鸭多杀性巴氏杆菌OmpH单克隆抗体的制备及阻断ELISA检测方法的建立
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摘要
鸭巴氏杆菌病(Duck pasteurellosis)是由多杀性巴氏杆菌(Pasteurella multocida, P.m)引起鸭的一种急性败血性传染病，又称为鸭出血性败血症或鸭霍乱。多杀性巴氏杆菌为革兰氏阴性菌，不但可以感染各年龄段的鸭只，而且传播途径广泛，几乎所有的家禽和野禽都是易感者。外膜蛋白H(OmpH)是多杀性巴氏杆菌主要外膜蛋白之一，各血清型间同源性较高。OmpH位于菌体表面，是一种孔道形成蛋白，属于非特异性细菌外膜脂蛋白超家族。用纯化的天然OmpH诱导的保护率与全菌的保护率相当；且OmpH和全菌都能诱导高水平的ELISA抗体。因此，以OmpH为免疫原，制备抗OmpH的特异性单克隆抗体，并在此基础上建立特异性较强的检测方法对鸭多杀性巴氏杆菌病的诊断与检测具有重要的意义。
本研究以多杀性巴氏杆菌OmpH重组蛋白为免疫原，免疫6周龄BALB/c小鼠，4次免疫后，小鼠的效价在1:64000～1:512000之间。取效价最高的小鼠脾细胞与SP2/0骨髓瘤细胞，在50%PEG3350的作用下进行融合，建立抗多杀性巴氏杆菌OmpH抗体的杂交瘤细胞。应用OmpH重组蛋白间接ELISA筛选及经过3次连续克隆，获得3株能稳定分泌抗OmpH单克隆抗体的杂交瘤细胞株，分别命名为1A9、1E9和3G7。杂交瘤细胞的腹水单克隆抗体间接ELISA效价1A9和1E9是1.6×10~4，3G7是3.2×10~4；单克隆抗体亚类鉴定表明，1A9、1E9和3G7均为IgG1亚类，轻链均为κ链。经Western blot及IFA证实，3株单克隆抗体与靶抗原具有良好的反应性，且不与鸭疫里默氏杆菌、鸭大肠杆菌病的阳性血清发生反应，说明都具有良好的特异性。将1A9、1E9和3G7体外培养连续传30代，每隔5代检测效价，说明其仍能稳定分泌抗体。
本研究在研制出多杀性巴氏杆菌特异性单克隆抗体的基础上以OmpH重组蛋白为包被抗原，被检血清为阻断剂，1A9、1E9和3G7单克隆抗体为一抗，羊抗鼠酶标抗体为二抗，进行阻断ELISA试验，确定以3G7单克隆抗体为最佳一抗。通过对阻断ELISA各条件的优化，最终确定了OmpH重组蛋白工作浓度为1.0μg/mL；待检血清的工作浓度为1:5；单克隆抗体的工作浓度为1:500；酶标二抗的工作浓度为1:30000建立了能检测多杀性巴氏杆菌特异性抗体的阻断ELISA方法。通过对60份鸭多杀性巴氏杆菌阴性血清样品阻断ELISA检测结果进行统计学分析，确定了该方法的判定标准：当阻断率PI≥37.6%时，血清样品判为阳性，阻断率PI <29.13%时，血清样品判为阴性。阻断率PI在两者之间时，判为可疑。应用该方法对鸭传染性浆膜炎、鸭大肠杆菌病、鸭肝炎、鸭H5N1流感、鸭网状内皮增生症、鸭呼肠孤病的阳性血清进行了检测，结果证明该方法具有良好的特异性。应用该方法对10份鸭阳性血清和10份鸭阴性血清进行检测，结果证明该方法具有良好的敏感性。批内、批间重复试验变异系数均小于10%，说明该方法具有良好的重复性。应用该方法对黑龙江省部分地区采集60份鸭血清样品进行检测，与Western blot检测结果对比，两者的符合率为91.38%。
Duck pasteurellois is caused an acute septic infectious disease by Pasteurella multocida, asduck hemorrhagic septicemia or cholera. Pasteurella multocida is a Gram-negative bacteria thatinfect ducks of all ages, almost all of the poultry and wild birds are susceptible, especiallychickens, ducks and geese. The outer membrane protein H is the major outer membrane proteinsof Pasteurella multocida, high homology among the serotypes. OmpH is a pore-forming proteinrelated to the superfamily of the nonspecific bacterial porins. Purified natural OmpH induced goodprotection rate as the protection rate of whole cell lysate induced; OmpH and whole cell lysatecould induce high levels of ELISA antibody. So Pasteurella multocida OmpH recombinantprotein as immunogen, preparation of OmpH monoclonal antibodies, Using this antibody, a blockELISA method for the detection of Pasteurella multocida was developed, that is of greatsignificance for diagnosis and detection of duck pasteurellois.
6-week-old female BALB/c mice were immunized by Pasteurella multocida OmpHrecombinant protein. spleen cells were fused with myeloma cell Sp2/0in50%polyethylene glycol3350after four immunizations, established hybridoma anti-Pasteurella multocida OmpH antibody.The test of indirect ELISA is used to screen hybridoma cells and limited dilution method wasperformed to subclone the positive clones. Three strains of hybridoma secreted MAb againstPasteurella multocida OmpH are obtained after three subcloning, and are designated as1A9,1E9and3G7. The titers of obtained MAbs from the mouse ascitic fluid are1.6×10~4～3.2×10~4inindirect ELISA test. By using the immunoglobulin subtypes kit, the antibodies are all IgG1with κlight chain. Western blot and IFA Confirme three Mabs are specific to Pasteurella multocidaOmpH.1A9,1E9and3G7in vitro continuous30generations, every5detection efficiency showthat they could secrete stably antibodies.
A blocking ELISA based on the3G7monoclonal antibody was developed to detectantibodies to Pasteurella multocida. The best concentration of antigen that was selected byphalanx experiment was1.0μg/mL; serum dilution and monoclonal antibody dilution were1:5and1:500; the best concentration dilution of second antibody marked with HRP was1:30,000. Thethreshold value of the blocking ELISA was conformed according to the detection result of60negative sera. It was positive when PI was more than37.6%, it was negative when PI was lessthan29.13%. Positive serums to6usual pathogens and of duck were tested by blocking ELISA,the results showed that the blocking ELISA with good specificity. Intro-batch test and Inter-batchtest were less than10%, with good reproducibility.10positive serums and10negative serumswere tested by blocking ELISA, the results showed that the blocking ELISA with good sensitivity.60sera samples collected in helongjiang province were tested by blocking ELISA, the coincidencerate between of ELISA and Western blot was91.3%.
本研究以多杀性巴氏杆菌OmpH重组蛋白为免疫原，免疫6周龄BALB/c小鼠，4次免疫后，小鼠的效价在1:64000～1:512000之间。取效价最高的小鼠脾细胞与SP2/0骨髓瘤细胞，在50%PEG3350的作用下进行融合，建立抗多杀性巴氏杆菌OmpH抗体的杂交瘤细胞。应用OmpH重组蛋白间接ELISA筛选及经过3次连续克隆，获得3株能稳定分泌抗OmpH单克隆抗体的杂交瘤细胞株，分别命名为1A9、1E9和3G7。杂交瘤细胞的腹水单克隆抗体间接ELISA效价1A9和1E9是1.6×10~4，3G7是3.2×10~4；单克隆抗体亚类鉴定表明，1A9、1E9和3G7均为IgG1亚类，轻链均为κ链。经Western blot及IFA证实，3株单克隆抗体与靶抗原具有良好的反应性，且不与鸭疫里默氏杆菌、鸭大肠杆菌病的阳性血清发生反应，说明都具有良好的特异性。将1A9、1E9和3G7体外培养连续传30代，每隔5代检测效价，说明其仍能稳定分泌抗体。
本研究在研制出多杀性巴氏杆菌特异性单克隆抗体的基础上以OmpH重组蛋白为包被抗原，被检血清为阻断剂，1A9、1E9和3G7单克隆抗体为一抗，羊抗鼠酶标抗体为二抗，进行阻断ELISA试验，确定以3G7单克隆抗体为最佳一抗。通过对阻断ELISA各条件的优化，最终确定了OmpH重组蛋白工作浓度为1.0μg/mL；待检血清的工作浓度为1:5；单克隆抗体的工作浓度为1:500；酶标二抗的工作浓度为1:30000建立了能检测多杀性巴氏杆菌特异性抗体的阻断ELISA方法。通过对60份鸭多杀性巴氏杆菌阴性血清样品阻断ELISA检测结果进行统计学分析，确定了该方法的判定标准：当阻断率PI≥37.6%时，血清样品判为阳性，阻断率PI <29.13%时，血清样品判为阴性。阻断率PI在两者之间时，判为可疑。应用该方法对鸭传染性浆膜炎、鸭大肠杆菌病、鸭肝炎、鸭H5N1流感、鸭网状内皮增生症、鸭呼肠孤病的阳性血清进行了检测，结果证明该方法具有良好的特异性。应用该方法对10份鸭阳性血清和10份鸭阴性血清进行检测，结果证明该方法具有良好的敏感性。批内、批间重复试验变异系数均小于10%，说明该方法具有良好的重复性。应用该方法对黑龙江省部分地区采集60份鸭血清样品进行检测，与Western blot检测结果对比，两者的符合率为91.38%。
Duck pasteurellois is caused an acute septic infectious disease by Pasteurella multocida, asduck hemorrhagic septicemia or cholera. Pasteurella multocida is a Gram-negative bacteria thatinfect ducks of all ages, almost all of the poultry and wild birds are susceptible, especiallychickens, ducks and geese. The outer membrane protein H is the major outer membrane proteinsof Pasteurella multocida, high homology among the serotypes. OmpH is a pore-forming proteinrelated to the superfamily of the nonspecific bacterial porins. Purified natural OmpH induced goodprotection rate as the protection rate of whole cell lysate induced; OmpH and whole cell lysatecould induce high levels of ELISA antibody. So Pasteurella multocida OmpH recombinantprotein as immunogen, preparation of OmpH monoclonal antibodies, Using this antibody, a blockELISA method for the detection of Pasteurella multocida was developed, that is of greatsignificance for diagnosis and detection of duck pasteurellois.
6-week-old female BALB/c mice were immunized by Pasteurella multocida OmpHrecombinant protein. spleen cells were fused with myeloma cell Sp2/0in50%polyethylene glycol3350after four immunizations, established hybridoma anti-Pasteurella multocida OmpH antibody.The test of indirect ELISA is used to screen hybridoma cells and limited dilution method wasperformed to subclone the positive clones. Three strains of hybridoma secreted MAb againstPasteurella multocida OmpH are obtained after three subcloning, and are designated as1A9,1E9and3G7. The titers of obtained MAbs from the mouse ascitic fluid are1.6×10~4～3.2×10~4inindirect ELISA test. By using the immunoglobulin subtypes kit, the antibodies are all IgG1with κlight chain. Western blot and IFA Confirme three Mabs are specific to Pasteurella multocidaOmpH.1A9,1E9and3G7in vitro continuous30generations, every5detection efficiency showthat they could secrete stably antibodies.
A blocking ELISA based on the3G7monoclonal antibody was developed to detectantibodies to Pasteurella multocida. The best concentration of antigen that was selected byphalanx experiment was1.0μg/mL; serum dilution and monoclonal antibody dilution were1:5and1:500; the best concentration dilution of second antibody marked with HRP was1:30,000. Thethreshold value of the blocking ELISA was conformed according to the detection result of60negative sera. It was positive when PI was more than37.6%, it was negative when PI was lessthan29.13%. Positive serums to6usual pathogens and of duck were tested by blocking ELISA,the results showed that the blocking ELISA with good specificity. Intro-batch test and Inter-batchtest were less than10%, with good reproducibility.10positive serums and10negative serumswere tested by blocking ELISA, the results showed that the blocking ELISA with good sensitivity.60sera samples collected in helongjiang province were tested by blocking ELISA, the coincidencerate between of ELISA and Western blot was91.3%.
引文
[1] Confer A W. lmmunogens of Pasteurella[J]. Vet Mierobiol,1993,37:353-368
[2] Bhasin,J L.,Can. J.. Serological types of Pasteurella multocida isolated from turkeys andchickens in Canada[J]. Microbiol.,1982,28:1078-1080.
[3] Hofacre C L., Clisson J R. A serotypic survey of Pasteurella multocida isolated frompoultry[J]. Avian Dis,1986,30:632-633.
[4] Rhoades K R, RimLer R B. Somatic serotypes of Pasteurella multocida strains isolatedfrom avian hosts (1976-1988)[J]. Avian Dis,1990,34:193-195.
[5] Turcotl,Stintzi A,Xu J, et al. Fast biological iron chelators:kinetics of iron removalfromhuman differric transferring by multidentate hydroxypyridonates[J]. J Biol. Inorg.Chem,2000,5:634-641.
[6] Moeck GS,Letellier L. Characterization of in vitro interactions between a truncated TonBProtein from Escherichia coli and the outer membrane receptors FhuA and FepA[J]. J Bacteriol,2001,183:2755-2764.
[7] Harper M,Boyce JD,Adler B. Pasteurella multocida pathogenesis:125years afterPasteur[J]. FEMS microbiology letters,2006,265(1):1-10.
[8] Leotta GA,Rivas M,Chinen I,et al. Avian cholera in a southern giant petrel(Macronectesgiganteus)from Antarctica[J]. Journal of wildlife diseases.2003,39(3):732-735.
[9]郭玉璞，王惠民．鸭病防治[M]．北京：金盾出版社，2003.
[10]刘亚刚，刘内生，岳华，等．西昌安宁河流域鸭霍乱的流行病学调查与诊治研究[J]．西南民族大学学报(自然科学版)，1991,17(3):91-95.
[11]龙虎，于学辉，罗薇，等．雏鸭鸭疫巴氏杆菌和P.m混合感染的诊治[J]．西南民族学院学报(自然科学版)，1996,22(4):432-434.
[12]郑明．我国111株由家禽分离的P.m的血清型[J]．畜牧兽医学报，1984,15(1):51-56.
[13]程安春，汪铭书，陈孝跃，等．256株鸭源P.m血清学鉴定及病原特性研究[J]．中国兽医科技，1997,27(7):13-15.
[14]陈佳．鸭常见病诊治要领[M]．合肥:安徽农业科学技术出版社，2004:58-59.
[15] Chung, A.C.K., Durica,D,S,.Clifton,S.W.. Cloning of crustacean ecdysteroid receptorand retinoid-X receptor gene homologs and elevation of retinoid-X-receptor mRNA by retinoicacid. Mol. Cell. Endocrinol,1998b,139:209-227.
[16] May BJ,Zhang Q,Li LL, et al. Complete genomic sequence of Pasteurella multocida,Pm70[J]. Proc Natl Acad Sci USA,2001,98:3460-3465.
[17] Snipes KP,Ghazikhanian GY,Hirsh DC. Fate of Pasteurella multocida in the bloodvascular system of turkeys following intravenous inoculation: comparison of an encapsulated,virulent strain with its avirulentacapsular variant [J]. Avian Dis,1987,31:254-259.
[18] Tsuji M,Matsumoto M. Pathogenesis of fowl cholera: influence of encapsulation on thefate of Pasteurella multocida after intravenous inoculation into turkeys[J]. Avian Dis.1989,33:238-247.
[19] Boyce JD,Adler B. The capsule is a virulence determinant in the pathogenesis ofPasteurella multocida M1404(B:2)[J]. Infect Immun,2000,68:3463-3468.
[20] Chung JY,Wilkie I, Boyce JD, et al. Role of capsule in the pathogenesis of fowl choleracaused by Pasteurella multocida serogroup A[J]. Infect Immun,2001,69:2487-2492.
[21] Chung JY, Wilkie I, Boyce JD, et al. Vaccination against fowl cholera with acapsularPasteurella multocida A:1[J]. Vaccine,2005,23:2751-2755.
[22] Harmon BG,Glisson JR,Latimer KS, et al. Resistance of Pasteurella multocida A:3,4tophagocytosis by turkey macrophages and heterophils[J]. Am J Vet Res,1991,52:1507-1511.
[23] Truscott WM,Hirsh DC. Demonstration of an outer membrane protein withantiphagocytic activity from Pasteurella multocida of avian origin[J]. Infect Immun,1988,56:1538-1544.
[24] Pruimboom IM, Rimler RB, Ackermann MR, et al. Capsular hyaluronic acid-mediatedadhesion of Pasteurella multocida to turkey air sac macrophages[J]. Avian Dis,1996,40:887-893.
[25] Wijewardana TG,Sutherland AD. Bactericidal activity in the sera of mice vaccinatedwith Pasteurella multocida type A[J]. Vet Microbiol,1990,24:55-62.
[26] Ramdani,Adler B. Opsonic monoclonal antibodies against lipopolysaccharide(LPS)antigens of Pasteurella multocida and the role of LPS in immunity[J]. Vet Microbiol,1991,26:335-347.
[27] Harper M,Cox AD,St Michael F, et al. A heptosyltransferase mutant of Pasteurellamultocida produces a truncated lipopolysaccharide structure and is attenuated in virulence[J].Infect Immun,2004,72:3436-3443.
[28] St Michael F,Li J,Cox AD. Structural analysis of the core oligosaccharide fromPasteurella multocida strain X73[J]. Carbohydr Res,2005a,340:1253-1257.
[29] Ram S,Cox AD,Wright JC, et al. Neisserial lipooligosaccharide is a target forcomplement component c4b. Inner core phosphoethanolamine residues define C4b klinkagespecificity[J]. J Biol Chem,2003,278:50853-50862.
[30] Ruffolo CG,Tennent JM,Michalski WP, et al. Identification, purification, andcharacterization of the type4fimbriae of Pasteurella multocida[J]. Infect Immun,1997,65:339-343.
[31] Doughty SW,Ruffolo CG,Adler B. The type4fimbrial subunit gene of Pasteurellamultocida[J]. Vet Microbiol,2000,72:79-90.
[32] Kachlany SC,Planet PJ,DeSalle R, et al. Flp-1, the first representative of a new pilingene subfamily, is required for non-specific adherence of Actinobacillusactinomycetemcomitans[J]. Mol Microbiol,2001b,40:542-554.
[33] Fuller TE,Kennedy MJ,Lowery DE. Identification of Pasteurella multocida virulencegenes in a septicemic mouse model using signature-tagged mutagenesis[J]. Microb Pathog,2000,29:25-38.
[34] Kachlany SC,Planet PJ,DeSalle R, et al. Genes for tight adherence of Actinobacillusactinomycetemcomitans: from plaque to plague to pond scum[J]. Trends Microbiol,2001a,9:429-437.
[35] Harper M,Boyce JD,Wilkie IW, et al. Signature-tagged mutagenesis of Pasteurellamultocida identifies mutants displaying differential virulence characteristics in mice andchickens[J]. Infect Immun,2003,71:5440-5446.
[36] Kimura A,Mountzouros KT,Relman DA, et al. Bordetella pertussis filamentoushemagglutinin:evaluation as a protective antigen and colonization factor in a mouse respiratoryinfection model[J]. Infect Immun,1990,58:7-16.
[37] Mooi FR,Jansen WH,Brunings H, et al. Construction and analysis of Bordetella pertussismutants defective in the production of fimbriae[J]. Microb Pathog,1992,12:127-135.
[38] Tatum FM,Yersin AG,Briggs RE. Construction and virulence of a Pasteurella multocidafhab2mutant in turkeys[J]. Microb Pathog,2005,39:9-17.
[39] Foged NT,Pedersen KB,Elling F. Characterization and biological effects of thePasteurella multocida toxin[J]. FEMS Microbiol Lett,1987,43:45-51.
[40] Lax AJ,Chanter N. Cloning of the toxin gene from Pasteurella multocida and its role inatrophic rhinitis[J]. J Gen Microbiol,1990,136:81-87.
[41] Murphy AC,Rozengurt E. Pasteurella multocida toxin selectively facilitatesphosphatidylinositol4,5-bisphosphate hydrolysis by bombesin, vasopressin, and endothelin.Requirement for a functional G protein[J]. J Biol Chem,1992,267:25296-25303.
[42] Wilson BA,Zhu X,Ho M, et al. Pasteurella multocida toxin activates the inositoltriphosphate signaling pathway in Xenopus oocytes via G(q)alpha-coupled phospholipaseC-beta1[J]. J Biol Chem,1997,272:1268-1275.
[43] Sterner-Kock A,Lanske B,Uberschar S, et al..Effects of the Pasteurella multocida toxinon osteoblastic cells in vitro[J]. Vet Pathol,1995,32:274-279.
[44] Mullan PB,Lax AJ. Pasteurella multocida toxin stimulates bone resorption by osteoclastsvia interaction with osteoblasts[J]. Calcif Tissue Int,1998,63:340-345.
[45] Zywietz A,Gohla A,Schmelz M, et al. Pleiotropic effects of Pasteurella multocida toxinare mediated by Gq-dependent and-independent mechanisms.Involvement of Gq but not G11[J].J Biol Chem,2001,276:3840-3845.
[46] Bagley KC,Abdelwahab SF,Tuskan RG, et al. Pasteurella multocida toxin activateshumanmonocyte-derived and murine bone marrow-derived dendritic cells in vitro butsuppresses antibody production in vivo[J]. Infect Immun,2005,73:413-421.
[47] Blocker D,Berod L,Fluhr JW,et al. Pasteurella multocida toxin(PMT)activatesRhoGTPases, induces actin polymerization and inhibits migration of human dendritic cells, butdoes not influence macropinocytosis[J]. Int Immunol,2006,18:459-464.
[48] Petersen SK,Foged NT,Bording A,et al. Recombinant derivatives of Pasteurellamultocida toxin: candidates for a vaccine against progressive atrophic rhinitis[J]. Infect Immun,1991,59:1387-1393.
[49] To H,Someno S,Nagai S. Development of a genetically modified nontoxigenicPasteurella multocida toxin as a candidate for use in vaccines against progressive atrophic rhinitisin pigs[J]. Am J Vet Res,2005,66:113-118.
[50] Snipes KP,Hansen LM,Hirsh DC.Plasma-and iron-regulated expression of highmolecular weight outer membrane proteins by Pasteurella multocida[J]. Am J Vet Res,1988,49:1336-1338.
[51] Choi-Kim K,Maheswaran SK,Felice LJ,et al. Relationship between the iron regulatedouter membrane proteins and the outer membrane proteins of in vivo grown Pasteurellamultocida[J]. Vet Microbiol,1991,28:75-92.
[52] Kennett L,Muniandy N,Mukkur TKS. Comparative Protective Potential of Non-livingIntact Cells and Purified Outer Membrane and Associated Proteins of Pasteurella multocida TypeB:6Grown Under Iron-Regulated Conditions[J]. Pasteurellosis in Production Animals,1993,(43)144-149.
[53] Glisson JR,Contreras MD,Cheng IHN,et al. Cross-protection studies with Pasteurellamultocida bacterins prepared from bacteria propagated in iron-depleted medium[J]. Avian Dis,1993,37:1074-1079.
[54] Ruffolo CG,Jost BH,Adler B. Iron-regulated outer membrane proteins of Pasteurellamultocida and their role in immunity[J]. Vet Microbiol,1998,59:123-137.
[55] Boyce JD,Cullen PA,Nguyen V,et al. Analysis of the Pasteurella multocida outermembrane sub-proteome and its response to the in vivo environment of the natural host[J].Proteomics,2006,6:870-880.
[56] Gray-Owen SD,Schryvers AB. Bacterial transferrin and lactoferrin receptors[J]. TrendsMicrobiol,1996,4:185-191.
[57] Ogunnariwo JA,Schryvers AB. Characterization of a novel transferrin receptor in bovinestrains of Pasteurella multocida[J]. J Bacteriol,2001,183:890-896.
[58] Ewers C,Lubke-Becker A,Bethe A,et al. Virulence genotype of Pasteurella multocidastrains isolated from different hosts with various disease status[J]. Vet Microbiol,2006,114:304-317.
[59] Fuller TE,Kennedy MJ,Lowery DE. Identification of Pasteurella multocida virulencegenes in a septicemic mouse model using signature-tagged mutagenesis[J]. Microb Pathog,2000,29:25-38.
[60] Bosch M,Garrido E,Llagostera M, et al. Pasteurella multocida exbB, exbD and tonBgenes are physically linked but independently transcribed[J]. FEMS Microbiol Lett,2002a,210:201-208.
[61] Carter GR, Chengappa MM. Hyaluronidase production by type B Pasteurella multocidafrom cases of hemorrhagic septicemia[J]. J Clin Microbiol,1980,11:94-96.
[62] Rimler RB,Rhoades KR. Hyaluronidase and chondroitinase activity of Pasteurellamultocida serotype B:2involved in hemorrhagic septicaemia[J]. Vet Rec,1994,134:67-68.
[63] Nikaido H. Molecular basis of bacterial outer membrane permeability revisited[J].Microbiol Mol Biol Rev,2003,67(4):593-656.
[64] Khalid A,Arshad M,Crowley DE. Accelerated decolorization of structurally different azodyes by newly isolated bacterial strains[J]. Microbiol Biotechnol,2008,78:361-369.
[65] Doug M. Smith,Stephen Cusack,Andrew W. Colman, et al. Improved SurfaceTemperature Prediction for the Coming Decade from a Global Climate Model[J].2007,317(5839):796-799.
[66] Dabo SM,Confer AW,Quijano-Blas RA. Molecular and immunological characterizationof Pasteurella multocida serotype A:3OmpA: evidence of its role in P. multocida interaction withextracellular matrix molecules[J]. Microb Pathog,2003,35:147-157.
[67] Ruffolo C.G.,Adler B. Cloning, sequencing, expression, and protective capacity of theoma87gene encoding the Pasteurella multocida87kDa outer membrane antigen[J]. Infect. Immun,1996,64:3161-3167.
[68] Loosmore S.M.,Yang Y.P., Coleman D.C., et al. Outer membrane protein D15isconserved among Haemophilus influenzae species and may represent a universal protectiveantigen against invasive disease[J]. Infect Immun,1997,65:3701-3707.
[69] Manning D.S.,Reschke D.K.,Judd R.C. Omp85Proteins of Neisseria gonorrhoeae andNeisseria meningitidis are similar to Haemophilus infuenzae D-15-Ag and Pasteurella multoeidaOma87[J]. Mierob.Pathog,1998,25:11-2.
[70] Yang Y., Thomas W.R., Chong P.,et al. A20-kDa N-terminal fragment of the D15proteincontains a protective epitope(s) against Haemophilus infuenzae type a and type b[J]. Infect.Immun,1998,66:3349-3354.
[71] Mitchison M., Wei L., Kwang J., et al. Overexpression and immunogenicity of theOma87outer membrane protein of Pasteurella multoeida[J]. Veterinary Microbiology,2000,72:91-96.
[72] Ranjith Babu,Jianying Zhang,Eric J. Beckman,et al. Antimicrobial activities of silverused as a polymerization catalyst for a wound-healing matrix[J]. Biomaterials,2006,27:4304-4314.
[73] Kasten U,Mullenders LH,Hartwig A. Cobalt(II) inhibits the incision and thepolymerization step of nucleotide excision repair in human fibroblasts[J]. Mutation Research,1997,383:81-90.
[74] Borrathybay E,Sawada T,Kataoka Y, et al. Capsule thickness and amounts of a39kDacapsular protein of avian Pasteurella multocida type A strains correlate with their pathogenicity forchickens[J]. Vet Microbiol,2003b,97:215-227.
[75] Ali HA,Sawada T,Hatakeyama H, et al. Characterization of a39kDa capsular protein ofavian Pasteurella multocida using monoclonal antibodies[J]. Vet Microbiol,2004a,100:43-53.
[76] Borrathybay E,Sawada T,Kataoka Y, et al. A39kDa protein mediates adhesion of avianPasteurella multocida to chicken embryo fibroblast cells[J]. Vet Microbiol,2003a,97:229-243.
[77] Al-haj Ali H,Sawada T,Hatakeyama H, et al. Invasion of chicken embryo fibroblast cellsby avian Pasteurella multocida[J]. Vet Microbiol,2004,104:55-62.
[78] Ali HA,Sawada T,Noda K. Protectivity of an immunoaffinity-purified39kDa capsularprotein of avian Pasteurella multocida in mice[J]. J Vet Med Sci,2004b,66:1603-1604.
[79] Merlin C,Gardiner G,Durand S, et al. The Escherichia coli metD locus encodes an ABCtransporter which includes Abc (MetN), YaeE (MetI), and YaeC (MetQ)[J]. J Bacteriol,2002,184:5513-5517.
[80] Luo Y,Glisson JR,Jaekwood MW, et al. Cloning and Chracterization of The Major OutMembrane Protein Gene(OmpH) of Pasteurella multoeida X-73[J]. Bacteriol,1997,179(6):7856-7864.
[81] Vasfi MM,Dubreuil JD,Mittal KR. The32KD Major Outer Membrane Protein ofPasteurella multoeida Capsular SerotypeD[J]. Mierobiology,1996,142(4):199-206.
[82] VasfiMM,Mittal KR. Rule of Outer Membrane Protein H(omPH)and OmpA-specificMonoclonal Antibodies from Hybridoma Turnorsin Protection of Mice Against Pasteurellamultoeida[J]. Infect lmmun,1997,65(l):4502-4508.
[83] Bosch E,Calafell F,Comas D, et al. High-Resolution Analysis of Human Y-ChromosomeVariation Shows a Sharp Discontinuity and Limited Gene Flow Between Northwestern Africa andthe Iberian Peninsula[J]. Am J Hum Genet,2001,68:1019-1029.
[84] Rimler RB. Purification of A Cross-Protective Antigen from Pasteurella multocidaGrown in vitro and in vivo[J]. Avian Dis,2001,45(2):572-580.
[85] Truscott WM.,Hirsh DC. Demonstration of An Outer Membrane Protein withAntiphagoeytic Activity from Pasteurella multocida of Avian Origin[J]. Infect Immun,1988,56(5):1538-1544.
[86]陈红梅，施少华，程龙飞，等．禽多杀性巴氏杆菌C48-1株Omp基因克隆及序列分析[J]．福建农业学报，2008,23(4):351-354.
[87]曹素芳，黄青云．禽多杀性巴氏杆菌C48-1成熟外膜蛋白H重组亚单位疫苗免疫效果的研究[J]．中国兽医科学，2006,36(06):464-467.
[88]李永明，罗阿东，徐景娥，等．猪源多杀性巴氏杆菌的生物学鉴定与荚膜PCR分型[J]．中国预防兽医学报，2007,(07):506-509.
[89] Kasten RW,Carpenter TE. Detection of Pasteurella multocida-specific DNA in turkeyflocks by use of the polymerase chain reaction[J]. Avian Dis,1997,41:676-682.
[90] Townsend KM,Boyce JD,ChungJ Y, et al. Genetic organization of Pasteurella multocidacap Loci and development of a multiplex capsular PCR typing system[J]. J Clin Microblol,2001,39:924-929.
[91]汤细彪，吴斌，刘国平，等．应用巢式PCR方法快速检测猪鼻拭子样品中产毒素多杀性巴氏杆菌[J]．畜牧兽医学报，2007,38(10):1083-1087.
[92] John H,Barnes,Gross W B. Colibacillosis, in Poultry of disease(tenth edition) edited byCalnek B W, John H, Barnes, Charles W. Beard[J]. Iowa State University Press,1997,131-141.
[93] Heddleston KL,Gallagher JE,Rebers PA. Fowl cholera: gel diffusion precipitin test forserotyping Pasteurella multocida from avian species[J]. Avian Disease,1972,16(04):925-936.
[94]吴范庚，钱心元．我国多杀性巴氏杆菌耐热抗原血清型[J]．中国兽医杂志，1987,(12):2-4.
[95]陈笃生．四川省畜禽多杀性巴氏杆菌菌体抗原的血清学鉴定[J]．中国兽医科技，1987,(4):28-29.
[96]胡东良．我国禽源多杀性巴氏杆菌血清型研究[J]．中国兽医杂志，1989,(8):4-6.
[97]毛跟年，许牡丹，杨秀芳．鸽源多杀性巴氏杆菌血清学鉴定[J]．西北农业学报，1999,(3):5-7.
[98]顾万钧．间接血凝试验测定鸡多杀性巴氏杆菌抗体[J]．中国预防兽医学报，1985,(04):35-37.
[99]毛春生，卢中华，崔保安．间接血凝试验检测鸡血清中多杀性巴氏体菌抗体[J]．中国兽医科技，1991,21(7)31-33.
[100] Sorensen KJ,Stralldbygaard B,B?tnerA, et al. Blocking ELISA’s for the distinctionbetween antibodies against European and American strains of porcine reproductive and resplratorysyndrome virus[J]. Veterinary Microbiology,1998,60(2-4):169-177.
[101] van Maanen C,de Boer-Luijtze E,Terpstra C. Development and validation of amonoclonal antibody blocking ELISA for the detection of antibodies against both equineherpesvirus type1(EHV1) and equine herpesvirus type4(EHV4)[J]. Vet Microbiol,2000,71(l-2):37-51.
[102] Carman S,Josephson G, McEwen B, et al. Field validation of a commercial blockingELISA to differentiate antibody to trallsmissible gastroenteritis virus(TGEV) and porcinerespiratory coronavirus and to identify TGEV-infected swine herds[J]. J Vet Diagn Invest,2002,14(2):97-105.
[103]张文文．副鸡嗜血杆菌单克隆抗体的制备及阻断ELISA检测方法的初步建立
[D]．扬州：扬州大学，2007.
[104] Gut-Winiarska M,Jaeobs L,Kerstens H, et al. A highly specific and sensitive sandwichblocking ELISA based on baculovirus expressed pseudorabies virus glycoprotein B[J]..J VirolMethods,2000,88(1):63-71.
[105]王蓓蕾．牛传染性鼻气管炎病毒gB单克隆抗体的制备与阻断EL1SA检测方法的建立[D]．长春：吉林农业大学，2011.
[106] Sun H,Miao D,Zhang P, et al. A comparison of a blocking ELISA and ahaemagglutination inhibition assay for the detection of antibodies to Avibacterium(Haemophilus)paragallinarurn in Sera from artificially infected chiekens[J]. Biologicals,2007,35(4):317-320.
[107] Cho HJ,Entz SC, Green GT, et al. A blocking ELISA with improved sensitivity for thedetection of passively acquired maternal antibodies to BHV-1[J]. Can Vet J,2002,43(l):43-45.
[108] Chenard G,Miedema K,Moonen P, et al. A solid-phase blocking ELISA for detection oftype O foot-and-mouth disease virus antibodies suitable for mass serology[J]. J Virol Methods,2003,107(l):89-98.
[109] Prabakaran M,Ho HT,Prabhu N, et al. Development of epitope-blocking ELISA foruniversal detection of antibodies to human H5N1infiuenza viruse[J]. PloS ONE,2009,4(2):e4566.
[110] Shien JH,Fu LF,Wu JR, et al. Development of blocking ELISA for detection ofantibodies against avian infiuenza virus of the H7subtype[J]. J Microbiol lmnlunol lnfect,2008,41(5):369-376.
[111]朱远茂．牛传染性鼻气管炎ELISA诊断方法的建立[J]．中国兽医科技，2005,35(12):959-963.
[112]胡春燕，刘全忠．单克隆抗体制备技术及应用研究进展[J]．安徽预防医学杂志，2008,02:116-118.
[113]董志伟．抗体工程[M]．北京：北京医科大学出版社，2000:1-18.
[114] Rosenberg SA. Progress in human tumor immunology and immunotherapy[J]. Nature,2001,411:380-384.
[115] Charles AJ. Immunobiology[M].4th ed.CB publication,1999:57-59.
[116] Zouoli M,Stollar B D. A rapid ELISA for measurement of antibodies to nucleic acidantigens using UV-treated polystyrene microtitres[J]. J. Immunol.Methods,1986,90(1):105-110.
[117] Campbell AM. Monoclonal antiboday and immunosensor techology[M]. ElsevierScience Publiser BV,1991,189-203.
[118]刘玉斌．动物免疫学试验技术[M]．长春：吉林科学技术出版社，1989.
[119]周宗安等译．单克隆抗体技术手册[M]．南京：南京大学出版社，1991.
[120]韦英益．猪瘟病毒SM株单克隆抗体的制备、初步鉴定及应用研究[D]．南宁：广西大学，2004,37.
[121]朱平，冯书章．抗体技术实验[M]．长春：长春出版社，1994.
[122] Yi-Cheng Chen, Chien-Hao Chen, Ching-Ho Wang. H5Antibody Detection byBlocking Enzyme-Linked Immunosorbent Assay Using a Monoclonal Antibody[J]. AVIANDISEASES,2008,(52):124-129.
[123] Tian Kegong,Yu Xiuling,ZhaoTiezhu, et al. Emergence of fatal PRRSV variantsUnparalleled outbreaks of atypical PRRS in China and molecular dissection of the uniquehallmark [J]. PLoS ONE,2007,(6):1-10.
[124]占春霞.诺氟沙星单克隆抗体的筛选制备及初步鉴定[D].暨南大学硕士学位论文,2007.
[125]梁荣，张耀相，刘再超，等．间接ELISA法对禽霍乱抗体的动态监测[J]．中国预防兽医学报，1999,21(6):469-470.
[2] Bhasin,J L.,Can. J.. Serological types of Pasteurella multocida isolated from turkeys andchickens in Canada[J]. Microbiol.,1982,28:1078-1080.
[3] Hofacre C L., Clisson J R. A serotypic survey of Pasteurella multocida isolated frompoultry[J]. Avian Dis,1986,30:632-633.
[4] Rhoades K R, RimLer R B. Somatic serotypes of Pasteurella multocida strains isolatedfrom avian hosts (1976-1988)[J]. Avian Dis,1990,34:193-195.
[5] Turcotl,Stintzi A,Xu J, et al. Fast biological iron chelators:kinetics of iron removalfromhuman differric transferring by multidentate hydroxypyridonates[J]. J Biol. Inorg.Chem,2000,5:634-641.
[6] Moeck GS,Letellier L. Characterization of in vitro interactions between a truncated TonBProtein from Escherichia coli and the outer membrane receptors FhuA and FepA[J]. J Bacteriol,2001,183:2755-2764.
[7] Harper M,Boyce JD,Adler B. Pasteurella multocida pathogenesis:125years afterPasteur[J]. FEMS microbiology letters,2006,265(1):1-10.
[8] Leotta GA,Rivas M,Chinen I,et al. Avian cholera in a southern giant petrel(Macronectesgiganteus)from Antarctica[J]. Journal of wildlife diseases.2003,39(3):732-735.
[9]郭玉璞，王惠民．鸭病防治[M]．北京：金盾出版社，2003.
[10]刘亚刚，刘内生，岳华，等．西昌安宁河流域鸭霍乱的流行病学调查与诊治研究[J]．西南民族大学学报(自然科学版)，1991,17(3):91-95.
[11]龙虎，于学辉，罗薇，等．雏鸭鸭疫巴氏杆菌和P.m混合感染的诊治[J]．西南民族学院学报(自然科学版)，1996,22(4):432-434.
[12]郑明．我国111株由家禽分离的P.m的血清型[J]．畜牧兽医学报，1984,15(1):51-56.
[13]程安春，汪铭书，陈孝跃，等．256株鸭源P.m血清学鉴定及病原特性研究[J]．中国兽医科技，1997,27(7):13-15.
[14]陈佳．鸭常见病诊治要领[M]．合肥:安徽农业科学技术出版社，2004:58-59.
[15] Chung, A.C.K., Durica,D,S,.Clifton,S.W.. Cloning of crustacean ecdysteroid receptorand retinoid-X receptor gene homologs and elevation of retinoid-X-receptor mRNA by retinoicacid. Mol. Cell. Endocrinol,1998b,139:209-227.
[16] May BJ,Zhang Q,Li LL, et al. Complete genomic sequence of Pasteurella multocida,Pm70[J]. Proc Natl Acad Sci USA,2001,98:3460-3465.
[17] Snipes KP,Ghazikhanian GY,Hirsh DC. Fate of Pasteurella multocida in the bloodvascular system of turkeys following intravenous inoculation: comparison of an encapsulated,virulent strain with its avirulentacapsular variant [J]. Avian Dis,1987,31:254-259.
[18] Tsuji M,Matsumoto M. Pathogenesis of fowl cholera: influence of encapsulation on thefate of Pasteurella multocida after intravenous inoculation into turkeys[J]. Avian Dis.1989,33:238-247.
[19] Boyce JD,Adler B. The capsule is a virulence determinant in the pathogenesis ofPasteurella multocida M1404(B:2)[J]. Infect Immun,2000,68:3463-3468.
[20] Chung JY,Wilkie I, Boyce JD, et al. Role of capsule in the pathogenesis of fowl choleracaused by Pasteurella multocida serogroup A[J]. Infect Immun,2001,69:2487-2492.
[21] Chung JY, Wilkie I, Boyce JD, et al. Vaccination against fowl cholera with acapsularPasteurella multocida A:1[J]. Vaccine,2005,23:2751-2755.
[22] Harmon BG,Glisson JR,Latimer KS, et al. Resistance of Pasteurella multocida A:3,4tophagocytosis by turkey macrophages and heterophils[J]. Am J Vet Res,1991,52:1507-1511.
[23] Truscott WM,Hirsh DC. Demonstration of an outer membrane protein withantiphagocytic activity from Pasteurella multocida of avian origin[J]. Infect Immun,1988,56:1538-1544.
[24] Pruimboom IM, Rimler RB, Ackermann MR, et al. Capsular hyaluronic acid-mediatedadhesion of Pasteurella multocida to turkey air sac macrophages[J]. Avian Dis,1996,40:887-893.
[25] Wijewardana TG,Sutherland AD. Bactericidal activity in the sera of mice vaccinatedwith Pasteurella multocida type A[J]. Vet Microbiol,1990,24:55-62.
[26] Ramdani,Adler B. Opsonic monoclonal antibodies against lipopolysaccharide(LPS)antigens of Pasteurella multocida and the role of LPS in immunity[J]. Vet Microbiol,1991,26:335-347.
[27] Harper M,Cox AD,St Michael F, et al. A heptosyltransferase mutant of Pasteurellamultocida produces a truncated lipopolysaccharide structure and is attenuated in virulence[J].Infect Immun,2004,72:3436-3443.
[28] St Michael F,Li J,Cox AD. Structural analysis of the core oligosaccharide fromPasteurella multocida strain X73[J]. Carbohydr Res,2005a,340:1253-1257.
[29] Ram S,Cox AD,Wright JC, et al. Neisserial lipooligosaccharide is a target forcomplement component c4b. Inner core phosphoethanolamine residues define C4b klinkagespecificity[J]. J Biol Chem,2003,278:50853-50862.
[30] Ruffolo CG,Tennent JM,Michalski WP, et al. Identification, purification, andcharacterization of the type4fimbriae of Pasteurella multocida[J]. Infect Immun,1997,65:339-343.
[31] Doughty SW,Ruffolo CG,Adler B. The type4fimbrial subunit gene of Pasteurellamultocida[J]. Vet Microbiol,2000,72:79-90.
[32] Kachlany SC,Planet PJ,DeSalle R, et al. Flp-1, the first representative of a new pilingene subfamily, is required for non-specific adherence of Actinobacillusactinomycetemcomitans[J]. Mol Microbiol,2001b,40:542-554.
[33] Fuller TE,Kennedy MJ,Lowery DE. Identification of Pasteurella multocida virulencegenes in a septicemic mouse model using signature-tagged mutagenesis[J]. Microb Pathog,2000,29:25-38.
[34] Kachlany SC,Planet PJ,DeSalle R, et al. Genes for tight adherence of Actinobacillusactinomycetemcomitans: from plaque to plague to pond scum[J]. Trends Microbiol,2001a,9:429-437.
[35] Harper M,Boyce JD,Wilkie IW, et al. Signature-tagged mutagenesis of Pasteurellamultocida identifies mutants displaying differential virulence characteristics in mice andchickens[J]. Infect Immun,2003,71:5440-5446.
[36] Kimura A,Mountzouros KT,Relman DA, et al. Bordetella pertussis filamentoushemagglutinin:evaluation as a protective antigen and colonization factor in a mouse respiratoryinfection model[J]. Infect Immun,1990,58:7-16.
[37] Mooi FR,Jansen WH,Brunings H, et al. Construction and analysis of Bordetella pertussismutants defective in the production of fimbriae[J]. Microb Pathog,1992,12:127-135.
[38] Tatum FM,Yersin AG,Briggs RE. Construction and virulence of a Pasteurella multocidafhab2mutant in turkeys[J]. Microb Pathog,2005,39:9-17.
[39] Foged NT,Pedersen KB,Elling F. Characterization and biological effects of thePasteurella multocida toxin[J]. FEMS Microbiol Lett,1987,43:45-51.
[40] Lax AJ,Chanter N. Cloning of the toxin gene from Pasteurella multocida and its role inatrophic rhinitis[J]. J Gen Microbiol,1990,136:81-87.
[41] Murphy AC,Rozengurt E. Pasteurella multocida toxin selectively facilitatesphosphatidylinositol4,5-bisphosphate hydrolysis by bombesin, vasopressin, and endothelin.Requirement for a functional G protein[J]. J Biol Chem,1992,267:25296-25303.
[42] Wilson BA,Zhu X,Ho M, et al. Pasteurella multocida toxin activates the inositoltriphosphate signaling pathway in Xenopus oocytes via G(q)alpha-coupled phospholipaseC-beta1[J]. J Biol Chem,1997,272:1268-1275.
[43] Sterner-Kock A,Lanske B,Uberschar S, et al..Effects of the Pasteurella multocida toxinon osteoblastic cells in vitro[J]. Vet Pathol,1995,32:274-279.
[44] Mullan PB,Lax AJ. Pasteurella multocida toxin stimulates bone resorption by osteoclastsvia interaction with osteoblasts[J]. Calcif Tissue Int,1998,63:340-345.
[45] Zywietz A,Gohla A,Schmelz M, et al. Pleiotropic effects of Pasteurella multocida toxinare mediated by Gq-dependent and-independent mechanisms.Involvement of Gq but not G11[J].J Biol Chem,2001,276:3840-3845.
[46] Bagley KC,Abdelwahab SF,Tuskan RG, et al. Pasteurella multocida toxin activateshumanmonocyte-derived and murine bone marrow-derived dendritic cells in vitro butsuppresses antibody production in vivo[J]. Infect Immun,2005,73:413-421.
[47] Blocker D,Berod L,Fluhr JW,et al. Pasteurella multocida toxin(PMT)activatesRhoGTPases, induces actin polymerization and inhibits migration of human dendritic cells, butdoes not influence macropinocytosis[J]. Int Immunol,2006,18:459-464.
[48] Petersen SK,Foged NT,Bording A,et al. Recombinant derivatives of Pasteurellamultocida toxin: candidates for a vaccine against progressive atrophic rhinitis[J]. Infect Immun,1991,59:1387-1393.
[49] To H,Someno S,Nagai S. Development of a genetically modified nontoxigenicPasteurella multocida toxin as a candidate for use in vaccines against progressive atrophic rhinitisin pigs[J]. Am J Vet Res,2005,66:113-118.
[50] Snipes KP,Hansen LM,Hirsh DC.Plasma-and iron-regulated expression of highmolecular weight outer membrane proteins by Pasteurella multocida[J]. Am J Vet Res,1988,49:1336-1338.
[51] Choi-Kim K,Maheswaran SK,Felice LJ,et al. Relationship between the iron regulatedouter membrane proteins and the outer membrane proteins of in vivo grown Pasteurellamultocida[J]. Vet Microbiol,1991,28:75-92.
[52] Kennett L,Muniandy N,Mukkur TKS. Comparative Protective Potential of Non-livingIntact Cells and Purified Outer Membrane and Associated Proteins of Pasteurella multocida TypeB:6Grown Under Iron-Regulated Conditions[J]. Pasteurellosis in Production Animals,1993,(43)144-149.
[53] Glisson JR,Contreras MD,Cheng IHN,et al. Cross-protection studies with Pasteurellamultocida bacterins prepared from bacteria propagated in iron-depleted medium[J]. Avian Dis,1993,37:1074-1079.
[54] Ruffolo CG,Jost BH,Adler B. Iron-regulated outer membrane proteins of Pasteurellamultocida and their role in immunity[J]. Vet Microbiol,1998,59:123-137.
[55] Boyce JD,Cullen PA,Nguyen V,et al. Analysis of the Pasteurella multocida outermembrane sub-proteome and its response to the in vivo environment of the natural host[J].Proteomics,2006,6:870-880.
[56] Gray-Owen SD,Schryvers AB. Bacterial transferrin and lactoferrin receptors[J]. TrendsMicrobiol,1996,4:185-191.
[57] Ogunnariwo JA,Schryvers AB. Characterization of a novel transferrin receptor in bovinestrains of Pasteurella multocida[J]. J Bacteriol,2001,183:890-896.
[58] Ewers C,Lubke-Becker A,Bethe A,et al. Virulence genotype of Pasteurella multocidastrains isolated from different hosts with various disease status[J]. Vet Microbiol,2006,114:304-317.
[59] Fuller TE,Kennedy MJ,Lowery DE. Identification of Pasteurella multocida virulencegenes in a septicemic mouse model using signature-tagged mutagenesis[J]. Microb Pathog,2000,29:25-38.
[60] Bosch M,Garrido E,Llagostera M, et al. Pasteurella multocida exbB, exbD and tonBgenes are physically linked but independently transcribed[J]. FEMS Microbiol Lett,2002a,210:201-208.
[61] Carter GR, Chengappa MM. Hyaluronidase production by type B Pasteurella multocidafrom cases of hemorrhagic septicemia[J]. J Clin Microbiol,1980,11:94-96.
[62] Rimler RB,Rhoades KR. Hyaluronidase and chondroitinase activity of Pasteurellamultocida serotype B:2involved in hemorrhagic septicaemia[J]. Vet Rec,1994,134:67-68.
[63] Nikaido H. Molecular basis of bacterial outer membrane permeability revisited[J].Microbiol Mol Biol Rev,2003,67(4):593-656.
[64] Khalid A,Arshad M,Crowley DE. Accelerated decolorization of structurally different azodyes by newly isolated bacterial strains[J]. Microbiol Biotechnol,2008,78:361-369.
[65] Doug M. Smith,Stephen Cusack,Andrew W. Colman, et al. Improved SurfaceTemperature Prediction for the Coming Decade from a Global Climate Model[J].2007,317(5839):796-799.
[66] Dabo SM,Confer AW,Quijano-Blas RA. Molecular and immunological characterizationof Pasteurella multocida serotype A:3OmpA: evidence of its role in P. multocida interaction withextracellular matrix molecules[J]. Microb Pathog,2003,35:147-157.
[67] Ruffolo C.G.,Adler B. Cloning, sequencing, expression, and protective capacity of theoma87gene encoding the Pasteurella multocida87kDa outer membrane antigen[J]. Infect. Immun,1996,64:3161-3167.
[68] Loosmore S.M.,Yang Y.P., Coleman D.C., et al. Outer membrane protein D15isconserved among Haemophilus influenzae species and may represent a universal protectiveantigen against invasive disease[J]. Infect Immun,1997,65:3701-3707.
[69] Manning D.S.,Reschke D.K.,Judd R.C. Omp85Proteins of Neisseria gonorrhoeae andNeisseria meningitidis are similar to Haemophilus infuenzae D-15-Ag and Pasteurella multoeidaOma87[J]. Mierob.Pathog,1998,25:11-2.
[70] Yang Y., Thomas W.R., Chong P.,et al. A20-kDa N-terminal fragment of the D15proteincontains a protective epitope(s) against Haemophilus infuenzae type a and type b[J]. Infect.Immun,1998,66:3349-3354.
[71] Mitchison M., Wei L., Kwang J., et al. Overexpression and immunogenicity of theOma87outer membrane protein of Pasteurella multoeida[J]. Veterinary Microbiology,2000,72:91-96.
[72] Ranjith Babu,Jianying Zhang,Eric J. Beckman,et al. Antimicrobial activities of silverused as a polymerization catalyst for a wound-healing matrix[J]. Biomaterials,2006,27:4304-4314.
[73] Kasten U,Mullenders LH,Hartwig A. Cobalt(II) inhibits the incision and thepolymerization step of nucleotide excision repair in human fibroblasts[J]. Mutation Research,1997,383:81-90.
[74] Borrathybay E,Sawada T,Kataoka Y, et al. Capsule thickness and amounts of a39kDacapsular protein of avian Pasteurella multocida type A strains correlate with their pathogenicity forchickens[J]. Vet Microbiol,2003b,97:215-227.
[75] Ali HA,Sawada T,Hatakeyama H, et al. Characterization of a39kDa capsular protein ofavian Pasteurella multocida using monoclonal antibodies[J]. Vet Microbiol,2004a,100:43-53.
[76] Borrathybay E,Sawada T,Kataoka Y, et al. A39kDa protein mediates adhesion of avianPasteurella multocida to chicken embryo fibroblast cells[J]. Vet Microbiol,2003a,97:229-243.
[77] Al-haj Ali H,Sawada T,Hatakeyama H, et al. Invasion of chicken embryo fibroblast cellsby avian Pasteurella multocida[J]. Vet Microbiol,2004,104:55-62.
[78] Ali HA,Sawada T,Noda K. Protectivity of an immunoaffinity-purified39kDa capsularprotein of avian Pasteurella multocida in mice[J]. J Vet Med Sci,2004b,66:1603-1604.
[79] Merlin C,Gardiner G,Durand S, et al. The Escherichia coli metD locus encodes an ABCtransporter which includes Abc (MetN), YaeE (MetI), and YaeC (MetQ)[J]. J Bacteriol,2002,184:5513-5517.
[80] Luo Y,Glisson JR,Jaekwood MW, et al. Cloning and Chracterization of The Major OutMembrane Protein Gene(OmpH) of Pasteurella multoeida X-73[J]. Bacteriol,1997,179(6):7856-7864.
[81] Vasfi MM,Dubreuil JD,Mittal KR. The32KD Major Outer Membrane Protein ofPasteurella multoeida Capsular SerotypeD[J]. Mierobiology,1996,142(4):199-206.
[82] VasfiMM,Mittal KR. Rule of Outer Membrane Protein H(omPH)and OmpA-specificMonoclonal Antibodies from Hybridoma Turnorsin Protection of Mice Against Pasteurellamultoeida[J]. Infect lmmun,1997,65(l):4502-4508.
[83] Bosch E,Calafell F,Comas D, et al. High-Resolution Analysis of Human Y-ChromosomeVariation Shows a Sharp Discontinuity and Limited Gene Flow Between Northwestern Africa andthe Iberian Peninsula[J]. Am J Hum Genet,2001,68:1019-1029.
[84] Rimler RB. Purification of A Cross-Protective Antigen from Pasteurella multocidaGrown in vitro and in vivo[J]. Avian Dis,2001,45(2):572-580.
[85] Truscott WM.,Hirsh DC. Demonstration of An Outer Membrane Protein withAntiphagoeytic Activity from Pasteurella multocida of Avian Origin[J]. Infect Immun,1988,56(5):1538-1544.
[86]陈红梅，施少华，程龙飞，等．禽多杀性巴氏杆菌C48-1株Omp基因克隆及序列分析[J]．福建农业学报，2008,23(4):351-354.
[87]曹素芳，黄青云．禽多杀性巴氏杆菌C48-1成熟外膜蛋白H重组亚单位疫苗免疫效果的研究[J]．中国兽医科学，2006,36(06):464-467.
[88]李永明，罗阿东，徐景娥，等．猪源多杀性巴氏杆菌的生物学鉴定与荚膜PCR分型[J]．中国预防兽医学报，2007,(07):506-509.
[89] Kasten RW,Carpenter TE. Detection of Pasteurella multocida-specific DNA in turkeyflocks by use of the polymerase chain reaction[J]. Avian Dis,1997,41:676-682.
[90] Townsend KM,Boyce JD,ChungJ Y, et al. Genetic organization of Pasteurella multocidacap Loci and development of a multiplex capsular PCR typing system[J]. J Clin Microblol,2001,39:924-929.
[91]汤细彪，吴斌，刘国平，等．应用巢式PCR方法快速检测猪鼻拭子样品中产毒素多杀性巴氏杆菌[J]．畜牧兽医学报，2007,38(10):1083-1087.
[92] John H,Barnes,Gross W B. Colibacillosis, in Poultry of disease(tenth edition) edited byCalnek B W, John H, Barnes, Charles W. Beard[J]. Iowa State University Press,1997,131-141.
[93] Heddleston KL,Gallagher JE,Rebers PA. Fowl cholera: gel diffusion precipitin test forserotyping Pasteurella multocida from avian species[J]. Avian Disease,1972,16(04):925-936.
[94]吴范庚，钱心元．我国多杀性巴氏杆菌耐热抗原血清型[J]．中国兽医杂志，1987,(12):2-4.
[95]陈笃生．四川省畜禽多杀性巴氏杆菌菌体抗原的血清学鉴定[J]．中国兽医科技，1987,(4):28-29.
[96]胡东良．我国禽源多杀性巴氏杆菌血清型研究[J]．中国兽医杂志，1989,(8):4-6.
[97]毛跟年，许牡丹，杨秀芳．鸽源多杀性巴氏杆菌血清学鉴定[J]．西北农业学报，1999,(3):5-7.
[98]顾万钧．间接血凝试验测定鸡多杀性巴氏杆菌抗体[J]．中国预防兽医学报，1985,(04):35-37.
[99]毛春生，卢中华，崔保安．间接血凝试验检测鸡血清中多杀性巴氏体菌抗体[J]．中国兽医科技，1991,21(7)31-33.
[100] Sorensen KJ,Stralldbygaard B,B?tnerA, et al. Blocking ELISA’s for the distinctionbetween antibodies against European and American strains of porcine reproductive and resplratorysyndrome virus[J]. Veterinary Microbiology,1998,60(2-4):169-177.
[101] van Maanen C,de Boer-Luijtze E,Terpstra C. Development and validation of amonoclonal antibody blocking ELISA for the detection of antibodies against both equineherpesvirus type1(EHV1) and equine herpesvirus type4(EHV4)[J]. Vet Microbiol,2000,71(l-2):37-51.
[102] Carman S,Josephson G, McEwen B, et al. Field validation of a commercial blockingELISA to differentiate antibody to trallsmissible gastroenteritis virus(TGEV) and porcinerespiratory coronavirus and to identify TGEV-infected swine herds[J]. J Vet Diagn Invest,2002,14(2):97-105.
[103]张文文．副鸡嗜血杆菌单克隆抗体的制备及阻断ELISA检测方法的初步建立
[D]．扬州：扬州大学，2007.
[104] Gut-Winiarska M,Jaeobs L,Kerstens H, et al. A highly specific and sensitive sandwichblocking ELISA based on baculovirus expressed pseudorabies virus glycoprotein B[J]..J VirolMethods,2000,88(1):63-71.
[105]王蓓蕾．牛传染性鼻气管炎病毒gB单克隆抗体的制备与阻断EL1SA检测方法的建立[D]．长春：吉林农业大学，2011.
[106] Sun H,Miao D,Zhang P, et al. A comparison of a blocking ELISA and ahaemagglutination inhibition assay for the detection of antibodies to Avibacterium(Haemophilus)paragallinarurn in Sera from artificially infected chiekens[J]. Biologicals,2007,35(4):317-320.
[107] Cho HJ,Entz SC, Green GT, et al. A blocking ELISA with improved sensitivity for thedetection of passively acquired maternal antibodies to BHV-1[J]. Can Vet J,2002,43(l):43-45.
[108] Chenard G,Miedema K,Moonen P, et al. A solid-phase blocking ELISA for detection oftype O foot-and-mouth disease virus antibodies suitable for mass serology[J]. J Virol Methods,2003,107(l):89-98.
[109] Prabakaran M,Ho HT,Prabhu N, et al. Development of epitope-blocking ELISA foruniversal detection of antibodies to human H5N1infiuenza viruse[J]. PloS ONE,2009,4(2):e4566.
[110] Shien JH,Fu LF,Wu JR, et al. Development of blocking ELISA for detection ofantibodies against avian infiuenza virus of the H7subtype[J]. J Microbiol lmnlunol lnfect,2008,41(5):369-376.
[111]朱远茂．牛传染性鼻气管炎ELISA诊断方法的建立[J]．中国兽医科技，2005,35(12):959-963.
[112]胡春燕，刘全忠．单克隆抗体制备技术及应用研究进展[J]．安徽预防医学杂志，2008,02:116-118.
[113]董志伟．抗体工程[M]．北京：北京医科大学出版社，2000:1-18.
[114] Rosenberg SA. Progress in human tumor immunology and immunotherapy[J]. Nature,2001,411:380-384.
[115] Charles AJ. Immunobiology[M].4th ed.CB publication,1999:57-59.
[116] Zouoli M,Stollar B D. A rapid ELISA for measurement of antibodies to nucleic acidantigens using UV-treated polystyrene microtitres[J]. J. Immunol.Methods,1986,90(1):105-110.
[117] Campbell AM. Monoclonal antiboday and immunosensor techology[M]. ElsevierScience Publiser BV,1991,189-203.
[118]刘玉斌．动物免疫学试验技术[M]．长春：吉林科学技术出版社，1989.
[119]周宗安等译．单克隆抗体技术手册[M]．南京：南京大学出版社，1991.
[120]韦英益．猪瘟病毒SM株单克隆抗体的制备、初步鉴定及应用研究[D]．南宁：广西大学，2004,37.
[121]朱平，冯书章．抗体技术实验[M]．长春：长春出版社，1994.
[122] Yi-Cheng Chen, Chien-Hao Chen, Ching-Ho Wang. H5Antibody Detection byBlocking Enzyme-Linked Immunosorbent Assay Using a Monoclonal Antibody[J]. AVIANDISEASES,2008,(52):124-129.
[123] Tian Kegong,Yu Xiuling,ZhaoTiezhu, et al. Emergence of fatal PRRSV variantsUnparalleled outbreaks of atypical PRRS in China and molecular dissection of the uniquehallmark [J]. PLoS ONE,2007,(6):1-10.
[124]占春霞.诺氟沙星单克隆抗体的筛选制备及初步鉴定[D].暨南大学硕士学位论文,2007.
[125]梁荣，张耀相，刘再超，等．间接ELISA法对禽霍乱抗体的动态监测[J]．中国预防兽医学报，1999,21(6):469-470.