗弧菌减毒活疫苗的基础上开发潜在的多价重组疫苗,将保护性抗原蛋白递呈到载体表面,需要引入高效的表面展示系统。本实验利用来源于鳗弧菌的外膜蛋白作为锚定元件,构建了一系列新型表面展示系统。该类系统的锚定元件由两部分组成：1)鳗弧菌外膜脂蛋白Wza信号肽及其N端的7个氨基酸或者大肠杆菌主要外膜脂蛋白Lpp信号肽以及N端的九个氨基酸；2)鳗弧菌外膜蛋白Omporfl, OmpU或Omp26La的跨膜区。将绿色荧光蛋白作为报告基因插入到这些系统中,构建的六个表面展示系统中有四个可以成功地将GFP展示到大肠杆菌细胞的表面；与Wza-Omporfl融合的来源于大黄鱼虹彩病毒的主要衣壳蛋白基因,也可以成功地展示到鳗弧菌减毒株MVAV6203的表面；通过将来源于迟钝爱德华氏菌的EseB抗原蛋白与六个展示系统融合,并转入MVAV6203中,可以将EseB展示到鳗弧菌表面,从而构建了一系列多价疫苗候选株。之后选用候选株AV/pW-26La-B,利用斑马鱼对其进行免疫保护评价,结果显示该菌株可以保护斑马鱼免受鳗弧菌感染,对迟钝爱德华氏菌感染的前期也能起到一定的保护作用。这一结果证明,可以通过在减毒鳗弧菌中利用表面展示将外源保护性抗原展示到细胞表面的方法开发多价疫苗。
同时我们还将MCP融合到大肠杆菌α-溶血素分泌系统中,在大肠杆菌和鳗弧菌MVAV6203中考察其分泌情况。在大肠杆菌宿主中,可以检测到约400μg／L的MCP被分泌出来,然而鳗弧菌中却没有分泌。
为了优化鳗弧菌疫苗株,以开发不含抗性标记的重组减毒鳗弧菌疫苗,我们尝试通过缺失relA、天冬氨酸半醛脱氢酶基因asd并插入TTaraC PBAD c2的方法在鳗弧菌中构建平衡-致死宿主载体系统。我们发现鳗弧菌中含有两个具有功能的asd基因asdA和asdB,且asdA和asdB都可以互补来源于其他菌种的asd突变株,但是asdA和asdB双突变对鳗弧菌却不致死。对AsdA和AsdB的生物学特性进行研究发现,AsdA与AsdB的3D结构有所不同,AsdB的结构更趋向于同一些革兰氏阳性菌的ASD结构相似,两个ASD的序列分析也显示了相同的结果。同时对鳗弧菌三磷酸甘油醛脱氢酶GAPDH的分析发现,该蛋白是被分泌到胞外的。
Vibrio anguillarum is an important bacterial fish pathogen, responsible for both marine and freshwater fish epizootics throughout the world. In our previous work, several attenuated V. anguillarum strains derived from wild-type V. anguillarum strain MVM425 were constructed with recombinant DNA technology and proved to be excellent live vaccine candidates against Vibrio pathogens.
For developing potential multivalent recombinant vaccines based on attenuated V. anguillarum live vaccine, efficient surface display systems are needed to display protective antigens onto the surface of live carrier. In this work, a series of novel cell surface display systems were examined by employing V. anguillarum outer membrane protein as anchoring motifs. These display systems consist of two parts. The first is the signal sequence and first eleven N-terminal amino acids of V. anguillarum outer membrane lipoprotein Wza, or the signal sequence and first nine N-terminal amino acids of the mature major Escherichia coli lipoprotein Lpp, and the second part is the transmembrane domains of V. anguillarum outer membrane proteins Omporfl, OmpU or Omp26La. Green fluorescent protein (GFP) was inserted to the systems and the results of GFP surface localization confirmed that four of the six surface display systems could successfully display GFP on Escherichia coli surface. The major capsid protein (MCP) of large yellow croaker iridovirus (LYCIV) was fused with Wza-Omporfl system and was displayed on the surface of an attenuated V. anguillarum MVAV6203, and a putative antigen protein EseB from pathogenic Edwardsiella tarda was successfully expressed on the surface of MVAV6203 to get multivalent vaccine candidates. The immune protection evaluation in zebra fish (Danio rerio) demonstrated that the V. anguillarum EseB-display strain AV/pW-26La-B could trigger a full protection against V. anguillarum infection and an early protection against E. tarda infection in the immunized fish. These results suggest that surface display of heterologous protective antigens in attenuated V. anguillarum could be used as a tool to develop potential V. anguillarum vector vaccine.
At the same time, the secretion of MCP based on the E. coliα-haemolysin transport system was investigated in both E. coli and attenuated V. anguillarum MVAV6203. With the aid of E. coli a-haemolysin transport system, about 400μg/L MCP was secreted into culture supernatant in E. coli, while intracellularly expressed MCP in MVAV6203 was not secreted.
In order to optimize the V. anguillarum vaccine strain and develop an antibiotic-sensitive recombinant attenuated Vibrio vaccine, we try to develop a balanced-lethal host-vector system by delete the V. anguillarum relA gene, aspartate-semialdehyde dehydrogenase gene asd and insert TTaraC PBAD c2. We found that V. anguillarum contained two asd genes, asdA and asdB. Each of two genes functioned to complement different asd mutants from other bacterial species, and double deletions of asdA and asdB in V. anguillarum were not lethal. In the 3D structure analysis of AsdA and AsdB it was shown that AsdB isoform is quite different from the AsdA isoform and in fact more closely resembles the Gram-positive spASD structure. The result was confirmed in their alignment analysis. We also found that in V. anguillarum the glyceraldehyde-3-phosphate dehydrogenase can be secreted to the supernatant.
引文
[1]Odd-Jarl B. New organizational forms within the aquaculture industry:the fish-farming enterprise as "virtual" organization. Aquac Econ Manag.1999,3:105-116
[2]王利,谢长勇.鱼类疫苗研究进展.动物医学进展.2003,24(5)：26-28
[3]Van Regennortel MHV, Fauquet CM, Bishop DHL, Carstens EB, Estes MK, Lemon SM, Maniloff J, Mayo MA, McGeoch DJ, Pringle CR, Wickner EB. Virus Taxonomy. Seventh Report of the International Committee on Taxonomy of Viruses. Academic Press, New York.2000
[4]Langdon JS, Humphrey JD, Williams LM, Hyatt AD, Westbury HA. First virus isolation from Australian fish:an iridovirus-like pathogen from redfin perch, Perca fluviatilis L. J Fish Dis.1986,9: 262-268
[5]Pozet F, Morand M, Moussa A, Torhy C, de Kinkelin P. Isolation and characterization of a pathogenic icosahedral deoxyribovirus from the catfish Ictalurus melas. Dis Aquat Org.1992,14:35-42
[6]Ahne W, Schlotfeldt HJ, Thomsen I. Fish viruses:isolation of an icosahedral cytoplasmic deoxyribovirus from sheatfish(Silurus glanis). J Vet Med B.1989,36:333-336
[7]Bloch B, Larsen JL. An iridovirus-like agent associated with systemic infection in cultured turbot Scophthalmus maximus fry in Denmark. Dis Aquat Org.1993,15:235-240
[8]Tapiovaara H, Olesen NJ, Linden J, Rimaila-Pamanen, E, Von Bonsdorff CH. Isolation of an iridovirus from pike-perch Stizostedion lucioperca. Dis Aquat Org.1998,32:185-193
[9]Plumb JA, Grizzle JM, Young HE, Noyes AD, Lamprecht S. An iridovirus isolated from wild largemouth bass. J Aquat Anim Health.1996,8:265-270
[10]Inouye K, Yamano K, Maeno Y, Nakajima K, Matsuoka M, Wada Y, Sorimachi M. Iridovirus infection of cultured red sea bream, Pagrus major. Fish Pathol.1992,27:19-27
[11]Chua FHC, Ng ML, Ng KL, Loo JJ, Wee JY. Investigation of outbreaks of a novel disease,'Sleepy Grouper Disease', affecting the brown-spotted grouper, Epinephelus tauvina Forskal. J Fish Dis.1994, 17:417-427
[12]Kasornchandra J, Khongpradit R. Isolation and preliminary characterization of a pathogenic iridovirus-like agent in nursing grouper Epinephelus malabaricus. Asian Aquat Health New.1995:4-5
[13]Qin QW, Chang SF, Ngoh GH, Gibson-Kueh S, Shi C, Lam TJ.Characterization of a novel ranavirus isolated from grouper, Epinephelus tauvina. Dis Aquat Org.2003,53:1-9
[14]Hyatt AD, Gould AR, Zupanovic Z, Cunningham AA, Hengstberger S, Whittington RJ, Kattenbelt J, Coupar BEH. Comparative studies of piscine and amphibian iridoviruses. Arch Virol.2000,145: 301-331
[15]Piaskoski TO, Plumb JA. Characterization of the largemouth bass virus in cell culture. J Aquat Ani. Health.1999,11:45-51
[16]Huang C, Zhang X, Karina YHG, Qin QW. In situ hybridization of a marine fish virus, Singapore grouper iridovirus with a nucleic acid probe of major capsid protein. J Virol Methods.2004,117: 123-128
[17]Williams T, Chinchar GD, Darai G, Hyatt AD, Kalmakoff J, Seligy VL. Family iridoviridae. In van Regenmortel MHV, Fauquet CM, Bishop DHL, Carstens EB, Estes MK, Lemon SM, Maniloff J, Mayo MA, McGeoch DJ, Pringle CR, Wickner RB. (Eds.) Virus Taxonomy. Seventh Report of the International Committee on Taxonomy of Viruses. Academic Press, San Diego/Wien, New York.2000, p167-182
[18]Monini M, Ruggeri FM. Antigenic peptides of the epizootic hematopoietic necrosis virus. Virology. 2002,297:8-18
[19]Willis DB, Granof A. Frog virus 3 DNA is heavily methylated at CpG sequence. Virology.1980,107: 250-257
[20]Tidona CA, Schnizler P, Kehm R. Identification of the gene encoding the DNA(cytosine-5) methyltransferase of lymphocystis disease virus. Virus Genes.1996,12:219-229
[21]Willis DB, Goorha R, Miles M, Granoff A. Macromolecular synthesis in cells infected by frog virus 3. VII. Transcriptional and post-transcriptional regulation of virus gene expression. J Virol.1977,24: 326-342
[22]Mao J, Tham TN, Gentry GA, Aubertin A, Chinchar VG. Cloning, sequence analysis, and expression of the major capsid protein of the iridovirus frog virus 3. Virology.1996,216:431-436
[23]Kaur K, Rohozinski J, Goorha R. Identification and characterization of the frog virus 3 DNA methyltransferase gene. J Gen Virol.1995,76:1937-1943
[24]张奇亚.我国水生动物病毒病研究进展.水生生物学报.2002,26：89-102
[25]陈翠珍.爱德华氏菌及鱼类爱德华氏菌病.河北科技师范学院学报.2004,18(3)：70-76
[26]Castro N, Toranzo AE, Barja JL, Nunez S, Magarinos B. Characterization of Edwardsiella tarda strains isolated from turbot, Psetta maxima (L.). J Fish Dis.2006,29:541-547
[27]Meyer FP, Bullock GL. Edwardsiella tarda, a new pathogen of channel catfish(Ictalurus punctatus). Appl Microbiol.1973,25:155-156
[28]Acharya M, Maiti NK, Mohanty S, Mishra P, Samanta M. Genotyping of Edwardsiella tarda isolated from freshwater fish culture system. Comp Immunol Microbiol Infect Dis.2007,30:33-40
[29]Grimont PAD, Grimont F, Richard C, Sakazaki R. Edwardsiella hoshinae, a new species of Enterobacteriaceae. Curr Microbiol.1980,4:347-351
[30]Fang H, Zhang X, Chen C, Jin X, Wang X. Studies on the edwardsiellosis and characterization of pathogenic bacteria from diseased flounder (Paralichthys olivaceus L.) and turbot (Scophthalmus maximus L.). Acta Oceanol Sin.2006,25:138-147
[31]张晓君,战文斌,陈翠珍,房海.牙鲆迟钝爱德华氏菌感染症及其病原的研究.水生生物学报.2005,29：31-37
[32]Austin B, Austin DA. Bacterial Fish Pathogens:Disease in Farmed and Wild Fish.3rd ed.1999. Chichester, UK:Parxis Publishing
[33]Actis LA, Tomlmasky ME, Crosa JH. Vibriosis. In:Woo PTK, Bruno DW (eds.). Fish Disease and Disorders. Volume 3. Viral, Bacterial, and Fugal Infections. Cab International Publishing Wallingford, Unites Kingdom.1999, p523-557
[34]Neilands JB. Microbial iron compounds. Annu Rev Microbiol.1981,50:715-731
[35]Neilands JB, Raymonds JB. Microbial envelope protein related to iron. Annu Rev Microbiol.1982,36: 285-309
[36]Harbell SO, Hodgins HO, Schiewe MH. Studies on the pathology of vibriosis in coho salmon. J Fish Dis.1979,2:527-535
[37]Crosa JH. A plasmid associated with virulence in the marine fish pathogen Vibrio anguillarum specifies and iron-sequestering system. Nature.1980,284:566-568
[38]Pedersen K, Gram L, Austin DA, Austin B. Pathogenicity of Vibrio anguillarum serogroup O1 strains compared to plasmids, outer membrane protein profiles and siderophore production. J Appl Microbiol. 1997,83:365-371
[39]Tolmasky ME, Actis LA, Crosa JH. Genetic analysis of the iron uptake region of the Vibrio anguillarum plasmid pJMl:molecular cloning of genetic determinants encoding a novel trans activator of siderophore biosynthesis. JBacteriol.1988,170:1913-1919
[40]Actis LA, Fish W, Crosa JH, Kellerman K, Ellenberger SR, Jauser FM, Sander-Loehr J. Characterization of anguibactin, a novel siderophore from Vibrio anguillarum 775 (pJMl). J Bacterial. 1986,167:57-65
'[41] Actis LA, Potter S, Crosa JH. Iron-regulated outer membrane protein OM2 of Vibrio anguillarum is encoded by virulence plasmid pJMl. J Bacteriol.1985,161:736-742
[42]Koter WL, Actis LA, Waldbeser LS, Tolmasky ME, Crosa JH. Molecular characterization of the iron transport system mediated by the pJMl plasmid in Vibrio anguillarum 775. J Biol Chem.1991,266: 23829-23833
[43]Crosa JH. Genetics and molecular biology of siderophore-mediated iron transport in bacteria. Microbiol Rev.1989,53:517-530
[44]Wu HZ, Ma Y, Zhang YX, Zhang HZ. Complete sequence of virulence plasmid pEIB1 from the marine fish pathogen Vibrio anguillarum strain MVM425 and location of its replication region. J Appl Microbiol.2004,97:1021-1028
[45]Liu Q, Ma Y, Wu HZ, Shao MF, Liu HF, Zhang YX. Cloning, identification and expression of an entE homologue angE from Vibrio anguillarum serotype O1. Arch Microbiol.2004,181:287-293
[46]Liu Q, Ma Y, Zhou LY, Zhang YX. Gene cloning, expression and functional characterization of a phosphopantetheinyl transferase from Vibrio anguillarum serotype O1. Arch Microbiol.2005,183: 37-44
[47].朱向军,张之伦.新疫苗的发展和应用展望.口岸卫生控制.2002,4：32-34
[48]Arnon R, Ben-Yedidia T. Old and new vaccine approaches. Int Immunopharmacol.2003,3:1195-1204
[49]Gudding R, Lillehaug A, Evensen O. Recent developments in fish vaccinology. Vet Immunol Immunopathol.1999,72:203-212
[50]Tang DC, Devit M, Johnston SA. Genetic immunization is a simple method for eliciting an immune response. Nature.1992,356:152-154
[51].罗晓春.鱼类疫苗最新进展.鱼类病害研究.2000,1：81-85
[52]Frey J. Biological safety concepts of genetically modified live bacterial vaccines. Vaccine.2007,25: 5598-5605
[53].郭文龙,朱瑞良.基因工程亚单位疫苗的研究现状及发展动态.猪与禽.2008,28：72-75
[54]夏春玲,胡栋友,郭学军,崔磊.DNA疫苗及其在动物传染病中的应用.中国家禽.2005,z1：185-188
[55]程相朝,张敏,王建军,王婷.DNA疫苗的主要意义与特点.河南科技大学学报(医学版).2005,23：236-238
[56]Medina E, Guzmain CA. Use of live bacterial vaccine vectors for antigen delivery:potential and limitations. Vaccine.2001,19:1573-1580
[57]Schodel F. Prospects for oral vaccination using recombinant bacteria expressing viral epitopes. Adv Virus Res.1992,41:409-446
[58]于力,童光志,赵雅芝.病毒活载体疫苗的构建原理及应用.中国预防兽医学报.1997,5：61-64
[59]Fath MJ, Kolter R. ABC transporters:bacterial exporters. Microbiol Rev.1993,12:995-1017
[60]Finlay BB, Falkow S. Common Themes in Microbial Pathogenicity Revisited. Microbiol Mol Biol Rev. 1997,61:136-169
[61]Salmond GP, Reeves PJ. Membrane traffic wardens and protein secretion in gram-negative bacteria. Trends Biochem Sci.1993,18:7-12
[62]Van Gijsegem F, Genin S, Boucher C. Conservation of secretion pathways for pathogenicity determinants of plant and animal bacteria. Trends Microbiol.1993,1:175-180
[63]Andersson K, Carballeira N, Magnusson KE, Persson C, Stendahl O, Wolf-Watz H, Fallman M. YopH of Yersinia pseudotuberculosis interrupts early phosphotyrosine signalling associated with phagocytosis. Mol Microbiol.1996,20:1057-1069
[64]Chervaux C, Holland IB. Random and directed mutagenesis to elucidate the functional importance of helix II and F-989 in the C-terminal secretion signal of Escherichia coli hemolysin. J Bacteriol.1996, 178:1232-1236
[65]Christoph JH. Type III protein secretion systems in bacterial pathogens of animals and plants. Microbiol Mol Biol Rev.1998,62:379-433
[66]Welch RA. Haemolysin contributes to virulence of extra-intestinal E. coli infections. Nature.1981, 284:665-667
[67]Saurin W. Getting in or out:early segregation between importers and exporters in the evolution of ATP-binding cassette (ABC) transporters. J Mol Evol.1999,48:22-41
[68]Dinh T. A family of extracytoplasmic proteins that allow transport of large molecules across the outer membranes of Gram-negative bacteria. J Bacteriol.1994,176:3825-3831
[69]Jennifer AL, Welch RA. Enhancing transcription through the Escherichia coli hemolysin operon, hlyCABD:RfaH and upstream JUMPStart DNA sequences function together via a postinitiation mechanism. J Bacteriol.1997,179:3519-3527
[70]Heinz PH, Bernd-Ulrich VS. Secretory delivery of recombinant proteins in attenuated Salmonella strains:potential and limitations of Type I protein transporters. FEMS Immunol Med Mic.2003,37: 87-98
[71]Gentschev I, Dietrich G, Goebel W. The E. coli a-hemolysin secretion system and its use in vaccine development. Trends Microbiol.2002,1:39-45
[72]Blight MA, Holland IB. Heterologous protein secretion and the versatile Escherichia coli haemolysin translocator. Trends Biotechnol.1994,12:450-455
[73]Mollenkopf H-J, Gentschev I, Bubert A, Schubert P, Goebel W. Extracellular PagC-HlyAs fusion protein for the generation and identification of Salmonella-specific antibodies. Appl Microbiol Biotechnol.1996,45:629-637
[74]Hess J, Gentschev I, Miko D, Welzel M, Ladel C, Goebel W, Kaufmann SHE. Superior efficacy of secreted over somatic antigen display in recombinant Salmonella vaccine induced protection against listeriosis. Proc Natl Acad Sci USA.1996,93:1458-1463
[75]Lee JS, Shin KS, Pan JG, Kim CJ. Surface-displayed viral antigens on Salmonella carrier vaccine. Nat Biotechnol.2000,18:645-648
[76]Freudl R, Maclntyre S, Degen M, Henning U. Cell surface exposure of the outer membrane protein OmpA of Escherichia coli K-12. J Mol Biol.1986,188:491-494
[77]Charbit A, Boulain JC, Ryter A, Hofnung M. Probing the topology of a bacterial membrane protein by genetic insertion of a foreign epitope; expression at the cell surface. EMBO J.1986,5:3029-3037
[78]Sang YL, Jong HC, Xu ZH. Microbial cell-surface display. Trends Biotechnol.2003,21:45-52
[79]李元刚,尹向东,王玉龙.细菌活载体疫苗的研究进展.黑龙江畜牧兽医.2004,2：63-65
[80]Sandkvist M, Bagdasarian M. Secretion of recombinant proteins by Gram-negative bacteria. Curr Opin Biotechnol.1996,7:505-511
[81]Francisco JA, Earhart CF, Georgiou G. Transport and anchoring of beta-lactamase to the external surface of Escherichia coli. Proc Natl Acad Sci USA.1992,89:2713-2717
[82]Wan HM, Chang BY, Lin SC. Anchorage of cyclodextrin glucanotransferase on the outer membrane of Escherichia coli. Biotechnol Bioeng.2002,79:457-464
[83]Prasher DC, Eckenrode VK, Ward WW, Prendergast FG, Cormier MJ. Primary structure of the Aequorea victoria green-fluorescent protein. Gene.1992,2:229-233
[84]Yang F, Moss LG, Phillips GN Jr. The molecular structure of green fluorescent protein. Nat Biotechnol. 1996,14:1246-1251
[85]Cubitt AB, Heim R, Adams SR, Boyd AE, Gross LA, Tsien RY. Understanding, improving and using green fluorescent proteins. Trends Biochem Sci.1995,11:448-455
[86]Reid BG, Flynn GC. Chromophore formation in green fluorescent protein. Biochemistry.1997,22: 6786-6791
[87]徐飞虎,龚兴国.绿色荧光蛋白应用研究进展.细胞生物学杂志.2002,24：332-334
[88]Caipang CMA, Takano T, Hirono I, Aoki T. Genetic vaccines protect red seabream, Pagrus major, upon challenge with red seabream iridovirus (RSIV). Fish Shellfish Immunol.2006,21:130-138
[89]Georgiou G, Stathopoulos C, Daugherty PS, Nayak AR, Iverson BL, Curtiss R. Display of heterologous proteins on the surface of microorganisms:from the screening of combinatorial libraries to live recombinant vaccines. Nat Biotechnol.1997.15:29-34
[90]Gentschev I, Mollenkopf H, Sokolovic Z, Hess J, Kaufmann SH, Goebel W. Development of antigen-delivery systems, based on the Escherichia coli hemolysin secretion pathway. Gene.1996,179: 133-140
[91]Shao M, Ma Y, Liu Q, Zhang Y. Secretory expression of recombinant proteins in an attenuated Vibrio anguillarum strain for potential use in vaccines. J Fish Dis.2005,28:723-728
[92]吴海珍.海洋鱼类病原菌鳗弧菌毒力质粒pEIB1的全序列测定及复制区的定位.华东理工大学博士学位论文.2005.
[93]Kern I, Ceglowski P. Secretion of streptokinase fusion proteins from Escherichia coli cells through the hemolysin transporter. Gene.1995,163:53-57
[94]Gentschev I, Sokolovic Z, Mollenkopf HJ, Hess J, Kaufmann SH, Kuhn M, Krohne GF, Goebel W. Salmonella strain secreting active listeriolysin changes its intracellular location. Infect Immun.1995,63: 4202-4205
[95]Li Y, Chen CX, von Specht BU, Hahn HP. Cloning and hemolysin-mediated secretory expression of a codon-optimized synthetic human interleukin-6 gene in Escherichia coli. Prot Expr Purif.2002,25: 437-447
[96]Hess J, Gentschev I, Szalay G, Ladel C, Bubert A, Goebel W, Kaufmann SH. Listeria monocytogenes p60 supports host cell invasion by and in vivo survival of attenuated Salmonella typhimurium. Infect Immun.1995,63:2047-2053
[97]Chang YF, Lauderdale TL, Lee WY, Shin SJ, Jacobson RH, Appel MJ, Lein, DH. Expression and secretion of outer surface protein (OSP-A) of Borrelia burgdorferi from Escherichia coli. FEMS Microbiol Lett.1993,109:297-302
[98]Mollenkopf HJ, Gentschev I, Goebel W. Conversion of bacterial gene products to secretion-competent fusion proteins. Biotechniques.1996,21:854-860
[99]Gentschev I, Glaser I, Goebel W, McKeever DJ, Musoke A, Heussler VT. Delivery of the p67 sporozoite antigen of Theileria parva using recombinant Salmonella:secretion of the product enhances specific antibody responses in cattle. Infect Immun.1998,66:2060-2064
[100]Su G, Brahmbhatt N, de Lorenzo V, Wehland J, Timmis K. Extracellular export of Shiga toxin B-subunit/haemolysin A (C-terminus) fusion protein expressed in Salmonella typhimurium aroA-mutant and stimulation of B-subunit specific antibody responses in mice. Microb Pathog.1992,13:465-476
[101]Orr N, Galen JE, Levine MM. Expression and immunogenicity of a mutant diphtheria toxin molecule, CRM (197), and its fragments in Salmonella typhi vaccine strain CVD 908-htrA. Infect Immun.1999, 67:4290-4294
[102]Li Y, Reichenstein K, Ullrich R, Danner T, von Specht BU, Hahn HP. Effect of in situ expression of human interleukin-6 on antibody responses against Salmonella typhimurium antigens. FEMS Immunol Med Microbiol.2003,37:135-145
[103]Xi CW, Lambrecht M, Vanderleyden J, Michiels J. Bi-functional gfp-and gusA-containing mini-Tn5 transposon derivatives for combined gene expression and bacterial localization studies. J Microb Methods.1999,35:85-92
[104]王蓬勃.溶藻弧菌铁摄取和调控机制的初步研究.华东理工大学硕士学位论文.2005.
[105]Sambrook J, Russell DW.黄培堂译.分子克隆实验指南,第三版.科学出版社,2002
[106]周丽,刘洪明,战文斌,宋微波.鳗弧菌、溶藻胶弧菌外膜蛋白的分离及特性.中国水产科学.2003,10：31-35
[107]Kawai K, Liu Y, Ohnishi K, Oshima SI. A conserved 37 kD outer membrane protein of Edwardsiella tarda is an effective vaccine candidate. Vaccine.2004,22:3411-3418
[108]Shi HD, Su WW. Display of fluorescent protein on Escherichia coli cell surface. Enzyme Microb Technol.2000,28:25-34
[109]Georgiou G, Stephens DL, Stathopoulos C. Display of beta-lactamase on the Escherichia coli surface outer membrane phenotypes conferred by Lpp-OmpA-beta-lactamase fusions. Protein Eng.1996,9: 239-247
[110]Whitfield C, Paiment A. Biosynthesis and assembly of Group 1 capsular polysaccharides in Escherichia coli and related extracellular polysaccharides in other bacteria. Carbohyd Res.2003,338: 2491-2502
[111]Kang SM, Rhee JK, Kim EJ, Han KH, Oh JW. Bacterial cell surface display for epitope mapping of hepatitis C virus core antigen. FEMS Microbiol Lett.2003,226:347-353
[112]Kim EJ, Yoo SK. Cell surface display of hepatitis B virus surface antigen by using Pseudomonas syringae ice nucleation protein. Lett Appl Microbiol.1999,29:292-297
[113]Tan YP, Zheng J, Tung SL, Rosenshine I, Leung KY. Role of type III secretion in Edwardsiella tarda virulence. Microbiology.2005,151:2301-2313
[114]Zheng J, Tung SL, Leung KY. Regulation of a type III and a putative secretion system in Edwardsiella tarda by EsrC is under the control of a two-component system, EsrA-EsrB. Infect Immun. 2005,73:4127-4137
[115]Rao PS, Yamada Y, Tan YP, Leung KY. Use of proteomics to identify novel virulence determinants that are required for Edwardsiella tarda pathogenesis. Mol Microbiol.2004,53:573-586
[116]Davey HM, Kell DB. Flow cytometry and cell sorting of heterogeneous microbial populations:the importance of single-cell analyses. Microbiol Rev.1996,60:641-696
[117]Grogan WM, Collins JM. Guide to Flow Cytometry Methods. New York:Marcel Dekker,1990, p29-190
[118]Porter J, Deere D, Hardman M, Edwards C, Pickup R. Go with the flow-use of flow cytometry in environmental microbiology. FEMS Microbiol Ecol.1997,24:93-101
[119]McSharry JJ. Uses of flow cytometry in virology. Clin Microbiol Rev.1994,7:576-604
[120]Wallner G, Amann R, Beisker W. Optimizing fluorescent in situ hybridization with rRNA-targeted oligonucleotide probes for flow cytometric identification of microorganisms. Cytometry.1993,14: 136-143
[121]Impert MBM, Nell R, Anne SP, Burel MC, Cantarovich D, Milpied N, Billaudel S. Development of a method direct quantification of cytomegalovirus antigenemia by flow cytometry. Clin Microbiol Infect. 1997,10:2665-2669
[122]Yurkow EJ, McKenzie MA. Characterization of hypoxiadependent peroxide production in cultures of Saccharomyces cerevisiae using flow cytometry:a model for ischemic tissue destruction. Cytometry. 1993,14:287-293
[123]Allman R, Hann AC, Manchee R, Lloyd D. Characterization of bacteria by multi parameter flow cytometry. J Appl Bacteriol.1992,73:438-444
[124]Amann R, Ludwig L, Scheleifer KH. Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol Rev.1995,59:143-169
[125]Islam D, Lindberg AA, Christensson B. Peripheral blood cell prepare influences the level of expression of leukocyte cell surface marshal assessed with quantitative multicolor flow cytometry. Cytometry.1996,22:128-134
[126]Pinder AC, Purdy PW, Poulter SA, Clark DC. Validation of flow cytometry for rapid enumeration of bacterial concentrations in pure cultures. J Appl Bacteriol.1990,69:92-100
[127]Kaprelyants A, Kell DB. Rapid assessment of bacterial viability and vitality using rhodamine 123 and flow cytometry. J Appl Bacteriol.1992,72:410-422
[128]Lebaron P, Parthuisot N, Catala P. Comparison of blue nucleic acid dyes for flow cytometric enumeration of bacteria in aquatic systems. Appl Environ Microbiol.1998,64:1725-1730
[129]Lopez-Amoros R, Castel S, Comas-Riu J, Vives-Rego J. Assessment of E. coli and Salmonella viability and starvation by confocal laser microscopy and flow cytometry using rhodamine 123, DiBAC4(3), propidium iodide, and CTC. Cytometry.1997,29:298-305
[130]Tidona CA, Schnitzler P, Kehm R, Darai G. Is the major capsid protein of iridoviruses a suitable target for the study of viral evolution? Virus Genes.1998,16:59-66
[131]许煜洲.基于冰核蛋白的鳗弧菌表面展示系统的构建.华东理工大学硕士学位论文.2008.
[132]杨路,马悦,张元兴.鳗弧菌MVAV6203基因缺失株的生物反应器培养及检测.水产科学.2007,26：191-194
[133]邵明非.鳗弧菌基于大肠杆菌α-溶血素分泌系统分泌表达外源蛋白的研究.华东理工大学硕士学位论文.2005.
[134]Nakayama K, Kelly SM, Curtiss III R. Construction of an ASD+expression-cloning vector:stable maintenance and high level expression of cloned genes in a Salmonella vaccine strain. Bio/Technol. 1988:6,693-697
[135]Viola RE. The central enzymes of the aspartate family of amino acid biosynthesis. Acc Chem Res. 2001,34:339-349
[136]Karsten WE, Viola RE. Chemical and kinetic mechanisms of aspartate-beta-semialdehyde dehydrogenase from Escherichia coli. Biochim Biophys Acta.1991,1077:209-219
[137]Blanco J, Moore RA, Kabaleeswaran V, Viola RE. A structural basis for the mechanism of aspartate-beta-semialdehyde dehydrogenase from Vibrio cholerae. Protein Sci.2003,12:27-33
[138]Faehnle CR, Ohren JF, Viola RE. A new branch in the family:structure of aspartate-beta-semialdehyde dehydrogenase from Methanococcus jannaschii. J Mol Biol.2005,353: 1055-1068
[139]Paidhungat M, Setlow B, Driks A, Setlow P. Characterization of spores of Bacillus subtilis which lack dipicolinic acid. J Bacteriol.2000,182:5505-5512
[140]Pavelka Jr MS, Jacobs Jr WR. Biosynthesis of diaminopimelate, the precursor of lysine and a component of peptidoglycan, is an essential function of Mycobacterium smegmatis. J Bacteriol.1996, 178:6496-6507
[141]Viola RE, Liu X, Ohren JF, Faehnle CR. The structure of a redundant enzyme:a second isoform of aspartate β-semialdehyde dehydrogenase in Vibrio cholerae. Acta Crystallogr D Biol Crystallogr.2008, 64:321-330
[142]Harb OS, Kwaik YA. Identification of the aspartate-beta-semialdehyde dehydrogenase gene of Legionella pneumophila and characterization of a null mutant. Infect Immun.1998,66:1898-1903
[143]Angeles TS, Hunsley JR, Viola RE. Reversal of enzyme regiospecificity with alternative substrates for aspartokinase I from Escherichia coli. Biochemistry.1992,31:799-805
[144]Kang HY, Srinivasan J, Curtiss III R. Immune responses to recombinant Pneumococcal PspA antigen delivered by live attenuated Salmonella enterica serovar Typhimurium vaccine. Infect Immun.2002,4: 1739-1749
[145]Tijhaar EJ, Zheng-Xin Y, Karlas JA, Meyer TF, Stukart MJ, Osterhaus ADME, Mooi FR. Construction and evaluation of an expression vector allowing the satable expression of foreign antigens in a Salmonella typhimurium vaccine strain. Vaccine.1994,12:1004-1111
[146]Chatfield S, Charles IQ Makoff AJ, Oxer MD, Dougan G, Pickard D, Slater D, Fairweather NF. Use of the nirB promoter to direct the stable expression of heterologous antigens in Salmonella oral vaccine strains:Development of a single-dose oral tetanus vaccine. Bio/Technol.1992,10:888-892
[147]Hohmann EL, Oletta CA, Miller SI. Evaluation of a phoP/phoQ-deleted, aroA-deleted live oral Salmonella typhi vaccine strain in human volunteers. Vaccine.1996,14:19-24
[148]Roberts M, Li J, Bacon A, Chatfield S. Oral vaccination against tetanus:Comparison of the immunogenicities of Salmonella strains expressing fragment C form the nirB and htrA promoters. Infect Immun.1998,66:3080-3087
[149]Orr N, Galen JE, Levine MM. Novel use of anaerobically induced promoter, dmsA, for controlled expressing of fragment C of tetanus toxin in live attenuated Salmonella enterica serovar Typhi strain CVC 908-htrA. Vaccine.2001,19:1694-1700
[150]Kong W, Wanda SY, Curtiss III R. Construction and application of hot-vector systems for DNA vaccine vector delivery [A-016]. In American Society for Microbiology,103rd General Meeting, Washington, DG:American Society for Microbiology,677
[151]Andrianopoulos K, Wang L, Reeves PR. Identification of the fucose synthetase gene in the colonic acid gene cluster of Escherichia coli K-12. J Bacteriol.1998,180:998-1001
[152]Moore RA, Bocik WE, Viola RE. Expression and purification of L-aspartate-beta-semi aldehyde dehydrogenase from infectious microorganisms. Protein Expr Purif.2002,25:189-194
[153]Gouet P, Courcelle E, Stuart DI, Metoz F. ESPript:analysis of multiple sequence alignments in PostScript. Bioinformatics.1999,15:305-308
[154]Liu Y, Oshima S, Kawai K. Glyceraldehyde-3-phosphate dehydrogenase of Edwardsiella tarda has protective antigenicity against Vibrio anguillarum in Japanese flounder. Dis Aquat Org.2007,75: 217-220
[2]王利,谢长勇.鱼类疫苗研究进展.动物医学进展.2003,24(5)：26-28
[3]Van Regennortel MHV, Fauquet CM, Bishop DHL, Carstens EB, Estes MK, Lemon SM, Maniloff J, Mayo MA, McGeoch DJ, Pringle CR, Wickner EB. Virus Taxonomy. Seventh Report of the International Committee on Taxonomy of Viruses. Academic Press, New York.2000
[4]Langdon JS, Humphrey JD, Williams LM, Hyatt AD, Westbury HA. First virus isolation from Australian fish:an iridovirus-like pathogen from redfin perch, Perca fluviatilis L. J Fish Dis.1986,9: 262-268
[5]Pozet F, Morand M, Moussa A, Torhy C, de Kinkelin P. Isolation and characterization of a pathogenic icosahedral deoxyribovirus from the catfish Ictalurus melas. Dis Aquat Org.1992,14:35-42
[6]Ahne W, Schlotfeldt HJ, Thomsen I. Fish viruses:isolation of an icosahedral cytoplasmic deoxyribovirus from sheatfish(Silurus glanis). J Vet Med B.1989,36:333-336
[7]Bloch B, Larsen JL. An iridovirus-like agent associated with systemic infection in cultured turbot Scophthalmus maximus fry in Denmark. Dis Aquat Org.1993,15:235-240
[8]Tapiovaara H, Olesen NJ, Linden J, Rimaila-Pamanen, E, Von Bonsdorff CH. Isolation of an iridovirus from pike-perch Stizostedion lucioperca. Dis Aquat Org.1998,32:185-193
[9]Plumb JA, Grizzle JM, Young HE, Noyes AD, Lamprecht S. An iridovirus isolated from wild largemouth bass. J Aquat Anim Health.1996,8:265-270
[10]Inouye K, Yamano K, Maeno Y, Nakajima K, Matsuoka M, Wada Y, Sorimachi M. Iridovirus infection of cultured red sea bream, Pagrus major. Fish Pathol.1992,27:19-27
[11]Chua FHC, Ng ML, Ng KL, Loo JJ, Wee JY. Investigation of outbreaks of a novel disease,'Sleepy Grouper Disease', affecting the brown-spotted grouper, Epinephelus tauvina Forskal. J Fish Dis.1994, 17:417-427
[12]Kasornchandra J, Khongpradit R. Isolation and preliminary characterization of a pathogenic iridovirus-like agent in nursing grouper Epinephelus malabaricus. Asian Aquat Health New.1995:4-5
[13]Qin QW, Chang SF, Ngoh GH, Gibson-Kueh S, Shi C, Lam TJ.Characterization of a novel ranavirus isolated from grouper, Epinephelus tauvina. Dis Aquat Org.2003,53:1-9
[14]Hyatt AD, Gould AR, Zupanovic Z, Cunningham AA, Hengstberger S, Whittington RJ, Kattenbelt J, Coupar BEH. Comparative studies of piscine and amphibian iridoviruses. Arch Virol.2000,145: 301-331
[15]Piaskoski TO, Plumb JA. Characterization of the largemouth bass virus in cell culture. J Aquat Ani. Health.1999,11:45-51
[16]Huang C, Zhang X, Karina YHG, Qin QW. In situ hybridization of a marine fish virus, Singapore grouper iridovirus with a nucleic acid probe of major capsid protein. J Virol Methods.2004,117: 123-128
[17]Williams T, Chinchar GD, Darai G, Hyatt AD, Kalmakoff J, Seligy VL. Family iridoviridae. In van Regenmortel MHV, Fauquet CM, Bishop DHL, Carstens EB, Estes MK, Lemon SM, Maniloff J, Mayo MA, McGeoch DJ, Pringle CR, Wickner RB. (Eds.) Virus Taxonomy. Seventh Report of the International Committee on Taxonomy of Viruses. Academic Press, San Diego/Wien, New York.2000, p167-182
[18]Monini M, Ruggeri FM. Antigenic peptides of the epizootic hematopoietic necrosis virus. Virology. 2002,297:8-18
[19]Willis DB, Granof A. Frog virus 3 DNA is heavily methylated at CpG sequence. Virology.1980,107: 250-257
[20]Tidona CA, Schnizler P, Kehm R. Identification of the gene encoding the DNA(cytosine-5) methyltransferase of lymphocystis disease virus. Virus Genes.1996,12:219-229
[21]Willis DB, Goorha R, Miles M, Granoff A. Macromolecular synthesis in cells infected by frog virus 3. VII. Transcriptional and post-transcriptional regulation of virus gene expression. J Virol.1977,24: 326-342
[22]Mao J, Tham TN, Gentry GA, Aubertin A, Chinchar VG. Cloning, sequence analysis, and expression of the major capsid protein of the iridovirus frog virus 3. Virology.1996,216:431-436
[23]Kaur K, Rohozinski J, Goorha R. Identification and characterization of the frog virus 3 DNA methyltransferase gene. J Gen Virol.1995,76:1937-1943
[24]张奇亚.我国水生动物病毒病研究进展.水生生物学报.2002,26：89-102
[25]陈翠珍.爱德华氏菌及鱼类爱德华氏菌病.河北科技师范学院学报.2004,18(3)：70-76
[26]Castro N, Toranzo AE, Barja JL, Nunez S, Magarinos B. Characterization of Edwardsiella tarda strains isolated from turbot, Psetta maxima (L.). J Fish Dis.2006,29:541-547
[27]Meyer FP, Bullock GL. Edwardsiella tarda, a new pathogen of channel catfish(Ictalurus punctatus). Appl Microbiol.1973,25:155-156
[28]Acharya M, Maiti NK, Mohanty S, Mishra P, Samanta M. Genotyping of Edwardsiella tarda isolated from freshwater fish culture system. Comp Immunol Microbiol Infect Dis.2007,30:33-40
[29]Grimont PAD, Grimont F, Richard C, Sakazaki R. Edwardsiella hoshinae, a new species of Enterobacteriaceae. Curr Microbiol.1980,4:347-351
[30]Fang H, Zhang X, Chen C, Jin X, Wang X. Studies on the edwardsiellosis and characterization of pathogenic bacteria from diseased flounder (Paralichthys olivaceus L.) and turbot (Scophthalmus maximus L.). Acta Oceanol Sin.2006,25:138-147
[31]张晓君,战文斌,陈翠珍,房海.牙鲆迟钝爱德华氏菌感染症及其病原的研究.水生生物学报.2005,29：31-37
[32]Austin B, Austin DA. Bacterial Fish Pathogens:Disease in Farmed and Wild Fish.3rd ed.1999. Chichester, UK:Parxis Publishing
[33]Actis LA, Tomlmasky ME, Crosa JH. Vibriosis. In:Woo PTK, Bruno DW (eds.). Fish Disease and Disorders. Volume 3. Viral, Bacterial, and Fugal Infections. Cab International Publishing Wallingford, Unites Kingdom.1999, p523-557
[34]Neilands JB. Microbial iron compounds. Annu Rev Microbiol.1981,50:715-731
[35]Neilands JB, Raymonds JB. Microbial envelope protein related to iron. Annu Rev Microbiol.1982,36: 285-309
[36]Harbell SO, Hodgins HO, Schiewe MH. Studies on the pathology of vibriosis in coho salmon. J Fish Dis.1979,2:527-535
[37]Crosa JH. A plasmid associated with virulence in the marine fish pathogen Vibrio anguillarum specifies and iron-sequestering system. Nature.1980,284:566-568
[38]Pedersen K, Gram L, Austin DA, Austin B. Pathogenicity of Vibrio anguillarum serogroup O1 strains compared to plasmids, outer membrane protein profiles and siderophore production. J Appl Microbiol. 1997,83:365-371
[39]Tolmasky ME, Actis LA, Crosa JH. Genetic analysis of the iron uptake region of the Vibrio anguillarum plasmid pJMl:molecular cloning of genetic determinants encoding a novel trans activator of siderophore biosynthesis. JBacteriol.1988,170:1913-1919
[40]Actis LA, Fish W, Crosa JH, Kellerman K, Ellenberger SR, Jauser FM, Sander-Loehr J. Characterization of anguibactin, a novel siderophore from Vibrio anguillarum 775 (pJMl). J Bacterial. 1986,167:57-65
'[41] Actis LA, Potter S, Crosa JH. Iron-regulated outer membrane protein OM2 of Vibrio anguillarum is encoded by virulence plasmid pJMl. J Bacteriol.1985,161:736-742
[42]Koter WL, Actis LA, Waldbeser LS, Tolmasky ME, Crosa JH. Molecular characterization of the iron transport system mediated by the pJMl plasmid in Vibrio anguillarum 775. J Biol Chem.1991,266: 23829-23833
[43]Crosa JH. Genetics and molecular biology of siderophore-mediated iron transport in bacteria. Microbiol Rev.1989,53:517-530
[44]Wu HZ, Ma Y, Zhang YX, Zhang HZ. Complete sequence of virulence plasmid pEIB1 from the marine fish pathogen Vibrio anguillarum strain MVM425 and location of its replication region. J Appl Microbiol.2004,97:1021-1028
[45]Liu Q, Ma Y, Wu HZ, Shao MF, Liu HF, Zhang YX. Cloning, identification and expression of an entE homologue angE from Vibrio anguillarum serotype O1. Arch Microbiol.2004,181:287-293
[46]Liu Q, Ma Y, Zhou LY, Zhang YX. Gene cloning, expression and functional characterization of a phosphopantetheinyl transferase from Vibrio anguillarum serotype O1. Arch Microbiol.2005,183: 37-44
[47].朱向军,张之伦.新疫苗的发展和应用展望.口岸卫生控制.2002,4：32-34
[48]Arnon R, Ben-Yedidia T. Old and new vaccine approaches. Int Immunopharmacol.2003,3:1195-1204
[49]Gudding R, Lillehaug A, Evensen O. Recent developments in fish vaccinology. Vet Immunol Immunopathol.1999,72:203-212
[50]Tang DC, Devit M, Johnston SA. Genetic immunization is a simple method for eliciting an immune response. Nature.1992,356:152-154
[51].罗晓春.鱼类疫苗最新进展.鱼类病害研究.2000,1：81-85
[52]Frey J. Biological safety concepts of genetically modified live bacterial vaccines. Vaccine.2007,25: 5598-5605
[53].郭文龙,朱瑞良.基因工程亚单位疫苗的研究现状及发展动态.猪与禽.2008,28：72-75
[54]夏春玲,胡栋友,郭学军,崔磊.DNA疫苗及其在动物传染病中的应用.中国家禽.2005,z1：185-188
[55]程相朝,张敏,王建军,王婷.DNA疫苗的主要意义与特点.河南科技大学学报(医学版).2005,23：236-238
[56]Medina E, Guzmain CA. Use of live bacterial vaccine vectors for antigen delivery:potential and limitations. Vaccine.2001,19:1573-1580
[57]Schodel F. Prospects for oral vaccination using recombinant bacteria expressing viral epitopes. Adv Virus Res.1992,41:409-446
[58]于力,童光志,赵雅芝.病毒活载体疫苗的构建原理及应用.中国预防兽医学报.1997,5：61-64
[59]Fath MJ, Kolter R. ABC transporters:bacterial exporters. Microbiol Rev.1993,12:995-1017
[60]Finlay BB, Falkow S. Common Themes in Microbial Pathogenicity Revisited. Microbiol Mol Biol Rev. 1997,61:136-169
[61]Salmond GP, Reeves PJ. Membrane traffic wardens and protein secretion in gram-negative bacteria. Trends Biochem Sci.1993,18:7-12
[62]Van Gijsegem F, Genin S, Boucher C. Conservation of secretion pathways for pathogenicity determinants of plant and animal bacteria. Trends Microbiol.1993,1:175-180
[63]Andersson K, Carballeira N, Magnusson KE, Persson C, Stendahl O, Wolf-Watz H, Fallman M. YopH of Yersinia pseudotuberculosis interrupts early phosphotyrosine signalling associated with phagocytosis. Mol Microbiol.1996,20:1057-1069
[64]Chervaux C, Holland IB. Random and directed mutagenesis to elucidate the functional importance of helix II and F-989 in the C-terminal secretion signal of Escherichia coli hemolysin. J Bacteriol.1996, 178:1232-1236
[65]Christoph JH. Type III protein secretion systems in bacterial pathogens of animals and plants. Microbiol Mol Biol Rev.1998,62:379-433
[66]Welch RA. Haemolysin contributes to virulence of extra-intestinal E. coli infections. Nature.1981, 284:665-667
[67]Saurin W. Getting in or out:early segregation between importers and exporters in the evolution of ATP-binding cassette (ABC) transporters. J Mol Evol.1999,48:22-41
[68]Dinh T. A family of extracytoplasmic proteins that allow transport of large molecules across the outer membranes of Gram-negative bacteria. J Bacteriol.1994,176:3825-3831
[69]Jennifer AL, Welch RA. Enhancing transcription through the Escherichia coli hemolysin operon, hlyCABD:RfaH and upstream JUMPStart DNA sequences function together via a postinitiation mechanism. J Bacteriol.1997,179:3519-3527
[70]Heinz PH, Bernd-Ulrich VS. Secretory delivery of recombinant proteins in attenuated Salmonella strains:potential and limitations of Type I protein transporters. FEMS Immunol Med Mic.2003,37: 87-98
[71]Gentschev I, Dietrich G, Goebel W. The E. coli a-hemolysin secretion system and its use in vaccine development. Trends Microbiol.2002,1:39-45
[72]Blight MA, Holland IB. Heterologous protein secretion and the versatile Escherichia coli haemolysin translocator. Trends Biotechnol.1994,12:450-455
[73]Mollenkopf H-J, Gentschev I, Bubert A, Schubert P, Goebel W. Extracellular PagC-HlyAs fusion protein for the generation and identification of Salmonella-specific antibodies. Appl Microbiol Biotechnol.1996,45:629-637
[74]Hess J, Gentschev I, Miko D, Welzel M, Ladel C, Goebel W, Kaufmann SHE. Superior efficacy of secreted over somatic antigen display in recombinant Salmonella vaccine induced protection against listeriosis. Proc Natl Acad Sci USA.1996,93:1458-1463
[75]Lee JS, Shin KS, Pan JG, Kim CJ. Surface-displayed viral antigens on Salmonella carrier vaccine. Nat Biotechnol.2000,18:645-648
[76]Freudl R, Maclntyre S, Degen M, Henning U. Cell surface exposure of the outer membrane protein OmpA of Escherichia coli K-12. J Mol Biol.1986,188:491-494
[77]Charbit A, Boulain JC, Ryter A, Hofnung M. Probing the topology of a bacterial membrane protein by genetic insertion of a foreign epitope; expression at the cell surface. EMBO J.1986,5:3029-3037
[78]Sang YL, Jong HC, Xu ZH. Microbial cell-surface display. Trends Biotechnol.2003,21:45-52
[79]李元刚,尹向东,王玉龙.细菌活载体疫苗的研究进展.黑龙江畜牧兽医.2004,2：63-65
[80]Sandkvist M, Bagdasarian M. Secretion of recombinant proteins by Gram-negative bacteria. Curr Opin Biotechnol.1996,7:505-511
[81]Francisco JA, Earhart CF, Georgiou G. Transport and anchoring of beta-lactamase to the external surface of Escherichia coli. Proc Natl Acad Sci USA.1992,89:2713-2717
[82]Wan HM, Chang BY, Lin SC. Anchorage of cyclodextrin glucanotransferase on the outer membrane of Escherichia coli. Biotechnol Bioeng.2002,79:457-464
[83]Prasher DC, Eckenrode VK, Ward WW, Prendergast FG, Cormier MJ. Primary structure of the Aequorea victoria green-fluorescent protein. Gene.1992,2:229-233
[84]Yang F, Moss LG, Phillips GN Jr. The molecular structure of green fluorescent protein. Nat Biotechnol. 1996,14:1246-1251
[85]Cubitt AB, Heim R, Adams SR, Boyd AE, Gross LA, Tsien RY. Understanding, improving and using green fluorescent proteins. Trends Biochem Sci.1995,11:448-455
[86]Reid BG, Flynn GC. Chromophore formation in green fluorescent protein. Biochemistry.1997,22: 6786-6791
[87]徐飞虎,龚兴国.绿色荧光蛋白应用研究进展.细胞生物学杂志.2002,24：332-334
[88]Caipang CMA, Takano T, Hirono I, Aoki T. Genetic vaccines protect red seabream, Pagrus major, upon challenge with red seabream iridovirus (RSIV). Fish Shellfish Immunol.2006,21:130-138
[89]Georgiou G, Stathopoulos C, Daugherty PS, Nayak AR, Iverson BL, Curtiss R. Display of heterologous proteins on the surface of microorganisms:from the screening of combinatorial libraries to live recombinant vaccines. Nat Biotechnol.1997.15:29-34
[90]Gentschev I, Mollenkopf H, Sokolovic Z, Hess J, Kaufmann SH, Goebel W. Development of antigen-delivery systems, based on the Escherichia coli hemolysin secretion pathway. Gene.1996,179: 133-140
[91]Shao M, Ma Y, Liu Q, Zhang Y. Secretory expression of recombinant proteins in an attenuated Vibrio anguillarum strain for potential use in vaccines. J Fish Dis.2005,28:723-728
[92]吴海珍.海洋鱼类病原菌鳗弧菌毒力质粒pEIB1的全序列测定及复制区的定位.华东理工大学博士学位论文.2005.
[93]Kern I, Ceglowski P. Secretion of streptokinase fusion proteins from Escherichia coli cells through the hemolysin transporter. Gene.1995,163:53-57
[94]Gentschev I, Sokolovic Z, Mollenkopf HJ, Hess J, Kaufmann SH, Kuhn M, Krohne GF, Goebel W. Salmonella strain secreting active listeriolysin changes its intracellular location. Infect Immun.1995,63: 4202-4205
[95]Li Y, Chen CX, von Specht BU, Hahn HP. Cloning and hemolysin-mediated secretory expression of a codon-optimized synthetic human interleukin-6 gene in Escherichia coli. Prot Expr Purif.2002,25: 437-447
[96]Hess J, Gentschev I, Szalay G, Ladel C, Bubert A, Goebel W, Kaufmann SH. Listeria monocytogenes p60 supports host cell invasion by and in vivo survival of attenuated Salmonella typhimurium. Infect Immun.1995,63:2047-2053
[97]Chang YF, Lauderdale TL, Lee WY, Shin SJ, Jacobson RH, Appel MJ, Lein, DH. Expression and secretion of outer surface protein (OSP-A) of Borrelia burgdorferi from Escherichia coli. FEMS Microbiol Lett.1993,109:297-302
[98]Mollenkopf HJ, Gentschev I, Goebel W. Conversion of bacterial gene products to secretion-competent fusion proteins. Biotechniques.1996,21:854-860
[99]Gentschev I, Glaser I, Goebel W, McKeever DJ, Musoke A, Heussler VT. Delivery of the p67 sporozoite antigen of Theileria parva using recombinant Salmonella:secretion of the product enhances specific antibody responses in cattle. Infect Immun.1998,66:2060-2064
[100]Su G, Brahmbhatt N, de Lorenzo V, Wehland J, Timmis K. Extracellular export of Shiga toxin B-subunit/haemolysin A (C-terminus) fusion protein expressed in Salmonella typhimurium aroA-mutant and stimulation of B-subunit specific antibody responses in mice. Microb Pathog.1992,13:465-476
[101]Orr N, Galen JE, Levine MM. Expression and immunogenicity of a mutant diphtheria toxin molecule, CRM (197), and its fragments in Salmonella typhi vaccine strain CVD 908-htrA. Infect Immun.1999, 67:4290-4294
[102]Li Y, Reichenstein K, Ullrich R, Danner T, von Specht BU, Hahn HP. Effect of in situ expression of human interleukin-6 on antibody responses against Salmonella typhimurium antigens. FEMS Immunol Med Microbiol.2003,37:135-145
[103]Xi CW, Lambrecht M, Vanderleyden J, Michiels J. Bi-functional gfp-and gusA-containing mini-Tn5 transposon derivatives for combined gene expression and bacterial localization studies. J Microb Methods.1999,35:85-92
[104]王蓬勃.溶藻弧菌铁摄取和调控机制的初步研究.华东理工大学硕士学位论文.2005.
[105]Sambrook J, Russell DW.黄培堂译.分子克隆实验指南,第三版.科学出版社,2002
[106]周丽,刘洪明,战文斌,宋微波.鳗弧菌、溶藻胶弧菌外膜蛋白的分离及特性.中国水产科学.2003,10：31-35
[107]Kawai K, Liu Y, Ohnishi K, Oshima SI. A conserved 37 kD outer membrane protein of Edwardsiella tarda is an effective vaccine candidate. Vaccine.2004,22:3411-3418
[108]Shi HD, Su WW. Display of fluorescent protein on Escherichia coli cell surface. Enzyme Microb Technol.2000,28:25-34
[109]Georgiou G, Stephens DL, Stathopoulos C. Display of beta-lactamase on the Escherichia coli surface outer membrane phenotypes conferred by Lpp-OmpA-beta-lactamase fusions. Protein Eng.1996,9: 239-247
[110]Whitfield C, Paiment A. Biosynthesis and assembly of Group 1 capsular polysaccharides in Escherichia coli and related extracellular polysaccharides in other bacteria. Carbohyd Res.2003,338: 2491-2502
[111]Kang SM, Rhee JK, Kim EJ, Han KH, Oh JW. Bacterial cell surface display for epitope mapping of hepatitis C virus core antigen. FEMS Microbiol Lett.2003,226:347-353
[112]Kim EJ, Yoo SK. Cell surface display of hepatitis B virus surface antigen by using Pseudomonas syringae ice nucleation protein. Lett Appl Microbiol.1999,29:292-297
[113]Tan YP, Zheng J, Tung SL, Rosenshine I, Leung KY. Role of type III secretion in Edwardsiella tarda virulence. Microbiology.2005,151:2301-2313
[114]Zheng J, Tung SL, Leung KY. Regulation of a type III and a putative secretion system in Edwardsiella tarda by EsrC is under the control of a two-component system, EsrA-EsrB. Infect Immun. 2005,73:4127-4137
[115]Rao PS, Yamada Y, Tan YP, Leung KY. Use of proteomics to identify novel virulence determinants that are required for Edwardsiella tarda pathogenesis. Mol Microbiol.2004,53:573-586
[116]Davey HM, Kell DB. Flow cytometry and cell sorting of heterogeneous microbial populations:the importance of single-cell analyses. Microbiol Rev.1996,60:641-696
[117]Grogan WM, Collins JM. Guide to Flow Cytometry Methods. New York:Marcel Dekker,1990, p29-190
[118]Porter J, Deere D, Hardman M, Edwards C, Pickup R. Go with the flow-use of flow cytometry in environmental microbiology. FEMS Microbiol Ecol.1997,24:93-101
[119]McSharry JJ. Uses of flow cytometry in virology. Clin Microbiol Rev.1994,7:576-604
[120]Wallner G, Amann R, Beisker W. Optimizing fluorescent in situ hybridization with rRNA-targeted oligonucleotide probes for flow cytometric identification of microorganisms. Cytometry.1993,14: 136-143
[121]Impert MBM, Nell R, Anne SP, Burel MC, Cantarovich D, Milpied N, Billaudel S. Development of a method direct quantification of cytomegalovirus antigenemia by flow cytometry. Clin Microbiol Infect. 1997,10:2665-2669
[122]Yurkow EJ, McKenzie MA. Characterization of hypoxiadependent peroxide production in cultures of Saccharomyces cerevisiae using flow cytometry:a model for ischemic tissue destruction. Cytometry. 1993,14:287-293
[123]Allman R, Hann AC, Manchee R, Lloyd D. Characterization of bacteria by multi parameter flow cytometry. J Appl Bacteriol.1992,73:438-444
[124]Amann R, Ludwig L, Scheleifer KH. Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol Rev.1995,59:143-169
[125]Islam D, Lindberg AA, Christensson B. Peripheral blood cell prepare influences the level of expression of leukocyte cell surface marshal assessed with quantitative multicolor flow cytometry. Cytometry.1996,22:128-134
[126]Pinder AC, Purdy PW, Poulter SA, Clark DC. Validation of flow cytometry for rapid enumeration of bacterial concentrations in pure cultures. J Appl Bacteriol.1990,69:92-100
[127]Kaprelyants A, Kell DB. Rapid assessment of bacterial viability and vitality using rhodamine 123 and flow cytometry. J Appl Bacteriol.1992,72:410-422
[128]Lebaron P, Parthuisot N, Catala P. Comparison of blue nucleic acid dyes for flow cytometric enumeration of bacteria in aquatic systems. Appl Environ Microbiol.1998,64:1725-1730
[129]Lopez-Amoros R, Castel S, Comas-Riu J, Vives-Rego J. Assessment of E. coli and Salmonella viability and starvation by confocal laser microscopy and flow cytometry using rhodamine 123, DiBAC4(3), propidium iodide, and CTC. Cytometry.1997,29:298-305
[130]Tidona CA, Schnitzler P, Kehm R, Darai G. Is the major capsid protein of iridoviruses a suitable target for the study of viral evolution? Virus Genes.1998,16:59-66
[131]许煜洲.基于冰核蛋白的鳗弧菌表面展示系统的构建.华东理工大学硕士学位论文.2008.
[132]杨路,马悦,张元兴.鳗弧菌MVAV6203基因缺失株的生物反应器培养及检测.水产科学.2007,26：191-194
[133]邵明非.鳗弧菌基于大肠杆菌α-溶血素分泌系统分泌表达外源蛋白的研究.华东理工大学硕士学位论文.2005.
[134]Nakayama K, Kelly SM, Curtiss III R. Construction of an ASD+expression-cloning vector:stable maintenance and high level expression of cloned genes in a Salmonella vaccine strain. Bio/Technol. 1988:6,693-697
[135]Viola RE. The central enzymes of the aspartate family of amino acid biosynthesis. Acc Chem Res. 2001,34:339-349
[136]Karsten WE, Viola RE. Chemical and kinetic mechanisms of aspartate-beta-semialdehyde dehydrogenase from Escherichia coli. Biochim Biophys Acta.1991,1077:209-219
[137]Blanco J, Moore RA, Kabaleeswaran V, Viola RE. A structural basis for the mechanism of aspartate-beta-semialdehyde dehydrogenase from Vibrio cholerae. Protein Sci.2003,12:27-33
[138]Faehnle CR, Ohren JF, Viola RE. A new branch in the family:structure of aspartate-beta-semialdehyde dehydrogenase from Methanococcus jannaschii. J Mol Biol.2005,353: 1055-1068
[139]Paidhungat M, Setlow B, Driks A, Setlow P. Characterization of spores of Bacillus subtilis which lack dipicolinic acid. J Bacteriol.2000,182:5505-5512
[140]Pavelka Jr MS, Jacobs Jr WR. Biosynthesis of diaminopimelate, the precursor of lysine and a component of peptidoglycan, is an essential function of Mycobacterium smegmatis. J Bacteriol.1996, 178:6496-6507
[141]Viola RE, Liu X, Ohren JF, Faehnle CR. The structure of a redundant enzyme:a second isoform of aspartate β-semialdehyde dehydrogenase in Vibrio cholerae. Acta Crystallogr D Biol Crystallogr.2008, 64:321-330
[142]Harb OS, Kwaik YA. Identification of the aspartate-beta-semialdehyde dehydrogenase gene of Legionella pneumophila and characterization of a null mutant. Infect Immun.1998,66:1898-1903
[143]Angeles TS, Hunsley JR, Viola RE. Reversal of enzyme regiospecificity with alternative substrates for aspartokinase I from Escherichia coli. Biochemistry.1992,31:799-805
[144]Kang HY, Srinivasan J, Curtiss III R. Immune responses to recombinant Pneumococcal PspA antigen delivered by live attenuated Salmonella enterica serovar Typhimurium vaccine. Infect Immun.2002,4: 1739-1749
[145]Tijhaar EJ, Zheng-Xin Y, Karlas JA, Meyer TF, Stukart MJ, Osterhaus ADME, Mooi FR. Construction and evaluation of an expression vector allowing the satable expression of foreign antigens in a Salmonella typhimurium vaccine strain. Vaccine.1994,12:1004-1111
[146]Chatfield S, Charles IQ Makoff AJ, Oxer MD, Dougan G, Pickard D, Slater D, Fairweather NF. Use of the nirB promoter to direct the stable expression of heterologous antigens in Salmonella oral vaccine strains:Development of a single-dose oral tetanus vaccine. Bio/Technol.1992,10:888-892
[147]Hohmann EL, Oletta CA, Miller SI. Evaluation of a phoP/phoQ-deleted, aroA-deleted live oral Salmonella typhi vaccine strain in human volunteers. Vaccine.1996,14:19-24
[148]Roberts M, Li J, Bacon A, Chatfield S. Oral vaccination against tetanus:Comparison of the immunogenicities of Salmonella strains expressing fragment C form the nirB and htrA promoters. Infect Immun.1998,66:3080-3087
[149]Orr N, Galen JE, Levine MM. Novel use of anaerobically induced promoter, dmsA, for controlled expressing of fragment C of tetanus toxin in live attenuated Salmonella enterica serovar Typhi strain CVC 908-htrA. Vaccine.2001,19:1694-1700
[150]Kong W, Wanda SY, Curtiss III R. Construction and application of hot-vector systems for DNA vaccine vector delivery [A-016]. In American Society for Microbiology,103rd General Meeting, Washington, DG:American Society for Microbiology,677
[151]Andrianopoulos K, Wang L, Reeves PR. Identification of the fucose synthetase gene in the colonic acid gene cluster of Escherichia coli K-12. J Bacteriol.1998,180:998-1001
[152]Moore RA, Bocik WE, Viola RE. Expression and purification of L-aspartate-beta-semi aldehyde dehydrogenase from infectious microorganisms. Protein Expr Purif.2002,25:189-194
[153]Gouet P, Courcelle E, Stuart DI, Metoz F. ESPript:analysis of multiple sequence alignments in PostScript. Bioinformatics.1999,15:305-308
[154]Liu Y, Oshima S, Kawai K. Glyceraldehyde-3-phosphate dehydrogenase of Edwardsiella tarda has protective antigenicity against Vibrio anguillarum in Japanese flounder. Dis Aquat Org.2007,75: 217-220