摘要
细菌性疾病是水产养殖业的重要疾病。迟缓爱德华氏菌（Edwardsiella tarda）是一种革兰氏阴性菌，能够感染多种经济鱼类并引起爱德华氏菌病，给水产养殖业造成巨大的经济损失。本论文主要是以牙鲆为实验动物，开展高效迟缓爱德华氏菌疫苗的设计与构建研究，并且探讨相关疫苗的免疫机制。具体如下：
首先通过理论推断与实际实验相结合，本研究筛选出一系列迟缓爱德华氏菌免疫保护原，包括Esa1、Eta2、Eta1等，并在此基础上构建了基于细菌传递的重组亚单位疫苗、DNA疫苗、减毒疫苗和交叉保护疫苗。
（1）基于Esa1构建重组亚单位疫苗、细菌传递的亚单位疫苗和DNA疫苗：Esa1是一个含有795个氨基酸残基的迟缓爱德华氏菌表面抗原，含有多个细菌表面抗原的保守位点且定位于迟缓爱德华氏菌的外膜。我们将Esa1在大肠杆菌中表达，获得了重组亚单位疫苗，发现其对迟缓爱德华氏菌引起的牙鲆感染有82%的免疫保护效应。随后我们利用肠道共生菌FP3构建了表面展示Esa1的细菌传递疫苗FP3/pJsa1。发现FP3/pJsa1作为口服微粒疫苗和注射疫苗分别能达到52%和79%的相对免疫保护率。我们进一步以Esa1为抗原，构建成了DNA疫苗pCEsa1。pCEsa1免疫牙鲆1个月和2个月后，其免疫保护率分别达到75%和71%。这些结果表明Esa1作为重组亚单位疫苗、鱼类肠道共生菌传递的表面展示型亚单位疫苗以及DNA疫苗皆具有良好的保护效应。
（2）基于Eta2构建重组亚单位疫苗和DNA疫苗：在鱼类疫苗研究中，疫苗的免疫机制以及不同类型疫苗的免疫比较研究十分缺乏。本研究筛选得到一个新的迟缓爱德华氏菌免疫保护原Eta2，研究了Eta2作为重组亚单位疫苗和DNA疫苗所诱发的免疫保护效应及免疫机制。我们发现Eta2作为重组亚单位疫苗在免疫牙鲆4周和8周后，可分别产生83%和78%的相对保护率；作为DNA疫苗在免疫牙鲆4周和8周后产生的相对保护率分别为67%和68%。免疫学研究表明此两类疫苗均可增强牙鲆头肾巨噬细胞的杀菌活性、引起特异性和非特异性免疫相关基因上调，以及刺激牙鲆产生特异性抗体，其中亚单位疫苗诱发产生的抗体水平较高。进一步研究表明，Eta2作为重组亚单位疫苗主要引起牙鲆的体液免疫，而作为DNA疫苗则同时引起了鱼体B细胞和T细胞的免疫应答。这些结果说明Eta2是良好的免疫保护原，可以作为重组亚单位疫苗和DNA疫苗激发宿主的免疫保护反应，但是不同形式疫苗的免疫机制存在差异。
（3）毒力因子Eta1及其免疫保护作用：在本研究中，我们发现了一个与细菌黏附素有同源性的体内诱导抗原Eta1。eta1基因在迟缓爱德华氏菌未侵染宿主时，其表达依赖于细菌的生长周期，而当迟缓爱德华氏菌侵染宿主时，eta1的mRNA水平显著上升。将eta1缺失突变后，缺失菌株在细胞和组织水平的侵染能力显著下降，说明Eta1是迟缓爱德华氏菌侵染所需的一个毒力因子。我们构建了Etal的重组亚单位疫苗rEta1，发现其能够诱导牙鲆产生高达83.3%的免疫保护率。rEtal免疫牙鲆后能够诱导鱼类产生特异性抗体，这些抗体在迟缓爱德华氏菌感染后能与表达于细菌表面的Eta1相结合。
（4）减毒疫苗的构建以及疫苗导入途径研究：本研究筛选得到一株迟缓爱德华氏菌的弱毒菌株TX5RM，利用该弱毒菌株作为减毒疫苗，探索了注射、口服、浸泡及口服浸泡叠加等不同导入方式对疫苗免疫保护效应的影响。结果表明，这四种免疫方式均产生了明显的免疫保护效应，其中以口服与浸泡叠加的方式效果最为显著，在免疫牙鲆5周和8周后的相对保护率分别达到了80.6%和69.4%。
（5）口服/浸泡DNA疫苗及其交叉免疫保护效应：我们将上述TX5RM作为DNA疫苗的导入载体，构建了携带海豚链球菌DNA疫苗pCS10的迟缓爱德华氏菌减毒疫苗TX5RMS10。免疫牙鲆后，TX5RMS10携带的DNA疫苗质粒能够在鱼体中稳定存在并表达海豚链球菌抗原Sia10。TX5RMS10经口服和浸泡方式免疫牙鲆1个月和2个月后，用迟缓爱德华氏菌和海豚链球菌分别攻毒，发现免疫组具有高达69~83%的存活率。这些结果说明TX5RMS10是一个高效的候选疫苗，其具有TX5RM和pCS10的免疫原性，因此可以诱导针对迟缓爱德华氏菌和海豚链球菌的交叉免疫保护效应。另外，该疫苗能够通过模拟自然感染的方式导入鱼体，从而避免了常规DNA疫苗的注射导入，因此在应用上具有非常好的优势。
综上所述，本论文研究建立了新型鱼类疫苗的构建技术和导入方法，获得了多种不同类型的高效疫苗，初步阐明了不同类型疫苗诱发的免疫反应机制。这些研究结果促进了迟缓爱德华氏菌疫苗研究的发展，为鱼类迟缓爱德华氏菌病的防控提供了新思路。
Bacterial pathogens are one of the major causes of aquaculture diseases.Edwardsiella tarda is a Gram-negative bacterial pathogen and the etiological agent ofa systematic fsh disease called edwardsiellosis, which affects a wide range of marineand freshwater fsh. The disease caused is often fatal and can lead to heavy economiclosses.
Based on this situation, we in this study have designed and constructed vaccinesagainst E. tarda and examined the protective effects of the vaccines in a Japaneseflounder (Paralichthys olivaceus) model.
We have so far constructed several different types of vaccines, which includeinactivated whole cell vaccines, attenuated live vaccines, DNA vaccines, cross-genusvaccines and recombinant subunit vaccines delivered by a live bacterial host. Thesevaccines proved to produce high levels (relative percentage survival greater than70%)of protection against E. tarda. We have identfied several E. tarda antigens, Esa1,Eta2and Eta1.
Esa1is a D15-like surface antigen, which, when used as a recombinant subunitvaccine and DNA vaccine, are able to induce protective immunity in Japanesefounder (Paralichthys olivaceus) against E. tarda challenge. Taking advantage of thesecretion capacity of Esa1and the natural gut-colonization ability of a fsh commensalstrain, we constructed an Esa1-expressing recombinant strain, FP3/pJsa1. Vaccinationanalyses showed that FP3/pJsa1as an intraperitoneal injection vaccine and an oralvaccine embedded in alginate microspheres produced relative percent survival rates of 79%and52%. We examined further the immunoprotective potential of Esa1as aDNA vaccine. The DNA vaccine plasmid pCEsa1was constructed. Hence, comparedto PBS-vaccinated fsh, pCEsa1-vaccinated fsh were signifcantly protected, with aRPS of75%and71%respectively, at one-and two-month post-vaccination (p.v.).Together these results indicate that Esa1is a protective immunogen and an effectiveoral vaccine when delivered by FP3/pJsa1as a surface-anchored antigen.
Eta2, a new E. tarda vaccine candidate, we have analyzed in a comparativemanner the immune response induced by Eta2in two different formfes:d purirecombinant subunit vaccine and DNA vaccine. Recombinant Eta2(rEta2) purfiedfrom Escherichia coli was highly protective against E. tarda challenge in a Japanesefounder model and produced relative percent of survival rates of83%and78%,respectively, at4-and8-week post-vaccination (p.v.). Eta2as a DNA vaccine in theform of plasmid pCEta2also induced strong protective immunity at4-and8-week p.v.Immunological analysis indicated that (i) rEta2and pCEta2enhanced head kidneymacrophageactivation;(ii) rEta2and pCEta2induced similar patterns of serumantibody production;(iii) both rEta2and pCEta2upregulated the expression ofspecifc and nonspecifc immune factors which include. Taken together, these resultsindicate that both rEta2and pCEta2induce sfpce ciand nonspecifc immunities,however, pCEta2induces both B cell and T cell responses, whereas rEta2inducesmainly humoral response.
Eta1, we identifed, via in vivo-induced antigen technology. eta1expression wasgrowth phase dependent and reached maximum at mid-logarithmic phase. Duringinfection offounder lymphocytes, eta1expression was drastically increased at theearly stage of infection. Compared to the wild type, the eta1-defective mutant, TXeta1,was unaffected in growth but exhibited attenuated overall virulence, reduced tissuedissemination and colonization capacity, and impaired ability to infvoaduenderlymphocytes and to block the immune response of host cells. Furthermore, when usedas a subunit vaccine, rEta1produced relative percent of survival rates of83.3%infounder against lethal E. tarda challenge. Taken together, these results indicate thatEta1is an in vivo-induced antigen that mediates pathogen-host nteraction and, as a result, is required for optimal bacterial infection.
In this study, we also have selected TX5RM, which is an attenuated E. tardastrain with good vaccine potential and that a combination of oral and immersionvaccinations may be a good choice for the administration of live attenuated vaccines.The results showed that TX5RM administered via all four approaches producedsignifcant protection, with the highest protection levels observed with TX5RMadministered via oral feeding plus immersion, which were, in terms of relative percentof survival (RPS),80.6%and69.4%at5-and8-week post-vaccination, respectively.
We have constructed a cross-genus vaccine by takeing advantage of theresidual infectivity of TX5RM and using it as a carrier host for the natural delivery ofa S. iniae DNA vaccine. The recombinant TX5RM, TX5RMS10, was created, whichharbours and retains stably the DNA vaccine plasmid pCS10that expresses Sia10.Following E. tarda and S. iniae challenge at one and2months p.v., the vaccinatedfsh exhibited relative percent survival rates of69–83%. Immunological analysisindicated that TX5RMS10-vaccinated fsh produced specifc serum antibodies andexhibited enhanced expression of a wide range of immune genes.
In addition, we also examined the immune response of the vaccines and foundthat DNA vaccines stimulate stronger systematic immunity than subunit vaccines.These findings promote the research on the marine fish vaccines and provide insightsto the protection mechanism of vaccines.
引文
李红艳.我国水产动物疫病与防控研究.牧业论坛.2012,8:50–51.
王波,莫照兰.迟缓爱德华氏菌及其致病机理.海洋科学集刊.2007,5:133–139.
王武.我国水产养殖业的现状与发展趋势.渔业致富指南.2009,7:12–17.
吴淑琴,王亚军.我国水产养殖病害控制技术现状与发展趋势.中国水产.2010,8:9–10.
曾令兵.我国水产养殖动物病害的现状及发展方向.科学养鱼.2010,3:1–2.
Agnew W, Barnes AC. Streptococcus iniae: an aquatic pathogen of global veterinary significanceand a challenging candidate for reliable vaccination. Vet. Microbiol.2007,122:1–15.
Ainsworth JA, Dexiang C. Differences in the phagocytosis of four bacteria by channel catfishneutrophils. Dev. Comp. Immunol.1990,14:201–209.
Alcaide E, Herraiz S, Esteve C. Occurrence of Edwardsiella tarda in wild European eels Anguillaanguilla from Mediterranean Spain. Dis. Aquat. Organ.2006,73:77–81.
Alice AF, Naka H, Crosa JH. Global gene expression as a function of the iron status of thebacterial cell: influence of differentially expressed genes in the virulence of the humanpathogen Vibrio vulnificus. Infect. Immun.2008,76:4019–4037.
Alvarez B, Alvarez J, Menendez A, Guijarro JA. A mutant in one of two exbD loci of a TonBsystem in Flavobacterium psychrophilum shows attenuated virulence and confers protectionagainst cold water disease. Microbiology.2008,154:1144–1151.
Amandi A, Hiu SF, Rohovec JS, Fryer JL. Isolation and characterization of Edwardsiella tardafrom fall chinook salmon （Oncorhynchus tshawytscha）. Appl. Environ. Microbiol.1982,43:1380–1384.
Amend DF. Potency testing fosfh vaccines. International Symposium on Fish Biologics:Serodiagnostics and Vaccines. Dev. Biol. Standard.1981,49:447–454.
Angelidis P, Karagiannis D, Crump EM. Efficacy of a Listonella anguillarum （syn. Vibrioanguillarum） vaccine for juvenile sea bass Dicentrarchus labrax. Dis. Aquat. Organ.2006,11;71:19–24.
Arambulo PV, Westerlund NC, Sarmiento RV: On the isolation of human enteric organisms fromthe bile of pigs and cattle. Acta Medica Philipp1968,5:84–86.
Arias CR, Shoemaker CA, Evans JJ, Klesius PH. A comparative study of Edwardsiella ictaluriparent （EILO） and E. ictaluri rifampicin-mutant （RE-33） isolates usinglipopolysaccharides, outer membrane proteins, fatty acids, Biolog, API20E and genomicanalyses. J. Fish. Dis.2003,26:415–421.
Austin B, Austin D.A. Bacterial fish pathogens: disease of farmed and wild fish. New York:Springer Verlag.2007.
Bader JA, Shoemaker CA, Klesius PH. Production, characterization and evaluation of virulence ofan adhesion defective mutant of Flavobacterium columnare produced by beta-lactamselection. Lett. Appl. Microbiol.2005,40:123–127.
Bhatnagar N, Getachew E, Straley S, Williams J, Meltzer M, Fortier A. Reduced virulence ofrifampicin-resistant mutants of Francisella tularensis. J. Infect. Dis.1994,170:841–847.
Billington OJ,McHughTD, Gillespie SH. Physiological cost of rifampin resistance induced in vitroin Mycobacterium tuberculosis. Antimicrob Agents Chemother.1999,43:1866–1869.
Blanch AR, Pinto RM, Jofre JT. Isolation and characterization of an Edwardsiella sp. strain,causative agent of mortalities in sea bass （Dicentrarchus labrax）. Aquaculture.1990,88:213–222.
Boudinot P, Blanco M, de Kinkelin P, Benmansour A. Combined DNA immunization with theglycoprotein gene of viral hemorrhagic septicemia virus and infectious hematopoieticnecrosis virus induces double-specific protective immunity and nonspecific response inrainbow trout. Virology.1998,249:297–306.
Bowden TJ, Menoyo-Luque D, Bricknell IR, Wergeland H. Efficacy of different administrationroutes for vaccination against Vibrio anguillarum in Atlantic halibut （Hippoglossushippoglossus L.）. Fish Shellfish Immunol.2002,12:283–285.
Bradford M. A rapid and sensitive method for the quantitation of microgram quantities of proteinutilizing the principle of protein-dye binding. Anal Biochem.1976,72:248–254.
Buchanan JT, Stannard JA, Lauth X, Ostland VE, Powell HC,Westerman ME, et al. Streptococcusiniae phosphoglucomutase is a virulence factor and a target for vaccine development. Infect.Immun.2005,73:6935–6944.
Byon JY, Ohira T, Hirono I, Aoki T. Use of a cDNA microarray to study immunity against viralhemorrhagic septicemia （VHS） in Japanese flounder （Paralichthys olivaceus） followingDNA vaccination. Fish Shellfish Immunol.2005,18:135–147.
Caipang CM, Hynes N, Puangkaew J, Brinchmann MF, Kiron V. Intraperitoneal vaccination ofAtlantic cod, Gadus morhua with heat-killed Listonella anguillarum enhances serumantibacterial activity and expression of immune response genes. Fish Shellfish Immunol.2008,24:314–322.
Carrington AC, Secombes CJ. A review of CpGs and their relevance to aquaculture. Vet.Immunol. Immunopathol.2006,112:87–101.
Castro N, Toranzo AE, Barja JL, Nunez S, Magarinos B. Characterization of Edwardsiella tardastrains isolated from turbot, Psetta maxima （L）. J. Fish. Dis.2006,29:541–547.
Castro N, Toranzo AE, Nunez S, Magarinos B. Development of an effective Edwardsiella tardavaccine for cultured turbot （Scophthalmus maximus）. Fish Shefllsh Immunol.2008.25:208–212.
Chang CI, Wu CC, Cheng TC, Tsai JM, Lin KJ. Multiplex nested-polymerase chain reaction forthe simultaneous detection of Aeromonas hydrophila, Edwardsiella tarda, Photobacteriumdamselae and Streptococcus iniae, four important fish pathogens in subtropical Asia.Aquacult. Res.2009,40:1182–1190.
Cheng S, Hu Y, Jiao X, Sun L. Identification and immunoprotective analysis of a Streptococcusiniae subunit vaccine candidate. Vaccine.2010,28:2636–2641.
Cheng S, Hu Y, Zhang M, Sun L. Analysis of the vaccine potential of a natural avirulentEdwardsiella tarda isolate. Vaccine.2010,28:2716–2721.
Cheng S, Zhang M, Sun L. The iron-cofactored superoxide dismutase of Edwardsiella tardainhibits macrophage-mediated innate immune response. Fish Shellfish Immunol.2010,28-29:972–978.
Chen JD, Huang SL1. Hemolysin from Edwardsiella tarda strain ET16isolated from eel Anguillajaponica identified as a hole-forming toxin. Fish Science.1996,62:638–542.
Chen JD, Lai SY, Huang SL. Molecular cloning, characterization, and sequencing of thehemolysin gene from Edwardsiella tarda. Arch. Microbiol.1996,165:9–17.
Chen K, Cerutti A. New insights into the enigma of immunoglobulin D. Immunol. Rev.2010,237:160–179.
Choi SH, Kim KH. Generation of two auxotrophic genes knock-out Edwardsiella tarda andassessment of its potential as a combined vaccine in olive flounder（Paralichthys olivaceus）.Fish Shellfish Immunol.2011,31:58–65.
Chung S, Secombes CJ. Analysis of events occurring with teleost macrophages during therespiratory burst. Comp. Biochem. Physiol.1988,89B:539–544.
Chung S, Secombes CJ. Activation of rainbow trout macrophages. J. Fish. Biol.1987,31:51–56.
Coban C, Koyama S, Takeshita F, Akira S, Ishii KJ. Molecular and cellular mechanisms of DNAvaccines. Hum. Vaccine.2008,4:453–456.
Corripio-Miyar Y, Mazorra de Quero C, Treasurer JW, Ford L, Smith PD, Secombes CJ.Vaccination experiments in the gadoid haddock, Melanogrammus aeglefinus L., against thebacterial pathogen Vibrio anguillarum. Vet. Immunol. Immunopathol.2007,15;118:147–153.
Crosbie PB, Nowak BF. Immune responses of barramundi, Lates calcarifer （Bloch）, afteradministration of an experimental Vibrio harveyi bacterin by intraperitoneal injection, analintubation and immersion. J. Fish. Dis.2004,27:623–632.
Cuesta A, Tafalla C. Transcription of immune genes upon challenge with viral hemorrhagicsepticemia virus （VHSV） in DNA vaccinated rainbow trout （Oncorhynchus mykiss）.Vaccine.2009,27:280–289.
Daly JG, Griffiths SG, Kew AK, Moore AR, Olivier G. Characterization of attenuatedRenibacterium salmoninarum strains and their use as live vaccines. Dis. Aquat. Org.2001,44:121–126.
Damme LRV, Vandepitte J. Frequent isolation of Edwardsiella tarda and Pleisiomonasshigelloides from healthy zairese freshwater fish: a possible source of sporadic diarrhea in thetropics. Appl. Environ. Microbiol.1980,39:475–479.
Damme LRV, Vandepitte J. Isolation of Edwardsiella tarda and plesiomonas shigelloides frommammals and birds in Zaire. Rev. Elev. Med. Vet. Pays. Trop.1984,37:145–151.
Dang W, Hu YH, Sun L. HtpG is involved in the pathogenesis of Edwardsiella tarda. Vet.Microbiol.2011,152:394–400.
Dang W, Zhang M, Sun L. Edwardsiella tarda DnaJ is a virulence-associated molecular chaperonewith immunoprotective potential. Fish Shellfish Immunol.2011,31:182–188.
Dumetz F, Duchaud E, Claverol S, Orieux N, Papillon S, Lapaillerie D, et al. Analysis of theFlavobacterium psychrophilum outer-membrane subproteome and identification of newantigenic targets for vaccine by immunomics. Microbiology.2008,154:1793–1801.
Edited by Evans JJ, Klesius PH, Plumb JA, Shoemaker CA. Wallingford. CABI International.2010,512–534.
Egusa S. Some bacterial diseases of freshwater fishes in Japan. Fish Pathol.1976,10:103–114.
Eldar A, Horovitcz A, Bercovier H. Development and efficacy of a vaccine against Streptococcusiniae infection in farmed rainbow trout. Vet. Immunol. Immunopathol.1997,56:175–183.
Entriken TL. Veterinary pharmaceuticals and biologicals.12th edition Montvale, NJ, USA:Veterinary Health Care Communications—Thomson Health Care.2002.
Evelyn TPT. A historical review of fish vaccinology. In: Fish Vaccinology, Developments inBiological Standardization. Gudding R, Lillehaug A, Midtlyng PJ, Brown F （Eds.）. Karger,Basel, Switzerland.1997,90:3–12.
Ewing WH, McWhorter AC, Escobar MR, Lubin AH. Edwardsiella, a new genus ofEnterobacteriaceae based on a new species, E. Tarda. Int. J. Syst. Evol. Microbiol.1965,15:33–38.
Fan WH, Huang J, Wang XH, Shi CY, Liu L. Identification and phylogenetic study of pathogenicbacteria causing ulcer disease of cultured Turbot（Scophthalmus maximus）. Acta. Microbiol.Sin.2005,45:665–670.
Flack FS, Loosmore S, Chong P, Thomas WR. The sequencing of the80-kDa D15protectivesurface antigen of Haemophilus infuenzae. Gene.1995.156:97–99.
Fletcher TC White A. Antibody production in the Plaice（Pleuronectes platessa L.）after oral andparenteral immunization with Vibrio anguillarum antigens. Aquaculture.1973,1:417–428.
Goldberg JB, Ganesan S, Comstock AT, Zhao Y, Sajjan US. Cable pili and the associated22kDaadhesin contribute to Burkholderia cenocepacia persistence in vivo. PLoS. One.2011,6:e22435.
Graham S, Jeffries AH, Secombes CJ. A novel assay to detect macrophage bactericidal activity infish: factors influencing the killing of Aeromonas salmonicida. J. Fish Dis.1988,11:389–396.
Grimont PAD, Grimont F, Richard C, Sakazaki R. Edwardsiella hoshinae, a new species ofenterobacteriaceae. Curr1. Microbiol.1980,14:345–351.
Gutierrez MA, Miyazaki T. Responses of Japanese eels to oral challenge with Edwardsiella tardaafter vaccination with formalin-killed cells or lipopolysaccharide of the bacterium. J. Aquat.Anim. Health.1994,6:110–117.
H stein T, Gudding R, Evensen. Bacterial vaccines for fish–an update of the current situationworldwide In: Fish Vaccinology, Developments in Biologicals. Midtlyng PJ （Ed.）, Karger,Basel, Switzerland.2005,121:55–74.
Hart S, Wrathmell AB, Harris JE, Grayson TH. Gut immunology in fish: a review. Dev. Comp.Immunol.1988,12:453–480.
Hawke JP, Mcwhorter A, Steigerwalt AG, Brenner DONJ. Edwardsiella ictaluri sp. nov., thecausative agent of enteric septicemia of catfish. Int. J. Syst. Evol. Microbiol.1981,31:396–400.
Heppell J, Davis HL. Application of DNA vaccine technology to aquaculture. Adv. Drug. Deliv.Rev.2000,43:29–43.
Hirono I, Tange N, Aoki T. Iron-regulated haemolysin gene from Edwardsiella tarda. Mol.Microbiol.1997,24:851–856.
Hu K, Chi Z, Li J, Zhang F, Li M, Yasoda HN, et al. The surface display of haemolysin fromVibrio harveyi on yeast cells and their potential applications as live vaccine in marinfes h.Vaccine.2006,24:6046–6052.
Hu Y, Dang W, Liu C, Hou J, Sun L. Analysis of the effect of copper on the virulence of apathogenic Edwardsiella tarda strain. Lett. Appl. Microbiol.2010,50:97–103.
Hu Y, Liu C, Hou J, Sun L. Idfecnattiion, characterizati on, and molecular application of avirulence-associated autotransporter from a pathogenic Pseudomonas fuorescens strain. Appl.Environ. Microbiol.2009,75:4333–4340.
Hu Y, Wang H, Zhang M, Sun L. Molecular analysis of the copperresponsive CopRSCD of apathogenic Pseudomonas fuorescens strain. J. Microbiol.2009,47:277–286.
Hoshina T. On a new bacterium, paracolobactrum anguillimortiferum n. sp. Bull. Jpn. Soc. Sci.Fish.1962,28:162–164.
Horenstein S, Smolowitz R, Uhlinger K, Roberts S. Diagnosis of Edwardsiella tarda infection inoyster toadfish （Opsanus tau） held at the Marine Resources Center. Biol. Bull.2004,207:171.
Hou JH, Zhang WW, Sun L. Immunoprotective analysis of two Edwardsiella tarda antigens. J.Gen. Appl. Microbiol.2009,55:57–61.
Ishibe K, Osatomi K, Hara K, Kanai K, Yamaguchi K, Oda T. Comparison of the responses ofperitoneal macrophages from Japanese flounder （Paralichthys olivaceus） against highvirulent and low virulent strains of Edwardsiella tarda. Fish Shellfish Immunol.2008,24:243–251.
Ishibe K, Yamanishi T, Wang Y, Osatomi K, Hara K, Kanai K, Yamaguchi K, Oda T. Comparativeanalysis of the production of nitric oxide （NO） and tumor necrosis factor-[alpha]（TNF-[alpha]） from macrophages exposed to high virulent and low virulent strains ofEdwardsiella tarda. Fish Shellfish Immunol.2009,27:386–389.
Ishihara S, Kusuda R. Growth and survival of Edwardsiella tarda bacteria in environmental water.Bull. Jpn. Soc. Sci. Fish.1982,48:483–488.
Iveson JB. Strontium chloride B and EE enrichment broth media for the isolation of Edwardsiella,Salmonella and Arizona species from tiger snakes. J. Hyg.（Lond）1971,69:323–330.
Janda JM, Abbott SL. Expression of an iron-regulated hemolysin by Edwardsiella tarda. FEMS.Microbiol. Lett.1993,111:275–280.
Janda JM, Abbott SL. Infections associated with the genus Edwardsiella: the role of Edwardsiellatarda in human disease. Clin. Infect. Dis.1993,17:742–748.
Janda JM, Abbott SL, Kroske-Bystrom S, Cheung, WKW, Powers C, Kokka RP, et al. Pathogenicproperties of Edwardsiella species. J. Clin. Microbiol.1991,29:1997–2001.
Jayasree L, Janakiram P, Madhavi R. Characterization of Vibrio spp. associated with diseasedshrimp from culture ponds of Andhra Pradesh （India）. J. World Aquac. Soc.2006,37:523–532.
Jiao X, Cheng S, Hu Y, Sun L. Comparative study of the effects of aluminum adjuvants andFreund’s incomplete adjuvant on the immune response to an Edwardsiella tarda majorantigen. Vaccine.2010,28:1832–1837.
Jiao X, Dang W, Hu Y, Sun L. Identification and immunoprotective analysis of an in vivo-inducedEdwardsiella tarda antigen. Fish Shellfish Immunol.2009,27:633–638.
Jiao X, Hu Y, Sun L. Dissection and localization of the immunostimulating domain ofEdwardsiella tarda FliC. Vaccine.2010,28:5635–5640.
Jiao X, Zhang M, Cheng S, Sun L. Analysis of Edwardsiella tarda DegP, a serine protease and aprotective immunogen. Fish Shellfish Immunol.2010,28:672–677.
Jiao X, Zhang M, Hu Y, Sun L. Construction and evaluation of DNA vaccines encodingEdwardsiella tarda antigens. Vaccine2009,27:5195–5202.
Kawai K, Liu Y, Ohnishi K, Oshima S. A conserved37kDa outer membranemprotein ofEdwardsiella tarda is an effective vaccine candidate. Vaccinem.2004,22:3411–3418.
Klesius PH, Shoemaker CA. Development and use of modified live Edwardsiella ictaluri vaccineagainst enteric septicemia of catfish. Adv. Vet. Med.1999,41:523–537.
Kumar SR, Parameswaran V, Ahmed VP, Musthaq SS, Hameed AS. Protective fficiency of DNAvaccination in Asian seabass （Lates calcarifer） against Vibrio nguillarum. Fish ShellfishImmunol.2007,23:316–326.
Kusuda R, Toyoshima T, Iwamura Y, Sako H. Edwardsiella tarda from an epizootic of mullets（Mugil cephalus） in Okitsu Bay. Bull. Jpn. Soc. Sci. Fish.1976,42:271–275.
Kusuda R, Itami T, Munekiyo M, Nakajima H. Characteristics of an Edwardsiella sp. from anepizootic of cultured crimson sea breams. Bull. Jpn. Soc. Sci. Fish.1977,43:129–134.
Kwon SR, Lee EH, Nam YK, Kim SK, Kim KH. Efficacy of oral immunizationmwithEdwardsiella tarda ghosts against edwardsiellosis in olive flounderm （Paralichthysolivaceus）. Aquaculture.2007,269:84–88.
Kwon SR, Nam YK, Kim SK, Kim KH. Protection of tilapia （Oreochromis mosambicus） fromedwardsiellosis by vaccination with Edwardsiella tarda ghosts. Fish Shellfish Immunol.2006,20:621–626.
LaFrentz BR, LaPatra SE, Call DR, Cain KD. Isolation of rifampicin resistant Flavobacteriumpsychrophilum strains and their potential as live attenuated vaccine candidates. Vaccine.2008,26:5582–5589.
Lahav D, Eyngor M, Hurvitz A, Ghittino C, Lublin A, Eldar A. Streptococcus iniae type IIinfections in rainbow trout Oncorhynchus mykiss. Dis. Aquat. Org.2004,62:177–180.
Lan MZ, Peng X, Xiang MY, Xia ZY, Bo W, Jie L, Li XY, Jun ZP. Construction andcharacterization of a live, attenuated esrB mutant of Edwardsiella tarda and its potential as avaccine against the haemorrhagic septicaemia in turbot, Scophthamus maximus （L.）. FishShellfish Immunol.2007,23:521–530.
Lau SKP, Woo PCY, Tse H, Leung KW, Wong SSY, Yuen KY. Invasive Streptococcus iniaeinfections outside North America. J. Clin. Microbiol.2003,41:1004–1009.
Lawrence ML, Banes MM, Williams ML. Phenotype and virulence of a transposon-derivedlipopolysaccharide O side-chain mutant strain of Edwardsiella ictaluri. J. Aquat. Anim.Health.2001,13:291–299.
Lawrence ML, Cooper RK, Thune RL. Attenuation, persistence, and vaccine potential of anEdwardsiella ictaluri purA mutant. Infect. Immun.1997,65:4642–4651.
Lee DJ, Kwon SR, Zenke K, Lee EH, Nam YK, Kim SK, et al. Generation of safety enhancedEdwardsiella tarda ghost vaccine. Dis. Aqua. Organ.2008,81:249–254.
Lee J, Kim YB, Kwon M. Outer membrane protein H for protective immunity against Pasteurellamultocida. J. Microbiol.2007,45:179–184.
Leo JC, Skurnik M. Adhesins of human pathogens from the genus Yersinia. Adv. Exp. Med. Biol.2011,715:1–15.
Leotta GA, Pi eyro P, Serena S, Vigo GB: Prevalence of Edwardsiella tarda in Antarctic wildlife.Polar. Biol.2009,32:809–812.
Leung KY, Siame BA, Tenkink BJ, Noort RJ, Mok Y.K. Edwardsiella tarda–Virulencemechanisms of an emerging gastroenteritis pathogen. Microbes Infect.2011.doi:10.1016/j.micinf.2011.08.005.
Limoli DH, Sladek JA, Fuller LA, Singh AK, King SJ. BgaA acts as an adhesin to mediateattachment of some pneumococcal strains to human epithelial cells. Microbiology.2011,157（Pt8）:2369–2381.
Ling SHM, Wang XH, Xie L, Lim TM, Leung KY. Use of green fluorescent protein （GFP） totrack the invasive pathways of Edwardsiella tarda in the in vivo and in vitro fish models.Microbiology.2000,146:7-19.
Liu CS, Sun Y, Hu YH, Sun L. Identification and analysis of the immune effects of CpG motifsthat protect Japanese flounder （Paralichthys olivaceus） against bacterial infection. FishShellfish Immunol.2010,29:279–285.
Liu Y, Bi Z. Potential use of a transposon Tn916-generated mutant of Aeromonas hydrophila J-1defective in some exoproducts as a live attenuated vaccine. Prev. Vet. Med.2007,78:79–84.
Liu Y, Oshima S, Kawai K.Glyceraldehyde-3-phosphate dehydrogenase of Edwardsiella tarda hasprotective antigenicity against Vibrio anguillarum in Japanese flounder. Dis. Aquat. Organ.2007,75:217–220.
Liu Y, Oshima S, Kurohara K, Ohnishi K, Kawai K. Vaccine efficacy of recombinant GAPDH ofEdwardsiella tarda against Edwardsiellosis. Microbiol. Immunol.2005,49:605–612.
Locke JB, Vicknair MR, Ostland VE, Nizet V, Buchanan JT. Evaluation of Streptococcus iniaekilled bacterin and live attenuated vaccines in hybrid striped bass through injection and bathimmersion. Dis. Aquat. Organ.2010,89:117–123.
Loosmore SM, Yang YP, Coleman DC, Shortreed JM, England DM, Klein MH. Outermembrane protein D15is conserved among Haemophfiluuesn zaien species andmay represent a universal protective antigen against invasive disease. Infect. Immun.1997,65:3701–3707.
Lorenzen N, Lorenzen E, Einer-Jensen K, LaPatra SE. DNA vaccines as a tool for nalysing theprotective immune response against rhabdoviruses in rainbow rout. Fish Shellfish Immunol.2002,12:439–453.
Maiti B, Shetty M, Shekar M, Karunasagar I, Karunasagar I. Recombinant outer membraneprotein A （OmpA） of Edwardsiella tarda, a potential vaccine candidate for fish, commoncarp. Microbiol. Res.2011,167:1–7.
Marandi MV, Mittal KR. Role of outer membrane proteinH（OmpH）-and OmpA specificmonoclonal antibodies from hybridoma tumors in protection of mice against Pasteurellamultocida. Infect. Immun.1997,65:4502–4508.
Mariam DH, Mengistu Y, Hoffner SE, Andersson DI. Effect of rpoB mutations conferringrifampin resistance on fitness of Mycobacterium tuberculosis. Antimicrob Agents Chemother.2004,48:1289–1294.
Martin P, Sun L, Hood D, Moxon ER. Involvement of genes of genome maintenance in theregulation of phase variation frequencies in Neisseria meningitides. Microbiology.2004,150:3001–3012.
Matsuyama T, Kamaishi T, Ooseko N, Kurohara K, Iida T. Pathogenicity of motile and non-motileEdwardsiella tarda to some marine fsh. Fish Pathol.2005,40:133–135.
Maughan H, Galeano B, Nicholson WL. Novel rpoB mutations conferring rifampin resistance onBacillus subtilis: global effects on growth, competence, sporulation, and germination. J.Bacteriol.2004,186:2481–2486.
Maurice S, Nussinovitch A, Jaffe N, Shoseyov O, Gertler A. Oral immunization f Carassiusauratus with modifed recombinant A-layer proteins entrapped in lginate beads. Vaccine.2004,23:450–459.
Mekuchi T, Kiyokawa T, Honda K, Nakai T, Muroga K. Vaccination trials in the Japanese flounderagainst Edwardsiellosis. Fish Pathol.1995,30:251–256.
Meyer FP, Bullock GL. Edwardsiella tarda, a new pathogen of channel catfish （Ictaluruspunctatus）. Appl. Microbiol.1973,25:155–156.
Mikamo H, Ninomiya M, Sawamura H, Tamaya T. Puerperal intrauterine infection caused byEdwardsiella tarda. J. Infect. Chemother.2003,9:341–343.
Mikkelsen H, Lund V, Larsen R, Seppola M. Vibriosis vaccines based on various sero-subgroupsof Vibrio anguillarum O2induce specific protection in Atlantic cod （Gadus morhua L.）juveniles. Fish Shellfish Immunol.2011,30:330–339.
Milton DL, O'Toole R, Horstedt P, Wolf-Watz H. Flagellin A is essential for the virulence of Vibrioanguillarum. J. Bacteriol.1996,178:1310–1319.
Mitchison M, Wei L, Kwang J, Wilkie I, Adler B. Overexpression and immunogenicity of theOma87outer membrane protein of Pasteurella multocida. Vet. Microbiol.2000,72:91–96.
Miyazaki T, Egusa S. Histopathological studies of edwardsiellosis of the Japanese eel （Anguillajaponica）,1: suppurative interstitial nephritis form. Fish Pathol.1976,11:33–43.
Miyazaki T, Kaige N. Comparative histopathology of edwardsiellosis in fishes. Fish Pathol.1985,20:219–227.
Mizunoe S, Yamasaki T, Tokimatsu I, Matsunaga N, Kushima H, Hashinaga K, Kadota JI. Acase of empyema caused by Edwardsiella tarda. J. Infect.2006,53:255–258.
Mohanty BR, Sahoo PK. Edwardsiellosis in fsh: a brief review. J. Biosci.2007,32:1331–1344.
Moorman DR, Mandell GL. Characteristics of rifampin-resistant variants obtained from clinicalisolates of Staphylococcus aureus. Antimicrob Agents Chemother.1981,20:709–713.
Moral CH, Castillo EFd, Fierro PL, Cortes AV, Castillo JA, Soriano AC, et al. Molecularcharacterization of the Aeromonas hydrophila aroA gene and potential use of an auxotrophicaroA mutant as a live attenuated vaccine. Infect. Immun.1998,66:1813–1821.
Mo ZL, Xiao P, Mao YX, Zou YX, Wang B, Li J, et al. Construction and characterization of a live,attenuated esrB mutant of Edwardsiella tarda and its potential as a vaccine against thehaemorrhagic septicaemia in turbot, Scophthamus maximus （L.） Fish Shellfish Immunol.2007,23:521–530.
Müller NF, Kaiser PO, Linke D, Schwarz H, Riess T, Sch fer A, Eble JA, Kempf VA. Trimericautotransporter adhesin-dependent adherence of Bartonella henselae, Bartonella quintana,and Yersinia enterocolitica to matrix components and endothelial cells under static anddynamic flow conditions. Infect Immun.2011,79:2544–53.
Nakatsugawa T. Edwardsiella tarda isolated from cultured young flounder. Fish Pathol.1983,18:99–101.
Nelson JJ, Nelson CA, Carter JE. Extraintestinal manifestations of Edwardsiella tarda infection: a10-year retrospective review. J. La. State. Med. Soc.2009,161:103–106.
Newman SG. Bacterial vaccines for fish. Annu. Rev. Fish Dis.1993,3:145–185.
Nguyen HT, Kana K, Yoshikoshi K. Ecological investigation of Streptococcus iniae in culturedJapanese flounder （Paralichthys olivaceus） using selective isolation procedures.Aquaculture.2002,205:7–17.
Norqvist A, Hagstrom A,Wolf-Watz H. Protection of rainbow trout against vibriosis andfurunculosis by the use of attenuated strains of Vibrio anguillarum. Appl. Environ. Microbiol.1989,55:1400–1405.
Nougayrede PH, Vuillaume A, Vigneulle M, Faivre B, Luengo S, Delprat J. First isolation ofEdwardsiella tarda from diseased turbot （Scophthalmus maximus） reared in a sea farm inthe Bay of Biscay. Bull. Eur. Assoc. Fish. Pathol.1994,14:128–129.
Okuda J, Kiriyama M, Yamanoi E, Nakai T. The type III secretion system-dependent repression ofNF-kappaB activation to the intracellular growth of Edwardsiella tarda in human epithelialcells. FEMS. Microbiol. Lett.2008,283:9–14.
Panangala VS, Shoemaker CA, McNulty ST, Arias CR, Klesius PH. Intra-and interspecificphenotypic characteristics of fish-pathogenic Edwardsiella ictaluri and E. tarda. Aquacult.Res.2006,37:49–60.
Park SI. Disease control in Korean aquaculture. Fish Pathol.2009,44:19–23.
Park S, Wakabayashi H, Watanabe Y. Serotype and virulence of Edwardsiella tarda isolated fromeel and their environment. Fish. Pathol.1983,18:85–89.
Pasnik DJ, Evans JJ, Klesius PH. Passive immunization of Nile tilapia （Oreochromis niloticus）provides signifcant protection against Streptococcus agalactiae. Fish Shellfsh Immunol.2006,21:365–371.
Pasnik DJ, Smith SA. Immune and histopathologic responses of DNA-vaccinated ybrid stripedbass Morone saxatilis_M. chrysops after acute Mycobacterium arinum infection. Dis. Aquat.Org.2006,73:33–41.
Pasnik DJ, Smith SA. Immunogenic and protective effects of a DNA vaccine for ycobacteriummarinum in fish. Vet. Immunol. Immunopathol.2005,103:195–206.
Pirarat N, Maita M, Endo M, Katagiri T. Lymphoid apoptosis in Edwardsiella tarda septicemia intilapia, Oreochromis niloticus. Fish Shellfish Immunol.2007,22:608–616.
Purcell MK, Nichols KM, Winton JR, Kurath G, Thorgaard GH, Wheeler P, et al.Comprehensive gene expression profiling following DNA vaccination of rainbow troutagainst infectious hematopoietic necrosis virus. Mol. Immunol.2006,43:2089–106.
Qin H, Jin X, Huang W, Liu Y. Production of an anti-idiotypic antibody single chain variablefragment vaccine against Edwardsiella tarda. Acta. Biochim. Biophys. Sin.（Shanghai）2010,42:129–136.
Quentel C, Vigneulle M. Antigen uptake and immune responses after immersion vaccination. In:Fish Vaccinology, Developments in Biological Standardization. Gudding R, Lillehaug A,Midtlyng PJ, Brown F （Ed.）. Karger, Basel, Switzerland.1997,90:69–78.
Rao PSS, Lim TM, Leung KY. Opsonized virulent Edwardsiella tarda strains are able to adhere toand survive and replicate within fish phagocytes but fail to stimulate reactive oxygenintermediates. Infect. Immun.2001,69:5689–5697.
Rashid MM, Honda K, Nakai T, Muroga K. An ecological study on Edwardsiella tarda in flounderfarms. Fish. Pathol.1994,29:221–227.
Rashid MM, Nakai T, Muroga K, Miyazaki T. Pathogenesis of experimental edwardsiellosis inJapanese founder Paralichthys olivaceus. Fish Sci.1997,63:384–387.
Rizzitello AE, Harper JR, Silhavy TJ. Genetic evidence for parallel pathways of chaperoneactivity in the periplasm of Escherichia coli. J. Bacteriol.2001,183:6794–6800.
Robinson A, Hudson MJ, Cranage MP. Vaccine potocols. In Overview of Vaccines.2nd edition.Edited by Ada G. Totowa: Humana Press.2003,1–17.
Romalde JL, Luzardo-Alvarez A, Ravelo C, Toranzo AE, Blanco-Mendez J. Oral mmunizationusing alginate microparticles as a useful strategy for booster accinationf sahgainstlactoccocosis. Aquaculture.2004.236:119–129.
Ruffolo CG, Adler B. Cloning, sequencing, expression, and protective capacity of the oma87geneencoding the Pasteurella multocida87-kilodalton outer membrane protein. Infect. Immun.1996,64:3161–3167.
Sae-Oui D, Muroga K, Nakai T. A case of Edwardsiella tarda infection in cultured colored carpCyprinus carpio. Fish. Pathol.1984,19:197–199.
Sahoo PK, Mukherjee SC, Sahoo SK. Aeromonas hydrophila versus Edwardsiella tarda: apathoanatomical study in Clarias batrachus. J. Aquacult.1998,6:57–66.
Sakai T, Matsuyama T, Sano M, Iida T. Identification of novel putative virulence factors, adhesinAIDA and type VI secretion system, in atypical strains of fish pathogenic Edwardsiella tardaby genomic subtractive hybridization. Microbiol. Immunol.2009,53:131–139.
Sakazaki R. A proposed group of the family Enterobacteriaceae, the asakusa group. Int. J. Syst.Evol. Microbiol.1965,15:45–47.
Salati F, Kawai K, Kusuda R. Immune response of eel to Edwardsiella tarda lipopolysaccharide.Fish Pathol.1984,19:187–192.
Salati F, Kusuda R. Vaccine preparations used for immunization of eel Anguilla japonica againstEdwardsiella tarda infection. Bull. Jpn. Soc. Sci. Fish.1985,51:1233–1237.
Sambrook J, Fritsch EF, Maniatis T. Molecular cloning: a laboratory manual. New York: ColdSpring Harbor Laboratory Press.1989.
Samuelson P, Gunneriusson E, Nygren PA, Stahl S. Display of proteins on bacteria. J. Biotechnol.2002,96:129–154.
Savan R, Kono T, Itami T, Sakai M. Loop-mediated isothermal amplification: an emergingtechnology for detection of fish and shellfish pathogens. J. Fish. Dis.2005,28:573–581.
Sekeyova Z, Kowalczewska M, Decloquement P, Pelletier N, Spitalska E, Raoult D.Identification of protein candidates for the serodiagnosis of Q fever endocarditis by animmunoproteomic approach. Eur. J. Clin. Microbiol.2009,28:287–295.
Seternes T, Tonheim TC, L voll M, B gwald J, Dalmo RA. Specific endocytosis and degradationof naked DNA in the endocardial cells of cod （Gadus morhua L.）. J Exp. Biol.2007,210:2091–2103.
Shoemaker CA, Klesius PH, Bricker JM. Efficacy of a modified live Edwardsiella ictaluri vaccinein channel catfish as young as7days post hatch. Aquaculture.1999,176:189–193.
Shoemaker CA, Klesius PH, Evans JJ. Immunization of eyed channel catfish, Ictalurus punctatus,eggs with monovalent Flavobacterium columnare vaccine and bivalent F. columnare andEdwardsiella ictaluri vaccine. Vaccine.2007,25:1126–1131.
Shoemaker CA, Klesius PH, Evans JJ. Prevalence of Streptococcus iniae in tilapia, hybrid stripedbass, and channel catfish on commercial fish farms in the United States. Am. J. Vet. Res.2001,62:174–177.
Shoemaker CA, Lafrentz BR, Klesius PH, Evans JJ. Protection against heterologous Streptococcusiniae isolates using a modified bacterin vaccine in Nile tilapia, Oreochromis niloticus （L.）.J. Fish. Dis.2010,33:537–544.
Slaven EM, Lopez FA, Hart SM, Sanders CV. Myonecrosis caused by Edwardsiella tarda: a casereport and case series of extraintestinal E. tarda infections. Clin. Infect. Dis.2001,32:1430–1433.
Sommerset I, Kross y B, Biering E, Frost P. Vaccines for fish in aquaculture. Expert. Rev.Vaccines.2005,4:89–101.
Srinivasa RPS, Lim TM, Leung KY. Functional genomics approach to the identification ofvirulence genes involved in Edwardsiella tarda pathogenesis. Infect. Immun.2003,71:1343–1351.
Srinivasa RPS, Lim TM, Leung KY. Opsonized virulent Edwardsiella tarda strains are able toadhere to and survive and replicate within fish phagocytes but fail to stimulate reactiveoxygen intermediates. Infect. Immun.2001,69:5689–5697.
Strauss EJ, Ghori N, Falkow S. An Edwardsiella tarda strain containing a mutation in a gene withhomology to shlB and hpmB is defective for entry into epithelial cells in culture. Infect.Immun.1997,65:3924–3932.
Strong PC, Hinchliffe SJ, Patrick H, Atkinson S, Champion OL, Wren BW. Identification andcharacterisation of a novel adhesin Ifp in Yersinia pseudotuberculosis. BMC. Microbiol.2011,28:11–85.
Sun K, Jiao X, Zhang M, Sun L. DNA adenine methylase is involved in the pathogenesis ofEdwardsiella tarda. Vet. Microbiol.2010,141:149–154.
Sun K, Wang H, Zhang M, Xiao Z, Sun L. Genetic mechanisms of multi antimicrobial resistancein a pathogenic Edwardsiella tarda strain. Aquaculture.2009,289:134–139.
Sun K, Zhang W, Hou J, Sun L. Immunoprotective analysis of VhhP2, a Vibrio harveyi vaccinecandidate. Vaccine.2009,27:2733–2740.
Sun Y, Liu CS, Sun L. Comparative study of the immune effect of an Edwardsiella tarda antigenin two forms: Subunit vaccine vs DNA vaccine. Vaccine.2011,29:2051–2057.
Sun Y, Liu C, Sun L. Construction and analysis of the immune effect of an Edwardsiella tardaDNA vaccine encoding a D15-like surface antigen. Fish Shellfish Immunol.2010,30:273–279.
Sun Y, Liu C, Sun L: Identification of an Edwardsiella tarda surface antigen and analysis of itsimmunoprotective potential as a purified recombinant subunit vaccine and a surface-anchoredsubunit vaccine expressed by a fish commensal strain. Vaccine.2010,28:6603–6608.
Swain P, Behura A, Dash S, Nayak SK. Serum antibody response of Indian ajor carp, Labeorohita to three species of pathogenic bacteria. Aeromonas ydrophila, Edwardsiella tarda andPseudomonas fluorescens. Vet. Immunol. Mmunopathol.2007,117:137–141.
Swain P, Nayak SK. Comparative sensitivity of different serological tests for seromonitoring andsurveillance of Edwardsiella tarda infection of Indian major carps. Fish Shellfish Immunol.2003,15:333–340.
Tacal JV Jr, Menez CF: The isolation of Edwardsiella tarda from a dog. Philipp. J. Vet. Med.1969,7:143–145.
Takano T, Matsuyama T, Oseko N, Sakai T, Kamaishi T, Nakayasu C, Sano M, Iida T. Theefficacy of five avirulent Edwardsiella tarda strains in a live vaccine against Edwardsiellosisin Japanese flounder, Paralichthys olivaceus. Fish Shellfish Immunol.2010,29:687–693.
Tamura K, Sakazaki R, McWhorter AC, Kosako Y. Edwardsiella tarda serotyping scheme forinternational use. J. Clin. Microbiol.1988,26:2343–2346.
Tan YP, Lin Q, Wang XH, Joshi S, Hew CL, Leung KY. Comparative proteomic analysis ofextracellular proteins of Edwardsiella tarda. Infect. Immun.2002,70:6475–6480.
Tan YP, Zheng J, Tung SL, Rosenshine I, Leung KY. Role of type III secretion in Edwardsiellatarda virulence. Microbiology.2005,151:2301–2313.
Tchesnokova V, Aprikian P, Kisiela D, Gowey S, Korotkova N, Thomas W, Sokurenko E. Type1Fimbrial Adhesin FimH Elicits an Immune Response That Enhances Cell Adhesion ofEscherichia coli. Infect. Immun.2011,79:3895–904.
TempranoA, Riano J,Yugueros J, Gonzalez P, de Castro L, VillenaA, et al. Potential use of aYersinia ruckeri O1auxotrophic aroA mutant as a live attenuated vaccine. J. Fish. Dis.2005,28:419–427.
Thomas KL, Leduc I, Olsen B, Thomas CE, Cameron DW, Elkins C. Cloning, overexpression,purifcation, and immunobiology of an85-kilodalton outer membrane protein fromHaemophilus ducreyi. Infect. Immun.2001,69:4438–4446.
Thornton JC, Garduno RA, KayWW. The development of live vaccines for furunculosis lackingthe A-layer and O-antigen of Aeromonas salmonicida. J. Fish. Dis.1994,17:195–204.
Thune RL, Fernandez DH, Battista JR. An aroA mutant of Edwardsiella ictaluri is safe andefficacious as a live, attenuated vaccine. J. Aquat. Anim. Health.1999,11:358–372.
Thune RL, Stanley LA, Cooper RK. Pathogenesis of Gram-negative bacterial infections in warmwater fsh.Annu. Rev. Fish Dis.1993,3:37–68.
Tonheim TC, Bogwald J, Dalmo RA. What happens to the DNA vaccine in fish? A review ofcurrent knowledge. Fish Shellfish Immunol.2008,25:1–18.
Tonheim TC, Dalmo RA, Bogwald J, Seternes T. Specific uptake of plasmid DNA withoutreporter gene expression in Atlantic salmon （Salmo salar L.） kidney after intramuscularadministration. Fish Shellfish Immunol.2008,24:90–101.
Uhland FC, Hélie P, Higgins R. Infections of Edwardsiella tarda among brook trout in Quebec. J.Aquat. Anim. Health.2000,12:74–77.
Ullah MA, Arai T. Exotoxic substances produced by Edwardsiella tarda. Fish Pathol.1983,18:71–75.
Utke K, Kock H, Schuetze H, Bergmann SM, Lorenzen N, Einer-Jensen K, et al. Combined DNAimmunization with the glycoprotein gene of viral hemorrhagic septicemia virus andinfectious hematopoietic necrosis virus induces double-specific protective immunity andnonspecific response in rainbow trout. Dev. Comp. Immunol.2008,32:239–252.
Valdezate S, Navarro A, Medina-Pascual MJ, Carrasco G, Saéz-Nieto JA. Molecular screening forrifampicin and fluoroquinolone resistance in a clinical population of Brucella melitensis. J.Antimicrob. Chemother.2010,65:51–53.
Vaughan LM, Smith PR, Foster TJ. An aromatic-dependent mutant of the fish pathogenAeromonas salmonicida is attenuated in fish and is effective as a live vaccine against thesalmonid disease furunculosis. Infect. Immun.1993,61:2172–2181.
Vazquez-Juarez RC, Gomez-Chiarri M, Barrera-Saldana H, Hernandez aavedra N, Dumas S,Ascencio F. Evaluation of DNA vaccination of spotted and bass （Paralabraxmaculatofasciatus） with two major outer-membrane rotein-encoding genes from Aeromonasveronii. Fish Shellfish Immunol.2005,19:153–163.
Verjan N, Hirono I, Aoki T. Genetic loci of major antigenic protein genes of Edwardsiella tarda.Appl. Environ. Microbiol.2005,71:5654–5658.
Vogler AJ, Busch JD, Percy-Fine S, Tipton-Hunton C, Smith KL, Keim P. Molecular Analysis ofRifampin Resistance in Bacillus anthracis and Bacillus cereus. Antimicrob Agents Chemother.2002,46:51–53.
Walton DT, Abbott SL, Janda JM. Sucrose-positive Edwardsiella tarda mimicking a biogroup1strain isolated from a patient with cholelithiasis. J. Clin. Microbiol.1993,31:155–156.
Walton TA, Sousa MC. Crystal structure of Skp, a prefoldin-like chaperone that protects solubleand membrane proteins from aggregation. Mol. Cell.2004,15:367–374.
Wang B, Mo ZL, Xiao P, Li J, Zou YX, Hao B, Li GY. EseD, a putative T3SS transloconcomponent of Edwardsiella tarda, contributes to virulence in fish and is a candidate forvaccine development. Mar. Biotechnol.（NY）2010,12:678–685.
Wang F, Cheng S, Sun K, Sun L. Molecular analysis of the fur （ferric uptake regulator） gene ofa pathogenic Edwardsiella tarda strain. J. Microbiol.2008,46:350–355.
Wang F, Zhang M, Hu Y, Zhang W, Sun L. Regulation of the Edwardsiella tarda haemolysin geneand luxS by EthR. J. Microbiol. Biotechnol.2009,19:765–773.
Wang H, Hu Y, Zhang W, Sun L. Construction of an attenuated Pseudomonas fuorescens strainand evaluation of its potential as a cross-protective vaccine. Vaccine.2009.27:4047–4055.
Wang X, Wang Q, Xiao J, Liu Q, Wu H, Xu L, Zhang Y. Edwardsiella tarda T6SS componentevpP is regulated by esrB and iron, and plays essential roles in the invasion of fish. FishShellfish Immunol.2009,27:469–477.
Wang X, Wang Q, Xiao J, Liu Q, Wu H, Xu L, Zhang Y. Hemolysin EthA in Edwardsiella tardais essential for fish invasion in vivo and in vitro and regulated by two-component systemEsrA-EsrB and nucleoid protein HhaEt.. Fish Shellfish Immunol.2010,29:1082–1091.
White FH, Simpson CF, Williams LE JR. Isolation of Edwardsiella tarda from aquatic animalspecies and surface waters in Florida. J. Wildl. Dis.1973,9:204–209.
Woo PTK, Bruno DW: Fish diseases and disorders. Volume3: viral, bacterial and fungalinfections. In Edwardsiella septicaemias.2ndedition.
Wyatt LE, Nickelson RI, Vanderzant C: Edwardsiella tarda in freshwater catfish and theirenvironment. Appl. Environ. Microbiol.1979,38:710–714.
Xie HX, Yu HB, Zheng J, Nie P, Foster LJ, Mok YK, Finlay BB, Leung KY. EseG, an effector ofthe type III secretion system of Edwardsiella tarda, triggers microtubule destabilization.Infect. Immun.2010,78:5011–5021.
Yang CZ, Wang XH, Huang J. Identification and phylogenetic analysis of athogen-Edwardsiellatarda from cultured turbot （Scophthalmus maximus）. J. hanghai. Fish Univ.2008,17:280–284.
Yang H, Chen J, Yang G, Zhang XH, Liu R, Xue X. Protection of Japanese lounder（Paralichthys olivaceus） against Vibrio anguillarum with a DNA accine containing themutated zinc-metalloprotease gene. Vaccine.2009,27:2150–2155.
Yang Z, Liu Q, Wang Q, Zhang Y. Novel bacterial surface display systems based on outermembrane anchoring elements from the marine bacterium Vibrio anguillarum. Appl. Environ.Microbiol.2008,74:4359–4365.
Yasunaga N, Ogawa S, Hatai K. Characteristics of the fish pathogen Edwardsiella isolated fromseveral species of cultured marine fishes. Bull. Nagasaki Pref. Inst. Fish.1982,8:57–65.
Yoo MH, Huh MD, Kim EH, Lee HH, Jeong HD. Characterization of chloramphenicolacetyltransferase gene by multiplex polymerase chain reaction inmultidrug-resistant strainsisolated from aquatic environments. Aquaculture.2003,217:11–221.
Yu JH, Han JJ, Park KS, Park KH, Park SW. Edwardsiella tarda infection in Korean catfish,silurus asotus, in a Korean fish farm. Aquacult. Res.2009,41:19–26.
Zhang M, Jiao X, Hu Y, Sun L. Attenuation of Edwardsiella tarda virulence by small peptides thatinterfere with LuxS/autoinducer type2quorum sensing. Appl. Environ. Microbiol.2009,75:3882–3890.
Zhang M, Sun K, Sun L. Regulation of autoinducer2production and luxS expression in apathogenic Edwardsiella tarda strain. Microbiology.2008,154:2060–2069.
Zhang M, Sun K, Sun L. Regulation of autoinducer2production and luxS expression in apathogenic Edwardsiella tarda strain. Microbiology.2008,154:2060–2069.
Zhang W, Sun K, Cheng S, Sun L. Characterization of DegQVh, a Serine Protease and aprotective immunogen from a pathogenic Vibrio harveyi strain. Appl. Environ. Microbiol.2008,74:6254–6262.
Zhang W, Sun L. Cloning, characterization and molecular application f a beta-agarase gene fromVibrio sp. Strain V134. Appl. Environ. Microbiol.2007,73:2825–2831.
Zhang W, Sun S, Cheng S, Sun L. Characterization of DegQVh, a serine protease and a protectiveimmunogen from a pathogenic Vibrio harveyi strain. Appl. Environ. Microbiol.2008,74:6254–62.
Zheng J, Leung KY. Dissection of a type VI secretion system in Edwardsiella tarda. Mol.Microbiol.2007,66:1192–1206.
Zheng J, Li N, Tan YP, Sivaraman J, Mok YK, Mo ZL, Leung KY. EscC is a chaperone for theEdwardsiella tarda type III secretion system putative translocon components EseB and EseD.Microbiology.2007,153:1953–1962.
Zheng J, Tung S L, Leung K Y. Regulation of a type III and a putative secretion system inEdwardsiella tarda by EsrC is under the control of a two-component system, EsrA-EsrB.Infect. Immun,2005,73:4127-4137.
