Construction and immunogenicity of a DNA vaccine coexpressing GP3 and GP5 of genotype-I porcine reproductive and respiratory syndrome virus

BMC Veterinary Research

Ning-Yi Jin⁴  Rui Du³  Shou-Wen Du⁴  Xing Chen³  Peng-Peng Xiao⁴  Cun-Xia Liu¹  Hao Liu²  Fu-Long Yan³  Jie Jing⁴  Shu-Bo Wen³  Hui-Jun Lu¹  Wen-Chao Sun³  Jing-Qiang Ren² 

[1] Institute of Military Veterinary, Key Laboratory of Jilin Province for Zoonosis Prevention and Control, Academy of Military Medical Sciences, Changchun 130122, China;Institute of Special Animal and Plant Sciences, Chinese Academy of Agricultural Sciences, Changchun 130122, China;College of Animal Science and Technology, Graduate School, Jilin Agricultural University, Changchun 130118, China;College of Veterinary Medicine, Jilin University, Changchun 130062, China

关键词: Quil A;    Immunogenicity;    DNA vaccine;    GP5;    GP3;    Genotype-I PRRSV;   

Others  :  866773
DOI  :  10.1186/1746-6148-10-128

 received in 2013-11-08, accepted in 2014-05-28,  发布年份 2014

【 摘 要 】

Background

The European (EU) genotype of porcine reproductive and respiratory syndrome virus (Genotype-I PRRSV) has recently emerged in China. The coexistence of Genotype-I and -II PRRSV strains could cause seriously affect PRRSV diagnosis and management. Current vaccines are not able to protect against PRRSV infection completely and have inherent drawbacks. Thus, genetically engineered vaccines, including DNA vaccine and live vector engineered vaccines, have been developed. This study aimed to determine the enhanced immune responses of mice inoculated with a DNA vaccine coexpressing GP3 and GP5 of a Genotype-I PRRSV.

Results

To evaluate the immunogenicity of GP3 and GP5 proteins from European-type PRRSV, three DNA vaccines, pVAX1-EU-ORF3-ORF5, pVAX1-EU-ORF3 and pVAX1-EU-ORF5, were constructed, which were based on a Genotype-I LV strain (GenBank ID: M96262). BALB/c mice were immunized with the DNA vaccines; delivered in the form of chitosan-DNA nanoparticles. To increase the efficiency of the vaccine, Quil A (Quillaja) was used as an adjuvant. GP3 and GP5-specific antibodies, neutralizing antibodies and cytokines (IL-2, IL-4, IL-10 and IFN gamma) from the immunized mice sera, and other immune parameters, were examined, including T-cell proliferation responses and subgroups of spleen T-lymphocytes. The results showed that ORF3 and ORF5 proteins of Genotype-I PRRSV induced GP3 and GP5-specific antibodies that could neutralize the virus. The levels of Cytokines IL-2, IL-4, IL-10, and IFN–γ of the experimental groups were significantly higher than those of control groups after booster vaccination (P < 0.05). The production of CD3⁺CD4⁺ and CD3⁺CD8⁺ T lymphocyte was also induced. T lymphocyte proliferation assays showed that the PRRSV LV strain virus could stimulate the proliferation of T lymphocytes in mice in the experimental group.

Conclusions

Using Quil A as adjuvant, Genotype-I PRRSV GP3 and GP5 proteins produced good immunogenicity and reactivity. More importantly, better PRRSV-specific neutralizing antibody titers and cell-mediated immune responses were observed in mice immunized with the DNA vaccine co-expressing GP3 and GP5 proteins than in mice immunized with a DNA vaccine expressing either protein singly. The results of this study demonstrated that co-immunization with GP3 and GP5 produced a better immune response in mice.

【 授权许可】

   
2014 Ren et al.; licensee BioMed Central Ltd.

【 图 表 】

【 参考文献 】

• [1]Bilodeau R, Dea S, Sauvageau RA, Martineau GP: Porcine reproductive and respiratory syndrome in Quebec. Vet Rec 1991, 129:102-103.
• [2]Wensvoort G, Terpstra C, Pol JM, ter Laak EA, Bloemraad M, de Kluyver EP, Kragten C, van Buiten L, den Besten A, Wagenaar F: Mystery swine disease in The Netherlands: the isolation of Lelystad virus. Vet Q 1991, 13(3):121-130.
• [3]Benfield DA, Nelson E, Collins JE, Harris L, Goyal SM, Robison D, Christianson WT, Morrison RB, Gorcyca D, Chladek D: Characterization of swine infertility and respiratory syndrome (SIRS) virus (isolate ATCC VR-2332). J Vet Diagn Invest 1992, 4(2):127-133.
• [4]Thanawongnuwech R, Thacker B, Halbur P, Thacker EL: Increased production of proinflammatory cytokines following infection with porcine reproductive and respiratory syndrome virus and mycoplasmahyop neumoniae. Clin Diagn Lab Immunol 2004, 11(5):901-908.
• [5]Psikal I, Moutelikova R, Kosinova E, Mojzis M, Smid B, Nejedla E, Indik S, Rodak L: Molecular identification and genotyping of porcine reproductive and respiratory syndrome virus (PRRSV) strains in the Czech and Slovak Republics. Proceedings of the 3rd International Symposium of PRRSV and Aujeszky 1999, 1:175-176.
• [6]Sun YJ, Sun YF: Quarantine and diagnosis of porcine reproductive and respiratory syndrome. Chinese Journal of Veterinary Medicine 1997, 23(2):8-9.
• [7]Zhang ZF, Li XL, Zhang Y, Cha YX, Fang WH: Establishment of RT-PCR method for diagnosing European-type PRRSV and sequencing and analysis of the ORF7 gene. Progress in Veterinary Medicine 2007, 28(5):26-29.
• [8]Huang MQ, Che YL, Chen SY, Jing B, Wei H, Wang LB, Chen SL, Zhou LJ, Zhuang XS: Isolation of European type porcine reproductive andrespiratory syndrome virus FJ0602 strain (PRRSV-FJ0602) and sequence analysis of the ORF7 gene. Chin J Vet Med 2008, 30(3):174-178.
• [9]Feng CY, Liu YH, Yan JH, Gao GF: An infectious clone of the highly pathogenic porcine reproductive and respiratory syndrome virus: Topology of glycoprotein 3 (GP3) addressing the intrachain disulfide bonds. Chinese Sci Bull 2011, 56:2785-2793.
• [10]Meulenberg JJ, Petersen-Den BA, De Kluyver EP, Moormann RJ, Schaaper WM, Wensvoort G: Characterization of proteins encoded by ORFs 2 to 7 of Lelystad virus. J Virol 1995, 206:155-163.
• [11]Miller LC, Fox JM: Apoptosis and porcine reproductive and respiratory syndrome virus. Vet Immunol Immunopathol 2004, 102:131-142.
• [12]Ansari IH, Kwon B, Osorio FA, Pattnaik AK: Influence of N-linked glycosylation of porcine reproductiveand respiratory syndrome virus GP5 on virus infectivity, antigenicity, and ability to induce neutralizing antibodies. J Virol 2006, 80(8):3994-4004.
• [13]Meng XJ: Heterogeneity of porcine reproductive and respiratory syndrome virus: implications for current vaccine efficacy and future vaccine development. Vet Microbiol 2000, 74:309-329.
• [14]Botner A, Strandbygaard B, Sorensen KJ, Have P, Madsen KG, Madsen ES, Alexandersen S: Appearance of acute PRRS-like symptoms in sow herds after vaccination with a modified live PRRS vaccine. Vet Rec 1997, 141:497-499.
• [15]Donnelly JJ, Ulmer JB, Liu MA: DNA vaccines. Life Sci 1997, 60:163-172.
• [16]Manickan E, Karem KL, Rouse BT: DNA vaccines–a modern gimmick or a boon to vaccinology? Crit Rev Immunol 1997, 17:139-154.
• [17]Whalen RG, Davis HL: Use of plasmid DNA for direct gene transfer and immunization. Ann N Y Acad Sci 1995, 772:21-29.
• [18]Mansouri S, Lavigne P, Corsi K, Benderdour M, Beaumont E, Fernandes JC: Chitosan-DNA nanoparticles as non-viral vectors in gene therapy: strategies to improve transfection efficacy. Eur J Pharm Biopharm 2004, 57:1-8.
• [19]Islam MA, Firdous J, Choi YJ, Yun CH, Cho CS: Design and application of chitosan microspheres as oral and nasal vaccine carriers: an updated review. Int J Nanomedicine 2012, 7:6077-6093.
• [20]Oda K, Matsuda H, Murakami T, Katayama S, Ohgitani T, Yoshikawa M: Adjuvant and haemolytic activities of 47 saponins derived from medicinal and food plants. Biol Chem 2000, 381:67-74.
• [21]Adams MM, Damani P, Perl NR, Won A, Hong F, Livingston PO, Ragupathi G, Gin DY: Design and synthesis of potent Quillaja saponin vaccine adjuvants. J Am Chem Soc 2010, 132:1939-1945.
• [22]Dewey C, Charbonneau G, Carman S, Hamel A, Nayar G, Friendship R, Eernisse K, Swenson S: Lelystad-like strain of porcine reproductive and respiratory syndrome virus (PRRSV) identied in Canadian swine. Can Vet J 2000, 41:493-494.
• [23]Kim SH, Roh IS, Choi EJ, Lee C, Lee CH, Lee KH, Lee KK, Song YK, Lee OS, Park CK: A molecular analysis of European porcine reproductive and respiratory syndrome virus isolated in South Korea. Vet Microbiol 2010, 143:394-400.
• [24]Amonsin A, Kedkovid R, Puranaveja S, Wongyanin P, Suradhat S, Thanawongnuwech R: Comparative analysis of complete nucleotide sequence of porcine reproductive and respiratory syndrome virus (PRRSV) isolates in Thailand (US and EU genotypes). J Virol 2009, 16(6):143.
• [25]Zhuang JS, Yuan SS, Zhang JW: Construction and analysis of full length genomic cDNA clone for European-type PRRSV low virulent strain. Chinese Veterinary Science 2008, 38(08):658-664.
• [26]Huang MQ, Zheng M, Chen SL, Che YL, Jiang B, Wang LB, Wei H, Zhou LJ, Zhuang XS, Chen SY: Differences in the physicochemical characteristics and pathogenicity between European type PRRSV and North Ametican type PRRSV. Fujian Journal of Agricultural Science 2012, 27(1):24-26.
• [27]Chen NH, Cao Z, Yu XL, Deng X, Zhao T, Wang L, Liu Q, Li X, Tian K: Emergence of novel European genotype porcine reproductive and respiratory syndrome virus in mainland China. J Gen Viro 2011, 92:880-892.
• [28]Zhao P, Tai C, Cui ZZ: Evolution of porcine reproductive and respiratory syndrome virus under antibody immune selective pressures. Sci China Life Sci 2010, 40(10):952-962.
• [29]Zhao P, Ma CT, Dong X, Cui Z: Evolution of quasispecies diversity for porcine reproductive and respiratory syndrome virus under antibody selective pressures. Sci China Life Sci 2012, 42(8):662-667.
• [30]Bernkop-Schnurch A: Chitosan and its derivatives: potential excipients for peroral peptide delivery systems. Int J Pharm 2000, 194:1-13.
• [31]Porporatto C, Bianco ID, Correa SG: Local and systemic activity of the polysaccharide chitosan at lymphoid tissues after oral administration. J Leukoc Biol 2005, 78:62-69.
• [32]Illum L, Jabbal-Gill I, Hinchcliffe M, Fisher AN, Davis SS: Chitosan as a novel nasal delivery system for vaccines. Adv Drug Deliv Rev 2001, 51:81-96.
• [33]Guang Liu W, De Yao K: Chitosan and its derivatives–a promising non-viral vector for gene transfection. J Control Release 2002, 83:1-11.
• [34]Mao HQ, Roy K, Troung-Le VL, Janes KA, Lin KY, Wang Y, August JT, Leong KW: Chitosan-DNA nanoparticles as gene carriers: synthesis, characterization and transfection efficiency. J Control Release 2001, 70:399-421.
• [35]Xu Q, Wang CH, Pack DW: Polymeric carriers for gene delivery: chitosan and poly(amidoamine) dendrimers. Curr Pharm Des 2010, 16:2350-2368.
• [36]Kensil CR: Saponins as vaccine adjuvants. Crit Rev Ther Drug Carrier Syst 1996, 13(1–2):1-55.
• [37]Kumar M, Behera AK, Matsuse H, Lockey RF, Mohapatra SS: Intranasal IFN-γ gene transfer protects BALB/c mice against respiratory syncytial virus infection. Vaccine 1999, 18(5–6):558-567.
• [38]Kim JJ, Maguire HC Jr, Nottingham LK, Morrison LD, Tsai A, Sin JI, Chalian AA, Weiner DB: Coadministration of IL-12 or IL-10 expression cassettes drives immune responses toward a Th1 phenotype. J Interferon Cytokine Res 1998, 18(7):537-547.
• [39]Martínez-Lobo FJ, Díez-Fuertes F, Simarro I, Castro JM, Prieto C: Porcine reproductive and respiratory syndrome virus isolates differ in their susceptibility to neutralization. Vaccine 2011, 29:6928-6940.
• [40]Lopez OJ, Osorio FA: Role of neutralizing antibodies in PRRSV protective immunity. Vet Immunol Immunopathol 2004, 102(3):155-163.
• [41]Martelli P, Gozio S, Ferrari L, Rosina S, De Angelis E, Quintavalla C, Bottarelli E, Borghetti P: Efficacy of a modified live porcine reproductive and respiratory syndrome virus (PRRSV) vaccine in pigs naturally exposed to a heterologous European (Italian cluster) field strain: clinical protection and cell-mediated immunity. Vaccine 2009, 27:3788-3799.
• [42]Lowe JE, Husmann R, Firkins LD, Zuckermann FA, Goldberg TL: Correlation of cell-mediated immunity against porcine reproductive and respiratory syndrome virus with protection against reproductive failure in sows during outbreaks of porcine reproductive and respiratory syndrome in commercial herds. J Am Vet Med Assoc 2005, 226:1707-1711.
• [43]Meier WA, Galeota J, Osorio FA, Husmann RJ, Schnitzlein WM, Zuckermann FA: Gradual development of the interferon-gamma response of swine to porcine reproductive and respiratory syndrome virus infection or vaccination. Virology 2003, 309:18-31.
• [44]Bassaganya-Riera J, Thacker BJ, Yu S, Strait E, Wannemuehler MJ, Thacker EL: Impact of immunizations with porcine reproductive and respiratory syndrome virus on lymphoproliferative recall responses of CD8+ T cells. Viral Immunol 2004, 17:25-37.
• [45]Parida R: Cell-Mediated Immunity in Porcine Reproductive and Respiratory Syndrome Virus. In Master thesis. University of Nebraska-Lincoln, Veterinary and Biomedical Sciences Department; 2012.
• [46]Gimeno M, Darwich L, Diaz I, de la Torre E, Pujols J, Martín M, Inumaru S, Cano E, Domingo M, Montoya M, Mateu E: Cytokine profiles and phenotype regulation of antigen presenting cells by genotype-I porcine reproductive and respiratory syndrome virus isolates. Vet Res 2011, 42:9. BioMed Central Full Text
• [47]Zhou Z, Liu Q, Hu D, Yue X, Yu X, Zhang Q, Gu X, Ni J, Li X, Zhai X, Tian K: Complete genome sequence of a European genotype porcine reproductive and respiratory syndrome virus in china. Genome Announc 2013, 1(3):175-13.
• [48]Reed LJ, Muench H: A simple method of estimating fifty percent end points. Am J Hyg 1938, 27:493-497.
• [49]Shen G, Jin N, Ma M, Jin K, Zheng M, Zhuang T, Lu H, Zhu G, Jin H, Jin M, Huo X, Qin X, Yin R, Li C, Li H, Li Y, Han Z, Chen Y, Jin M: Immune responses of pigs inoculated with a recombinant fowlpox virus coexpressing GP5/GP3 of porcine reproductive and respiratory syndrome virus and swine IL-18. Vaccine 2007, 25:4193-4202.
• [50]Han S, Jin NY, Lu HJ, Jin KS, Nan WL, Mu WF, Zhao CQ: Immune responses of mice inoculated with a recombinant fowlpox virus expressing ORF5/ORF6 of porcine reproductive and respiratory syndrome virus. Chin J Vet Sci 2010, 30(1):1-5.