Production of a Bacteria-like Particle Vaccine Targeting Rock Bream (Oplegnathus fasciatus) Iridovirus Using Nicotiana benthamiana

다국어 초록 (Multilingual Abstract)

Viral diseases are extremely widespread infections that change constantly through mutations. To produce vaccines against viral diseases, transient expression systems are employed, and Nicotiana benthamiana (tobacco) plants are a rapidly expanding platform. In this study, we developed a recombinant protein vaccine targeting the major capsid protein (MCP) of iridovirus fused with the lysine motif (LysM) and coiled-coil domain of coronin 1 (ccCor1) for surface display using Lactococcus lactis. The protein was abundantly produced in N. benthamiana in its N-glycosylated form. Total soluble proteins isolated from infiltrated N. benthamiana leaves were treated sequentially with increasing ammonium sulfate solution, and recombinant MCP mainly precipitated at 40–60%. Additionally, affinity chromatography using Ni–NTA resin was applied for further purification. Native structure analysis using size exclusion chromatography showed that recombinant MCP existed in a large oligomeric form. A minimum OD600 value of 0.4 trichloroacetic acid (TCA)-treated L. lactis was required for efficient recombinant MCP display. Immunogenicity of recombinant MCP was assessed in a mouse model through enzyme-linked immunosorbent assay (ELISA) with serum-injected recombinant MCP-displaying L. lactis. In summary, we developed a plant-based recombinant vaccine production system combined with surface display on L. lactis.

참고문헌 (Reference)

1 Trevor Williams, "The Iridoviruses" Elsevier 345-412, 1996

2 J G He, "Systemic disease caused by an iridovirus-like agent in cultured mandarinfish, Siniperca chuatsi (Basilewsky), in China" Wiley 23 (23): 219-222, 2000

3 K Mahardika, "Susceptibility of juvenile humpback grouper Cromileptes altivelis to grouper sleepy disease iridovirus (GSDIV)" Inter-Research Science Center 59 : 1-9, 2004

4 C S Wang, "Studies on epizootic iridovirus infection among red sea bream, Pagrus major (Temminck & Schlegel), cultured in Taiwan" Wiley 26 (26): 127-133, 2003

5 Bao van Nguyen ; 이동욱 ; 이상민 ; 황인환 ; 정강원, "Structural Analysis of Tha4, a Twin-arginine Translocase Protein Localized in Plant Thylakoid Membranes" 한국식물학회 62 (62): 129-136, 2019

6 J. W. Do, "Sequence variation in the gene encoding the major capsid protein of Korean fish iridoviruses" Springer Science and Business Media LLC 150 (150): 351-359, 2004

7 Grigore Pintilie, "Segmentation and Comparative Modeling in an 8.6-Å Cryo-EM Map of the Singapore Grouper Iridovirus" Elsevier BV 27 (27): 1561-1569.e4, 2019

8 V. G. Chinchar, "Ranaviruses (family Iridoviridae): emerging cold-blooded killers" Springer Science and Business Media LLC 147 (147): 447-470, 2002

9 Hajime Shimmoto, "Protection of red sea bream Pagrus major against red sea bream iridovirus infection by vaccination with a recombinant viral protein" Wiley 54 (54): 135-142, 2010

10 박기영 ; 위수진, "Potential of plants to produce recombinant protein products" Springer Science and Business Media LLC 59 (59): 559-568, 2016

11 Shi‐Jian Song, "Plant-based, adjuvant-free, potent multivalent vaccines for avian infuenza virus via Lactococcus surface display" Wiley 63 (63): 1505-1520, 2021

12 Edward P Rybicki, "Plant-based vaccines against viruses" Springer Science and Business Media LLC 11 (11): 2014

13 Hongmin Kim, "Plant-Produced N-glycosylated Ag85A Exhibits Enhanced Vaccine Efficacy Against Mycobacterium tuberculosis HN878 Through Balanced Multifunctional Th1 T Cell Immunity" MDPI AG 8 (8): 189-, 2020

14 Hajime Shinmoto, "Phenotypic Diversity of Infectious Red Sea Bream Iridovirus Isolates from Cultured Fish in Japan" American Society for Microbiology 75 (75): 3535-3541, 2009

15 Y.J. Shin, "Oral immunization of fish against iridovirus infection using recombinant antigen produced from rice callus" Elsevier BV 31 (31): 5210-5215, 2013

16 Se-Ra Park, "Optimization of Ammonium Sulfate Concentration for Purification of Colorectal Cancer Vaccine Candidate Recombinant Protein GA733-FcK Isolated from Plants" Frontiers Media SA 6 : 2015

17 Tjibbe Bosma, "Novel Surface Display System for Proteins on Non-Genetically Modified Gram-Positive Bacteria" American Society for Microbiology 72 (72): 880-889, 2006

18 Y. Q. Wang, "Molecular epidemiology and phylogenetic analysis of a marine fish infectious spleen and kidney necrosis virus-like (ISKNV-like) virus" Springer Science and Business Media LLC 152 (152): 763-773, 2006

19 Murali S, "Molecular characterization and pathogenicity of a grouper iridovirus(GIV)isolated from yellow grouper, Epinephelus awoara(Temminck and Schlegel)" 25 : 91-100, 2002

20 F. H. C. CHUA, "Investigation of outbreaks of a novel disease, 'Sleepy Grouper Disease', affecting the brown-spotted grouper, Epinephelus tauvina Forskal" Wiley 17 (17): 417-427, 1994

21 Zhijian Huang, "Immunological evaluation of Vibrio alginolyticus, Vibrio harveyi, Vibrio vulnificus and infectious spleen and kidney necrosis virus (ISKNV) combined-vaccine efficacy in Epinephelus coioides" Elsevier BV 150 (150): 61-68, 2012

22 Stéphane Pillet, "Immunogenicity and safety of a quadrivalent plant-derived virus like particle influenza vaccine candidate—Two randomized Phase II clinical trials in 18 to 49 and ≥50 years old adults" Public Library of Science (PLoS) 14 (14): e0216533-, 2019

23 Abbas Tanhaeian, "Generation of an engineered food-grade Lactococcus lactis strain for production of an antimicrobial peptide: in vitro and in silico evaluation" Springer Science and Business Media LLC 20 (20): 2020

24 Ganesh Ram R. Visweswaran, "Exploiting the peptidoglycan-binding motif, LysM, for medical and industrial applications" Springer Science and Business Media LLC 2014

25 C. Michon, "Display of recombinant proteins at the surface of lactic acid bacteria: strategies and applications" Springer Science and Business Media LLC 15 (15): 2016

26 Inhwan Hwang ; Eun-Ju Sohn ; Yongjik Lee ; Namjo Park; Minhee Park; Nam Hyung Kim; Soohong Park; Kyungmin Min ; Sungmin Gu ; Youngmin Park ; Jae-Young Song ; Dong-Jun An, "Development of Plant-produced E2 Protein for Use as a Green Vaccine Against Classical Swine Fever Virus" Springer Science and Business Media LLC 61 (61): 241-252, 2018

27 Kawakami H, "Cultured fsh species afected by red sea bream iridoviral disease from 1996 to 2000" 37 : 45-48, 2002

28 Ahran Kim, "Co-Expression Network Analysis of Spleen Transcriptome in Rock Bream (Oplegnathus fasciatus) Naturally Infected with Rock Bream Iridovirus (RBIV)" MDPI AG 21 (21): 1707-, 2020

29 Hsin-I Liu, "Cloning of the Major Capsid Protein (MCP) of Grouper Iridovirus of Taiwan (TGIV) and Preliminary Evaluation of a Recombinant MCP Vaccine against TGIV" MDPI AG 16 (16): 28647-28656, 2015

30 Adelene Ai-Lian Song, "A review on Lactococcus lactis: from food to factory" Springer Science and Business Media LLC 16 (16): 2017

31 Richard A. Kammerer, "A conserved trimerization motif controls the topology of short coiled coils" Proceedings of the National Academy of Sciences 102 (102): 13891-13896, 2005

32 Youngmin Park, "A Novel E2 Glycoprotein Subunit Marker Vaccine Produced in Plant Is Able to Prevent Classical Swine Fever Virus Vertical Transmission after Double Vaccination" MDPI AG 9 (9): 418-, 2021

33 Trevor Williams, "A Decade of Advances in Iridovirus Research" Elsevier 173-248, 2005