鸡传染性法氏囊病病毒VP2与白喉毒素截短体DT390融合蛋白的真核表达及免疫原性检测

doi:10.16431/j.cnki.1671-7236.2021.12.033

Abstract

Abstract: In order to explore whether truncated diphtheria toxin DT390 had immune enhancing effect on VP2 protein of Infectious bursal disease virus (IBDV), in this study, DT390 and VP2 were fused and expressed by Pichia pastoris expression system with diphtheria toxin resistance.The expressed fusion protein DT390-VP2 was identified by SDS-PAGE and Western blotting.Twenty four 21 days old SPF chicks were randomly divided into four groups:VP2 group, DT390-VP2 group, PBS (control group) and B87 group (positive control group).The chicks in each group were immunized on the the 0 and 14th days, respectively.On the 28th day after the first immunization, each chick was inoculated with IBDV BC6/85 strain of 100 BID₅₀.The serum VP2 specific antibody levels of chicks in each group at different time before and after immunization were detected by ELISA.HE staining and pathological damage score were used to evaluate the damage degree of Fabricius, and the immunogenicity and protective function of fusion protein DT390-VP2 in chicks.SDS-PAGE and Western blotting analysis showed that the expressed and purified target protein was glycosylated fusion protein DT390-VP2 (91.26 ku).The results of in vivo test showed that 21 days after the first immunization, the titer of VP2 specific antibody in DT390-VP2 group was significantly and extremely significantly higher than that in B87 group (P<0.05) and VP2 group (P<0.01), respectively.28 days after the first immunization, the titer of VP2 specific antibody in DT390-VP2 group was extremely significantly higher than that in VP2 group (P<0.01).Compared with VP2 protein, the fusion protein DT390-VP2 reduced the atrophy of bursal lymph follicles in chicks.In conclusion, truncated diphtheria toxin DT390 enhanced the humoral immunogenicity of VP2 by fusion expression, the fusion protein DT390-VP2 reduced the damage of tissue caused by IBDV BC6/85 strain to a certain extent.

Key words: Infectious bursal disease virus (IBDV); VP2 protein; DT390; immunogenicity

CLC Number:

S852.65⁺7

MENG Weijin, FAN Kuohai, CHANG Xiongxiong, YIN Wei, LI Hongquan, WANG Zhirui. Eukaryotic Expression and Immunogenicity Detection of Fusion Protein of Chicken Infectious Bursal Disease Virus VP2 and Truncated Diphtheria Toxin DT390[J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(12): 4652-4660.

References

[1] DEY S,PATHAK D C,RAMAMURTHY N,et al.Infectious bursal disease virus in chickens:Prevalence,impact,and management strategies[J].Veterinary Medicine:Research and Reports,2019,10:85-97.
[2] YILMAZ A,TURAN N,BAYRAKTAR E,et al.Phylogeny and evolution of Infectious bursal disease virus circulating in Turkish broiler flocks[J].Poultry Science,2019,98(5):1976-1984.
[3] MYINT O,SUWANRUENGSRI M,ARAKI K,et al.Bursa atrophy at 28 days old caused by variant Infectious bursal disease virus has a negative economic impact on broiler farms in Japan[J].Avian Pathology:Journal of the WVPA,2021,50(1):6-17.
[4] LEE C C,KO T P,CHOU C C,et al.Crystal structure of Infectious bursal disease virus VP2 subviral particle at 2.6A resolution:Implications in virion assembly and immunogenicity[J].Journal of Structural Biology,2006,155(1):74-86.
[5] WU H,YANG H,JI G,et al.SUMO1 modification facilitates Avibirnavirus replication by stabilizing polymerase VP1[J].Journal of Virology,2019,93(10):e02227-18.
[6] LUQUE D,RIVAS G,ALFONSO C,et al.Infectious bursal disease virus is an icosahedral polyploid dsRNA virus[J].Proceedings of the National Academy of Sciences of the United States of America,2009,106(7):2148-2152.
[7] KIBENGE F S,QIAN B,CLEGHORN J R,et al.Infectious bursal disease virus polyprotein processing does not involve cellular proteases[J].Archives of Virology,1997,142(12):2401-2419.
[8] RAUTENSCHLEIN S,ALKIE T N.Infectious bursal disease virus in poultry:Current status and future prospects[J].Veterinary Medicine Research and Reports,2016,7:9-18.
[9] MATA C P,MERTENS J,FONTANA J,et al.The RNA-binding protein of a double-stranded RNA virus acts like a scaffold protein[J].Journal of Virology,2018,92(19):e00968-18.
[10] WANG Z,MI J,WANG Y,et al.Recombinant Lactococcus expressing a novel variant of Infectious bursal disease virus VP2 protein can induce unique specific neutralizing antibodies in chickens and provide complete protection[J].Viruses,2020,12(12):1350.
[11] FAHEY K J,ERNY K,CROOKS J.A conformational immunogen on VP-2 of Infectious bursal disease virus that induces virus-neutralizing antibodies that passively protect chickens[J].The Journal of General Virology,1989,70(Pt 6):1473-1481.
[12] MULLER H,MUNDT E,ETERRADOSSI N,et al.Current status of vaccines against infectious bursal disease[J].Avian Pathology,2012,41(2):133-139.
[13] 王彦伟,张素玲,吴芃,等.鸡传染性法氏囊病病毒rVP2蛋白高效组装亚病毒颗粒的鉴定[J].中国畜牧兽医,2020,47(6):1677-1684. WANG Y W,ZHANG S L,WU P,et al.Identification of subviral particles efficiently self-assembled by Infectious bursal disease virus rVP2 protein[J].China Animal Husbandry & Veterinary Medicine,2020,47(6):1677-1684.(in Chinese)
[14] MISHRA R P N,YADAV R S P,JONES C,et al.Structural and immunological characterization of E.coli derived recombinant CRM(197) protein used as carrier in conjugate vaccines[J].Bioscience Reports,2018,38(5):BSR20180238.
[15] ROBERTSON C A,GREENBERG D P,HEDRICK J,et al.Safety and immunogenicity of a booster dose of meningococcal(groups A,C,W,and Y) polysaccharide diphtheria toxoid conjugate vaccine[J].Vaccine,2016,34(44):5273-5278.
[16] PECETTA S,TONTINI M,FAENZI E,et al.Carrier priming effect of CRM197 is related to an enhanced B and T cell activation in meningococcal serogroup A conjugate vaccination.Immunological comparison between CRM197 and diphtheria toxoid[J].Vaccine,2016,34(20):2334-2341.
[17] COLLIER R J.Diphtheria toxin:Mode of action and structure[J].Bacteriological Reviews,1975,39(1):54-85.
[18] WANG Z,DURAN-STRUUCK R,CREPEAU R,et al.Development of a diphtheria toxin based antiporcine CD3 recombinant immunotoxin[J].Bioconjugate Chemistry,2011,22(10):2014-2020.
[19] 闫伟,杨娜,毛若雨,等.噬菌体裂解酶Lysep3的毕赤酵母重组表达和活性分析[J].中国畜牧兽医,2020,47(4):1018-1027. YAN W,YANG N,MAO R Y,et al.Recombinant expression and activity analysis of phage lyase Lysep3[J].China Animal Husbandry & Veterinary Medicine,2020,47(4):1018-1027(in Chinese)
[20] 尹训南,司昌德.IBDV地方毒株毒力测定的病理组织学检验标准探讨[J].中国预防兽医学报,1997,2:52-54. YIN X N,SI C D.Discussion on histopathological test criteria for virulence determination of local strains of IBDV[J].Chinese Journal of Preventive Veterinary Medicine,1997,2:52-54.(in Chinese)
[21] MARUSIC C,DRISSI TOUZANI C,BORTOLAMI A,et al.The expression in plants of an engineered VP2 protein of Infectious bursal disease virus induces formation of structurally heterogeneous particles that protect from a very virulent viral strain[J].PLoS One,2021,16(2):e0247134.
[22] TAGHAVIAN O,SPIEGEL H,HAUCK R,et al.Protective oral vaccination against Infectious bursal disease virus using the major viral antigenic protein VP2 produced in Pichia pastoris[J].PLoS One,2013,8(12):e83210.
[23] 段马魁.传染性法氏囊病病毒VP2蛋白的体外表达及免疫保护效果评价[D].太谷:山西农业大学,2018. DUAN M K.Expression of Infectious bursal disease virus VP2 protein in vitro and evaluation of its immunoprotective effect[D].Taigu:Shanxi Agricultural University,2018.(in Chinese)
[24] DELIC M,VALLI M,GRAF A B,et al.The secretory pathway:Exploring yeast diversity[J].FEMS Microbiology Reviews,2013,37(6):872-914.
[25] MATTANOVICH D,BRANDUARDI P,DATO L,et al.Recombinant protein production in yeasts[J].Methods in Molecular Biology,2012,824:329-358.
[26] CASADEVALL A,PIROFSKI L A.A reappraisal of humoral immunity based on mechanisms of antibody-mediated protection against intracellular pathogens[J].Advances in Immunology,2006,91:1-44.
[27] BADAHDAH A M,RASHID H,KHATAMI A.Update on the use of meningococcal serogroup C CRM197-conjugate vaccine (Meningitec) against meningitis[J].Expert Review of Vaccines,2016,15(1):9-29.
[28] PARIJS B A,MALINOSKI F J.Post-marketing effectiveness of Prevnar [pneumococcal 7-valent conjugate vaccine (diphtheria CRM197 protein)] and implications for adult immunization[J].Mechanisms of Ageing and Development,2004,125(2):147-148.
[29] 于雷,王秀丽,杨承槐,等.传染性法氏囊病病毒BC6/85株全基因组克隆及其序列分析[J].中国兽药杂志,2013,47(12):17-21. YU L,WANG X L,YANG C H,et al.Sequence analysis of the complete genome of IBDV BC6/85[J].Chinese Journal of Veterinary Drug,2013,47(12):17-21.(in Chinese)

Eukaryotic Expression and Immunogenicity Detection of Fusion Protein of Chicken Infectious Bursal Disease Virus VP2 and Truncated Diphtheria Toxin DT390

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